Clinical trial NCT03881163
Conditions: Respiratory Tract Disorders
Sponsor: Zambon SpA
Start date: 2019-11-11; Completion date: 2020-01-18
Enrollment: 24 participants; Allocation: NON_RANDOMIZED; Intervention model: FACTORIAL

Brief summary:
This is a single and multiple dose, single centre, open-label, one-way, pharmacokinetics, safety and tolerability clinical trial of Phase I to be performed in Chinese healthy male and female volunteers. Twenty-four (24) healthy male and female Chinese volunteers will be included in the study. Drop-out subjects will not be replaced. The study has been designed in agreement with the Chinese Technical Guideline on Clinical Pharmacokinetic Research of Chemical Drugs, 18 March 2005 and the European Guideline on the Investigation of Bioequivalence. No randomisation will take place in this study. All the participant will receive the same treatment with the investigational medicinal product (IMP), i.e. NAC, 300 mg/ 3 mL solution for injection.

Detailed description:

Arms: arm 1: On day 1 at 08:00 ±1 hours, one dose of 600 mg of NAC (300 + 300 mg ampoule) will be administered under fasting conditions. arm 2: On days 4 and 5 at 08:00 ±1 hours and 20:00 ±1 hours and at 08:00 ±1 on day 6, 5 doses of 600 mg of NAC (300 + 300 mg ampoule) will be administered.
Interventions: intervention 1: N-acetylcysteine (NAC)
Intervention descriptions: intervention 1: Two ampoules of IMP (300 + 300 mg) corresponding to a total dose of 600 mg of NAC diluted in 10 mL of NaCl 0.9% sterile saline solution, will be administered by a 5-minute i.v. infusion.

Protocol documents:
Prot_000.pdf: 
• Protocol number: Z7244J01 
• Document title: Phase I Study to Evaluate Pharmacokinetics, Safety, and Tolerability 
of Single and Multiple i.v. Doses of N-acetylcysteine (NAC) in Chinese Healthy 
Volunteers 
• NCT number: NCT03881163 
• Version number: 3.0 
• Date of the document: 03 December 2019 


Clinical Study Report Appendices 
Final 1.0 
20 Nov 2020 
16.1 Study Information 
16.1.1 
Protocol and Protocol Amendments 
The following documents are included: 

Clinical Study Protocol, Version 1.0, dated 30 Aug 2018

Clinical Study Protocol, Version 2.0, dated 19 Dec 2018

Clinical Study Protocol, Version 3.0, dated 03 Dec 2019
Zambon S.p.A., Italy 
Z7244J01 


CONFIDENTIAL 
Sponsor code Z7244J01 
NAC 300 mg/3 mL i.v. bioavailability 
Final version 1.0, 30AUG2018 
Page 1 
CLINICAL STUDY PROTOCOL 
Sponsor code Z7244J01 
Phase I study to evaluate pharmacokinetics, safety and tolerability 
of single and multiple i.v. doses of N-acetylcysteine (NAC) in 
Chinese healthy volunteers 
Single and multiple dose, single centre, open-label, one-way, pharmacokinetics, safety and 
tolerability clinical trial 
Test formulation: 
NAC 300 mg/ 3 mL solution for injection, Zambon S.p.A., 
Italy.
Sponsor: 
Zambon S.p.A., via Lillo del Duca 10, I-20091 Bresso, Italy 
Phone: +39.02.66.52.41 
Fax:
+39.02.66.50.14.92
Investigator: 
“Name Surname”, MD - Principal investigator 
XXXXXXX, Phase I Unit, “Address” 
China 
Phone: +xx.xx.xxxxx 
Fax: 
+xx.xx.xxxxx 
Email: xxx@xxxx
Development phase:
I
Version and date:
Final version 1.0, 30 August 2018 
This study will be conducted in accordance with the current version of Good Clinical Practice (GCP), 
ICH topic E6 (R2 and CFDA GCP) 
Property of the sponsor 
May not be used, divulged, published or otherwise disclosed without the consent of the sponsor 
This document comprises 63 pages 


PPD
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CONFIDENTIAL 
 
 
Sponsor code Z7244J01 
NAC 300 mg/3 mL i.v. bioavailability 
Final version 1.0, 30AUG2018 
 
Page 4 
2 
STUDY SYNOPSIS 
 
Title: Phase I study to evaluate pharmacokinetics, safety and tolerability of single and multiple i.v. doses of 
N-acetylcysteine (NAC) in Chinese healthy volunteers
 
Protocol number: Z7244J01 
 
Clinical phase: Phase I 
 
Study design: Single and multiple dose, single centre, open-label, one-way, pharmacokinetics, safety and 
tolerability clinical trial 
 
Planned nr. of centres / countries: 1/China 
 
Investigator and centre: Principal investigator: “Name Surname”, MD, “Institution name”, Phase I Unit, 
“Address”…., China 
 
Investigational medicinal product (IMP):  
Test product:
NAC, 300 mg/ 3 mL solution for injection, Zambon S.p.A., Italy
 
Dose regimen:  
Two ampoules (300 + 300 mg) corresponding to a total dose of 600 mg of NAC diluted in 10 mL of NaCl 0.9% 
saline solution, will be administered by a 5-minute intravenous (i.v.) infusion. 
Single dose: 
One (1) dose of the investigational product will be administered under fasting conditions on day 1 at 08:00 ±1 h 
Multiple dose regime: 
Five (5) doses of the investigational product will be administered twice a day (b.i.d.) on days 4 and 5 at 08:00 ±1 
h and 20:00 ±1 h and one dose will be administered on day 6 at 08:00 ±1. 
 
Objective: To evaluate the NAC pharmacokinetics, safety and tolerability after single and multiple dose i.v. 
administration.
 
End-points:  
Primary end-point:  
 To evaluate pharmacokinetic parameters of NAC in plasma after single and multiple dose administration of 
the investigational product.  
Secondary end-points:  
 To collect safety and tolerability data after single and multiple dose administration of the investigational 
product. 
 
Study variables: 
Primary variables:  
After single dose: 
 Cmax: maximum NAC plasma concentration 
 tmax: time to achieve Cmax 
 kel: terminal elimination rate constant, calculated, if feasible, by log-linear regression using at least 3 points 
 t1/2: NAC half-life, calculated, if feasible, as ln2/kel 
 AUC(0-t): area under the concentration-time curve from single dose to the last observed concentration time t, 
calculated with the linear up/log down trapezoidal method 
 AUC(0-): area under the concentration-time curve extrapolated to infinity, calculated, if feasible, as AUC(0-t) 
+ Ct/kel, where Ct is the last measurable drug concentration 
 AUC(0-12): area under the concentration-time curve at steady-state in the tau interval (from single dose to 12 
h post-dose), calculated with the linear up/log down trapezoidal method 
 Vd: volume of distribution associated with the terminal slope, calculated, if feasible, as Dose/(AUC(0-)*kel) 
 CLt: total body clearance, calculated, if feasible, as Dose/ AUC(0-) 
 Ae(0-t): total amount of NAC excreted in urine from single dose up to 32 h  
 Fe(0-t): total fraction of NAC dose excreted in urine from single dose up to 32 h 
 CLr: renal clearance, calculated, if feasible, as Ae(0-t)/ AUC(0-) 
 
 


CONFIDENTIAL 
 
 
Sponsor code Z7244J01 
NAC 300 mg/3 mL i.v. bioavailability 
Final version 1.0, 30AUG2018 
 
Page 5 
STUDY SYNOPSIS (cont.) 
 
Study variables (continued): 
After multiple dose: 
 Css_max: maximum NAC plasma concentration at steady-state 
 tss_max: time to achieve Css_max 
 Css_min: trough NAC plasma concentration at steady-state, measured as concentration at t=12 h  
 AUCss(0-t): area under the concentration-time curve at steady-state from the last multiple dose to the last 
observed concentration time t, calculated with the linear up/log down trapezoidal method 
 AUCss(0-12): area under the concentration-time curve at steady-state in the tau interval (from the last multiple 
dose to 12 h post-dose), calculated with the linear up/log down trapezoidal method 
 Css_av: average NAC plasma concentration at steady-state, calculated as AUCss(0-12) /tau  
 R: accumulation ratio, calculated as AUCss(0-12)/ AUC(0-12) 
 DF%: 
degree 
of 
fluctuation 
over 
one 
dosing 
interval 
at 
steady-state, 
calculated 
as 
(Css_max - Css_min)/ Css_av*100 
 Aess(0-t): total amount of NAC excreted in urine from the last multiple dose to 32 h at steady-state 
Secondary variables:  
 Treatment emergent adverse events (TEAEs), vital signs (blood pressure, heart rate), body weight, physical 
examinations, clinical laboratory parameters. 
 
Analytics: total NAC will be determined in plasma and urine at a certified bioanalytical laboratory to be 
designated, using a LC-MS/MS validated method with a Lower Quantification Limit (LQL) of 10 ng/mL. 
Analytical facilities and procedures are in compliance with the general principles of GLP regulations 
 
Sample size: Twenty-four (24) healthy male and female Chinese volunteers will be included in the study. Drop-
out subjects will not be replaced.
Main selection criteria:  
Inclusion criteria: 
1. Informed consent: signed written informed consent before inclusion in the study  
2. Ethnicity, Sex and Age: Chinese males and females, 18-45 year old inclusive 
3. Weight: body weight ≥50 kg; 
4. Body Mass Index: 19-24 kg/m2 inclusive 
5. Vital signs: systolic blood pressure 100-139 mmHg, diastolic blood pressure 50-89 mmHg, heart rate 50-90 
bpm, measured after 5 min at rest in the sitting/supine position (to be chosen according to the usual 
procedure at the clinical site) 
6. Full comprehension: ability to comprehend the full nature and purpose of the study, including possible risks 
and side effects; ability to co-operate with the investigator and to comply with the requirements of the entire 
study 
7. Nicotine addiction (smoker subjects only): ability to abstain from smoking for the duration of the clinical 
study 
8. Contraception and fertility (women only): women of child-bearing potential must be using at least one of the 
following reliable methods of contraception: 
a. Hormonal oral, implantable, transdermal, or injectable contraceptives for at least 60 calendar days before 
the screening visit 
b. A non-hormonal intrauterine device or female condom with spermicide or contraceptive sponge with 
spermicide or diaphragm with spermicide or cervical cap with spermicide for at least 60 calendar days 
before the screening visit 
c. A male sexual partner who agrees to use a male condom with spermicide 
d. A sterile sexual partner 
Women of non-child-bearing potential or in post-menopausal status for at least 1 year will be admitted. 
Women of childbearing potential should be willing to adopt abstinence or contraception measures during the 
study and two weeks post-dose. 
For all women, pregnancy test result must be negative at screening and day -1.
 
 


CONFIDENTIAL 
 
 
Sponsor code Z7244J01 
NAC 300 mg/3 mL i.v. bioavailability 
Final version 1.0, 30AUG2018 
 
Page 6 
STUDY SYNOPSIS (cont.) 
 
Exclusion criteria: 
1. Electrocardiogram (12-lead ECG in supine position): clinically significant abnormalities 
2. Physical findings: clinically significant abnormal physical findings which could interfere with the objectives 
of the study 
3. Laboratory analyses: clinically significant abnormal laboratory values indicative of physical illness, in 
particular significant laboratory abnormality indicative of hepatic condition (more than 3 times the upper 
limit) 
4. Allergy: ascertained or presumptive hypersensitivity to the active principle and/or formulations' ingredients; 
history of anaphylaxis to drugs or allergic reactions in general, which the investigator considers may affect 
the outcome of the study 
5. Diseases: significant history of renal, hepatic, gastrointestinal, cardiovascular, respiratory, skin, 
haematological, endocrine, urologic, metabolic, neurological or psychiatric diseases, as determined by the 
investigator, that may interfere with the aim of the study; history of carcinoma in situ and malignant disease; 
active bacterial or viral infection and fever >38° C within 48 h prior to study treatment administration 
6. Virology: positive result of HIV, hepatitis B (HBV), hepatitis C (HCV) or Treponema pallidum (TP) assays 
7. Surgery: any surgery within 2 months of screening (excluding diagnostic surgery) 
8. Medications: medications, including over the counter (OTC) medications, herbal remedies and traditional 
Chinese remedies for 2 weeks before the start of the study. Hormonal contraceptives for women will be 
allowed 
9. Investigative drug studies: participation in the evaluation of any investigational product for 30 calendar days 
before this study 
10. Blood donation: blood donations for 90 calendar days before this study 
11. Drug, alcohol, caffeine, tobacco: history of drug, alcohol [>1 drink/day for females and >2 drinks/day for 
males, defined according to the USDA Dietary Guidelines 2015-2020] caffeine (>5 cups coffee/tea/day) or 
tobacco abuse (≥10 cigarettes/day) 
12. Abuse drug test: positive urine abuse drug test at screening or day -1 
13. Alcohol test: positive alcohol breath test at day -1 
14. Diet: abnormal diets (<1600 or >3500 kcal/day) or substantial changes in eating habits in the 4 weeks before 
this study; vegetarians; consumption of alcohol, grapefruit, products containing grapefruit, or beverages 
containing xanthines (coffee, tea, soda, coffee with milk, energy drinks) within 48 hours prior to the 
enrolment  
15. Pregnancy (females only): positive or missing pregnancy test at screening or day -1, pregnant or lactating 
women 
16. Vaccination within 4 weeks of study treatment 
17. Other unspecified reasons that, in the opinion of the investigator, make the subject unsuitable for enrolment.
 
Schedule:
 
Day 
Procedures/Assessments 
Notes 
Screening – Visit 1 
From day -14 to 
day -2 
 Explanation to the subject of study aims, procedures 
and possible risks 
 Informed consent signature 
 Screening number (as 
, etc.) 
 Demographic data and life style recording 
 Medical/surgical history 
 Previous/concomitant medications 
 Full physical examination (body weight, height, vital 
signs, physical abnormalities) 
 ECG recording 
 Clinical laboratory analyses: haematology, blood 
chemistry, urinalysis, serum virology, bacteriology 
and pregnancy test (women) 
 Urine multi-drug kit test 
 Adverse event (AE) monitoring 
 Inclusion/exclusion criteria evaluation 
 Eligibility evaluation 
 
The subject screening 
numbers will be in 
running 
order 
(e.g. 
, etc.). 
 
The 
subject 
study 
numbers (those who 
meet the eligibility at 
day-1) 
will 
be 
in 
running order e.g. 
 
, 
etc. 
to 
be 
identified 
as 
the 
subjects. 
CCI
CCI
CCI


CONFIDENTIAL 
 
 
Sponsor code Z7244J01 
NAC 300 mg/3 mL i.v. bioavailability 
Final version 1.0, 30AUG2018 
 
Page 7 
STUDY SYNOPSIS (cont.) 
 
Schedule (continued):
 
Day 
Procedures/Assessments 
Notes 
Visit 2 
Day -1 
 Alcohol breath test 
 Urine multi-drug kit test 
 Urine pregnancy test (women) 
 Inclusion/exclusion criteria evaluation 
 Eligibility evaluation 
 Enrolment 
 Subject study number (e.g. 
, etc.) 
 Check of AEs and concomitant medications  
 
Arrival 
at 
the 
clinic 
before 
the 
evening 
(before 18:30)  
Confinement until the 
morning of day 8 
Standardised dinner 
Fasting for at least 10 h 
(overnight) before first 
dosing
Visit 3 
Day 1  
 Vital sign measurement at pre-dose 
 IMP administration at 08:00 ± 1 h; intravenous 
infusion for 5 min 
 Vital signs measurement at 1 h post-dose 
 Blood sample collection for pharmacokinetic analysis 
at: pre-dose (0) and 5 (at the end of the infusion), 8, 
12, 15, 20, 25, 30, 60 min, 2, 4, 6, 8, 10 and 12 h post-
dose 
 Urine sample collection for pharmacokinetic analysis 
at: 0-4; 4-8; 8-12 and 12-24 h post-dose 
 Check of AEs and concomitant medications  
Standardized lunch at 
13:00  
standardized dinner at 
21:00  
 
Day 2 
 Blood sample collection for pharmacokinetic analysis 
at 24 and 32 h post-dose 
 Urine sample collection for pharmacokinetic analysis 
at 24-32 h post-dose 
 Check of AEs and concomitant medications 
Standardized breakfast 
around 9:00  
Standardized 
lunch 
around 13:00  
Standardized 
dinner 
around 21:00 
Day 3 
 Vital sign measurement at 48 h post-dose 
 Physical 
examination 
(body 
weight, 
physical 
abnormalities) at 48 h post-dose 
 Clinical laboratory analyses: haematology, blood 
chemistry, urinalysis at 48 h post-dose  
 Check of AEs and concomitant medications 
Standardized breakfast 
around 9:00  
Standardized 
lunch 
around 13:00  
Standardized 
dinner 
around 21:00
Visit 4 
From day 4 to 
day 5 
 IMP administration at 08:00 ± 1 h; infusion for 5 min 
 IMP administration at 20:00 ± 1 h; infusion for 5 min 
 Blood sample collection for pharmacokinetic analysis 
at: pre-dose (0) before the morning infusion  
 Check of AEs and concomitant medications 
Standardized breakfast 
around 9:00  
Standardized 
lunch 
around 13:00  
Standardized 
dinner 
around 21:00 
 
 
 
CCI


CONFIDENTIAL 
 
 
Sponsor code Z7244J01 
NAC 300 mg/3 mL i.v. bioavailability 
Final version 1.0, 30AUG2018 
 
Page 8 
STUDY SYNOPSIS (cont.) 
 
Schedule (continued):
 
Day 
Procedures/Assessments 
Notes 
Visit 5 
Day 6 
 Vital sign measurement at pre-dose 
 IMP administration at 08:00 ± 1 h; infusion for 5 min 
 Vital signs measurement at 1 h post-dose 
 Blood sample collection for pharmacokinetic analysis 
at: pre-dose (0) and 5 (at the end of the infusion), 8, 
12, 15, 20, 25, 30, 60 min, 2, 4, 6, 8, 10 and 12 h post-
dose 
 Urine sample collection for pharmacokinetic analysis 
at: 0-4; 4-8; 8-12 and 12-24 h post-dose 
 Check of AEs and concomitant medications  
Standardized lunch at 
13:00 (about 5 h post-
dose) 
Standardized dinner at 
21:00 (about 13 h post-
dose)  
 
Day 7 
 Blood sample collection for pharmacokinetic analysis 
at 24 and 32 h post-dose 
 Urine sample collection for pharmacokinetic analysis 
at 24-32 h post-dose 
 Check of AEs and concomitant medications 
Standardized breakfast 
around 9:00  
Standardized 
lunch 
around 13:00  
Standardized 
dinner 
around 21:00 
Final Visit/ETV 
Day 8 or early 
termination visit 
(ETV) in case of 
discontinuation 
 Vital sign measurement at 48 h post-dose (or at ETV) 
 Physical 
examination 
(body 
weight, 
physical 
abnormalities) 
 Clinical laboratory analyses: haematology, blood 
chemistry, urinalysis  
 Check of AEs and concomitant medications 
 
In case of clinically significant results at the final visit, the 
subjects will be followed-up by the investigator until the 
normalisation of the concerned clinical parameter(s) 
 
Discharge 
from 
the 
clinical centre in the 
morning of Day 8 after 
vital sign measurement, 
blood 
sampling 
for 
clinical laboratory assays 
and physical examination 
or in case of ETV. Upon 
leaving, the subjects will 
be instructed to contact 
immediately 
the 
investigator in case of 
occurrence 
of 
any 
untoward 
medical 
occurrence
Life style and constraints: 
During the study, the subjects will be confined from the evening preceding the first drug administration (study 
day -1) until the morning of day 8 (after the final visit). 
The subjects will not take any food or drinks (except water) for at least 10 h (i.e. overnight) before the first drug 
administration (Day 1) and will remain fasted up to 5 h post-dose.  
During the study, water will be allowed as desired. Coffee, tea or food containing xanthines (i.e. coffee, tea, 
soda, coffee with milk, energy drinks coke, chocolate, etc.), alcohol and grapefruit will be forbidden starting 48 
h prior to the enrolment until the end of the study. Smoking is not allowed for the whole study duration. 
 
 
 
 


CONFIDENTIAL 
 
 
Sponsor code Z7244J01 
NAC 300 mg/3 mL i.v. bioavailability 
Final version 1.0, 30AUG2018 
 
Page 9 
STUDY SYNOPSIS (cont.) 
 
Withdrawal of subjects: 
It will be documented whether or not each subject completes the clinical study. In case of premature 
discontinuation of any subject, the primary reason for discontinuation will be recorded. 
 Adverse event: Any (significant) adverse event that in the opinion of the investigator or concerned subject is 
not compatible with study continuation 
 death: the absence of life or state of being dead 
 lost to follow-up: the loss or lack of continuation of a subject up to follow-up 
 non-compliance with study drug: an indication that a subject has not agreed with or followed the instructions 
related to the study medication 
 physician decision: a position, opinion or judgment reached after consideration by a physician with 
reference to the subject 
 pregnancy: pregnancy is the state or condition of having a developing embryo or fetus in the body (uterus), 
after union of an ovum and spermatozoon, during the period from conception to birth 
 protocol deviation: an event or decision that stands in contrast to the guidelines set out by the protocol 
 study terminated by sponsor: an indication that a clinical study was stopped by its sponsor 
 technical problems: a problem with some technical aspect of a clinical study, usually related to an 
instrument 
 withdrawal by subject: study discontinuation requested by a subject for whatever reason 
For any subject discontinuing the study, the investigator will ask the subject to undergo, as far as possible, a final 
medical visit (ETV) to examine the subject's health conditions and perform the required blood sampling for the 
laboratory assays. This examination will verify that all values tested at screening have remained within a 
clinically acceptable range (i.e. not clinically significant changes compared to screening) and report in the case 
report form (CRF) date and time of the last dose administration, and date and primary reason of study 
discontinuation.
Data analysis:  
The data documented in this trial and the measured clinical parameters will be presented using classic descriptive 
statistics (i.e. total number of subjects treated [N], number of observations [n], mean standard deviation [SD], 
minimum [Min], median, maximum [Max]) for quantitative variables and frequencies (i.e. count and 
percentages) for qualitative variables if not stated otherwise. 
Analysis set:  
Enrolled set: all enrolled subjects. This analysis set will be used for demographic, baseline and background 
characteristics 
Safety set: all subjects who receive at least one dose of IMP. This analysis set will be used for the safety analyses
PK set: all enrolled subjects who fulfil the study protocol requirements in terms of IMP administration and have 
evaluable pharmacokinetic data readouts for the planned analysis, with no major deviations that may affect the 
pharmacokinetic results. This analysis set will be used for the statistical analysis of the pharmacokinetic results 
Safety Assessments: 
The statistical analysis of demographic and safety data will be performed using the analysis software SAS®   . 
The toxicity grading of laboratory tests is determined based on the NCI CTCAE V4.03. All AEs, adverse drug 
reactions and serious adverse events will be summarised by system organ class, severity and relationship to study 
drug. 
Pharmacokinetics: 
The statistical analysis of PK parameters will be performed using the validated software Phoenix® WinNonlin® 
version 6.3 or higher (Certara USA, Inc., Princeton, NJ) (actual version will be stated in the final report).   
The individual plasma/urine concentration and pharmacokinetic parameters will be presented in listings and their 
descriptive statistics summarised in tables. 
 
 
 
 
 


CONFIDENTIAL 
 
 
Sponsor code Z7244J01 
NAC 300 mg/3 mL i.v. bioavailability 
Final version 1.0, 30AUG2018 
 
Page 10 
STUDY SYNOPSIS (cont.) 
Background:  
N-acetyl-L-cysteine (NAC) is the N-acetyl derivative of the naturally occurring amino acid L-cysteine.  
NAC was introduced into the Italian pharmaceutical market in 1965 and it is now manufactured and marketed by 
Zambon S.p.A. in Europe, Asia, South America, Central America, and the Middle East, under various trade 
names (Ventresca 1989). 
NAC is widely known as a mucolytic agent, with a well-established use in the treatment of respiratory tract 
disorders. However, after more than 35 years from its introduction in the clinical practice, this molecule 
continues to be the object of scientific interest due to its potential in the treatment of several conditions other 
than those related to the respiratory system. NAC can also act as a direct antioxidant agent due to the -SH group 
(Auroma 1988) and can easily penetrate into the cells where it is deacetylated to L-cysteine, thus supporting the 
biosynthesis of glutathione (Ziment 1988). 
Clinical pharmacology and pharmacokinetics 
Several pharmacokinetic (PK) studies were carried out in healthy volunteers (Borgstrom 1986, Olsson 1988, 
Burgunder 1989, De Caro 1989, Borgstrom 1990, Frascio, Crestani 2002, Rusca 2014, Rusca 2010, Rusca 2002). 
Further PK studies were carried out in patients with chronic liver damage (Jones 1997), with liver damage 
caused by paracetamol overdose (Prescott 1989), in subjects under long-term treatment for chemoprevention 
(Pendyala 1995), in patients with respiratory disorders (Rodenstein 1978) and in patients with End Stage Renal 
Diseases (Internal report 2002). 
In the clinical studies, NAC was administered orally and intravenously both as single and repeated dose. Oral 
NAC was administered in different dosage forms, such as capsules, granulate, effervescent tablets, fast 
dissolving tablets, slow-release tablets. 
Clinical trials and safety concerns 
NAC has been available to patients as a mucolytic agent for many years and has a well-established and 
satisfactory safety and tolerability profile (see IB).  
Independently of route of administration (inhalation, oral, intravenous) adverse events are reported rarely 
following NAC administration and are usually mild and transient.  
The most frequent adverse events associated with NAC oral administration are gastrointestinal (e.g., vomiting, 
diarrhoea, stomatitis, abdominal pain and nausea). Hypersensitivity reactions (e.g. anaphylactoid reactions, 
urticaria, rash, pruritus, and bronchospasm) have been reported less frequently after oral administration, while 
with the IV administration they were reported more frequently compared to gastrointestinal symptoms. Some of 
the hypersensitivity reactions were associated to higher infusion rate. Most anaphylactoid reactions can be 
managed by temporarily suspending the acetylcysteine infusion, administering appropriate supportive care. A 
cumulative analysis of all safety reports received by the Sponsor up to September 2017 for NAC IV, confirmed 
the nature and severity of the events were in line with the above profile of the product. 
Intravenous NAC has been studied in clinical trials and published case series both as mucolytic agent and as an 
antidote against intoxication by paracetamol and other toxic agents including Amanita phalloides and chlorinated 
hydrocarbons. Overall, in published studies in humans, a total of 745 patients were exposed to i.v. NAC at doses 
ranging from 500 mg/day to 980mg/kg/day for a duration of 1 to 18 days.  Details of safety results are included 
in the IB. 
In published studies, whereas i.v. NAC as mucolytic agent typically was given at doses up to 1000 mg/day, 
much higher i.v. doses of NAC as antidote were given over multiple days, maintaining a satisfactory safety 
profile. 
Locatelli and colleagues published two case series, one of 86 patients with acute intoxication from Amanita 
phalloides and the other of 35 patients with acute intoxication from chlorinated hydrocarbons. Patients received 
i.v. NAC 300 mg/kg/day for 3 to 14 days and 3 to 18 days respectively. The safety and tolerability profile were 
overall benign, with 4 out of 121 exposed patients suffering from anaphylactoid reaction or rash (due to the rapid 
administration of the bolus according to the authors), none of which caused discontinuation of treatment. No 
other AEs were reported. (Locatelli 1996).
 
Rationale 
NAC is widely known as a mucolytic agent, with a well-established use in the treatment of respiratory tract 
disorders. In the present study, the kinetic profile and the bioavailability of NAC will be investigated in Chinese 
healthy male and female volunteers all receiving the same single i.v. dose, i.e. a total dose of 600 mg of NAC, 
and the same multiple dose administrations, i.e. five i.v. doses of 600 mg of NAC administered b.i.d. for 2 days 
and once on the last day. The safety and tolerability will be monitored as well.  
The present study will be part of the clinical development plan of NAC i.v. for registration in China of this 
formulation. 


CONFIDENTIAL 
 
 
Sponsor code Z7244J01 
NAC 300 mg/3 mL i.v. bioavailability 
Final version 1.0, 30AUG2018 
 
Page 11 
3 
STUDY SCHEDULE 
 
ACTIVITIES 
Screening 
Single Dose 
Multiple Dose 
Final 
visit/ETV1 
Visit 
V1 
V2 
V3 
V4 
V5 
Day -14/-2 
Day -1 
Day 1 
Day 2 
Day 3 
Day 4 
Day 5 
Day 6 
Day 7 
Day 82 
Informed consent 
x
Demography and lifestyle 
x
Medical and surgical history  
x
Physical examination 
x
x
 
 
x 
Prior and concomitant medications 
x
x
x
x
x
x
x
x
x
x
Height 
x
Body Weight 
x
x
x
Clinical laboratory analysis 
(haematology, blood chemistry, 
urinalysis) 
x 
 
 
 
x 
 
 
 
 
x 
Virology and bacteriology  
x
Serum pregnancy test (women) 
x
Urine multi-drug kit test 
x
x
Blood pressure and heart rate 
x
x3
x
x3
x4
Alcohol breath test 
x
Urine pregnancy test (women)
x
ECG 
X
Inclusion/exclusion criteria 
X
x
Subject eligibility
X
x
Enrolment 
x
Confinement 
x
x
x
x
x
x
x
x
Discharge 
x
Investigational product administration
x5
x6
x6
x5
Blood sampling  
x7 
x7
x8
x8
x7
x7
Urine sampling  
x9 
x9
x9
x9
Standardised meals 
x10
x11
x12
x12
x12
x12
x11
x12
Adverse event monitoring13 
x
x
x
x
x
x
x
x
x
x
1. 
Early termination visit (ETV) 


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2. 
Final visit on day 8 or in case of ETV 
3. 
At pre-dose and 1 h post-dose 
4. 
The vital signs check at 48 h post-dose (day 8), will correspond to the final measurement 
5. 
At 8:00 ± 1 h: 5-min infusion 
6. 
At 8:00 ± 1 h and at 20:00 ± 1 h; 5-min infusion 
7. 
At pre-dose (0) and 5 (at the end of the infusion), 8, 12, 15, 20, 25, 30, 60 min and 2, 4, 6, 8, 10, 12, 24 and 32 h post-dose  
8. 
At pre-dose (0) 
9. 
0-4; 4-8; 8-12, 12-24 and 24-32 h post-dose  
10. Standardised dinner 
11. Standardised lunch and dinner 
12. Standardised breakfast, lunch and dinner  
13. Adverse events monitored starting at the screening visit, immediately after informed consent, up to the final visit/ETV 
 


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4 
TABLE OF CONTENTS 
 
Page 
1 
CLINICAL STUDY PROTOCOL 
1 
2 
STUDY SYNOPSIS 
4 
3 
STUDY SCHEDULE 
11 
4 
TABLE OF CONTENTS 
13 
4.1 
TABLES 
15 
5 
LIST OF ABBREVIATIONS 
16 
6 
Introduction 
18 
6.1 
Background 
18 
6.2 
Clinical pharmacology and pharmacokinetics 
18 
6.3 
Clinical trials and safety concerns 
19 
6.4 
Rationale 
20 
6.5 
Risk and benefits 
20 
7 
STUDY OBJECTIVES 
22 
7.1 
Primary end-point 
22 
7.2 
Secondary end-point 
22 
8 
CLINICAL SUPPLIES 
23 
8.1 
Treatment 
23 
8.1.1 
Description of products 
23 
8.1.1.1 
Test product 
23 
8.1.2 
Dose regimen 
24 
8.1.3 
Route and method of administration 
24 
8.1.4 
Investigational product distribution 
24 
8.2 
Packaging and labelling 
24 
8.3 
Storage conditions 
25 
8.4 
Drug accountability 
25 
9 
INVESTIGATIONAL PLAN 
26 
9.1 
Overall study design 
26 
9.2 
Discussion of design 
26 
10 
STUDY POPULATION 
27 
10.1 
Target population 
27 
10.2 
Inclusion criteria 
27 
10.3 
Exclusion criteria 
27 
10.3.1 
Not allowed treatments 
28 
11 
STUDY SCHEDULE 
30 
11.1 
Study visits and procedures 
30 
11.2 
Diet and lifestyle 
32 
11.2.1 
Restrictions 
33 
12 
DESCRIPTION OF SPECIFIC PROCEDURES 
34 
12.1 
Physical examination 
34 
12.1.1 
Body weight 
34 
12.1.2 
Vital signs 
34 
12.1.3 
ECGs 
34 
12.2 
Clinical laboratory assays 
34 
12.3 
Sampling for pharmacokinetic analysis 
35 
12.3.1 
Venous blood sampling 
35 
12.3.2 
Urine collection 
36 
12.3.3 
Analytics 
37 
12.3.4 
Labelling, storage and transport of samples 
37 
12.3.4.1 
Samples labelling 
37 
12.3.4.2 
Samples storage and transport 
37 
13 
ASSIGNMENT OF STUDY TREATMENT 
39 
13.1 
Randomisation 
39 


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13.2 
Blinding 
39 
14 
EVALUATION PARAMETERS 
40 
14.1 
Study variables 
40 
14.1.1 
Primary variables 
40 
14.1.2 
Secondary variables 
40 
14.2 
Pharmacokinetic assessments 
40 
14.2.1 
Pharmacokinetic parameters 
40 
14.3 
Safety assessments 
42 
15 
STATISTICAL METHODS 
43 
15.1 
Analysis Sets 
43 
15.1.1 
Definitions 
43 
15.1.2 
Reasons for exclusion from the PK set before bioanalysis 
44 
15.2 
Sample size and power considerations 
44 
15.3 
Demographic, baseline and background characteristics 
44 
15.4 
Analysis of pharmacokinetic parameters 
44 
15.4.1 
Descriptive pharmacokinetics 
44 
15.5 
Safety and tolerability evaluation 
44 
16 
DEFINITION AND HANDLING OF AEs AND SAEs 
46 
16.1 
Applicable SOPs 
46 
16.2 
Definition of Adverse Event (AE) 
46 
16.3 
Definition of Adverse Drug Reaction (ADR) 
46 
16.4 
Definition of Serious Adverse Events or Serious Adverse Drug Reaction 
47 
16.5 
Definition of Severity of Adverse Events 
47 
16.6 
Definition of Adverse Event causality 
47 
16.7 
Adverse Events recording 
48 
16.8 
AEs monitoring window 
48 
16.9 
Adverse Events reporting 
48 
16.9.1 
SAEs reporting 
49 
16.10 
Follow-up for Adverse Events 
49 
16.11 
SUSARs management 
49 
16.12 
Other events qualified for expedited reporting 
50 
16.13 
SAEs: contacts 
51 
16.14 
Pregnancy 
51 
17 
DATA MANAGEMENT PROCEDURES 
52 
17.1 
Data collection – CRFs 
52 
17.2 
Database management 
52 
17.2.1 
Coding dictionaries 
52 
18 
STUDY MONITORING, QUALITY CONTROL AND QUALITY ASSURANCE 
53 
18.1 
Monitoring 
53 
18.2 
Quality Control and Quality Assurance 
53 
18.3 
Applicable SOPs 
54 
18.4 
Data access 
54 
18.5 
Audits and inspections 
54 
19 
ETHICAL CONSIDERATIONS 
55 
19.1 
Ethics and Good Clinical Practice (GCP) 
55 
19.2 
Informed consent 
55 
19.3 
Insurance policy 
56 
19.4 
Withdrawal of subjects 
56 
19.4.1 
Primary reason for discontinuation 
56 
19.5 
Study termination 
57 
20 
ADMINISTRATIVE PROCEDURES 
58 
20.1 
Material supplied to the clinical centre 
58 
20.2 
Protocol amendments 
58 
20.3 
Study documentation and record keeping 
58 
20.4 
Study subjects’ recruitment 
59 
20.5 
Confidentiality and data protection 
59 
20.6 
Publication policy 
59 
21 
STUDY RESPONSIBLE PERSONS 
61 


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21.1 
Sponsor 
61 
21.2 
Drug assay 
61 
21.3 
Project management, data analysis & reporting, monitoring 
61 
22 
REFERENCES 
62 
 
 
 
4.1 
TABLES 
Page 
 
Table 11.1.1 
Schedule of study activities and procedures 
31 
Table 12.3.1.1 
Tolerance ranges for the scheduled sampling times 
36 
 
 


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5 
LIST OF ABBREVIATIONS 
 
-HCG
human chorionic gonadotropin 
-GT
-Glutamyl transpeptidase
ADR
Adverse Drug Reaction
AE
Adverse Event
ALT
Alanine aminotransferase
Ae0-t
Total amount of NAC excreted in urine 
Aess(0-t)
Total amount of NAC excreted in urine at steady-state
ANOVA
Analysis of Variance
AST
Aspartate aminotransferase
BB
Blue book
BLQL
Below Lower Quantification Limit
BUN
Blood Urea Nitrogen
AUC(0-t) 
Area under the concentration-time curve from single dose to the last observed concentration 
time t
AUC(0-)
Area under the concentration-time curve extrapolated to infinity 
AUC(0-12)
Area under the concentration-time curve at steady-state in the tau interval 
AUCss(0-t) 
Area under the concentration-time curve at steady-state from the last multiple dose to the 
last observed concentration time t
AUCss(0-12)
Area under the concentration-time curve at steady-state in the tau interval 
BMI
Body Mass Index
BP
Blood Pressure
CA
Competent Authority
CDISC
Clinical Data Interchange Standards Consortium
CFDA
China Food And Drug Administration
CI
Confidence Interval
Cmax
Peak drug concentration
CLt
Total body clearance
CLr
Renal clearance
CPL
Clinical Project Leader
CRA
Clinical Research Associate
CRF
Case Report Form
CRO
Contract Research Organisation
CS
Clinically Significant
CSP
Clinical Study Protocol
CSR
Clinical Study Report
Css_av
Average NAC plasma concentration at steady-state
Css_max
Maximum plasma concentration at steady-state
Css_min
Trough plasma concentration at steady-state
CV
Coefficient of Variation
DBP
Diastolic Blood Pressure
DF
Degree of fluctuation 
DSU
Drug safety unit
DTT
Dithiothreitol
EC
Ethics Committee
ECG
Electrocardiogram
ETV
Early Termination Visit
FDA
Food and Drug Administration
Fe0-t
Total fraction of NAC dose excreted in urine 
Frel
Relative Bioavailability
FSFV
First Subject First Visit
GCP
Good Clinical Practice
GLP
Good Laboratory Practice
HBs Ag
Hepatitis B virus surface antigen
HCV Ab
Hepatitis C virus antibodies
HIV
Human Immunodeficiency Virus


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HR
Heart Rate
IB
Investigator’s Brochure
ICH
International Conference on Harmonisation
IRB/IEC
Institutional Review Board/Independent Ethics Committee
i.m.
Intramuscular
IMP
Investigational Medicinal Product
IUD
Intra-Uterine Device
i.v.
Intravenous
kel
Terminal elimination rate constant
LQL
Lower Quantification Limit
LSLV
Last Subject Last Visit
MCH
Mean Cell Haemoglobin
MCHC
Mean Cell Haemoglobin Concentration
MCV
Mean Cell Volume
MedDRA
Medical Dictionary for Regulatory Activities
MRT
Mean Residence Time
MW
Molecular Weight
N
Normal
NA
Not Applicable
NAC
N-acetyl-L-cysteine
NC
Not calculated
NCS
Not clinically significant
NSAIDs
Non-Steroidal Anti-Inflammatory Drugs
OTC
Over The Counter
PE
Point Estimate
PK
Pharmacokinetics
PT
Preferred Term
PTAE
Pre-Treatment Adverse Event
R
Accumulation ratio
RBC 
Red Blood Cells
RSI
Reference safety information
SAE
Serious Adverse Event
SBP
Systolic Blood Pressure
SD
Standard Deviation
SOC
System Organ Class
SOP
Standard Operating Procedure
SDTM
Study Data Tabulation Model
SUSAR
Suspected Unexpected Serious Adverse Reaction
T
Test
TEAE
Treatment-Emergent Adverse Event
THC
delta-9-tetrahydrocannabinol
t1/2
Half-life
tmax
Time to achieve Cmax
tss_max
Time to achieve Css_max
USDA
United States Department of Agriculture
Vd
Volume of distribution 
WBC
White Blood Cells
WHODDE
World Health Organisation Drug Dictionary Enhanced
 
 
 
 


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6 
INTRODUCTION 
 
6.1 
Background 
 
N-acetyl-L-cysteine (NAC) is the N-acetyl derivative of the naturally occurring amino acid L-
cysteine.  
 
NAC was introduced into the Italian pharmaceutical market in 1965 and it is now 
manufactured and marketed by Zambon S.p.A. in Europe, Asia, South America, Central 
America, and the Middle East, under various trade names (1). 
 
NAC is widely known as a mucolytic agent, with a well-established use in the treatment of 
respiratory tract disorders. However, after more than 35 years from its introduction in the 
clinical practice, this molecule continues to be the object of scientific interest due to its 
potential in the treatment of several conditions other than those related to the respiratory 
system. NAC can also act as a direct antioxidant agent due to the -SH group (2) and can easily 
penetrate into the cells where it is deacetylated to L-cysteine, thus supporting the biosynthesis 
of glutathione (3). 
 
6.2 
Clinical pharmacology and pharmacokinetics 
 
Several pharmacokinetic (PK) studies were carried out in healthy volunteers (4, 5, 6, 7, 8, 9, 
10, 11, 12, 13, 14). Further PK studies were carried out in patients with chronic liver damage 
(15), with liver damage caused by paracetamol overdosage (16), in subjects under long-term 
treatment for chemoprevention (17), in patients with respiratory disorders (18) and in patients 
with End Stage Renal Diseases (19). 
In the clinical studies, NAC was administered orally and intravenously both as single and 
repeated doses. Oral NAC was administered in different dosage forms, such as capsules, 
granulate, effervescent tablets, fast dissolving tablets, slow-release tablets. 
 
In particular, Borgström et al. (4) studied for the first time the PK profile of NAC after i.v. 
dose of 600 mg of NAC infused over 5 min in 10 healthy male and female volunteers in 
comparison with 3 other oral dosage forms. The PK parameters obtained by the authors after 
i.v. infusion are summarised in the following table.  
 
Table 6.2.1 
PK parameters of plasma free NAC (N=10) 
Cmax 
(µmol/L) 
tmax 
(h) 
CL 
(L/kgxh) 
CLr 
(L/kgxh) 
t½ 
(h) 
Ae0-12 
(% of dose) 
16.0±7.9 
0.65±0.33 
0.207±0.017 
0.058±0.011 
2.27±0.32 
29.0±3.2* 
mean±SD is reported; *: N=9; Source: 4 
 
However, the authors analysed NAC in deproteinised plasma thus missing the protein-bound 
NAC in their measurements.  
 
Olsson et al. (5) improved the bioanalytical method and were able to measure the total NAC 
by reduction of the disulphide bonds in plasma before precipitating proteins. After single i.v. 


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dose of 200 mg of NAC to 6 healthy volunteers, the authors found the following PK 
parameters: 
 
Table 6.2.2 
PK parameters of plasma total NAC (N=6) 
 
Cmax 
(µM) 
Vss 
(L/kg) 
CL 
(L/kgxh) 
T 
121 (82.9 – 162) 
0.47 (0.46 – 0.55) 
0.11 (0.09 – 1.13) 
median (range) is reported; Source: 5 
 
 
Jones et al. (15) studied the bioavailability of NAC after single i.v. dose of 600 mg of NAC 
infused over 3 min to patients with chronic liver disease, but also to 6 healthy male and 
female volunteers as control subjects. The authors found the following PK parameters for 
total NAC: 
 
Table 6.2.3 
PK parameters of plasma total NAC (N=6) 
AUC 
(mg/Lxh) 
CLr 
(L/h) 
t½ 
(h) 
Vdss 
(L) 
93.9±9.6 
6.5±0.8 
2.6±0.3 
17.4±2.8 
mean±SD is reported; Source: 15 
 
Brown et al. (14) investigated the PK of NAC infused i.v. to 24 healthy men at rest and 
during exercise. NAC was infused at the dose of 125 mg/kgxh for 15 min followed by 25 
mg/kgxh for 35 min to healthy men at rest. Then, the infusion continued during exercise until 
fatigue. The authors found NAC peak concentrations in plasma at the end of the initial 
loading infusion. Mean peak concentration of total NAC was 205.1±68.3 mg/L, while mean 
CL was 0.164 L/kgxh.  
 
It is worth noting that, while scientific literature shows that PK profiles of NAC after oral 
administration are very consistent (in terms of half-life, rate and extent of exposure, 
bioavailability, dose-linearity, etc.) among various publications, this is not the case for the 
data obtained after i.v. dosing. In fact, for this administration route, published studies differ in 
several key aspects, i.e., for example, type of administration, applied bioanalytical methods, 
drug doses, calculation methods for PK parameters. Furthermore the number of subjects 
enrolled in each published study is very small. For these reasons, the comparability of the 
literature data is low among the published studies and potentially also to the expected results 
of the present study.   
 
6.3 
Clinical trials and safety concerns 
 
NAC has been available to patients as a mucolytic agent for many years and has a well-
established and satisfactory safety and tolerability profile (see IB).  
Independently of route of administration (inhalation, oral, intravenous) adverse events are 
reported rarely following NAC administration and are usually mild and transient.  
The most frequent adverse events associated with NAC oral administration are 
gastrointestinal (e.g., vomiting, diarrhoea, stomatitis, abdominal pain and nausea). 
Hypersensitivity reactions (e.g. anaphylactoid reactions, urticaria, rash, pruritus, and 


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Sponsor code Z7244J01 
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bronchospasm) have been reported less frequently after oral administration, while with the i.v. 
administration they were reported more frequently compared to gastrointestinal symptoms. 
Some of the hypersensitivity reactions were associated to higher infusion rate. Most 
anaphylactoid reactions can be managed by temporarily suspending the acetylcysteine 
infusion, administering appropriate supportive care. A cumulative analysis of all safety 
reports received by the Sponsor up to September 2017 for NAC IV, confirmed the nature and 
severity of the events were in line with the above profile of the product. 
Intravenous NAC has been studied in clinical trials and published case series both as 
mucolytic agent and as an antidote against intoxication by paracetamol and other toxic agents 
including Amanita phalloides and chlorinated hydrocarbons. Overall, in published studies in 
humans, a total of 745 patients were exposed to i.v. NAC at doses ranging from 500 mg/day 
to 980mg/kg/day for a duration of 1 to 18 days.  Details of safety results are included in the 
IB. 
In published studies, whereas i.v. NAC as mucolytic agent typically was given at doses up to 
1000 mg/day, much higher i.v. doses of NAC as antidote were given over multiple days, 
maintaining a satisfactory safety profile. 
Locatelli and colleagues published two case series, one of 86 patients with acute intoxication 
from Amanita phalloides and the other of 35 patients with acute intoxication from chlorinated 
hydrocarbons. Patients received i.v. NAC 300 mg/kg/day for 3 to 14 days and 3 to 18 days 
respectively. The safety and tolerability profile were overall benign, with 4 out of 121 
exposed patients suffering from anaphylactoid reaction or rash (due to the rapid 
administration of the bolus according to the authors), none of which caused discontinuation of 
treatment. No other AEs were reported (21). 
 
 
6.4 
Rationale 
 
NAC is widely known as a mucolytic agent, with a well-established use in the treatment of 
respiratory tract disorders. 
In the present study, the kinetic profile and the bioavailability of NAC will be investigated in 
Chinese healthy male and female volunteers all receiving the same single i.v. dose, i.e. a total 
dose of 600 mg of NAC, and the same multiple dose administrations, i.e. five i.v. doses of 
600 mg of NAC administered b.i.d. for 2 days and once on the last day. The safety and 
tolerability will be monitored as well.  
The present study will be part of the clinical development plan of NAC i.v.  for registration in 
China of this formulation. 
 
6.5 
Risk and benefits 
 
NAC is a well-known drug which has been used for decades.  
 
Undesired effects which may occur during treatment with NAC include: anaphylactic shock, 
anaphylactic reaction, anaphylactoid reaction, hypersensitivity, tachycardia, bronchospasm, 
dyspnoea, vomiting, nausea, angioedema, urticaria, flushing rash, pruritus, face oedema, 
blood pressure decreased, prothrombin time prolonged (for details refer to IB; 22).  
 
Blood sampling with cannula insertion may cause minor discomfort. The risks associated with 
blood draws include pain, bleeding and bruising. 


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No specific benefits for the participants in the current study are foreseen. Their remuneration 
will be paid after study completion. The remuneration covers loss of time and any 
inconvenience caused by the participation in the study. 


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7 
STUDY OBJECTIVES 
 
The objective of the study is to evaluate the NAC pharmacokinetics, safety and tolerability 
after single and multiple dose i.v. administration. 
 
7.1 
Primary end-point 
 
 To evaluate pharmacokinetic parameters of NAC in plasma after single and multiple dose 
administration of the investigational product.  
 
7.2 
Secondary end-point 
 
 To collect safety and tolerability data after single and multiple dose administration of the 
investigational product. 


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8 
CLINICAL SUPPLIES 
 
8.1 
Treatment 
 
All the subjects enrolled in the study will receive the same treatment with the investigational 
medicinal product (IMP), i.e. NAC, 300 mg/ 3 mL solution for injection, as follows: 
 
 on day 1 at 08:00 ±1 h, one dose of 600 mg of NAC (300 + 300 mg ampoule) will be 
administered under fasting conditions; 
 
After a wash-out of 3 days: 
 
 on days 4 and 5 at 08:00 ±1 h and 20:00 ±1 h and at 08:00 ±1 on day 6, 5 doses of 600 mg 
of NAC (300 + 300 mg ampoule) will be administered. 
 
8.1.1 
Description of products 
 
8.1.1.1 
Test product 
 
IMP
NAC 300 mg/ 3 mL solution for injection
Active substance
N-acetyl-L-cysteine
Manufacturer 
(active substance) 
F.I.S. Fabbrica Italiana Sintetici S.p.A.,  
Via Dovaro, 36045 Lonigo (Vicenza), Italy 
(GMP compliant)
Manufacturer 
(finished product) 
Zambon S.p.A.,  
Via della Chimica 9, 36100 Vicenza, Italy 
(GMP compliant)
Pharmaceutical form
Solution for injection
Dose
300 mg/ 3 mL
Administration route
Parenteral 
 
The analytical certificates will be supplied with the IMP. Quali-quantitative formulation is as 
follows: 
 
Each 3 mL vial (10%) contains: 
 N-acetyl-L-cysteine (NAC) 300 mg 
 sodium hydroxide 74 mg 
 disodium edetate 3 mg 
 water for injections q.s. to 3 mL 
 
 
 


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8.1.2 
Dose regimen 
 
Two ampoules of IMP (300 + 300 mg) corresponding to a total dose of 600 mg of NAC 
diluted in 10 mL of NaCl 0.9% sterile saline solution, will be administered by a 5-minute i.v. 
infusion. 
 
Single dose: 
 
One (1) dose of IMP will be administered under fasting conditions on day 1 at 08:00 ±1 h. 
 
Multiple dose regime: 
 
Five (5) doses of IMP will be administered twice a day (b.i.d.) on days 4 and 5 at 08:00 ±1 h 
and 20:00 ±1 h and one dose will be administered on day 6 at 08:00 ±1.  
 
8.1.3 
Route and method of administration 
 
Each dose of IMP will be prepared by diluting the content of 2 ampoules of IMP in 10 mL of 
saline, as described above (§ 8.1.2), and administered to each study subject at the clinical site 
only by the investigator or his/her deputy. 
 
Each dose will be infused intravenously over 5 min.  
 
The start of infusion (not the end) will be considered as time 0. 
 
8.1.4 
Investigational product distribution 
 
The IMP will be exclusively used for the present clinical study and will only be administered 
to the subjects enrolled in the study.  
 
8.2 
Packaging and labelling 
 
The IMP primary packaging will be glass vials. 
 
The formulation labelling will report all the information requested according to the Annex 13 
to the Good Manufacturing Practice (published by the Commission in The rules governing 
medicinal products in the European Community, Volume 4; 23) and in compliance with 
applicable laws and regulations as follows: 
a. 
Name, address and telephone number of the sponsor, contract research organisation or 
investigator (the main contact for information on the product, clinical study and any 
emergency ) 
b. Pharmaceutical dosage form, route of administration, quantity of dosage units, and the 
name and strength 
c. 
The batch and/or code number to identify the contents and packaging operation 
d. A study reference code allowing identification of the study, site, investigator and sponsor 
if not given elsewhere 


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e. 
The study subject identification number/treatment number and where relevant, the visit 
number 
f. 
The name of the investigator (if not included in (a) or (d)) 
g. Directions for use (reference may be made to a leaflet or other explanatory document 
intended for the study subject or person administering the product) 
h. “For clinical study use only” or similar wording 
i. 
The storage conditions 
j. 
Period of use (use-by date, expiry date or re-test date as applicable), in month/year format 
and in a manner that avoids any ambiguity 
k. “Keep out of reach of children” 
 
Labels will be in local language. 
 
8.3 
Storage conditions 
 
The IMP will be stored at ≤25° C in a dry locked place, sheltered from light.  
 
8.4 
Drug accountability 
 
The IMP will be provided directly to the investigator by the Sponsor or designee, in excess of 
the amount necessary for the study (at least 25% excess). 
 
After receipt of the IMP supply, the pharmacist will confirm in writing by signing and dating 
standard drug accountability forms. 
 
At the end of the study, used, unused and partially used supplies of IMP provided by the 
Sponsor or  designee will either be destroyed on site (upon written authorisation) or returned 
to the Sponsor or  designee, after assessment of drug accountability. 
 
 


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NAC 300 mg/3 mL i.v. bioavailability 
Final version 1.0, 30AUG2018 
 
Page 26 
9 
INVESTIGATIONAL PLAN 
 
9.1 
Overall study design 
 
This is a single and multiple dose, single centre, open-label, one-way, pharmacokinetics, 
safety and tolerability clinical trial of Phase I to be performed in Chinese healthy male and 
female volunteers. 
 
9.2 
Discussion of design 
 
The study has been designed in agreement with the Chinese Technical Guideline on Clinical 
Pharmacokinetic Research of Chemical Drugs, 18 March 2005 and the European Guideline on 
the Investigation of Bioequivalence (CPMP/QWP/EWP/1401/98 Rev. 1, 20 January 2010) 
(24).  
 
An open-label design is used since the primary end-point of the study is based on objective 
measurements of NAC in blood. The outcome variables are not influenced by the subjects or 
investigator being aware of the administered products. 
 
Blood sampling time-points were selected on the basis of the known PK profile of NAC (4, 5, 
6, 7, 8, 9, 10, 11, 12, 13, 14). 
 
The dose of 600 mg has been selected since this is the efficacious, safe and well tolerated 
dose used in clinical practice (see also 22).  
 
The bioanalysis will be performed in compliance with GCP and CFDA GCP regulations and 
in accordance with the applicable principles of GLP, as defined by OECD, in a GLP 
compliant facility. Moreover, sample analysis will be conducted in compliance with the 
Chinese Guidance on Bioanalysis: method validation and analysis of study samples (2015).  
 


CONFIDENTIAL 
 
 
Sponsor code Z7244J01 
NAC 300 mg/3 mL i.v. bioavailability 
Final version 1.0, 30AUG2018 
 
Page 27 
10 
STUDY POPULATION 
 
10.1 
Target population 
 
Chinese healthy male and female volunteers will be included in the study.  
 
10.2 
Inclusion criteria 
 
To be enrolled in this study, subjects must fulfil all these criteria: 
 
1. Informed consent: signed written informed consent before inclusion in the study  
2. Ethnicity, Sex and Age: Chinese males and females, 18-45 year old inclusive 
3. Weight: body weight ≥50 kg; 
4. Body Mass Index: 19-24 kg/m2 inclusive 
5. Vital signs: systolic blood pressure 100-139 mmHg, diastolic blood pressure 50-89 
mmHg, heart rate 50-90 bpm, measured after 5 min at rest in the sitting/supine position (to 
be chosen according to the usual procedure at the clinical site) 
6. Full comprehension: ability to comprehend the full nature and purpose of the study, 
including possible risks and side effects; ability to co-operate with the investigator and to 
comply with the requirements of the entire study 
7. Nicotine addiction (smoker subjects only): ability to abstain from smoking for the duration 
of the clinical study 
8. Contraception and fertility (women only): women of child-bearing potential must be using 
at least one of the following reliable methods of contraception: 
a. Hormonal oral, implantable, transdermal, or injectable contraceptives for at least 60 
calendar days before the screening visit 
b. A non-hormonal intrauterine device or female condom with spermicide or 
contraceptive sponge with spermicide or diaphragm with spermicide or cervical cap 
with spermicide for at least 60 calendar days before the screening visit 
c. A male sexual partner who agrees to use a male condom with spermicide 
d. A sterile sexual partner 
Women of non-child-bearing potential or in post-menopausal status for at least 1 year will 
be admitted. 
Women of childbearing potential should be willing to adopt abstinence or contraception 
measures during the study and two weeks post-dose. 
For all women, pregnancy test result must be negative at screening and day -1. 
 
10.3 
Exclusion criteria 
 
Subjects meeting any of these criteria will not be enrolled in the study: 
 


CONFIDENTIAL 
 
 
Sponsor code Z7244J01 
NAC 300 mg/3 mL i.v. bioavailability 
Final version 1.0, 30AUG2018 
 
Page 28 
1. Electrocardiogram (12-lead ECG in supine position): clinically significant abnormalities 
2. Physical findings: clinically significant abnormal physical findings which could interfere 
with the objectives of the study 
3. Laboratory analyses: clinically significant abnormal laboratory values indicative of 
physical illness, in particular significant laboratory abnormality indicative of hepatic 
condition (more than 3 times the upper limit) 
4. Allergy: ascertained or presumptive hypersensitivity to the active principle and/or 
formulations' ingredients; history of anaphylaxis to drugs or allergic reactions in general, 
which the investigator considers may affect the outcome of the study 
5. Diseases: significant history of renal, hepatic, gastrointestinal, cardiovascular, respiratory, 
skin, haematological, endocrine, urologic, metabolic, neurological or psychiatric diseases, 
as determined by the investigator, that may interfere with the aim of the study; history of 
carcinoma in situ and malignant disease; active bacterial or viral infection and fever >38° 
C within 48 h prior to study treatment administration 
6. Virology: positive result of HIV, hepatitis B (HBV), hepatitis C (HCV) or Treponema 
pallidum (TP) assays 
7. Surgery: any surgery within 60 calendar days of screening (excluding diagnostic surgery) 
8. Medications: medications, including over the counter (OTC) medications, herbal remedies 
and traditional Chinese remedies for 2 weeks before the start of the study. Hormonal 
contraceptives for women will be allowed 
9. Investigative drug studies: participation in the evaluation of any investigational product 
for 1 month before this study 
10. Blood donation: blood donations for 90 calendar days before this study 
11. Drug, alcohol, caffeine, tobacco: history of drug, alcohol [>1 drink/day for females and 
>2 drinks/day for males, defined according to the USDA Dietary Guidelines 2015-2020] 
caffeine (>5 cups coffee/tea/day) or tobacco abuse (≥10 cigarettes/day) 
12. Abuse drug test: positive urine abuse drug test at screening or day -1 
13. Alcohol test: positive alcohol breath test at day -1 
14. Diet: abnormal diets (<1600 or >3500 kcal/day) or substantial changes in eating habits in 
the 4 weeks before this study; vegetarians; consumption of alcohol, grapefruit, products 
containing grapefruit, or beverages containing xanthines (coffee, tea, soda, coffee with 
milk, energy drinks) within 48 hours prior to the enrolment  
15. Pregnancy (females only): positive or missing pregnancy test at screening or day -1, 
pregnant or lactating women 
16. Vaccination within 4 weeks of study treatment 
17. Other unspecified reasons that, in the opinion of the investigator, make the subject 
unsuitable for enrolment. 
 
10.3.1 
Not allowed treatments 
 


CONFIDENTIAL 
 
 
Sponsor code Z7244J01 
NAC 300 mg/3 mL i.v. bioavailability 
Final version 1.0, 30AUG2018 
 
Page 29 
No medication, including OTC, traditional Chinese medicine and herbal remedies, will be 
allowed for 2 weeks before the start of the study and during the whole study duration. 
Paracetamol will be allowed as therapeutic counter-measure for adverse events (AEs) 
according to the investigator’s opinion. Hormonal contraceptives will be allowed.  
 
The intake of any other medication will be reported as a protocol deviation.  
 
 


CONFIDENTIAL 
 
 
Sponsor code Z7244J01 
NAC 300 mg/3 mL i.v. bioavailability 
Final version 1.0, 30AUG2018 
 
Page 30 
11 
STUDY SCHEDULE 
 
The schedule of the study is summarised at page 11. 
 
11.1 
Study visits and procedures 
 
Each study subject will undergo 6 visits. 
 
The study protocol foresees a screening visit and a confinement lasting from Visit 2 to the 
Final Visit/early termination visit (ETV), i.e. from Day -1 to Day 8. Maximum study duration 
will be 22 days, screening visit included. A written informed consent will be obtained before 
any study assessment or procedure. 
 
The first subject first visit (FSFV) is defined as the 1st visit performed at the clinical centre on 
the 1st screened subject. The last subject last visit (LSLV) is defined as the last visit performed 
at the clinical centre (or the telephonic follow-up, if applicable) on the last subject, i.e. the last 
visit foreseen by the study protocol, independently of the fact that the subject is a completer or 
a withdrawn subject. 
 
The following phases, visits and procedures will be performed: 
 
 Screening phase 
 Screening – visit 1: between day -14 and day -2 
 Visit 2: day -1 
 
 Interventional phase 
 Visit 3: days 1-3 
 Visit 4: days 4-5 
 Visit 5: days 6-7 
 
 Final phase 
 Visit 6: day 8 - Final visit/ETV. In case of early discontinuation, discontinued subjects 
will undergo an ETV 
 
Activities to be performed are listed by visit in the following table:  


CONFIDENTIAL 
 
 
Sponsor code Z7244J01 
NAC 300 mg/3 mL i.v. bioavailability 
Final version 1.0, 30AUG2018 
 
Page 31 
Table 11.1.1 Schedule of study activities and procedures 
 
Day 
Procedures/Assessments 
Notes 
Screening – Visit 1 
From day -14 to 
day -2 
 Explanation to the subject of study aims, procedures 
and possible risks 
 Informed consent signature 
 Screening number (as 
, etc.) 
 Demographic data and life style recording 
 Medical/surgical history 
 Previous/concomitant medications 
 Full physical examination (body weight, height, vital 
signs, physical abnormalities) 
 ECG recording 
 Clinical laboratory analyses: haematology, blood 
chemistry, urinalysis, serum virology, bacteriology 
and pregnancy test (women) 
 Urine multi-drug kit test 
 AE monitoring 
 Inclusion/exclusion criteria evaluation 
 Eligibility evaluation 
 
The subject screening 
numbers (e.g. 
 
, etc.) and the 
subject study numbers 
(e.g. 
, etc.) 
separated by a slash, 
will be used to identify 
the subjects 
Visit 2 
Day -1 
 Alcohol breath test 
 Urine multi-drug kit test 
 Urine pregnancy test (women) 
 Inclusion/exclusion criteria evaluation 
 Eligibility evaluation 
 Enrolment 
 Study subject number (e.g. 
, etc.) 
 Check of AEs and concomitant medications  
 
Arrival 
at 
the 
clinic 
before 
the 
evening 
(before 18:30)  
Confinement until the 
morning of day 8 
Standardised dinner 
Fasting for at least 10 h 
(overnight) before first 
dosing 
Visit 3 
Day 1  
 Vital sign measurement at pre-dose 
 IMP administration at 08:00 ± 1 h; intravenous 
infusion for 5 min 
 Vital signs measurement at 1 h post-dose 
 Blood sample collection for pharmacokinetic analysis 
at: pre-dose (0) and 5 (at the end of the infusion), 8, 
12, 15, 20, 25, 30, 60 min, 2, 4, 6, 8, 10 and 12 h 
post-dose 
 Urine sample collection for pharmacokinetic analysis 
at: 0-4; 4-8; 8-12 and 12-24 h post-dose 
 Check of AEs and concomitant medications  
Standardized lunch at 
13:00  
standardized dinner at 
21:00  
 
Day 2 
 Blood sample collection for pharmacokinetic analysis 
at 24 and 32 h post-dose 
 Urine sample collection for pharmacokinetic analysis 
at 24-32 h post-dose 
 Check of AEs and concomitant medications 
Standardized breakfast 
around 9:00  
Standardized 
lunch 
around 13:00  
Standardized 
dinner 
around 21:00  
Day 3 
 Vital sign measurement at 48 h post-dose 
 Physical 
examination 
(body 
weight, 
physical 
abnormalities) at 48 h post-dose 
 Clinical laboratory analyses: haematology, blood 
chemistry, urinalysis at 48 h post-dose  
 Check of AEs and concomitant medications 
 
Standardized breakfast 
around 9:00  
Standardized 
lunch 
around 13:00  
Standardized 
dinner 
around 21:00 
CCI
CCI
CCI
CCI


CONFIDENTIAL 
 
 
Sponsor code Z7244J01 
NAC 300 mg/3 mL i.v. bioavailability 
Final version 1.0, 30AUG2018 
 
Page 32 
 
Day 
Procedures/Assessments 
Notes 
Visit 4 
From day 4 to 
day 5 
 IMP administration at 08:00 ± 1 h; infusion for 5 min 
 IMP administration at 20:00 ± 1 h; infusion for 5 min 
 Blood sample collection for pharmacokinetic analysis 
at: pre-dose (0) before the morning infusion  
 Check of AEs and concomitant medications 
Standardized breakfast 
around 9:00  
Standardized 
lunch 
around 13:00  
Standardized 
dinner 
around 21:00  
 
Visit 5 
Day 6 
 Vital sign measurement at pre-dose 
 IMP administration at 08:00 ± 1 h; infusion for 5 min 
 Vital signs measurement at 1 h post-dose 
 Blood sample collection for pharmacokinetic analysis 
at: pre-dose (0) and 5 (at the end of the infusion), 8, 
12, 15, 20, 25, 30, 60 min, 2, 4, 6, 8, 10 and 12 h post-
dose 
 Urine sample collection for pharmacokinetic analysis 
at: 0-4; 4-8; 8-12 and 12-24 h post-dose 
 Check of AEs and concomitant medications  
Standardized lunch at 
13:00 (about 5 h post-
dose) 
Standardized dinner at 
21:00 (about 13 h post-
dose)  
 
Day 7 
 Blood sample collection for pharmacokinetic analysis 
at 24 and 32 h post-dose 
 Urine sample collection for pharmacokinetic analysis 
at 24-32 h post-dose 
 Check of AEs and concomitant medications 
Standardized breakfast 
around 9:00  
Standardized 
lunch 
around 13:00  
Standardized 
dinner 
around 21:00  
Final Visit/ETV 
Day 8 or early 
termination visit 
(ETV) in case of 
discontinuation 
 Vital sign measurement at 48 h post-dose (or at ETV) 
 Physical 
examination 
(body 
weight, 
physical 
abnormalities) 
 Clinical laboratory analyses: haematology, blood 
chemistry, urinalysis  
 Check of AEs and concomitant medications 
 
In case of clinically significant results at the final visit, the 
subjects will be followed-up by the investigator until the 
normalisation of the concerned clinical parameter(s) 
 
Discharge 
from 
the 
clinical centre in the 
morning of Day 8 after 
vital sign measurement, 
blood 
sampling 
for 
clinical 
laboratory 
assays 
and 
physical 
examination or in case 
of ETV. Upon leaving, 
the subjects will be 
instructed to contact 
immediately 
the 
investigator in case of 
occurrence 
of 
any 
untoward 
medical 
occurrence 
 
11.2 
Diet and lifestyle 
 
The subjects will not take any food or drinks (except water) for at least 10 h (i.e. overnight) 
before the first drug administration (Day 1) and will remain fasted up to 5 h post-dose.  
During the study, water will be allowed as desired. In order to maintain an adequate 
hydration, the subjects will be encouraged to drink at least 120 mL of mineral water every 2 h 
for 6 h post-dose on Days 1 and 6. 
 
Coffee, tea or food containing xanthines (i.e. coffee, tea, soda, coffee with milk, energy 
drinks, etc.), alcohol and grapefruit will be forbidden starting 48 h before the enrolment until 
the end of the study. Smoking is not allowed for the whole study duration. 
 


CONFIDENTIAL 
 
 
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NAC 300 mg/3 mL i.v. bioavailability 
Final version 1.0, 30AUG2018 
 
Page 33 
During confinement, routine ambulant daily activities will be strongly recommended. 
 
The timing of each meal to be served during confinement is shown in Table11.1.1 above.  
In the evening of day -1, upon confinement, the subjects will receive a light dinner before 
fasting overnight.  
On days 1 and 6 (PK sampling collection days), standardised lunch and dinner will be served.  
On days 2, 3, 4, 5 and 7, standardised breakfast, lunch and dinner will be served.  
 
11.2.1 
Restrictions 
 
During the study, the subjects will be confined from the evening preceding the first drug 
administration (study day -1) until the morning of day 8 (after the final visit). They will attend 
the clinic in the evening of day -1 not later than 18:30.  
 
During confinement, hazardous, strenuous or athletic activities will not be permitted. 


CONFIDENTIAL 
 
 
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NAC 300 mg/3 mL i.v. bioavailability 
Final version 1.0, 30AUG2018 
 
Page 34 
12 
DESCRIPTION OF SPECIFIC PROCEDURES 
 
12.1 
Physical examination 
 
Full physical examinations will be performed at the screening visit, visit 3, day 3 and final 
visit/ ETV. Information about the physical examination will be recorded by the investigator. 
Any abnormalities will be recorded. 
 
Significant findings/illnesses, reported after the start of the study and that meet the definition 
of an AE (see § 16), will be recorded in the subject source documents. 
 
Date of the physical examination, overall investigator’s interpretation (as normal or abnormal 
and, if abnormal, clinically significant or not clinically significant) and clinically significant 
abnormalities (if any) will be reported in the individual CRFs. 
 
12.1.1 
Body weight 
 
Body weight will be recorded during each physical examination. 
Subjects will be weighed (kg) lightly clothed without shoes. Height will be measured at 
screening only and BMI will be recorded. BMI will be calculated as weight [kg]/(height [m] x 
height [m]). 
 
12.1.2 
Vital signs 
 
Subjects blood pressure (BP) and heart rate (HR) will be measured by the investigator or 
his/her deputy after 5 min at rest (in sitting position) at:  
 
 Screening  
 Visit 3, Day 1: at pre-dose and 1 h post-dose 
 Visit 3, Day 3: at 48 h post-dose  
 Visit 5, Day 6: at pre-dose and 1 h post-dose 
 Final visit (at 48 h post-dose)/ETV  
 
12.1.3 
ECGs 
 
One 12-lead ECG will be performed (in sitting/supine position) at screening only. 
 
12.2 
Clinical laboratory assays 
 
Samples of blood and urine will be collected. The following laboratory analyses will be 
performed at the screening visit: 
 
Haematology 
Leukocytes and leukocyte differential count (percentage values and absolute values), 
erythrocytes, haemoglobin (conv. units), haemoglobin (IS units), haematocrit, MCV, MCH, 
MCHC, thrombocytes. 
 
 


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NAC 300 mg/3 mL i.v. bioavailability 
Final version 1.0, 30AUG2018 
 
Page 35 
Blood chemistry 
Electrolytes: sodium, potassium, calcium, chloride, inorganic phosphorus 
Enzymes: alkaline phosphatase, γ-GT, AST, ALT 
Substrates/metabolites: total bilirubin, creatinine, glucose, BUN or  urea, uric acid, total 
cholesterol, triglycerides 
Proteins: total proteins 
 
Serum pregnancy test (women). 
 
Urine analysis 
Urine chemical analysis (stick): pH, specific weight, appearance, colour, nitrites, proteins, 
glucose, urobilinogen, bilirubin, ketones, haematic pigments, leukocytes 
Urine sediment (analysis performed only if positive): leukocytes, erythrocytes, flat cells, 
round cells, crystals, cylinders, mucus, bacteria 
 
Serum virology and bacteriology 
Hepatitis B (HBs antigen), Hepatitis C (HCV antibodies), HIV 1/2 (HIV Ag/Ab combo), 
Treponema pallidum.  
 
The same analyses, with the exception of virology, bacteriology and serum pregnancy test, 
will be performed also at Visit 3, Day 3 (48 h post-dose after the single dose) and at the final 
visit/ETV. 
 
A urine drug test will be performed at the clinical centre at screening, using a urine multi-drug 
kit. The following drugs will be assessed: cocaine, amphetamine, methamphetamine, 
cannabinoids (delta-9-tetrahydrocannabinol - THC), opiates and ecstasy. The same test will be 
repeated upon confinement at Visit 2, day -1.  
 
A serum pregnancy test will be performed by the laboratory at screening. A urine pregnancy 
test will be performed at Visit 2, day -1 at the clinical centre, upon confinement.  
 
Date/time of samples collection, overall investigator’s interpretation (as normal or abnormal 
and, if abnormal, clinically significant or not clinically significant) and clinically significant 
findings (if any) will be reported in the individual CRFs. All clinically significant 
abnormalities after the screening visit will be recorded as AEs. Hard copies of the laboratory 
print-outs will be attached to the CRFs. 
 
12.3 
Sampling for pharmacokinetic analysis 
 
12.3.1 
Venous blood sampling 
 
Venous blood samples (up to 10 mL) will be collected from a forearm vein at the following 
times, both on Day 1, 2, after the single dose, and on Day 6, 7, after the last multiple dose:  
 at pre-dose (0) and 5 (at the end of the infusion), 8, 12, 15, 20, 25, 30, 60 min, 2, 4, 6, 8, 
10, 12, 24 and 32 h post-dose 
 
The start of infusion (not the end) will be considered as time 0. 
 


CONFIDENTIAL 
 
 
Sponsor code Z7244J01 
NAC 300 mg/3 mL i.v. bioavailability 
Final version 1.0, 30AUG2018 
 
Page 36 
Actual sampling times for each subject will be recorded in the individual case report forms 
(CRFs). The actual sampling times should not exceed the recommended tolerance ranges 
presented in the following table. Any deviation outside the recommended ranges will be 
verified through Data Clarification Forms and, if confirmed, will be reported as protocol 
deviation, although it will not automatically lead to the exclusion of the concerned subjects 
from the PK analysis set. 
 
Table 12.3.1.1 
Tolerance ranges for the scheduled sampling times 
Sampling time 
Tolerance range 
Pre-dose (0)
Within 30 minutes before IMP administration
0.0833 h (5 min), 0.1333 (8 min) 
0.200 h (12 min), 0.250 h (15 min)
20 seconds 
0.3333 h (20 min), 0.4167 (25 min), 0.5 h (30 min)
± 1 min
1 h
± 3 min
2, 4 h
± 5 min
6, 8, 10, 12, 24, 32 h
± 10 min
 
Blood samples for PK analysis will be collected using an indwelling catheter with switch 
valve. The cannula will be rinsed, after each sampling, with about 1 mL of sterile saline 
solution. The first mL of blood will be discarded at each collection time. 
 
The remaining blood sample will be collected into heparinised tubes (Li-heparin). 
 
Storage on ice will not last longer than 10 min until centrifugation at 4° C for 10 min at 
2500xg to obtain plasma. Plasma will be collected into separate tubes, where an appropriate 
volume of DTT solution (50 µL DTT solution/mL plasma, i.e. 5% of the plasma volume 
according to the ratio 75 µL of DTT solution for 1.5 mL of plasma) will be added. The 
samples will be mixed shortly (Vortex, 20 sec), divided into two aliquots, P1 and P2 and 
transferred to -70° C as soon as possible. The DTT solution (5 mg DTT/mL of water) will be 
freshly prepared in the evening before each sampling day and kept refrigerated until use. The 
preparation of the solution will be recorded in appropriate forms. 
 
If any clinical assessment, such as vital signs measurement, is foreseen at the same time-point 
as blood sampling for PK analysis, blood collection will be performed at the scheduled time. 
However, vital signs can be influenced by the blood sampling. Therefore, these assessments 
can be performed within 30 min before the pre-dose PK time point (0 h) and within 10 min 
before the other scheduled PK time-points. Any deviations outside the recommended time 
will be verified through Data Clarification Forms. However, since vital signs measurements 
will be performed for safety reasons only, deviations from the planned time schedule will be 
considered not relevant. 
 
12.3.2 
Urine collection 
 
Urine for PK will be collected in the following time intervals both on Day 1, 2, after the 
single dose, and on Day 6, 7 after the last multiple dose:  
 
 0-4; 4-8; 8-12, 12-24 and 24-32 h post-dose 
 


CONFIDENTIAL 
 
 
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NAC 300 mg/3 mL i.v. bioavailability 
Final version 1.0, 30AUG2018 
 
Page 37 
Bladder must be emptied before the end of each collection period. During each interval, urine 
will be collected into containers, kept refrigerated at approximately 4° C and containing DTT 
solution. At the end of each collection interval, urine volume will be measured and after 
thorough mixing, two aliquots of 10 mL each (U1 and U2) will be prepared in polypropylene 
tubes. Urine volume collected in each collection interval will be accurately recorded in 
appropriate forms and in the CRFs. The two aliquots will be stored at -70° C. The DTT 
solution (5 mg DTT/mL of water) will be freshly prepared in the evening before each 
sampling day and kept refrigerated until use. The preparation of the solution will be recorded 
in appropriate forms. 
 
12.3.3 
Analytics  
 
The concentration of total NAC in plasma and urine will be determined at a certified 
bioanalytical laboratory to be designated in China, using a fully validated LC-MS/MS 
method, with a lower quantification limit (LQL) of 10 ng/mL.  
 
Analyses will be performed according to the general Principles of “OECD Good Laboratory 
Practices for testing of chemicals" C(81) 30 (final) and GCP/ CFDA GCP (see also § 9.2). 
 
The method validation report and the analytical report will be attached to the final report. 
 
12.3.4 
Labelling, storage and transport of samples 
 
12.3.4.1 
Samples labelling 
 
Each sample tube will be clearly and unequivocally identified with a label resistant to the 
storage temperature and reporting: 
Study code 
Sponsor code Z7244J01
Subject number 
Subject or Process or Unique Subject Identification number, e.g. 
 etc.
Tube identification
Study day 
1 or 6
Scheduled sampling time
as min and h; see § 12.3.1
 
12.3.4.2 
Samples storage and transport 
 
During the study the samples will be stored at ≤-70° C. At the end of each collection day, 
aliquots 1 and 2 will be stored in separate freezers. 
 
All aliquots 1, packed in sufficient solid CO2, will be shipped by an authorised courier from 
the clinical site, China, to Eurofins Central Laboratory, Shanghai, China. Aliquots 1 will 
remain stored at Eurofins Central Laboratory or a subcontracted bioanalytical laboratory to be 
designated until finalisation of the bioanalytical report. Afterwards, the samples will be 
destroyed and a certificate of destruction will be provided to the sponsor. 
 
The counter-samples (aliquot 2) will remain stored at the clinical site, China. These samples 
could either be: 
CCI
CCI


CONFIDENTIAL 
 
 
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NAC 300 mg/3 mL i.v. bioavailability 
Final version 1.0, 30AUG2018 
 
Page 38 
 sent to the laboratory for reanalysis should this become necessary for analytical reasons 
or if any problems occur during the delivery of aliquots 1, or 
 destroyed at an authorised site, or 
 transferred to the sponsor upon written request. 
 
No analyses different from those stated in this protocol and agreed by the subjects when 
signing the informed consent form will be performed unless a new informed consent and a 
new approval from the Ethical Committee is obtained. The subjects may ask to destroy their 
own samples at any time. 
 
 


CONFIDENTIAL 
 
 
Sponsor code Z7244J01 
NAC 300 mg/3 mL i.v. bioavailability 
Final version 1.0, 30AUG2018 
 
Page 39 
13 
ASSIGNMENT OF STUDY TREATMENT 
 
13.1 
Randomisation 
 
No randomisation will take place in the present study. All the subjects will receive the same 
treatment.  
 
13.2 
Blinding 
 
This is an open study. No masking procedure will be applied.  
 
 
 
 


CONFIDENTIAL 
 
 
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NAC 300 mg/3 mL i.v. bioavailability 
Final version 1.0, 30AUG2018 
 
Page 40 
14 
EVALUATION PARAMETERS 
 
14.1 
Study variables 
 
14.1.1 
Primary variables 
 
After single dose: 
 
 Cmax, tmax, kel, t1/2, AUC(0-t), AUC(0-), AUC(0-12), Vd, CLt, Ae(0-t), Fe(0-t) and CLr 
 
After multiple doses: 
 
 Css_max, tss_max, Css_min, AUCss(0-t), AUCss(0-12), Css_av, R, DF% and Aess(0-t). 
 
14.1.2 
Secondary variables 
 
 TEAEs, vital signs (BP, HR), body weight, physical examinations, laboratory parameters. 
 
14.2 
Pharmacokinetic assessments 
 
14.2.1 
Pharmacokinetic parameters 
 
The following PK parameters will be measured and/or calculated for plasma NAC, using the 
validated software Phoenix® WinNonlin® version 6.3 or higher (Certara USA, Inc., Princeton, 
NJ) (actual version will be stated in the final report), after single dose of IMP: 
 
Cmax: 
Maximum NAC plasma concentration 
tmax: 
Time to achieve Cmax 
kel: 
Terminal elimination rate constant, calculated, if feasible, by log-linear 
regression using at least 3 points 
t1/2: 
Half-life, calculated, if feasible, as ln2/kel 
AUC(0-t): 
Area under the concentration-time curve from single dose to the last observed 
concentration time t, calculated with the linear up/log down trapezoidal 
method 
AUC(0-): 
Area under the concentration-time curve extrapolated to infinity, calculated, if 
feasible, as AUC(0-t) + Ct/kel, where Ct is the last measurable drug 
concentration 
AUC(0-12): 
Area under the concentration-time curve at steady-state in the tau interval 
(from single dose to 12 h post-dose), calculated with the linear up/log down 
trapezoidal method


CONFIDENTIAL 
 
 
Sponsor code Z7244J01 
NAC 300 mg/3 mL i.v. bioavailability 
Final version 1.0, 30AUG2018 
 
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Vd: 
Volume of distribution associated with the terminal slope, calculated, if 
feasible, as Dose/(AUC(0-)*kel) 
CLt: 
Total body clearance, calculated, if feasible, as Dose/ AUC(0-) 
Ae(0-t): 
Total amount of NAC excreted in urine from single dose up to 32 h 
Fe(0-t): 
Total fraction of NAC dose excreted in urine from single dose up to 32 h 
CLr: 
Renal clearance, calculated, if feasible, as Ae(0-t)/ AUC(0-) 
 
The following PK parameters will be measured and/or calculated for plasma NAC, using the 
same software, after multiple doses of IMP: 
 
Css_max: 
Maximum NAC plasma concentration at steady-state 
tss_max: 
Time to achieve Css_max 
Css_min: 
Trough NAC plasma concentration at steady-state, measured as concentration 
at t=12 h 
AUCss(0-t): 
Area under the concentration-time curve at steady-state from the last multiple 
dose to the last observed concentration time t, calculated with the linear 
up/log down trapezoidal method 
AUCss(0-12): 
Area under the concentration-time curve at steady-state in the tau interval 
(from the last multiple dose to 12 h post-dose), calculated with the linear 
up/log down trapezoidal method 
Css_av: 
Average NAC plasma concentration at steady-state, calculated as AUCss(0-12) 
/tau 
R: 
Accumulation ratio, calculated as AUCss(0-12)/ AUC(0-12) 
Aess(0-t): 
Total amount of NAC excreted in urine from the last multiple dose to 32 h at 
steady-state 
DF%: 
Degree of fluctuation over one dosing interval at steady-state, calculated as 
(Css_max - Css_min)/ Css_av*100 
 
The sampling schedule is considered adequate if the ratio AUC(0-t)/ AUC(0-) equals or 
exceeds a factor of 0.8 (i.e. if %AUCextra is <20%) for more than 80% of the individual PK 
profiles. This assures that the primary variable AUC(0-t) covers a sufficient percentage of the 
theoretical total extent of exposure.  
 


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The quality of log-linear regression (and, consequently, the reliability of the extrapolated PK 
parameters) should be demonstrated by a determination coefficient R2 ≥ 0.8. Individual 
extrapolated parameters, when considered unreliable, will be reported as NC (not calculated).  
 
14.3 
Safety assessments 
 
Safety and general tolerability of the IMP will be based on TEAEs, physical examinations 
including body weight, vital signs and routine haematology, blood chemistry and urinalysis 
laboratory tests. 
 


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15 
STATISTICAL METHODS 
 
The data documented in this trial and the measured clinical parameters will be presented using 
classic descriptive statistics (i.e. total number of subjects treated [N], number of observations 
[n], mean standard deviation [SD], minimum [Min], median, maximum [Max]) for 
quantitative variables and frequencies (i.e. count and percentages) for qualitative variables if 
not stated otherwise. 
 
Not available data will be evaluated as “missing values”. The statistical analysis of 
demographic and safety data will be performed using the analysis software SAS®   . 
 
The statistical analysis of PK parameters will be performed using the validated software 
Phoenix® WinNonlin® version 6.3 or higher (Certara USA, Inc., Princeton, NJ) (actual 
version will be stated in the final report). 
 
15.1 
Analysis Sets 
 
15.1.1 
Definitions 
 
A subject will be defined as screened after the signature of the informed consent, regardless of 
the completion of all the screening procedures. 
 
A subject will be defined as eligible if he/she meets all the inclusion/exclusion criteria. 
Otherwise he/she will be defined as a screen failure.  
 
A subject will be defined as enrolled in the study if he/she is included into the interventional 
phase of the study.  
 
The following analysis sets are defined: 
 Enrolled set: all enrolled subjects. This analysis set will be used for demographic, 
baseline and background characteristics 
 Safety set: all subjects who receive at least one dose of IMP. This analysis set will be 
used for the safety analyses 
 PK set: all enrolled subjects who fulfil the study protocol requirements in terms of IMP 
intake and have evaluable PK data readouts for the planned analyses, with no major 
deviations that may affect the PK results. This analysis set will be used for the statistical 
analysis of the PK parameters. 
 
Each subject will be coded as valid or not valid for the safety and the PK set. Subjects will be 
evaluated according to the treatment they actually receive. 
 


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15.1.2 
Reasons for exclusion from the PK set before bioanalysis 
 
Reasons for the exclusion of subjects from the PK set are the following: 
 
 intake of concomitant medications which could render the plasma concentration-time 
profile unreliable 
 AEs which could render the plasma concentration-time profile unreliable 
 administration errors which could render the plasma concentration-time profile unreliable 
 other events which could render the plasma concentration-time profile unreliable 
If one of these events occurs, it will be noted in the CRF during the study. 
 
The samples from the subjects excluded from the PK set should still be assayed and the 
results listed. Subjects should not be excluded from the PK set if the AUC0-t covers less than 
80% of the AUC0-∞. 
 
15.2 
Sample size and power considerations 
 
The sample size was not calculated through any statistical calculation. A sample size of 24 
subjects was estimated as sufficient for the descriptive purposes of the present study. 
Drop-out subjects will not be replaced.  
 
15.3 
Demographic, baseline and background characteristics 
 
Critical demographic characteristics will be examined according to qualitative or quantitative 
data. Qualitative data will be summarised in contingency tables. Quantitative data will be 
summarised using classic descriptive statistics. 
 
15.4 
Analysis of pharmacokinetic parameters 
 
15.4.1 
Descriptive pharmacokinetics 
 
A descriptive PK will be presented. The results will be displayed and summarised in tables 
and figures. Individual and mean curves (+SD at sampling times), indicating inter-subject 
variability, will be plotted. Data below the lower quantification limit (BLQL) will be 
considered as 0 in the calculations and presented as BLQL in listings and tables. As a 
consequence of BLQL (i.e. 0) values, calculated geometric means (if requested) could be null. 
For this reason, in the presence of any null value, the geometric mean will be reported as not 
calculated (NC). 
 
15.5 
Safety and tolerability evaluation 
 
 AEs 
AEs will be coded by System Organ Class (SOC) and Preferred Term (PT), using the Medical 
Dictionary for Regulatory Activities (MedDRA). 
AEs will be classified as pre-treatment AEs (PTAEs) and TEAEs, according to the period of 
occurrence, as follows:  


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 PTAEs: all AEs occurring before the first dose of IMP and not worsening after the 
first dose of IMP 
 TEAEs: all AEs occurring or worsening after the first dose of IMP 
Individual PTAEs and TEAEs will be listed in subject data listings. No summary table will be 
provided for PTAEs. TEAEs will be summarised by treatment and overall. The number and 
percentage of subjects with any TEAE and the number of TEAEs will be tabulated by SOC 
and PT, seriousness, relationship to treatment and severity.  
 
 Physical examination 
Date of the physical examination, overall investigator’s interpretation (as normal or abnormal 
and, if abnormal, clinically significant or not clinically significant) and clinically significant 
abnormalities (if any) will be listed. 
 
 Laboratory data  
Date/time of samples collection, overall investigator’s interpretation (as normal or abnormal 
and, if abnormal, clinically significant or not clinically significant) and clinically significant 
findings (if any) will be listed. All laboratory results will be listed and a table of all the 
abnormal values will be presented. The overall investigator’s interpretation will be 
summarised using tables of frequency.  
The toxicity grading of laboratory tests is determined based on the NCI CTCAE V4.03. 
 
 Vital signs 
Vital signs values will be listed and summarised by descriptive statistics.  
 
 Body weight  
Body weight values will be listed and summarised by descriptive statistics. 
 


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16 
DEFINITION AND HANDLING OF AEs AND SAEs 
 
16.1 
Applicable SOPs 
 
AEs definition, classification and management will follow the CRO’s SOPs, based upon 
applicable local and international regulations. The full SOPs or an operative summary will be 
made available to the clinical centre. 
 
A brief summary of AE definition, classification and management is reported below. 
 
16.2 
Definition of Adverse Event (AE) 
 
An Adverse Event is “any untoward medical occurrence in a patient or clinical trial subject 
administered a medicinal product and which does not necessarily have a causal relationship 
with this treatment”. 
AEs include: 
 worsening (change in nature, severity or frequency) of conditions present at the onset 
of the trial 
 patient/subject deterioration due to the primary illness 
 intercurrent illnesses 
 drug interactions 
 events related or possibly related to concomitant medications 
 abnormal laboratory values, as well as significant shifts from baseline within the range 
of normal, which the investigator considers to be clinically significant. 
An AE can therefore be any unfavourable and unintended sign (including an abnormal 
laboratory finding), symptom, or disease temporally associated with the use of an 
investigational medicinal product, whether or not considered related to the investigational 
medicinal product. 
 
16.3 
Definition of Adverse Drug Reaction (ADR) 
 
An Adverse Reaction is “any untoward and unintended response to an investigational 
medicinal product related to any dose administered”. 
All AEs judged by either the reporting investigator or the sponsor as having a reasonable 
causal relationship to a medicinal product qualify as adverse reactions. The expression 
reasonable causal relationship means to convey in general that there is evidence or matter to 
suggest a causal relationship. 
 
 Unexpected Adverse Drug Reaction 
 
An unexpected ADR is: “An adverse reaction, the nature, or severity of which is not 
consistent with the applicable product information (Reference Safety Information [RSI]).  
 
 Reference Safety Information (RSI): in order to assess whether an adverse reaction is 
expected, the IB will be used. 


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16.4 
Definition of Serious Adverse Events or Serious Adverse Drug Reaction 
 
A Serious Adverse Event (SAE) or Serious Adverse Reaction (SAR) is: “any untoward 
medical occurrence or effect that at any dose: 
 results in death 
 is life-threatening (i.e. the subject was at risk of death at the time of the event/reaction; it 
does not refer to an event/reaction which hypothetically might have caused death if it were 
more severe) 
 requires inpatient hospitalization or prolongation of existing hospitalization 
 results in persistent or significant disability/incapacity (where disability is defined as a 
permanent or substantial disruption of ability to carry out normal life functions, either 
reported or defined as per clinical judgement) 
 is a congenital anomaly/birth defect 
 is an important medical event that may not result in death, be life-threatening, or require 
hospitalization but, according to appropriate medical judgment, it may jeopardize the 
subject and may require medical or surgical intervention to prevent any of the outcomes 
listed in the definition above.  Examples of such events include allergic bronchospasm 
requiring intensive treatment in an emergency room or at home, and blood dyscrasias or 
convulsions that do not result in hospitalization, or development of drug dependency or 
drug abuse.  
 
Any suspected transmission via a medicinal product of an infectious agent is also considered a 
serious adverse event. 
 
A Suspected Unexpected Serious Adverse Reaction (SUSAR) is an ADR that is both 
unexpected (not consistent with the applicable product information, e.g. IB) and also meets 
the definition of a SAE. 
 
A non-serious adverse event is any adverse event that does not meet the criteria listed above 
for a serious adverse event. 
 
 
16.5 
Definition of Severity of Adverse Events 
 
The term “severe” is used to describe the intensity (severity) of a specific event: 
- 
Mild: causing no limitation of usual activities; the subject may experience slight 
discomfort. 
- 
Moderate: causing some limitation of usual activities; the subject may experience 
annoying discomfort. 
- 
Severe: causing inability to carry out usual activities; the subject may experience 
intolerable discomfort or pain. 
 
16.6 
Definition of Adverse Event causality 
 
Causality shall be determined according to the definition of ADR given in 16.3. 
 


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All AE judged by either the investigator or the sponsor as having a reasonable suspected 
causal relationship to an investigational medicinal product qualify as adverse reactions. The 
causality assessment given by the investigator should not be downgraded by the sponsor. 
 
The following binary decision for causality will be used: 
 Reasonable possibility that the IMP caused the event 
 No reasonable possibility that the IMP caused the event 
Features supportive of an association include: 
 temporal plausibility 
 pharmacological properties of the drug or of the substance class 
 course of the adverse event after dechallenge and, if applicable, after rechallenge 
 specific tests indicating involvement of the drug in the occurrence/worsening of the 
adverse event  
 alternative explanations 
 
16.7 
Adverse Events recording 
 
Each AE occurring to a subject, either spontaneously revealed by the subject or observed by 
the investigator, whether believed by the investigator to be related or unrelated to the IMP, 
must be recorded on the AE information page of the CRF (for SAEs information must be 
recorded also on the “Serious Adverse Event Form”).  
The Investigator will also perform an evaluation with respect to seriousness and causality 
(relationship of any AE to IMP) of the AEs. and record it on the appropriate section of the 
CRF and on the ”Serious Adverse Event Form” (if appropriate).  
 
16.8 
AEs monitoring window 
 
 Start of monitoring: from immediately after the signature of the informed consent 
 End of monitoring: final study visit/ETV (observation window) 
An AE occurring after the final visit/ETV and coming to knowledge of the investigator (e.g. 
during a follow-up assessment or by spontaneous reporting by study subjects) must be 
recorded only if it is an ADR, according to the investigator’s judgment. 
 
16.9 
Adverse Events reporting 
 
The official language for reporting is English. The investigator and clinical staff of the present 
study are familiar with English language. 
 
The investigator must report to the CRO all AEs which occur during the study, regardless of 
their relationship to the IMP. Protocol specific AEs or laboratory abnormalities critical to 
safety evaluations are to be identified in the protocol and reported to the sponsor according to 
reporting requirements and within the time periods specified. 
 


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All AEs are recorded by the investigator on the AE information page of the CRF. 
 
In addition, SAE will have to be reported according to the following detailed procedure. 
 
16.9.1 
SAEs reporting 
 
With the exception of those SAEs that are identified as not requiring immediate reporting in 
the protocol, the investigator must report the SAEs to the CRO immediately and no later than 
24 hours from when he/she becomes aware of the SAE, by faxing the “Serious Adverse Event 
Form” (back up plan) or e-mailing as scanned attachment (backup plan) or by Electronic Data 
Capture to the Drug Safety Unit (DSU) personnel (preferred method), as stated in the "List of 
CRO/ personnel" in § 16.13 of this protocol. 
 
The community standards of confidentiality must always be maintained and any relevant 
national legislation on data protection must be followed. 
 
Another copy of the “Serious Adverse Event Form” will be retained by the Investigator for 
the Investigator’s file. 
 
 
If the investigator becomes aware of any SAE occurred to a subject within the follow-up 
window established in the protocol, he/she will report the SAE as above. The SAE will be 
also reported in the CRF. 
 
If outside the follow-up window established in the protocol the investigator becomes aware of 
a SAE, it is the investigator’s responsibility to report the SAE to the CRO. The Investigator 
might use the “Serious Adverse Event Form” via email or fax, but the SAE must not to be 
reported in the CRF, as it is not an event occurred within the study period. 
 
16.10 
Follow-up for Adverse Events 
 
A follow-up “Serious Adverse Event Form” will be filled in if important follow-up 
information (i.e. diagnosis, outcome, causality assessment, results of specific investigations) 
is made available after submission of the initial AE Form for immediate reporting. Follow-up 
“Serious Adverse Event Form” will be reported to the Sponsor as above-described, under 
Section 16.9.1. 
 
In any case of an AE that, in the opinion of the investigator, requires the subject’s 
discontinuation, follow-up information relating to the subjects subsequent course must be 
collected until the event has subsided or the condition stabilised or when the subject is lost to 
follow up and uncontactable. 
 
When follow-up data on non-serious AE are collected, information should be reported under 
“Comments” in the Final report of the CRF. 
 
16.11 
SUSARs management 
 


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The clock for initial expedited reporting starts as soon as the information containing the 
minimum reporting criteria has been received by the sponsor (day 0).  
For fatal and life-threatening SUSARs the EC and Competent Authority (CA) should be 
informed as soon as possible and in any case within 7 days. 
If the initial report is incomplete, e.g. not all the information/assessments were available, a 
complete report should be sent within an additional 8 days. Complete follow-up report should 
be sent within 15 calendar days. 
SUSARs which are not fatal and not life-threatening are to be reported within 15 days. 
The minimum information to be reported includes: 
 Sponsor study number 
 One identifiable coded subject 
 One identifiable reporter 
 One SUSAR 
 One suspect IMP (including active substance name, code) 
 A causality assessment (a reasonable possibility of a causal relationship with the study 
drug can be excluded only if there is information supporting this decision, otherwise it 
cannot be excluded). 
In addition, in order to properly process the report electronically, the following administrative 
information should be provided: 
 
 the sender’s (case) safety report unique identifier, 
 the receipt date of the initial information from the primary source, 
 the receipt date of the most recent information, 
 the worldwide unique case identification number, 
 the sender identifier. 
 
16.12 
Other events qualified for expedited reporting 
 
Other safety issues also qualify for expedited reporting when they might materially alter the 
current benefit-risk assessment of a medicinal product or would be sufficient to consider 
changes in the medicinal product administration or in the overall conduct of the trial, for 
instance: 
 single case reports of an expected serious adverse reaction with an unexpected 
outcome (e.g. a fatal outcome) 
 an increase in the rate of occurrence of an expected serious adverse reaction, which is 
judged to be clinically important. 
 post-study SUSARs that occur after the subject has completed a clinical trial and are 
reported to the investigator by the subject. 
 new events relating to the conduct of the trial or the development of the medicinal 
product likely to affect the safety of the subjects, such as : 
 a SAE which could be associated with the trial procedures and which could modify 
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 a significant hazard to the subject population such as lack of efficacy of a medicinal 
product used for the treatment of a life-threatening disease 
 a major safety finding from a newly completed animal study (such as 
carcinogenicity) or from other clinical trials. 
 
16.13 
SAEs: contacts 
 
 
The investigator will report any SAE to the CRO. The CRO’s details for SAEs are the 
following: 
 
 
 
16.14 
Pregnancy 
 
Subjects must be instructed that known or suspected pregnancy occurring during the study 
should be confirmed and reported to the Investigator, who must then withdraw the subject 
from the study without delay. The Investigator should also be notified of pregnancy occurring 
during the study but confirmed after its completion. In the event that a subject is subsequently 
found to be pregnant after inclusion in the study, then any pregnancy will be followed to term 
and the status of mother and child will be reported to the Sponsor after delivery through the 
Pregnancy Report Form provided by the Sponsor. The Investigator will send pregnancy 
reports to DSU within the timeframes of SAEs, with follow-up information to be actively 
sought for the outcome of pregnancy.  
- 
Part I of the Form is completed at the time of awareness of foetal exposure during 
pregnancy.  
- 
Part II of the Form is filled in when information on pregnancy outcome becomes 
available. 
If pregnancy results in abnormal outcome that the investigator considers to be due to the IMP, 
this will be treated as an expedited ADR report. 
 
CCI
CCI


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17 
DATA MANAGEMENT PROCEDURES 
 
17.1 
Data collection – CRFs 
 
The investigator must ensure that the clinical data required by the study protocol are carefully 
reported in the CRFs. He/she must also check that the data reported in the CRFs correspond to 
those in the subject’s source documents.  
To ensure legibility, the CRFs should be filled out in English, in block capitals with a ball-
point pen (not pencil, felt tip or fountain pen). Any correction to the CRFs' entries must be 
carried out by the investigator or a designated member of staff. Incorrect entries must not be 
covered with correcting fluid, or obliterated, or made illegible in any way. A single stroke 
must be drawn through the original entry. Corrections have to be dated and initialled. In the 
interest of completeness of data acquisition, the questions which are repeated in each section 
of the CRFs should be answered in full, even if there are no changes from a previous 
examination. The investigator must provide a reasonable explanation for all missing data. 
The CRFs will be completed, signed by the investigator, sent to the CRO for data 
management procedures and finally sent to the sponsor.  
 
 
17.2 
Database management 
 
The CRO will provide a data entry and will update and verify the database and create the final 
data sets. The final data file will be transferred to the sponsor in the agreed format with all the 
other study documentation. 
 
17.2.1 
Coding dictionaries 
 
Medical/surgical history and underlying diseases, clinically significant physical examination 
abnormalities and AEs will be coded using the Medical Dictionary for Regulatory Activities 
(MedDRA™). 
Previous and concomitant medications will be coded using the WHO Drug Dictionary 
Enhanced (WHODDE). The version of the coding dictionaries will be stated in the study 
report. 


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STUDY MONITORING, QUALITY CONTROL AND 
QUALITY ASSURANCE 
 
18.1 
Monitoring 
 
The monitoring visits will be conducted by appropriate staff of PAREXEL China Co. Ltd 
according to PAREXEL SOPs.  
 
Monitoring activities, including monitoring purpose, selection and qualifications of monitors, 
extent and nature of monitoring, monitoring procedures, monitoring reports will comply with 
ICH-GCP chapter 5.18, CFDA GCP requirements and, when appropriate, further national 
regulatory guidelines. 
 
Adequate time and availability for monitoring activities should be ensured by the investigator 
and key study personnel. 
 
Data verification is required and will be done by direct comparison with source documents, 
always giving due consideration to data protection and medical confidentiality. In this respect 
the investigator will assure support to the monitor at all times. 
 
The investigator agrees, by written consent to this protocol, to fully co-operate with 
compliance checks by allowing authorised individuals to have access to all the study 
documentation. In addition to the monitoring activities performed by the study monitor, the 
sponsor could perform some quality control activities to verify the compliance with the study 
procedures, the ICH-GCP guidelines and CFDA GCP.  
 
18.2 
Quality Control and Quality Assurance  
 
The CRO has implemented and maintains a Quality System that includes quality controls and 
audits at different study steps with written SOPs to ensure that the study is conducted in 
compliance with the protocol and all effective amendments, ICH-GCP, and the applicable 
regulatory requirement(s) and that data have been reliably and correctly generated, recorded, 
processed and reported, in agreement with the ALCOAC principles (Attributable-Legible-
Contemporaneous-Original-Accurate-Complete). 
 
The clinical site is responsible for implementing and maintaining quality assurance and a 
quality control system to ensure that the study is conducted, and data are generated, 
documented (recorded), and reported in compliance with the protocol, ICH-GCP, CFDA GCP 
and any applicable regulatory requirement(s). 
This protocol has been audited by the Sponsor QA.   
The CRO and the sponsor will be responsible for their respective activities. 
 
The sponsor may transfer any or all of the sponsor’s trial-related duties and functions to a 
CRO, but the ultimate responsibility for the quality and integrity of the trial data always 
resides with the sponsor. 
 
 


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18.3 
Applicable SOPs 
 
The sponsor, the clinical centre and the CRO will follow their respective SOPs in the conduct 
of the respective activities, unless otherwise stated in written agreements. CRO/Sponsor's 
SOPs will be used for AE and SAE definition and management. SOPs will be made available 
for Sponsor’s review, if required. 
 
18.4 
Data access 
 
The investigator and the CRO will ensure that all source data, medical records, CRFs and all 
other documentation that is relevant to this study will be made accessible for monitoring 
activities, audits, IEC review, and regulatory inspections. 
 
18.5 
Audits and inspections 
 
The sponsor, independent bodies acting on behalf of the sponsor and the CRO have the right 
to perform audits according to ICH-GCP, CFDA GCP and CFDA responsibilities. 
 
The study may also be inspected by regulatory authorities.  
 
The investigator and the CRO agree, by written consent to this protocol, to fully co-operate 
and support audits and inspections compliance checks by allowing authorised individuals to 
have access to all the study documentation. 
 
 
 


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19 
ETHICAL CONSIDERATIONS 
 
19.1 
Ethics and Good Clinical Practice (GCP) 
 
The study will be performed in accordance with the relevant guidelines of the Declaration of 
Helsinki. 
The approval of the study protocol by the local IEC and by the Chinese Health Authorities 
will be obtained before the start of the study.  
 
The present clinical study will be carried out according to the current revision of Good 
Clinical Practice (GCP), ICH topic E6 (R2), CFDA GCP and any applicable local law 
requirements. 
 
19.2 
Informed consent 
 
Before being enrolled into the clinical study, the subjects must have expressed their consent to 
participate, after the investigator has explained to them, clearly and in details, the scope, the 
procedures and the possible consequences of the clinical study. Information will be given in 
both oral and written form. The information sheet and informed consent form will be prepared 
in the local language by the CRO and must be approved by the EC and regulatory authorities. 
It will include all the elements required by law according to the ICH-GCP and CFDA GCP 
recommendations.  
In addition to the standard requirements that physicians are currently obliged to observe when 
providing information, the following points must also be covered: 
 a description of the aims of the study and how it will be organised 
 the type of treatment (information on the IMP and treatment procedures, as applicable) 
 any potential negative effects attributable to the study product or treatment 
 the freedom to ask for further information at any time 
 the subjects’ right to withdraw from the clinical study at any time without giving reasons 
and without jeopardising their further course of medical treatment  
 the existence of a subject insurance cover and obligations following from this cover 
Adequate time and opportunity to satisfy questions will be given to the subjects and the time 
will be recorded. 
 
The investigator will be supplied with an adequate number of blank informed consent forms 
to be used. The forms will be signed and dated by both the investigator and the subject.  
A copy of the signed form will be given to the subject. 
 
To ensure medical confidentiality and data protection, the signed informed consent forms will 
be stored in the investigator's study file according to the regulatory requirements (see § 20.3). 
The investigator will allow inspection of the forms by authorised representatives of the 
sponsor, EC members and regulatory authorities. He/she will confirm, by signing and dating 
the forms, that informed consent has been obtained. 
 
 


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19.3 
Insurance policy 
 
An insurance cover has been issued in favour of the subjects participating in this clinical 
study. The insurance is in compliance with the local regulation and with the requirements of 
the Health Authorities. 
 
19.4 
Withdrawal of subjects 
 
It will be documented whether or not each subject completed the clinical study. If, for a 
subject, study treatment or observations are discontinued, the primary reason for 
discontinuation will be recorded. 
 
19.4.1 
Primary reason for discontinuation 
 
 Adverse event: Any (significant) adverse event that in the opinion of the investigator or 
concerned subject is not compatible with study continuation. For the definition of AE, 
please refer to § 16.2. 
 death: the absence of life or state of being dead 
 lost to follow-up: the loss or lack of continuation of a subject to follow-up 
 non-compliance with study drug: an indication that a subject has not agreed with or 
followed the instructions related to the study medication 
 physician decision: a position, opinion or judgment reached after consideration by a 
physician with reference to the subject 
 pregnancy: pregnancy is the state or condition of having a developing embryo or fetus in 
the body (uterus), after union of an ovum and spermatozoon, during the period from 
conception to birth 
 protocol deviation: an event or decision that stands in contrast to the guidelines set out by 
the protocol 
 study terminated by sponsor: an indication that a clinical study was stopped by its 
sponsor 
 technical problems: a problem with some technical aspect of a clinical study, usually 
related to an instrument 
 withdrawal by subject: study discontinuation requested by a subject for whatever reason 
 other: different than the ones previously specified 
 
For any subject discontinuing the study, the investigator will:  
 ask the subject to undergo, as far as possible, a final medical visit (ETV) to examine the 
subject's health conditions and perform the required blood sampling for the laboratory 
assays. This examination will verify that all values tested at screening have remained 
within a clinically acceptable range (i.e. not clinically significant changes compared to 
screening) 
 arrange for alternative medical care of the withdrawn subject, if necessary 
 record the subject decision about the use of collected biological samples 


CONFIDENTIAL 
 
 
Sponsor code Z7244J01 
NAC 300 mg/3 mL i.v. bioavailability 
Final version 1.0, 30AUG2018 
 
Page 57 
 report in the CRF date and time of the last dose administration, and date and primary 
reason of study discontinuation 
 record in the CRF any follow-up, if the subject is withdrawn for an AE 
Discontinued subjects will not be replaced.  
 
19.5 
Study termination 
 
The study will be considered terminated at the date of the last visit of the last subject or upon 
completion of any follow-up procedure described in protocol. The investigator and the 
sponsor have the right to discontinue the study at any time for reasonable medical and/or 
administrative reasons. As far as possible, this should occur after mutual consultation. 
Reasons for discontinuation have to be documented appropriately. 


CONFIDENTIAL 
 
 
Sponsor code Z7244J01 
NAC 300 mg/3 mL i.v. bioavailability 
Final version 1.0, 30AUG2018 
 
Page 58 
20 
ADMINISTRATIVE PROCEDURES 
 
20.1 
Material supplied to the clinical centre 
 
Beside IMP, the following study material will be supplied to the clinical centre: 
 final version of the study protocol 
 CRF for each subject plus some spare copies 
 copy of the investigator’s brochure (IB) relative to the IMP 
 informed consent forms 
Moreover, before the start of the study, the investigator will be provided with the following 
documents: CFDA GCP, ICH guidelines, confidentiality agreement (if applicable), protocol 
amendments (if any), declaration of Helsinki, insurance statement, SAE forms, financial 
agreement (if applicable), confidential subject identification code list form, drug 
accountability forms, investigator and study staff list form. 
 
20.2 
Protocol amendments 
 
In order to obtain interpretable results, neither the investigator nor the sponsor will alter the 
study conditions agreed upon and set out in this protocol. Amendments should be made by 
mutual agreement between the investigator and the sponsor. Any amendment must be set out 
in writing, giving the reasons, and being signed by all concerned parties. The amendment 
becomes then part of the protocol. 
 
20.3 
Study documentation and record keeping 
 
The investigator should ensure the accuracy, completeness, legibility, and timeliness of the 
data reported to the sponsor in the CRFs and in all required reports. 
 
The investigator must keep source documents for each subject in the study. All information 
on the CRFs must be traceable to these source documents, which are generally stored in the 
subject's medical file. The source documents should contain all demographic and medical 
information, including laboratory data, ECGs, etc., and the original signed informed consent 
forms. 
 
Data reported on the CRF that are derived from source documents should be consistent with 
the source documents or the discrepancies should be explained. 
 
The investigator and the sponsor should maintain the study documents as specified in the 
“Essential Documents for the Conduct of a Clinical Trial” chapter 8 of ICH-GCP and as 
required by CFDA GCP and the applicable regulatory requirement(s).  
 
These are documents which individually and collectively permit evaluation of a study and the 
quality of the data produced and include groups of documents, generated before the study 
commences, during the clinical study, and after termination of the study and include but are 
not limited to, study protocol, amendments, submission and approval of EC, raw data of 
subjects including lab tests and ECG tracing, insurance contracts, certificate of analysis of the 


CONFIDENTIAL 
 
 
Sponsor code Z7244J01 
NAC 300 mg/3 mL i.v. bioavailability 
Final version 1.0, 30AUG2018 
 
Page 59 
IMP(s), drug accountability records, signed informed consent forms, confidential subjects 
identification code, CRFs, curricula vitae of the investigator and other participants in the 
study, study staff lists and responsibilities, monitoring reports and final study report. 
 
The investigator and the sponsor should take measures to prevent accidental or premature 
destruction of these documents. 
 
Study documents must be retained by the investigator and the sponsor as long as needed to 
comply with ICH-GCP, CFDA GCP, national and international regulations. By signing the 
protocol, the investigator and the sponsor agree to adhere to these requirements. 
 
20.4 
Study subjects’ recruitment 
 
Study participants will be recruited from the volunteers’ database. The study site may also use 
the site management organisation to recruit the study participants.  
 
In addition, they may also use recruitment posters in other departments within the hospital, 
community outside or newspapers to raise awareness of the study that is going on. 
 
Other departments within the hospital will be involved to recommend study participants as 
well.    
 
The clinical site has detailed SOPs on the recruitment process. 
 
20.5 
Confidentiality and data protection 
 
By signing this protocol, the investigator and the CRO agree to keep all the information 
provided by the sponsor in strict confidentiality and to request the same confidentiality from 
his/her staff. Study documents provided by the sponsor (protocols, IB, CRFs and other 
materials) will be stored appropriately to ensure confidentiality. The information provided by 
the sponsor to the investigator and to the CRO cannot be disclosed to others without direct 
written authorisation from the sponsor, except for the extent necessary to obtain the informed 
consent from the subjects wishing to participate in the study. 
 
Data on subjects collected in the CRFs during the study will be documented in an anonymous 
way. If, as an exception, for safety or regulatory reasons identification of a subject becomes 
necessary, the monitor, the sponsor and the investigator will be bound to keep this 
information confidential. 
 
20.6 
Publication policy 
 
The sponsor agrees that the study results (including negative and inconclusive as well as 
positive results) can be made publicly available by the investigator publishing in peer 
reviewed journals, presenting results at scientific congresses and posting information and 
results on internet-based public registers and databases. 
Study results will be communicated in full to the competent Health Authorities by the 
submission of a complete clinical study report. 
 


CONFIDENTIAL 
 
 
Sponsor code Z7244J01 
NAC 300 mg/3 mL i.v. bioavailability 
Final version 1.0, 30AUG2018 
 
Page 60 
As the sponsor agrees that the study results can be published by the investigator(s), the 
investigator agrees to submit any manuscript (abstract, publication, paper, etc.) to the sponsor 
before any public disclosure. 
This will be done in order to ensure that clinical study results are reported in an objective, 
accurate and balanced manner. The sponsor reviews the proposed manuscripts, before 
submission, within a reasonable period of time (30-90 days in relation with the complexity of 
the work). 
 
The investigator will also be provided by the sponsor with the clinical study report and the 
results of any additional analysis, tables, figures, etc. undertaken for the purposes of the 
article, in order to take responsibility for the content of the publication(s). 
On an exceptional basis, the sponsor may temporarily delay registration of certain data 
elements (e.g. compound, name, outcome, measures, etc.) to seek necessary intellectual 
property protection. This is because early disclosure of such data could, in some 
circumstances, prevent or negatively impact patentability. 
 
 
 


CONFIDENTIAL 
 
 
Sponsor code Z7244J01 
NAC 300 mg/3 mL i.v. bioavailability 
Final version 1.0, 30AUG2018 
 
Page 61 
21 
STUDY RESPONSIBLE PERSONS 
 
21.1 
Sponsor 
 
Zambon S.p.A., via Lillo del Duca 10, I-20091 Bresso, Italy 
Phone: 
+39.02.66.52.41 
Fax: 
+39.02.66.50.14.92 
 
 
Protocol Review Committee Chairman  
 
 
Sponsor’s representatives 
 
 
 
 
 
21.2 
Drug assay 
 
Eurofins Central Laboratory Shanghai, 395 Jiang Chang West Road, 7th Floor, Shanghai, 
200436 China 
 
 
 
21.3 
Project management, data analysis & reporting, monitoring  
 
PAREXEL International (IRL) Limited (“PAREXEL”), a corporation organized under the 
laws of Ireland with a registered address at 70 Sir John Rogerson’s Quay, Dublin 2, Ireland. 
PPD
PPD


CONFIDENTIAL 
 
 
Sponsor code Z7244J01 
NAC 300 mg/3 mL i.v. bioavailability 
Final version 1.0, 30AUG2018 
 
Page 62 
22 
REFERENCES 
 
1. Ventresca GP, Cicchetti V, Ferrari V. Drugs in Bronchial Mucology. 1989. Cap. 4.1 Thiols 77-102. Edited 
by PC Braga and Allegra. Raven Press Ltd 
2. Aruoma OI, Halliwell B, Hoey BM, Butler J. The antioxidant action of N-acetylcysteine: Its reaction with 
hydrogen peroxide, hydroxyl radical, superoxide and hypochlorous acid. Free Radical Biol Med 1988; 
6:593-597 
3. Ziment I. Acetylcysteine: a drug that is much more than a mucokinetic. Biomed & Pharmacother 1988; 42: 
513-520 
4. Borgstrom L, Kagedal B, Paulsen O. Pharmacokinetics of N-acetylcysteine in man. Eur J Clin Pharmacol 
1986; 31: 217-222 
5. Olsson B, Johansson M, Gabrielsson J, Bolme P. Pharmacokinetics and bioavailability of reduced and 
oxidized N-acetylcysteine. Eur J Clin Pharmacol 1988; 34: 77-82 
6. Burgunder JM, Varriale A, Lauterburg BH. Effect of N-acetylcysteine on plasma cysteine and glutathione 
following paracetamol administration. Eur J Clin Pharmacol 1989; 36: 127-131 
7. De Caro L, Ghizzi A, Costa R, Longo A, Ventresca GP, Lodola L.  Pharmacokinetics and bioavailability of 
oral acetylcysteine in healthy volunteers. Arzneim-Forsh/Drug Res 1989; 39: 382-386 
8. Borgstrom L, Kagedal B. Dose dependent pharmacokinetics of N-acetylcysteine after oral dosing to man. 
Biopharmaceutics & Drug Disposition 1990; 11: 131-136 
9. Frascio M. Pharmacokinetic study after single dose (1, 2, and 4 g) of a new N-acetylcysteine (NAC) oral 
formulation. Zambon Group Internal Report, code ZPM/94/34 
10. Crestani S. Open label, randomized, cross-over, pharmacokinetic study to assess the bioequivalence of two 
oral N-Acetylcysteine preparations: 600 mg film coated tablets versus 600 mg effervescent tablets. 2002 
Report in Zambon Group’s file 
11. Rusca A. Study CRO-PK-14-286 – Sponsor code Z7169J02. Comparative bioavailability study of N-acetyl-
cysteine (NAC) 600 mg uncoated tablets vs. NAC 600 mg film-coated tablets in healthy male and female 
volunteers. CROSS Research S.A., Switzerland 2014 
12. Rusca A. Study CRO-PK-07-189 – Sponsor code 7169J01. Comparative bioavailability study of a new oral 
N-acetyl-cysteine (NAC) formulation vs. Fluimucil®, Zambon in healthy volunteers. CROSS Research S.A., 
Switzerland, 2008 
13. Rusca A. Study CRO-PK-09-225 – Sponsor code 7203J01. Pilot bioavailability study of a new 1200 mg N-
acetyl-cysteine (NAC) effervescent tablet formulation vs. Fluimucil® 600 mg effervescent tablet (Zambon 
S.p.A., Italy) in healthy male and female volunteers. CROSS Research S.A., Switzerland, 2010 
14. Brown M, Bjorksten A, Medved I and M McKenna. Pharmacokinetics of intravenous N-acetylcysteine in 
men at rest and during exercise. Eur J Clin Pharmacol 2004;60:717-723 
15. Jones AL, Jarvier D. Simpson D. Pharmacokinetics of N-acetylcysteine are altered in patients with chronic 
liver diseases. Aliment Pharmacol Ther. 1997; 787-791 
16. Prescott LF, Donovan JW, Jarvie DR, Proudfoot AT. The disposition and kinetics of intravenous N-
acetylcysteine in patients with paracetamol overdosage. Eur J Clin Pharmacol 1989; 37: 501-506 
17. Pendyala L, Creaven PJ. Pharmacokinetic and pharmacodynamic studies of N-acetylcysteine, a potential 
chemopreventing agent during a phase I trial. Cancer Epidemiology, Biomarkers and Prevention 1995; 4: 
245-251 
18. Rodenstein D, De Coster A, Gazzaniga A. Pharmacokinetics of oral acetylcysteine: absorption, binding and 
metabolism in patients with respiratory disorders. Clinical Pharmacokinetics 1978; 3: 247-254 
19. Pharmacokinetics of N-Acetylcysteine following repeated i.v. infusions to hemodialysed chronic renal 
failure patients. Internal Report 2002. Zambon Group’s file 


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Sponsor code Z7244J01 
NAC 300 mg/3 mL i.v. bioavailability 
Final version 1.0, 30AUG2018 
 
Page 63 
20. Sjodin K, Nilsson E, Hallemberg A, Tunek A. Metabolism of N-acetyl-l-cysteine Biochem. Pharmacol 38 
1989; 22: 3981-3985 
21. Locatelli C. Fluimucil 20% injectable (N-Acetylcysteine). Expert Report on the clinical documentation 
1996. Zambon Group’s file 
22. N-acetyl-L-cysteine (NAC) intravenous Investigator’s Brochure, edition number 1, 20 April 2018 
23. Guidelines on GMP, published by the Commission in The rules governing medicinal products in the 
European Community, Volume 4 
24. Guidance on the investigation of bioequivalence. CPMP/EWP/QWP/1401/98 Rev. 1/Corr **, 20 January 
2010  
25. U.S. Department of Health and Human Services and U.S. Department of Agriculture, Nutrition and your 
health: 2015-2020 Dietary Guidelines, Appendix 9 
 


CONFIDENTIAL 
Sponsor code Z7244J01 
NAC 300 mg/3 mL i.v. bioavailability 
Final version 2.0, 19Dec2018 
Page 1 
CLINICAL STUDY PROTOCOL 
Sponsor code Z7244J01 
Phase I study to evaluate pharmacokinetics, safety and tolerability 
of single and multiple i.v. doses of N-acetylcysteine (NAC) in 
Chinese healthy volunteers 
Single and multiple dose, single centre, open-label, one-way, pharmacokinetics, safety and 
tolerability clinical trial
Test formulation:
NAC 300 mg/ 3 mL solution for injection, Zambon S.p.A., Italy.
Sponsor: 
Zambon S.p.A., via Lillo del Duca 10, I-20091 Bresso, Italy 
Phone: +39.02.66.52.41 
Fax:
+39.02.66.50.14.92
Investigator: 
 - Principal investigator 
Ruijin Hospital Affiliated to Shanghai Jiao Tong University 
School of Medicine, Phase I Unit, No.197, Rui Jin Er Road, 
Shanghai 
China 
Email 
Development phase:
I
Version and date:
Final version 2.0, 19 Dec 2018 
This study will be conducted in accordance with the current version of Good Clinical Practice (GCP),  
ICH topic E6 (R2 and CFDA GCP) 
Property of the sponsor 
May not be used, divulged, published or otherwise disclosed without the consent of the sponsor 
PPD
PPD


PPD
PPD
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PPD


PPD
PPD
PPD


CONFIDENTIAL 
Sponsor code Z7244J01 
NAC 300 mg/3 mL i.v. bioavailability 
Final version 2.0, 19Dec2018 
Page 4 
2
STUDY SYNOPSIS 
Title: Phase I study to evaluate pharmacokinetics, safety and tolerability of single and multiple i.v. doses of 
N-acetylcysteine (NAC) in Chinese healthy volunteers
Protocol number: Z7244J01
Clinical phase: Phase I
Study design: Single and multiple dose, single centre, open-label, one-way, pharmacokinetics, safety and 
tolerability clinical trial
Planned nr. of centres / countries: 1/China
Investigator and centre: : Principal investigator: 
 Ruijin Hospital Affiliated to Shanghai 
Jiao Tong University School of Medicine, Phase I Unit, No.197,Rui Jin Er Road, Shanghai, China
Investigational medicinal product (IMP):
Test product:
NAC, 300 mg/ 3 mL solution for injection, Zambon S.p.A., Italy
Dose regimen:  
Two ampoules (300 + 300 mg) corresponding to a total dose of 600 mg of NAC diluted in 10 mL of NaCl 0.9% 
saline solution, will be administered by a 5-minute intravenous (i.v.) infusion. 
Single dose: 
One (1) dose of the investigational product will be administered under fasting conditions on day 1 at 08:00 ±1 h 
Multiple dose regime: 
Five (5) doses of the investigational product will be administered twice a day (b.i.d.) on days 4 and 5 at 08:00 ±1 
h and 20:00 ±1 h and one dose will be administered on day 6 at 08:00 ±1.
Objective: To evaluate the NAC pharmacokinetics, safety and tolerability after single and multiple dose i.v. 
administration.
End-points:  
Primary end-point:

To evaluate pharmacokinetic parameters of NAC in plasma after single and multiple dose administration of 
the investigational product.  
Secondary end-points:  

To collect safety and tolerability data after single and multiple dose administration of the investigational 
product.
Study variables: 
Primary variables:  
After single dose: 

Cmax: maximum NAC plasma concentration

tmax: time to achieve Cmax

kel: terminal elimination rate constant, calculated, if feasible, by log-linear regression using at least 3 points 

t1/2: NAC half-life, calculated, if feasible, as ln2/kel

AUC(0-t): area under the concentration-time curve from single dose to the last observed concentration time t, 
calculated with the linear up/log down trapezoidal method 

AUC(0-∞): area under the concentration-time curve extrapolated to infinity, calculated, if feasible, as AUC(0-t)
+ Ct/kel, where Ct is the last measurable drug concentration 

AUC(0-12): area under the concentration-time curve at steady-state in the tau interval (from single dose to 12 h 
post-dose), calculated with the linear up/log down trapezoidal method 

Vd: volume of distribution associated with the terminal slope, calculated, if feasible, as Dose/(AUC(0-∞)*kel) 

CLt: total body clearance, calculated, if feasible, as Dose/ AUC(0-∞)

Ae(0-t): total amount of NAC excreted in urine from single dose up to 32 h  

Fe(0-t): total fraction of NAC dose excreted in urine from single dose up to 32 h 

CLr: renal clearance, calculated, if feasible, as Ae(0-t)/ AUC(0-∞)
PPD


CONFIDENTIAL 
Sponsor code Z7244J01 
NAC 300 mg/3 mL i.v. bioavailability 
Final version 2.0, 19Dec2018 
Page 5 
STUDY SYNOPSIS (cont.) 
Study variables (continued): 
After multiple dose: 

Css_max: maximum NAC plasma concentration at steady-state 

tss_max: time to achieve Css_max

Css_min: trough NAC plasma concentration at steady-state, measured as concentration at t=12 h  

AUCss(0-t): area under the concentration-time curve at steady-state from the last multiple dose to the last 
observed concentration time t, calculated with the linear up/log down trapezoidal method 

AUCss(0-12): area under the concentration-time curve at steady-state in the tau interval (from the last multiple 
dose to 12 h post-dose), calculated with the linear up/log down trapezoidal method 

Css_av: average NAC plasma concentration at steady-state, calculated as AUCss(0-12) /tau  

R: accumulation ratio, calculated as AUCss(0-12)/ AUC(0-12)

DF%: 
degree 
of 
fluctuation 
over 
one 
dosing 
interval 
at 
steady-state, 
calculated 
as 
(Css_max - Css_min)/ Css_av*100 

Aess(0-t): total amount of NAC excreted in urine from the last multiple dose to 32 h at steady-state 
Secondary variables:  

Treatment emergent adverse events (TEAEs), vital signs (blood pressure, heart rate), body weight, physical 
examinations, clinical laboratory parameters.
Analytics: total NAC will be determined in plasma and urine at a certified bioanalytical laboratory to be 
designated, using a LC-MS/MS validated method with a Lower Quantification Limit (LQL) of 10 ng/mL. 
Analytical facilities and procedures are in compliance with the general principles of GLP regulations
Sample size: Twenty-four (24) healthy male and female Chinese volunteers will be included in the study. Drop-
out subjects will not be replaced.
Main selection criteria:  
Inclusion criteria: 
1.
Informed consent: signed written informed consent before inclusion in the study 
2.
Ethnicity, Sex and Age: Chinese males and females, 18-45 year old inclusive
3.
Weight: body weight ≥50 kg;
4.
Body Mass Index: 19-26 kg/m2 inclusive
5.
Vital signs: systolic blood pressure 100-139 mmHg, diastolic blood pressure 50-89 mmHg, heart rate 50-90 
bpm, measured after 5 min at rest in the sitting/supine position (to be chosen according to the usual procedure 
at the clinical site) 
6.
Full comprehension: ability to comprehend the full nature and purpose of the study, including possible risks 
and side effects; ability to co-operate with the investigator and to comply with the requirements of the entire 
study
7.
Nicotine addiction (smoker subjects only): ability to abstain from smoking for the duration of the clinical study
8.
Contraception and fertility (women only): women of child-bearing potential must be using at least one of the 
following reliable methods of contraception: 
a. Hormonal oral, implantable, transdermal, or injectable contraceptives for at least 60 calendar days before 
the screening visit 
b. A non-hormonal intrauterine device or female condom with spermicide or contraceptive sponge with 
spermicide or diaphragm with spermicide or cervical cap with spermicide for at least 60 calendar days 
before the screening visit 
c. A male sexual partner who agrees to use a male condom with spermicide 
d. A sterile sexual partner 
Women of non-child-bearing potential or in post-menopausal status for at least 1 year will be admitted. 
Women of childbearing potential should be willing to adopt abstinence or contraception measures during the 
study and two weeks post-dose. 
For all women, pregnancy test result must be negative at screening and day -1.


CONFIDENTIAL 
Sponsor code Z7244J01 
NAC 300 mg/3 mL i.v. bioavailability 
Final version 2.0, 19Dec2018 
Page 6 
STUDY SYNOPSIS (cont.) 
Exclusion criteria: 
1.
Electrocardiogram (12-lead ECG in supine position): clinically significant abnormalities 
2.
Physical findings: clinically significant abnormal physical findings which could interfere with the objectives 
of the study 
3.
Laboratory analyses: clinically significant abnormal laboratory values indicative of physical illness, in 
particular significant laboratory abnormality indicative of hepatic condition (more than 3 times the upper 
limit) 
4.
Allergy: ascertained or presumptive hypersensitivity to the active principle and/or formulations' ingredients; 
history of anaphylaxis to drugs or allergic reactions in general, which the investigator considers may affect 
the outcome of the study 
5.
Diseases: significant history of renal, hepatic, gastrointestinal, cardiovascular, respiratory, skin, 
haematological, endocrine, urologic, metabolic, neurological or psychiatric diseases, as determined by the 
investigator, that may interfere with the aim of the study; history of carcinoma in situ and malignant disease; 
active bacterial or viral infection and fever >38° C within 48 h prior to study treatment administration 
6.
Virology: positive result of HIV, hepatitis B (HBV), hepatitis C (HCV) or Treponema pallidum (TP) assays 
7.
Surgery: any surgery within 2 months of screening (excluding diagnostic surgery) 
8.
Medications: medications, including over the counter (OTC) medications, herbal remedies and traditional 
Chinese remedies for 2 weeks before the start of the study. Hormonal contraceptives for women will be 
allowed 
9.
Investigative drug studies: participation in the evaluation of any investigational product for 30 calendar days 
before this study 
10. Blood donation: blood donations for 90 calendar days before this study 
11. Drug, alcohol, caffeine, tobacco: history of drug, alcohol [>1 drink/day for females and >2 drinks/day for 
males, defined according to the USDA Dietary Guidelines 2015-2020] caffeine (>5 cups coffee/tea/day) or 
tobacco abuse (≥10 cigarettes/day)
12. Abuse drug test: positive urine abuse drug test at screening or day -1
13. Alcohol test: positive alcohol breath test at day -1
14. Diet: abnormal diets (<1600 or >3500 kcal/day) or substantial changes in eating habits in the 4 weeks before 
this study; vegetarians; consumption of alcohol, grapefruit, products containing grapefruit, or beverages 
containing xanthines (coffee, tea, soda, coffee with milk, energy drinks) within 48 hours prior to the 
enrolment  
15. Pregnancy (females only): positive or missing pregnancy test at screening or day -1, pregnant or lactating 
women 
16. Vaccination within 4 weeks of study treatment 
17. Other unspecified reasons that, in the opinion of the investigator, make the subject unsuitable for enrolment.
Schedule:
Day 
Procedures/Assessments 
Notes 
Screening – Visit 1
From day -14 to 
day -2 

Explanation to the subject of study aims, procedures 
and possible risks 

Informed consent signature 

Screening number (e.g. 
, etc.) 

Demographic data and life style recording 

Medical/surgical history 

Previous/concomitant medications 

Full physical examination (body weight, height, vital 
signs, physical abnormalities) 

ECG recording 

Clinical laboratory analyses: haematology, blood 
chemistry, urinalysis, serum virology, bacteriology and 
pregnancy test (women) 

Urine multi-drug kit test 

Adverse event (AE) monitoring 

Inclusion/exclusion criteria evaluation 

Eligibility evaluation 
The subject screening 
numbers will be in 
running 
order 
(e.g. 
 
, 
etc.). 
CCI
CCI


CONFIDENTIAL 
Sponsor code Z7244J01 
NAC 300 mg/3 mL i.v. bioavailability 
Final version 2.0, 19Dec2018 
Page 7 
STUDY SYNOPSIS (cont.) 
Schedule (continued):
Day 
Procedures/Assessments 
Notes 
Visit 2
Day -1 

Alcohol breath test 

Urine multi-drug kit test 

Urine pregnancy test (women) 

Inclusion/exclusion criteria evaluation 

Eligibility evaluation 

Enrolment 

Subject study number will be the screening number if 
successfully enrolled (e.g. 
, etc.) 

Check of AEs and concomitant medications  
Arrival 
at 
the 
clinic 
before 
the 
evening 
(before 18:30)  
Confinement until the 
morning of day 8 
Standardised dinner 
Fasting for at least 10 h 
(overnight) before first 
dosing
Visit 3
Day 1  

Vital sign measurement at pre-dose 

IMP administration at 08:00 ± 1 h; intravenous infusion 
for 5 min 

Vital signs measurement at 1 h post-dose 

Blood sample collection for pharmacokinetic analysis 
at: pre-dose (0) and 5 (at the end of the infusion), 8, 12, 
15, 20, 25, 30, 60 min, 2, 4, 6, 8, 10 and 12 h post-dose 

Urine sample collection for pharmacokinetic analysis 
at: 0-4; 4-8; 8-12 and 12-24 h post-dose 

Check of AEs and concomitant medications  
Standardized lunch at 
13:00  
standardized dinner at 
21:00  
Day 2 

Blood sample collection for pharmacokinetic analysis 
at 24 and 32 h post-dose 

Urine sample collection for pharmacokinetic analysis at 
24-32 h post-dose 

Check of AEs and concomitant medications 
Standardized breakfast 
around 9:00  
Standardized 
lunch 
around 13:00  
Standardized 
dinner 
around 21:00 
Day 3 

Vital sign measurement at 48 h post-dose 

Physical 
examination 
(body 
weight, 
physical 
abnormalities) at 48 h post-dose 

Clinical laboratory analyses: haematology, blood 
chemistry, urinalysis at 48 h post-dose  

Check of AEs and concomitant medications 
Standardized breakfast 
around 9:00  
Standardized 
lunch 
around 13:00  
Standardized 
dinner 
around 21:00
Visit 4
From day 4 to 
day 5 

IMP administration at 08:00 ± 1 h; infusion for 5 min 

IMP administration at 20:00 ± 1 h; infusion for 5 min 

Blood sample collection for pharmacokinetic analysis 
at: pre-dose (0) before the morning infusion  

Check of AEs and concomitant medications 
Standardized breakfast 
around 9:00  
Standardized 
lunch 
around 13:00  
Standardized 
dinner 
around 21:00 
CCI


CONFIDENTIAL 
Sponsor code Z7244J01 
NAC 300 mg/3 mL i.v. bioavailability 
Final version 2.0, 19Dec2018 
Page 8 
STUDY SYNOPSIS (cont.) 
Schedule (continued):
Day 
Procedures/Assessments 
Notes 
Visit 5
Day 6 

Vital sign measurement at pre-dose 

IMP administration at 08:00 ± 1 h; infusion for 5 min 

Vital signs measurement at 1 h post-dose 

Blood sample collection for pharmacokinetic analysis 
at: pre-dose (0) and 5 (at the end of the infusion), 8, 12, 
15, 20, 25, 30, 60 min, 2, 4, 6, 8, 10 and 12 h post-dose 

Urine sample collection for pharmacokinetic analysis 
at: 0-4; 4-8; 8-12 and 12-24 h post-dose 

Check of AEs and concomitant medications  
Standardized lunch at 
13:00 (about 5 h post-
dose) 
Standardized dinner at 
21:00 (about 13 h post-
dose)  
Day 7 

Blood sample collection for pharmacokinetic analysis 
at 24 and 32 h post-dose 

Urine sample collection for pharmacokinetic analysis at 
24-32 h post-dose 

Check of AEs and concomitant medications 
Standardized breakfast 
around 9:00  
Standardized 
lunch 
around 13:00  
Standardized 
dinner 
around 21:00 
Final Visit/ETV
Day 8 or early 
termination visit 
(ETV) in case of 
discontinuation 

Vital sign measurement at 48 h post-dose (or at ETV) 

Physical 
examination 
(body 
weight, 
physical 
abnormalities) 

Clinical laboratory analyses: haematology, blood 
chemistry, urinalysis  

Check of AEs and concomitant medications 
In case of clinically significant results at the final visit, the 
subjects will be followed-up by the investigator until the 
normalisation of the concerned clinical parameter(s) 
Discharge 
from 
the 
clinical centre in the 
morning of Day 8 after 
vital sign measurement, 
blood 
sampling 
for 
clinical laboratory assays 
and physical examination 
or in case of ETV. Upon 
leaving, the subjects will 
be instructed to contact 
immediately 
the 
investigator in case of 
occurrence 
of 
any 
untoward 
medical 
occurrence
Life style and constraints: 
During the study, the subjects will be confined from the evening preceding the first drug administration (study day 
-1) until the morning of day 8 (after the final visit). 
The subjects will not take any food or drinks (except water) for at least 10 h (i.e. overnight) before the first drug 
administration (Day 1) and will remain fasted up to 5 h post-dose.  
During the study, water will be allowed as desired. Coffee, tea or food containing xanthines (i.e. coffee, tea, soda, 
coffee with milk, energy drinks coke, chocolate, etc.), alcohol and grapefruit will be forbidden starting 48 h prior 
to the enrolment until the end of the study. Smoking is not allowed for the whole study duration. 


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STUDY SYNOPSIS (cont.) 
Withdrawal of subjects: 
It will be documented whether or not each subject completes the clinical study. In case of premature 
discontinuation of any subject, the primary reason for discontinuation will be recorded. 

Adverse event: Any (significant) adverse event that in the opinion of the investigator or concerned subject is 
not compatible with study continuation 

death: the absence of life or state of being dead 

lost to follow-up: the loss or lack of continuation of a subject up to follow-up 

non-compliance with study drug: an indication that a subject has not agreed with or followed the instructions 
related to the study medication 

physician decision: a position, opinion or judgment reached after consideration by a physician with 
reference to the subject 

pregnancy: pregnancy is the state or condition of having a developing embryo or fetus in the body (uterus), 
after union of an ovum and spermatozoon, during the period from conception to birth 

protocol deviation: an event or decision that stands in contrast to the guidelines set out by the protocol 

study terminated by sponsor: an indication that a clinical study was stopped by its sponsor 

technical problems: a problem with some technical aspect of a clinical study, usually related to an 
instrument 

withdrawal by subject: study discontinuation requested by a subject for whatever reason 
For any subject discontinuing the study, the investigator will ask the subject to undergo, as far as possible, a final 
medical visit (ETV) to examine the subject's health conditions and perform the required blood sampling for the 
laboratory assays. This examination will verify that all values tested at screening have remained within a clinically 
acceptable range (i.e. not clinically significant changes compared to screening) and report in the case report form 
(CRF) date and time of the last dose administration, and date and primary reason of study discontinuation.
Data analysis:  
The data documented in this trial and the measured clinical parameters will be presented using classic descriptive 
statistics (i.e. total number of subjects treated [N], number of observations [n], mean standard deviation [SD], 
minimum [Min], median, maximum [Max]) for quantitative variables and frequencies (i.e. count and percentages) 
for qualitative variables if not stated otherwise. 
Analysis set:  
Enrolled set: all enrolled subjects. This analysis set will be used for demographic, baseline and background 
characteristics 
Safety set: all subjects who receive at least one dose of IMP. This analysis set will be used for the safety analyses
PK set: all enrolled subjects who fulfil the study protocol requirements in terms of IMP administration and have 
evaluable pharmacokinetic data readouts for the planned analysis, with no major deviations that may affect the 
pharmacokinetic results. This analysis set will be used for the statistical analysis of the pharmacokinetic results 
Safety Assessments: 
The statistical analysis of demographic and safety data will be performed using the analysis software SAS®   . 
The toxicity grading of laboratory tests is determined based on the NCI CTCAE V4.03. All AEs, adverse drug 
reactions and serious adverse events will be summarised by system organ class, severity and relationship to study 
drug. 
Pharmacokinetics: 
The statistical analysis of PK parameters will be performed using the validated software Phoenix® WinNonlin®
version 6.3 or higher (Certara USA, Inc., Princeton, NJ) (actual version will be stated in the final report).   
The individual plasma/urine concentration and pharmacokinetic parameters will be presented in listings and their 
descriptive statistics summarised in tables.


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STUDY SYNOPSIS (cont.) 
Background:  
N-acetyl-L-cysteine (NAC) is the N-acetyl derivative of the naturally occurring amino acid L-cysteine.  
NAC was introduced into the Italian pharmaceutical market in 1965 and it is now manufactured and marketed by 
Zambon S.p.A. in Europe, Asia, South America, Central America, and the Middle East, under various trade names 
(Ventresca 1989). 
NAC is widely known as a mucolytic agent, with a well-established use in the treatment of respiratory tract 
disorders. However, after more than 35 years from its introduction in the clinical practice, this molecule continues 
to be the object of scientific interest due to its potential in the treatment of several conditions other than those 
related to the respiratory system. NAC can also act as a direct antioxidant agent due to the -SH group (Auroma 
1988) and can easily penetrate into the cells where it is deacetylated to L-cysteine, thus supporting the biosynthesis 
of glutathione (Ziment 1988). 
Clinical pharmacology and pharmacokinetics 
Several pharmacokinetic (PK) studies were carried out in healthy volunteers (Borgstrom 1986, Olsson 1988, 
Burgunder 1989, De Caro 1989, Borgstrom 1990, Frascio, Crestani 2002, Rusca 2014, Rusca 2010, Rusca 2002). 
Further PK studies were carried out in patients with chronic liver damage (Jones 1997), with liver damage caused 
by paracetamol overdose (Prescott 1989), in subjects under long-term treatment for chemoprevention (Pendyala 
1995), in patients with respiratory disorders (Rodenstein 1978) and in patients with End Stage Renal Diseases 
(Internal report 2002). 
In the clinical studies, NAC was administered orally and intravenously both as single and repeated dose. Oral NAC 
was administered in different dosage forms, such as capsules, granulate, effervescent tablets, fast dissolving tablets, 
slow-release tablets. 
Clinical trials and safety concerns 
NAC has been available to patients as a mucolytic agent for many years and has a well-established and satisfactory 
safety and tolerability profile (see IB).  
Independently of route of administration (inhalation, oral, intravenous) adverse events are reported rarely 
following NAC administration and are usually mild and transient.  
The most frequent adverse events associated with NAC oral administration are gastrointestinal (e.g., vomiting, 
diarrhoea, stomatitis, abdominal pain and nausea). Hypersensitivity reactions (e.g. anaphylactoid reactions, 
urticaria, rash, pruritus, and bronchospasm) have been reported less frequently after oral administration, while with 
the IV administration they were reported more frequently compared to gastrointestinal symptoms. Some of the 
hypersensitivity reactions were associated to higher infusion rate. Most anaphylactoid reactions can be managed 
by temporarily suspending the acetylcysteine infusion, administering appropriate supportive care. A cumulative 
analysis of all safety reports received by the Sponsor up to September 2017 for NAC IV, confirmed the nature and 
severity of the events were in line with the above profile of the product. 
Intravenous NAC has been studied in clinical trials and published case series both as mucolytic agent and as an 
antidote against intoxication by paracetamol and other toxic agents including Amanita phalloides and chlorinated 
hydrocarbons. Overall, in published studies in humans, a total of 745 patients were exposed to i.v. NAC at doses 
ranging from 500 mg/day to 980mg/kg/day for a duration of 1 to 18 days.  Details of safety results are included in 
the IB. 
In published studies, whereas i.v. NAC as mucolytic agent typically was given at doses up to 1000 mg/day, much 
higher i.v. doses of NAC as antidote were given over multiple days, maintaining a satisfactory safety profile. 
Locatelli and colleagues published two case series, one of 86 patients with acute intoxication from Amanita 
phalloides and the other of 35 patients with acute intoxication from chlorinated hydrocarbons. Patients received 
i.v. NAC 300 mg/kg/day for 3 to 14 days and 3 to 18 days respectively. The safety and tolerability profile were 
overall benign, with 4 out of 121 exposed patients suffering from anaphylactoid reaction or rash (due to the rapid 
administration of the bolus according to the authors), none of which caused discontinuation of treatment. No other 
AEs were reported. (Locatelli 1996).
Rationale 
NAC is widely known as a mucolytic agent, with a well-established use in the treatment of respiratory tract 
disorders. In the present study, the kinetic profile and the bioavailability of NAC will be investigated in Chinese 
healthy male and female volunteers all receiving the same single i.v. dose, i.e. a total dose of 600 mg of NAC, and 
the same multiple dose administrations, i.e. five i.v. doses of 600 mg of NAC administered b.i.d. for 2 days and 
once on the last day. The safety and tolerability will be monitored as well.  
The present study will be part of the clinical development plan of NAC i.v. for registration in China of this 
formulation.


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3
STUDY SCHEDULE 
ACTIVITIES 
Screening 
Single Dose 
Multiple Dose 
Final 
visit/ETV1
Visit 
V1 
V2 
V3 
V4 
V5 
Day -14/-2 
Day -1 
Day 1 
Day 2 
Day 3 
Day 4 
Day 5 
Day 6 
Day 7 
Day 82
Informed consent 
x
Demography and lifestyle 
x
Medical and surgical history  
x
Physical examination 
x
x
x
Prior and concomitant medications 
x
x
x
x
x
x
x
x
x
x
Height 
x
Body Weight 
x
x
x
Clinical laboratory analysis 
(haematology, blood chemistry, 
urinalysis) 
x 
x 
x
Virology and bacteriology  
x
Serum pregnancy test (women) 
x
Urine multi-drug kit test 
x
x
Blood pressure and heart rate 
x
x3
x
x3
x4
Alcohol breath test 
x
Urine pregnancy test (women)
x
ECG 
X
Inclusion/exclusion criteria 
X
x
Subject eligibility
X
x
Enrolment 
x
Confinement 
x
x
x
x
x
x
x
x
Discharge 
x
Investigational product administration
x5
x6
x6
x5
Blood sampling  
x7
x7
x8
x8
x7
x7
Urine sampling  
x9
x9
x9
x9
Standardised meals 
x10
x11
x12
x12
x12
x12
x11
x12
Adverse event monitoring13
x
x
x
x
x
x
x
x
x
x


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1.
Early termination visit (ETV) 
2.
Final visit on day 8 or in case of ETV 
3.
At pre-dose and 1 h post-dose 
4.
The vital signs check at 48 h post-dose (day 8), will correspond to the final measurement 
5.
At 8:00 ± 1 h: 5-min infusion 
6.
At 8:00 ± 1 h and at 20:00 ± 1 h; 5-min infusion 
7.
At pre-dose (0) and 5 (at the end of the infusion), 8, 12, 15, 20, 25, 30, 60 min and 2, 4, 6, 8, 10, 12, 24 and 32 h post-dose  
8.
At pre-dose (0) 
9.
0-4; 4-8; 8-12, 12-24 and 24-32 h post-dose  
10. Standardised dinner 
11. Standardised lunch and dinner 
12. Standardised breakfast, lunch and dinner  
13. Adverse events monitored starting at the screening visit, immediately after informed consent, up to the final visit/ETV 


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4
TABLE OF CONTENTS 
Page 
1 
CLINICAL STUDY PROTOCOL 
1
2
STUDY SYNOPSIS 
4
3
STUDY SCHEDULE 
11
4
TABLE OF CONTENTS 
13
4.1
TABLES 
15
5
LIST OF ABBREVIATIONS 
16
6
Introduction 
18
6.1
Background 
18
6.2
Clinical pharmacology and pharmacokinetics 
18
6.3
Clinical trials and safety concerns 
19
6.4
Rationale 
20
6.5
Risk and benefits 
20
7
STUDY OBJECTIVES 
21
7.1
Primary end-point 
21
7.2
Secondary end-point 
21
8
CLINICAL SUPPLIES 
22
8.1
Treatment 
22
8.1.1
Description of products 
22
8.1.1.1
Test product 
22
8.1.2
Dose regimen 
23
8.1.3
Route and method of administration 
23
8.1.4
Investigational product distribution 
23
8.2
Packaging and labelling 
23
8.3
Storage conditions 
24
8.4
Drug accountability 
24
9
INVESTIGATIONAL PLAN 
25
9.1
Overall study design 
25
9.2
Discussion of design 
25
10
STUDY POPULATION 
26
10.1
Target population 
26
10.2
Inclusion criteria 
26
10.3
Exclusion criteria 
27
10.3.1
Not allowed treatments 
28
11
STUDY SCHEDULE 
29
11.1
Study visits and procedures 
29
11.2
Diet and lifestyle 
31
11.2.1
Restrictions 
32
12
DESCRIPTION OF SPECIFIC PROCEDURES 
33
12.1
Physical examination 
33
12.1.1
Body weight 
33
12.1.2
Vital signs 
33
12.1.3
ECGs 
33
12.2
Clinical laboratory assays 
33
12.3
Sampling for pharmacokinetic analysis 
34
12.3.1
Venous blood sampling 
34
12.3.2
Urine collection 
35
12.3.3
Analytics 
36
12.3.4
Labelling, storage and transport of samples 
36
12.3.4.1
Samples labelling 
36
12.3.4.2
Samples storage and transport 
36
13
ASSIGNMENT OF STUDY TREATMENT 
38
13.1
Randomisation 
38


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13.2
Blinding 
38
14
EVALUATION PARAMETERS 
39
14.1
Study variables 
39
14.1.1
Primary variables 
39
14.1.2
Secondary variables 
39
14.2
Pharmacokinetic assessments 
39
14.2.1
Pharmacokinetic parameters 
39
14.3
Safety assessments 
41
15
STATISTICAL METHODS 
42
15.1
Analysis Sets 
42
15.1.1
Definitions 
42
15.1.2
Reasons for exclusion from the PK set before bioanalysis 
43
15.2
Sample size and power considerations 
43
15.3
Demographic, baseline and background characteristics 
43
15.4
Analysis of pharmacokinetic parameters 
43
15.4.1
Descriptive pharmacokinetics 
43
15.5
Safety and tolerability evaluation 
43
16
DEFINITION AND HANDLING OF AEs AND SAEs 
45
16.1
Applicable SOPs 
45
16.2
Definition of Adverse Event (AE) 
45
16.3
Definition of Adverse Drug Reaction (ADR) 
45
16.4
Definition of Serious Adverse Events or Serious Adverse Drug Reaction 
46
16.5
Definition of Severity of Adverse Events 
46
16.6
Definition of Adverse Event causality 
46
16.7
Adverse Events recording 
47
16.8
AEs monitoring window 
47
16.9
Adverse Events reporting 
47
16.9.1
SAEs reporting 
48
16.10
Follow-up for Adverse Events 
48
16.11
SUSARs management 
49
16.12
Other events qualified for expedited reporting 
49
16.13
SAEs: contacts 
50
16.14
Pregnancy 
50
17
DATA MANAGEMENT PROCEDURES 
51
17.1
Data collection – CRFs 
51
17.2
Database management 
51
17.2.1
Coding dictionaries 
51
18
STUDY MONITORING, QUALITY CONTROL AND QUALITY ASSURANCE 
52
18.1
Monitoring 
52
18.2
Quality Control and Quality Assurance 
52
18.3
Applicable SOPs 
53
18.4
Data access 
53
18.5
Audits and inspections 
53
19
ETHICAL CONSIDERATIONS 
54
19.1
Ethics and Good Clinical Practice (GCP) 
54
19.2
Informed consent 
54
19.3
Insurance policy 
55
19.4
Withdrawal of subjects 
55
19.4.1
Primary reason for discontinuation 
55
19.5
Study termination 
56
20
ADMINISTRATIVE PROCEDURES 
57
20.1
Material supplied to the clinical centre 
57
20.2
Protocol amendments 
57
20.3
Study documentation and record keeping 
57
20.4
Study subjects’ recruitment 
58
20.5
Confidentiality and data protection 
58
20.6
Publication policy 
58
21
STUDY RESPONSIBLE PERSONS 
60


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21.1
Sponsor 
60
21.2
Drug assay 
60
21.3
Project management, data analysis & reporting, monitoring 
60
22
REFERENCES 
61
4.1
TABLES 
Page 
Table 11.1.1
Schedule of study activities and procedures 
30
Table 12.3.1.1
Tolerance ranges for the scheduled sampling times 
35


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5
LIST OF ABBREVIATIONS 
β-HCG
human chorionic gonadotropin β
γ-GT
γ-Glutamyl transpeptidase
ADR
Adverse Drug Reaction
AE
Adverse Event
ALT
Alanine aminotransferase
Ae0-t
Total amount of NAC excreted in urine 
Aess(0-t)
Total amount of NAC excreted in urine at steady-state
ANOVA
Analysis of Variance
AST
Aspartate aminotransferase
BB
Blue book
BLQL
Below Lower Quantification Limit
BUN
Blood Urea Nitrogen
AUC(0-t)
Area under the concentration-time curve from single dose to the last observed concentration 
time t
AUC(0-∞)
Area under the concentration-time curve extrapolated to infinity 
AUC(0-12)
Area under the concentration-time curve at steady-state in the tau interval 
AUCss(0-t)
Area under the concentration-time curve at steady-state from the last multiple dose to the last 
observed concentration time t
AUCss(0-12)
Area under the concentration-time curve at steady-state in the tau interval 
BMI
Body Mass Index
BP
Blood Pressure
CA
Competent Authority
CDISC
Clinical Data Interchange Standards Consortium
CFDA
China Food And Drug Administration
CI
Confidence Interval
Cmax
Peak drug concentration
CLt
Total body clearance
CLr
Renal clearance
CPL
Clinical Project Leader
CRA
Clinical Research Associate
CRF
Case Report Form
CRO
Contract Research Organisation
CS
Clinically Significant
CSP
Clinical Study Protocol
CSR
Clinical Study Report
Css_av
Average NAC plasma concentration at steady-state
Css_max
Maximum plasma concentration at steady-state
Css_min
Trough plasma concentration at steady-state
CV
Coefficient of Variation
DBP
Diastolic Blood Pressure
DF
Degree of fluctuation 
DSU
Drug safety unit
DTT
Dithiothreitol
EC
Ethics Committee
ECG
Electrocardiogram
ETV
Early Termination Visit
FDA
Food and Drug Administration
Fe0-t
Total fraction of NAC dose excreted in urine 
Frel
Relative Bioavailability
FSFV
First Subject First Visit
GCP
Good Clinical Practice
GLP
Good Laboratory Practice
HBs Ag
Hepatitis B virus surface antigen
HCV Ab
Hepatitis C virus antibodies
HIV
Human Immunodeficiency Virus


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HR
Heart Rate
IB
Investigator’s Brochure
ICH
International Conference on Harmonisation
IRB/IEC
Institutional Review Board/Independent Ethics Committee
i.m.
Intramuscular
IMP
Investigational Medicinal Product
IUD
Intra-Uterine Device
i.v.
Intravenous
kel
Terminal elimination rate constant
LQL
Lower Quantification Limit
LSLV
Last Subject Last Visit
MCH
Mean Cell Haemoglobin
MCHC
Mean Cell Haemoglobin Concentration
MCV
Mean Cell Volume
MedDRA
Medical Dictionary for Regulatory Activities
MRT
Mean Residence Time
MW
Molecular Weight
N
Normal
NA
Not Applicable
NAC
N-acetyl-L-cysteine
NC
Not calculated
NCS
Not clinically significant
NSAIDs
Non-Steroidal Anti-Inflammatory Drugs
OTC
Over The Counter
PE
Point Estimate
PK
Pharmacokinetics
PT
Preferred Term
PTAE
Pre-Treatment Adverse Event
R
Accumulation ratio
RBC
Red Blood Cells
RSI
Reference safety information
SAE
Serious Adverse Event
SBP
Systolic Blood Pressure
SD
Standard Deviation
SOC
System Organ Class
SOP
Standard Operating Procedure
SDTM
Study Data Tabulation Model
SUSAR
Suspected Unexpected Serious Adverse Reaction
T
Test
TEAE
Treatment-Emergent Adverse Event
THC
delta-9-tetrahydrocannabinol
t1/2
Half-life
tmax
Time to achieve Cmax
tss_max
Time to achieve Css_max
USDA
United States Department of Agriculture
Vd
Volume of distribution 
WBC
White Blood Cells
WHODDE
World Health Organisation Drug Dictionary Enhanced


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6
INTRODUCTION 
6.1
Background 
N-acetyl-L-cysteine (NAC) is the N-acetyl derivative of the naturally occurring amino acid L-
cysteine.  
NAC was introduced into the Italian pharmaceutical market in 1965 and it is now manufactured 
and marketed by Zambon S.p.A. in Europe, Asia, South America, Central America, and the 
Middle East, under various trade names (1). 
NAC is widely known as a mucolytic agent, with a well-established use in the treatment of 
respiratory tract disorders. However, after more than 35 years from its introduction in the 
clinical practice, this molecule continues to be the object of scientific interest due to its potential 
in the treatment of several conditions other than those related to the respiratory system. NAC 
can also act as a direct antioxidant agent due to the -SH group (2) and can easily penetrate into 
the cells where it is deacetylated to L-cysteine, thus supporting the biosynthesis of glutathione 
(3). 
6.2
Clinical pharmacology and pharmacokinetics 
Several pharmacokinetic (PK) studies were carried out in healthy volunteers (4, 5, 6, 7, 8, 9, 
10, 11, 12, 13, 14). Further PK studies were carried out in patients with chronic liver damage 
(15), with liver damage caused by paracetamol overdosage (16), in subjects under long-term 
treatment for chemoprevention (17), in patients with respiratory disorders (18) and in patients 
with End Stage Renal Diseases (19). 
In the clinical studies, NAC was administered orally and intravenously both as single and 
repeated doses. Oral NAC was administered in different dosage forms, such as capsules, 
granulate, effervescent tablets, fast dissolving tablets, slow-release tablets. 
In particular, Borgström et al. (4) studied for the first time the PK profile of NAC after i.v. dose 
of 600 mg of NAC infused over 5 min in 10 healthy male and female volunteers in comparison 
with 3 other oral dosage forms. The PK parameters obtained by the authors after i.v. infusion 
are summarised in the following table.  
Table 6.2.1 
PK parameters of plasma free NAC (N=10) 
Cmax
(µmol/L) 
tmax
(h) 
CL 
(L/kgxh) 
CLr
(L/kgxh) 
t½
(h) 
Ae0-12
(% of dose) 
16.0±7.9 
0.65±0.33 
0.207±0.017
0.058±0.011 
2.27±0.32 
29.0±3.2* 
mean±SD is reported; *: N=9; Source: 4
However, the authors analysed NAC in deproteinised plasma thus missing the protein-bound 
NAC in their measurements.  
Olsson et al. (5) improved the bioanalytical method and were able to measure the total NAC by 
reduction of the disulphide bonds in plasma before precipitating proteins. After single i.v. dose 
of 200 mg of NAC to 6 healthy volunteers, the authors found the following PK parameters: 


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Table 6.2.2 
PK parameters of plasma total NAC (N=6) 
Cmax
(µM) 
Vss
(L/kg) 
CL 
(L/kgxh) 
T
121 (82.9 – 162) 
0.47 (0.46 – 0.55) 
0.11 (0.09 – 1.13)
median (range) is reported; Source: 5
Jones et al. (15) studied the bioavailability of NAC after single i.v. dose of 600 mg of NAC 
infused over 3 min to patients with chronic liver disease, but also to 6 healthy male and female 
volunteers as control subjects. The authors found the following PK parameters for total NAC: 
Table 6.2.3 
PK parameters of plasma total NAC (N=6) 
AUC 
(mg/Lxh) 
CLr
(L/h) 
t½
(h) 
Vdss
(L) 
93.9±9.6
6.5±0.8 
2.6±0.3 
17.4±2.8 
mean±SD is reported; Source: 15
Brown et al. (14) investigated the PK of NAC infused i.v. to 24 healthy men at rest and during 
exercise. NAC was infused at the dose of 125 mg/kgxh for 15 min followed by 25 mg/kgxh for 
35 min to healthy men at rest. Then, the infusion continued during exercise until fatigue. The 
authors found NAC peak concentrations in plasma at the end of the initial loading infusion. 
Mean peak concentration of total NAC was 205.1±68.3 mg/L, while mean CL was 0.164 
L/kgxh.  
It is worth noting that, while scientific literature shows that PK profiles of NAC after oral 
administration are very consistent (in terms of half-life, rate and extent of exposure, 
bioavailability, dose-linearity, etc.) among various publications, this is not the case for the data 
obtained after i.v. dosing. In fact, for this administration route, published studies differ in 
several key aspects, i.e., for example, type of administration, applied bioanalytical methods, 
drug doses, calculation methods for PK parameters. Furthermore, the number of subjects 
enrolled in each published study is very small. For these reasons, the comparability of the 
literature data is low among the published studies and potentially also to the expected results of 
the present study.   
6.3
Clinical trials and safety concerns 
NAC has been available to patients as a mucolytic agent for many years and has a well-
established and satisfactory safety and tolerability profile (see IB).  
Independently of route of administration (inhalation, oral, intravenous) adverse events are 
reported rarely following NAC administration and are usually mild and transient.  
The most frequent adverse events associated with NAC oral administration are gastrointestinal 
(e.g., vomiting, diarrhoea, stomatitis, abdominal pain and nausea). Hypersensitivity reactions 
(e.g. anaphylactoid reactions, urticaria, rash, pruritus, and bronchospasm) have been reported 
less frequently after oral administration, while with the i.v. administration they were reported 
more frequently compared to gastrointestinal symptoms. Some of the hypersensitivity reactions 
were associated to higher infusion rate. Most anaphylactoid reactions can be managed by 
temporarily suspending the acetylcysteine infusion, administering appropriate supportive care. 


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Sponsor code Z7244J01 
NAC 300 mg/3 mL i.v. bioavailability 
Final version 2.0, 19Dec2018 
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A cumulative analysis of all safety reports received by the Sponsor up to September 2017 for 
NAC IV, confirmed the nature and severity of the events were in line with the above profile of 
the product. 
Intravenous NAC has been studied in clinical trials and published case series both as mucolytic 
agent and as an antidote against intoxication by paracetamol and other toxic agents including 
Amanita phalloides and chlorinated hydrocarbons. Overall, in published studies in humans, a 
total of 745 patients were exposed to i.v. NAC at doses ranging from 500 mg/day to 
980mg/kg/day for a duration of 1 to 18 days.  Details of safety results are included in the IB. 
In published studies, whereas i.v. NAC as mucolytic agent typically was given at doses up to 
1000 mg/day, much higher i.v. doses of NAC as antidote were given over multiple days, 
maintaining a satisfactory safety profile. 
Locatelli and colleagues published two case series, one of 86 patients with acute intoxication 
from Amanita phalloides and the other of 35 patients with acute intoxication from chlorinated 
hydrocarbons. Patients received i.v. NAC 300 mg/kg/day for 3 to 14 days and 3 to 18 days 
respectively. The safety and tolerability profile were overall benign, with 4 out of 121 exposed 
patients suffering from anaphylactoid reaction or rash (due to the rapid administration of the 
bolus according to the authors), none of which caused discontinuation of treatment. No other 
AEs were reported (21). 
6.4
Rationale 
NAC is widely known as a mucolytic agent, with a well-established use in the treatment of 
respiratory tract disorders. 
In the present study, the kinetic profile and the bioavailability of NAC will be investigated in 
Chinese healthy male and female volunteers all receiving the same single i.v. dose, i.e. a total 
dose of 600 mg of NAC, and the same multiple dose administrations, i.e. five i.v. doses of 600 
mg of NAC administered b.i.d. for 2 days and once on the last day. The safety and tolerability 
will be monitored as well.  
The present study will be part of the clinical development plan of NAC i.v. for registration in 
China of this formulation. 
6.5
Risk and benefits 
NAC is a well-known drug which has been used for decades.  
Undesired effects which may occur during treatment with NAC include: anaphylactic shock, 
anaphylactic reaction, anaphylactoid reaction, hypersensitivity, tachycardia, bronchospasm, 
dyspnoea, vomiting, nausea, angioedema, urticaria, flushing rash, pruritus, face oedema, blood 
pressure decreased, prothrombin time prolonged (for details refer to IB; 22).  
Blood sampling with cannula insertion may cause minor discomfort. The risks associated with 
blood draws include pain, bleeding and bruising. 
No specific benefits for the participants in the current study are foreseen. Their remuneration 
will be paid after study completion. The remuneration covers loss of time and any 
inconvenience caused by the participation in the study. 


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Sponsor code Z7244J01 
NAC 300 mg/3 mL i.v. bioavailability 
Final version 2.0, 19Dec2018 
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7
STUDY OBJECTIVES 
The objective of the study is to evaluate the NAC pharmacokinetics, safety and tolerability after 
single and multiple dose i.v. administration. 
7.1
Primary end-point 
To evaluate pharmacokinetic parameters of NAC in plasma after single and multiple dose 
administration of the investigational product.  
7.2
Secondary end-point 
To collect safety and tolerability data after single and multiple dose administration of the 
investigational product. 


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Sponsor code Z7244J01 
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Final version 2.0, 19Dec2018 
Page 22 
8
CLINICAL SUPPLIES 
8.1
Treatment 
All the subjects enrolled in the study will receive the same treatment with the investigational 
medicinal product (IMP), i.e. NAC, 300 mg/ 3 mL solution for injection, as follows: 
on day 1 at 08:00 ±1 h, one dose of 600 mg of NAC (300 + 300 mg ampoule) will be 
administered under fasting conditions; 
After a wash-out of 3 days: 
on days 4 and 5 at 08:00 ±1 h and 20:00 ±1 h and at 08:00 ±1 on day 6, 5 doses of 600 mg 
of NAC (300 + 300 mg ampoule) will be administered. 
8.1.1
Description of products 
8.1.1.1
Test product 
IMP
NAC 300 mg/ 3 mL solution for injection
Active substance
N-acetyl-L-cysteine
Manufacturer 
(active substance) 
F.I.S. Fabbrica Italiana Sintetici S.p.A.,  
Via Dovaro, 36045 Lonigo (Vicenza), Italy 
(GMP compliant)
Manufacturer 
(finished product) 
Zambon S.p.A.,  
Via della Chimica 9, 36100 Vicenza, Italy 
(GMP compliant)
Pharmaceutical form
Solution for injection
Dose
300 mg/ 3 mL
Administration route
Parenteral 
The analytical certificates will be supplied with the IMP. Quali-quantitative formulation is as 
follows: 
Each 3 mL vial (10%) contains: 
N-acetyl-L-cysteine (NAC) 300 mg 
sodium hydroxide 74 mg 
disodium edetate 3 mg 
water for injections q.s. to 3 mL 


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8.1.2
Dose regimen 
Two ampoules of IMP (300 + 300 mg) corresponding to a total dose of 600 mg of NAC diluted 
in 10 mL of NaCl 0.9% sterile saline solution, will be administered by a 5-minute i.v. infusion. 
Single dose: 
One (1) dose of IMP will be administered under fasting conditions on day 1 at 08:00 ±1 h. 
Multiple dose regime: 
Five (5) doses of IMP will be administered twice a day (b.i.d.) on days 4 and 5 at 08:00 ±1 h 
and 20:00 ±1 h and one dose will be administered on day 6 at 08:00 ±1.  
8.1.3
Route and method of administration 
Each dose of IMP will be prepared by diluting the content of 2 ampoules of IMP in 10 mL of 
saline, as described above (§ 8.1.2), and administered to each study subject at the clinical site 
only by the investigator or his/her deputy. 
Each dose will be infused intravenously over 5 min.  
The start of infusion (not the end) will be considered as time 0. 
8.1.4
Investigational product distribution 
The IMP will be exclusively used for the present clinical study and will only be administered 
to the subjects enrolled in the study.  
8.2
Packaging and labelling 
The IMP primary packaging will be glass vials. 
The formulation labelling will report all the information requested according to the Annex 13 
to the Good Manufacturing Practice (published by the Commission in The rules governing 
medicinal products in the European Community, Volume 4; 23) and in compliance with 
applicable laws and regulations as follows: 
a.
Name, address and telephone number of the sponsor, contract research organisation or 
investigator (the main contact for information on the product, clinical study and any 
emergency) 
b.
Pharmaceutical dosage form, route of administration, quantity of dosage units, and the 
name and strength 
c.
The batch and/or code number to identify the contents and packaging operation 
d.
A study reference code allowing identification of the study, site, investigator and sponsor 
if not given elsewhere 


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e.
The study subject identification number/treatment number and where relevant, the visit 
number 
f.
The name of the investigator (if not included in (a) or (d)) 
g.
Directions for use (reference may be made to a leaflet or other explanatory document 
intended for the study subject or person administering the product) 
h.
“For clinical study use only” or similar wording 
i.
The storage conditions 
j.
Period of use (use-by date, expiry date or re-test date as applicable), in month/year format 
and in a manner that avoids any ambiguity 
k.
“Keep out of reach of children” 
Labels will be in local language. 
8.3
Storage conditions 
The IMP will be stored at ≤25° C in a dry locked place, sheltered from light.  
8.4
Drug accountability 
The IMP will be provided directly to the investigator by the Sponsor or designee, in excess of 
the amount necessary for the study (at least 25% excess). 
After receipt of the IMP supply, the pharmacist will confirm in writing by signing and dating 
standard drug accountability forms. 
At the end of the study, used, unused and partially used supplies of IMP provided by the Sponsor 
or designee will either be destroyed on site (upon written authorisation) or returned to the 
Sponsor or designee, after assessment of drug accountability. 


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9
INVESTIGATIONAL PLAN 
9.1
Overall study design 
This is a single and multiple dose, single centre, open-label, one-way, pharmacokinetics, safety 
and tolerability clinical trial of Phase I to be performed in Chinese healthy male and female 
volunteers. 
9.2
Discussion of design 
The study has been designed in agreement with the Chinese Technical Guideline on Clinical 
Pharmacokinetic Research of Chemical Drugs, 18 March 2005 and the European Guideline on 
the Investigation of Bioequivalence (CPMP/QWP/EWP/1401/98 Rev. 1, 20 January 2010) (24). 
An open-label design is used since the primary end-point of the study is based on objective 
measurements of NAC in blood. The outcome variables are not influenced by the subjects or 
investigator being aware of the administered products. 
Blood sampling time-points were selected on the basis of the known PK profile of NAC (4, 5, 
6, 7, 8, 9, 10, 11, 12, 13, 14). 
The dose of 600 mg has been selected since this is the efficacious, safe and well tolerated dose 
used in clinical practice (see also 22).  
The bioanalysis will be performed in compliance with GCP and CFDA GCP regulations and in 
accordance with the applicable principles of GLP, as defined by OECD, in a GLP compliant 
facility. Moreover, sample analysis will be conducted in compliance with the Chinese Guidance 
on Bioanalysis: method validation and analysis of study samples (2015).  


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10
STUDY POPULATION 
10.1
Target population 
Chinese healthy male and female volunteers will be included in the study.  
10.2
Inclusion criteria 
To be enrolled in this study, subjects must fulfil all these criteria: 
1. Informed consent: signed written informed consent before inclusion in the study 
2. Ethnicity, Sex and Age: Chinese males and females, 18-45 year old inclusive
3. Weight: body weight ≥50 kg;
4. Body Mass Index: 19-26 kg/m2 inclusive
5. Vital signs: systolic blood pressure 100-139 mmHg, diastolic blood pressure 50-89 
mmHg, heart rate 50-90 bpm, measured after 5 min at rest in the sitting/supine position (to 
be chosen according to the usual procedure at the clinical site) 
6. Full comprehension: ability to comprehend the full nature and purpose of the study, 
including possible risks and side effects; ability to co-operate with the investigator and to 
comply with the requirements of the entire study
7. Nicotine addiction (smoker subjects only): ability to abstain from smoking for the duration 
of the clinical study
8. Contraception and fertility (women only): women of child-bearing potential must be using 
at least one of the following reliable methods of contraception: 
a. Hormonal oral, implantable, transdermal, or injectable contraceptives for at least 60 
calendar days before the screening visit 
b. A non-hormonal intrauterine device or female condom with spermicide or 
contraceptive sponge with spermicide or diaphragm with spermicide or cervical cap 
with spermicide for at least 60 calendar days before the screening visit 
c. A male sexual partner who agrees to use a male condom with spermicide 
d. A sterile sexual partner 
Women of non-child-bearing potential or in post-menopausal status for at least 1 year will 
be admitted. 
Women of childbearing potential should be willing to adopt abstinence or contraception 
measures during the study and two weeks post-dose. 
For all women, pregnancy test result must be negative at screening and day -1. 


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10.3
Exclusion criteria 
Subjects meeting any of these criteria will not be enrolled in the study: 
1. Electrocardiogram (12-lead ECG in supine position): clinically significant abnormalities 
2. Physical findings: clinically significant abnormal physical findings which could interfere 
with the objectives of the study 
3. Laboratory analyses: clinically significant abnormal laboratory values indicative of 
physical illness, in particular significant laboratory abnormality indicative of hepatic 
condition (more than 3 times the upper limit) 
4. Allergy: ascertained or presumptive hypersensitivity to the active principle and/or 
formulations' ingredients; history of anaphylaxis to drugs or allergic reactions in general, 
which the investigator considers may affect the outcome of the study 
5. Diseases: significant history of renal, hepatic, gastrointestinal, cardiovascular, respiratory, 
skin, haematological, endocrine, urologic, metabolic, neurological or psychiatric diseases, 
as determined by the investigator, that may interfere with the aim of the study; history of 
carcinoma in situ and malignant disease; active bacterial or viral infection and fever >38° 
C within 48 h prior to study treatment administration 
6. Virology: positive result of HIV, hepatitis B (HBV), hepatitis C (HCV) or Treponema 
pallidum (TP) assays 
7. Surgery: any surgery within 60 calendar days of screening (excluding diagnostic surgery) 
8. Medications: medications, including over the counter (OTC) medications, herbal remedies 
and traditional Chinese remedies for 2 weeks before the start of the study. Hormonal 
contraceptives for women will be allowed 
9. Investigative drug studies: participation in the evaluation of any investigational product for 
1 month before this study 
10. Blood donation: blood donations for 90 calendar days before this study 
11. Drug, alcohol, caffeine, tobacco: history of drug, alcohol [>1 drink/day for females and >2 
drinks/day for males, defined according to the USDA Dietary Guidelines 2015-2020] 
caffeine (>5 cups coffee/tea/day) or tobacco abuse (≥10 cigarettes/day)
12. Abuse drug test: positive urine abuse drug test at screening or day -1
13. Alcohol test: positive alcohol breath test at day -1
14. Diet: abnormal diets (<1600 or >3500 kcal/day) or substantial changes in eating habits in 
the 4 weeks before this study; vegetarians; consumption of alcohol, grapefruit, products 
containing grapefruit, or beverages containing xanthines (coffee, tea, soda, coffee with 
milk, energy drinks) within 48 hours prior to the enrolment  
15. Pregnancy (females only): positive or missing pregnancy test at screening or day -1, 
pregnant or lactating women 
16. Vaccination within 4 weeks of study treatment 
17. Other unspecified reasons that, in the opinion of the investigator, make the subject 
unsuitable for enrolment. 


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10.3.1
Not allowed treatments 
No medication, including OTC, traditional Chinese medicine and herbal remedies, will be 
allowed for 2 weeks before the start of the study and during the whole study duration. 
Paracetamol will be allowed as therapeutic counter-measure for adverse events (AEs) according 
to the investigator’s opinion. Hormonal contraceptives will be allowed.  
The intake of any other medication will be reported as a protocol deviation.  


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11
STUDY SCHEDULE 
The schedule of the study is summarised at page 11. 
11.1
Study visits and procedures 
Each study subject will undergo 6 visits. 
The study protocol foresees a screening visit and a confinement lasting from Visit 2 to the Final 
Visit/early termination visit (ETV), i.e. from Day -1 to Day 8. Maximum study duration will be 
22 days, screening visit included. A written informed consent will be obtained before any study 
assessment or procedure. 
The first subject first visit (FSFV) is defined as the 1st visit performed at the clinical centre on 
the 1st screened subject. The last subject last visit (LSLV) is defined as the last visit performed 
at the clinical centre (or the telephonic follow-up, if applicable) on the last subject, i.e. the last 
visit foreseen by the study protocol, independently of the fact that the subject is a completer or 
a withdrawn subject. 
The following phases, visits and procedures will be performed: 
Screening phase 
Screening – visit 1: between day -14 and day -2 
Visit 2: day -1 
Interventional phase 
Visit 3: days 1-3 
Visit 4: days 4-5 
Visit 5: days 6-7 
Final phase 
Visit 6: day 8 - Final visit/ETV. In case of early discontinuation, discontinued subjects 
will undergo an ETV 
Activities to be performed are listed by visit in the following table:  


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Table 11.1.1 Schedule of study activities and procedures 
Day 
Procedures/Assessments 
Notes 
Screening – Visit 1
From day -14 to 
day -2 

Explanation to the subject of study aims, procedures 
and possible risks 

Informed consent signature 

Screening number (e.g. 
, etc.) 

Demographic data and life style recording 

Medical/surgical history 

Previous/concomitant medications 

Full physical examination (body weight, height, vital 
signs, physical abnormalities) 

ECG recording 

Clinical laboratory analyses: haematology, blood 
chemistry, urinalysis, serum virology, bacteriology and 
pregnancy test (women) 

Urine multi-drug kit test 

AE monitoring 

Inclusion/exclusion criteria evaluation 

Eligibility evaluation 
The subject screening 
numbers (e.g. 
 
, etc.) will be 
used to identify the 
subjects 
Visit 2
Day -1 

Alcohol breath test 

Urine multi-drug kit test 

Urine pregnancy test (women) 

Inclusion/exclusion criteria evaluation 

Eligibility evaluation 

Enrolment 

Study subject number will be the same as the screening 
number that was initially assigned when the subject is 
successfully enrolled (e.g screening number .) 

Check of AEs and concomitant medications  
Arrival 
at 
the 
clinic 
before 
the 
evening 
(before 18:30)  
Confinement until the 
morning of day 8 
Standardised dinner 
Fasting for at least 10 h 
(overnight) before first 
dosing 
Visit 3
Day 1  

Vital sign measurement at pre-dose 

IMP administration at 08:00 ± 1 h; intravenous infusion 
for 5 min 

Vital signs measurement at 1 h post-dose 

Blood sample collection for pharmacokinetic analysis 
at: pre-dose (0) and 5 (at the end of the infusion), 8, 12, 
15, 20, 25, 30, 60 min, 2, 4, 6, 8, 10 and 12 h 
post-dose 

Urine sample collection for pharmacokinetic analysis 
at: 0-4; 4-8; 8-12 and 12-24 h post-dose 

Check of AEs and concomitant medications  
Standardized lunch at 
13:00  
standardized dinner at 
21:00  
Day 2 

Blood sample collection for pharmacokinetic analysis 
at 24 and 32 h post-dose 

Urine sample collection for pharmacokinetic analysis at 
24-32 h post-dose 

Check of AEs and concomitant medications 
Standardized breakfast 
around 9:00  
Standardized 
lunch 
around 13:00  
Standardized 
dinner 
around 21:00  
Day 3 

Vital sign measurement at 48 h post-dose 

Physical 
examination 
(body 
weight, 
physical 
abnormalities) at 48 h post-dose 

Clinical laboratory analyses: haematology, blood 
chemistry, urinalysis at 48 h post-dose  

Check of AEs and concomitant medications
Standardized breakfast 
around 9:00  
Standardized 
lunch 
around 13:00  
Standardized 
dinner 
around 21:00 
CCI
CCI


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Day 
Procedures/Assessments 
Notes 
Visit 4
From day 4 to 
day 5 

IMP administration at 08:00 ± 1 h; infusion for 5 min 

IMP administration at 20:00 ± 1 h; infusion for 5 min 

Blood sample collection for pharmacokinetic analysis 
at: pre-dose (0) before the morning infusion  

Check of AEs and concomitant medications 
Standardized breakfast 
around 9:00  
Standardized 
lunch 
around 13:00  
Standardized 
dinner 
around 21:00  
Visit 5
Day 6 

Vital sign measurement at pre-dose 

IMP administration at 08:00 ± 1 h; infusion for 5 min 

Vital signs measurement at 1 h post-dose 

Blood sample collection for pharmacokinetic analysis 
at: pre-dose (0) and 5 (at the end of the infusion), 8, 12, 
15, 20, 25, 30, 60 min, 2, 4, 6, 8, 10 and 12 h post-dose 

Urine sample collection for pharmacokinetic analysis 
at: 0-4; 4-8; 8-12 and 12-24 h post-dose 

Check of AEs and concomitant medications  
Standardized lunch at 
13:00 (about 5 h post-
dose) 
Standardized dinner at 
21:00 (about 13 h post-
dose)  
Day 7 

Blood sample collection for pharmacokinetic analysis 
at 24 and 32 h post-dose 

Urine sample collection for pharmacokinetic analysis at 
24-32 h post-dose 

Check of AEs and concomitant medications 
Standardized breakfast 
around 9:00  
Standardized 
lunch 
around 13:00  
Standardized 
dinner 
around 21:00  
Final Visit/ETV
Day 8 or early 
termination visit 
(ETV) in case of 
discontinuation 

Vital sign measurement at 48 h post-dose (or at ETV) 

Physical 
examination 
(body 
weight, 
physical 
abnormalities) 

Clinical laboratory analyses: haematology, blood 
chemistry, urinalysis  

Check of AEs and concomitant medications 
In case of clinically significant results at the final visit, the 
subjects will be followed-up by the investigator until the 
normalisation of the concerned clinical parameter(s) 
Discharge 
from 
the 
clinical centre in the 
morning of Day 8 after 
vital sign measurement, 
blood 
sampling 
for 
clinical 
laboratory 
assays 
and 
physical 
examination or in case 
of ETV. Upon leaving, 
the subjects will be 
instructed to contact 
immediately 
the 
investigator in case of 
occurrence 
of 
any 
untoward 
medical 
occurrence 
11.2
Diet and lifestyle 
The subjects will not take any food or drinks (except water) for at least 10 h (i.e. overnight) 
before the first drug administration (Day 1) and will remain fasted up to 5 h post-dose.  
During the study, water will be allowed as desired. In order to maintain an adequate hydration, 
the subjects will be encouraged to drink at least 120 mL of mineral water every 2 h for 6 h post-
dose on Days 1 and 6. 
Coffee, tea or food containing xanthines (i.e. coffee, tea, soda, coffee with milk, energy drinks, 
etc.), alcohol and grapefruit will be forbidden starting 48 h before the enrolment until the end 
of the study. Smoking is not allowed for the whole study duration. 
During confinement, routine ambulant daily activities will be strongly recommended. 


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The timing of each meal to be served during confinement is shown in Table11.1.1 above.  
In the evening of day -1, upon confinement, the subjects will receive a light dinner before 
fasting overnight.  
On days 1 and 6 (PK sampling collection days), standardised lunch and dinner will be served.  
On days 2, 3, 4, 5 and 7, standardised breakfast, lunch and dinner will be served.  
11.2.1
Restrictions 
During the study, the subjects will be confined from the evening preceding the first drug 
administration (study day -1) until the morning of day 8 (after the final visit). They will attend 
the clinic in the evening of day -1 not later than 18:30.  
During confinement, hazardous, strenuous or athletic activities will not be permitted. 


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12
DESCRIPTION OF SPECIFIC PROCEDURES 
12.1
Physical examination 
Full physical examinations will be performed at the screening visit, visit 3, day 3 and final visit/ 
ETV. Information about the physical examination will be recorded by the investigator. Any 
abnormalities will be recorded. 
Significant findings/illnesses, reported after the start of the study and that meet the definition 
of an AE (see § 16), will be recorded in the subject source documents. 
Date of the physical examination, overall investigator’s interpretation (as normal or abnormal 
and, if abnormal, clinically significant or not clinically significant) and clinically significant 
abnormalities (if any) will be reported in the individual CRFs. 
12.1.1
Body weight 
Body weight will be recorded during each physical examination. 
Subjects will be weighed (kg) lightly clothed without shoes. Height will be measured at 
screening only and BMI will be recorded. BMI will be calculated as weight [kg]/(height [m] x 
height [m]). 
12.1.2
Vital signs 
Subjects blood pressure (BP) and heart rate (HR) will be measured by the investigator or his/her 
deputy after 5 min at rest (in sitting position) at:  
Screening  
Visit 3, Day 1: at pre-dose and 1 h post-dose 
Visit 3, Day 3: at 48 h post-dose  
Visit 5, Day 6: at pre-dose and 1 h post-dose 
Final visit (at 48 h post-dose)/ETV  
12.1.3
ECGs 
One 12-lead ECG will be performed (in sitting/supine position) at screening only. 
12.2
Clinical laboratory assays 
Samples of blood and urine will be collected. The following laboratory analyses will be 
performed at the screening visit: 
Haematology 
Leukocytes and leukocyte differential count (percentage values and absolute values), 
erythrocytes, haemoglobin (conv. units), haemoglobin (IS units), haematocrit, MCV, MCH, 
MCHC, thrombocytes. 


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Blood chemistry 
Electrolytes: sodium, potassium, calcium, chloride, inorganic phosphorus 
Enzymes: alkaline phosphatase, γ-GT, AST, ALT 
Substrates/metabolites: total bilirubin, creatinine, glucose, BUN or urea, uric acid, total 
cholesterol, triglycerides 
Proteins: total proteins 
Serum pregnancy test (women). 
Urine analysis 
Urine chemical analysis (stick): pH, specific weight, appearance, colour, nitrites, proteins, 
glucose, urobilinogen, bilirubin, ketones, haematic pigments, leukocytes 
Urine sediment (analysis performed only if positive): leukocytes, erythrocytes, flat cells, round 
cells, crystals, cylinders, mucus, bacteria 
Serum virology and bacteriology 
Hepatitis B (HBs antigen), Hepatitis C (HCV antibodies), HIV 1/2 (HIV Ag/Ab combo), 
Treponema pallidum.  
The same analyses, with the exception of virology, bacteriology and serum pregnancy test, will 
be performed also at Visit 3, Day 3 (48 h post-dose after the single dose) and at the final 
visit/ETV. 
A urine drug test will be performed at the clinical centre at screening, using a urine multi-drug 
kit. The following drugs will be assessed: cocaine, amphetamine, methamphetamine, 
cannabinoids (delta-9-tetrahydrocannabinol - THC), opiates. The same test will be repeated 
upon confinement at Visit 2, day -1. 
A serum pregnancy test will be performed by the laboratory at screening. A urine pregnancy 
test will be performed at Visit 2, day -1 at the clinical centre, upon confinement. 
Date/time of samples collection, overall investigator’s interpretation (as normal or abnormal 
and, if abnormal, clinically significant or not clinically significant) and clinically significant 
findings (if any) will be reported in the individual CRFs. All clinically significant abnormalities 
after the screening visit will be recorded as AEs. Hard copies of the laboratory print-outs will 
be attached to the CRFs. 
12.3
Sampling for pharmacokinetic analysis 
12.3.1
Venous blood sampling 
Venous blood samples (up to 10 mL) will be collected from a forearm vein at the following 
times, both on Day 1, 2, after the single dose, and on Day 6, 7, after the last multiple dose:  
at pre-dose (0) and 5 (at the end of the infusion), 8, 12, 15, 20, 25, 30, 60 min, 2, 4, 6, 8, 
10, 12, 24 and 32 h post-dose 
The start of infusion (not the end) will be considered as time 0. 


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Actual sampling times for each subject will be recorded in the individual case report forms 
(CRFs). The actual sampling times should not exceed the recommended tolerance ranges 
presented in the following table. Any deviation outside the recommended ranges will be verified 
through Data Clarification Forms and, if confirmed, will be reported as protocol deviation, 
although it will not automatically lead to the exclusion of the concerned subjects from the PK 
analysis set. 
Table 12.3.1.1 
Tolerance ranges for the scheduled sampling times 
Sampling time 
Tolerance range 
Pre-dose (0)
Within 30 minutes before IMP administration
0.0833 h (5 min), 0.1333 (8 min) 
0.200 h (12 min), 0.250 h (15 min)
20 seconds 
0.3333 h (20 min), 0.4167 (25 min), 0.5 h (30 min)
± 1 min
1 h
± 3 min
2, 4 h
± 5 min
6, 8, 10, 12, 24, 32 h
± 10 min
Blood samples for PK analysis will be collected using an indwelling catheter with switch valve. 
The cannula will be rinsed, after each sampling, with about 1 mL of sterile saline solution. The 
first mL of blood will be discarded at each collection time. 
The remaining blood sample will be collected into blood collection tube. 
Storage on ice will not last longer than 10 min until centrifugation at 4° C for 10 min at 2500xg 
to obtain plasma. Plasma will be collected into separate tubes, where an appropriate volume of 
DTT solution (50 µL DTT solution/mL plasma, i.e. 5% of the plasma volume according to the 
ratio 75 µL of DTT solution for 1.5 mL of plasma) will be added. The samples will be mixed 
shortly (Vortex, 20 sec), divided into two aliquots, P1 and P2 and transferred to ≤-70° C as soon 
as possible. The DTT solution (5 mg DTT/mL of water) will be freshly prepared in the evening 
before each sampling day and kept refrigerated until use. The preparation of the solution will 
be recorded in appropriate forms. 
If any clinical assessment, such as vital signs measurement, is foreseen at the same time-point 
as blood sampling for PK analysis, blood collection will be performed at the scheduled time. 
However, vital signs can be influenced by the blood sampling. Therefore, these assessments 
can be performed within 30 min before the pre-dose PK time point (0 h) and within 10 min 
before the other scheduled PK time-points. Any deviations outside the recommended time will 
be verified through Data Clarification Forms. However, since vital signs measurements will be 
performed for safety reasons only, deviations from the planned time schedule will be considered 
not relevant. 
12.3.2
Urine collection 
Urine for PK will be collected in the following time intervals both on Day 1, 2, after the single 
dose, and on Day 6, 7 after the last multiple dose:  
0-4; 4-8; 8-12, 12-24 and 24-32 h post-dose 


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Bladder must be emptied before the end of each collection period. During each interval, urine 
will be collected into containers, kept refrigerated at approximately 4° C and containing DTT 
solution. At the end of each collection interval, urine volume will be measured and after 
thorough mixing, two aliquots of 10 mL each (U1 and U2) will be prepared in polypropylene 
tubes. Urine volume collected in each collection interval will be accurately recorded in 
appropriate forms and in the CRFs. The two aliquots will be stored at ≤-70° C. The DTT 
solution (5 mg DTT/mL of water) will be freshly prepared in the evening before each sampling 
day and kept refrigerated until use. The preparation of the solution will be recorded in 
appropriate forms. 
12.3.3
Analytics  
The concentration of total NAC in plasma and urine will be determined at a certified 
bioanalytical laboratory to be designated in China, using a fully validated LC-MS/MS method, 
with a lower quantification limit (LQL) of 10 ng/mL.  
Analyses will be performed according to the general Principles of “OECD Good Laboratory 
Practices for testing of chemicals" C(81) 30 (final) and GCP/ CFDA GCP (see also § 9.2). 
The method validation report and the analytical report will be attached to the final report. 
12.3.4
Labelling, storage and transport of samples 
12.3.4.1
Samples labelling 
Each sample tube will be clearly and unequivocally identified with a label resistant to the 
storage temperature and reporting: 
Study code
Sponsor code Z7244J01
Subject number 
Subject or Process or Unique Subject Identification number, e.g. 
, etc.
Tube identification
Study day
1 or 6
Scheduled sampling time as min and h; see § 12.3.1
12.3.4.2
Samples storage and transport 
During the study the samples will be stored at ≤-70° C. At the end of each collection day, 
aliquots 1 and 2 will be stored in separate freezers. 
All aliquots 1, packed in sufficient solid CO2, will be shipped by an authorised courier from the 
clinical site, China, to Eurofins Central Laboratory, Shanghai, China. Aliquots 1 will remain 
stored at Eurofins Central Laboratory or a subcontracted bioanalytical laboratory to be 
designated until finalisation of the bioanalytical report. Afterwards, the samples will be 
destroyed and a certificate of destruction will be provided to the sponsor.
The counter-samples (aliquot 2) will remain stored at the clinical site, China. These samples 
could either be: 
CCI
CCI


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
sent to the laboratory for reanalysis should this become necessary for analytical reasons or 
if any problems occur during the delivery of aliquots 1, or 

destroyed at an authorised site, or 

transferred to the sponsor upon written request. 
No analyses different from those stated in this protocol and agreed by the subjects when signing 
the informed consent form will be performed unless a new informed consent and a new approval 
from the Ethical Committee is obtained. The subjects may ask to destroy their own samples at 
any time. 


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13
ASSIGNMENT OF STUDY TREATMENT 
13.1
Randomisation 
No randomisation will take place in the present study. All the subjects will receive the same 
treatment.  
13.2
Blinding 
This is an open study. No masking procedure will be applied. 


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14
EVALUATION PARAMETERS 
14.1
Study variables 
14.1.1
Primary variables 
After single dose: 
Cmax, tmax, kel, t1/2, AUC(0-t), AUC(0-∞), AUC(0-12), Vd, CLt, Ae(0-t), Fe(0-t) and CLr
After multiple doses: 
Css_max, tss_max, Css_min, AUCss(0-t), AUCss(0-12), Css_av, R, DF% and Aess(0-t). 
14.1.2
Secondary variables 
TEAEs, vital signs (BP, HR), body weight, physical examinations, laboratory parameters. 
14.2
Pharmacokinetic assessments 
14.2.1
Pharmacokinetic parameters 
The following PK parameters will be measured and/or calculated for plasma NAC, using the 
validated software Phoenix® WinNonlin® version 6.3 or higher (Certara USA, Inc., Princeton, 
NJ) (actual version will be stated in the final report), after single dose of IMP: 
Cmax: 
Maximum NAC plasma concentration 
tmax: 
Time to achieve Cmax 
kel: 
Terminal elimination rate constant, calculated, if feasible, by log-linear 
regression using at least 3 points 
t1/2: 
Half-life, calculated, if feasible, as ln2/kel
AUC(0-t): 
Area under the concentration-time curve from single dose to the last observed 
concentration time t, calculated with the linear up/log down trapezoidal method
AUC(0-∞): 
Area under the concentration-time curve extrapolated to infinity, calculated, if 
feasible, as AUC(0-t) + Ct/kel, where Ct is the last measurable drug concentration
AUC(0-12): 
Area under the concentration-time curve at steady-state in the tau interval (from 
single dose to 12 h post-dose), calculated with the linear up/log down 
trapezoidal method 


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Vd: 
Volume of distribution associated with the terminal slope, calculated, if 
feasible, as Dose/(AUC(0-∞)*kel) 
CLt: 
Total body clearance, calculated, if feasible, as Dose/ AUC(0-∞)
Ae(0-t): 
Total amount of NAC excreted in urine from single dose up to 32 h 
Fe(0-t): 
Total fraction of NAC dose excreted in urine from single dose up to 32 h 
CLr: 
Renal clearance, calculated, if feasible, as Ae(0-t)/ AUC(0-∞)
The following PK parameters will be measured and/or calculated for plasma NAC, using the 
same software, after multiple doses of IMP: 
Css_max: 
Maximum NAC plasma concentration at steady-state 
tss_max: 
Time to achieve Css_max 
Css_min: 
Trough NAC plasma concentration at steady-state, measured as concentration 
at t=12 h 
AUCss(0-t): 
Area under the concentration-time curve at steady-state from the last multiple 
dose to the last observed concentration time t, calculated with the linear up/log 
down trapezoidal method 
AUCss(0-12): 
Area under the concentration-time curve at steady-state in the tau interval (from 
the last multiple dose to 12 h post-dose), calculated with the linear up/log down 
trapezoidal method 
Css_av: 
Average NAC plasma concentration at steady-state, calculated as AUCss(0-12)
/tau 
R: 
Accumulation ratio, calculated as AUCss(0-12)/ AUC(0-12)
Aess(0-t): 
Total amount of NAC excreted in urine from the last multiple dose to 32 h at 
steady-state 
DF%: 
Degree of fluctuation over one dosing interval at steady-state, calculated as 
(Css_max - Css_min)/ Css_av*100 
The sampling schedule is considered adequate if the ratio AUC(0-t)/ AUC(0-∞) equals or exceeds 
a factor of 0.8 (i.e. if %AUCextra is <20%) for more than 80% of the individual PK profiles. This 
assures that the primary variable AUC(0-t) covers a sufficient percentage of the theoretical total 
extent of exposure.  


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The quality of log-linear regression (and, consequently, the reliability of the extrapolated PK 
parameters) should be demonstrated by a determination coefficient R2 ≥ 0.8. Individual 
extrapolated parameters, when considered unreliable, will be reported as NC (not calculated).  
14.3
Safety assessments 
Safety and general tolerability of the IMP will be based on TEAEs, physical examinations 
including body weight, vital signs and routine haematology, blood chemistry and urinalysis 
laboratory tests. 


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15
STATISTICAL METHODS 
The data documented in this trial and the measured clinical parameters will be presented using 
classic descriptive statistics (i.e. total number of subjects treated [N], number of observations 
[n], mean standard deviation [SD], minimum [Min], median, maximum [Max]) for quantitative 
variables and frequencies (i.e. count and percentages) for qualitative variables if not stated 
otherwise. 
Not available data will be evaluated as “missing values”. The statistical analysis of demographic 
and safety data will be performed using the analysis software SAS®  . 
The statistical analysis of PK parameters will be performed using the validated software 
Phoenix® WinNonlin® version 6.3 or higher (Certara USA, Inc., Princeton, NJ) (actual version 
will be stated in the final report). 
15.1
Analysis Sets 
15.1.1
Definitions 
A subject will be defined as screened after the signature of the informed consent, regardless of 
the completion of all the screening procedures. 
A subject will be defined as eligible if he/she meets all the inclusion/exclusion criteria. 
Otherwise he/she will be defined as a screen failure.  
A subject will be defined as enrolled in the study if he/she is included into the interventional 
phase of the study.
The following analysis sets are defined: 

Enrolled set: all enrolled subjects. This analysis set will be used for demographic, baseline 
and background characteristics 

Safety set: all subjects who receive at least one dose of IMP. This analysis set will be used 
for the safety analyses 

PK set: all enrolled subjects who fulfil the study protocol requirements in terms of IMP 
intake and have evaluable PK data readouts for the planned analyses, with no major 
deviations that may affect the PK results. This analysis set will be used for the statistical 
analysis of the PK parameters. 
Each subject will be coded as valid or not valid for the safety and the PK set. Subjects will be 
evaluated according to the treatment they actually receive. 


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15.1.2
Reasons for exclusion from the PK set before bioanalysis 
Reasons for the exclusion of subjects from the PK set are the following: 

intake of concomitant medications which could render the plasma concentration-time 
profile unreliable 

AEs which could render the plasma concentration-time profile unreliable 

administration errors which could render the plasma concentration-time profile unreliable 

other events which could render the plasma concentration-time profile unreliable 
If one of these events occurs, it will be noted in the CRF during the study. 
The samples from the subjects excluded from the PK set should still be assayed and the results 
listed. Subjects should not be excluded from the PK set if the AUC0-t covers less than 80% of 
the AUC0-∞. 
15.2
Sample size and power considerations 
The sample size was not calculated through any statistical calculation. A sample size of 24 
subjects was estimated as sufficient for the descriptive purposes of the present study. 
Drop-out subjects will not be replaced.  
15.3
Demographic, baseline and background characteristics 
Critical demographic characteristics will be examined according to qualitative or quantitative 
data. Qualitative data will be summarised in contingency tables. Quantitative data will be 
summarised using classic descriptive statistics. 
15.4
Analysis of pharmacokinetic parameters 
15.4.1
Descriptive pharmacokinetics 
A descriptive PK will be presented. The results will be displayed and summarised in tables and 
figures. Individual and mean curves (+SD at sampling times), indicating inter-subject 
variability, will be plotted. Data below the lower quantification limit (BLQL) will be considered 
as 0 in the calculations and presented as BLQL in listings and tables. As a consequence of 
BLQL (i.e. 0) values, calculated geometric means (if requested) could be null. For this reason, 
in the presence of any null value, the geometric mean will be reported as not calculated (NC). 
15.5
Safety and tolerability evaluation 

AEs
AEs will be coded by System Organ Class (SOC) and Preferred Term (PT), using the Medical 
Dictionary for Regulatory Activities (MedDRA). 
AEs will be classified as pre-treatment AEs (PTAEs) and TEAEs, according to the period of 
occurrence, as follows:  


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PTAEs: all AEs occurring before the first dose of IMP and not worsening after the first 
dose of IMP 
TEAEs: all AEs occurring or worsening after the first dose of IMP 
Individual PTAEs and TEAEs will be listed in subject data listings. No summary table will be 
provided for PTAEs. TEAEs will be summarised by treatment and overall. The number and 
percentage of subjects with any TEAE and the number of TEAEs will be tabulated by SOC and 
PT, seriousness, relationship to treatment and severity.  

Physical examination 
Date of the physical examination, overall investigator’s interpretation (as normal or abnormal 
and, if abnormal, clinically significant or not clinically significant) and clinically significant 
abnormalities (if any) will be listed. 

Laboratory data  
Date/time of samples collection, overall investigator’s interpretation (as normal or abnormal 
and, if abnormal, clinically significant or not clinically significant) and clinically significant 
findings (if any) will be listed. All laboratory results will be listed and a table of all the abnormal 
values will be presented. The overall investigator’s interpretation will be summarised using 
tables of frequency.  
The toxicity grading of laboratory tests is determined based on the NCI CTCAE V4.03. 

Vital signs 
Vital signs values will be listed and summarised by descriptive statistics. 

Body weight 
Body weight values will be listed and summarised by descriptive statistics. 


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16
DEFINITION AND HANDLING OF AEs AND SAEs 
16.1
Applicable SOPs 
AEs definition, classification and management will follow the CRO’s SOPs, based upon 
applicable local and international regulations. The full SOPs or an operative summary will be 
made available to the clinical centre. 
A brief summary of AE definition, classification and management is reported below. 
16.2
Definition of Adverse Event (AE) 
An Adverse Event is “any untoward medical occurrence in a patient or clinical trial subject 
administered a medicinal product and which does not necessarily have a causal relationship 
with this treatment”. 
AEs include: 
worsening (change in nature, severity or frequency) of conditions present at the onset 
of the trial 
patient/subject deterioration due to the primary illness 
intercurrent illnesses 
drug interactions 
events related or possibly related to concomitant medications 
abnormal laboratory values, as well as significant shifts from baseline within the range 
of normal, which the investigator considers to be clinically significant. 
An AE can therefore be any unfavourable and unintended sign (including an abnormal 
laboratory finding), symptom, or disease temporally associated with the use of an 
investigational medicinal product, whether or not considered related to the investigational 
medicinal product. 
16.3
Definition of Adverse Drug Reaction (ADR) 
An Adverse Reaction is “any untoward and unintended response to an investigational 
medicinal product related to any dose administered”. 
All AEs judged by either the reporting investigator or the sponsor as having a reasonable causal 
relationship to a medicinal product qualify as adverse reactions. The expression reasonable 
causal relationship means to convey in general that there is evidence or matter to suggest a 
causal relationship. 
Unexpected Adverse Drug Reaction 
An unexpected ADR is: “An adverse reaction, the nature, or severity of which is not consistent 
with the applicable product information (Reference Safety Information [RSI]).  
Reference Safety Information (RSI): in order to assess whether an adverse reaction is 
expected, the IB will be used.


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16.4
Definition of Serious Adverse Events or Serious Adverse Drug Reaction 
A Serious Adverse Event (SAE) is: “any untoward medical occurrence or effect that at any 
dose: 
results in death 
is life-threatening (i.e. the subject was at risk of death at the time of the event/reaction; it 
does not refer to an event/reaction which hypothetically might have caused death if it were 
more severe) 
requires inpatient hospitalization or prolongation of existing hospitalization 
results in persistent or significant disability/incapacity (where disability is defined as a 
permanent or substantial disruption of ability to carry out normal life functions, either 
reported or defined as per clinical judgement) 
is a congenital anomaly/birth defect 
is an important medical event that may not result in death, be life-threatening, or require 
hospitalization but, according to appropriate medical judgment, it may jeopardize the 
subject and may require medical or surgical intervention to prevent any of the outcomes 
listed in the definition above.  Examples of such events include allergic bronchospasm 
requiring intensive treatment in an emergency room or at home, and blood dyscrasias or 
convulsions that do not result in hospitalization, or development of drug dependency or 
drug abuse.  
Any suspected transmission via a medicinal product of an infectious agent is also considered a 
serious adverse event. 
A Suspected Unexpected Serious Adverse Reaction (SUSAR) is an ADR that is both 
unexpected (not consistent with the applicable product information, e.g. IB) and also meets the 
definition of a SAE. 
A non-serious adverse event is any adverse event that does not meet the criteria listed above for 
a serious adverse event. 
16.5
Definition of Severity of Adverse Events 
The term “severe” is used to describe the intensity (severity) of a specific event: 
-
Mild: causing no limitation of usual activities; the subject may experience slight 
discomfort. 
-
Moderate: causing some limitation of usual activities; the subject may experience 
annoying discomfort. 
-
Severe: causing inability to carry out usual activities; the subject may experience 
intolerable discomfort or pain. 
16.6
Definition of Adverse Event causality 
Causality shall be determined according to the definition of ADR given in 16.3. 


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All AE judged by either the investigator or the sponsor as having a reasonable suspected causal 
relationship to an investigational medicinal product qualify as adverse reactions. The causality 
assessment given by the investigator should not be downgraded by the sponsor. 
The following binary decision for causality will be used: 

Reasonable possibility that the IMP caused the event 

No reasonable possibility that the IMP caused the event 
Features supportive of an association include: 
•
temporal plausibility 
•
pharmacological properties of the drug or of the substance class 
•
course of the adverse event after dechallenge and, if applicable, after rechallenge 
•
specific tests indicating involvement of the drug in the occurrence/worsening of the 
adverse event  
•
alternative explanations 
16.7
Adverse Events recording 
Each AE occurring to a subject, either spontaneously revealed by the subject or observed by 
the investigator, whether believed by the investigator to be related or unrelated to the IMP, must 
be recorded on the AE information page of the eCRF (for SAEs information must be recorded 
also on the AE information page of the eCRF).  
The Investigator will also perform an evaluation with respect to seriousness and causality 
(relationship of any AE to IMP) of the AEs. and record it on the appropriate section of the eCRF 
and records it on the appropriate section of the eCRF. 
16.8
AEs monitoring window 
Start of monitoring: from immediately after the signature of the informed consent 
End of monitoring: final study visit/ETV (observation window) 
An AE occurring after the final visit/ETV and coming to knowledge of the investigator (e.g. 
during a follow-up assessment or by spontaneous reporting by study subjects) must be recorded 
only if it is an ADR, according to the investigator’s judgment. 
16.9
Adverse Events reporting 
The official language for reporting is English. The investigator and clinical staff of the present 
study are familiar with English language. 
The investigator must report to the CRO all AEs which occur during the study, regardless of 
their relationship to the IMP. Protocol specific AEs or laboratory abnormalities critical to safety 
evaluations are to be identified in the protocol and reported to the sponsor according to reporting 
requirements and within the time periods specified. 


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All AEs are recorded by the investigator on the AE information page of the eCRF. 
In addition, SAE will have to be reported according to the following detailed procedure. 
16.9.1
SAEs reporting 
With the exception of those SAEs that are identified as not requiring immediate reporting in the 
protocol, the investigator must report the SAEs to the CRO immediately and no later than 24 
hours from when he/she becomes aware of the SAE, by faxing the “Serious Adverse Event 
Form” (back up plan) or e-mailing as scanned attachment (backup plan) or by Electronic Data 
Capture to the Drug Safety Unit (DSU) personnel (preferred method), as stated in the "List of 
CRO/ personnel" in § 16.13 of this protocol. 
The community standards of confidentiality must always be maintained and any relevant 
national legislation on data protection must be followed. 
Another copy of the “Serious Adverse Event Form” will be retained by the Investigator for the 
Investigator’s file. 
If the investigator becomes aware of any SAE occurred to a subject within the follow-up 
window established in the protocol, he/she will report the SAE as above. The SAE will be also 
reported in the eCRF. 
If outside the follow-up window established in the protocol the investigator becomes aware of 
a SAE, if the investigator judges that the SAE is related to the study drug, it should be reported 
to the Sponsor. The Investigator might use the “Serious Adverse Event Form” via email or fax, 
but the SAE must not to be reported in the eCRF, as it is not an event occurred within the study 
period. 
The investigator must report all SAEs that occur to the subjects to National Medical Products 
Administration/Local Ethics Committee/Provincial Food and Drug Administration/National 
Health and Family Planning Commission immediately no later than 24hours from when he/she 
becomes aware of SAE. Any SAEs that happen to the subjects outside the follow-up window 
should be reported by investigator to National Medical Products Administration/Local Ethics 
Committee/Provincial Food and Drug Administration/National Health and Family Planning 
Commission immediately. 
16.10
Follow-up for Adverse Events 
A follow-up “Serious Adverse Event Form” will be filled in if important follow-up information 
(i.e. diagnosis, outcome, causality assessment, results of specific investigations) is made 
available after submission of the initial AE Form for immediate reporting. Follow-up “Serious 
Adverse Event Form” will be reported to the Sponsor as above-described, under Section 16.9.1. 
In any case of an AE that, in the opinion of the investigator, requires the subject’s 
discontinuation, follow-up information relating to the subject’s subsequent course must be 
collected until the event has subsided or the condition stabilised or when the subject is lost to 
follow up or death. 


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When follow-up data on non-serious AE are collected, information should be reported under 
“Comments” in the Final report of the CRF. 
16.11
SUSARs management 
The clock for initial expedited reporting starts as soon as the information containing the 
minimum reporting criteria has been received by the sponsor (day 0).  
For fatal and life-threatening SUSARs the EC and Competent Authority (CA) should be 
informed as soon as possible and in any case within 7 days. 
If the initial report is incomplete, e.g. not all the information/assessments were available, a 
complete report should be sent within an additional 8 days. Complete follow-up report should 
be sent within 15 calendar days. 
SUSARs which are not fatal and not life-threatening are to be reported within 15 days. 
The minimum information to be reported includes: 
Sponsor study number 
One identifiable coded subject 
One identifiable reporter 
One SUSAR 
One suspect IMP (including active substance name, code) 
A causality assessment (a reasonable possibility of a causal relationship with the study 
drug can be excluded only if there is information supporting this decision, otherwise it 
cannot be excluded). 
16.12
Other events qualified for expedited reporting 
Other safety issues also qualify for expedited reporting when they might materially alter the 
current benefit-risk assessment of a medicinal product or would be sufficient to consider 
changes in the medicinal product administration or in the overall conduct of the trial, for 
instance: 
single case reports of an expected serious adverse reaction with an unexpected outcome 
(e.g. a fatal outcome) 
an increase in the rate of occurrence of an expected serious adverse reaction, which is 
judged to be clinically important. 
post-study SUSARs that occur after the subject has completed a clinical trial and are 
reported to the investigator by the subject. 
new events relating to the conduct of the trial or the development of the medicinal 
product likely to affect the safety of the subjects, such as : 
−a SAE which could be associated with the trial procedures and which could modify 
the conduct of the trial 
−a significant hazard to the subject population such as lack of efficacy of a medicinal 
product used for the treatment of a life-threatening disease 
−a major safety finding from a newly completed animal study (such as carcinogenicity) 
or from other clinical trials. 


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16.13
SAEs: contacts 
The investigator will report any SAE to the CRO. The CRO’s details for SAEs are the 
following: 
Email: 
The sponsor’s details for SAEs are the following: 
Email: 
 
16.14
Pregnancy 
Subjects must be instructed that known or suspected pregnancy occurring during the study 
should be confirmed and reported to the Investigator, who must then withdraw the subject from 
the study without delay. The Investigator should also be notified of pregnancy occurring during 
the study but confirmed after its completion. In the event that a subject is subsequently found 
to be pregnant after inclusion in the study, then any pregnancy will be followed to the end of 
pregnancy or pregnancy termination and the status of mother and child will be reported to the 
Sponsor after delivery through the Pregnancy Report Form provided by the Sponsor. The 
Investigator will send pregnancy reports to DSU within the timeframes of SAEs, with follow-
up information to be actively sought for the outcome of pregnancy.  
-
Part I of the Form is completed at the time of awareness of foetal exposure during 
pregnancy.  
-
Part II of the Form is filled in when information on pregnancy outcome becomes available. 
If pregnancy results in abnormal outcome (spontaneous miscarriage, stillbirth and congenital 
anomalies) that the investigator considers to be due to the IMP, this will be treated as an 
expedited ADR report. 
CCI
CCI


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DATA MANAGEMENT PROCEDURES 
17.1
Data collection – CRFs 
The investigator must ensure that the clinical data required by the study protocol are carefully 
reported in the eCRFs. He/she must also check that the data reported in the eCRFs correspond 
to those in the subject’s source documents.  
ECG and laboratory results must be printed and signed by the Investigator and kept as source 
data on site after entering outcome into the eCRF. 
All other data has to be documented in the subject file as source data first and then entered into 
the eCRF.  
Data must be entered into eCRFs in English by the designated site personnel as soon as possible 
after a subject visit, and monitors will have access to data recorded. These data will be reviewed 
versus source documents by trial monitors for completeness and acceptability during 
monitoring visits.  
If data correction is required for an eCRF after initial entry, the site personnel can make 
necessary corrections on their own or as a response to an automated query by the eCRF system. 
Monitor and Clinical Data Manager review the eCRF for accuracy and can also generate queries 
to the investigational staff for resolution.  
Any correction to the eCRFs' entries must be carried out by the Investigator or a designated 
member of staff. Corrections are recorded in an audit trail that records the old information, the 
new information, and identification of the person making the changes, date of correction made 
and reason for change. In the interests of completeness of data acquisition, the questions which 
are repeated in each section of the eCRFs should be answered in full, even if there are no 
changes from a previous examination. A reasonable explanation must be given by the 
Investigator for all missing data. The Investigator or his/her designees named in the clinical 
staff list will review the eCRF for accuracy and completeness. The Investigator must 
electronically sign and date the eCRF pages as indicated.  
17.2
Database management 
The CRO will update and verify the database and create the final SAS data sets. The final data 
file will be transferred to the sponsor in the agreed format with all the other study 
documentation. 
17.2.1
Coding dictionaries 
Medical/surgical history and underlying diseases, clinically significant physical examination 
abnormalities and AEs will be coded using the Medical Dictionary for Regulatory Activities 
(MedDRA™). 
Previous and concomitant medications will be coded using the WHO Drug Dictionary 
Enhanced (WHODDE). The version of the coding dictionaries will be stated in the study report. 


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STUDY MONITORING, QUALITY CONTROL AND 
QUALITY ASSURANCE 
18.1
Monitoring 
The monitoring visits will be conducted by appropriate staff of PAREXEL China Co. Ltd 
according to PAREXEL SOPs.  
Monitoring activities, including monitoring purpose, selection and qualifications of monitors, 
extent and nature of monitoring, monitoring procedures, monitoring reports will comply with 
ICH-GCP chapter 5.18, CFDA GCP requirements and, when appropriate, further national 
regulatory guidelines.
Adequate time and availability for monitoring activities should be ensured by the investigator 
and key study personnel. 
Data verification is required and will be done by direct comparison with source documents, 
always giving due consideration to data protection and medical confidentiality. In this respect 
the investigator will assure support to the monitor at all times. 
The investigator agrees, by written consent to this protocol, to fully co-operate with compliance 
checks by allowing authorised individuals to have access to all the study documentation. In 
addition to the monitoring activities performed by the study monitor, the sponsor could perform 
some quality control activities to verify the compliance with the study procedures, the ICH-
GCP guidelines and CFDA GCP.  
18.2
Quality Control and Quality Assurance  
The CRO has implemented and maintains a Quality System that includes quality controls and 
audits at different study steps with written SOPs to ensure that the study is conducted in 
compliance with the protocol and all effective amendments, ICH-GCP, and the applicable 
regulatory requirement(s) and that data have been reliably and correctly generated, recorded, 
processed and reported, in agreement with the ALCOAC principles (Attributable-Legible-
Contemporaneous-Original-Accurate-Complete). 
The clinical site is responsible for implementing and maintaining quality assurance and a 
quality control system to ensure that the study is conducted, and data are generated, documented 
(recorded), and reported in compliance with the protocol, ICH-GCP, CFDA GCP and any 
applicable regulatory requirement(s). 
This protocol has been audited by the Sponsor QA.   
The CRO and the sponsor will be responsible for their respective activities. 
The sponsor may transfer any or all of the sponsor’s trial-related duties and functions to a CRO, 
but the ultimate responsibility for the quality and integrity of the trial data always resides with 
the sponsor. 


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18.3
Applicable SOPs 
The sponsor, the clinical centre and the CRO will follow their respective SOPs in the conduct 
of the respective activities, unless otherwise stated in written agreements. CRO/Sponsor's SOPs 
will be used for AE and SAE definition and management. SOPs will be made available for 
Sponsor’s review, if required. 
18.4
Data access 
The investigator and the CRO will ensure that all source data, medical records, CRFs and all 
other documentation that is relevant to this study will be made accessible for monitoring 
activities, audits, IEC review, and regulatory inspections. 
18.5
Audits and inspections 
The sponsor, independent bodies acting on behalf of the sponsor and the CRO have the right to 
perform audits according to ICH-GCP, CFDA GCP and CFDA responsibilities. 
The study may also be inspected by regulatory authorities.  
The investigator and the CRO agree, by written consent to this protocol, to fully co-operate and 
support audits and inspections compliance checks by allowing authorised individuals to have 
access to all the study documentation. 


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19
ETHICAL CONSIDERATIONS 
19.1
Ethics and Good Clinical Practice (GCP) 
The study will be performed in accordance with the relevant guidelines of the Declaration of 
Helsinki. 
The approval of the study protocol by the local IEC and by the Chinese Health Authorities will 
be obtained before the start of the study.  
The present clinical study will be carried out according to the current revision of Good Clinical 
Practice (GCP), ICH topic E6 (R2), CFDA GCP and any applicable local law requirements. 
19.2
Informed consent 
Before being enrolled into the clinical study, the subjects must have expressed their consent to 
participate, after the investigator has explained to them, clearly and in details, the scope, the 
procedures and the possible consequences of the clinical study. Information will be given in 
both oral and written form. The information sheet and informed consent form will be prepared 
in the local language by the CRO and must be approved by the EC and regulatory authorities. 
It will include all the elements required by law according to the ICH-GCP and CFDA GCP 
recommendations.  
In addition to the standard requirements that physicians are currently obliged to observe when 
providing information, the following points must also be covered: 

a description of the aims of the study and how it will be organised 

the type of treatment (information on the IMP and treatment procedures, as applicable) 

any potential negative effects attributable to the study product or treatment 

the freedom to ask for further information at any time 

the subjects’ right to withdraw from the clinical study at any time without giving reasons 
and without jeopardising their further course of medical treatment  

the existence of a subject insurance cover and obligations following from this cover 
Adequate time and opportunity to satisfy questions will be given to the subjects and the time 
will be recorded. 
The investigator will be supplied with an adequate number of blank informed consent forms to 
be used. The forms will be signed and dated by both the investigator and the subject.  
A copy of the signed form will be given to the subject. 
To ensure medical confidentiality and data protection, the signed informed consent forms will 
be stored in the investigator's study file according to the regulatory requirements (see § 20.3). 
The investigator will allow inspection of the forms by authorised representatives of the sponsor, 
EC members and regulatory authorities. He/she will confirm, by signing and dating the forms, 
that informed consent has been obtained. 


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19.3
Insurance policy 
An insurance cover has been issued in favour of the subjects participating in this clinical study. 
The insurance is in compliance with the local regulation and with the requirements of the Health 
Authorities. 
19.4
Withdrawal of subjects 
It will be documented whether or not each subject completed the clinical study. If, for a subject, 
study treatment or observations are discontinued, the primary reason for discontinuation will 
be recorded. 
19.4.1
Primary reason for discontinuation 
Adverse event: Any (significant) adverse event that in the opinion of the investigator or 
concerned subject is not compatible with study continuation. For the definition of AE, 
please refer to § 16.2.
death: the absence of life or state of being dead
lost to follow-up: the loss or lack of continuation of a subject to follow-up
non-compliance with study drug: an indication that a subject has not agreed with or 
followed the instructions related to the study medication
physician decision: a position, opinion or judgment reached after consideration by a 
physician with reference to the subject
pregnancy: pregnancy is the state or condition of having a developing embryo or fetus in 
the body (uterus), after union of an ovum and spermatozoon, during the period from 
conception to birth
protocol deviation: an event or decision that stands in contrast to the guidelines set out by 
the protocol
study terminated by sponsor: an indication that a clinical study was stopped by its sponsor
technical problems: a problem with some technical aspect of a clinical study, usually 
related to an instrument
withdrawal by subject: study discontinuation requested by a subject for whatever reason
other: different than the ones previously specified
For any subject discontinuing the study, the investigator will:  

ask the subject to undergo, as far as possible, a final medical visit (ETV) to examine the 
subject's health conditions and perform the required blood sampling for the laboratory 
assays. This examination will verify that all values tested at screening have remained within 
a clinically acceptable range (i.e. not clinically significant changes compared to screening) 

arrange for alternative medical care of the withdrawn subject, if necessary 

record the subject decision about the use of collected biological samples 


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
report in the CRF date and time of the last dose administration, and date and primary reason 
of study discontinuation 

record in the CRF any follow-up, if the subject is withdrawn for an AE 
Discontinued subjects will not be replaced.  
19.5
Study termination 
The study will be considered terminated at the date of the last visit of the last subject or upon 
completion of any follow-up procedure described in protocol. The investigator and the sponsor 
have the right to discontinue the study at any time for reasonable medical and/or administrative 
reasons. As far as possible, this should occur after mutual consultation. Reasons for 
discontinuation have to be documented appropriately. 


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20
ADMINISTRATIVE PROCEDURES 
20.1
Material supplied to the clinical centre 
Beside IMP, the following study material will be supplied to the clinical centre: 
final version of the study protocol 
CRF for each subject plus some spare copies 
copy of the investigator’s brochure (IB) relative to the IMP 
informed consent forms 
Moreover, before the start of the study, the investigator will be provided with the following 
documents: CFDA GCP, ICH guidelines, confidentiality agreement (if applicable), protocol 
amendments (if any), declaration of Helsinki, insurance statement, SAE forms, financial 
agreement (if applicable), confidential subject identification code list form, drug accountability 
forms, investigator and study staff list form. 
20.2
Protocol amendments 
In order to obtain interpretable results, neither the investigator nor the sponsor will alter the 
study conditions agreed upon and set out in this protocol. Amendments should be made by 
mutual agreement between the investigator and the sponsor. Any amendment must be set out 
in writing, giving the reasons, and being signed by all concerned parties. The amendment 
becomes then part of the protocol. 
20.3
Study documentation and record keeping 
The investigator should ensure the accuracy, completeness, legibility, and timeliness of the data 
reported to the sponsor in the CRFs and in all required reports. 
The investigator must keep source documents for each subject in the study. All information on 
the CRFs must be traceable to these source documents, which are generally stored in the 
subject's medical file. The source documents should contain all demographic and medical 
information, including laboratory data, ECGs, etc., and the original signed informed consent 
forms. 
Data reported on the CRF that are derived from source documents should be consistent with the 
source documents or the discrepancies should be explained. 
The investigator and the sponsor should maintain the study documents as specified in the 
“Essential Documents for the Conduct of a Clinical Trial” chapter 8 of ICH-GCP and as 
required by CFDA GCP and the applicable regulatory requirement(s).  
These are documents which individually and collectively permit evaluation of a study and the 
quality of the data produced and include groups of documents, generated before the study 
commences, during the clinical study, and after termination of the study and include but are not 
limited to, study protocol, amendments, submission and approval of EC, raw data of subjects 
including lab tests and ECG tracing, insurance contracts, certificate of analysis of the IMP(s), 


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drug accountability records, signed informed consent forms, confidential subjects identification 
code, CRFs, curricula vitae of the investigator and other participants in the study, study staff 
lists and responsibilities, monitoring reports and final study report. 
The investigator and the sponsor should take measures to prevent accidental or premature 
destruction of these documents. 
Study documents must be retained by the investigator and the sponsor as long as needed to 
comply with ICH-GCP, CFDA GCP, national and international regulations. By signing the 
protocol, the investigator and the sponsor agree to adhere to these requirements. 
20.4
Study subjects’ recruitment 
Study participants will be recruited from the volunteers’ database. The study site may also use 
the site management organisation to recruit the study participants.  
In addition, they may also use recruitment posters in other departments within the hospital, 
community outside or newspapers to raise awareness of the study that is going on. 
Other departments within the hospital will be involved to recommend study participants as well.    
The clinical site has detailed SOPs on the recruitment process. 
20.5
Confidentiality and data protection 
By signing this protocol, the investigator and the CRO agree to keep all the information 
provided by the sponsor in strict confidentiality and to request the same confidentiality from 
his/her staff. Study documents provided by the sponsor (protocols, IB, CRFs and other 
materials) will be stored appropriately to ensure confidentiality. The information provided by 
the sponsor to the investigator and to the CRO cannot be disclosed to others without direct 
written authorisation from the sponsor, except for the extent necessary to obtain the informed 
consent from the subjects wishing to participate in the study. 
Data on subjects collected in the CRFs during the study will be documented in an anonymous 
way. If, as an exception, for safety or regulatory reasons identification of a subject becomes 
necessary, the monitor, the sponsor and the investigator will be bound to keep this information 
confidential. 
20.6
Publication policy 
The sponsor agrees that the study results (including negative and inconclusive as well as 
positive results) can be made publicly available by the investigator publishing in peer reviewed 
journals, presenting results at scientific congresses and posting information and results on 
internet-based public registers and databases. 
Study results will be communicated in full to the competent Health Authorities by the 
submission of a complete clinical study report. 


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As the sponsor agrees that the study results can be published by the investigator(s), the 
investigator agrees to submit any manuscript (abstract, publication, paper, etc.) to the sponsor 
before any public disclosure. 
This will be done in order to ensure that clinical study results are reported in an objective, 
accurate and balanced manner. The sponsor reviews the proposed manuscripts, before 
submission, within a reasonable period of time (30-90 days in relation with the complexity of 
the work). 
The investigator will also be provided by the sponsor with the clinical study report and the 
results of any additional analysis, tables, figures, etc. undertaken for the purposes of the article, 
in order to take responsibility for the content of the publication(s). 
On an exceptional basis, the sponsor may temporarily delay registration of certain data elements 
(e.g. compound, name, outcome, measures, etc.) to seek necessary intellectual property 
protection. This is because early disclosure of such data could, in some circumstances, prevent 
or negatively impact patentability. 


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STUDY RESPONSIBLE PERSONS 
21.1
Sponsor 
Zambon S.p.A., via Lillo del Duca 10, I-20091 Bresso, Italy 
Phone: 
+39.02.66.52.41 
Fax: 
+39.02.66.50.14.92 
Protocol Review Committee Chairman 
 
Sponsor’s representatives 
 
21.2
Drug assay 
Eurofins Central Laboratory Shanghai, 395 Jiang Chang West Road, 7th Floor, Shanghai, 
200436 China 
Analytical facility: 
United-Power Pharma Tech Co., Ltd. (UP-Pharma) 
2F, Tower B, No. 33 Science Park Road, 
Changping District, Beijing, P.R.China 
ZIP Code: 102206 
21.3
Project management, data analysis & reporting, monitoring  
PAREXEL International (IRL) Limited (“PAREXEL”), a corporation organized under the 
laws of Ireland with a registered address at 70 Sir John Rogerson’s Quay, Dublin 2, Ireland. 
PPD
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22
REFERENCES 
1.
Ventresca GP, Cicchetti V, Ferrari V. Drugs in Bronchial Mucology. 1989. Cap. 4.1 Thiols 77-102. Edited by 
PC Braga and Allegra. Raven Press Ltd 
2.
Aruoma OI, Halliwell B, Hoey BM, Butler J. The antioxidant action of N-acetylcysteine: Its reaction with 
hydrogen peroxide, hydroxyl radical, superoxide and hypochlorous acid. Free Radical Biol Med 1988; 6:593-
597 
3.
Ziment I. Acetylcysteine: a drug that is much more than a mucokinetic. Biomed & Pharmacother 1988; 42: 
513-520 
4.
Borgstrom L, Kagedal B, Paulsen O. Pharmacokinetics of N-acetylcysteine in man. Eur J Clin Pharmacol 
1986; 31: 217-222 
5.
Olsson B, Johansson M, Gabrielsson J, Bolme P. Pharmacokinetics and bioavailability of reduced and 
oxidized N-acetylcysteine. Eur J Clin Pharmacol 1988; 34: 77-82 
6.
Burgunder JM, Varriale A, Lauterburg BH. Effect of N-acetylcysteine on plasma cysteine and glutathione 
following paracetamol administration. Eur J Clin Pharmacol 1989; 36: 127-131 
7.
De Caro L, Ghizzi A, Costa R, Longo A, Ventresca GP, Lodola L.  Pharmacokinetics and bioavailability of 
oral acetylcysteine in healthy volunteers. Arzneim-Forsh/Drug Res 1989; 39: 382-386 
8.
Borgstrom L, Kagedal B. Dose dependent pharmacokinetics of N-acetylcysteine after oral dosing to man. 
Biopharmaceutics & Drug Disposition 1990; 11: 131-136 
9.
Frascio M. Pharmacokinetic study after single dose (1, 2, and 4 g) of a new N-acetylcysteine (NAC) oral 
formulation. Zambon Group Internal Report, code ZPM/94/34 
10. Crestani S. Open label, randomized, cross-over, pharmacokinetic study to assess the bioequivalence of two 
oral N-Acetylcysteine preparations: 600 mg film coated tablets versus 600 mg effervescent tablets. 2002 
Report in Zambon Group’s file 
11. Rusca A. Study CRO-PK-14-286 – Sponsor code Z7169J02. Comparative bioavailability study of N-acetyl-
cysteine (NAC) 600 mg uncoated tablets vs. NAC 600 mg film-coated tablets in healthy male and female 
volunteers. CROSS Research S.A., Switzerland 2014 
12. Rusca A. Study CRO-PK-07-189 – Sponsor code 7169J01. Comparative bioavailability study of a new oral 
N-acetyl-cysteine (NAC) formulation vs. Fluimucil®, Zambon in healthy volunteers. CROSS Research S.A., 
Switzerland, 2008 
13. Rusca A. Study CRO-PK-09-225 – Sponsor code 7203J01. Pilot bioavailability study of a new 1200 mg N-
acetyl-cysteine (NAC) effervescent tablet formulation vs. Fluimucil® 600 mg effervescent tablet (Zambon 
S.p.A., Italy) in healthy male and female volunteers. CROSS Research S.A., Switzerland, 2010 
14. Brown M, Bjorksten A, Medved I and M McKenna. Pharmacokinetics of intravenous N-acetylcysteine in men 
at rest and during exercise. Eur J Clin Pharmacol 2004;60:717-723 
15. Jones AL, Jarvier D. Simpson D. Pharmacokinetics of N-acetylcysteine are altered in patients with chronic 
liver diseases. Aliment Pharmacol Ther. 1997; 787-791 
16. Prescott LF, Donovan JW, Jarvie DR, Proudfoot AT. The disposition and kinetics of intravenous N-
acetylcysteine in patients with paracetamol overdosage. Eur J Clin Pharmacol 1989; 37: 501-506 
17. Pendyala L, Creaven PJ. Pharmacokinetic and pharmacodynamic studies of N-acetylcysteine, a potential 
chemopreventing agent during a phase I trial. Cancer Epidemiology, Biomarkers and Prevention 1995; 4: 245-
251 
18. Rodenstein D, De Coster A, Gazzaniga A. Pharmacokinetics of oral acetylcysteine: absorption, binding and 
metabolism in patients with respiratory disorders. Clinical Pharmacokinetics 1978; 3: 247-254 
19. Pharmacokinetics of N-Acetylcysteine following repeated i.v. infusions to hemodialysed chronic renal failure 
patients. Internal Report 2002. Zambon Group’s file 


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20. Sjodin K, Nilsson E, Hallemberg A, Tunek A. Metabolism of N-acetyl-l-cysteine Biochem. Pharmacol 38 
1989; 22: 3981-3985 
21. Locatelli C. Fluimucil 20% injectable (N-Acetylcysteine). Expert Report on the clinical documentation 1996. 
Zambon Group’s file 
22. N-acetyl-L-cysteine (NAC) intravenous Investigator’s Brochure, edition number 1, 20 April 2018 
23. Guidelines on GMP, published by the Commission in The rules governing medicinal products in the European 
Community, Volume 4 
24. Guidance on the investigation of bioequivalence. CPMP/EWP/QWP/1401/98 Rev. 1/Corr **, 20 January 2010  
25. U.S. Department of Health and Human Services and U.S. Department of Agriculture, Nutrition and your 
health: 2015-2020 Dietary Guidelines, Appendix 9 


CONFIDENTIAL 
 
 
Sponsor code Z7244J01 
NAC 300 mg/3 mL i.v. bioavailability 
Final version 3.0, 3Dec2019 
 
Page 1 
 
CLINICAL STUDY PROTOCOL 
 
 
Sponsor code Z7244J01 
 
 
 
 
Phase I study to evaluate pharmacokinetics, safety and tolerability 
of single and multiple i.v. doses of N-acetylcysteine (NAC) in 
Chinese healthy volunteers 
 
Single and multiple dose, single centre, open-label, one-way, pharmacokinetics, safety and 
tolerability clinical trial 
 
 
 
Test formulation: 
NAC 300 mg/ 3 mL solution for injection, Zambon S.p.A., Italy. 
 
 
Sponsor: 
Zambon S.p.A., via Lillo del Duca 10, I-20091 Bresso, Italy 
Phone: +39.02.66.52.41 
Fax: 
+39.02.66.50.14.92 
 
 
Investigator: 
- Principal investigator 
Ruijin Hospital Affiliated to Shanghai Jiao Tong University 
School of Medicine, Phase I Unit, No.197, Rui Jin Er Road, 
Shanghai 
China 
Email 
 
 
 
Development phase: 
I 
 
 
Version and date: 
Final version 3.0, 3 Dec 2019  
 
 
 
This study will be conducted in accordance with the current version of Good Clinical Practice (GCP),  
ICH topic E6 (R2 and CFDA GCP) 
 
Property of the sponsor 
May not be used, divulged, published or otherwise disclosed without the consent of the sponsor 
 
 
 
PPD
PPD


PPD
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NAC 300 mg/3 mL i.v. bioavailability 
Final version 3.0, 3Dec2019 
 
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2 
STUDY SYNOPSIS 
 
Title: Phase I study to evaluate pharmacokinetics, safety and tolerability of single and multiple i.v. doses of  
N-acetylcysteine (NAC) in Chinese healthy volunteers  
 
Protocol number: Z7244J01 
 
Clinical phase: Phase I 
 
Study design: Single and multiple dose, single centre, open-label, one-way, pharmacokinetics, safety and 
tolerability clinical trial 
 
Planned nr. of centres / countries: 1/China 
 
Investigator and centre: Principal investigator: 
, Ruijin Hospital Affiliated to Shanghai 
Jiao Tong University School of Medicine, Phase I Unit, No.197, Rui Jin Er Road, Shanghai, China 
 
Investigational medicinal product (IMP):  
Test product: 
NAC, 300 mg/ 3 mL solution for injection, Zambon S.p.A., Italy 
 
Dose regimen:  
Two ampoules (300 + 300 mg) corresponding to a total dose of 600 mg of NAC diluted in 10 mL of NaCl 0.9% 
saline solution, will be administered by a 5-minute intravenous (i.v.) infusion. 
Single dose: 
One (1) dose of the investigational product will be administered under fasting conditions on day 1 at 08:00 ±1 h 
Multiple dose regime: 
Five (5) doses of the investigational product will be administered twice a day (b.i.d.) on days 4 and 5 at 08:00 ±1 
h and 20:00 ±1 h and one dose will be administered on day 6 at 08:00 ±1. 
 
Objective: To evaluate the NAC pharmacokinetics, safety and tolerability after single and multiple dose i.v. 
administration. 
 
End-points:  
Primary end-point:  
➢ To evaluate pharmacokinetic parameters of NAC in plasma after single and multiple dose administration of 
the investigational product.  
Secondary end-points:  
➢ To collect safety and tolerability data after single and multiple dose administration of the investigational 
product. 
 
Study variables: 
Primary variables:  
After single dose: 
➢ Cmax: maximum NAC plasma concentration 
➢ tmax: time to achieve Cmax 
➢ kel: terminal elimination rate constant, calculated, if feasible, by log-linear regression using at least 3 points 
➢ t1/2: NAC half-life, calculated, if feasible, as ln2/kel 
➢ AUC(0-t): area under the concentration-time curve from single dose to the last observed concentration time t, 
calculated with the linear up/log down trapezoidal method 
➢ AUC(0-): area under the concentration-time curve extrapolated to infinity, calculated, if feasible, as AUC(0-t) 
+ Ct/kel, where Ct is the last measurable drug concentration 
➢ AUC(0-12): area under the concentration-time curve at steady-state in the tau interval (from single dose to 12 h 
post-dose), calculated with the linear up/log down trapezoidal method 
➢ Vd: volume of distribution associated with the terminal slope, calculated, if feasible, as Dose/(AUC(0-)*kel) 
➢ CLt: total body clearance, calculated, if feasible, as Dose/ AUC(0-) 
➢ Ae(0-t): total amount of NAC excreted in urine from single dose up to 32 h  
➢ Fe(0-t): total fraction of NAC dose excreted in urine from single dose up to 32 h 
➢ CLr: renal clearance, calculated, if feasible, as Ae(0-t)/ AUC(0-) 
 
 
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Final version 3.0, 3Dec2019 
 
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STUDY SYNOPSIS (cont.) 
 
Study variables (continued): 
After multiple dose: 
➢ Css_max: maximum NAC plasma concentration at steady-state 
➢ tss_max: time to achieve Css_max 
➢ Css_min: trough NAC plasma concentration at steady-state, measured as concentration at t=12 h  
➢ AUCss(0-t): area under the concentration-time curve at steady-state from the last multiple dose to the last 
observed concentration time t, calculated with the linear up/log down trapezoidal method 
➢ AUCss(0-12): area under the concentration-time curve at steady-state in the tau interval (from the last multiple 
dose to 12 h post-dose), calculated with the linear up/log down trapezoidal method 
➢ Css_av: average NAC plasma concentration at steady-state, calculated as AUCss(0-12) /tau  
➢ R: accumulation ratio, calculated as AUCss(0-12)/ AUC(0-12) 
➢ DF%: 
degree 
of 
fluctuation 
over 
one 
dosing 
interval 
at 
steady-state, 
calculated 
as  
(Css_max - Css_min)/ Css_av*100 
➢ Aess(0-t): total amount of NAC excreted in urine from the last multiple dose to 32 h at steady-state 
Secondary variables:  
➢ Treatment emergent adverse events (TEAEs), vital signs (blood pressure, heart rate), body weight, physical 
examinations, clinical laboratory parameters. 
 
Analytics: total NAC will be determined in plasma and urine at a certified bioanalytical laboratory to be 
designated, using a LC-MS/MS validated method with a Lower Quantification Limit (LQL) of 10 ng/mL. 
Analytical facilities and procedures are in compliance with the general principles of GLP regulations 
 
Sample size: Twenty-four (24) healthy male and female Chinese volunteers will be included in the study. Drop-
out subjects will not be replaced. 
 
Main selection criteria:  
Inclusion criteria: 
1. Informed consent: signed written informed consent before inclusion in the study  
2. Ethnicity, Sex and Age: Chinese males and females, 18-45 year old inclusive 
3. Weight: body weight ≥50 kg; 
4. Body Mass Index: 19-26 kg/m2 inclusive 
5. Vital signs: systolic blood pressure 100-139 mmHg, diastolic blood pressure 50-89 mmHg, heart rate 50-90 
bpm, measured after 5 min at rest in the sitting/supine position (to be chosen according to the usual procedure 
at the clinical site) 
6. Full comprehension: ability to comprehend the full nature and purpose of the study, including possible risks 
and side effects; ability to co-operate with the investigator and to comply with the requirements of the entire 
study 
7. Nicotine addiction (smoker subjects only): ability to abstain from smoking for the duration of the clinical study 
8. Contraception and fertility (women only): women of child-bearing potential must be using at least one of the 
following reliable methods of contraception: 
a. Hormonal oral, implantable, transdermal, or injectable contraceptives for at least 60 calendar days before 
the screening visit 
b. A non-hormonal intrauterine device or female condom with spermicide or contraceptive sponge with 
spermicide or diaphragm with spermicide or cervical cap with spermicide for at least 60 calendar days 
before the screening visit 
c. A male sexual partner who agrees to use a male condom with spermicide 
d. A sterile sexual partner 
Women of non-child-bearing potential or in post-menopausal status for at least 1 year will be admitted. 
Women of childbearing potential should be willing to adopt abstinence or contraception measures during the 
study and two weeks post-dose. 
For all women, pregnancy test result must be negative at screening and day -1. 
 
 


CONFIDENTIAL 
 
 
Sponsor code Z7244J01 
NAC 300 mg/3 mL i.v. bioavailability 
Final version 3.0, 3Dec2019 
 
Page 6 
STUDY SYNOPSIS (cont.) 
 
Exclusion criteria: 
1. Electrocardiogram (12-lead ECG in supine position): clinically significant abnormalities 
2. Physical findings: clinically significant abnormal physical findings which could interfere with the objectives 
of the study 
3. Laboratory analyses: clinically significant abnormal laboratory values indicative of physical illness, in 
particular significant laboratory abnormality indicative of hepatic condition (more than 3 times the upper 
limit) 
4. Allergy: ascertained or presumptive hypersensitivity to the active principle and/or formulations' ingredients; 
history of anaphylaxis to drugs or allergic reactions in general, which the investigator considers may affect 
the outcome of the study 
5. Diseases: significant history of renal, hepatic, gastrointestinal, cardiovascular, respiratory, skin, 
haematological, endocrine, urologic, metabolic, neurological or psychiatric diseases, as determined by the 
investigator, that may interfere with the aim of the study; history of carcinoma in situ and malignant disease; 
active bacterial or viral infection and fever >38° C within 48 h prior to study treatment administration 
6. Virology: positive result of HIV, hepatitis B (HBV), hepatitis C (HCV) or Treponema pallidum (TP) assays 
7. Surgery: any surgery within 2 months of screening (excluding diagnostic surgery) 
8. Medications: medications, including over the counter (OTC) medications, herbal remedies and traditional 
Chinese remedies for 2 weeks before the start of the study. Hormonal contraceptives for women will be 
allowed 
9. Investigative drug studies: participation in the evaluation of any investigational product for 30 calendar days 
before this study 
10. Blood donation: blood donations for 90 calendar days before this study 
11. Drug, alcohol, caffeine, tobacco: history of drug, alcohol [>1 drink/day for females and >2 drinks/day for 
males, defined according to the USDA Dietary Guidelines 2015-2020] caffeine (>5 cups coffee/tea/day) or 
tobacco abuse (≥10 cigarettes/day) 
12. Abuse drug test: positive urine abuse drug test at screening or day -1 
13. Alcohol test: positive alcohol breath test at day -1 
14. Diet: abnormal diets (<1600 or >3500 kcal/day) or substantial changes in eating habits in the 4 weeks before 
this study; vegetarians; consumption of alcohol, grapefruit, products containing grapefruit, or beverages 
containing xanthines (coffee, tea, soda, coffee with milk, energy drinks) within 48 hours prior to the 
enrolment  
15. Pregnancy (females only): positive or missing pregnancy test at screening or day -1, pregnant or lactating 
women 
16. Vaccination within 4 weeks of study treatment 
17. Other unspecified reasons that, in the opinion of the investigator, make the subject unsuitable for enrolment. 
 
Schedule: 
 
Day 
Procedures/Assessments 
Notes 
Screening – Visit 1 
From day -14 to 
day -2 
➢ Explanation to the subject of study aims, procedures 
and possible risks 
➢ Informed consent signature 
➢ Screening number (e.g. 
, etc.) 
➢ Demographic data and life style recording 
➢ Medical/surgical history 
➢ Previous/concomitant medications 
➢ Full physical examination (body weight, height, vital 
signs, physical abnormalities) 
➢ ECG recording 
➢ Clinical laboratory analyses: haematology, blood 
chemistry, urinalysis, serum virology, bacteriology and 
pregnancy test (women) 
➢ Urine multi-drug kit test 
➢ Adverse event (AE) monitoring 
➢ Inclusion/exclusion criteria evaluation 
➢ Eligibility evaluation 
 
The subject screening 
numbers will be in 
running 
order 
(e.g. 
 
, 
etc.). 
 
 
CCI
CCI


CONFIDENTIAL 
 
 
Sponsor code Z7244J01 
NAC 300 mg/3 mL i.v. bioavailability 
Final version 3.0, 3Dec2019 
 
Page 7 
STUDY SYNOPSIS (cont.) 
 
Schedule (continued): 
 
Day 
Procedures/Assessments 
Notes 
Visit 2 
Day -1 
➢ Alcohol breath test 
➢ Urine multi-drug kit test 
➢ Urine pregnancy test (women) 
➢ Inclusion/exclusion criteria evaluation 
➢ Eligibility evaluation 
➢ Enrolment 
➢ Subject study number will be the screening number if 
successfully enrolled (e.g. 
, etc.) 
➢ Check of AEs and concomitant medications  
Arrival 
at 
the 
clinic 
before 
the 
evening 
(before 18:30)  
Confinement until the 
morning of day 8 
Standardised dinner 
Fasting for at least 10 h 
(overnight) before first 
dosing 
Visit 3 
Day 1  
➢ Vital sign measurement at pre-dose 
➢ IMP administration at 08:00 ± 1 h; intravenous infusion 
for 5 min 
➢ Vital signs measurement at 1 h post-dose 
➢ Blood sample collection for pharmacokinetic analysis 
at: pre-dose (0) and 5 (at the end of the infusion), 8, 12, 
15, 20, 25, 30, 60 min, 2, 4, 6, 8, 10 and 12 h post-dose 
➢ Urine sample collection for pharmacokinetic analysis 
at: 0-4; 4-8; 8-12 and 12-24 h post-dose 
➢ Check of AEs and concomitant medications  
Standardized lunch at 
13:00  
standardized dinner at 
21:00  
Day 2 
➢ Blood sample collection for pharmacokinetic analysis 
at 24 and 32 h post-dose 
➢ Urine sample collection for pharmacokinetic analysis at 
24-32 h post-dose 
➢ Check of AEs and concomitant medications 
Standardized breakfast 
around 9:00  
Standardized 
lunch 
around 13:00  
Standardized 
dinner 
around 21:00  
Day 3 
➢ Vital sign measurement at 48 h post-dose 
➢ Physical 
examination 
(body 
weight, 
physical 
abnormalities) at 48 h post-dose 
➢ Clinical laboratory analyses: haematology, blood 
chemistry, urinalysis at 48 h post-dose  
➢ Check of AEs and concomitant medications 
Standardized breakfast 
around 9:00  
Standardized 
lunch 
around 13:00  
Standardized 
dinner 
around 21:00 
Visit 4 
From day 4 to 
day 5 
➢ IMP administration at 08:00 ± 1 h; infusion for 5 min 
➢ IMP administration at 20:00 ± 1 h; infusion for 5 min 
➢ Blood sample collection for pharmacokinetic analysis 
at: pre-dose (0) before the morning infusion  
➢ Check of AEs and concomitant medications 
Standardized breakfast 
around 9:00  
Standardized 
lunch 
around 13:00  
Standardized 
dinner 
around 21:00  
 
 
 
CCI


CONFIDENTIAL 
 
 
Sponsor code Z7244J01 
NAC 300 mg/3 mL i.v. bioavailability 
Final version 3.0, 3Dec2019 
 
Page 8 
STUDY SYNOPSIS (cont.) 
 
Schedule (continued): 
 
Day 
Procedures/Assessments 
Notes 
Visit 5 
Day 6 
➢ Vital sign measurement at pre-dose 
➢ IMP administration at 08:00 ± 1 h; infusion for 5 min 
➢ Vital signs measurement at 1 h post-dose 
➢ Blood sample collection for pharmacokinetic analysis 
at: pre-dose (0) and 5 (at the end of the infusion), 8, 12, 
15, 20, 25, 30, 60 min, 2, 4, 6, 8, 10 and 12 h post-dose 
➢ Urine sample collection for pharmacokinetic analysis 
at: 0-4; 4-8; 8-12 and 12-24 h post-dose 
➢ Check of AEs and concomitant medications  
Standardized lunch at 
13:00 (about 5 h post-
dose) 
Standardized dinner at 
21:00 (about 13 h post-
dose)  
Day 7 
➢ Blood sample collection for pharmacokinetic analysis 
at 24 and 32 h post-dose 
➢ Urine sample collection for pharmacokinetic analysis at 
24-32 h post-dose 
➢ Check of AEs and concomitant medications 
Standardized breakfast 
around 9:00  
Standardized 
lunch 
around 13:00  
Standardized 
dinner 
around 21:00  
Final Visit/ETV 
Day 8 or early 
termination visit 
(ETV) in case of 
discontinuation 
➢ Vital sign measurement at 48 h post-dose (or at ETV) 
➢ Physical 
examination 
(body 
weight, 
physical 
abnormalities) 
➢ Clinical laboratory analyses: haematology, blood 
chemistry, urinalysis  
➢ Check of AEs and concomitant medications 
 
In case of clinically significant results at the final visit, the 
subjects will be followed-up by the investigator until the 
normalisation of the concerned clinical parameter(s) 
Discharge 
from 
the 
clinical centre in the 
morning of Day 8 after 
vital sign measurement, 
blood 
sampling 
for 
clinical laboratory assays 
and physical examination 
or in case of ETV. Upon 
leaving, the subjects will 
be instructed to contact 
immediately 
the 
investigator in case of 
occurrence 
of 
any 
untoward 
medical 
occurrence 
Life style and constraints: 
During the study, the subjects will be confined from the evening preceding the first drug administration (study day 
-1) until the morning of day 8 (after the final visit). 
The subjects will not take any food or drinks (except water) for at least 10 h (i.e. overnight) before the first drug 
administration (Day 1) and will remain fasted up to 5 h post-dose.  
During the study, water will be allowed as desired. Coffee, tea or food containing xanthines (i.e. coffee, tea, soda, 
coffee with milk, energy drinks coke, chocolate, etc.), alcohol and grapefruit will be forbidden starting 48 h prior 
to the enrolment until the end of the study. Smoking is not allowed for the whole study duration.  
 
 
 
 


CONFIDENTIAL 
 
 
Sponsor code Z7244J01 
NAC 300 mg/3 mL i.v. bioavailability 
Final version 3.0, 3Dec2019 
 
Page 9 
STUDY SYNOPSIS (cont.) 
 
Withdrawal of subjects: 
It will be documented whether or not each subject completes the clinical study. In case of premature 
discontinuation of any subject, the primary reason for discontinuation will be recorded. 
➢ Adverse event: Any (significant) adverse event that in the opinion of the investigator or concerned subject is 
not compatible with study continuation 
➢ death: the absence of life or state of being dead 
➢ lost to follow-up: the loss or lack of continuation of a subject up to follow-up 
➢ non-compliance with study drug: an indication that a subject has not agreed with or followed the instructions 
related to the study medication 
➢ physician decision: a position, opinion or judgment reached after consideration by a physician with 
reference to the subject 
➢ pregnancy: pregnancy is the state or condition of having a developing embryo or fetus in the body (uterus), 
after union of an ovum and spermatozoon, during the period from conception to birth 
➢ protocol deviation: an event or decision that stands in contrast to the guidelines set out by the protocol 
➢ study terminated by sponsor: an indication that a clinical study was stopped by its sponsor 
➢ technical problems: a problem with some technical aspect of a clinical study, usually related to an 
instrument 
➢ withdrawal by subject: study discontinuation requested by a subject for whatever reason 
For any subject discontinuing the study, the investigator will ask the subject to undergo, as far as possible, a final 
medical visit (ETV) to examine the subject's health conditions and perform the required blood sampling for the 
laboratory assays. This examination will verify that all values tested at screening have remained within a clinically 
acceptable range (i.e. not clinically significant changes compared to screening) and report in the case report form 
(CRF) date and time of the last dose administration, and date and primary reason of study discontinuation. 
 
Data analysis:  
The data documented in this trial and the measured clinical parameters will be presented using classic descriptive 
statistics (i.e. total number of subjects treated [N], number of observations [n], mean standard deviation [SD], 
minimum [Min], median, maximum [Max]) for quantitative variables and frequencies (i.e. count and percentages) 
for qualitative variables if not stated otherwise. 
Analysis set:  
Enrolled set: all enrolled subjects. This analysis set will be used for demographic, baseline and background 
characteristics 
Safety set: all subjects who receive at least one dose of IMP. This analysis set will be used for the safety analyses 
PK set: all enrolled subjects who fulfil the study protocol requirements in terms of IMP administration and have 
evaluable pharmacokinetic data readouts for the planned analysis, with no major deviations that may affect the 
pharmacokinetic results. This analysis set will be used for the statistical analysis of the pharmacokinetic results 
Safety Assessments: 
The statistical analysis of demographic and safety data will be performed using the analysis software SAS®   . 
The toxicity grading of laboratory tests is determined based on the NCI CTCAE V4.03. All AEs, adverse drug 
reactions and serious adverse events will be summarised by system organ class, severity and relationship to study 
drug. 
Pharmacokinetics: 
The statistical analysis of PK parameters will be performed using the validated software Phoenix® WinNonlin® 
version 6.3 or higher (Certara USA, Inc., Princeton, NJ) (actual version will be stated in the final report).   
The individual plasma/urine concentration and pharmacokinetic parameters will be presented in listings and their 
descriptive statistics summarised in tables. 
 
 
 
 
 


CONFIDENTIAL 
 
 
Sponsor code Z7244J01 
NAC 300 mg/3 mL i.v. bioavailability 
Final version 3.0, 3Dec2019 
 
Page 10 
STUDY SYNOPSIS (cont.) 
Background:  
N-acetyl-L-cysteine (NAC) is the N-acetyl derivative of the naturally occurring amino acid L-cysteine.  
NAC was introduced into the Italian pharmaceutical market in 1965 and it is now manufactured and marketed by 
Zambon S.p.A. in Europe, Asia, South America, Central America, and the Middle East, under various trade names 
(Ventresca 1989). 
NAC is widely known as a mucolytic agent, with a well-established use in the treatment of respiratory tract 
disorders. However, after more than 35 years from its introduction in the clinical practice, this molecule continues 
to be the object of scientific interest due to its potential in the treatment of several conditions other than those 
related to the respiratory system. NAC can also act as a direct antioxidant agent due to the -SH group (Auroma 
1988) and can easily penetrate into the cells where it is deacetylated to L-cysteine, thus supporting the biosynthesis 
of glutathione (Ziment 1988). 
Clinical pharmacology and pharmacokinetics 
Several pharmacokinetic (PK) studies were carried out in healthy volunteers (Borgstrom 1986, Olsson 1988, 
Burgunder 1989, De Caro 1989, Borgstrom 1990, Frascio, Crestani 2002, Rusca 2014, Rusca 2010, Rusca 2002). 
Further PK studies were carried out in patients with chronic liver damage (Jones 1997), with liver damage caused 
by paracetamol overdose (Prescott 1989), in subjects under long-term treatment for chemoprevention (Pendyala 
1995), in patients with respiratory disorders (Rodenstein 1978) and in patients with End Stage Renal Diseases 
(Internal report 2002). 
In the clinical studies, NAC was administered orally and intravenously both as single and repeated dose. Oral NAC 
was administered in different dosage forms, such as capsules, granulate, effervescent tablets, fast dissolving tablets, 
slow-release tablets. 
Clinical trials and safety concerns 
NAC has been available to patients as a mucolytic agent for many years and has a well-established and satisfactory 
safety and tolerability profile (see IB).  
Independently of route of administration (inhalation, oral, intravenous) adverse events are reported rarely 
following NAC administration and are usually mild and transient.  
The most frequent adverse events associated with NAC oral administration are gastrointestinal (e.g., vomiting, 
diarrhoea, stomatitis, abdominal pain and nausea). Hypersensitivity reactions (e.g. anaphylactoid reactions, 
urticaria, rash, pruritus, and bronchospasm) have been reported less frequently after oral administration, while with 
the IV administration they were reported more frequently compared to gastrointestinal symptoms. Some of the 
hypersensitivity reactions were associated to higher infusion rate. Most anaphylactoid reactions can be managed 
by temporarily suspending the acetylcysteine infusion, administering appropriate supportive care. A cumulative 
analysis of all safety reports received by the Sponsor up to September 2017 for NAC IV, confirmed the nature and 
severity of the events were in line with the above profile of the product. 
Intravenous NAC has been studied in clinical trials and published case series both as mucolytic agent and as an 
antidote against intoxication by paracetamol and other toxic agents including Amanita phalloides and chlorinated 
hydrocarbons. Overall, in published studies in humans, a total of 745 patients were exposed to i.v. NAC at doses 
ranging from 500 mg/day to 980mg/kg/day for a duration of 1 to 18 days.  Details of safety results are included in 
the IB. 
In published studies, whereas i.v. NAC as mucolytic agent typically was given at doses up to 1000 mg/day, much 
higher i.v. doses of NAC as antidote were given over multiple days, maintaining a satisfactory safety profile. 
Locatelli and colleagues published two case series, one of 86 patients with acute intoxication from Amanita 
phalloides and the other of 35 patients with acute intoxication from chlorinated hydrocarbons. Patients received 
i.v. NAC 300 mg/kg/day for 3 to 14 days and 3 to 18 days respectively. The safety and tolerability profile were 
overall benign, with 4 out of 121 exposed patients suffering from anaphylactoid reaction or rash (due to the rapid 
administration of the bolus according to the authors), none of which caused discontinuation of treatment. No other 
AEs were reported. (Locatelli 1996). 
 
Rationale 
NAC is widely known as a mucolytic agent, with a well-established use in the treatment of respiratory tract 
disorders. In the present study, the kinetic profile and the bioavailability of NAC will be investigated in Chinese 
healthy male and female volunteers all receiving the same single i.v. dose, i.e. a total dose of 600 mg of NAC, and 
the same multiple dose administrations, i.e. five i.v. doses of 600 mg of NAC administered b.i.d. for 2 days and 
once on the last day. The safety and tolerability will be monitored as well.  
The present study will be part of the clinical development plan of NAC i.v. for registration in China of this 
formulation. 


CONFIDENTIAL 
 
 
Sponsor code Z7244J01 
NAC 300 mg/3 mL i.v. bioavailability 
Final version 3.0, 3Dec2019 
 
Page 11 
3 
STUDY SCHEDULE 
 
ACTIVITIES 
Screening 
Single Dose 
Multiple Dose 
Final 
visit/ETV1 
Visit 
V1 
V2 
V3 
V4 
V5 
 Day -14/-2 
Day -1 
Day 1 
Day 2 
Day 3 
Day 4 
Day 5 
Day 6 
Day 7 
Day 82 
Informed consent 
x 
 
 
 
 
 
 
 
 
 
Demography and lifestyle 
x 
 
 
 
 
 
 
 
 
 
Medical and surgical history  
x 
 
 
 
 
 
 
 
 
 
Physical examination 
x 
 
 
 
x 
 
 
 
 
x 
Prior and concomitant medications 
x 
x 
x 
x 
x 
x 
x 
x 
x 
x 
Height 
x 
 
 
 
 
 
 
 
 
 
Body Weight 
x 
 
 
 
x 
 
 
 
 
x 
Clinical laboratory analysis 
(haematology, blood chemistry, 
urinalysis)  
x 
 
 
 
x 
 
 
 
 
x 
Virology and bacteriology  
x 
 
 
 
 
 
 
 
 
 
Serum pregnancy test (women) 
x 
 
 
 
 
 
 
 
 
 
Urine multi-drug kit test 
x 
x 
 
 
 
 
 
 
 
 
Blood pressure and heart rate 
x 
 
x3 
 
x 
 
 
x3 
 
x4 
Alcohol breath test 
 
x 
 
 
 
 
 
 
 
 
Urine pregnancy test (women) 
 
x 
 
 
 
 
 
 
 
 
ECG 
X 
 
 
 
 
 
 
 
 
 
Inclusion/exclusion criteria  
X 
x 
 
 
 
 
 
 
 
 
Subject eligibility 
X 
x 
 
 
 
 
 
 
 
 
Enrolment  
 
x 
 
 
 
 
 
 
 
 
Confinement 
 
x 
x 
x 
x 
x 
x 
x 
x 
 
Discharge 
 
 
 
 
 
 
 
 
 
x 
Investigational product administration 
 
 
x5 
 
 
x6 
x6 
x5 
 
 
Blood sampling  
 
 
x7 
x7 
 
x8 
x8 
x7 
x7 
 
Urine sampling  
 
 
x9 
x9 
 
 
 
x9 
x9 
 
Standardised meals 
 
x10 
x11 
x12 
x12 
x12 
x12 
x11 
x12 
 
Adverse event monitoring13 
x 
x 
x 
x 
x 
x 
x 
x 
x 
x 
 


CONFIDENTIAL 
 
 
Sponsor code Z7244J01 
NAC 300 mg/3 mL i.v. bioavailability 
Final version 3.0, 3Dec2019 
 
Page 12 
1. 
Early termination visit (ETV) 
2. 
Final visit on day 8 or in case of ETV 
3. 
At pre-dose and 1 h post-dose 
4. 
The vital signs check at 48 h post-dose (day 8), will correspond to the final measurement 
5. 
At 8:00 ± 1 h: 5-min infusion 
6. 
At 8:00 ± 1 h and at 20:00 ± 1 h; 5-min infusion 
7. 
At pre-dose (0) and 5 (at the end of the infusion), 8, 12, 15, 20, 25, 30, 60 min and 2, 4, 6, 8, 10, 12, 24 and 32 h post-dose  
8. 
At pre-dose (0) 
9. 
0-4; 4-8; 8-12, 12-24 and 24-32 h post-dose  
10. Standardised dinner 
11. Standardised lunch and dinner 
12. Standardised breakfast, lunch and dinner  
13. Adverse events monitored starting at the screening visit, immediately after informed consent, up to the final visit/ETV 
 


CONFIDENTIAL 
 
 
Sponsor code Z7244J01 
NAC 300 mg/3 mL i.v. bioavailability 
Final version 3.0, 3Dec2019 
 
Page 13 
 
4 
TABLE OF CONTENTS 
 
Page 
1 
CLINICAL STUDY PROTOCOL 
1 
2 
STUDY SYNOPSIS 
4 
3 
STUDY SCHEDULE 
11 
4 
TABLE OF CONTENTS 
13 
4.1 
TABLES 
15 
5 
LIST OF ABBREVIATIONS 
16 
6 
Introduction 
18 
6.1 
Background 
18 
6.2 
Clinical pharmacology and pharmacokinetics 
18 
6.3 
Clinical trials and safety concerns 
19 
6.4 
Rationale 
20 
6.5 
Risk and benefits 
20 
7 
STUDY OBJECTIVES 
21 
7.1 
Primary end-point 
21 
7.2 
Secondary end-point 
21 
8 
CLINICAL SUPPLIES 
22 
8.1 
Treatment 
22 
8.1.1 
Description of products 
22 
8.1.1.1 
Test product 
22 
8.1.2 
Dose regimen 
23 
8.1.3 
Route and method of administration 
23 
8.1.4 
Investigational product distribution 
23 
8.2 
Packaging and labelling 
23 
8.3 
Storage conditions 
24 
8.4 
Drug accountability 
24 
9 
INVESTIGATIONAL PLAN 
25 
9.1 
Overall study design 
25 
9.2 
Discussion of design 
25 
10 
STUDY POPULATION 
26 
10.1 
Target population 
26 
10.2 
Inclusion criteria 
26 
10.3 
Exclusion criteria 
27 
10.3.1 
Not allowed treatments 
28 
11 
STUDY SCHEDULE 
29 
11.1 
Study visits and procedures 
29 
11.2 
Diet and lifestyle 
31 
11.2.1 
Restrictions 
32 
12 
DESCRIPTION OF SPECIFIC PROCEDURES 
33 
12.1 
Physical examination 
33 
12.1.1 
Body weight 
33 
12.1.2 
Vital signs 
33 
12.1.3 
ECGs 
33 
12.2 
Clinical laboratory assays 
33 
12.3 
Sampling for pharmacokinetic analysis 
34 
12.3.1 
Venous blood sampling 
34 
12.3.2 
Urine collection 
35 
12.3.3 
Analytics 
36 
12.3.4 
Labelling, storage and transport of samples 
36 
12.3.4.1 Samples labelling 
36 
12.3.4.2 Samples storage and transport 
36 
13 
ASSIGNMENT OF STUDY TREATMENT 
38 
13.1 
Randomisation 
38 


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Final version 3.0, 3Dec2019 
 
Page 14 
13.2 
Blinding 
38 
14 
EVALUATION PARAMETERS 
39 
14.1 
Study variables 
39 
14.1.1 
Primary variables 
39 
14.1.2 
Secondary variables 
39 
14.2 
Pharmacokinetic assessments 
39 
14.2.1 
Pharmacokinetic parameters 
39 
14.3 
Safety assessments 
41 
15 
STATISTICAL METHODS 
42 
15.1 
Analysis Sets 
42 
15.1.1 
Definitions 
42 
15.1.2 
Reasons for exclusion from the PK set before bioanalysis 
43 
15.2 
Sample size and power considerations 
43 
15.3 
Demographic, baseline and background characteristics 
43 
15.4 
Analysis of pharmacokinetic parameters 
43 
15.4.1 
Descriptive pharmacokinetics 
43 
15.5 
Safety and tolerability evaluation 
43 
16 
DEFINITION AND HANDLING OF AEs AND SAEs 
45 
16.1 
Applicable SOPs 
45 
16.2 
Definition of Adverse Event (AE) 
45 
16.3 
Definition of Adverse Drug Reaction (ADR) 
45 
16.4 
Definition of Serious Adverse Events or Serious Adverse Drug Reaction 
46 
16.5 
Definition of Severity of Adverse Events 
46 
16.6 
Definition of Adverse Event causality 
46 
16.7 
Adverse Events recording 
47 
16.8 
AEs monitoring window 
47 
16.9 
Adverse Events reporting 
47 
16.9.1 
SAEs reporting 
48 
16.10 
Follow-up for Adverse Events 
48 
16.11 
SUSARs management 
49 
16.12 
Other events qualified for expedited reporting 
49 
16.13 
SAEs: contacts 
50 
16.14 
Pregnancy 
50 
17 
DATA MANAGEMENT PROCEDURES 
51 
17.1 
Data collection – CRFs 
51 
17.2 
Database management 
51 
17.2.1 
Coding dictionaries 
51 
18 
STUDY MONITORING, QUALITY CONTROL AND QUALITY ASSURANCE 
52 
18.1 
Monitoring 
52 
18.2 
Quality Control and Quality Assurance 
52 
18.3 
Applicable SOPs 
53 
18.4 
Data access 
53 
18.5 
Audits and inspections 
53 
19 
ETHICAL CONSIDERATIONS 
54 
19.1 
Ethics and Good Clinical Practice (GCP) 
54 
19.2 
Informed consent 
54 
19.3 
Insurance policy 
55 
19.4 
Withdrawal of subjects 
55 
19.4.1 
Primary reason for discontinuation 
55 
19.5 
Study termination 
56 
20 
ADMINISTRATIVE PROCEDURES 
57 
20.1 
Material supplied to the clinical centre 
57 
20.2 
Protocol amendments 
57 
20.3 
Study documentation and record keeping 
57 
20.4 
Study subjects’ recruitment 
58 
20.5 
Confidentiality and data protection 
58 
20.6 
Publication policy 
58 
21 
STUDY RESPONSIBLE PERSONS 
60 


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Final version 3.0, 3Dec2019 
 
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21.1 
Sponsor 
60 
21.2 
Drug assay 
60 
21.3 
Project management, data analysis & reporting, monitoring 
60 
22 
REFERENCES 
61 
 
 
 
4.1 
TABLES 
Page 
 
Table 11.1.1 
Schedule of study activities and procedures 
30 
Table 12.3.1.1 
Tolerance ranges for the scheduled sampling times 
35 
 
 


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5 
LIST OF ABBREVIATIONS 
 
-HCG 
human chorionic gonadotropin  
-GT 
-Glutamyl transpeptidase 
ADR 
Adverse Drug Reaction 
AE 
Adverse Event 
ALT 
Alanine aminotransferase 
Ae0-t 
Total amount of NAC excreted in urine  
Aess(0-t) 
Total amount of NAC excreted in urine at steady-state 
ANOVA 
Analysis of Variance 
AST 
Aspartate aminotransferase 
BB 
Blue book 
BLQL 
Below Lower Quantification Limit 
BUN 
Blood Urea Nitrogen 
AUC(0-t) 
Area under the concentration-time curve from single dose to the last observed concentration 
time t 
AUC(0-) 
Area under the concentration-time curve extrapolated to infinity 
AUC(0-12) 
Area under the concentration-time curve at steady-state in the tau interval  
AUCss(0-t) 
Area under the concentration-time curve at steady-state from the last multiple dose to the last 
observed concentration time t 
AUCss(0-12) 
Area under the concentration-time curve at steady-state in the tau interval  
BMI 
Body Mass Index 
BP 
Blood Pressure 
CA 
Competent Authority 
CDISC 
Clinical Data Interchange Standards Consortium 
CFDA 
China Food And Drug Administration 
CI 
Confidence Interval 
Cmax 
Peak drug concentration 
CLt 
Total body clearance 
CLr 
Renal clearance 
CPL 
Clinical Project Leader 
CRA 
Clinical Research Associate 
CRF 
Case Report Form 
CRO 
Contract Research Organisation 
CS 
Clinically Significant 
CSP 
Clinical Study Protocol 
CSR 
Clinical Study Report 
Css_av 
Average NAC plasma concentration at steady-state 
Css_max 
Maximum plasma concentration at steady-state 
Css_min 
Trough plasma concentration at steady-state 
CV 
Coefficient of Variation 
DBP 
Diastolic Blood Pressure 
DF 
Degree of fluctuation  
DSU 
Drug safety unit 
DTT 
Dithiothreitol 
EC 
Ethics Committee 
ECG 
Electrocardiogram 
ETV 
Early Termination Visit 
FDA 
Food and Drug Administration 
Fe0-t 
Total fraction of NAC dose excreted in urine  
Frel 
Relative Bioavailability 
FSFV 
First Subject First Visit 
GCP 
Good Clinical Practice 
GLP 
Good Laboratory Practice 
HBs Ag 
Hepatitis B virus surface antigen 
HCV Ab 
Hepatitis C virus antibodies 
HIV 
Human Immunodeficiency Virus 


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HR 
Heart Rate 
IB 
Investigator’s Brochure 
ICH 
International Conference on Harmonisation 
IRB/IEC 
Institutional Review Board/Independent Ethics Committee 
i.m. 
Intramuscular 
IMP 
Investigational Medicinal Product 
IUD 
Intra-Uterine Device 
i.v. 
Intravenous 
kel 
Terminal elimination rate constant 
LQL 
Lower Quantification Limit 
LSLV 
Last Subject Last Visit 
MCH 
Mean Cell Haemoglobin 
MCHC 
Mean Cell Haemoglobin Concentration 
MCV 
Mean Cell Volume 
MedDRA 
Medical Dictionary for Regulatory Activities 
MRT 
Mean Residence Time 
MW 
Molecular Weight 
N 
Normal 
NA 
Not Applicable 
NAC 
N-acetyl-L-cysteine 
NC 
Not calculated 
NCS 
Not clinically significant 
NSAIDs 
Non-Steroidal Anti-Inflammatory Drugs 
OTC 
Over The Counter 
PE 
Point Estimate 
PK 
Pharmacokinetics 
PT 
Preferred Term 
PTAE 
Pre-Treatment Adverse Event 
R 
Accumulation ratio 
RBC 
Red Blood Cells 
RSI 
Reference safety information 
SAE 
Serious Adverse Event 
SBP 
Systolic Blood Pressure 
SD 
Standard Deviation 
SOC 
System Organ Class 
SOP 
Standard Operating Procedure 
SDTM 
Study Data Tabulation Model 
SUSAR 
Suspected Unexpected Serious Adverse Reaction 
T 
Test 
TEAE 
Treatment-Emergent Adverse Event 
THC 
delta-9-tetrahydrocannabinol 
t1/2 
Half-life 
tmax 
Time to achieve Cmax 
tss_max 
Time to achieve Css_max 
USDA 
United States Department of Agriculture 
Vd 
Volume of distribution  
WBC 
White Blood Cells 
WHODDE 
World Health Organisation Drug Dictionary Enhanced 
 
 
 
 


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6 
INTRODUCTION 
 
6.1 
Background 
 
N-acetyl-L-cysteine (NAC) is the N-acetyl derivative of the naturally occurring amino acid L-
cysteine.  
 
NAC was introduced into the Italian pharmaceutical market in 1965 and it is now manufactured 
and marketed by Zambon S.p.A. in Europe, Asia, South America, Central America, and the 
Middle East, under various trade names (1). 
 
NAC is widely known as a mucolytic agent, with a well-established use in the treatment of 
respiratory tract disorders. However, after more than 35 years from its introduction in the 
clinical practice, this molecule continues to be the object of scientific interest due to its potential 
in the treatment of several conditions other than those related to the respiratory system. NAC 
can also act as a direct antioxidant agent due to the -SH group (2) and can easily penetrate into 
the cells where it is deacetylated to L-cysteine, thus supporting the biosynthesis of glutathione 
(3). 
 
6.2 
Clinical pharmacology and pharmacokinetics 
 
Several pharmacokinetic (PK) studies were carried out in healthy volunteers (4, 5, 6, 7, 8, 9, 
10, 11, 12, 13, 14). Further PK studies were carried out in patients with chronic liver damage 
(15), with liver damage caused by paracetamol overdosage (16), in subjects under long-term 
treatment for chemoprevention (17), in patients with respiratory disorders (18) and in patients 
with End Stage Renal Diseases (19). 
In the clinical studies, NAC was administered orally and intravenously both as single and 
repeated doses. Oral NAC was administered in different dosage forms, such as capsules, 
granulate, effervescent tablets, fast dissolving tablets, slow-release tablets. 
 
In particular, Borgström et al. (4) studied for the first time the PK profile of NAC after i.v. dose 
of 600 mg of NAC infused over 5 min in 10 healthy male and female volunteers in comparison 
with 3 other oral dosage forms. The PK parameters obtained by the authors after i.v. infusion 
are summarised in the following table.  
 
Table 6.2.1 
PK parameters of plasma free NAC (N=10) 
Cmax 
(µmol/L) 
tmax 
(h) 
CL 
(L/kgxh) 
CLr 
(L/kgxh) 
t½ 
(h) 
Ae0-12 
(% of dose) 
16.0±7.9 
0.65±0.33 
0.207±0.017 
0.058±0.011 
2.27±0.32 
29.0±3.2* 
mean±SD is reported; *: N=9; Source: 4 
 
However, the authors analysed NAC in deproteinised plasma thus missing the protein-bound 
NAC in their measurements.  
 
Olsson et al. (5) improved the bioanalytical method and were able to measure the total NAC by 
reduction of the disulphide bonds in plasma before precipitating proteins. After single i.v. dose 
of 200 mg of NAC to 6 healthy volunteers, the authors found the following PK parameters: 


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Table 6.2.2 
PK parameters of plasma total NAC (N=6) 
 
Cmax 
(µM) 
Vss 
(L/kg) 
CL 
(L/kgxh) 
T 
121 (82.9 – 162) 
0.47 (0.46 – 0.55) 
0.11 (0.09 – 1.13) 
median (range) is reported; Source: 5 
 
Jones et al. (15) studied the bioavailability of NAC after single i.v. dose of 600 mg of NAC 
infused over 3 min to patients with chronic liver disease, but also to 6 healthy male and female 
volunteers as control subjects. The authors found the following PK parameters for total NAC: 
 
Table 6.2.3 
PK parameters of plasma total NAC (N=6) 
AUC 
(mg/Lxh) 
CLr 
(L/h) 
t½ 
(h) 
Vdss 
(L) 
93.9±9.6 
6.5±0.8 
2.6±0.3 
17.4±2.8 
mean±SD is reported; Source: 15 
 
Brown et al. (14) investigated the PK of NAC infused i.v. to 24 healthy men at rest and during 
exercise. NAC was infused at the dose of 125 mg/kgxh for 15 min followed by 25 mg/kgxh for 
35 min to healthy men at rest. Then, the infusion continued during exercise until fatigue. The 
authors found NAC peak concentrations in plasma at the end of the initial loading infusion. 
Mean peak concentration of total NAC was 205.1±68.3 mg/L, while mean CL was 0.164 
L/kgxh.  
 
It is worth noting that, while scientific literature shows that PK profiles of NAC after oral 
administration are very consistent (in terms of half-life, rate and extent of exposure, 
bioavailability, dose-linearity, etc.) among various publications, this is not the case for the data 
obtained after i.v. dosing. In fact, for this administration route, published studies differ in 
several key aspects, i.e., for example, type of administration, applied bioanalytical methods, 
drug doses, calculation methods for PK parameters. Furthermore, the number of subjects 
enrolled in each published study is very small. For these reasons, the comparability of the 
literature data is low among the published studies and potentially also to the expected results of 
the present study.   
 
6.3 
Clinical trials and safety concerns 
 
NAC has been available to patients as a mucolytic agent for many years and has a well-
established and satisfactory safety and tolerability profile (see IB).  
Independently of route of administration (inhalation, oral, intravenous) adverse events are 
reported rarely following NAC administration and are usually mild and transient.  
The most frequent adverse events associated with NAC oral administration are gastrointestinal 
(e.g., vomiting, diarrhoea, stomatitis, abdominal pain and nausea). Hypersensitivity reactions 
(e.g. anaphylactoid reactions, urticaria, rash, pruritus, and bronchospasm) have been reported 
less frequently after oral administration, while with the i.v. administration they were reported 
more frequently compared to gastrointestinal symptoms. Some of the hypersensitivity reactions 
were associated to higher infusion rate. Most anaphylactoid reactions can be managed by 
temporarily suspending the acetylcysteine infusion, administering appropriate supportive care. 


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A cumulative analysis of all safety reports received by the Sponsor up to September 2017 for 
NAC IV, confirmed the nature and severity of the events were in line with the above profile of 
the product. 
Intravenous NAC has been studied in clinical trials and published case series both as mucolytic 
agent and as an antidote against intoxication by paracetamol and other toxic agents including 
Amanita phalloides and chlorinated hydrocarbons. Overall, in published studies in humans, a 
total of 745 patients were exposed to i.v. NAC at doses ranging from 500 mg/day to 
980mg/kg/day for a duration of 1 to 18 days.  Details of safety results are included in the IB. 
In published studies, whereas i.v. NAC as mucolytic agent typically was given at doses up to 
1000 mg/day, much higher i.v. doses of NAC as antidote were given over multiple days, 
maintaining a satisfactory safety profile. 
Locatelli and colleagues published two case series, one of 86 patients with acute intoxication 
from Amanita phalloides and the other of 35 patients with acute intoxication from chlorinated 
hydrocarbons. Patients received i.v. NAC 300 mg/kg/day for 3 to 14 days and 3 to 18 days 
respectively. The safety and tolerability profile were overall benign, with 4 out of 121 exposed 
patients suffering from anaphylactoid reaction or rash (due to the rapid administration of the 
bolus according to the authors), none of which caused discontinuation of treatment. No other 
AEs were reported (21). 
 
6.4 
Rationale 
 
NAC is widely known as a mucolytic agent, with a well-established use in the treatment of 
respiratory tract disorders. 
In the present study, the kinetic profile and the bioavailability of NAC will be investigated in 
Chinese healthy male and female volunteers all receiving the same single i.v. dose, i.e. a total 
dose of 600 mg of NAC, and the same multiple dose administrations, i.e. five i.v. doses of 600 
mg of NAC administered b.i.d. for 2 days and once on the last day. The safety and tolerability 
will be monitored as well.  
The present study will be part of the clinical development plan of NAC i.v. for registration in 
China of this formulation. 
 
6.5 
Risk and benefits 
 
NAC is a well-known drug which has been used for decades.  
 
Undesired effects which may occur during treatment with NAC include: anaphylactic shock, 
anaphylactic reaction, anaphylactoid reaction, hypersensitivity, tachycardia, bronchospasm, 
dyspnoea, vomiting, nausea, angioedema, urticaria, flushing rash, pruritus, face oedema, blood 
pressure decreased, prothrombin time prolonged (for details refer to IB; 22).  
 
Blood sampling with cannula insertion may cause minor discomfort. The risks associated with 
blood draws include pain, bleeding and bruising. 
 
No specific benefits for the participants in the current study are foreseen. Their remuneration 
will be paid after study completion. The remuneration covers loss of time and any 
inconvenience caused by the participation in the study. 


CONFIDENTIAL 
 
 
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Final version 3.0, 3Dec2019 
 
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7 
STUDY OBJECTIVES 
 
The objective of the study is to evaluate the NAC pharmacokinetics, safety and tolerability after 
single and multiple dose i.v. administration. 
 
7.1 
Primary end-point 
 
➢ To evaluate pharmacokinetic parameters of NAC in plasma after single and multiple dose 
administration of the investigational product.  
 
7.2 
Secondary end-point 
 
➢ To collect safety and tolerability data after single and multiple dose administration of the 
investigational product. 


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8 
CLINICAL SUPPLIES 
 
8.1 
Treatment 
 
All the subjects enrolled in the study will receive the same treatment with the investigational 
medicinal product (IMP), i.e. NAC, 300 mg/ 3 mL solution for injection, as follows: 
 
➢ on day 1 at 08:00 ±1 h, one dose of 600 mg of NAC (300 + 300 mg ampoule) will be 
administered under fasting conditions; 
 
After a wash-out of 3 days: 
 
➢ on days 4 and 5 at 08:00 ±1 h and 20:00 ±1 h and at 08:00 ±1 on day 6, 5 doses of 600 mg 
of NAC (300 + 300 mg ampoule) will be administered. 
 
8.1.1 
Description of products 
 
8.1.1.1 
Test product 
 
IMP 
NAC 300 mg/ 3 mL solution for injection  
Active substance 
N-acetyl-L-cysteine 
Manufacturer 
(active substance) 
F.I.S. Fabbrica Italiana Sintetici S.p.A.,  
Via Dovaro, 36045 Lonigo (Vicenza), Italy 
(GMP compliant)  
Manufacturer 
(finished product) 
Zambon S.p.A.,  
Via della Chimica 9, 36100 Vicenza, Italy 
(GMP compliant)  
Pharmaceutical form 
Solution for injection 
Dose 
300 mg/ 3 mL 
Administration route 
Parenteral  
 
The analytical certificates will be supplied with the IMP. Quali-quantitative formulation is as 
follows: 
 
Each 3 mL vial (10%) contains: 
➢ N-acetyl-L-cysteine (NAC) 300 mg 
➢ sodium hydroxide 74 mg 
➢ disodium edetate 3 mg 
➢ water for injections q.s. to 3 mL 
 
 
 


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8.1.2 
Dose regimen 
 
Two ampoules of IMP (300 + 300 mg) corresponding to a total dose of 600 mg of NAC diluted 
in 10 mL of NaCl 0.9% sterile saline solution, will be administered by a 5-minute i.v. infusion. 
 
Single dose: 
 
One (1) dose of IMP will be administered under fasting conditions on day 1 at 08:00 ±1 h. 
 
Multiple dose regime: 
 
Five (5) doses of IMP will be administered twice a day (b.i.d.) on days 4 and 5 at 08:00 ±1 h 
and 20:00 ±1 h and one dose will be administered on day 6 at 08:00 ±1.  
 
8.1.3 
Route and method of administration 
 
Each dose of IMP will be prepared by diluting the content of 2 ampoules of IMP in 10 mL of 
saline, as described above (§ 8.1.2), and administered to each study subject at the clinical site 
only by the investigator or his/her deputy. 
 
Each dose will be infused intravenously over 5 min.  
 
The start of infusion (not the end) will be considered as time 0. 
 
8.1.4 
Investigational product distribution 
 
The IMP will be exclusively used for the present clinical study and will only be administered 
to the subjects enrolled in the study.  
 
8.2 
Packaging and labelling 
 
The IMP primary packaging will be glass vials. 
 
The formulation labelling will report all the information requested according to the Annex 13 
to the Good Manufacturing Practice (published by the Commission in The rules governing 
medicinal products in the European Community, Volume 4; 23) and in compliance with 
applicable laws and regulations as follows: 
a. 
Name, address and telephone number of the sponsor, contract research organisation or 
investigator (the main contact for information on the product, clinical study and any 
emergency) 
b. Pharmaceutical dosage form, route of administration, quantity of dosage units, and the 
name and strength 
c. 
The batch and/or code number to identify the contents and packaging operation 
d. A study reference code allowing identification of the study, site, investigator and sponsor 
if not given elsewhere 


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e. 
The study subject identification number/treatment number and where relevant, the visit 
number 
f. 
The name of the investigator (if not included in (a) or (d)) 
g. Directions for use (reference may be made to a leaflet or other explanatory document 
intended for the study subject or person administering the product) 
h. “For clinical study use only” or similar wording 
i. 
The storage conditions 
j. 
Period of use (use-by date, expiry date or re-test date as applicable), in month/year format 
and in a manner that avoids any ambiguity 
k. “Keep out of reach of children” 
 
Labels will be in local language. 
 
8.3 
Storage conditions 
 
The IMP will be stored at ≤25° C in a dry locked place, sheltered from light.  
 
8.4 
Drug accountability 
 
The IMP will be provided directly to the investigator by the Sponsor or designee, in excess of 
the amount necessary for the study (at least 25% excess). 
 
After receipt of the IMP supply, the pharmacist will confirm in writing by signing and dating 
standard drug accountability forms. 
 
At the end of the study, used, unused and partially used supplies of IMP provided by the Sponsor 
or designee will either be destroyed on site (upon written authorisation) or returned to the 
Sponsor or designee, after assessment of drug accountability. 
 
 


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9 
INVESTIGATIONAL PLAN 
 
9.1 
Overall study design 
 
This is a single and multiple dose, single centre, open-label, one-way, pharmacokinetics, safety 
and tolerability clinical trial of Phase I to be performed in Chinese healthy male and female 
volunteers. 
 
9.2 
Discussion of design 
 
The study has been designed in agreement with the Chinese Technical Guideline on Clinical 
Pharmacokinetic Research of Chemical Drugs, 18 March 2005 and the European Guideline on 
the Investigation of Bioequivalence (CPMP/QWP/EWP/1401/98 Rev. 1, 20 January 2010) (24).  
 
An open-label design is used since the primary end-point of the study is based on objective 
measurements of NAC in blood. The outcome variables are not influenced by the subjects or 
investigator being aware of the administered products. 
 
Blood sampling time-points were selected on the basis of the known PK profile of NAC (4, 5, 
6, 7, 8, 9, 10, 11, 12, 13, 14). 
 
The dose of 600 mg has been selected since this is the efficacious, safe and well tolerated dose 
used in clinical practice (see also 22).  
 
The bioanalysis will be performed in compliance with GCP and CFDA GCP regulations and in 
accordance with the applicable principles of GLP, as defined by OECD, in a GLP compliant 
facility. Moreover, sample analysis will be conducted in compliance with the Chinese Guidance 
on Bioanalysis: method validation and analysis of study samples (2015).  
 


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10 
STUDY POPULATION 
 
10.1 
Target population 
 
Chinese healthy male and female volunteers will be included in the study.  
 
10.2 
Inclusion criteria 
 
To be enrolled in this study, subjects must fulfil all these criteria: 
 
1. Informed consent: signed written informed consent before inclusion in the study  
2. Ethnicity, Sex and Age: Chinese males and females, 18-45 year old inclusive 
3. Weight: body weight ≥50 kg; 
4. Body Mass Index: 19-26 kg/m2 inclusive 
5. Vital signs: systolic blood pressure 100-139 mmHg, diastolic blood pressure 50-89 
mmHg, heart rate 50-90 bpm, measured after 5 min at rest in the sitting/supine position (to 
be chosen according to the usual procedure at the clinical site) 
6. Full comprehension: ability to comprehend the full nature and purpose of the study, 
including possible risks and side effects; ability to co-operate with the investigator and to 
comply with the requirements of the entire study 
7. Nicotine addiction (smoker subjects only): ability to abstain from smoking for the duration 
of the clinical study 
8. Contraception and fertility (women only): women of child-bearing potential must be using 
at least one of the following reliable methods of contraception: 
a. Hormonal oral, implantable, transdermal, or injectable contraceptives for at least 60 
calendar days before the screening visit 
b. A non-hormonal intrauterine device or female condom with spermicide or 
contraceptive sponge with spermicide or diaphragm with spermicide or cervical cap 
with spermicide for at least 60 calendar days before the screening visit 
c. A male sexual partner who agrees to use a male condom with spermicide 
d. A sterile sexual partner 
Women of non-child-bearing potential or in post-menopausal status for at least 1 year will 
be admitted. 
Women of childbearing potential should be willing to adopt abstinence or contraception 
measures during the study and two weeks post-dose. 
For all women, pregnancy test result must be negative at screening and day -1. 
 
 


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10.3 
Exclusion criteria 
 
Subjects meeting any of these criteria will not be enrolled in the study: 
 
1. Electrocardiogram (12-lead ECG in supine position): clinically significant abnormalities 
2. Physical findings: clinically significant abnormal physical findings which could interfere 
with the objectives of the study 
3. Laboratory analyses: clinically significant abnormal laboratory values indicative of 
physical illness, in particular significant laboratory abnormality indicative of hepatic 
condition (more than 3 times the upper limit) 
4. Allergy: ascertained or presumptive hypersensitivity to the active principle and/or 
formulations' ingredients; history of anaphylaxis to drugs or allergic reactions in general, 
which the investigator considers may affect the outcome of the study 
5. Diseases: significant history of renal, hepatic, gastrointestinal, cardiovascular, respiratory, 
skin, haematological, endocrine, urologic, metabolic, neurological or psychiatric diseases, 
as determined by the investigator, that may interfere with the aim of the study; history of 
carcinoma in situ and malignant disease; active bacterial or viral infection and fever >38° 
C within 48 h prior to study treatment administration 
6. Virology: positive result of HIV, hepatitis B (HBV), hepatitis C (HCV) or Treponema 
pallidum (TP) assays 
7. Surgery: any surgery within 60 calendar days of screening (excluding diagnostic surgery) 
8. Medications: medications, including over the counter (OTC) medications, herbal remedies 
and traditional Chinese remedies for 2 weeks before the start of the study. Hormonal 
contraceptives for women will be allowed 
9. Investigative drug studies: participation in the evaluation of any investigational product for 
1 month before this study 
10. Blood donation: blood donations for 90 calendar days before this study 
11. Drug, alcohol, caffeine, tobacco: history of drug, alcohol [>1 drink/day for females and >2 
drinks/day for males, defined according to the USDA Dietary Guidelines 2015-2020] 
caffeine (>5 cups coffee/tea/day) or tobacco abuse (≥10 cigarettes/day) 
12. Abuse drug test: positive urine abuse drug test at screening or day -1 
13. Alcohol test: positive alcohol breath test at day -1 
14. Diet: abnormal diets (<1600 or >3500 kcal/day) or substantial changes in eating habits in 
the 4 weeks before this study; vegetarians; consumption of alcohol, grapefruit, products 
containing grapefruit, or beverages containing xanthines (coffee, tea, soda, coffee with 
milk, energy drinks) within 48 hours prior to the enrolment  
15. Pregnancy (females only): positive or missing pregnancy test at screening or day -1, 
pregnant or lactating women 
16. Vaccination within 4 weeks of study treatment 
17. Other unspecified reasons that, in the opinion of the investigator, make the subject 
unsuitable for enrolment. 


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10.3.1 
Not allowed treatments 
 
No medication, including OTC, traditional Chinese medicine and herbal remedies, will be 
allowed for 2 weeks before the start of the study and during the whole study duration. 
Paracetamol will be allowed as therapeutic counter-measure for adverse events (AEs) according 
to the investigator’s opinion. Hormonal contraceptives will be allowed.  
 
The intake of any other medication will be reported as a protocol deviation.  
 
 


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11 
STUDY SCHEDULE 
 
The schedule of the study is summarised at page 11. 
 
11.1 
Study visits and procedures 
 
Each study subject will undergo 6 visits. 
 
The study protocol foresees a screening visit and a confinement lasting from Visit 2 to the Final 
Visit/early termination visit (ETV), i.e. from Day -1 to Day 8. Maximum study duration will be 
22 days, screening visit included. A written informed consent will be obtained before any study 
assessment or procedure. 
 
The first subject first visit (FSFV) is defined as the 1st visit performed at the clinical centre on 
the 1st screened subject. The last subject last visit (LSLV) is defined as the last visit performed 
at the clinical centre (or the telephonic follow-up, if applicable) on the last subject, i.e. the last 
visit foreseen by the study protocol, independently of the fact that the subject is a completer or 
a withdrawn subject. 
 
The following phases, visits and procedures will be performed: 
 
➢ Screening phase 
▪ Screening – visit 1: between day -14 and day -2 
▪ Visit 2: day -1 
 
➢ Interventional phase 
▪ Visit 3: days 1-3 
▪ Visit 4: days 4-5 
▪ Visit 5: days 6-7 
 
➢ Final phase 
▪ Visit 6: day 8 - Final visit/ETV. In case of early discontinuation, discontinued subjects 
will undergo an ETV 
 
Activities to be performed are listed by visit in the following table:  


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Table 11.1.1 Schedule of study activities and procedures 
 
Day 
Procedures/Assessments 
Notes 
Screening – Visit 1 
From day -14 to 
day -2 
➢ Explanation to the subject of study aims, procedures 
and possible risks 
➢ Informed consent signature 
➢ Screening number (e.g. 
, etc.) 
➢ Demographic data and life style recording 
➢ Medical/surgical history 
➢ Previous/concomitant medications 
➢ Full physical examination (body weight, height, vital 
signs, physical abnormalities) 
➢ ECG recording 
➢ Clinical laboratory analyses: haematology, blood 
chemistry, urinalysis, serum virology, bacteriology and 
pregnancy test (women) 
➢ Urine multi-drug kit test 
➢ AE monitoring 
➢ Inclusion/exclusion criteria evaluation 
➢ Eligibility evaluation 
The subject screening 
numbers (e.g. 
 
, etc.) will be 
used to identify the 
subjects 
Visit 2 
Day -1 
➢ Alcohol breath test 
➢ Urine multi-drug kit test 
➢ Urine pregnancy test (women) 
➢ Inclusion/exclusion criteria evaluation 
➢ Eligibility evaluation 
➢ Enrolment 
➢ Study subject number will be the same as the screening 
number that was initially assigned when the subject is 
successfully enrolled (e.g screening number .) 
➢ Check of AEs and concomitant medications  
Arrival 
at 
the 
clinic 
before 
the 
evening 
(before 18:30)  
Confinement until the 
morning of day 8 
Standardised dinner 
Fasting for at least 10 h 
(overnight) before first 
dosing 
Visit 3 
Day 1  
➢ Vital sign measurement at pre-dose 
➢ IMP administration at 08:00 ± 1 h; intravenous infusion 
for 5 min 
➢ Vital signs measurement at 1 h post-dose 
➢ Blood sample collection for pharmacokinetic analysis 
at: pre-dose (0) and 5 (at the end of the infusion), 8, 12, 
15, 20, 25, 30, 60 min, 2, 4, 6, 8, 10 and 12 h  
post-dose 
➢ Urine sample collection for pharmacokinetic analysis 
at: 0-4; 4-8; 8-12 and 12-24 h post-dose 
➢ Check of AEs and concomitant medications  
Standardized lunch at 
13:00  
standardized dinner at 
21:00  
 
Day 2 
➢ Blood sample collection for pharmacokinetic analysis 
at 24 and 32 h post-dose 
➢ Urine sample collection for pharmacokinetic analysis at 
24-32 h post-dose 
➢ Check of AEs and concomitant medications 
Standardized breakfast 
around 9:00  
Standardized 
lunch 
around 13:00  
Standardized 
dinner 
around 21:00  
Day 3 
➢ Vital sign measurement at 48 h post-dose 
➢ Physical 
examination 
(body 
weight, 
physical 
abnormalities) at 48 h post-dose 
➢ Clinical laboratory analyses: haematology, blood 
chemistry, urinalysis at 48 h post-dose  
➢ Check of AEs and concomitant medications 
 
Standardized breakfast 
around 9:00  
Standardized 
lunch 
around 13:00  
Standardized 
dinner 
around 21:00 
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NAC 300 mg/3 mL i.v. bioavailability 
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Day 
Procedures/Assessments 
Notes 
Visit 4 
From day 4 to 
day 5 
➢ IMP administration at 08:00 ± 1 h; infusion for 5 min 
➢ IMP administration at 20:00 ± 1 h; infusion for 5 min 
➢ Blood sample collection for pharmacokinetic analysis 
at: pre-dose (0) before the morning infusion  
➢ Check of AEs and concomitant medications 
Standardized breakfast 
around 9:00  
Standardized 
lunch 
around 13:00  
Standardized 
dinner 
around 21:00  
Visit 5 
Day 6 
➢ Vital sign measurement at pre-dose 
➢ IMP administration at 08:00 ± 1 h; infusion for 5 min 
➢ Vital signs measurement at 1 h post-dose 
➢ Blood sample collection for pharmacokinetic analysis 
at: pre-dose (0) and 5 (at the end of the infusion), 8, 12, 
15, 20, 25, 30, 60 min, 2, 4, 6, 8, 10 and 12 h post-dose 
➢ Urine sample collection for pharmacokinetic analysis 
at: 0-4; 4-8; 8-12 and 12-24 h post-dose 
➢ Check of AEs and concomitant medications  
Standardized lunch at 
13:00 (about 5 h post-
dose) 
Standardized dinner at 
21:00 (about 13 h post-
dose)  
Day 7 
➢ Blood sample collection for pharmacokinetic analysis 
at 24 and 32 h post-dose 
➢ Urine sample collection for pharmacokinetic analysis at 
24-32 h post-dose 
➢ Check of AEs and concomitant medications 
Standardized breakfast 
around 9:00  
Standardized 
lunch 
around 13:00  
Standardized 
dinner 
around 21:00  
Final Visit/ETV 
Day 8 or early 
termination visit 
(ETV) in case of 
discontinuation 
➢ Vital sign measurement at 48 h post-dose (or at ETV) 
➢ Physical 
examination 
(body 
weight, 
physical 
abnormalities) 
➢ Clinical laboratory analyses: haematology, blood 
chemistry, urinalysis  
➢ Check of AEs and concomitant medications 
 
In case of clinically significant results at the final visit, the 
subjects will be followed-up by the investigator until the 
normalisation of the concerned clinical parameter(s) 
Discharge 
from 
the 
clinical centre in the 
morning of Day 8 after 
vital sign measurement, 
blood 
sampling 
for 
clinical 
laboratory 
assays 
and 
physical 
examination or in case 
of ETV. Upon leaving, 
the subjects will be 
instructed to contact 
immediately 
the 
investigator in case of 
occurrence 
of 
any 
untoward 
medical 
occurrence 
 
11.2 
Diet and lifestyle 
 
The subjects will not take any food or drinks (except water) for at least 10 h (i.e. overnight) 
before the first drug administration (Day 1) and will remain fasted up to 5 h post-dose.  
During the study, water will be allowed as desired. In order to maintain an adequate hydration, 
the subjects will be encouraged to drink at least 120 mL of mineral water every 2 h for 6 h post-
dose on Days 1 and 6. 
 
Coffee, tea or food containing xanthines (i.e. coffee, tea, soda, coffee with milk, energy drinks, 
etc.), alcohol and grapefruit will be forbidden starting 48 h before the enrolment until the end 
of the study. Smoking is not allowed for the whole study duration. 
 
During confinement, routine ambulant daily activities will be strongly recommended. 
 


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NAC 300 mg/3 mL i.v. bioavailability 
Final version 3.0, 3Dec2019 
 
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The timing of each meal to be served during confinement is shown in Table11.1.1 above.  
In the evening of day -1, upon confinement, the subjects will receive a light dinner before 
fasting overnight.  
On days 1 and 6 (PK sampling collection days), standardised lunch and dinner will be served.  
On days 2, 3, 4, 5 and 7, standardised breakfast, lunch and dinner will be served.  
 
11.2.1 
Restrictions 
 
During the study, the subjects will be confined from the evening preceding the first drug 
administration (study day -1) until the morning of day 8 (after the final visit). They will attend 
the clinic in the evening of day -1 not later than 18:30.  
 
During confinement, hazardous, strenuous or athletic activities will not be permitted. 


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12 
DESCRIPTION OF SPECIFIC PROCEDURES 
 
12.1 
Physical examination 
 
Full physical examinations will be performed at the screening visit, visit 3, day 3 and final visit/ 
ETV. Information about the physical examination will be recorded by the investigator. Any 
abnormalities will be recorded. 
 
Significant findings/illnesses, reported after the start of the study and that meet the definition 
of an AE (see § 16), will be recorded in the subject source documents. 
 
Date of the physical examination, overall investigator’s interpretation (as normal or abnormal 
and, if abnormal, clinically significant or not clinically significant) and clinically significant 
abnormalities (if any) will be reported in the individual CRFs. 
 
12.1.1 
Body weight 
 
Body weight will be recorded during each physical examination. 
Subjects will be weighed (kg) lightly clothed without shoes. Height will be measured at 
screening only and BMI will be recorded. BMI will be calculated as weight [kg]/(height [m] x 
height [m]). 
 
12.1.2 
Vital signs 
 
Subjects blood pressure (BP) and heart rate (HR) will be measured by the investigator or his/her 
deputy after 5 min at rest (in sitting position) at:  
 
➢ Screening  
➢ Visit 3, Day 1: at pre-dose and 1 h post-dose 
➢ Visit 3, Day 3: at 48 h post-dose  
➢ Visit 5, Day 6: at pre-dose and 1 h post-dose 
➢ Final visit (at 48 h post-dose)/ETV  
 
12.1.3 
ECGs 
 
One 12-lead ECG will be performed (in sitting/supine position) at screening only. 
 
12.2 
Clinical laboratory assays 
 
Samples of blood and urine will be collected. The following laboratory analyses will be 
performed at the screening visit: 
 
Haematology 
Leukocytes and leukocyte differential count (percentage values and absolute values), 
erythrocytes, haemoglobin (conv. units), haemoglobin (IS units), haematocrit, MCV, MCH, 
MCHC, thrombocytes. 
 
 


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Final version 3.0, 3Dec2019 
 
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Blood chemistry 
Electrolytes: sodium, potassium, calcium, chloride, inorganic phosphorus 
Enzymes: alkaline phosphatase, γ-GT, AST, ALT 
Substrates/metabolites: total bilirubin, creatinine, glucose, BUN or urea, uric acid, total 
cholesterol, triglycerides 
Proteins: total proteins 
 
Serum pregnancy test (women). 
 
Urine analysis 
pH, specific gravity, appearance, colour, nitrites, proteins, glucose, urobilinogen, bilirubin, 
ketones, blood, leukocytes, erythrocytes, epithelial cells, crystals, cylinders. 
 
Serum virology and bacteriology 
Hepatitis B (HBs antigen), Hepatitis C (HCV antibodies), HIV 1/2 (HIV Ag/Ab combo), 
Treponema pallidum.  
 
The same analyses, with the exception of virology, bacteriology and serum pregnancy test, will 
be performed also at Visit 3, Day 3 (48 h post-dose after the single dose) and at the final 
visit/ETV. 
 
A urine drug test will be performed at the clinical centre at screening, using a urine multi-drug 
kit. The following drugs will be assessed: cocaine, amphetamine, methamphetamine, 
cannabinoids (delta-9-tetrahydrocannabinol - THC), opiates. The same test will be repeated 
upon confinement at Visit 2, day -1.  
 
A serum pregnancy test will be performed by the laboratory at screening. A urine pregnancy 
test will be performed at Visit 2, day -1 at the clinical centre, upon confinement.  
 
Date/time of samples collection, overall investigator’s interpretation (as normal or abnormal 
and, if abnormal, clinically significant or not clinically significant) and clinically significant 
findings (if any) will be reported in the individual CRFs. All clinically significant abnormalities 
after the screening visit will be recorded as AEs.    
 
12.3 
Sampling for pharmacokinetic analysis 
 
12.3.1 
Venous blood sampling 
 
Venous blood samples (up to 10 mL) will be collected from a forearm vein at the following 
times, both on Day 1, 2, after the single dose, and on Day 6, 7, after the last multiple dose:  
➢ at pre-dose (0) and 5 (at the end of the infusion), 8, 12, 15, 20, 25, 30, 60 min, 2, 4, 6, 8, 
10, 12, 24 and 32 h post-dose 
 
The start of infusion (not the end) will be considered as time 0. 
 
Actual sampling times for each subject will be recorded in the individual case report forms 
(CRFs). The actual sampling times should not exceed the recommended tolerance ranges 
presented in the following table. Any deviation outside the recommended ranges will be verified 


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Final version 3.0, 3Dec2019 
 
Page 35 
through Data Clarification Forms and, if confirmed, will be reported as protocol deviation, 
although it will not automatically lead to the exclusion of the concerned subjects from the PK 
analysis set. 
 
Table 12.3.1.1 
Tolerance ranges for the scheduled sampling times 
Sampling time 
Tolerance range 
Pre-dose (0) 
Within 30 minutes before IMP administration 
0.0833 h (5 min), 0.1333 (8 min) 
0.200 h (12 min), 0.250 h (15 min) 
20 seconds 
0.3333 h (20 min), 0.4167 (25 min), 0.5 h (30 min) 
± 1 min 
1 h 
± 3 min 
2, 4 h 
± 5 min 
6, 8, 10, 12, 24, 32 h 
± 10 min 
 
Blood samples for PK analysis will be collected using an indwelling catheter with switch valve. 
The cannula will be rinsed, after each sampling, with about 1 mL of sterile saline solution. The 
first mL of blood will be discarded at each collection time. 
 
The remaining blood sample will be collected into blood collection tube. 
 
Storage on ice will not last longer than 60 min until centrifugation at 4° C for 10 min at 2500xg 
to obtain plasma. Plasma will be collected into separate tubes, where an appropriate volume of 
DTT solution (50 µL DTT solution/mL plasma, i.e. 5% of the plasma volume according to the 
ratio 75 µL of DTT solution for 1.5 mL of plasma) will be added. The samples will be mixed 
shortly (example Vortex, 20 sec or relevant method to mix evenly inverting the tube up and 
down gently ~ about 20 times), divided into two aliquots, P1 and P2 and transferred to -70° C 
as soon as possible. The DTT solution (5 mg DTT/mL of water) will be freshly prepared in the 
evening before each sampling day and kept refrigerated until use. The preparation of the 
solution will be recorded in appropriate forms. 
 
If any clinical assessment, such as vital signs measurement, is foreseen at the same time-point 
as blood sampling for PK analysis, blood collection will be performed at the scheduled time. 
However, vital signs can be influenced by the blood sampling. Therefore, these assessments 
can be performed within 30 min before the pre-dose PK time point (0 h) and within 10 min 
before the other scheduled PK time-points. Any deviations outside the recommended time will 
be verified through Data Clarification Forms. However, since vital signs measurements will be 
performed for safety reasons only, deviations from the planned time schedule will be considered 
not relevant. 
 
12.3.2 
Urine collection 
 
Urine for PK will be collected in the following time intervals both on Day 1, 2, after the single 
dose, and on Day 6, 7 after the last multiple dose:  
 
➢ 0-4; 4-8; 8-12, 12-24 and 24-32 h post-dose 
 
Bladder must be emptied before the end of each collection period. During each interval, urine 
will be collected into containers, kept refrigerated at approximately 4° C and containing DTT 
solution. At the end of each collection interval, urine volume will be measured and after 


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NAC 300 mg/3 mL i.v. bioavailability 
Final version 3.0, 3Dec2019 
 
Page 36 
thorough mixing, two aliquots of  1 mL each (U1 and U2) will be prepared in polypropylene 
tubes. Urine volume collected in each collection interval will be accurately recorded in 
appropriate forms and in the CRFs. The two aliquots will be stored at -70° C. The DTT 
solution (5 mg DTT/mL of water) will be freshly prepared in the evening before each sampling 
day and kept refrigerated until use. The preparation of the solution will be recorded in 
appropriate forms. 
 
12.3.3 
Analytics  
 
The concentration of total NAC in plasma and urine will be determined at a certified 
bioanalytical laboratory to be designated in China, using a fully validated LC-MS/MS method, 
with a lower quantification limit (LQL) of 10 ng/mL.  
 
Analyses will be performed according to the general Principles of “OECD Good Laboratory 
Practices for testing of chemicals" C(81) 30 (final) and GCP/ CFDA GCP (see also § 9.2). 
 
The method validation report and the analytical report will be attached to the final report. 
 
12.3.4 
Labelling, storage and transport of samples 
 
12.3.4.1 
Samples labelling 
 
Each sample tube will be clearly and unequivocally identified with a label resistant to the 
storage temperature and reporting: 
Study code 
Sponsor code Z7244J01 
Subject number 
Subject or Process or Unique Subject Identification number, e.g. 
, etc. 
Tube identification 
 
Study day 
1 or 6 
Scheduled sampling time as min and h; see § 12.3.1 
 
12.3.4.2 
Samples storage and transport 
 
During the study the samples will be stored at ≤-70° C. At the end of each collection day, 
aliquots 1 and 2 will be stored in separate freezers. 
 
All aliquots 1, packed in sufficient solid CO2, will be shipped by an authorised courier from the 
clinical site, China, to Eurofins Central Laboratory, Shanghai, China. Aliquots 1 will remain 
stored at Eurofins Central Laboratory or a subcontracted bioanalytical laboratory to be 
designated until finalisation of the bioanalytical report. Afterwards, the samples will be 
destroyed and a certificate of destruction will be provided to the sponsor. 
 
The counter-samples (aliquot 2) will remain stored at the clinical site, China. These samples 
could either be: 
➢ sent to the laboratory for reanalysis should this become necessary for analytical reasons or 
if any problems occur during the delivery of aliquots 1, or 
➢ destroyed at an authorised site, or 
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➢ transferred to the sponsor upon written request. 
 
No analyses different from those stated in this protocol and agreed by the subjects when signing 
the informed consent form will be performed unless a new informed consent and a new approval 
from the Ethical Committee is obtained. The subjects may ask to destroy their own samples at 
any time. 
 
 


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NAC 300 mg/3 mL i.v. bioavailability 
Final version 3.0, 3Dec2019 
 
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13 
ASSIGNMENT OF STUDY TREATMENT 
 
13.1 
Randomisation 
 
No randomisation will take place in the present study. All the subjects will receive the same 
treatment.  
 
13.2 
Blinding 
 
This is an open study. No masking procedure will be applied.  
 
 
 
 


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Final version 3.0, 3Dec2019 
 
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14 
EVALUATION PARAMETERS 
 
14.1 
Study variables 
 
14.1.1 
Primary variables 
 
After single dose: 
 
➢ Cmax, tmax, kel, t1/2, AUC(0-t), AUC(0-), AUC(0-12), Vd, CLt, Ae(0-t), Fe(0-t) and CLr 
 
After multiple doses: 
 
➢ Css_max, tss_max, Css_min, AUCss(0-t), AUCss(0-12), Css_av, R, DF% and Aess(0-t). 
 
14.1.2 
Secondary variables 
 
➢ TEAEs, vital signs (BP, HR), body weight, physical examinations, laboratory parameters. 
 
14.2 
Pharmacokinetic assessments 
 
14.2.1 
Pharmacokinetic parameters 
 
The following PK parameters will be measured and/or calculated for plasma NAC, using the 
validated software Phoenix® WinNonlin® version 6.3 or higher (Certara USA, Inc., Princeton, 
NJ) (actual version will be stated in the final report), after single dose of IMP: 
 
Cmax: 
Maximum NAC plasma concentration 
 
tmax: 
Time to achieve Cmax 
 
kel: 
Terminal elimination rate constant, calculated, if feasible, by log-linear 
regression using at least 3 points 
 
t1/2: 
Half-life, calculated, if feasible, as ln2/kel 
 
AUC(0-t): 
Area under the concentration-time curve from single dose to the last observed 
concentration time t, calculated with the linear up/log down trapezoidal method 
 
AUC(0-): 
Area under the concentration-time curve extrapolated to infinity, calculated, if 
feasible, as AUC(0-t) + Ct/kel, where Ct is the last measurable drug concentration 
 
AUC(0-12): 
Area under the concentration-time curve at steady-state in the tau interval (from 
single dose to 12 h post-dose), calculated with the linear up/log down 
trapezoidal method 
 


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Vd: 
Volume of distribution associated with the terminal slope, calculated, if 
feasible, as Dose/(AUC(0-)*kel) 
 
CLt: 
Total body clearance, calculated, if feasible, as Dose/ AUC(0-) 
 
Ae(0-t): 
Total amount of NAC excreted in urine from single dose up to 32 h 
 
Fe(0-t): 
Total fraction of NAC dose excreted in urine from single dose up to 32 h 
 
CLr: 
Renal clearance, calculated, if feasible, as Ae(0-t)/ AUC(0-) 
 
 
The following PK parameters will be measured and/or calculated for plasma NAC, using the 
same software, after multiple doses of IMP: 
 
Css_max: 
Maximum NAC plasma concentration at steady-state 
 
tss_max: 
Time to achieve Css_max 
 
Css_min: 
Trough NAC plasma concentration at steady-state, measured as concentration 
at t=12 h 
 
AUCss(0-t): 
Area under the concentration-time curve at steady-state from the last multiple 
dose to the last observed concentration time t, calculated with the linear up/log 
down trapezoidal method 
 
AUCss(0-12): 
Area under the concentration-time curve at steady-state in the tau interval (from 
the last multiple dose to 12 h post-dose), calculated with the linear up/log down 
trapezoidal method 
 
Css_av: 
Average NAC plasma concentration at steady-state, calculated as AUCss(0-12) 
/tau 
 
R: 
Accumulation ratio, calculated as AUCss(0-12)/ AUC(0-12) 
 
Aess(0-t): 
Total amount of NAC excreted in urine from the last multiple dose to 32 h at 
steady-state 
 
DF%: 
Degree of fluctuation over one dosing interval at steady-state, calculated as 
(Css_max - Css_min)/ Css_av*100 
 
 
The sampling schedule is considered adequate if the ratio AUC(0-t)/ AUC(0-) equals or exceeds 
a factor of 0.8 (i.e. if %AUCextra is <20%) for more than 80% of the individual PK profiles. This 
assures that the primary variable AUC(0-t) covers a sufficient percentage of the theoretical total 
extent of exposure.  
 


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The quality of log-linear regression (and, consequently, the reliability of the extrapolated PK 
parameters) should be demonstrated by a determination coefficient R2 ≥ 0.8. Individual 
extrapolated parameters, when considered unreliable, will be reported as NC (not calculated).  
 
14.3 
Safety assessments 
 
Safety and general tolerability of the IMP will be based on TEAEs, physical examinations 
including body weight, vital signs and routine haematology, blood chemistry and urinalysis 
laboratory tests. 
 


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15 
STATISTICAL METHODS 
 
The data documented in this trial and the measured clinical parameters will be presented using 
classic descriptive statistics (i.e. total number of subjects treated [N], number of observations 
[n], mean standard deviation [SD], minimum [Min], median, maximum [Max]) for quantitative 
variables and frequencies (i.e. count and percentages) for qualitative variables if not stated 
otherwise. 
 
Not available data will be evaluated as “missing values”. The statistical analysis of demographic 
and safety data will be performed using the analysis software SAS®  . 
 
The statistical analysis of PK parameters will be performed using the validated software 
Phoenix® WinNonlin® version 6.3 or higher (Certara USA, Inc., Princeton, NJ) (actual version 
will be stated in the final report). 
 
15.1 
Analysis Sets 
 
15.1.1 
Definitions 
 
A subject will be defined as screened after the signature of the informed consent, regardless of 
the completion of all the screening procedures. 
 
A subject will be defined as eligible if he/she meets all the inclusion/exclusion criteria. 
Otherwise he/she will be defined as a screen failure.  
 
A subject will be defined as enrolled in the study if he/she is included into the interventional 
phase of the study.  
 
The following analysis sets are defined: 
➢ Enrolled set: all enrolled subjects. This analysis set will be used for demographic, baseline 
and background characteristics 
➢ Safety set: all subjects who receive at least one dose of IMP. This analysis set will be used 
for the safety analyses 
➢ PK set: all enrolled subjects who fulfil the study protocol requirements in terms of IMP 
intake and have evaluable PK data readouts for the planned analyses, with no major 
deviations that may affect the PK results. This analysis set will be used for the statistical 
analysis of the PK parameters. 
 
Each subject will be coded as valid or not valid for the safety and the PK set. Subjects will be 
evaluated according to the treatment they actually receive. 
 


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15.1.2 
Reasons for exclusion from the PK set before bioanalysis 
 
Reasons for the exclusion of subjects from the PK set are the following: 
 
➢ intake of concomitant medications which could render the plasma concentration-time 
profile unreliable 
➢ AEs which could render the plasma concentration-time profile unreliable 
➢ administration errors which could render the plasma concentration-time profile unreliable 
➢ other events which could render the plasma concentration-time profile unreliable 
If one of these events occurs, it will be noted in the CRF during the study. 
 
The samples from the subjects excluded from the PK set should still be assayed and the results 
listed. Subjects should not be excluded from the PK set if the AUC0-t covers less than 80% of 
the AUC0-∞. 
 
15.2 
Sample size and power considerations 
 
The sample size was not calculated through any statistical calculation. A sample size of 24 
subjects was estimated as sufficient for the descriptive purposes of the present study. 
Drop-out subjects will not be replaced.  
 
15.3 
Demographic, baseline and background characteristics 
 
Critical demographic characteristics will be examined according to qualitative or quantitative 
data. Qualitative data will be summarised in contingency tables. Quantitative data will be 
summarised using classic descriptive statistics. 
 
15.4 
Analysis of pharmacokinetic parameters 
 
15.4.1 
Descriptive pharmacokinetics 
 
A descriptive PK will be presented. The results will be displayed and summarised in tables and 
figures. Individual and mean curves (+SD at sampling times), indicating inter-subject 
variability, will be plotted. Data below the lower quantification limit (BLQL) will be considered 
as 0 in the calculations and presented as BLQL in listings and tables. As a consequence of 
BLQL (i.e. 0) values, calculated geometric means (if requested) could be null. For this reason, 
in the presence of any null value, the geometric mean will be reported as not calculated (NC). 
 
15.5 
Safety and tolerability evaluation 
 
➢ AEs 
AEs will be coded by System Organ Class (SOC) and Preferred Term (PT), using the Medical 
Dictionary for Regulatory Activities (MedDRA). 
AEs will be classified as pre-treatment AEs (PTAEs) and TEAEs, according to the period of 
occurrence, as follows:  


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Final version 3.0, 3Dec2019 
 
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➢ PTAEs: all AEs occurring before the first dose of IMP and not worsening after the first 
dose of IMP 
➢ TEAEs: all AEs occurring or worsening after the first dose of IMP 
Individual PTAEs and TEAEs will be listed in subject data listings. No summary table will be 
provided for PTAEs. TEAEs will be summarised by treatment and overall. The number and 
percentage of subjects with any TEAE and the number of TEAEs will be tabulated by SOC and 
PT, seriousness, relationship to treatment and severity.  
 
➢ Physical examination 
Date of the physical examination, overall investigator’s interpretation (as normal or abnormal 
and, if abnormal, clinically significant or not clinically significant) and clinically significant 
abnormalities (if any) will be listed. 
 
➢ Laboratory data  
Date/time of samples collection, overall investigator’s interpretation (as normal or abnormal 
and, if abnormal, clinically significant or not clinically significant) and clinically significant 
findings (if any) will be listed. All laboratory results will be listed and a table of all the abnormal 
values will be presented. The overall investigator’s interpretation will be summarised using 
tables of frequency.  
The toxicity grading of laboratory tests is determined based on the NCI CTCAE V4.03. 
 
➢ Vital signs 
Vital signs values will be listed and summarised by descriptive statistics.  
 
➢ Body weight  
Body weight values will be listed and summarised by descriptive statistics. 
 


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Final version 3.0, 3Dec2019 
 
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16 
DEFINITION AND HANDLING OF AEs AND SAEs 
 
16.1 
Applicable SOPs 
 
AEs definition, classification and management will follow the CRO’s SOPs, based upon 
applicable local and international regulations. The full SOPs or an operative summary will be 
made available to the clinical centre. 
 
A brief summary of AE definition, classification and management is reported below. 
 
16.2 
Definition of Adverse Event (AE) 
 
An Adverse Event is “any untoward medical occurrence in a patient or clinical trial subject 
administered a medicinal product and which does not necessarily have a causal relationship 
with this treatment”. 
AEs include: 
➢ worsening (change in nature, severity or frequency) of conditions present at the onset 
of the trial 
➢ patient/subject deterioration due to the primary illness 
➢ intercurrent illnesses 
➢ drug interactions 
➢ events related or possibly related to concomitant medications 
➢ abnormal laboratory values, as well as significant shifts from baseline within the range 
of normal, which the investigator considers to be clinically significant. 
An AE can therefore be any unfavourable and unintended sign (including an abnormal 
laboratory finding), symptom, or disease temporally associated with the use of an 
investigational medicinal product, whether or not considered related to the investigational 
medicinal product. 
 
16.3 
Definition of Adverse Drug Reaction (ADR) 
 
An Adverse Reaction is “any untoward and unintended response to an investigational 
medicinal product related to any dose administered”. 
All AEs judged by either the reporting investigator or the sponsor as having a reasonable causal 
relationship to a medicinal product qualify as adverse reactions. The expression reasonable 
causal relationship means to convey in general that there is evidence or matter to suggest a 
causal relationship. 
 
➢ Unexpected Adverse Drug Reaction 
 
An unexpected ADR is: “An adverse reaction, the nature, or severity of which is not consistent 
with the applicable product information (Reference Safety Information [RSI]).  
 
➢ Reference Safety Information (RSI): in order to assess whether an adverse reaction is 
expected, the IB will be used. 


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16.4 
Definition of Serious Adverse Events or Serious Adverse Drug Reaction 
 
A Serious Adverse Event (SAE) is: “any untoward medical occurrence or effect that at any 
dose: 
➢ results in death 
➢ is life-threatening (i.e. the subject was at risk of death at the time of the event/reaction; it 
does not refer to an event/reaction which hypothetically might have caused death if it were 
more severe) 
➢ requires inpatient hospitalization or prolongation of existing hospitalization 
➢ results in persistent or significant disability/incapacity (where disability is defined as a 
permanent or substantial disruption of ability to carry out normal life functions, either 
reported or defined as per clinical judgement) 
➢ is a congenital anomaly/birth defect 
➢ is an important medical event that may not result in death, be life-threatening, or require 
hospitalization but, according to appropriate medical judgment, it may jeopardize the 
subject and may require medical or surgical intervention to prevent any of the outcomes 
listed in the definition above.  Examples of such events include allergic bronchospasm 
requiring intensive treatment in an emergency room or at home, and blood dyscrasias or 
convulsions that do not result in hospitalization, or development of drug dependency or 
drug abuse.  
 
Any suspected transmission via a medicinal product of an infectious agent is also considered a 
serious adverse event. 
 
A Suspected Unexpected Serious Adverse Reaction (SUSAR) is an ADR that is both 
unexpected (not consistent with the applicable product information, e.g. IB) and also meets the 
definition of a SAE. 
 
A non-serious adverse event is any adverse event that does not meet the criteria listed above for 
a serious adverse event. 
 
16.5 
Definition of Severity of Adverse Events 
 
The term “severe” is used to describe the intensity (severity) of a specific event: 
- 
Mild: causing no limitation of usual activities; the subject may experience slight 
discomfort. 
- 
Moderate: causing some limitation of usual activities; the subject may experience 
annoying discomfort. 
- 
Severe: causing inability to carry out usual activities; the subject may experience 
intolerable discomfort or pain. 
 
16.6 
Definition of Adverse Event causality 
 
Causality shall be determined according to the definition of ADR given in 16.3. 
 


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All AE judged by either the investigator or the sponsor as having a reasonable suspected causal 
relationship to an investigational medicinal product qualify as adverse reactions. The causality 
assessment given by the investigator should not be downgraded by the sponsor. 
 
The following binary decision for causality will be used: 
▪ Reasonable possibility that the IMP caused the event 
▪ No reasonable possibility that the IMP caused the event 
Features supportive of an association include: 
• temporal plausibility 
• pharmacological properties of the drug or of the substance class 
• course of the adverse event after dechallenge and, if applicable, after rechallenge 
• specific tests indicating involvement of the drug in the occurrence/worsening of the 
adverse event  
• alternative explanations 
 
16.7 
Adverse Events recording 
 
Each AE occurring to a subject, either spontaneously revealed by the subject or observed by 
the investigator, whether believed by the investigator to be related or unrelated to the IMP, must 
be recorded on the AE information page of the eCRF (for SAEs information must be recorded 
also on the AE information page of the eCRF).  
The Investigator will also perform an evaluation with respect to seriousness and causality 
(relationship of any AE to IMP) of the AEs. and record it on the appropriate section of the eCRF 
and records it on the appropriate section of the eCRF. 
 
16.8 
AEs monitoring window 
 
➢ Start of monitoring: from immediately after the signature of the informed consent 
➢ End of monitoring: final study visit/ETV (observation window) 
An AE occurring after the final visit/ETV and coming to knowledge of the investigator (e.g. 
during a follow-up assessment or by spontaneous reporting by study subjects) must be recorded 
only if it is an ADR, according to the investigator’s judgment. 
 
16.9 
Adverse Events reporting 
 
The official language for reporting is English. The investigator and clinical staff of the present 
study are familiar with English language. 
 
The investigator must report to the CRO all AEs which occur during the study, regardless of 
their relationship to the IMP. Protocol specific AEs or laboratory abnormalities critical to safety 
evaluations are to be identified in the protocol and reported to the sponsor according to reporting 
requirements and within the time periods specified. 
 


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All AEs are recorded by the investigator on the AE information page of the eCRF. 
 
In addition, SAE will have to be reported according to the following detailed procedure. 
 
16.9.1 
SAEs reporting 
 
With the exception of those SAEs that are identified as not requiring immediate reporting in the 
protocol, the investigator must report the SAEs to the CRO immediately and no later than 24 
hours from when he/she becomes aware of the SAE, by faxing the “Serious Adverse Event 
Form” (back up plan) or e-mailing as scanned attachment (backup plan) or by Electronic Data 
Capture to the Drug Safety Unit (DSU) personnel (preferred method), as stated in the "List of 
CRO/ personnel" in § 16.13 of this protocol. 
 
The community standards of confidentiality must always be maintained and any relevant 
national legislation on data protection must be followed. 
 
Another copy of the “Serious Adverse Event Form” will be retained by the Investigator for the 
Investigator’s file. 
 
If the investigator becomes aware of any SAE occurred to a subject within the follow-up 
window established in the protocol, he/she will report the SAE as above. The SAE will be also 
reported in the eCRF. 
 
If outside the follow-up window established in the protocol the investigator becomes aware of 
a SAE, if the investigator judges that the SAE is related to the study drug, it should be reported 
to the Sponsor. The Investigator might use the “Serious Adverse Event Form” via email or fax, 
but the SAE must not to be reported in the eCRF, as it is not an event occurred within the study 
period. 
 
The investigator must report all SAEs that occur to the subjects to National Medical Products 
Administration/Local Ethics Committee/Provincial Food and Drug Administration/National 
Health and Family Planning Commission immediately no later than 24hours from when he/she 
becomes aware of SAE. Any SAEs that happen to the subjects outside the follow-up window 
should be reported by investigator to National Medical Products Administration/Local Ethics 
Committee/Provincial Food and Drug Administration/National Health and Family Planning 
Commission immediately. 
 
16.10 
Follow-up for Adverse Events 
 
A follow-up “Serious Adverse Event Form” will be filled in if important follow-up information 
(i.e. diagnosis, outcome, causality assessment, results of specific investigations) is made 
available after submission of the initial AE Form for immediate reporting. Follow-up “Serious 
Adverse Event Form” will be reported to the Sponsor as above-described, under Section 16.9.1. 
 
In any case of an AE that, in the opinion of the investigator, requires the subject’s 
discontinuation, follow-up information relating to the subject’s subsequent course must be 
collected until the event has subsided or the condition stabilised or when the subject is lost to 
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When follow-up data on non-serious AE are collected, information should be reported under 
“Comments” in the Final report of the CRF. 
 
16.11 
SUSARs management 
 
The clock for initial expedited reporting starts as soon as the information containing the 
minimum reporting criteria has been received by the sponsor (day 0).  
For fatal and life-threatening SUSARs the EC and Competent Authority (CA) should be 
informed as soon as possible and in any case within 7 days. 
If the initial report is incomplete, e.g. not all the information/assessments were available, a 
complete report should be sent within an additional 8 days. Complete follow-up report should 
be sent within 15 calendar days. 
SUSARs which are not fatal and not life-threatening are to be reported within 15 days. 
The minimum information to be reported includes: 
➢ Sponsor study number 
➢ One identifiable coded subject 
➢ One identifiable reporter 
➢ One SUSAR 
➢ One suspect IMP (including active substance name, code) 
➢ A causality assessment (a reasonable possibility of a causal relationship with the study 
drug can be excluded only if there is information supporting this decision, otherwise it 
cannot be excluded). 
 
16.12 
Other events qualified for expedited reporting 
 
Other safety issues also qualify for expedited reporting when they might materially alter the 
current benefit-risk assessment of a medicinal product or would be sufficient to consider 
changes in the medicinal product administration or in the overall conduct of the trial, for 
instance: 
➢ single case reports of an expected serious adverse reaction with an unexpected outcome 
(e.g. a fatal outcome) 
➢ an increase in the rate of occurrence of an expected serious adverse reaction, which is 
judged to be clinically important. 
➢ post-study SUSARs that occur after the subject has completed a clinical trial and are 
reported to the investigator by the subject. 
➢ new events relating to the conduct of the trial or the development of the medicinal 
product likely to affect the safety of the subjects, such as : 
− a SAE which could be associated with the trial procedures and which could modify 
the conduct of the trial 
− a significant hazard to the subject population such as lack of efficacy of a medicinal 
product used for the treatment of a life-threatening disease 
− a major safety finding from a newly completed animal study (such as carcinogenicity) 
or from other clinical trials. 
 


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16.13 
SAEs: contacts 
 
The investigator will report any SAE to the CRO. The CRO’s details for SAEs are the 
following: 
Email: 
 
 
The sponsor’s details for SAEs are the following: 
Email: 
 
 
16.14 
Pregnancy 
 
Subjects must be instructed that known or suspected pregnancy occurring during the study 
should be confirmed and reported to the Investigator, who must then withdraw the subject from 
the study without delay. The Investigator should also be notified of pregnancy occurring during 
the study but confirmed after its completion. In the event that a subject is subsequently found 
to be pregnant after inclusion in the study, then any pregnancy will be followed to the end of 
pregnancy or pregnancy termination and the status of mother and child will be reported to the 
Sponsor after delivery through the Pregnancy Report Form provided by the Sponsor. The 
Investigator will send pregnancy reports to DSU within the timeframes of SAEs, with follow-
up information to be actively sought for the outcome of pregnancy.  
- 
Part I of the Form is completed at the time of awareness of foetal exposure during 
pregnancy.  
- 
Part II of the Form is filled in when information on pregnancy outcome becomes available. 
If pregnancy results in abnormal outcome (spontaneous miscarriage, stillbirth and congenital 
anomalies) that the investigator considers to be due to the IMP, this will be treated as an 
expedited ADR report. 
 
CCI
CCI


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17 
DATA MANAGEMENT PROCEDURES 
 
17.1 
Data collection – CRFs 
 
The investigator must ensure that the clinical data required by the study protocol are carefully 
reported in the eCRFs. He/she must also check that the data reported in the eCRFs correspond 
to those in the subject’s source documents.  
ECG and laboratory results must be printed and signed by the Investigator and kept as source 
data on site after entering outcome into the eCRF. 
All other data has to be documented in the subject file as source data first and then entered into 
the eCRF.  
Data must be entered into eCRFs in English by the designated site personnel as soon as possible 
after a subject visit, and monitors will have access to data recorded. These data will be reviewed 
versus source documents by trial monitors for completeness and acceptability during 
monitoring visits.  
If data correction is required for an eCRF after initial entry, the site personnel can make 
necessary corrections on their own or as a response to an automated query by the eCRF system. 
Monitor and Clinical Data Manager review the eCRF for accuracy and can also generate queries 
to the investigational staff for resolution.  
Any correction to the eCRFs' entries must be carried out by the Investigator or a designated 
member of staff. Corrections are recorded in an audit trail that records the old information, the 
new information, and identification of the person making the changes, date of correction made 
and reason for change. In the interests of completeness of data acquisition, the questions which 
are repeated in each section of the eCRFs should be answered in full, even if there are no 
changes from a previous examination. A reasonable explanation must be given by the 
Investigator for all missing data. The Investigator or his/her designees named in the clinical 
staff list will review the eCRF for accuracy and completeness. The Investigator must 
electronically sign and date the eCRF pages as indicated.  
 
17.2 
Database management 
 
The CRO will update and verify the database and create the final SAS data sets. The final data 
file will be transferred to the sponsor in the agreed format with all the other study 
documentation. 
 
17.2.1 
Coding dictionaries 
 
Medical/surgical history and underlying diseases, clinically significant physical examination 
abnormalities and AEs will be coded using the Medical Dictionary for Regulatory Activities 
(MedDRA™). 
Previous and concomitant medications will be coded using the WHO Drug Dictionary 
Enhanced (WHODDE). The version of the coding dictionaries will be stated in the study report. 


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18 
STUDY MONITORING, QUALITY CONTROL AND 
QUALITY ASSURANCE 
 
18.1 
Monitoring 
 
The monitoring visits will be conducted by appropriate staff of PAREXEL China Co. Ltd 
according to PAREXEL SOPs.  
 
Monitoring activities, including monitoring purpose, selection and qualifications of monitors, 
extent and nature of monitoring, monitoring procedures, monitoring reports will comply with 
ICH-GCP chapter 5.18, CFDA GCP requirements and, when appropriate, further national 
regulatory guidelines. 
 
Adequate time and availability for monitoring activities should be ensured by the investigator 
and key study personnel. 
 
Data verification is required and will be done by direct comparison with source documents, 
always giving due consideration to data protection and medical confidentiality. In this respect 
the investigator will assure support to the monitor at all times. 
 
The investigator agrees, by written consent to this protocol, to fully co-operate with compliance 
checks by allowing authorised individuals to have access to all the study documentation. In 
addition to the monitoring activities performed by the study monitor, the sponsor could perform 
some quality control activities to verify the compliance with the study procedures, the ICH-
GCP guidelines and CFDA GCP.  
 
18.2 
Quality Control and Quality Assurance  
 
The CRO has implemented and maintains a Quality System that includes quality controls and 
audits at different study steps with written SOPs to ensure that the study is conducted in 
compliance with the protocol and all effective amendments, ICH-GCP, and the applicable 
regulatory requirement(s) and that data have been reliably and correctly generated, recorded, 
processed and reported, in agreement with the ALCOAC principles (Attributable-Legible-
Contemporaneous-Original-Accurate-Complete). 
 
The clinical site is responsible for implementing and maintaining quality assurance and a 
quality control system to ensure that the study is conducted, and data are generated, documented 
(recorded), and reported in compliance with the protocol, ICH-GCP, CFDA GCP and any 
applicable regulatory requirement(s). 
This protocol has been audited by the Sponsor QA.   
The CRO and the sponsor will be responsible for their respective activities. 
 
The sponsor may transfer any or all of the sponsor’s trial-related duties and functions to a CRO, 
but the ultimate responsibility for the quality and integrity of the trial data always resides with 
the sponsor. 
 
 


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18.3 
Applicable SOPs 
 
The sponsor, the clinical centre and the CRO will follow their respective SOPs in the conduct 
of the respective activities, unless otherwise stated in written agreements. CRO/Sponsor's SOPs 
will be used for AE and SAE definition and management. SOPs will be made available for 
Sponsor’s review, if required. 
 
18.4 
Data access 
 
The investigator and the CRO will ensure that all source data, medical records, CRFs and all 
other documentation that is relevant to this study will be made accessible for monitoring 
activities, audits, IEC review, and regulatory inspections. 
 
18.5 
Audits and inspections 
 
The sponsor, independent bodies acting on behalf of the sponsor and the CRO have the right to 
perform audits according to ICH-GCP, CFDA GCP and CFDA responsibilities. 
 
The study may also be inspected by regulatory authorities.  
 
The investigator and the CRO agree, by written consent to this protocol, to fully co-operate and 
support audits and inspections compliance checks by allowing authorised individuals to have 
access to all the study documentation. 
 
 
 


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19 
ETHICAL CONSIDERATIONS 
 
19.1 
Ethics and Good Clinical Practice (GCP) 
 
The study will be performed in accordance with the relevant guidelines of the Declaration of 
Helsinki. 
The approval of the study protocol by the local IEC and by the Chinese Health Authorities will 
be obtained before the start of the study.  
 
The present clinical study will be carried out according to the current revision of Good Clinical 
Practice (GCP), ICH topic E6 (R2), CFDA GCP and any applicable local law requirements. 
 
19.2 
Informed consent 
 
Before being enrolled into the clinical study, the subjects must have expressed their consent to 
participate, after the investigator has explained to them, clearly and in details, the scope, the 
procedures and the possible consequences of the clinical study. Information will be given in 
both oral and written form. The information sheet and informed consent form will be prepared 
in the local language by the CRO and must be approved by the EC and regulatory authorities. 
It will include all the elements required by law according to the ICH-GCP and CFDA GCP 
recommendations.  
In addition to the standard requirements that physicians are currently obliged to observe when 
providing information, the following points must also be covered: 
➢ a description of the aims of the study and how it will be organised 
➢ the type of treatment (information on the IMP and treatment procedures, as applicable) 
➢ any potential negative effects attributable to the study product or treatment 
➢ the freedom to ask for further information at any time 
➢ the subjects’ right to withdraw from the clinical study at any time without giving reasons 
and without jeopardising their further course of medical treatment  
➢ the existence of a subject insurance cover and obligations following from this cover 
Adequate time and opportunity to satisfy questions will be given to the subjects and the time 
will be recorded. 
 
The investigator will be supplied with an adequate number of blank informed consent forms to 
be used. The forms will be signed and dated by both the investigator and the subject.  
A copy of the signed form will be given to the subject. 
 
To ensure medical confidentiality and data protection, the signed informed consent forms will 
be stored in the investigator's study file according to the regulatory requirements (see § 20.3). 
The investigator will allow inspection of the forms by authorised representatives of the sponsor, 
EC members and regulatory authorities. He/she will confirm, by signing and dating the forms, 
that informed consent has been obtained. 
 
 


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19.3 
Insurance policy 
 
An insurance cover has been issued in favour of the subjects participating in this clinical study. 
The insurance is in compliance with the local regulation and with the requirements of the Health 
Authorities. 
 
19.4 
Withdrawal of subjects 
 
It will be documented whether or not each subject completed the clinical study. If, for a subject, 
study treatment or observations are discontinued, the primary reason for discontinuation will 
be recorded. 
 
19.4.1 
Primary reason for discontinuation 
 
➢ Adverse event: Any (significant) adverse event that in the opinion of the investigator or 
concerned subject is not compatible with study continuation. For the definition of AE, 
please refer to § 16.2. 
➢ death: the absence of life or state of being dead 
➢ lost to follow-up: the loss or lack of continuation of a subject to follow-up 
➢ non-compliance with study drug: an indication that a subject has not agreed with or 
followed the instructions related to the study medication 
➢ physician decision: a position, opinion or judgment reached after consideration by a 
physician with reference to the subject 
➢ pregnancy: pregnancy is the state or condition of having a developing embryo or fetus in 
the body (uterus), after union of an ovum and spermatozoon, during the period from 
conception to birth 
➢ protocol deviation: an event or decision that stands in contrast to the guidelines set out by 
the protocol 
➢ study terminated by sponsor: an indication that a clinical study was stopped by its sponsor 
➢ technical problems: a problem with some technical aspect of a clinical study, usually 
related to an instrument 
➢ withdrawal by subject: study discontinuation requested by a subject for whatever reason 
➢ other: different than the ones previously specified 
 
For any subject discontinuing the study, the investigator will:  
➢ ask the subject to undergo, as far as possible, a final medical visit (ETV) to examine the 
subject's health conditions and perform the required blood sampling for the laboratory 
assays. This examination will verify that all values tested at screening have remained within 
a clinically acceptable range (i.e. not clinically significant changes compared to screening) 
➢ arrange for alternative medical care of the withdrawn subject, if necessary 
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➢ report in the CRF date and time of the last dose administration, and date and primary reason 
of study discontinuation 
➢ record in the CRF any follow-up, if the subject is withdrawn for an AE 
Discontinued subjects will not be replaced.  
 
19.5 
Study termination 
 
The study will be considered terminated at the date of the last visit of the last subject or upon 
completion of any follow-up procedure described in protocol. The investigator and the sponsor 
have the right to discontinue the study at any time for reasonable medical and/or administrative 
reasons. As far as possible, this should occur after mutual consultation. Reasons for 
discontinuation have to be documented appropriately. 


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ADMINISTRATIVE PROCEDURES 
 
20.1 
Material supplied to the clinical centre 
 
Beside IMP, the following study material will be supplied to the clinical centre: 
➢ final version of the study protocol 
➢ CRF for each subject plus some spare copies 
➢ copy of the investigator’s brochure (IB) relative to the IMP 
➢ informed consent forms 
Moreover, before the start of the study, the investigator will be provided with the following 
documents: CFDA GCP, ICH guidelines, confidentiality agreement (if applicable), protocol 
amendments (if any), declaration of Helsinki, insurance statement, SAE forms, financial 
agreement (if applicable), confidential subject identification code list form, drug accountability 
forms, investigator and study staff list form. 
 
20.2 
Protocol amendments 
 
In order to obtain interpretable results, neither the investigator nor the sponsor will alter the 
study conditions agreed upon and set out in this protocol. Amendments should be made by 
mutual agreement between the investigator and the sponsor. Any amendment must be set out 
in writing, giving the reasons, and being signed by all concerned parties. The amendment 
becomes then part of the protocol. 
 
20.3 
Study documentation and record keeping 
 
The investigator should ensure the accuracy, completeness, legibility, and timeliness of the data 
reported to the sponsor in the CRFs and in all required reports. 
 
The investigator must keep source documents for each subject in the study. All information on 
the CRFs must be traceable to these source documents, which are generally stored in the 
subject's medical file. The source documents should contain all demographic and medical 
information, including laboratory data, ECGs, etc., and the original signed informed consent 
forms. 
 
Data reported on the CRF that are derived from source documents should be consistent with the 
source documents or the discrepancies should be explained. 
 
The investigator and the sponsor should maintain the study documents as specified in the 
“Essential Documents for the Conduct of a Clinical Trial” chapter 8 of ICH-GCP and as 
required by CFDA GCP and the applicable regulatory requirement(s).  
 
These are documents which individually and collectively permit evaluation of a study and the 
quality of the data produced and include groups of documents, generated before the study 
commences, during the clinical study, and after termination of the study and include but are not 
limited to, study protocol, amendments, submission and approval of EC, raw data of subjects 
including lab tests and ECG tracing, insurance contracts, certificate of analysis of the IMP(s), 


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drug accountability records, signed informed consent forms, confidential subjects identification 
code, CRFs, curricula vitae of the investigator and other participants in the study, study staff 
lists and responsibilities, monitoring reports and final study report. 
 
The investigator and the sponsor should take measures to prevent accidental or premature 
destruction of these documents. 
 
Study documents must be retained by the investigator and the sponsor as long as needed to 
comply with ICH-GCP, CFDA GCP, national and international regulations. By signing the 
protocol, the investigator and the sponsor agree to adhere to these requirements. 
 
20.4 
Study subjects’ recruitment 
 
Study participants will be recruited from the volunteers’ database. The study site may also use 
the site management organisation to recruit the study participants.  
 
In addition, they may also use recruitment posters in other departments within the hospital, 
community outside or newspapers to raise awareness of the study that is going on. 
 
Other departments within the hospital will be involved to recommend study participants as well.    
 
The clinical site has detailed SOPs on the recruitment process. 
 
20.5 
Confidentiality and data protection 
 
By signing this protocol, the investigator and the CRO agree to keep all the information 
provided by the sponsor in strict confidentiality and to request the same confidentiality from 
his/her staff. Study documents provided by the sponsor (protocols, IB, CRFs and other 
materials) will be stored appropriately to ensure confidentiality. The information provided by 
the sponsor to the investigator and to the CRO cannot be disclosed to others without direct 
written authorisation from the sponsor, except for the extent necessary to obtain the informed 
consent from the subjects wishing to participate in the study. 
 
Data on subjects collected in the CRFs during the study will be documented in an anonymous 
way. If, as an exception, for safety or regulatory reasons identification of a subject becomes 
necessary, the monitor, the sponsor and the investigator will be bound to keep this information 
confidential. 
 
20.6 
Publication policy 
 
The sponsor agrees that the study results (including negative and inconclusive as well as 
positive results) can be made publicly available by the investigator publishing in peer reviewed 
journals, presenting results at scientific congresses and posting information and results on 
internet-based public registers and databases. 
Study results will be communicated in full to the competent Health Authorities by the 
submission of a complete clinical study report. 
 


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As the sponsor agrees that the study results can be published by the investigator(s), the 
investigator agrees to submit any manuscript (abstract, publication, paper, etc.) to the sponsor 
before any public disclosure. 
This will be done in order to ensure that clinical study results are reported in an objective, 
accurate and balanced manner. The sponsor reviews the proposed manuscripts, before 
submission, within a reasonable period of time (30-90 days in relation with the complexity of 
the work). 
 
The investigator will also be provided by the sponsor with the clinical study report and the 
results of any additional analysis, tables, figures, etc. undertaken for the purposes of the article, 
in order to take responsibility for the content of the publication(s). 
On an exceptional basis, the sponsor may temporarily delay registration of certain data elements 
(e.g. compound, name, outcome, measures, etc.) to seek necessary intellectual property 
protection. This is because early disclosure of such data could, in some circumstances, prevent 
or negatively impact patentability. 
 
 
 


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STUDY RESPONSIBLE PERSONS 
 
21.1 
Sponsor 
 
Zambon S.p.A., via Lillo del Duca 10, I-20091 Bresso, Italy 
Phone: 
+39.02.66.52.41 
Fax: 
+39.02.66.50.14.92 
 
Protocol Review Committee Chairman  
, Global Chief Medical Officer and Patient’s Access Head 
 
Sponsor’s representatives 
, Global Medical Affairs Head Established and Respiratory Medicine 
 
 
21.2 
Drug assay 
 
Eurofins Central Laboratory Shanghai, 395 Jiang Chang West Road, 7th Floor, Shanghai, 
200436 China 
 
Analytical facility: 
United-Power Pharma Tech Co., Ltd. (UP-Pharma) 
2F, Tower B, No. 33 Science Park Road, 
Changping District, Beijing, P.R.China 
ZIP Code: 102206 
 
21.3 
Project management, data analysis & reporting, monitoring  
 
PAREXEL International (IRL) Limited (“PAREXEL”), a corporation organized under the 
laws of Ireland with a registered address at 70 Sir John Rogerson’s Quay, Dublin 2, Ireland. 
PPD
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CONFIDENTIAL 
 
 
Sponsor code Z7244J01 
NAC 300 mg/3 mL i.v. bioavailability 
Final version 3.0, 3Dec2019 
 
Page 61 
22 
REFERENCES 
 
1. Ventresca GP, Cicchetti V, Ferrari V. Drugs in Bronchial Mucology. 1989. Cap. 4.1 Thiols 77-102. Edited by 
PC Braga and Allegra. Raven Press Ltd 
2. Aruoma OI, Halliwell B, Hoey BM, Butler J. The antioxidant action of N-acetylcysteine: Its reaction with 
hydrogen peroxide, hydroxyl radical, superoxide and hypochlorous acid. Free Radical Biol Med 1988; 6:593-
597 
3. Ziment I. Acetylcysteine: a drug that is much more than a mucokinetic. Biomed & Pharmacother 1988; 42: 
513-520 
4. Borgstrom L, Kagedal B, Paulsen O. Pharmacokinetics of N-acetylcysteine in man. Eur J Clin Pharmacol 
1986; 31: 217-222 
5. Olsson B, Johansson M, Gabrielsson J, Bolme P. Pharmacokinetics and bioavailability of reduced and 
oxidized N-acetylcysteine. Eur J Clin Pharmacol 1988; 34: 77-82 
6. Burgunder JM, Varriale A, Lauterburg BH. Effect of N-acetylcysteine on plasma cysteine and glutathione 
following paracetamol administration. Eur J Clin Pharmacol 1989; 36: 127-131 
7. De Caro L, Ghizzi A, Costa R, Longo A, Ventresca GP, Lodola L.  Pharmacokinetics and bioavailability of 
oral acetylcysteine in healthy volunteers. Arzneim-Forsh/Drug Res 1989; 39: 382-386 
8. Borgstrom L, Kagedal B. Dose dependent pharmacokinetics of N-acetylcysteine after oral dosing to man. 
Biopharmaceutics & Drug Disposition 1990; 11: 131-136 
9. Frascio M. Pharmacokinetic study after single dose (1, 2, and 4 g) of a new N-acetylcysteine (NAC) oral 
formulation. Zambon Group Internal Report, code ZPM/94/34 
10. Crestani S. Open label, randomized, cross-over, pharmacokinetic study to assess the bioequivalence of two 
oral N-Acetylcysteine preparations: 600 mg film coated tablets versus 600 mg effervescent tablets. 2002 
Report in Zambon Group’s file 
11. Rusca A. Study CRO-PK-14-286 – Sponsor code Z7169J02. Comparative bioavailability study of N-acetyl-
cysteine (NAC) 600 mg uncoated tablets vs. NAC 600 mg film-coated tablets in healthy male and female 
volunteers. CROSS Research S.A., Switzerland 2014 
12. Rusca A. Study CRO-PK-07-189 – Sponsor code 7169J01. Comparative bioavailability study of a new oral 
N-acetyl-cysteine (NAC) formulation vs. Fluimucil®, Zambon in healthy volunteers. CROSS Research S.A., 
Switzerland, 2008 
13. Rusca A. Study CRO-PK-09-225 – Sponsor code 7203J01. Pilot bioavailability study of a new 1200 mg N-
acetyl-cysteine (NAC) effervescent tablet formulation vs. Fluimucil® 600 mg effervescent tablet (Zambon 
S.p.A., Italy) in healthy male and female volunteers. CROSS Research S.A., Switzerland, 2010 
14. Brown M, Bjorksten A, Medved I and M McKenna. Pharmacokinetics of intravenous N-acetylcysteine in men 
at rest and during exercise. Eur J Clin Pharmacol 2004;60:717-723 
15. Jones AL, Jarvier D. Simpson D. Pharmacokinetics of N-acetylcysteine are altered in patients with chronic 
liver diseases. Aliment Pharmacol Ther. 1997; 787-791 
16. Prescott LF, Donovan JW, Jarvie DR, Proudfoot AT. The disposition and kinetics of intravenous N-
acetylcysteine in patients with paracetamol overdosage. Eur J Clin Pharmacol 1989; 37: 501-506 
17. Pendyala L, Creaven PJ. Pharmacokinetic and pharmacodynamic studies of N-acetylcysteine, a potential 
chemopreventing agent during a phase I trial. Cancer Epidemiology, Biomarkers and Prevention 1995; 4: 245-
251 
18. Rodenstein D, De Coster A, Gazzaniga A. Pharmacokinetics of oral acetylcysteine: absorption, binding and 
metabolism in patients with respiratory disorders. Clinical Pharmacokinetics 1978; 3: 247-254 
19. Pharmacokinetics of N-Acetylcysteine following repeated i.v. infusions to hemodialysed chronic renal failure 
patients. Internal Report 2002. Zambon Group’s file 


CONFIDENTIAL 
 
 
Sponsor code Z7244J01 
NAC 300 mg/3 mL i.v. bioavailability 
Final version 3.0, 3Dec2019 
 
Page 62 
20. Sjodin K, Nilsson E, Hallemberg A, Tunek A. Metabolism of N-acetyl-l-cysteine Biochem. Pharmacol 38 
1989; 22: 3981-3985 
21. Locatelli C. Fluimucil 20% injectable (N-Acetylcysteine). Expert Report on the clinical documentation 1996. 
Zambon Group’s file 
22. N-acetyl-L-cysteine (NAC) intravenous Investigator’s Brochure, edition number 1, 20 April 2018 
23. Guidelines on GMP, published by the Commission in The rules governing medicinal products in the European 
Community, Volume 4 
24. Guidance on the investigation of bioequivalence. CPMP/EWP/QWP/1401/98 Rev. 1/Corr **, 20 January 2010  
25. U.S. Department of Health and Human Services and U.S. Department of Agriculture, Nutrition and your 
health: 2015-2020 Dietary Guidelines, Appendix 9 
 

SAP_001.pdf: 
• Protocol number: Z7244J01 
• Document title: Phase I Study to Evaluate Pharmacokinetics, Safety, and Tolerability 
of Single and Multiple i.v. Doses of N-acetylcysteine (NAC) in Chinese Healthy 
Volunteers 
• NCT number: NCT03881163 
• Version number: 4.0 
• Date of the document: 15 June 2020 


Clinical Study Report Appendices 
Zambon S.p.A., Italy 
Z7244J01 
Final 1.0 
20 Nov 2020 
16.1.9 Documentation of Statistical Methods 
The following documents are included: 

Statistical Analysis Plan, Version 4.0, dated 15 Jun 2020
Previous versions of Statistical Analysis Plan (Version 1.0, dated 18 Apr 2019, Version 2.0, 
dated 04 Nov 2019, and Version 3.0, dated 21 Apr 2020) and Statistical Analysis Report 
(Version 1.0, dated 10 Jul 2020) are in 
 and are available upon request. 
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PAREXEL International 
Zambon S.p.A. 
Z7244J01 
Statistical Analysis Plan 
TP-GDO-WW-016-06 
CONFIDENTIAL 
Project Document Version No. 4.0 
Effective Date:  28 Jun 17 
Project Document Effective Date: 15 Jun 2020 
Related to:  
 
Page 1 of 31 
 PAREXEL International 
Zambon S.p.A. 
Z7244J01 
Phase I Study to Evaluate Pharmacokinetics, Safety, and Tolerability of Single and 
Multiple i.v. Doses of N-acetylcysteine (NAC) in Chinese Healthy Volunteers 
Single and Multiple Dose, Single Centre, Open-Label, One-Way, Pharmacokinetics, Safety and 
Tolerability Clinical Trial 
Statistical Analysis Plan 
PAREXEL Project Number: 
 
CCI
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PPD
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Zambon S.p.A. 
 
Z7244J01 
Statistical Analysis Plan 
 
TP-GDO-WW-016-06 
CONFIDENTIAL 
Project Document Version No. 4.0 
Effective Date:  28 Jun 17 
Project Document Effective Date: 15 Jun 2020 
Related to:  
 
Page 3 of 31 
 
 
PAREXEL SIGNATURE PAGE 
 
 
Signatures below confirm that the review process has been completed in accordance with 
 
 
This document has been approved and signed electronically on the final page by the 
following: 
 
 
 
 
Signatory 
Author  
Project Role: 
  
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Zambon S.p.A. 
 
Z7244J01 
Statistical Analysis Plan 
 
TP-GDO-WW-016-06 
CONFIDENTIAL 
Project Document Version No. 4.0 
Effective Date:  28 Jun 17 
Project Document Effective Date: 15 Jun 2020 
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Page 4 of 31 
TABLE OF CONTENTS 
 
1 
INTRODUCTION ....................................................................................................... 9 
2 
STUDY OBJECTIVES ............................................................................................... 9 
2.1 Primary End-Points ............................................................................................... 9 
2.2 Secondary End-Points ........................................................................................... 9 
3 
INVESTIGATIONAL PLAN ..................................................................................... 9 
3.1 Overall Study Design and Plan ............................................................................. 9 
3.2 Pharmacokinetics and Safety Variables .............................................................. 10 
3.2.1 Pharmacokinetics Variables ...................................................................... 10 
3.2.2 Safety Variables ........................................................................................ 12 
4 
STATISTICAL METHODS ..................................................................................... 13 
4.1 Data Quality Assurance ...................................................................................... 13 
4.2 General Presentation Considerations .................................................................. 13 
4.3 Study Subjects .................................................................................................... 14 
4.3.1 Disposition of Subjects ............................................................................. 14 
4.3.2 Protocol Deviations ................................................................................... 14 
4.4 Analysis Populations .......................................................................................... 15 
4.4.1 Definitions................................................................................................. 15 
4.4.2 Reasons for exclusion from the PK set before statistical analysis ............ 15 
4.4.3 Analysis..................................................................................................... 15 
4.5 Demographic and Other Baseline Characteristics .............................................. 16 
4.6 Medical and Surgical History ............................................................................. 16 
4.7 Prior and Concomitant Medications/Procedures ................................................ 16 
4.8 Treatment Compliance ........................................................................................ 17 
4.9 Pharmacokinetics Analysis ................................................................................. 17 
4.9.1 Pharmacokinetic Concentrations .............................................................. 17 
4.9.2 Handling of Values Below the Limit of Qualification (BQL) .................. 19 
4.9.3 Pharmacokinetic Parameters ..................................................................... 19 
4.10 
Safety Evaluation ...................................................................................... 20 
4.10.1 
Adverse Events ................................................................................. 20 
4.10.2 
Clinical Laboratory Evaluation ......................................................... 22 
4.10.3 
Vital Signs, body weight, 12-Lead Electrocardiogram (ECG), and 
Physical Examinations .............................................................................. 22 
4.11 
Determination of Sample Size .................................................................. 23 
4.12 
Changes in the Conduct of the Study or Planned Analysis ...................... 23 
5 
REFERENCES .......................................................................................................... 23 
6 
APPENDIX ............................................................................................................... 24 
APPENDIX 1 Study Schedule .................................................................................. 24 
APPENDIX 2 Partial Date Conventions ................................................................... 26 
Algorithm for Medical History, Prior/Concomitant Medications, Therapies or 
Procedures: ................................................................................................ 26 
Algorithm for Treatment-Emergent Adverse Events: ........................................ 28 
APPENDIX 3 CTC Grading for Laboratory Parameters .......................................... 30 
 
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REVISION HISTORY 
 
Version No. Effective 
Date 
Summary of Change(s) 
Draft 0.1 
29 Aug 2018 
New document 
Draft 0.2 
01 Mar 2019 
Revised from draft version for CTA submission (draft 
0.1), based on final protocol 2.0, 19 Dec 2018. 
Draft 0.3 
12 Mar 2019 
Revised based on internal review. 
Draft 0.4 
08 Apr 2019 
Revised based on sponsor review and internal review on 
TLF Shells: 
1) Move screening lab tests analysis from Safety 
section to Demographic and Baseline section. 
2) Add shifting table to present change from 
baseline for categorical safety evaluations 
(clinically significance and CTCAE Grade). 
3) Add 
summary 
of 
cumulative 
urine 
PK 
concentration. 
Final 1.0 
18 Apr 2019 
Upgraded version to Final 1.0. 
Draft 1.1 
28 Oct 2019 
1) Update urinalysis parameters in section 3.2.2 
Safety Variables according to protocol v2.0 
memorandum. 
2) Update section 4.7: concomitant procedures are 
analyzed in Safety set. 
3) Update section 4.10.1: The summary of adverse 
events is analyzed by period as well. 
Draft 1.2 
30 Oct 2019 
1) Adding PK relevant plots during multiple dose. 
2) Remove physical examination on Day 1. 
3) Alcohol breath test is added in section 4.5.  
4) The surgical history is added in section 4.6. 
2.0 
04 Nov 2019 
Upgraded version to Final 2.0. 
Draft 2.1 
07 Feb 2020 
The major change includes following: 
1. Section 4.7: Updated WHODrug and MedDRA 
version. 
2. Section 4.9.3: removed "all other descriptive 
statistics will be reported to three significant 
digits" to show more significant digits in 
AUCmin and AUCmax; updated statistical outlier 
testing rule in PK concentration data. 
3. Section 4.10.1: Updated MedDRA version; 
updated "Any TEAE leading to dose reduced" as 
"Any TEAE leading to dose interruption". 
4. APPENDIX 3: Remove duplicate rule to grade 
Hyper Serum creatinine based on BL (baseline) 
value; deleted coagulation. 
3.0 
21 Apr 2020 
Upgraded version to Final 3.0. 
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3.1 
12 Jun 2020 
After database locked, SAP appendix updated to be 
clearer after final TFL output reviewed by Sponsor: 
1. APPENDIX 
2 
Partial 
Date 
Conventions: 
The imputation rule of partial date for dates of 
ECG, laboratory data, standardized meals, 
physical examination, substance usage and vital 
sign is specified clearly in Appendix. This update 
in SAP doesn’t affect final TFL results, just to 
ensure consistency in ADaM spec derivation, 
datasets programming and final TFL outputs. 
2. APPENDIX 3 CTC Grading for Laboratory 
Parameters: update wording of LLC as LLN and 
ULC as ULN, to ensure consistency between 
SAP and final TFL outputs. Also updated in List 
of Abbreviation. 
3. PK results of BLQ is corrected as BQL to ensure 
consistency between SAP and outputs. Also 
updated in List of Abbreviation. 
 
4.0 
15 Jun 2020 
Upgraded version to Final 4.0. 
 
 
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LIST OF ABBREVIATIONS 
 
Abbreviation / Acronym 
Definition / Expansion 
AE 
Adverse Event 
ALT 
Alanine aminotransferase 
Ae0-t 
Total amount of NAC excreted in urine  
Aess(0-t) 
Total amount of NAC excreted in urine at steady-state 
AST 
Aspartate aminotransferase 
AUC(0-t) 
Area under the concentration-time curve from single dose to the last 
observed concentration time t 
AUC(0-) 
Area under the concentration-time curve extrapolated to infinity 
AUC(0-12) 
Area under the concentration-time curve from single dose to 12h 
postdose 
AUCss(0-t) 
Area under the concentration-time curve at steady-state from the last 
multiple dose to the last observed concentration time t 
AUCss(0-12) 
Area under the concentration-time curve at steady-state in the tau 
interval  
BQL 
Below Quantification Limit  
BMI 
Body Mass Index 
BP 
Blood Pressure 
Cmax 
Peak drug concentration 
CLt 
Total body clearance 
CLr 
Renal clearance 
CRF 
Case Report Form 
CS 
Clinically Significant 
CSP 
Clinical Study Protocol 
CSR 
Clinical Study Report 
Css_av 
Average NAC plasma concentration at steady-state 
Css_max 
Maximum plasma concentration at steady-state 
Css_min 
Trough plasma concentration at steady-state 
CTCAE 
Common Terminology Criteria for Adverse Events 
CV 
Coefficient of Variation 
DBL 
Database Lock 
DBP 
Diastolic Blood Pressure 
DF 
Degree of fluctuation  
DTT 
Dithiothreitol 
ECG 
Electrocardiogram 
ETV 
Early Termination Visit 
Fe0-t 
Total fraction of NAC dose excreted in urine  
Frel 
Relative Bioavailability 
HBs Ag 
Hepatitis B virus surface antigen 
HCV Ab 
Hepatitis C virus antibodies 
HIV 
Human Immunodeficiency Virus 
HR 
Heart Rate 
ICH 
International Conference on Harmonisation 
i.m. 
Intramuscular 
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IMP 
Investigational Medicinal Product 
i.v. 
Intravenous 
kel 
Terminal elimination rate constant 
LQL 
Lower Quantification Limit 
LLN 
Lower limit of Normal 
MCH 
Mean Cell Haemoglobin 
MCHC 
Mean Cell Haemoglobin Concentration 
MCV 
Mean Cell Volume 
MedDRA 
Medical Dictionary for Regulatory Activities 
NAC 
N-acetyl-L-cysteine 
NC 
Not calculated 
NCI 
National Cancer Institute 
NCS 
Not clinically significant 
PK 
Pharmacokinetics 
PT 
Preferred Term 
PTAE 
Pre-Treatment Adverse Event 
R 
Accumulation ratio 
RBC 
Red Blood Cells 
SAE 
Serious Adverse Event 
SAP 
Statistical Analysis Plan 
SBP 
Systolic Blood Pressure 
SD 
Standard Deviation 
SOC 
System Organ Class 
SOP 
Standard Operating Procedure 
SDTM 
Study Data Tabulation Model 
TEAE 
Treatment-Emergent Adverse Event 
THC 
delta-9-tetrahydrocannabinol 
t1/2 
Half-life 
tmax 
Time to achieve Cmax 
tss_max 
Time to achieve Css_max 
ULN 
Upper Limit of Normal 
Vd 
Volume of distribution  
WBC 
White Blood Cells 
 
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Page 9 of 31 
1 
INTRODUCTION 
This SAP is based upon the following study documents: 
• Study Protocol, Version 3.0 (Dec 03, 2019) 
• electronic Case Report Form (eCRF), Version 3.0 (Oct 14, 2019) 
 
The Statistical Analysis Plan (SAP) details the statistical methodology to be used in 
analyzing study data and outlines the statistical programming specifications for the 
Tables, Listings, and Figures. It describes the variables and populations, anticipated data 
transformations and manipulations and other details of the analyses not provided in the 
Clinical Study Protocol (CSP). 
The SAP will be finalized prior to database lock (DBL) and describes the statistical 
analysis as it is foreseen when the study is being planned. If circumstances should arise 
during the study rendering this analysis inappropriate, or if improved methods of analysis 
should arise, updates to the analyses may be made. Any deviations from the SAP after 
DBL, reasons for such deviations and all alternative or additional statistical analyses that 
may be performed, will be described in a SAP Addendum. 
2 
STUDY OBJECTIVES 
The objective of the study is to evaluate the NAC pharmacokinetics (PK), safety and 
tolerability after single and multiple dose i.v. administration. 
 
2.1 
Primary End-Points 
To evaluate pharmacokinetic parameters of NAC in plasma after single and multiple dose 
administration of the investigational product.  
 
2.2 
Secondary End-Points 
To collect safety and tolerability data after single and multiple dose administration of the 
investigational product. 
3 
INVESTIGATIONAL PLAN 
3.1 
Overall Study Design and Plan 
This is a single and multiple dose, single centre, open-label, one-way, pharmacokinetics, 
safety and tolerability clinical trial of Phase I to be performed in Chinese healthy male and 
female volunteers. Study schedule is described as Appendix 1. 
 
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3.2 
Pharmacokinetics and Safety Variables 
3.2.1 Pharmacokinetics Variables 
The derivation of PK parameters will be the responsibility of Quantitative Clinical 
Development (QCD), PAREXEL International.  PK parameters will be calculated by non-
compartmental analysis methods following these guidelines; 
 
-  Actual sampling times relative to dosing rather than nominal times will be used 
in the calculation of all derived PK parameters. 
- There will be no imputation of missing data.  
- Any subjects with missing concentration data will be included in the analysis 
provided that at least Cmax and AUC(0-t) can be reliably calculated 
 
The following PK parameters will be measured and/or calculated for plasma and urine 
NAC data, using the validated software Phoenix® WinNonlin® version 8.0 or higher 
(Certara USA, Inc., Princeton, NJ) (actual version will be stated in the final report), after a 
single dose of the Investigational Medicinal Product (IMP): 
 
 
Cmax: 
Maximum NAC plasma concentration will be obtained directly from the 
concentration data 
 
tmax: 
Time to achieve Cmax 
 
kel: 
Terminal elimination rate constant, calculated, by linear regression s  
- 
Only those data points that are judged to describe the terminal 
log-linear decline will be used in the regression.  
− A minimum number of three data points in the terminal phase will 
be used in calculating Kel with the line of regression starting at 
any post-Cmax data point (Cmax should not be part of the regression 
slope) and including Clast, tlast. 
− The correlation coefficient (R2) in general should be greater than 
0.80.  
 
t1/2: 
Half-life, calculated, if feasible, as ln2/kel 
 
AUC(0-t): 
Area under the concentration-time curve from single dose to the last 
observed concentration time t, calculated with the linear up/log down 
trapezoidal method 
 
AUC(0-): 
Area under the concentration-time curve extrapolated to infinity, 
calculated, if feasible, as AUC(0-t) + Ct/kel, where Ct is the last measurable 
drug concentration 
 
AUC(0-12): 
Area under the concentration-time curve from single dose to 12 h post-
dose, calculated with the linear up/log down trapezoidal method 
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%AUCextra: 
Percentage of the AUC(0-) obtained by extrapolation, calculated as;  (1－
[AUC(0-t)/AUC(0-∞)] )×100 
 
Vd: 
Volume of distribution associated with the terminal slope, calculated, if 
feasible, as Dose/(AUC(0-)*kel) 
 
CLt: 
 
Total body clearance, calculated, if feasible, as Dose/ AUC(0-) 
 
Vur: 
Measured overall volume of urine collected during each collection 
interval. 
 
Ae(t1-t2): 
 
Amount excreted in the urine at each interval collection 
 
Ae(0-tz): 
Cumulative amount excreted up to tz hours, where tz is 4, 8, 12, 24 or 32 
will be calculated as the sum of the products of concentration and urine 
volume over the appropriate collection intervals 
 
Ae(0-t): 
 
 
Total amount of NAC excreted in urine from single dose up to 32 h 
 
 
Fe(0-tz): 
 
The fraction of unchanged drug excreted in urine will be calculated as 
fe=Ae/Dose, up to tz hours, where tz is 4, 8, 12, 24 or 32 hours 
 
Fe(0-t): 
Total fraction of NAC dose excreted in urine from single dose up to 32 h 
 
CLr: 
Renal clearance, calculated, if feasible, as Ae(0-t)/ AUC(0-) 
 
 
The following PK parameters will be measured and/or calculated for plasma NAC, after 
multiple doses of IMP: 
 
Css_max: 
Maximum NAC plasma concentration at steady-state will be obtained 
directly from the concentration data 
 
tss_max: 
Time to achieve Css_max 
 
Css_min: 
Trough NAC plasma concentration at steady-state, measured as the 
concentration at t=12 h 
 
AUCss(0-t): 
Area under the concentration-time curve at steady-state from the last 
multiple dose to the last observed concentration time t, calculated with the 
linear up/log down trapezoidal method 
 
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AUCss(0-12): 
Area under the concentration-time curve at steady-state in the tau interval 
(from the last multiple dose to 12 h post-dose), calculated with the linear 
up/log down trapezoidal method 
 
Css_av: 
Average NAC plasma concentration at steady-state, calculated as 
AUCss(0-12) /tau 
 
R: 
Accumulation ratio, calculated as AUCss(0-12)/ AUC(0-12) 
 
Aess(0-t): 
Total amount of NAC excreted in urine from the last multiple dose to 32 
h at steady-state 
 
DF%: 
Degree of fluctuation over one dosing interval at steady-state, calculated 
as (Css_max - Css_min)/ Css_av*100 
 
 
The sampling schedule is considered adequate if the ratio AUC(0-t)/ AUC(0-) equals or 
exceeds a factor of 0.8 (i.e. if %AUCextra is <20%) for more than 80% of the individual PK 
profiles. This assures that the primary variable AUC(0-t) covers a sufficient percentage of 
the theoretical total extent of exposure.  
 
The quality of log-linear regression (and, consequently, the reliability of the extrapolated 
PK parameters) should be demonstrated by a determination coefficient R2 ≥ 0.8. Individual 
extrapolated parameters, when considered unreliable, will be reported as NC (not 
calculated).  
 
BQL values will be imputed in the PK concentration dataset used for the derivation of PK 
parameters. The following rules will be applied: 
 
• BQLs at the beginning of a subject profile (i.e., before the first incidence of a 
measurable concentration) will be assigned to zero. 
• BQLs at the end of a subject profile (i.e., after the last incidence of a measurable 
concentration) will be set to missing. 
• Single BQLs which fall between two measurable concentrations and consecutive BQLs 
which fall between measurable concentrations will be set to missing. 
3.2.2 Safety Variables 
Safety and general tolerability of the IMP variables include: 
• Treatment-Emergent Adverse Event (TEAEs) 
• Physical examinations including body weight at screening, day 3 and final 
visit/ETV. 
• Vital signs at screening, day 1 (pre and post dose), day 3 (post dose), day 6 (pre and 
post dose) and Final visit (post dose)/ETV. 
• 12-lead ECG at screening. 
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• Laboratory tests at screening, day 3 and the final visit/ETV:  
o Haematology:  
Leukocytes and leukocyte differential count (percentage values and 
absolute values), erythrocytes, haemoglobin (conv. units), haemoglobin (IS 
units), haematocrit, MCV, MCH, MCHC, thrombocytes. 
o Blood chemistry: 
Electrolytes: sodium, potassium, calcium, chloride, inorganic phosphorus 
Enzymes: alkaline phosphatase, γ-GT, AST, ALT. 
Substrates/metabolites: total bilirubin, creatinine, glucose, BUN or urea, 
uric acid, total cholesterol, triglycerides. 
Proteins: total proteins. 
o Urine analysis: 
Urine chemical analysis: pH, specific gravity, appearance, colour, nitrites, 
proteins, glucose, urobilinogen, bilirubin, ketones, blood, leukocytes, 
erythrocytes, epithelial, crystals, cylinders. 
4 
STATISTICAL METHODS 
4.1 
Data Quality Assurance 
All tables, figures and data listings to be included in the report will be independently 
checked for consistency, integrity and in accordance with standard PAREXEL procedures. 
 
4.2 
General Presentation Considerations 
‘Baseline’ is defined as the last available pre-treatment assessment.  ‘End of Study’ is 
defined as the last available post-treatment assessment.  ‘Treatment Day’ will be calculated 
relative to the date of first dose i.e. Treatment Day = Assessment Date -First Dose Date + 
1.  
 
Continuous data will be summarized in terms of the mean, standard deviation (SD), 
median, minimum, maximum and number of observations, unless otherwise stated. 
Continuous data that are expected to be skewed will be presented in terms of the maximum, 
upper quartile, median, lower quartile, minimum and number of observations.  The 
minimum and maximum will be reported to the same number of decimal places as the raw 
data recorded in the database.  The mean, median, lower quartile and upper quartile will be 
reported to one more decimal place than the raw data recorded in the database.  The SD 
will be reported to two more decimal places than the raw data recorded in the database.  In 
general, the maximum number of decimal places reported shall be four for any summary 
statistic. 
 
Categorical data will be summarized in terms of the number of subjects providing data at 
the relevant time point (n), frequency counts and percentages.  Any planned collapsing of 
categories will be detailed in the SAP text and the data displays.   
 
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Percentages will be presented to one decimal place. Percentages will not be presented for 
zero counts.  Percentages will be calculated using n as the denominator.  If sample sizes 
are small, the data displays will show the percentages, but any textual report will describe 
frequencies only.   
 
Changes from baseline in categorical data will be summarized using shift tables where 
appropriate. 
 
P-values greater than or equal to 0.001, in general, will be presented to three decimal 
places.  P-values less than 0.001 will be presented as “<0.001”. 
 
Confidence intervals will be presented to one more decimal place than the raw data. 
 
No available data will be evaluated as “missing values”.  
 
All report outputs will be produced using SAS® version [9.4] or a later version in a secure 
and validated environment.  All report outputs will be provided to the Sponsor in a single 
Microsoft Word document.   
  
4.3 
Study Subjects 
4.3.1 Disposition of Subjects 
Subject disposition will be listed and summarized. The summary will include the number 
and percentage of subjects: 
• Screened 
• Screen failure and primary reason 
• Treated 
• Discontinued treatment and primary reason 
• Discontinued the study and primary reason 
4.3.2 Protocol Deviations 
 
Major protocol deviations and any action to be taken regarding the exclusion of subjects or 
affected data from PK set are defined in the project-specific Protocol Deviation 
Specification. 
 
• 
A summary of the number and percentage of subjects with a major protocol deviation 
by type of deviation will be provided for Enrolled set. 
 
• 
A summary of the number and percentage of subjects excluded from PK set by type of 
reason will be provided for Enrolled set. 
 
• 
A by-subject listing of protocol deviations will be provided. 
 
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4.4 
Analysis Populations 
4.4.1 
Definitions 
 
A subject will be defined as screened after the signature of the informed consent, regardless 
of the completion of all the screening procedures. 
 
A subject will be defined as eligible if he/she meets all the inclusion/exclusion criteria. 
Otherwise he/she will be defined as a screen failure.  
 
A subject will be defined as enrolled in the study if he/she is included into the interventional 
phase of the study.  
 
The following analysis sets are defined: 
• 
Enrolled set: all enrolled subjects. This analysis set will be used for demographic, 
baseline and background characteristics 
• 
Safety set: all subjects who receive at least one dose of IMP. This analysis set will be 
used for the safety analyses 
• 
PK set: all enrolled subjects who fulfil the study protocol requirements in terms of IMP 
intake and have evaluable PK data readouts for the planned analyses, with no major 
deviations that may affect the PK results. This analysis set will be used for the statistical 
analysis of the PK parameters. 
 
Each subject will be coded as valid or not valid for the safety and the PK set. Subjects will 
be evaluated according to the treatment they actually receive. 
 
4.4.2 
Reasons for exclusion from the PK set before statistical analysis 
 
Reasons for the exclusion of subjects from the PK set are the following: 
• 
intake of concomitant medications which could render the plasma concentration-time 
profile unreliable 
• 
Adverse events (AEs) which could render the plasma concentration-time profile 
unreliable 
• 
administration errors which could render the plasma concentration-time profile 
unreliable 
• 
other events which could render the plasma concentration-time profile unreliable 
If one of these events occurs, it will be noted in the Case report form (CRF) during the 
study. 
 
The samples from the subjects excluded from the PK set should still be assayed and the 
results listed. Subjects should not be excluded from the PK set if the AUC0-t covers less 
than 80% of the AUC0-∞. 
 
4.4.3 
Analysis 
 
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• 
A summary of subjects included in each analysis set will be provided for Enrolled Set. 
• 
By-subject listing including analysis set flag (Yes or No), and reason of exclusion from 
safety and PK set will be provided. 
 
4.5 
Demographic and Other Baseline Characteristics 
The following summaries will be provided for demographic and other baseline 
characteristics based on Enrolled set. 
 
• 
A summary of demographic variables and baseline characteristics:  
Age (years), sex, ethnicity, height (cm), weight (kg), BMI (kg/m2). 
BMI (kg/m2) = weight (kg) / [height (cm)]2 
 
• 
A summary of other possibly relevant variables: 
Alcohol consumption, tobacco consumption, caffeine consumption, and history of 
drugs. 
 
• 
By-subject listings of demographic data and other baseline characteristics (as 
summarized above) will be provided. 
 
Other lab tests performed during screening and baseline visits will also be listed: 
• Serum virology and bacteriology at screening: Hepatitis B (HBs antigen), Hepatitis 
C (HCV antibodies), HIV 1/2 (HIV Ag/Ab combo), Treponema pallidum. 
• Urine drug test at screening and Visit 2, day -1: cocaine, amphetamine, 
methamphetamine, cannabinoids (delta-9-tetrahydrocannabinol - THC), opiates 
• Alcohol breath test. 
• Serum pregnancy test at screening.  
• Urine pregnancy test at Visit 2, day -1.  
 
4.6 
Medical and Surgical History 
Medical and surgical history will be coded using the Medical Dictionary for Regulatory 
Activities (MedDRA version 22.0). 
 
A summary of medical and surgical history by system organ class (SOC) and preferred 
term (PT) on Enrolled Set will be provided.  
 
By-subject listings of medical and surgical history will also be provided. 
 
4.7 
Prior and Concomitant Medications/Procedures 
Prior and Concomitant medications will be coded using the WHO Drug Dictionary 
Enhanced (WHODrug Global version March 2019). 
 
Prior and Concomitant procedures will be coded using the Medical Dictionary for 
Regulatory Activities (MedDRA version 22.0). 
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Medications will be classified as “Prior” or “Concomitant” based on the comparison 
between start and stop dates of the medication and the date of first dose of study drug. 
Medications that stop prior to the date of first dose of study drug will be classified as “Prior 
Medications”. 
Medications that stop on or after the date of first dose of study drug will be classified as 
“Concomitant Medications”. Imputation of partial dates of medication records for 
classification of “prior” and “concomitant” is described in Appendix 2. 
Prior and concomitant procedures will be classified in the same way as medications. 
 
• 
A summary of concomitant medications by ATC level 3 term and preferred name will 
be provided for Enrolled Set. 
• 
A summary of concomitant procedures by SOC and PT will be provided for Safety Set. 
• 
By-subject listing of concomitant medications and procedures will be provided. 
• 
By-subject listing of prior medications and procedures will be provided. 
 
4.8 
Treatment Compliance 
Exposure and compliance data will be summarized and listed based on Safety Set. 
• 
A descriptive summary of treatment compliance variables will be provided: 
o Treatment duration (days) = last dose date – first dose date + 1 
o Total amount of exposure doses (ml)  
o Treatment compliance (%) = (total amount of actual dosage infused/total amount 
of prescribed dosage)*100  
o Treatment compliance by categories (<80%, 80%-120%, >120%) 
Detailed IMP administration data and the treatment compliance variables will be listed. 
Detailed standardized meals will also be listed. 
4.9 
Pharmacokinetics Analysis 
4.9.1 Pharmacokinetic Concentrations 
Pharmacokinetic plasma and urine concentration data for NAC will be listed by subject. 
Listings will include actual sampling times relative to dose administration, in addition the 
urine data listing will include the volume of urine collected, the concentration of NAC and 
the amount of NAC excreted per collection interval. 
Plasma concentrations and urine amounts of NAC will be summarized descriptively. The 
following descriptive statistics will be presented for plasma and urine data obtained at each 
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nominal time point: n, arithmetic mean, SD, coefficient of variation (CV%), geometric 
mean, geometric SD, geometric CV% (calculated as:  gCV% =SQRT(es²-1)*100; where s 
is the standard deviation of the log-transformed values), median, minimum and maximum 
values. 
The following rules will be followed with regards to the number of decimal places and 
presentation of data in the tables and listings of concentration data:  
 
• Source data shall be used in all derived PK concentrations without prior rounding  
• The mean, standard deviation (SD), geometric mean and median will be tabulated 
to one more significant digit compared to the source data, but with a maximum of 
four significant digits.  
• Minimum and maximum values will be tabulated to the same precision as the 
source data, but with a maximum of four significant digits.  
• Geometric coefficient of variation (CV) % and coefficient of variation (CV%) 
will be presented to one decimal place.  
 
The following descriptive pharmacokinetic graphs will be generated: 
 
• Subject Profiles for NAC Plasma Concentration vs. Time Data for Single Dose – 
Linear Scale 
• Subject Profiles for NAC Plasma Concentration vs. Time Data for Multiple Dose 
– Linear Scale 
• Subject Profiles for NAC Plasma Concentration vs. Time Data for Single Dose – 
Semi-Logarithmic Scale 
• Subject Profiles for NAC Plasma Concentration vs. Time Data for Multiple Dose 
– Semi- Logarithmic Scale  
• Arithmetic Mean (± SD) NAC Plasma Concentration vs. Time Data for Single 
Dose – Linear Scale 
• Arithmetic Mean (± SD) NAC Plasma Concentration vs. Time Data for Multiple 
Dose – Linear Scale 
• Arithmetic Mean NAC Plasma Concentration vs. Time Data for Single Dose – 
Semi-Logarithmic Scale 
• Arithmetic Mean NAC Plasma Concentration vs. Time Data for Multiple Dose – 
Semi-Logarithmic Scale 
• Subject Profiles for Cumulative NAC Urine Amount Excreted vs. Time Data for 
Single Dose – Linear Scale 
• Subject Profiles for Cumulative NAC Urine Amount Excreted vs. Time Data for 
Multiple Dose – Linear Scale 
• Subject Profiles for Cumulative NAC Urine Amount Excreted vs. Time Data for 
Single Dose – Semi-Logarithmic Scale 
• Subject Profiles for Cumulative NAC Urine Amount Excreted vs. Time Data for 
Multiple Dose – Semi-Logarithmic Scale 
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• Arithmetic Mean (± SD) Cumulative NAC Urine Amount Excreted vs. Time Data 
for Single Dose – Linear Scale 
• Arithmetic Mean (± SD) Cumulative NAC Urine Amount Excreted vs. Time Data 
for Multiple Dose – Linear Scale 
• Arithmetic Mean Cumulative NAC Urine Amount Excreted vs. Time Data for 
Single Dose – Semi-Logarithmic Scale 
• Arithmetic Mean Cumulative NAC Urine Amount Excreted vs. Time Data for 
Multiple Dose – Semi-Logarithmic Scale 
 
4.9.2 Handling of Values Below the Limit of Qualification (BQL) 
For listings, all concentrations below the limit of quantification (BQL) will be labeled as 
“BQL” in the concentration data listings. Missing data will not be imputed. 
For calculation of descriptive statistics of concentration by time point and graphs of 
arithmetic means, values that are BQL will be substituted with zero. 
For calculation of geometric means, as a consequence of BQL values, calculated geometric 
means could be null. In the presence of any null values, the geometric mean will be reported 
as not calculated (NC). 
4.9.3 Pharmacokinetic Parameters 
 
Pharmacokinetic parameters will be listed by subject. Descriptive statistics for calculated 
PK parameters will include: n, arithmetic mean, SD, CV%, geometric mean, geometric 
CV%, median, minimum and maximum values. For tmax, only median, minimum and 
maximum values will be presented. No descriptive statistics will be determined when fewer 
than three individual PK parameters are available. 
The following rules will be followed with regards to the number of decimal places and 
presentation of data in the tables and listings for PK parameters:  
• Individual PK parameters will be presented to four significant digits, with the 
exception of tmax, which will be presented to two decimal places.  
• Parameters derived directly from source data (e.g. Cmax,) shall be reported with 
the same precision as the source data (if this is not four significant digits).  
• The mean, geometric mean, median and SD values will be reported to four 
significant digits except for CV% which will be presented to one decimal place.  
• For tmax the minimum and maximum will be presented to two decimals places and 
all other descriptive statistics will be presented to three decimal places.  
• Estimates and confidence intervals in the form of percentages will be presented to 
two decimal places.  
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After receiving Plasma and urine PK concentration data from Vendor, statistical outlier 
testing will be performed by using boxplots to identify outliers as described below: 
o Q1: 25th percentiles; Q3: 75th percentiles; Interquartile Range (IQR) = Q3 – Q1; 
o Lower Far outlier: value < Q1 − 3 × IQR; 
o Upper Far outlier: value < Q3 + 3 × IQR. 
The far outliers will be identified, reviewed and discussed during the Data Review 
Meeting prior to Database lock. Outliers that may have an impact on the concentration-
time profiles will be discussed and it will be determined whether to exclude them from 
PK concentrations, PK parameters or PK set. As a general rule, outliers will be ruled out 
from tabulations but kept in data listings.  
 
If a subject or outlier is required to be excluded from PK analysis, details will be 
documented in Data Review Meeting minutes, including an external excel file that 
documents the required Analysis Flag on applicable analysis datasets level. 
 
4.10 Safety Evaluation 
All safety summaries and analyses will be based upon the Safety Set. 
4.10.1 Adverse Events 
 
AEs will be coded by System Organ Class (SOC) and Preferred Term (PT), using the 
Medical Dictionary for Regulatory Activities (MedDRA version 22.0). 
AEs will be classified as pre-treatment AEs (PTAEs) and TEAEs, according to the period 
of occurrence, as follows:  
• 
PTAEs: all AEs occurring before the first dose of IMP and not worsening after the 
first dose of IMP 
• 
TEAEs: all AEs occurring or worsening after the first dose of IMP 
 
Where dates are missing or partially missing, adverse events will be assumed to be 
treatment-emergent, unless there is clear evidence (through comparison of partial dates) to 
suggest that the adverse event started prior to the first dose of IMP.  The detailed rules are 
described in Appendix 2. 
Individual PTAEs and TEAEs will be listed in subject data listings. No summary table will 
be provided for PTAEs.  
Serious adverse event (SAEs) occurring or worsening after the first dose of IMP are defined 
as TESAEs. 
For all TEAE tables, a subject will be counted only once for each System Organ Class 
(SOC) and each PT, even if the subject reported more than one event under each 
subcategory. Unless specified otherwise, TEAE tables will be ordered in terms of 
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decreasing number of subjects for SOC and then PT within the SOC in the column of “NAC 
600 mg”, and then alphabetically for SOC and PT within the SOC if the frequency is tied. 
For each subject and each adverse event, the worst severity recorded will be attributed and 
used in the by-severity summaries.  Similarly, the worst causality (most related to 
treatment) will be attributed and used in the by-causality summaries.  If severity or 
causality is missing, the worst case will be assumed. 
An overview table will summarize the number and percentage on subject level of the 
following categories and by period (single and multiple dose): 
• Any TEAE 
• Any TEAE related to IMP 
• Any severe TEAE 
• Any severe TEAE, related to IMP 
• Any TEAE with outcome of death 
• Any TEAE with outcome of death, related to IMP 
• Any TESAE  
• Any TESAE, related to IMP 
• Any TEAE leading to discontinuation of IMP 
• Any TESAE leading to discontinuation of IMP 
• Any TEAE leading to discontinuation of IMP, related to IMP 
• Any TEAE leading to dose interruption 
TEAE summaries will present the number and percentage of subjects reporting at least one 
TEAE. The following summaries will be provided: 
• TEAEs by SOC and PT 
• TEAEs by SOC and PT, by maximum severity 
• TEAEs by SOC and PT, by maximum relationship 
• TEAEs related to IMP, by SOC and PT 
• TESAEs by SOC and PT 
• TESAEs by SOC and PT, by maximum severity 
• TESAEs by SOC and PT, by maximum relationship 
• TESAEs related to IMP, by SOC, PT 
• TEAE leading to IMP permanently stopped, by SOC and PT 
• TEAE leading to death, by SOC and PT 
The number and percentage of deaths will also be summarized. 
The following by-subject AE listings will be provided: 
• TEAEs 
• TESAEs 
• TEAEs related to IMP 
• TEAEs leading to drug permanently stopped 
• TEAEs leading to death 
• PTAEs 
• Deaths 
 
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The listings will include: subject identifier, age, sex, race, adverse event (SOC, PT, and 
verbatim term), period, date of onset, date of resolution, duration, severity, seriousness, 
action taken, outcome and causality.  
4.10.2 Clinical Laboratory Evaluation 
Laboratory values (hematology, biochemistry, and urinalysis) will be listed by subject and 
study time point including changes from baseline for continuous variables. The baseline 
for the laboratory values will be the results obtained on the last measurement before the 
first dose. 
All values outside the clinical reference ranges will be flagged in the data listings. The 
abnormal values will be flagged with ‘L’ for values below the lower limit of the clinical 
reference range and ‘H’ for values above the upper limit of the clinical reference range and 
included in the listings. The Investigator will assess whether the values outside the clinical 
reference range are clinically significant and these will be reported as “abnormal, not 
clinically significant (NCS)” or “abnormal, clinically significant (CS)”. Clinically 
significant laboratory values will be recorded by the Investigator as AEs. 
Quantitative laboratory assessment reported as “<X”, i.e. BQL, or “>X”, i.e. above the 
upper limit of quantification, will be converted to X for the purpose of quantitative 
summaries, but will be presented as recorded, i.e. as “<X” or “>X” in the listings. 
The toxicity grading of laboratory tests is determined based on the NCI CTCAE V4.03 
(Appendix 3). 
The following summaries will be provided for laboratory data: 
• 
Absolute value and change from baseline for continuous data by scheduled time point. 
• 
The number and percentage of subjects in each test result category for categorical data, 
by scheduled time point. 
• 
Shifting from baseline of subjects with lab results evaluation as Normal, NCS, and CS, 
by scheduled time point. 
• 
Shifting from baseline of subjects with lab results evaluated by NCI CTCAE V4.03 by 
toxicity grading and scheduled time point. 
The following laboratory data listings will be provided: 
• 
Listing of laboratory test results 
• 
Listing of abnormal laboratory test results 
• 
Listing of other lab tests and pregnancy tests. 
4.10.3 Vital Signs, body weight, 12-Lead Electrocardiogram (ECG), and Physical 
Examinations 
 
The following summaries and listings will be provided for vital signs and body weight: 
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• 
Absolute value and change from baseline of vital signs parameters and body weight 
with descriptive statistics, by scheduled time point. 
• 
Listing of vital signs and body weight data. 
12-Lead ECG will be performed at screening only and will not be summarized. A listing 
will be provided for ECG performance and interpretation. 
Physical examinations will not be summarized. A listing of physical examination data will 
be provided with investigator’s interpretation.  
4.11 Determination of Sample Size 
The sample size was not calculated through any statistical calculation. A sample size of 24 
subjects was estimated as sufficient for the descriptive purposes of the present study. 
Drop-out subjects will not be replaced.  
 
4.12 Changes in the Conduct of the Study or Planned Analysis 
Not Applicable. 
5 
REFERENCES 
No Reference. 
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                                     Project Document Version No. 3.1 
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Project Document Effective Date: Date of last signature 
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                                                                                                                                                                                   Page 24 of 31 
6 
APPENDIX 
APPENDIX 1 Study Schedule 
ACTIVITIES 
Screening 
Single Dose 
Multiple Dose 
Final 
visit/ETV1 
Visit 
V1 
V2 
V3 
V4 
V5 
 Day -14/-2 Day -1 
Day 1 
Day 2 
Day 3 
Day 4 
Day 5 
Day 6 
Day 7 
Day 82 
Informed consent 
x 
 
 
 
 
 
 
 
 
 
Demography and lifestyle 
x 
 
 
 
 
 
 
 
 
 
Medical and surgical history  
x 
 
 
 
 
 
 
 
 
 
Physical examination 
x 
 
 
 
x 
 
 
 
 
x 
Prior and concomitant 
medications 
x 
x 
x 
x 
x 
x 
x 
x 
x 
x 
Height 
x 
 
 
 
 
 
 
 
 
 
Body Weight 
x 
 
 
 
x 
 
 
 
 
x 
Clinical laboratory analysis 
(haematology, blood chemistry, 
urinalysis)  
x 
 
 
 
x 
 
 
 
 
x 
Virology and bacteriology  
x 
 
 
 
 
 
 
 
 
 
Serum pregnancy test (women) 
x 
 
 
 
 
 
 
 
 
 
Urine multi-drug kit test 
x 
x 
 
 
 
 
 
 
 
 
Blood pressure and heart rate 
x 
 
x3 
 
x 
 
 
x3 
 
x4 
Alcohol breath test 
 
x 
 
 
 
 
 
 
 
 
Urine pregnancy test (women) 
 
x 
 
 
 
 
 
 
 
 
ECG 
X 
 
 
 
 
 
 
 
 
 
Inclusion/exclusion criteria  
X 
x 
 
 
 
 
 
 
 
 
Subject eligibility 
X 
x 
 
 
 
 
 
 
 
 
Enrolment  
 
x 
 
 
 
 
 
 
 
 
Confinement 
 
x 
x 
x 
x 
x 
x 
x 
x 
 
Discharge 
 
 
 
 
 
 
 
 
 
x 
Investigational product 
administration 
 
 
x5 
 
 
x6 
x6 
x5 
 
 
Blood sampling  
 
 
x7 
x7 
 
x8 
x8 
x7 
x7 
 
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                                                                                                                                                                                   Page 25 of 31 
Urine sampling  
 
 
x9 
x9 
 
 
 
x9 
x9 
 
Standardised meals 
 
x10 
x11 
x12 
x12 
x12 
x12 
x11 
x12 
 
Adverse event monitoring13 
x 
x 
x 
x 
x 
x 
x 
x 
x 
x 
1. 
Early termination visit (ETV) 
2. 
Final visit on day 8 or in case of ETV 
3. 
At pre-dose and 1 h post-dose 
4. 
The vital signs check at 48 h post-dose (day 8), will correspond to the final measurement 
5. 
At 8:00 ± 1 h: 5-min infusion 
6. 
At 8:00 ± 1 h and at 20:00 ± 1 h; 5-min infusion 
7. 
At pre-dose (0) and 5 (at the end of the infusion), 8, 12, 15, 20, 25, 30, 60 min and 2, 4, 6, 8, 10, 12, 24 and 32 h post-dose  
8. 
At pre-dose (0) 
9. 
0-4; 4-8; 8-12, 12-24 and 24-32 h post-dose  
10. Standardised dinner 
11. Standardised lunch and dinner 
12. Standardised breakfast, lunch and dinner  
13. Adverse events monitored starting at the screening visit, immediately after informed consent, up to the final visit/ETV 
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APPENDIX 2 Partial Date Conventions 
Imputed dates will NOT be presented in the listings. However, in general, when calculating relative 
days, partial dates of ECG, laboratory data, standardized meals, physical examination, substance 
usage and vital sign with missing day only will be assumed to be 15th of the month, and partial 
dates with both missing day and month will be assumed to be June 30.  
Otherwise, for partial dates of medical history, concomitant medication and procedures the 
following rules in the given table will be applied for each case. 
Algorithm for Medical History, Prior/Concomitant Medications, Therapies or 
Procedures: 
Start Date 
Stop Date 
Action 
Known 
Known 
If stop date is prior to the date of first dose of study drug, considered as 
prior; 
if start date is on or after the date of first dose of study drug, considered 
as concomitant. 
 
Partial 
The last day of the month and the last month (i.e. December) will be 
used if the day/month of stop date is missing. 
If the impuated stop date is prior to the date of first dose of study drug, 
considered as prior; 
if the imputed stop date is on or after the date of first dose of study drug, 
considered as concomitant. 
 
Missing 
Considered as concomitant. 
 
Partial 
Known 
The first day of the month and January will be used if the start 
day/month is missing. 
If stop date is prior to the date of first dose of study drug, considered as 
prior; 
if the stop date is on or after the date of first dose of study drug, 
considered as concomitant. 
 
Partial 
The first day of the month and January will be used if the start 
day/month is missing. 
The last day of the month and the last month (i.e. December) will be 
used if the day/month of stop date is missing. 
If the imputed stop date is prior to the date of first dose of study drug, 
considered as prior; 
if the imputed stop date is on or after the date of first dose of study drug, 
considered as concomitant. 
 
Missing 
The first day of the month and January will be used if the start 
day/month is missing. 
Considered as concomitant. 
 
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Start Date 
Stop Date 
Action 
Missing 
Known 
If stop date is prior to the date of first dose of study drug, considered as 
prior;  
if stop date is on or after the date of first dose of study drug, considered 
as concomitant; 
 
Partial 
The last day of the month and the last month (i.e. December) will be 
used if the day/month of stop date is missing. 
If the impuated stop date is prior to the date of first dose of study drug, 
considered as prior; 
if the imputed stop date is on or after the date of first dose of study drug, 
considered as concomitant. 
 
Missing 
Considered as concomitant. 
 
 
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Algorithm for Treatment-Emergent Adverse Events: 
Start/ Increase Severity Date 
Stop Date 
Action 
Known 
Known 
Considered as a TEAE if start/increase 
severity date on or after the date of the 
first dose of study drug. 
 
Partial 
Considered as a TEAE if start/increase 
severity date on or after the date of the 
first dose of study drug and on or before 
the last dose of study drug + 30 days. The 
last day of the month and the last month 
(i.e. December) will be used if the stop 
day/month is missing. 
 
Missing 
Considered as a TEAE if start/increase 
severity date on or after the date of the 
first dose of study drug and on or before 
the last dose of study drug + 30 days. 
 
Partial, but known components 
show that it cannot be on or after 
first study drug taken date. 
Known 
Not a TEAE. The first day of the month 
and January will be used if the start 
day/month or the increase severity 
day/month is missing. 
 
Partial 
Not a TEAE. The first day of the month 
and January will be used if the start 
day/month or the increase severity 
day/month is missing. The last day of the 
month and the last month (i.e. December) 
will be used if the stop day/month is 
missing. 
 
Missing 
 Not a TEAE. The first day of the month 
and January will be used if the start 
day/month or the increase severity 
day/month  is missing. 
 
Partial, could be on or after first 
study drug taken date 
Known 
Considered as TEAE, if stop date is after 
first study drug taken date. The first study 
drug taken date will be used if 
start/increase severity date is in the same 
month/year with first study drug taken 
date, or the first day of the month and 
January will be used if the start day/month 
or the increase severity day/month is after 
first of study drug taken date. 
Considered as not TEAE, if stop date is 
prior to first of study drug taken date. The 
first day of the month and January will be 
used if the start day/month or the increase 
CCI


Zambon S.p.A. 
Z7244J01 
Statistical Analysis Plan 
 
TP-GDO-WW-016-06 
CONFIDENTIAL 
Project Document Version No. 4.0 
Effective Date:  28 Jun 17 
Project Document Effective Date: 15 Jun 2020 
Related to:  
 
Page 29 of 31 
  
Start/ Increase Severity Date 
Stop Date 
Action 
severity day/month is missing. 
 
Partial 
Considered as TEAE. The first of study 
drug taken date will be used if start/ 
increase severity date is in the same 
month/year with first study drug taken 
date, or the first day of the month and 
January will be used if the start day/month 
or the increase severity day/month is after 
first study drug taken date. The last day of 
the month and the last month (i.e. 
December) will be used if the stop 
day/month is missing. 
 
Missing 
Considered as TEAE. The first study drug 
taken date will be used if start/ increase 
severity date is in the same month/year 
with first study drug taken date, or the first 
day of the month and January will be used 
if the start day/month or the increase 
severity day/month is after first study 
drug taken date. 
 
Missing 
Known 
Considered as TEAE if stop date is on or 
after the date of the first dose of study 
drug. 
 
Partial 
The last day of the month and the last 
month (i.e. December) will be used if the 
stop day/month is missing. If the imputed 
stop date is on or after the first dose of 
study drug considered as a TEAE; if the 
year is missing, considered as a TEAE. 
 
Missing 
Considered as a TEAE 
CCI


Zambon S.p.A. 
Z7244J01 
Statistical Analysis Plan 
 
TP-GDO-WW-016-06 
CONFIDENTIAL 
Project Document Version No. 4.0 
Effective Date:  28 Jun 17 
Project Document Effective Date: 15 Jun 2020 
Related to:  
 
Page 30 of 31 
  
APPENDIX 3 CTC Grading for Laboratory Parameters 
Laboratory measurements will be graded using the NCI-CTCAE v4.03. 
 
  
 
  
  
Grade 
Analytic 
 
Direction 
Unit 
0 
1 
2 
3 
4 
 
Hematology 
Hemoglobin 
 
Hypo 
g/L 
LLN =< 
Value  
100 =< Value 
< LLN 
80 =< Value < 
100 
Value < 80 
 
 
Hypo 
g/dL 
LLN =< 
Value  
10 =< Value < 
LLN 
8 =< Value < 10 
Value < 8 
 
 
Hypo 
mmol/L 
LLN =< 
Value 
6.2 =< Value 
< LLN 
4.9 =< Value < 
6.2 
Value < 4.9 
 
WBC  
 
Hypo 
10^9/L 
LLN =< 
Value  
3 =< Value < 
LLN 
2 =< Value < 3 
1 =< Value < 
2 
Value < 1 
 
Hypo 
/mm3 
LLN =< 
Value  
3000 =< 
Value < LLN 
2000 =< Value < 
3000 
1000 =< 
Value < 
2000 
Value < 
1000 
Neutrophils 
 
Hypo 
10^9/L 
LLN =< 
Value  
1.5 =< Value 
< LLN 
1 =< Value < 1.5 
0.5 =< Value 
< 1 
Value < 
0.5 
 
Hypo 
/mm3 
LLN =< 
Value 
1500 =< 
Value < LLN 
1000 =< Value < 
1500 
500 =< 
Value < 
1000 
Value < 
500 
Lymphocytes 
 
Hypo 
10^9/L 
LLN =< 
Value 
0.8 =< Value 
< LLN 
0.5 =< Value < 
0.8 
0.2 =< Value 
< 0.5 
Value < 
0.2 
 
Hypo 
/mm3 
LLN =< 
Value 
800 =< Value 
< LLN 
500 =< Value < 
800 
200 =< 
Value < 500 
Value < 
200 
 
Hyper 
10^9/L 
Value <= 
ULC 
 
4 <  Value =< 20 
20 < Value 
 
 
Hyper 
/mm3 
Value <= 
ULN 
 
4000 <  Value =< 
20,000 
20,000 < 
Value 
 
Monocytes 
 
NA 
Eosinophils 
 
NA 
Basophils 
 
NA 
Platelet count 
 
Hypo 
10^9/L 
LLN =< 
Value 
75 =< Value < 
LLN 
50 =< Value < 75 
25 =< Value 
< 50 
Value < 
25 
 
Hypo 
/mm3 
LLN =< 
Value 
75,000 =< 
Value < LLN 
50,000 =< Value 
< 75,000 
25,000 =< 
Value < 
50,000 
Value < 
25,000 
 
Biochemistry 
  
Serum creatinine 
 
Hyper 
 
umol/L 
 
Value <= 
ULN 
 
ULN<  Value 
=< 1.5 × ULN 
 
1.5 × ULN<  
Value =< 3 × 
ULN 
 
3 × ULN<  
Value =< 6 × 
ULN 
 
6 × ULN<  
Value 
BUN 
 
NA 
Alanine 
aminotransferase 
(ALT) 
 
Hyper 
U/L 
Value <= 
ULN 
ULN <  Value 
=< 3 × ULN 
3 × ULN<  Value 
=< 5 × ULN 
5 × ULN<  
Value =< 30 
× ULN 
20 × 
ULN<  
Value 
Aspartate 
aminotransferase 
(AST) 
 
Hyper 
U/L 
 Value <= 
ULN 
 
ULN <  Value 
=< 3 × ULN 
3 × ULN<  Value 
=< 5 × ULN 
5 × ULN<  
Value =< 30 
× ULN 
20 × 
ULN<  
Value 
Alkaline 
phosphatase 
(ALP) 
 
Hyper 
U/L 
Value <= 
ULN 
ULN <  Value 
=< 2.5 × ULN 
2.5 × ULN<  
Value =< 5 × 
ULN 
5 × ULN<  
Value =< 20 
× ULN 
20 × 
ULN<  
Value 
Total bilirubin 
 
Hyper 
umol/L 
Value <= 
ULN 
ULN <  Value 
=< 1.5 × ULN 
1.5 × ULN<  
Value =< 3 × 
ULN 
3 × ULN<  
Value =< 10 
× ULN 
10 × 
ULN<  
Value 
Albumin 
 
Hypo 
g/L 
LLN =< 
Value  
30 =< Value < 
LLN 
20 =< Value < 30 
Value < 20 
 
CCI


Zambon S.p.A. 
Z7244J01 
Statistical Analysis Plan 
 
TP-GDO-WW-016-06 
CONFIDENTIAL 
Project Document Version No. 4.0 
Effective Date:  28 Jun 17 
Project Document Effective Date: 15 Jun 2020 
Related to:  
 
Page 31 of 31 
  
  
 
  
  
Grade 
Analytic 
 
Direction 
Unit 
0 
1 
2 
3 
4 
 
Hypo 
g/dL 
LLN =< 
Value  
3 =< Value < 
LLN 
2 =< Value < 3 
Value < 2 
 
Sodium 
 
Hypo 
mmol/L 
LLN =< 
Value 
130 =< Value 
< LLN 
 
120 =< 
Value < 130 
Value < 
120 
 
Hyper 
mmol/L 
LLN =< 
Value 
ULN <  Value 
=< 150 
150 <  Value =< 
155 
155 <  Value 
=< 160 
160 <  
Value 
Potassium 
 
Hypo 
mmol/L 
LLN =< 
Value 
3 =< Value < 
LLN 
 
2.5 =< Value 
< 3 
Value < 
2.5 
 
Hyper 
mmol/L 
Value <= 
ULN 
ULN <  Value 
=< 5.5 
5.5 <  Value =< 6 
6 <  Value 
=< 7 
7 <  Value 
Chloride 
 
NA 
 
Serology 
  
HBV 
 
NA 
HCV 
 
NA 
 
Urinalysis 
  
Urine protein 
 
Hyper 
  
Negative 
1+ 
2+ 
  
  
 
Hyper 
g/24hrs 
Value <= 
ULN 
ULN <  Value 
< 1 
1 =<  Value =< 
3.4 
3.5= <  
Value 
  
 
Pregnancy test 
  
Pregnancy test 
 
 NA  
LLN: lower limit of normal; ULN: upper limit of normal. 
 
CCI