Detection of Pseudomonas. Aeruginosa in the Airways of Patients With Cystic Fibrosis Using of Aerosolized 13C-urea
Phase: NA
Conditions: Cystic Fibrosis
Sponsor: University of New Mexico

University of New Mexico 
Clinical Study Protocol 
 
Study Title: 
A Dose-escalation Study to Detect Urease-producing Bacteria in Lungs of 
Healthy Volunteers and Subjects with Cystic Fibrosis Using Aerosolized 13C-urea 
Test Product: 
 
13C-urea Breath Test Kit (IDE# G120190) 
 
HRPO Number: 
12-521 
Principal Investigator: 
Hengameh Raissy, PharmD 
Research Associate Professor of Pediatrics 
University of New Mexico 
Department of Pediatrics, MSC 10 5590 
1 University of New Mexico 
Albuquerque, NM  87131 
505-272-5484 
Co-investigators: 
Lea Davies, MD 
Assistant Professor of Pediatrics 
Division Chair, Pediatric Pulmonary 
The University of New Mexico 
Department of Pediatrics, MSC10-5590 
1 University of New Mexico 
Albuquerque, NM  87131 
(505) 272-5551 
Michelle Harkins, MD  
Associate Professor of Medicine 
The University of New Mexico 
Department of Internal Medicine, Pulmonary Division, MSC10-5590 
1 University of New Mexico 
Albuquerque, NM  87131 
(505) 272-4751 
 
 
Date: 
29, November 2012 
Report Status: 
Version 3 
 
 
 
 

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STUDY SYNOPSIS 
Location of study: 
University of New Mexico Hospital, 
Pediatric/Adult Pulmonary Clinic, 
Ambulatory Care Center, Albuquerque, NM 
Tabulated Study Protocol 
 
Finished product:   
13C-urea Breath Test Kit 
Active ingredient: 
13C-urea 
Protocol date 
29 November 2012 
HRPO# 
12-521 
Planned study period 
January1, 2013-March 31,2013 
Title of Study: 
A Dose-escalation Study to Detect Urease-producing Bacteria in Lungs of Healthy Volunteers and 
Subjects with Cystic Fibrosis Using Aerosolized 13C-urea 
Indication:  Use of a 13C-urea breath test in the qualitative detection of urease-containing bacteria in the 
respiratory tract and as an aid in the initial and post-treatment detection of respiratory tract infections 
Study Design:  This is a single-center, single-administration, dose-escalation study designed to determine 
the safety and dose response of aerosolized 13C-urea 20 mg and 50 mg in detecting urease-producing 
bacteria in the lungs of healthy volunteers (those without and those diagnosed with Helicobacter pylori 
infection) and in subjects with cystic fibrosis (CF).  Subjects will be contacted by telephone the next day 
and asked a non-leading question about adverse events. 
The dose will not be escalated until at least 24 hours after all subjects in the cohort have been tested at the 
current dose.  If a subject has an adverse event or new symptom that remains unresolved at 48 hours, no 
new subjects will be enrolled until the relationship between the AE and symptom can be evaluated.  If any 
subject experiences an adverse event considered by the data safety monitor to be dose limiting, then three 
additional subjects will be enrolled at that current dose.  Dose escalation will stop if more than one 
additional subject experiences dose-limiting toxicity.  
After the safety of the maximum dose in the cohort of healthy volunteers without H. pylori infection has 
been reviewed by a data safety monitor, healthy volunteers diagnosed with H. pylori infection and 
subjects with CF will be enrolled.  Three subjects diagnosed with H. pylori infection will be tested at the 
maximum dose to assess potential interference of urease producing bacteria in the stomach; subjects 
infected with H. pylori can be enrolled at any time after the safety of the maximum dose in healthy 
volunteers without H. pylori infection has been evaluated, but they will meet the entrance criteria for 
healthy volunteers without H. pylori infection. 
Sputum will be collected from each subject with CF before administering study drug in order to identify 
bacterial flora using two standard microbiologic methods. The first is a quantitative method in which 
colonies are stained and counted, the second is a colorimetric method in which a sample is plated in a 
medium that changes to a blue color as its pH is increased by the production of ammonia, a by-product of 
the urease-catalyzed degradation of urea. 
Study Duration:  The maximum duration of study participation for each subject is approximately 9 days:  
up to 7 days between the screening visit and the study visit plus a follow-up visit 24 hours after study 
drug administration. 

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Location of study: 
University of New Mexico Hospital, 
Pediatric/Adult Pulmonary Clinic, 
Ambulatory Care Center, Albuquerque, NM 
Tabulated Study Protocol 
 
Finished product:   
13C-urea Breath Test Kit 
Active ingredient: 
13C-urea 
Protocol date 
29 November 2012 
HRPO# 
12-521 
Planned study period 
January1, 2013-March 31,2013 
Study Objectives:  The primary objective is to assess the safety of 13C-urea administered by inhalation. 
The secondary objective is to assess the kinetics of 13C-carbon dioxide production by measuring the 
isotopic ratio of 13C to 12C in exhaled carbon dioxide up to 15 minutes after administration and to assess 
potential interference of urease-producing bacteria in the stomach. 
Study Endpoints:  The primary endpoint is the safety of inhaled 13C-urea assessed by adverse events 
including pulmonary findings and changes from baseline in physical examination findings, pulse 
oximetry results, pulmonary function test results, and vital signs.  Secondary endpoints are the isotopic 
ratios of 13C to 12C in exhaled carbon dioxide measured with the POCone detector (Meretek Diagnostics 
Group of Otsuka America Pharmaceutical, Rockville, MD) at 5, 10, and 15 minutes after study drug 
administration. 
Study Population: 
Subjects who meet all the following inclusion and exclusion criteria are eligible to enroll in the study. 
Inclusion Criteria for Healthy Volunteers Without H. pylori infection: 
Healthy volunteers who meet all the following inclusion criteria are eligible to enroll. 
• 
At least 18 years old at the time of providing informed consent in a manner approved by the 
University of New Mexico, Institutional Review Board and willing to comply with the requirements 
of the study  
• 
In generally good health as determined by the investigator, with no respiratory illness within 2 weeks 
before the screening visit and no pulmonary history 
• 
No history of allergy or asthma 
• 
Has not smoked tobacco within 6 months before the screening visit and agrees not to smoke until 
after the follow up visit 
• 
Forced expiratory volume in 1 second (FEV1) at least 80% at the time of providing informed consent 
Inclusion Criteria for Healthy Volunteers Diagnosed With H. pylori infection: 
Healthy volunteers diagnosed with H. pylori infection who meet all the inclusion criteria for healthy 
volunteers and additionally who have been diagnosed with H. pylori infection are eligible to enroll. 
Inclusion Criteria for Subjects With Cystic Fibrosis: 
Subjects with CF who meet all the following inclusion criteria are eligible to enroll. 
• 
At least 18 years old at the time of providing informed consent in a manner approved by the 
University of New Mexico, Institutional Review Board and willing to comply with the requirements 
of the study 
• 
Diagnosed with CF at least 24 months before the screening visit 

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Location of study: 
University of New Mexico Hospital, 
Pediatric/Adult Pulmonary Clinic, 
Ambulatory Care Center, Albuquerque, NM 
Tabulated Study Protocol 
 
Finished product:   
13C-urea Breath Test Kit 
Active ingredient: 
13C-urea 
Protocol date 
29 November 2012 
HRPO# 
12-521 
Planned study period 
January1, 2013-March 31,2013 
• 
Documented presence of Pseudomonas organisms in at least three sputum cultures within 2 years 
before the screening visit, one of which within 6 months before the screening visit 
• 
Is able to produce sputum at the time of the screening visit 
• 
FEV1>60% or more than 1.5 L at the time of providing informed consent 
• 
Has not smoked tobacco within 6 months before the screening visit and agrees not to smoke until 
after the follow up visit 
Exclusion Criteria 
Subjects who meet any of the following exclusion criteria are not eligible to enroll. 
• 
Positive H. pylori serology, except for subjects in the H. pylori cohort 
• 
Subjects intolerant of albuterol 
• 
Subjects diagnosed with asthma 
• 
Subjects with evidence of upper respiratory tract infection within 14 days before the screening visit 
• 
Use of inhaled antibiotics within 12 hours before the administration of the breath test on Day 1 or 
required use of inhaled antibiotics during the study 
• 
Has smoked tobacco within 6 months before the screening visit 
• 
Females who are pregnant or nursing or of child-bearing potential who are not using a medically 
acceptable form of contraception (acceptable forms of contraception are hormonal birth control, 
intrauterine device, or barrier method plus a spermicidal agent) or abstinence 
• 
Any condition or history that, in the judgment of the investigator, would compromise the ability of 
the subject to comply with the study protocol or to complete the study 
• 
Subjects who have used an investigational agent within 28 days before Day 1 
Study Product, Dose, and Mode of Administration: 
13C-urea:  20 mg and 50 mg administered by inhalation 
Reference Product, Dose, and Mode of Administration:  Not applicable 
Efficacy Variables:  Breath test results 
Safety Variables:  Adverse events (frequency, intensity, seriousness, and relationship to study drug), 
spirometry test results (FEV1 and FVC), and respiratory examinations (respiratory rate, auscultation 
[presence of wheezing and crackles], pulse oximetry, and related symptoms, such as cough) 
Pharmacokinetics:  Production of carbon dioxide 
Sample Size and Statistical Rationale:  Approximately 15 subjects will be enrolled, 9 healthy 
volunteers and 6 subjects with CF.  Three healthy volunteers without H. pylori infection will each receive 

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Location of study: 
University of New Mexico Hospital, 
Pediatric/Adult Pulmonary Clinic, 
Ambulatory Care Center, Albuquerque, NM 
Tabulated Study Protocol 
 
Finished product:   
13C-urea Breath Test Kit 
Active ingredient: 
13C-urea 
Protocol date 
29 November 2012 
HRPO# 
12-521 
Planned study period 
January1, 2013-March 31,2013 
a single 20 mg dose and 3 will each receive a single 50 mg dose.  After the safety of the 50 mg dose has 
been evaluated, 3 subjects with CF will each receive a single 20 mg dose and 3 subjects with CF will each 
receive a single 50 mg dose.  Three healthy volunteers diagnosed with H. pylori infection will also each 
receive a 50 mg dose to assess potential interference of urease producing bacteria in the stomach.  Three 
additional subjects will be added to every dose level at which a dose-limiting adverse event occurs; no 
more than 30 subjects will be enrolled.  This sample size is based upon a dose-escalation model, not a 
calculation of statistical power. 
Study Sites and Location:  The study will be conducted at a single center at the University of New 
Mexico Hospital, Pediatric/Adult Pulmonary Clinic, Ambulatory Care Center, Albuquerque, NM. 

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TABLE OF CONTENTS 
 
STUDY SYNOPSIS ........................................................................................................................2 
TABLE OF CONTENTS .................................................................................................................6 
LIST OF ABBREVIATIONS AND DEFINITIONS OF TERMS ..................................................9 
1. 
INTRODUCTION .............................................................................................................10 
1.1 
Cystic Fibrosis .......................................................................................................10 
1.2 
13C-Urea Breath Test ..............................................................................................10 
1.3 
Urea ........................................................................................................................11 
1.4 
Rationale ................................................................................................................12 
1.5 
Benefit-to-risk Assessment ....................................................................................12 
2. 
OBJECTIVES ....................................................................................................................13 
2.1 
Primary Objective ..................................................................................................13 
2.2 
Secondary Objectives .............................................................................................13 
3. 
INVESTIGATIONAL PLAN ............................................................................................13 
3.1 
Overall Description of Study Design and Plan ......................................................13 
3.1.1 
Primary Endpoint .................................................................................13 
3.1.2 
Secondary Endpoint .............................................................................13 
3.1.3 
Study Design ........................................................................................13 
3.2 
Discussion of Study Design ...................................................................................15 
3.2.1 
Number of Subjects..............................................................................16 
3.2.2 
Duration of Study .................................................................................16 
3.2.3 
Premature Termination of Study ..........................................................17 
4. 
SELECTION OF STUDY POPULATION .......................................................................17 
4.1 
Inclusion Criteria ...................................................................................................17 
4.1.1 
Healthy Volunteers Without Helicobacter pylori infection .................17 
4.1.2 
Healthy Volunteers Diagnosed With Helicobacter pylori infection ....17 
4.1.3 
Inclusion Criteria for Subjects With Cystic Fibrosis ...........................17 
4.2 
Exclusion Criteria ..................................................................................................18 
4.3 
Removal of Subjects ..............................................................................................18 
4.4 
Replacement of Subjects ........................................................................................18 
5. 
PROCEDURES FOR ADMINISTRATION OF TREATMENTS....................................19 
5.1 
Treatments to be Administered ..............................................................................19 
5.1.1 
Identification of Investigational Product .............................................19 
5.1.2 
13C-urea Breath Test Kit ......................................................................19 
5.1.3 
Administration of Study Drug .............................................................19 
5.1.4 
Pre-treatment ........................................................................................20 
5.1.5 
Rescue Medication ...............................................................................20 
5.1.6 
Methods of Assigning Subjects to Treatment Group ...........................20 
5.1.7 
Selection of Doses in the Study ...........................................................20 
5.1.8 
Selection and Timing of Doses for Each Subject ................................20 
5.1.9 
Blinding................................................................................................21 
5.1.10 
Packaging and Labeling .......................................................................21 

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5.1.11 
Storage .................................................................................................21 
5.1.12 
Accountability and Destruction of Study Drug ...................................21 
5.2 
Concomitant Therapy.............................................................................................21 
5.3 
Treatment compliance ............................................................................................21 
6. 
VISIT SCHEDULE ...........................................................................................................22 
6.1 
Screening (Day –7 to Day –1) ...............................................................................24 
6.2 
Active Treatment (Day 1) ......................................................................................24 
6.3 
Follow-up ...............................................................................................................24 
6.4 
Discontinuation ......................................................................................................25 
7. 
STUDY VARIABLES AND METHODS OF ASSESSMENT ........................................25 
7.1 
Efficacy Variable ...................................................................................................25 
7.1.1 
Overview of Variable ...........................................................................25 
7.1.2 
Methods of Assessment .......................................................................25 
7.1.3 
Microbial Analysis of Sputum .............................................................26 
7.2 
Safety variables ......................................................................................................26 
7.2.1 
Spirometry............................................................................................26 
7.2.2 
Respiratory Examination .....................................................................27 
7.3 
Pharmacokinetics ...................................................................................................27 
7.4 
Appropriateness of Measurements .........................................................................27 
8. 
ADVERSE EVENTS, SERIOUS ADVERSE EVENTS, AND SUSPECTED 
ADVERSE REACTIONS ..................................................................................................27 
8.1 
Definition of an Adverse Event .............................................................................27 
8.2 
Definition of a Suspected Adverse Reaction .........................................................28 
8.3 
Definition of a Serious Adverse Event or Serious Suspected Adverse Reaction ..29 
8.3.1 
Serious Adverse Events That Occur Before Administration of 
Study Medication .................................................................................29 
8.3.2 
Serious Adverse Events That Occur After Last Administration of 
Study Drug ...........................................................................................29 
8.4 
Recording and Evaluating Adverse Events and Serious Adverse Events ..............30 
8.4.1 
Assessment of Intensity .......................................................................30 
8.4.2 
Assessment of Causality ......................................................................30 
8.4.3 
Assessment of Outcome .......................................................................31 
8.5 
Data Safety Monitor ...............................................................................................31 
8.6 
Follow-up of Adverse Events and Serious Adverse Events ..................................31 
8.7 
Prompt Reporting of Serious Adverse Events to the Data safety monitor ............32 
8.8 
Regulatory Reporting Requirements ......................................................................33 
8.9 
Precautions .............................................................................................................33 
9. 
STATISTICS .....................................................................................................................33 
9.1 
Determination of Sample Size ...............................................................................33 
9.2 
Randomization and Blinding .................................................................................34 
9.3 
Analysis Populations ..............................................................................................34 
9.4 
Statistical Analyses and Methods ..........................................................................34 
10. 
STUDY ADMINISTRATION ..........................................................................................35 
10.1 
Regulatory and Ethical Considerations ..................................................................35 
10.1.1 
Regulatory Authority Approval ...........................................................35 

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10.1.2 
Ethical Conduct of the Study and Ethics Approval .............................35 
10.1.3 
Informed Consent.................................................................................36 
10.1.4 
Investigator Reporting Requirements ..................................................37 
10.2 
Study Monitoring ...................................................................................................37 
10.3 
Quality Assurance ..................................................................................................37 
10.4 
Study and Site Closure ...........................................................................................38 
10.5 
Records Retention ..................................................................................................38 
10.5.1 
Health Insurance Portability and Accountability Act of 1996 .............38 
10.5.2 
Financial Disclosure .............................................................................38 
10.5.3 
Access to Original Records ..................................................................38 
10.5.4 
Archiving of Study-Related Documents ..............................................39 
10.6 
Subject Tracking ....................................................................................................39 
11. 
REFERENCES ..................................................................................................................39 
 
 
 
LIST OF IN-TEXT TABLES 
Table 1 
Dose Escalation Rules........................................................................................... 14 
Table 2 
Identification of Investigational Product .............................................................. 19 
Table 3 
Identification of Pre-treatment .............................................................................. 20 
Table 4 
Schedule of Procedures and Assessments for Dose-ranging Study ...................... 23 
 

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LIST OF ABBREVIATIONS AND DEFINITIONS OF TERMS 
 
Abbreviation 
Term 
AE 
adverse event 
CF 
cystic fibrosis 
CFR 
Code of Federal Regulations 
COPD 
chronic obstructive pulmonary disease 
CRF 
case report form 
DLT 
dose-limiting toxicity 
FDA 
Food and Drug Administration 
FEV1 
forced expiratory volume in 1 second 
FVC 
forced vital capacity 
GCP 
Good Clinical Practice 
ICH 
International Conference on Harmonization 
IEC 
Independent Ethics Committee 
IRB 
Institutional Review Board 
MedDRA 
Medical Dictionary for Regulatory Activities 
SAE 
serious adverse event 
SAR 
suspected adverse reaction 
SSAR 
serious suspected adverse reaction 
TEAE 
treatment-emergent adverse event 
WFI 
water for injection 
 

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1. 
INTRODUCTION 
1.1 
CYSTIC FIBROSIS 
Cystic fibrosis (CF) is a genetic disease which affects approximately 30,000 people in the United 
States.  Prior to the development of therapies, most people with CF died in childhood.  New 
advances in therapies have increased the average lifespan to about 35 years with respiratory 
failure continuing to be the primary cause of death. 
The CF gene that codes the CF protein (cystic fibrosis transmembrane conductance regulator 
[CFTR]) was discovered in 1989.  CFTR is a protein that regulates the flows of ions, including 
chloride and bicarbonate, across membranes. 
One consequence of abnormal CFTR is that glandular fluids thicken, which leads to obstruction 
of the ducts.  In the pancreas, this obstruction blocks the release of digestive enzymes into the 
gastrointestinal tract, resulting in poor absorption of nutrients.  Patients with CF take 
replacement pancreatic enzymes before every meal to aid digestion and absorption. 
In the lungs of patients with CF, thickened layers of fluid lining the airways obstruct the removal 
of bacteria.  Bacteria that are normally present in healthy lungs, and that are normally washed out 
by movement of thin layers of surface fluid, are not cleared easily from lungs of patients with 
CF.  Urease-producing bacteria Staphylococcus aureus and pseudomonas are the most prominent 
bacteria found in increased numbers in lungs of patients with CF.  As bacterial loads increase, 
patients with CF start to develop mildly increased symptoms such as cough, but they generally 
do not develop the dramatic signs of lung infection seen in healthy patients who have sudden 
increases in bacteria that develop into pneumonia. 
Patients with CF are followed in CF centers where the insidious nature of increasing bacterial 
loads is followed by routine sputum or throat cultures at least every three months.  The bacterial 
profile of each patient is known and used to guide aggressive antibiotic therapy for 
exacerbations.  All bacteria are not cleared with antibiotic therapy, but it is believed that 
antibiotic therapy decreases bacterial burden.  It is further believed that patients with CF would 
experience fewer or less severe exacerbations if more timely antibiotic therapy was administered 
in advance of exacerbations.  To date, however, the timing of these exacerbations cannot be 
predicted. 
1.2 
13C-UREA BREATH TEST 
The 13C-urea breath test is based upon a modification of existing breath testing technology that 
detects and monitors a biomarker associated with peptic ulcers.  The mechanism of action for 
detecting urease-containing bacteria in the lungs is the same as for detecting another 
urease-containing bacterium, Helicobacter pylori, in the stomachs of patients with peptic ulcers. 

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For the intended indication, inhaled 13C-urea is catalytically hydrolyzed into 13C-carbon dioxide 
by urease, an enzyme associated with certain bacterial pathogens common to lung infections in 
patients who develop CF (I). 
 
13CO(NH2)2 + 3H2O 
urease
→     13CO2 + 2NH4OH 
(I) 
By measuring the ratio of two stable and naturally occurring isotopes of carbon in exhaled 
carbon dioxide before and after inhalation of 13C-urea, the presence of urease-containing bacteria 
can be confirmed.  In the absence of urease-containing bacteria in the lungs, the isotopic ratio of 
13C to 12C in the baseline and test breath samples will be approximately the same.  If 
urease-containing bacteria are present within the lungs, the concentration of exhaled 13C will be 
increased measurably in the test breath sample relative to baseline.  It is not anticipated that the 
identity or numbers of organisms in the lungs can be deduced from the numerical δ value 
reported by the breath test, but it is anticipated that the magnitude of the numerical δ value will 
reflect the underlying bacterial load.  It is anticipated that urease-producing bacteria can be 
detected within 15 minutes of initiating the breath test. 
1.3 
UREA 
Urea has been approved by the Food and Drug Administration (FDA) for oral administration 
(75 mg when labeled with 13C) in the diagnosis of H. pylori infection in the stomach and for 
intravenous administration in high doses (500 to 1500 mg/kg/day, up to 120 g/day) as an osmotic 
diuretic to treat patients with cerebral edema and glaucoma.  There is a significant amount of 
safety and toxicology data of urea administration via these routes (Crews and Davidson 1961, 
Crews and Davidson 1962, Gisbert and Pajares 2005, Pathak et al 2004, Pathak et al 2010, 
Soupart and Decaux 1996). 
Urea liquefies viscid mucus when injected into the auditory canal (Bauer 1970) and is capable of 
reducing mucus viscosity and increasing mucus expectoration in lung diseases such as chronic 
obstructive pulmonary disease (COPD), bronchiectasis, CF, and asthma when administered by 
inhalation. 
No adverse effects on lung function were reported when 2 to 14 g of urea was administered daily 
by inhalation with an intermittent positive-pressure ventilator over “short intervals” for 14 to 
30 days to 32 patients with chronic bronchitis, cystic fibrosis, asthma, bronchiectasis, and 
emphysema; and no adverse effects were reported when approximately 7.3 g of urea was 
administered by inhalation over 10 minutes to 6 patients with CF using a DeVilbis nebulizer 
(assuming 6 mL 2M urea/min) (Waldron-Edward and Skoryna 1966). 
The mucolytic properties of urea have not been pursued, however, because hypertonic solutions 
were subsequently shown to be a broncho-constrictor in patients with asthma (Cade and Pain 
1972, Pain and Denborough 1967).  In one such study, mean forced expiratory volume in 
1 second (FEV1) decreased 12% ± 20% in 56 adults who inhaled urea aerosolized using a Wright 
nebulizer (Cade and Pain 1972).  It is estimated that approximately 32 mg of urea was inhaled by 
each patient in this study assuming typical values of parameters of operation (output, 

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0.13 mL/min; airflow, 9 L/min) and reported experimental detail (urea concentration, 4M; 
inhalation time, 10 min). 
It is now known that inhalation of osmotic agents such as urea, mannitol, and hypertonic saline 
cause the release of mast cell mediators (histamine and leukotrienes) in patients with asthma, 
resulting in bronchospasm (Anderson 2010, Brannan et al 2003, Brannan et al 2009, Sverrild et 
al 2009).  Hypertonic saline has been used as a test for bronchial hyper-responsiveness in 
patients with asthma, and is widely used, nonetheless, with preventive bronchodilator 
administration to treat hyper-responsive patients with CF. 
Overall, evidence points to the safety of inhaled urea.  As with all inhaled solutions, it is possible 
that it will trigger some cough and bronchospasm; this does not occur in all subjects, however, 
and when it does occur, it is mild and transient.  Further, pretreatment with a bronchodilator 
(albuterol) has been shown to prevent the response in most subjects.  When bronchospasm does 
occur, it also can be rapidly reversed with albuterol therapy. 
1.4 
RATIONALE 
It is anticipated that rapid detection and monitoring of lung pathogens will provide an early 
warning signal of the onset of bacterial load and early confirmation of the therapeutic effect of 
the treatment regimen.  In laboratory experiments, urease-producing bacteria were detected 
within approximately 15 to 20 minutes after exposure to urea. 
1.5 
BENEFIT-TO-RISK ASSESSMENT 
The safety profile of 13C-urea 20 mg and 50 mg administered by inhalation will be assessed in a 
dose escalation study conducted in two parts at a single center located at 
University of New Mexico Hospital 
Pediatric/Adult Pulmonary Clinic 
Ambulatory Care Center 
2211 Lomas Blvd NE 
Albuquerque, NM  87106 
This facility has the required equipment/protocol on site to respond to an emergent adverse effect 
of treatment.  Co-investigators Dr. Harkins or Davies will be available on site should an 
emergent adverse event (such as severe bronchospasm or cardiopulmonary arrest) occur during 
study administration. 
This is the first study in humans of this 13C-urea breath test.  In part 1, a conventional 
dose-escalation study design will assess the safety of inhaled 13C-urea in healthy volunteers.  The 
maximum dose administered will not exceed 50 mg, which approximates the lowest estimated 
dose (32 mg) studied in humans (Cade and Pain 1972).  In part 2, the same dose-escalation study 
design will assess the safety of inhaled 13C-urea in subjects with CF. 

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Thus, the risk to subjects enrolled in this study is small; there is no direct benefit to subjects who 
enrolled.  The potential benefit of an early warning of accelerated colonization of bacteria that 
compromises or destroys lung function that may extend or save lives is significant. 
2. 
OBJECTIVES 
2.1 
PRIMARY OBJECTIVE 
The primary objective is to assess the safety of 13C-urea administered to healthy volunteers and 
subjects with CF by inhalation. 
2.2 
SECONDARY OBJECTIVES 
The secondary objective is to assess the kinetics of 13C-carbon dioxide production by measuring 
the isotopic ratio of 13C to 12C in exhaled carbon dioxide up to 15 minutes after administration. 
3. 
INVESTIGATIONAL PLAN 
3.1 
OVERALL DESCRIPTION OF STUDY DESIGN AND PLAN 
3.1.1 
Primary Endpoint 
The primary endpoint is the safety of inhaled 13C-urea in healthy volunteers and in subjects with 
CF assessed by adverse events, spirometry test results (FEV1 and forced vital capacity [FVC]), 
and respiratory examinations (respiratory rate, auscultation [presence of wheezing and crackles], 
pulse oximetry, and related symptoms, such as cough). 
3.1.2 
Secondary Endpoint 
The secondary endpoint is the isotopic ratio of 13C to 12C in exhaled carbon dioxide of healthy 
volunteers and in subjects with CF measured with the POCone detector (Meretek Diagnostics 
Group of Otsuka America Pharmaceutical, Rockville, MD) before study drug administration and 
at 5, 10, and 15 minutes after study drug administration. 
3.1.3 
Study Design 
This is a single-center, single-administration, dose-escalation study designed to determine the 
safety and dose response of aerosolized 13C-urea 20 mg and 50 mg in detecting urease-producing 
bacteria in the lungs of healthy volunteers (those without and those diagnosed with H. pylori 
infection) and in subjects with CF.  Subjects will be contacted by telephone the next day and 
asked a non-leading question about adverse events. 
The dose will not be escalated until at least 24 hours after all subjects in the cohort have been 
tested at the current dose.  If a subject has an adverse event or new symptom that remains 
unresolved at 48 hours, no new subjects will be enrolled until the relationship between the AE 
and symptom can be evaluated.  If any subject experiences an adverse event considered by the 
data safety monitor to be dose limiting, then three additional subjects will be enrolled at that 

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current dose.  Dose escalation will stop as described in Table 1 if more than one additional 
subject experiences dose-limiting toxicity (DLT) (Section 3.1.3.1).  
After the safety of the maximum dose in the cohort of healthy volunteers without H. pylori 
infection has been reviewed by a data safety monitor, healthy volunteers diagnosed with H. 
pylori infection and subjects with CF will be enrolled.  Three subjects diagnosed with H. pylori 
infection will be tested at the maximum dose to assess potential interference of urease producing 
bacteria in the stomach; subjects infected with H. pylori can be enrolled at any time after the 
safety of the maximum dose in healthy volunteers without H. pylori infection has been evaluated, 
but they will meet the entrance criteria for healthy volunteers without H pylori infection.  
Subjects will be discharged from the study site after 30 minutes or thereafter when oxygen 
saturation levels and FEV1 values return to baseline and coughing symptoms are resolved.  
Supportive treatment for symptom resolution, if needed, will be recorded on the case report form 
(CRF). 
Table 1 
Dose Escalation Rules 
Condition within 24 hours after 
administration of study drug 
Number of DLTs/ 
Subjects in Cohort 
Action 
If no subject experiences DLT 
0/3 
Enroll three subjects at the 
50 mg dose level 
If one subject experiences DLT 
1/3 
Enroll three additional subjects 
at the 20 mg dose 
 If no additional subject experiences DLT 
1/6 
Enroll three subjects at the 
50 mg dose level 
 If more than one subject experiences 
DLT 
>1/6 
Stop enrollment 
Subjects will be receive 13C-urea 20 mg and 50 mg unless enrollment stops because of DLT. 
DLT  Dose-limiting toxicity. 
 
Sputum will be collected from each subject with CF before administering study drug in order to 
identify bacterial flora using two standard microbiologic methods. The first is a quantitative 
method in which colonies are stained and counted, the second is a colorimetric method in which 
a sample is plated in a medium that changes to a blue color as its pH is increased by the 
production of ammonia, a by-product of the urease-catalyzed degradation of urea. 
The pharmacokinetics of the production of 13C-carbon dioxide will be characterized by 
determining the isotopic ratio of 13C to 12C in exhaled carbon dioxide of subjects in each part 
before study drug administration and at 5, 10, and 15 minutes. 

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3.1.3.1 
Dose-limiting Toxicity 
The following significant respiratory events will be considered DLTs (Rosenfeld et al 2011): 
• 
an absolute decrease from baseline FEV1 of 20% of baseline or greater 
• 
an absolute decrease from baseline oxygen saturation of at least 10% 
• 
an increase from baseline respiratory rate of 10 breaths per minute for more than 
2 minutes during quiet breathing 
• 
new wheezes or rales on examination. 
In addition to these events that are mandated DLT, the data safety monitor may determine any 
serious adverse event attributed to study drug to be dose limiting. 
3.1.3.2 
Maximum Tolerated Dose 
The maximum tolerated dose is either the dose level administered before enrollment was stopped 
because of DLT or a lower dose if, in the data safety monitor’s opinion, significant evidence 
exists to reasonably predict an adverse safety profile at that dose for the study drug. 
3.2 
DISCUSSION OF STUDY DESIGN 
Aerosolized urea has been studied in doses ranging from approximately 32 mg (Cade and Pain 
1972) to approximately over 7000 mg (Waldron-Edward and Skoryna 1966).  The present study 
investigates doses up to 50 mg, which, based upon laboratory studies discussed below, is 
anticipated to evidence early indication of changes in the number of urease-containing bacteria in 
the lungs.   
Subjects with CF are selected because they have large amounts of urease-producing bacteria in 
their lungs, are not acutely ill, and are familiar with procedures used in this study.  Subjects with 
CF who are known to be colonized with Pseudomonas will be selected because pseudomonas has 
well documented urease activity.  It is likely that subjects with CF will have additional lung 
pathogens that may also contribute to the signal; the breath test is intended to be specific for 
urease-producing bacteria.  Pathogens in sputum cultures from subjects with CF will also be 
identified and tested for urease activity. 
Healthy volunteers diagnosed with H. pylori infection are enrolled to determine whether a 
urease-producing pathogen in the stomach can produce 13C-carbon dioxide in exhaled breath 
within 15 minutes after administering study drug by inhalation. 
The dose-escalation model minimizes subject risk by incrementally increasing the dose of study 
drug administered to healthy volunteers, beginning at a dose that approximates 0.1% of one that 
produced no adverse effects in patients with CF and ending with a dose that approximates the 
minimum amount administered to patients with asthma. 
A maximum dose of 50 mg of 13C-urea was selected because it can be reconstituted to provide an 
isotonic solution in a volume that can be inhaled in a sufficiently short time window using a 
standard nebulizer.  The urease reaction is rapid, and the direct delivery of 13C-urea to the lung 

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combined with immediate release of 13C-carbon  dioxide enables the production of a signal.  
Because a 75 mg oral dose of 13C-urea is sufficient to reliably produce a measurable amount of 
exhaled 13C-carbon dioxide in patients infected with stomach H. pylori, which requires the 
carbon dioxide produced by the catalyzed hydrolysis of urea in the stomach to be absorbed into 
the blood stream and transported to the lung, it is likely that a 50 mg dose administered to the 
lung directly will produce a reliably measurable signal.  The suitability of a lower dose, 20 mg 
dose, 13C-urea will also be tested. 
In addition to mandatory rules that stop dose escalation in the event of DLT, dose escalation may 
also be stopped at the discretion of the data safety monitor. 
Albuterol is standard therapy for relief of bronchospasm (Eng et al 1996) and also for the 
prevention of bronchospasm in diseases such as exercise-induced asthma (Raissy et al 2008).  
Patients with CF are frequently pretreated with albuterol if a therapy may produce 
bronchospasm, e.g., inhalation of 7% saline.  Although nebulized isotonic solutions are not 
expected to produce bronchospasm, subjects with CF will be pretreated with albuterol because of 
the potential of mild bronchospasm induced from irritation of lung pathogens at the time of study 
drug administration. 
The characterization of the pharmacokinetics of 13C-carbon dioxide will inform the useful time 
interval during which the isotopic ratio of 13C to 12C can be measured after administration of 
study drug in patients with lung infections (Otsuka America Pharmaceutical, Rockville, 
Maryland).  Multiple breath specimens will be obtained at 15 minutes to estimate intra-patient 
variability. 
3.2.1 
Number of Subjects 
Approximately 15 subjects will be enrolled, 9 healthy volunteers and 6 subjects with CF.  Three 
healthy volunteers without H. pylori infection will each receive a single 20 mg dose and 3 will 
each receive a single 50 mg dose.  After the safety of the 50 mg dose has been evaluated, 
3 subjects with CF will each receive a single 20 mg dose and 3 subjects with CF will each 
receive a single 50 mg dose.  Three subjects diagnosed with H. pylori infection will also each 
receive a 50 mg dose to assess potential interference of urease-producing bacteria in the stomach.  
Three additional subjects will be added to every dose level at which a dose-limiting adverse 
event occurs.  If one subject at each dose level experiences DLT, then a maximum of 30 subjects 
will enroll. 
3.2.2 
Duration of Study 
Subjects will participate in the study for approximately 9 days, up to 7 days between the 
screening visit and the study visit plus a follow-up visit 24 hours after study drug administration.  
Subjects will receive a single dose of study drug and remain in the clinic for at 30 minutes for 
evaluation and observation.  Subjects will be discharged from the study site after 30 minutes or 
thereafter when oxygen saturation levels and FEV1 values return to baseline and coughing 
symptoms are resolved.  Twenty-four hours after administration of study drug, subjects will be 
contacted by telephone for a safety evaluation by study personnel.  If an adverse event is 

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reported at the telephone contact, the subject will be asked to return to the clinic for an 
evaluation. 
A 24-hour period will separate dose escalations.  Thus, the study is anticipated to be completed 
within approximately 4 to 8 weeks after the first subject enrolls. 
3.2.3 
Premature Termination of Study 
Dose escalation will be stopped if more than one subject experiences DLT at any dose level.  The 
data safety monitor may terminate the study early by stopping enrollment based upon safety 
concerns. 
4. 
SELECTION OF STUDY POPULATION 
4.1 
INCLUSION CRITERIA 
4.1.1 
Healthy Volunteers Without Helicobacter pylori infection 
Healthy volunteers who meet all the following inclusion criteria are eligible to enroll: 
1. At least 18 years old at the time of providing informed consent in a manner approved by the 
University of New Mexico, Institutional Review Board and willing to comply with the 
requirements of the study 
2. In generally good health as determined by the investigator, with no respiratory illness within 
2 weeks before the screening visit and no pulmonary history 
3. No history of allergy or asthma 
4. Has not smoked tobacco within 6 months before the screening visit and agrees not to smoke 
until after the follow up visit 
5. FEV1 at least 80% at the time of providing informed consent 
4.1.2 
Healthy Volunteers Diagnosed With Helicobacter pylori infection 
Healthy volunteers diagnosed with H. pylori infection who meet all the inclusion criteria for 
healthy volunteers in Section 4.1.1 and additionally who have diagnosed with H. pylori infection 
are eligible to enroll. 
4.1.3 
Inclusion Criteria for Subjects With Cystic Fibrosis 
Subjects who meet all the following inclusion criteria are eligible to enroll. 
1. At least 18 years old at the time of providing informed consent in a manner approved by the 
University of New Mexico, Institutional Review Board and willing to comply with the 
requirements of the study 
2. Diagnosed with CF at least 24 months before the screening visit 
3. Documented presence of Pseudomonas aeruginosa in at least three sputum cultures within 
2 years before the screening visit, one of which within 6 months before the screening visit 
4. Is able to produce sputum at the time of the screening visit 
5. FEV1>60%  or 1.5 L at the time of providing informed consent 

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6. Has not smoked tobacco within 6 months before the screening visit and agrees not to smoke 
until after the follow up visit 
4.2 
EXCLUSION CRITERIA 
Subjects who meet any of the following exclusion criteria are not eligible to enroll. 
1. Positive H. pylori serology, except for subjects in the H. pylori cohort 
2. Subjects intolerant of albuterol 
3. Subjects diagnosed with asthma 
4. Subjects with evidence of upper respiratory tract infection within 14 days before the 
screening visit 
5. Use of inhaled antibiotics within 12 hours before the administration of the breath test on 
Day 1 or required use of inhaled antibiotics during the study 
6. Has smoked tobacco within 6 months before the screening visit 
7. Females who are pregnant or nursing or of child-bearing potential who are not using a 
medically acceptable form of contraception (acceptable forms of contraception are hormonal 
birth control, intrauterine device, or barrier method plus a spermicidal agent) or abstinence 
8. Any condition or history that, in the judgment of the investigator, would compromise the 
ability of the subject to comply with the study protocol or to complete the study 
9. Subjects who have used an investigational agent within 28 days before Day 1 
4.3 
REMOVAL OF SUBJECTS 
Subjects may be removed from the study by the investigator at any time for any of the following 
reasons: 
1. Adverse event 
2. Non-compliance with study protocol 
3. Any condition that, in the judgment of the investigator, would compromise the ability of the 
subject to comply with the study protocol or to complete the study 
In addition, subjects may remove themselves from the study at any time for any reason without 
consequence to further treatment by the investigator. 
All subjects who are removed prematurely from the study should be assessed according to the 
procedures and assessments scheduled for the end-of-study visit in Section 6.3. 
4.4 
REPLACEMENT OF SUBJECTS 
Subjects who are removed from the study prematurely will be replaced in order to maintain the 
required number of subjects in each cohort. 

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5. 
PROCEDURES FOR ADMINISTRATION OF TREATMENTS 
All investigational medications will be prepared and dispensed by the investigational pharmacist 
or delegate at the University of New Mexico Hospital in accordance with procedures in this 
protocol. 
5.1 
TREATMENTS TO BE ADMINISTERED 
5.1.1 
Identification of Investigational Product 
The active pharmaceutical ingredient is lyophilized 13C-urea; the carbon content is enriched in 
13C (at least 99%), a naturally-occurring, non-radioactive isotope of carbon (Table 2). 
Table 2 
Identification of Investigational Product 
 
Study Product 
Active Substance (INN) 
13C-urea 
Trade Name (if applicable) 
Not applicable 
Formulation (including dosage form and strength) 
Lyophilisate to be reconstituted with 3 mL of 
sterile water for injection to form isotonic solutions 
of pH 7.2 containing 20 mg or 50 mg of active 
ingredient 
Route/Mode of Administration  
Inhalation 
Manufacturer 
Coldstream Laboratories, Lexington, Kentucky 
USA 
 
5.1.2 
13C-urea Breath Test Kit 
The 13C-urea breath test kit in this study contains 13C-urea lyophilized in a 10 mL glass vial with 
an aluminum crimp closure, 3 mL of sterile water for injection (WFI), a nebulizer cup (LC Sprint 
Star reusable nebulizer cup, PARI, Midlothian, Virginia, part number 023F35), 12 300-mL mylar 
breath collection bags (Meretek, Rockville, Maryland), and plastic tubing to connect the 
nebulizer cup to the compressor (Vios™ Adult Aerosol Delivery System, PARI, Midlothian, 
Virginia, part number 310F35-LCS).  Additionally, each kit will contain 24 self-adhesive 
bar-code stickers used to identify collection bags and to maintain integrity of data collection.  
The lyophilisate will be reconstituted by transferring WFI using a 3 mL syringe (Becton 
Dickinson, Franklin Lakes, New Jersey, part number 309595).  All components will be supplied 
sterile; all are intended for single use. 
5.1.3 
Administration of Study Drug 
The lyophilisate will be reconstituted in WFI (3 mL) supplied with the breath test kit and 
administered by inhalation.).  If all the reconstituted study drug does not go into solution, or if 

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the reconstituted solution is cloudy or contains particulate material, the vial will be discarded.  
The nebulizer cup is connected to the compressor with the supplied tubing, and the entire clear, 
reconstituted solution of 13C-urea is transferred into the nebulizer cup  After turning on the 
compressor, the subject will inhale the study drug by breathing through the mouthpiece of the 
nebulizer cup until all the solution has been aerosolized and inhaled (Appendix A). 
5.1.4 
Pre-treatment 
Subjects with CF will receive two puffs of albuterol (90 μg albuterol base per puff) from a 
standard holding chamber (Aerochamber, Lupin Pharmaceuticals, Baltimore, Maryland) 
10 minutes before the administration of study drug (Table 3). 
Table 3 
Identification of Pre-treatment 
 
Study Product 
Active Substance (INN) 
Albuterol 
Trade Name (if applicable) 
Ventolin HFA 
Formulation (including dosage form and strength) 
Aerosol (108 μg albuterol sulfate, 90 μg albuterol 
base from mouthpiece per actuation) 
Route/Mode of Administration  
Inhalation 
Source 
GlaxoSmithKline 
 
5.1.5 
Rescue Medication 
In the event of wheezing or bronchospasm following administration of study drug, albuterol may 
be administered as rescue medication. 
5.1.6 
Methods of Assigning Subjects to Treatment Group 
Subjects without H. pylori infection will be assigned to a single, open-label treatment, beginning 
with the lower dose of study drug, in the order in which they enter the study according to the 
dose escalation rules in Table 1 (Section 3.1.3).  Healthy volunteers diagnosed with H. pylori 
infection will received the MTD. 
5.1.7 
Selection of Doses in the Study 
Enrolled subjects will receive single doses of 13C-urea 20 mg or 50 mg. 
5.1.8 
Selection and Timing of Doses for Each Subject 
Subjects will receive single doses of 13C-urea in increasing amounts, 20 mg in the first cohort 
and 50 mg in the second cohort as detailed in Table 1.  Study drug may be administered in the 
fasted or as-fed state, but subjects should not ingest food until after the last breath sample is 

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collected; subjects may receive water ad libitum.  Subjects may be administered study drug at 
any time of day with the provision they are required to be available 24 hours after the study drug 
is administered for a follow-up evaluation. 
5.1.9 
Blinding 
This is an open-label study. 
5.1.10 
Packaging and Labeling 
Lyophilized 13C-urea 20 mg and 50 mg will be supplied in 10 mL stoppered vials sealed with 
aluminum closures and labeled for investigational use only.  Study drug will be reconstituted in 
WFI (3 mL) supplied with the breath test kit and administered by inhalation (see Table 2 for the 
identification of the investigational product). 
5.1.11 
Storage 
13C-urea should be stored at ambient conditions (from 15º to 30º C [59º to 86º F]) in an area 
restricted to investigational staff or the pharmacy at the study center. 
5.1.12 
Accountability and Destruction of Study Drug 
All investigational medications will be signed for by the investigational pharmacist at the 
University of New Mexico (or designee) when they are received.  The study drug must be 
handled and stored as described and dispensed only to those subjects formally entered into the 
study. 
At the completion of the study, and after reconciliation of all delivery and usage records, any 
unused study drug will be destroyed. 
5.2 
CONCOMITANT THERAPY 
With the exception of inhaled antibiotics, subjects are allowed concomitant medications for 
pre-existing conditions as deemed necessary by the investigator from the time of study drug 
inhalation until the last breath sample is collected.  Drugs taken from the beginning of inhalation 
of study drug until the follow-up visit will be recorded on the CRF.  The generic name of the 
drug, the dose, and the time and reason for administration will be recorded on the CRF. 
Inhaled antibiotics are disallowed from the beginning of the inhalation of study medication until 
the collection of the final breath sample at 2 hours. 
5.3 
TREATMENT COMPLIANCE 
As all subjects will receive study drug in the clinic under supervision of study personnel, each 
subject will be considered treatment compliant who inhales the full volume of reconstituted 
study drug. 

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6. 
VISIT SCHEDULE 
The schedule of assessments and procedures are summarized in Table 4.

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Table 4 
Schedule of Procedures and Assessments for Dose-ranging Study 
Assessment/Procedure 
 
Screening 
(Day –7 to Day –1) 
Day 1 
Follow-upa 
Informed consent 
X 
 
 
Inclusion/exclusion criteria 
X 
X 
 
Medical history 
X 
 
 
Concomitant medications, including all medications taken within the last 48 hours 
X 
X 
X 
Twelve-lead electrocardiogram 
X 
 
 
Helicobacter pylori serology 
X 
 
 
Urine pregnancy test (child-bearing females) 
 
X 
 
Five-minute vital signs measurementsc 
 
X 
 
Respiratory examination (respiration rate, auscultation, pulse oximetry, and related 
symptoms, such as cough) 
 
X 
 
Perform spirometry 
Xb 
X 
 
Administer albuterol (subjects with cystic fibrosis only) 
 
X 
 
Perform spirometryd 
 
X 
 
Obtain sputum specimen 
 
Xe 
 
Perform breath testf 
 
X 
 
Assess adverse events 
 
X 
Xg 
a Subjects will be contacted by telephone 24 hours after administration of study drug for a follow-up assessment. 
b Spirometry does not need to be performed at the screening visit if performed within the past 14 days. 
c Heart rate, systolic and diastolic blood pressure, and body temperature will be recorded before study drug administration to assist in the determination that enrolled subjects 
are in generally good health. 
d Spirometry will be performed before and at 10 and 30 min after the end of study drug administration.  In addition, only for subjects with cystic fibrosis, spirometry will be 
performed before and 10 ± 2 minutes after pre-treatment with albuterol. 
e Sputum specimens will be collected immediately after performing baseline spirometry only from subjects with cystic fibrosis.  Sputum specimens will be cultured to 
determine the identity and number of pathogens, and will also be qualitatively analyzed nonselectively for the presence of urease-producing bacteria using a colorimetric 
assay. 
f Study drug will be administered immediately following spirometry, or for subjects with cystic fibrosis, the collection of sputum.  Five baseline exhaled breath specimens 
will be obtained immediately before administration of 13C-urea.    Exhaled breath specimens will be collected at 5 ± 1, 10 ± 2, 15 ± 2 (in triplicate at this timepoint only) 
after administration of 13C-urea and sent within 24 h after collection to the central laboratory for analysis. 
g Subjects who report adverse events at the follow-up assessment will be asked to return to the clinic for evaluation.

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6.1 
SCREENING (DAY –7 TO DAY –1) 
The following procedures and assessments will be performed at the screening visit (Day –7 to 
Day –1). 
1. Acquire written informed consent. 
2. Review inclusion and exclusion criteria. 
3. Document medical history, including all medications taken within 48 hours before the 
administration of study drug. 
4. Obtain a 12-lead electrocardiogram (ECG). 
5. Collect a blood specimen (5 mL) to assess H. pylori infection.  
6. Perform spirometry, if not performed within the previous 14 days (Section 7.2.1). 
6.2 
ACTIVE TREATMENT (DAY 1) 
1. Review inclusion and exclusion criteria. 
2. Document all medications taken within the last 48 hours. 
3. Perform a urine pregnancy test on women of child-bearing potential. 
4. Record 5-minute sitting vital signs measurements (heart rate, systolic and diastolic blood 
pressure, and body temperature). 
5. Perform respiratory examination (respiration rate, auscultation [presence of wheezing and 
crackles], pulse oximetry, and related symptoms, such as cough) (Section 7.2.1). 
6. Perform spirometry (Section 7.2.1). 
7. Administer albuterol, only to subjects with CF (Section 5.1.4). 
8. Perform baseline spirometry 10 ± 2 minutes after administering albuterol to subjects with CF 
only (Section 7.2.1). 
9. For subjects with CF only, collect sputum specimen. 
10. Collect five breath specimens.  Record the clock time at which the sample was collected on 
two self-adhesive stickers.  Place one sticker on the collection bag and the other on the CRF 
(Section 7.1.2.1). 
11. Administer study drug by inhalation (Section 5.1.3).  Record the clock time at the beginning 
and end of study drug administration. 
12. Measuring time from the end of the inhalation of study drug, collect breath specimens at 
5 ± 1, 10 ± 2, 15 ± 2 minutes (three specimens at this time point only) after the inhalation of 
study drug is completed.  Record the clock time of each breath collection on two 
self-adhesive stickers.  Place one sticker on the collection bag and the other on the CRF 
(Section 7.1.2.1). 
13. Perform spirometry after collecting the breath specimen at 10 and 30 minutes after the end of 
study drug administration (Section 7.2.1). 
14. Assess adverse events (Section 8). 
6.3 
FOLLOW-UP 
Twenty-four hours after administration of study drug, subjects will be contacted by telephone 
and asked a non-leading question to assess adverse events such as “How do you feel after 
participating in the study?”  Additionally, subjects will be asked if they have taken any 

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medications since receiving study drug.  All medications taken from the time of study drug 
administration until the follow-up assessment will be recorded on the CRF.  Subjects who have 
adverse events at the follow-up visit will be asked to return to the clinic for evaluation. 
6.4 
DISCONTINUATION 
Subjects who discontinue from the study prematurely will receive a follow-up assessment 
(Section 6.3). 
7. 
STUDY VARIABLES AND METHODS OF ASSESSMENT 
7.1 
EFFICACY VARIABLE 
7.1.1 
Overview of Variable 
The 13C-urea breath test will be administered to all subjects after performing baseline spirometry. 
7.1.2 
Methods of Assessment 
7.1.2.1 
Collection of Breath Specimen 
Samples of exhaled breath will be collected immediately before and at 5 ± 1, 10 ± 2, 15 ± 2 
minutes after the inhalation of study drug is completed.  Five breath specimens will be obtained 
before the inhalation of study drug; one specimen will be obtained at each subsequent assessment 
except for the 15-minute assessment at which time three exhaled breath specimens will be 
obtained.  Step-wise instructions for performing the breath test are provided in Appendix A. 
1. Record the clock time, intended sample time (e.g., baseline, 5-minute), and subject 
demographic information on two self-adhesive stickers in the 13C-urea breath test kit.  Place 
one of the stickers on the breath collection bag and the other sticker on the CRF. 
2. Remove the pull-off cap from the mouthpiece of the breath collection bag. 
3. Instruct the subject to breath normally, then take a deep breath and pause momentarily before 
exhaling into the mouthpiece of the bag. 
4. Place the cap on the mouthpiece of the bag until it clicks into place. 
5. Store the breath specimen at 15º to 30º C (59º to 86º F) until analyzed. 
7.1.2.2 
Analysis of Breath Specimen 
After collection, all breath test specimens will be picked up within 24 hours by the central 
laboratory to be analyzed within 7 days of collection.  The isotopic ratio of 13C to 12C is 
calculated from breath samples collected before and after administration of 13C-urea using a 
POCone® Infrared Spectrophotometer (Otsuka America Pharmaceutical, Rockville, Maryland).  
The relative increase in isotopic ratio, δ, is determined automatically by the spectrophotometer 
(I). 

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1000
R
R
R
δ
std
std
x







−
=
, 
(I) 
where Rx is the ratio of the abundance of 13C to 12C in the post-administration sample and Rstd is 
the corresponding ratio at baseline. 
7.1.3 
Microbial Analysis of Sputum 
7.1.3.1 
Overview of Variables 
Sputum specimens will be obtained from subjects with CF after the performance of baseline 
spirometry and before administration of study drug.  Specimens will be obtained using standard 
practices at the study site, and in all cases will be expectorated into a sterile sputum cup, labeled 
with the time of day, date, and unique study identifier for that patient. 
7.1.3.2 
 Methods of Assessment 
The presence of urease-containing bacteria will be determined locally using a colorimetric 
method in which a sample is plated in a medium that changes to a blue color as its pH is 
increased by the production of ammonia, a by-product of the urease-catalyzed degradation of 
urea (Cox et al 1977, Damato et al 1982, Isenberg 2004). 
Additionally, bacterial flora in collected sputum will be assessed using standard quantitative 
microbiologic methods employed at the study site in which colony forming units (CFU) are 
identified and counted, and in all cases organisms in numbers greater than 104 and appearing in 
more than one CFU will be identified and counted. 
7.2 
SAFETY VARIABLES 
The safety of 13C-urea will be assessed from adverse events (Section 8), spirometry test results 
(FEV1 and FVC), and respiratory examinations (respiratory rate, auscultation [presence of 
wheezing and crackles], pulse oximetry, and related symptoms, such as cough). 
7.2.1 
Spirometry 
7.2.1.1 
Overview of Variables 
Spirometry will be performed at the initial respiratory examination (baseline, Day 1) and after 
collecting breath specimens at 10 and 30 minutes.  If the last FEV1 measurement is less than 10% 
of baseline, 4 puffs of albuterol will be administered and spirometry will be repeated in 15 
minutes. Patients will be discharged from the clinic when FEV1 is within 10% of baseline 
measurements and patients are symptom free.  For subjects with CF, spirometry will be 
performed before and 10 minutes after administration of albuterol on Day 1.  

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7.2.1.2 
Methods of Assessment 
Spirometry will be performed using standard techniques specified by the American Thoracic 
Society and the European Respiratory Society (Miller et al 2005). 
7.2.2 
Respiratory Examination 
7.2.2.1 
Overview of Variables 
Respiratory rate, pulse oximetry data, and related symptoms, such as cough, will be recorded 
before performing each spirometry (at baseline and after collecting the breath specimen at 10 and 
30 minutes).  Auscultation, noting the presence of wheezing and crackles, will be performed at 
baseline and at any assessment at which FEV1 decreases at least 20% from baseline. 
7.2.2.2 
Methods of Assessment 
Respiratory rate will be determined by counting the number of breaths in one minute.  Arterial 
blood saturation will be determined by oximetry; the same location will be used for each 
individual subject.  The number of coughs in the 90-second interval following the end of the 
administration of study drug will be recorded in the CRF (Koskela et al 2005).  Wheezing and 
crackles will be noted when auscultation is performed. 
7.3 
PHARMACOKINETICS 
The pharmacokinetics of 13C-carbon dioxide production will be determined from isotopic ratios 
of 13C to 12C as described in Section 7.1.2.2. 
7.4 
APPROPRIATENESS OF MEASUREMENTS 
Data will be collected using standard methods, widely used and generally regarded as reliable, 
accurate, and relevant. 
8. 
ADVERSE EVENTS, SERIOUS ADVERSE EVENTS, AND 
SUSPECTED ADVERSE REACTIONS 
The investigator is responsible for the detection and documentation of events meeting the criteria 
and definition of an adverse event (AE) or serious adverse event (SAE) as provided in this 
protocol.  During the study, when there is a safety evaluation, the investigator or site staff will be 
responsible for detecting AEs and SAEs as detailed in this section of the protocol.  All AEs will 
be reviewed by a data safety monitor. 
8.1 
DEFINITION OF AN ADVERSE EVENT 
An AE is defined as any untoward medical occurrence associated with the use of a drug in 
humans, whether or not considered drug related. 

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An AE may be any unfavorable and unintended sign (including an abnormal laboratory finding), 
symptom, or disease temporally associated with the use of a study medication, whether or not 
considered causally associated with the use of the study medication.  Any abnormal laboratory 
value deemed clinically significant by the investigator, regardless of causal relationship, must be 
reported as an AE. 
Examples of an AE include the following: 
• 
significant or unexpected worsening or exacerbation of the condition or indication 
under study 
• 
exacerbation of a chronic or intermittent preexisting condition, including either an 
increase in frequency or intensity of the condition (e.g., abnormal physical 
examination finding) 
• 
signs, symptoms, or clinical sequelae of a suspected interaction 
• 
signs, symptoms, or clinical sequelae of a suspected overdose of the study 
medication or a concurrent medication (overdose per se should not be reported as an 
AE or SAE, unless nonserious or serious sequelae occur) 
The following examples are not considered AEs: 
• 
medical or surgical procedure (e.g., endoscopy, appendectomy), although the 
condition that leads to the procedure is an AE 
• 
anticipated day to day fluctuations of preexisting disease(s) or condition(s) 
(including laboratory values) present or detected at the start of the study that do not 
worsen 
• 
the disease or disorder being studied, or expected progression, signs, or symptoms of 
the disease or disorder being studied, unless they become more severe or occur with 
a greater frequency than expected for the subject’s condition 
All AEs, whether volunteered, elicited, or noted on physical examination, and regardless of 
causality or seriousness, will be assessed and recorded on the CRF beginning after administration 
of study medication through the final follow-up assessment.  SAEs will be assessed and recorded 
after administration of study medication through 30 days after study drug administration 
(Sections 8.2 and 8.4). 
8.2 
DEFINITION OF A SUSPECTED ADVERSE REACTION 
A suspected adverse reaction (SAR) is defined as any adverse event for which there is a 
reasonable possibility that the adverse event was caused by the study drug.  ‘Reasonable 
possibility’ means there is evidence to suggest a causal relationship between the drug and the 
adverse event.  A suspected adverse reaction implies a lesser degree of certainty about causality 
than adverse reaction, which means any adverse event caused by a drug. 

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8.3 
DEFINITION OF A SERIOUS ADVERSE EVENT OR SUSPECTED 
ADVERSE REACTION 
An SAE or serious suspected adverse reaction (SSAR) is defined as any event that meets the 
following criteria: 
• 
It results in death or is life-threatening (i.e., presents an immediate risk of death from 
the event as it occurred).  (This criterion is not intended to include an AE that, had it 
occurred in a more severe form, might have caused death.) 
• 
It results in persistent or substantial disability or incapacitation.  (This criterion is not 
intended to include experiences of relatively minor medical significance, such as 
uncomplicated headache, diarrhea, or sprained ankle.) 
• 
It results in hospitalization. 
• 
It results in prolongation of an existing hospitalization. 
• 
It is a congenital anomaly or birth defect. 
• 
It requires medical or surgical intervention to prevent any of the above outcomes. 
Medical and scientific judgment should be exercised in determining whether an AE is serious 
when considering important medical events that may not be immediately life-threatening or 
result in death or hospitalization, but may jeopardize the subject or may require intervention to 
prevent any of the other outcomes listed.  Examples of such medical events that may also be 
considered serious include, but are not limited to, intensive treatment in an emergency room or at 
home for allergic bronchospasm, blood dyscrasias, or convulsions that do not result in 
hospitalization. 
Hospitalization for elective treatment of a preexisting condition that did not worsen from 
baseline does not meet the definition of an SAE. 
Social or convenience admission to a hospital or prolongation of a hospitalization for social or 
convenience reasons not associated with the occurrence of an AE does not meet the definition of 
an SAE. 
8.3.1 
Serious Adverse Events That Occur Before Administration of Study Medication 
Before administration of study medication, only SAEs assessed by the investigator as related to 
study participation (e.g., related to study procedures or a change in existing therapy) will be 
transcribed onto the SAE reporting form and reported to the data safety monitor. 
8.3.2 
Serious Adverse Events That Occur After Last Administration of Study Drug 
If an investigator becomes aware of an SAE or death occurring within 30 days after a subject 
receives the last administration of study medication, the investigator should report this to the data 
safety monitor. 
If an investigator becomes aware of an SAE or death occurring more than 30 days after a subject 
receives study medication and the investigator considers the event to be related to the study 
medication, the investigator is obligated to report the SAE to the data safety monitor. 

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8.4 
RECORDING AND EVALUATING ADVERSE EVENTS AND SERIOUS 
ADVERSE EVENTS 
The investigator will attempt to establish a diagnosis of the event on the basis of signs, 
symptoms, or other clinical information.  In such cases, the diagnosis, not the individual signs or 
symptoms, should be documented as the AE or SAE. 
8.4.1 
Assessment of Intensity 
The investigator will assess the intensity of each AE and SAE reported during the study.  The 
intensity of each AE and SAE recorded on the CRF should be assigned to one of the following 
categories: 
• 
mild:  an event that is easily tolerated by the subject, causes minimal discomfort, and 
does not interfere with everyday activities 
• 
moderate:  an event that is sufficiently discomforting to interfere with normal 
everyday activities 
• 
severe:  an event that prevents normal everyday activities 
An AE that is assessed as severe should not be confused with an SAE.  Severity is a term used to 
describe the intensity of a specific event, and both AEs and SAEs can be assessed as severe.  The 
event itself, however, may be of relatively minor medical significance (such as a severe 
headache).  This is not the same as serious, which is based on the subject’s or event’s outcome or 
on action criteria usually associated with events that pose a threat to a subject’s life or 
functioning (Section 8.2). 
8.4.2 
Assessment of Causality 
The investigator is obligated to use his or her clinical judgment to assess the relationship 
between the study medication and the occurrence of each AE or SAE.  The investigator will 
assess the relationship to the study medication by using the following criteria: 
• 
Definitely Related:  An AE has a strong temporal relationship to the study drug.  The 
AE is most likely explained by study drug.  Dechallenge and rechallenge (if possible) 
are positive.  The AE is consistent with a known response to the study drug.  Another 
etiology is unlikely or significantly less likely. 
• 
Probably Related:  An AE has a strong temporal relationship to the study drug.  The 
AE is more likely explained by study drug than by another cause.  Dechallenge (if 
performed) is positive. 
• 
Possibly Related:  An AE has a reasonable temporal relationship to study drug.  The 
AE could have been due to another equally likely cause.  Dechallenge is positive. 
• 
Not Related:  The subject did not receive the study drug OR the AE has no temporal 
relationship to study drug OR the AE has a much more likely alternate etiology OR 
the AE is due to an underlying or concurrent illness or effect of another drug. 
Even in situations in which minimal information is available for the initial SAE report, it is 
important that the investigator always make an assessment of causality for every event before 

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transmitting the SAE reporting form and AE CRF page(s) to the data safety monitor.  The 
causality assessment is one of the criteria used when determining regulatory reporting 
requirements.  The investigator may change his or her opinion of causality in light of follow-up 
information and amend the SAE reporting form and AE CRF page(s) accordingly. 
8.4.3 
Assessment of Outcome 
The investigator will assess the outcome of the event by using the following terms: 
• 
Resolved:  The event resolved or the subject recovered without sequelae.  An event 
(either serious or nonserious) occurred and had an endpoint, and the subject 
experienced no restrictions.  For example, if an AE led to surgery that resulted in a 
postoperative wound infection, the infection would not be considered a sequela. 
• 
Resolved with sequelae:  The event has at least one secondary outcome that may 
result in permanent disability, functional limitation, or both.  Such sequelae are 
usually limited to SAEs.  For example, if an AE of stroke resulted in paralysis, or 
emboli formation after an AE of bacterial infection resulted in a renal infarct and loss 
of renal function, then paralysis and loss of renal function would be considered 
sequelae. 
• 
Not resolved:  At the end of the study, a nonserious event either has not changed in 
intensity or may not have recovered to baseline values.  Examples include headache, 
low-grade fever, or nausea. 
• 
Unknown: The status of the event is unknown. 
• 
Death 
8.5 
DATA SAFETY MONITOR 
Clinical data obtained from each subject will be presented daily to the data safety monitor, who 
will adjudicate AEs and determine when dose levels may be increased and when subjects may be 
enrolled into the study.  All the adverse events will be reported to IRB as soon as possible but no 
later than 5 days from the event date or the reported date by the patients or as specified in 
Section 8.7, whichever is earlier. 
8.6 
FOLLOW-UP OF ADVERSE EVENTS AND SERIOUS ADVERSE EVENTS 
After the occurrence of an AE or SAE, the investigator is required to follow each subject 
proactively and provide further information on the subject’s condition.  All AEs and SAEs 
documented at a previous visit or contact are designated as ongoing and will be reviewed at 
subsequent visits or contacts. 
Nonserious AEs will be followed after the last scheduled study visit, until an appropriate 
resolution can be documented. 
SAEs will be followed until the event resolves, the condition stabilizes, the event is otherwise 
explained, or the subject is lost to follow-up.  The investigator will ensure that follow-up 
information provided to the data safety monitor includes results of any additional laboratory tests 

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or investigations, histopathologic examinations, or consultations with other healthcare 
professionals that serve to clarify the nature of the event, the cause of the event, or both. 
For SAEs only, new or updated information will be recorded on the originally completed SAE 
reporting form and CRF pages, with all changes signed and dated by the investigator.  The 
updated SAE reporting form and CRF pages should be resubmitted to the data safety monitor 
within the time frames outlined in Section 8.7. 
8.7 
PROMPT REPORTING OF SERIOUS ADVERSE EVENTS TO THE DATA 
SAFETY MONITOR 
Once the investigator determines that an event meets the protocol definition of an SAE, the 
investigator must notify the data safety monitor within 24 hours. 
Complete the SAE details reporting form and forward by e-mail to the following contact: 
Mark Schuyler, MD 
mschulyer@salud.unm.edu 
In the initial e-mail, the investigator must provide to the data safety monitor the following CRF 
pages, completed to the greatest extent possible: 
• 
AE record 
• 
medical history 
• 
prior and concomitant medications 
Also, the following documents are to be forwarded:  any laboratory results, diagnostic test 
results, and medical reports relevant to the SAE. 
E-mail transmission is the preferred method to transmit SAE information.  In rare circumstances 
and in the absence of e-mail capacity, notification by fax (505-272-8700) or telephone (505-272-
4751) is acceptable, with a copy of the SAE reporting form and CRF pages sent by overnight 
mail to: 
Mark Schuyler, MD 
Data Safety Monitor  
Professor of Medicine 
The University of New Mexico 
Department of Internal Medicine, Pulmonary Division, MSC10-5590 
1 University of New Mexico 
Albuquerque, NM  87131 
Initial notification via telephone does not replace the need for the investigator to complete the 
SAE reporting form and CRF pages within the time frames outlined. 
The investigator must not wait to have all information regarding an SAE before notifying the 
data safety monitor of the event.  The SAE details reporting form must be updated when 

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additional information is received.  Follow-up information received on all SAEs must be 
forwarded to the data safety monitor using the same procedure and in the same timeframe as for 
an initial report. 
8.8 
REGULATORY REPORTING REQUIREMENTS 
The investigator must promptly report all SAEs to the data safety monitor in accordance with the 
procedures detailed in Section 8.7, “Prompt Reporting of Serious Adverse Events to the Data 
safety monitor.”  The data safety monitor has a legal responsibility to notify, as appropriate, both 
the local regulatory authority and other regulatory agencies about the safety of a product under 
clinical investigation. 
Prompt notification of SAEs by the investigator to the appropriate project contact for SAE 
receipt is essential so that SSARs that are either unexpected or observed with increasing 
occurrence, be reported and legal obligations and ethical responsibilities regarding the safety of 
other subjects are met. 
The investigator, or responsible person according to local requirements, must comply with 
requirements related to the reporting of SAEs to the Institutional Review Board (IRB) or 
Independent Ethics Committee (IEC). 
8.9 
PRECAUTIONS 
Any subject who becomes pregnant during the study must be discontinued immediately, but 
should be followed through delivery or termination of the pregnancy.  A subject should also 
notify the investigator if she becomes pregnant within 30 days after receiving study medication.  
The data safety monitor must be notified of all pregnancies reported to the investigator (see 
Section 8.7 for contact information). 
9. 
STATISTICS 
9.1 
DETERMINATION OF SAMPLE SIZE 
The sample size is not based upon inferential analyses, but is deemed appropriate based on 
standard designs of Phase 1 dose-escalation studies in which increasing doses of study drug are 
administered to cohorts of three subjects.  Approximately 15 subjects will be enrolled, 9 healthy 
volunteers and 6 subjects with CF.  Three healthy volunteers without H. pylori infection will 
each receive a single 20 mg dose and 3 will each receive a single 50 mg dose.  After the safety of 
the 50 mg dose has been evaluated, 3 subjects with CF will each receive a single 20 mg dose and 
3 subjects with CF will each receive a single 50 mg dose.  Three subjects diagnosed with H. 
pylori infection will also each receive a 50 mg dose to assess potential interference of 
urease-producing bacteria in the stomach.  Three additional subjects will be added to every dose 
level at which a dose-limiting adverse event occurs.  If one subject at each dose level experiences 
DLT, then a maximum of 30 subjects will enroll. 

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9.2 
RANDOMIZATION AND BLINDING 
This is an open-label study.  Subjects are assigned to treatment based upon their sequential order 
of enrollment. 
9.3 
ANALYSIS POPULATIONS 
Three populations will be analyzed: 
• The intention-to-treat (ITT) population is those subjects who enroll in the study. 
• The safety population is the subset of the ITT population who are exposed to study drug 
and receive at least one safety evaluation. 
• The pharmacodynamics population is the subset of the safety population who contribute 
at least one post-administration evaluable breath specimen. 
9.4 
STATISTICAL ANALYSES AND METHODS 
This section outlines the nature of and rationale for the statistical methods to be used for the 
analysis of data from the study.  A separate Statistical Analysis Plan will describe the data 
handling and statistical techniques in full detail. 
All subjects who meet the inclusion criteria for the study, sign the informed consent form, and 
are enrolled and exposed to the study medication will be accounted for in the analyses.  Subjects 
who are deemed screen failures will not be accounted for in the data presentation of response or 
safety. 
Demographic variables and subject characteristics will be summarized descriptively by cohort 
and overall.  Demographic variables will include age, weight, height, and race/ethnicity.  
Continuous demographic parameters such as the subject’s age at the time of enrollment will be 
summarized for the ITT population using descriptive statistics (N, mean, median, standard 
deviation, minimum and maximum value, and 95% two-sided confidence limits).  Categorical 
demographic parameters, such as sex, will be summarized as a proportion of the ITT population.  
Other important subject characteristics for the ITT population will also be summarized using 
counts and percentages and presented in a listing format.  These tabular summaries will be 
prepared by cohort and overall. 
Heart rate, systolic and diastolic blood pressure, and body temperature recorded before study 
drug administration will be summarized descriptively by cohort and overall for the ITT 
population.  Spirometry measurements will summarized descriptively by cohort at each time 
point, including the change from baseline, for the ITT population.  The number of coughs in the 
90-second interval following the end of the administration of study drug will summarized 
descriptively by cohort. 
The incidence of all reported adverse events and treatment-emergent adverse events (TEAE) will 
be tabulated.  A TEAE is defined as an event that first occurs or worsens in intensity after the 
administration of study drug.  Adverse events will be classified by system organ class and 

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preferred term using the Medical Dictionary for Regulatory Activities (MedDRA).  For 
incidence reporting, if a subject reports more than one adverse event that is coded to the same 
system organ class or preferred term, the subject will be counted only once for that specific 
system organ class or preferred term. 
An overview of adverse events, which includes incidence of TEAEs, treatment-related adverse 
events, adverse events related to study procedures, SAEs, deaths, and adverse events leading to 
discontinuation, will be presented.  For adverse events presented by severity, the worst severity 
during the study will be presented for each subject.  The subject incidence of TEAEs will be 
summarized by system organ class and preferred term.  The subject incidence of 
treatment-related adverse events will be summarized by preferred term. 
Missing values will not be substituted using estimated values, but treated as missing in the 
statistical evaluation.  All data from all subjects who receive 13C-urea in the study will be 
included in all listings, plots, summary tables, and statistical analyses.  All analyses will be 
performed using SAS (version 9.2). 
10. 
STUDY ADMINISTRATION 
10.1 
REGULATORY AND ETHICAL CONSIDERATIONS 
10.1.1 
Regulatory Authority Approval 
The investigator will obtain approval to conduct the study from the appropriate regulatory 
agency in accordance with any applicable country-specific regulatory requirements before any 
site may initiate the study in that country. 
10.1.2 
Ethical Conduct of the Study and Ethics Approval 
This study will be conducted according to GCP; US 21 Code of Federal Regulations (CFR) Part 
50 (Protection of Human Subjects); US 21 CFR Part 56 (IRBs); US 21 CFR Part 54 (Financial 
Disclosure); International Conference on Harmonization (ICH) Guidance for Industry, E6 GCP: 
Consolidated Guidance; the Nuremberg Code; and, where applicable, the principles of the 
Declaration of Helsinki (Recommendations guiding Medical Doctors in Biomedical Research 
Involving Human Subjects). 
10.1.2.1 
Ethics Committees 
The investigator is responsible for ensuring that this protocol, the site’s informed consent form, 
and any other information that will be presented to potential subjects (e.g., advertisements or 
information that supports or supplements the informed consent form) are reviewed and approved 
by the appropriate IRB or IEC.  The investigator agrees to allow the IRB or IEC direct access to 
all relevant documents.  The IRB or IEC must be constituted in accordance with all applicable 
regulatory requirements.  The investigator will provide relevant documents or data needed for 
IRB or IEC review and approval of the study.  Before investigational products and CRFs can be 
shipped to the site, the investigator must receive copies of the IRB or IEC approval, the approved 

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informed consent form, and any other information that the IRB or IEC has approved for 
presentation to potential subjects. 
If the protocol, the informed consent form, or any other information that the IRB or IEC has 
approved for presentation to potential subjects is amended during the study, the investigator is 
responsible for ensuring that the IRB or IEC reviews and approves, where applicable, these 
amended documents.  The investigator must follow all applicable regulatory requirements 
pertaining to the use of an amended informed consent form, including obtaining IRB or IEC 
approval of the amended form, before new subjects consent to take part in the study using the 
new version of the form.  IRB or IEC approval of the consent forms must be obtained in addition 
to the approval given for the clinical study.  Regulatory review and approval may be required in 
some countries before IRB or IEC approval can be sought. 
10.1.2.2 
General Considerations 
The ethical standards defined within GCP are intended to ensure the following: 
• 
Human subjects are provided with an adequate understanding of the possible risks of 
their participation in the study, and they have a free choice to participate or not. 
• 
The study is conducted with diligence and in conformance with the protocol in such 
a way as to ensure the integrity of the findings. 
• 
The potential benefits of the research justify the risks. 
This is an investigator-sponsored study.  The investigator is responsible for all of the following: 
• 
selecting qualified co-investigators 
• 
providing co-investigators with the information they need to conduct the 
investigation properly 
• 
ensuring proper monitoring of the investigation 
• 
ensuring that appropriate regulatory agencies and all participating co-investigators 
are properly informed of significant new information regarding AEs or risks 
associated with 13C-urea. 
10.1.3 
Informed Consent 
The investigator will provide an informed consent form for this study to suit the needs of the 
institution (although it must reflect the required elements of informed consent specified in 21 
CFR Part 50.25).  The final informed consent form must be approved by the IRB or IEC.  If any 
new information becomes available that might affect subjects’ willingness to participate in the 
study, or if any amendments to the protocol require changes to the informed consent form, the 
investigator will revise the informed consent form.  The revised informed consent form must be 
approved by the IRB or IEC in advance of its use. 
Investigators must provide subjects with all the information necessary to make an informed 
decision about their participation in the study, including the nature and intended purpose of the 
study, possible benefits, and possible risks. 

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All information in the informed consent form should be provided in a language (whether written 
or spoken) that is as nontechnical as practical and that is understandable to the subjects. 
Before written informed consent is obtained, the subject should be given ample time and 
opportunity to inquire about the details of the study.  All questions must be answered to the 
satisfaction of the subject or the subject’s legally authorized representative. 
Before a subject undergoes procedures specific to the protocol, the informed consent form must 
be signed and dated by the subject or the subject’s legally authorized representative and any 
other signatories as required by the IRB or IEC in accordance with 21 CFR 50.27 (b2). 
After all required signatures have been obtained, a copy of the informed consent form should be 
provided to the subject, and the original must be kept on file at the site and made available for 
review by the IRB or IEC.  Documentation of the informed consent discussion must be noted in 
the subject’s CRF. 
10.1.4 
Investigator Reporting Requirements 
The investigator is responsible for completing and maintaining adequate and accurate CRFs and 
source documentation.  Source documentation constitutes original records (first point of entry, 
either hard copy or electronic), which may include progress notes, medication administration 
records, operation reports, laboratory reports, discharge summaries, and so on.  All CRFs should 
be completed contemporaneously in their entirety and stored in a confidential and locked 
location. 
10.2 
STUDY MONITORING 
The investigator is responsible for ensuring the proper conduct of the study with regard to 
subject protection, ethics, protocol adherence, site procedures, and integrity of the data.  The IRB 
or IEC will review study progress and CRF completion and address any concerns or questions 
regarding the study conduct.  The IRB or IEC may monitor documentation and procedures 
including, but not limited to, subjects’ informed consent documents, subject recruitment 
procedures, subjects’ compliance with the study procedures, source-data verification, drug 
accountability, use of concomitant therapy by subjects, AE and SAE documentation and 
reporting, and quality of data. 
10.3 
QUALITY ASSURANCE 
A regulatory authority or an IRB representative may visit the study site at any time during the 
study or after completion of the study to perform audits or inspections.  The purpose of an audit 
or regulatory inspection is to examine systematically and independently all study-related 
activities and documents to determine whether these activities were conducted according to the 
protocol, GCP, ICH guidelines, and any other applicable regulatory requirements.  Investigators 
should contact the IRB or IEC immediately if contacted by a regulatory agency about an 
inspection at their site. 

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10.4 
STUDY AND SITE CLOSURE 
If the investigator, IRB or IEC, or officials from regulatory agencies discover conditions arising 
during the study that indicate that the study should be halted or that the study site should be 
closed, this action may be taken after appropriate consultation between the IRB or IEC and 
investigator.  Conditions that may warrant termination of the study include, but are not limited 
to, the following: 
• 
discovery of an unexpected, serious, or unacceptable risk to the subjects enrolled in 
the study 
• 
submission of knowingly false information from the research facility to the data 
safety monitor, IRB or IEC or regulatory agencies 
• 
failure of the investigator to comply with GCP (e.g., ICH guidelines, regulatory 
agency guidelines) 
• 
insufficient adherence to protocol requirements or an unacceptably high rate of 
missing, erroneous, or improperly collected data 
• 
evidence from the blinded data of sufficient technical problems with the study that 
one could believe with a high degree of certainty that subjects are being exposed to 
the investigational drug without a realistic expectation of evaluable data 
• 
a decision on the part of the data safety monitor to suspend or discontinue testing 
evaluation or development of the product 
• 
failure of the investigator to enroll subjects into the study at an acceptable rate 
10.5 
RECORDS RETENTION 
10.5.1 
Health Insurance Portability and Accountability Act of 1996 
The investigator agrees to comply with all applicable federal, state, and local laws and 
regulations relating to the privacy of subjects’ health information, including, but not limited to, 
the Standards for Individually Identifiable Health Information, 45 CFR Parts 160 and 164 (the 
Health Insurance Portability and Accountability Act of 1996 privacy regulation).  The 
investigator shall ensure that study subjects authorize the use and disclosure of protected health 
information in accordance with the privacy regulations of the Health Insurance Portability and 
Accountability Act and in a form satisfactory to the University of New Mexico. 
10.5.2 
Financial Disclosure 
Financial disclosure is not required for this study. 
10.5.3 
Access to Original Records 
Regulatory authorities expect that monitors, auditors, and representatives of national and 
international government regulatory agency bodies have access to original source documentation 
(see examples in Section 10.1.4) to ensure data integrity.  “Original” in this context is defined as 
the first documentation of an observation and does not differentiate between hard copy and 
electronic records. 

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10.5.4 
Archiving of Study-Related Documents 
Records related to this clinical study must be retained either for at least 2 years after the last 
approval of a marketing application in an ICH region and until there are no pending or 
contemplated marketing applications in an ICH region or until at least 2 years have elapsed since 
the formal discontinuation of clinical development of the investigational product.  The 
University of New Mexico will notify the investigator as to when these documents no longer 
need to be retained for this use. 
10.6 
SUBJECT TRACKING 
Drug accountability logs, a subject identification log (to be retained by the investigator only), 
and a subject enrollment log will be used to track subject participation in the study. 
11. 
REFERENCES 
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Bauer F. The use of urea in exudative otitis media. Otolaryngologic clinics of North America. 
1970 Feb;3(1):67-78. 
Brannan JD, Gulliksson M, Anderson SD, Chew N, Kumlin M. Evidence of mast cell activation 
and leukotriene release after mannitol inhalation. Eur Respir J. 2003 Sep;22(3):491-6. 
Brannan JD, Porsbjerg C, Anderson SD. Inhaled mannitol as a test for bronchial hyper-
responsiveness. Expert Rev Respir Med. 2009 Oct;3(5):457-68. 
Cade JF, Pain MC. Lung function in provoked asthma: responses to inhaled urea, methacholine 
and isoprenaline. Clin Sci. 1972 Dec;43(6):759-69. 
Cox FR, Cox ME, Martin JR. Rapid urease test for mycobacteria: preliminary observations. J 
Clin Microbiol. 1977 Jun;5(6):656-7. 
Crews SJ, Davidson SI. Intravenous urea therapy in glaucoma. The British journal of 
ophthalmology. 1961 Dec;45(12):769-77. 
Crews SJ, Davidson SI. Biochemical observations during intravenous urea therapy. The British 
journal of ophthalmology. 1962 Oct;46(10):597-600. 
Damato JJ, Collins MT, McClatchy JK. Urease testing of mycobacteria with BACTEC 
radiometric instrumentation. J Clin Microbiol. 1982 Mar;15(3):478-80. 
Eng PA, Morton J, Douglass JA, Riedler J, Wilson J, Robertson CF. Short-term efficacy of 
ultrasonically nebulized hypertonic saline in cystic fibrosis. Pediatr Pulmonol. 1996 
Feb;21(2):77-83. 

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Appendix A 
13C-urea Breath Test Kit Instructions for Use 
Warnings and Precautions: 
1. The use of this kit is restricted by federal law to investigational use only.  The 13C-urea 
breath test should be administered only by trained personnel to patients who have 
enrolled in clinical studies approved by a local institutional review board. 
2. Reconstituted 13C-urea should not be used if the resulting solution is cloudy. 
3. No information is available on the use of 13C-urea in pregnant or nursing women. 
4. The performance characteristics for persons under the age of 18 have not been 
established for this test. 
5. The specimen integrity of breath samples and reference gases stored in breath bags under 
ambient conditions has not been determined beyond 7 days. 
6. A correlation between the number of organisms in the lungs and the 13C-urea breath test 
result has not been established. 
Preparation of 13C-urea: 
1. Remove the overseals on the vial of 13C-urea and WFI to expose the septa. 
2. Using the supplied 3 mL syringe, withdraw 3 mL of WFI and inject it into the vial of 
13C-urea. 
3. While the syringe needle remains in the vial of 13C-urea, gently swirl the 10 mL vial in 
order to ensure complete dissolution of 13C-urea.  Do not use the solution if it is cloudy or 
undissolved material is present. 
4. Draw the reconstituted 13C-urea into the syringe for subsequent charging of the nebulizer 
cup. 
Preparation of the nebulizer: 
1. Ensure availability of PARI compressor for the nebulizer. 
2. Connect the tubing from the air source to the nebulizer cup. 
3. Express the 13C-urea solution into the nebulizer cup. 
Collection of baseline sample: 
1. Place a self-adhesive bar-code sticker supplied with the kit onto the blue Mylar collection 
bag and the case report form. 
2. On the blue Mylar collection bag, record the subject ID, initials, sex, and age, collection 
date, and clock time at which the baseline sample is being taken. 
3. Remove the pull-off cap from the breath collection bag. 
4. Instruct the subject to breath normally, to take a deep breath and pause momentarily, and 
then to exhale into the mouthpiece of the collection bag. 

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5. Immediately place the cap over the inlet port of the bag to prevent sample loss. 
6. Store the breath sample at 15º to 30º C (59º to 86º F) until analyzed. 
Delivery of 13C-urea: 
1. Attach the mouthpiece to the nebulizer outlet with the expiratory valve facing up. 
2. Seat the subject in a relaxed, upright position. 
3. Ensure that the 13C-urea solution has been expressed into the nebulizer cup. 
4. Turn on power to the compressor. 
5. Instruct the subject to place the mouthpiece on top of the tongue, closing the lips around 
the mouthpiece.  The subject should take slow, deep breaths, inhaling and exhaling 
though the mouth.  The valve on the mouthpiece will open during exhalation to allow 
exhaled mist to escape. 
6. The subject should continue to breath through the mouthpiece of the nebulizer cup until 
the study drug is emptied from the nebulizer cup or until a slight sputtering sound is 
heard. 
7. Turn power off to the compressor. 
8. Dispose of nebulizer cup. 
Collection of test sample: 
1. Place a self-adhesive bar-code sticker supplied with the kit onto the pink Mylar collection 
bag. 
2. On a pink Mylar collection bag, record the subject ID, initials, sex, and age, collection 
date, and clock time at which the test sample is being taken. 
3. At the specified times after stopping inhalation of 13C-urea, remove the pull-off cap. 
4. Instruct the subject to breath normally, to take a deep breath and pause momentarily, and 
then to exhale into the mouthpiece of the collection bag. 
5. Immediately place the cap over the inlet port of the bag to prevent sample loss. 
6. Store the sample at 15º to 30º C (59º to 86º F) until analyzed.