Clinical trial NCT04725682
Conditions: Healthy Volunteers
Keywords: generic drug tacrolimus bioequivalence reference listed drug fasting condition
Sponsor: Food and Drug Administration (FDA)
Start date: 2021-01-05; Completion date: 2021-06-17
Enrollment: 67 participants; Allocation: RANDOMIZED; Intervention model: CROSSOVER

Brief summary:
This is an in-vivo study to investigate the bioequivalence of generic tacrolimus and its reference listed drug (RLD). The objective of this study is to investigate the bioequivalence of generic Tacrolimus and RLD in healthy male and non-pregnant, non-lactating female volunteers under fasting conditions. The outcome of this study will help further understanding about pharmacokinetic (PK) performance of tacrolimus in a healthy volunteer population and improve review standards for bioequivalence of narrow therapeutic index (NTI) drugs.

Detailed description:
This is a single-dose, randomized, open-label, fully replicate crossover, four-period, two- treatment, two-sequence, bioequivalence study to investigate the bioequivalence of generic tacrolimus and its reference listed drug (RLD).

Tacrolimus, a calcineurin inhibitor (CNI), has been widely used in solid organ and bone marrow transplants for more than two decades. Calcineurin, a calcium-dependent phosphatase, is instrumental for signal transduction for activation of T cell and B cell, which in turn cause production of autoinflammatory cytokines such as such as interleukin 2 (IL- 2), tumor necrosis factor-alpha (TNF-α), and interferon-gamma (IFN-γ). CNIs prevents the dephosphorylation and translocation of various factors such as the nuclear factor of activated T-cells (NF-AT), and nuclear factor kappa-light-chain enhancer of activated B-cells (NF-κB), thereby inhibiting the signal transduction responsible for growth and proliferation of activated T cells and expression of autoinflammatory cytokines. This mechanism of action results in immunosuppression and prevents organ rejection.

However, therapeutic drug monitoring is required for tacrolimus since the range between tacrolimus therapeutic and toxic tacrolimus whole blood concentrations is narrow and some toxicities are serious and/or irreversible. The objective of this study is to investigate the bioequivalence of generic Tacrolimus and RLD in healthy male and non-pregnant, non-lactating female volunteers under fasting conditions.

The outcome of this study will help further understanding about pharmacokinetic (PK) performance of tacrolimus in a healthy volunteer population and improve review standards for bioequivalence of narrow therapeutic index (NTI) drugs.

Arms: arm 1: Each subject is scheduled to receive each treatment twice by the end of the study in the order of TRTR.

Treatment T (Test): 1 × 1mg Test tacrolimus capsules (Applicant holder: Accord Healthcare, Inc.); Treatment R (Reference): 1 × 1mg RLD tacrolimus capsules arm 2: Each subject is scheduled to receive each treatment twice by the end of the study in the order of RTRT.

Treatment T (Test): 1 × 1mg Test tacrolimus capsules (Applicant holder: Accord Healthcare, Inc.); Treatment R (Reference): 1 × 1mg RLD capsules
Interventions: intervention 1: Tacrolimus
Intervention descriptions: intervention 1: a single dose of 1 mg capsule per period

Protocol documents:
Prot_SAP_000.pdf: 


BPSUSA Protocol Number: 2521
Protocol Version Number: 3.0, Date: Jun 22, 2021
Tacrolimus Capsule
Fasting Conditions
Page 2 of 68
CLINICAL STUDY PROTOCOL
A Single Dose, Open-Label, Randomized, Four-Way Crossover, Fully Replicate, 
Bioequivalence Study of Generic Tacrolimus Capsules and Prograf® Capsules in 
Healthy Volunteers under Fasting Conditions
BPSUSA Protocol Number:
2521
Protocol Version Number:
3.0
Protocol Version Date:
Jun 22, 2021
Sponsor:
U.S. Food and Drug Administration
PROTOCOL APPROVAL SIGNATORY PAGE
I am aware of, and agree to comply with, all of the procedures contained within this 
protocol and requirements of applicable regulatory agencies:
_____
________________________________
Sanjida Mahjabeen, Ph.D.
Date (Mmm dd, yyyy)


BPSUSA Protocol Number: 2521 
Protocol Version Number: 3.0, Date: Jun 22, 2021 
Tacrolimus Capsule 
Fasting Conditions 
 
Page 3 of 68 
REVISION HISTORY 
Version # 
(Version 
Date) 
Affected Sections 
1.0 (Aug 07, 
2020) 
N/A 
2.0 (Dec 23, 
2020) 
As per discussion between sponsor and BPSI/BPSUSA, the 
following sections were revised from the previously approved 
protocol: 
i) 
Protocol Title Pages and Headers – Protocol Version and 
Date revised to Version 2.0 and Dec 23, 2020, 
respectively. 
ii) 
Updated BPSUSA telephone and fax number. Following 
sections have been updated: 
• Title page 
• Section 
SPONSOR, 
INVESTIGATORS 
AND 
FACILITIES 
• Section INVESTIGATOR CREDENTIALS 
iii) 
Section Table of Contents – updated pagination 
iv) 
Tuberculosis (TB) Test and/or Questionnaire have been 
removed from the study. Following sections have been 
updated: 
• Section 2.0 
TABLE OF STUDY PROCEDURES 
• Section 5.1.1 Screening (Visit 1) 
• Section 7.3 
Exclusion Criteria 
• Section 8.6 Tuberculin Test (has been deleted) 
• APPENDIX A – SCHEDULE OF EVENTS 
v) 
Included light smokers to this study and removed cotinine 
test. Following sections have been updated:  
• Section 1.0 SYNOPSIS 
• Section 2.0 TABLE OF STUDY PROCEDURES 
• Section 4.1 Study Objective 
• Section 5.0 STUDY DESIGN 
• Section 6.0 STUDY POPULATION 
• Section 
7.0 
SUBJECT 
SELECTION 
AND 
WITHDRAWAL/TERMINATION 
• APPENDIX A – SCHEDULE OF EVENTS 


BPSUSA Protocol Number: 2521 
Protocol Version Number: 3.0, Date: Jun 22, 2021 
Tacrolimus Capsule 
Fasting Conditions 
 
Page 4 of 68 
• APPENDIX B – CLINICAL LABORATORY 
ASSESSMENT  
vi) 
The test for SARS-CoV-2 at screening has been removed, 
and the test at check-in will be done if applicable, related 
wordings have been updated. Following sections have 
been updated:  
• Section 1.0 SYNOPSIS 
• Section 2.0 TABLE OF STUDY PROCEDURES 
• Section 8.5.5 SARS-CoV-2 Test 
• APPENDIX A – SCHEDULE OF EVENTS 
vii) 
Section 
SPONSOR, 
INVESTIGATORS 
AND 
FACILITIES has been updated for the IRB official name. 
viii) 
Section 7.3 Exclusion Criteria has been updated regarding 
Systolic blood pressure and temperature normal range, 
timeframe for the restriction of beverages containing 
grapefruit and/or pomelo consumption. 
ix) 
Section 7.5 Study Restrictions has been updated regarding 
timeframe for the restriction of beverages containing 
grapefruit and/or pomelo consumption. 
x) 
Section 8.5.4 
Pulse Oximetry and Respiratory Rate 
has been updated to add normal range. 
xi) 
Section 8.6 Vital Signs has been updated regarding vital 
sign normal ranges. 
xii) 
End-of-Study (EOS) now has one-week ±3 days window. 
Following sections have been updated:  
• Section 2.0 TABLE OF STUDY PROCEDURES 
• Section 5.1.3 Post-Study [End-of-Study (EOS)]/Early 
Termination 
xiii) 
Section 8.5.3 Temperature Check. Temperature will be 
measured using an oral thermometer instead of a non-
contact thermometer.     
xiv) 
Section 9.3 Drug Inventory has been updated regarding 
the purchase of study drug. 
xv) 
Section 11.1.1 Treatment-emergent adverse events 
(TEAEs) added sentence “The PI/sub-I will be 
responsible for determining whether a TEAE is 
possibly related to the SARS-CoV-2 and document 
accordingly.” 


BPSUSA Protocol Number: 2521 
Protocol Version Number: 3.0, Date: Jun 22, 2021 
Tacrolimus Capsule 
Fasting Conditions 
 
Page 5 of 68 
xvi) 
Section 11.3 Adverse Events and Serious Adverse Events 
Reporting has been updated regarding the responsibility 
of notifying the local IRB or regulatory agency and SAE 
contact.  
xvii) 
In Section 13.1 Safety Data Set, related sentences have 
been updated to: “In a case where SARS-CoV-2 related 
TEAEs lead to the withdrawal or dismissal of 
approximately 5 % or more of the dosed subjects, a 
subgroup analysis related to Adverse Events (AE) may 
be performed. And the safety listings and tables for AE 
with subgroup and the total safety population will 
both be presented.” 
xviii) Section 13.2.2 Statistical Analysis of Pharmacokinetic 
Data, updated statistical model regarding dosing in 
groups. 
xix) 
In Section 15.3 Study Completion/Termination, related 
sentences have been updated to “BioPharma Services 
USA Inc. and/or the Sponsor reserve the right to terminate 
the study at any time, and for any reason related to the 
safety of subjects or due to any other non-study 
specific reason.” 
xx) 
Minor administrative/grammatical revisions were also 
made. 
The study ICF was also revised to reflect applicable changes. 
3.0 (Jun 22, 
2021) 
As per discussion between sponsor and BPSI/BPSUSA, the 
following sections were revised from the previously approved 
protocol: 
i) 
Protocol Title Pages and Headers – Protocol Version and 
Date revised to Version 3.0 and Jun 22, 2021, 
respectively. 
ii) 
Non or light smoking has been removed from Section 1.0. 
4.1, 5.1.2.2 and 6.0.  
iii) 
Confinement has been updated to read as: “From at least 
12   hours prior to dosing until at least 72 hours post dose, 
for a total of approximately 84 hours for each study 
period.” And Footer in Section 2.0 and 5.2 has been 
revised accordingly. 
iv) 
Additional measurements of the heart rate and blood 
pressure have been added at 48- and 72-hour post-dose for 


BPSUSA Protocol Number: 2521 
Protocol Version Number: 3.0, Date: Jun 22, 2021 
Tacrolimus Capsule 
Fasting Conditions 
 
Page 6 of 68 
additional safety monitoring. Therefore, Sections 1.0, 2.0, 
8.6 and Appendix A have been updated. 
 
v) 
Section 7.1 sample size has been updated for the 
clarification. 
vi) 
MedWatch has been replaced with “FDA Adverse Event 
Reporting System (FAERS)” in Section 11.3  
vii) 
Section 12.1.1 and 13.1 has been updated to further clarify 
the samples to be assayed and data to be included in the 
pharmacokinetic analysis. 
viii) 
AUC72 is going to be measured as a primary endpoint in 
this study. Therefore, Sections 1.0, 13.2.1, 13.2.2, 13.4 
and Appendix C have been updated.  
ix) 
Section 13.3.2 was updated to include the listing for 
magnesium, creatinine kinase and lipid panel test.  
x) 
Section 13.3.3 has been updated to include the following 
statements in line with the SAP: Descriptive statistics of 
all quantitative measurements from all clinical laboratory 
parameters will be presented by the type of laboratory 
test, laboratory parameter, and visit. 


BPSUSA Protocol Number: 2521 
Protocol Version Number: 3.0, Date: Jun 22, 2021 
Tacrolimus Capsule 
Fasting Conditions 
 
Page 7 of 68 
TABLE OF CONTENTS 
TITLE PAGE .................................................................................................................... 1 
REVISION HISTORY ..................................................................................................... 3 
TABLE OF CONTENTS ................................................................................................. 7 
SPONSOR, INVESTIGATORS AND FACILITIES .................................................. 10 
INVESTIGATOR CREDENTIALS ............................................................................. 11 
1.0 
SYNOPSIS ............................................................................................................. 12 
2.0 
TABLE OF STUDY PROCEDURES .................................................................. 15 
3.0 
INTRODUCTION: BACKGROUND INFORMATION AND SCIENTIFIC 
RATIONALE ......................................................................................................... 17 
3.1 
Background Information ...................................................................................... 17 
3.2 
Rationale ................................................................................................................ 18 
3.3 
Product Information ............................................................................................. 18 
4.0 
STUDY OBJECTIVE ........................................................................................... 21 
4.1 
Study Objective ....................................................................................................... 21 
5.0 
STUDY DESIGN ................................................................................................... 21 
5.1 
Discussion of Study Design .................................................................................... 21 
5.1.1 Screening (Visit 1) .................................................................................................. 22 
5.1.2 Treatment Phase (Visit 2 to 14) ............................................................................... 23 
5.1.3 Post-Study [End-of-Study (EOS)]/Early Termination ............................................ 24 
5.2 
Study Duration and Confinement ............................................................................ 24 
5.3 
Randomization and Blinding ................................................................................... 24 
6.0 
STUDY POPULATION ........................................................................................ 25 
7.0 
SUBJECT SELECTION AND WITHDRAWAL/TERMINATION ................ 25 
7.1 
Sample Size ............................................................................................................. 25 
7.2 
Inclusion Criteria ..................................................................................................... 26 
7.3 
Exclusion Criteria .................................................................................................... 27 
7.4 
Informed Consent Procedure ................................................................................... 31 
7.5 
Study Restrictions ................................................................................................... 31 
7.6 
Withdrawal and Dismissal ...................................................................................... 34 
8.0 
STUDY PROCEDURES ....................................................................................... 35 
8.1 
Demographics and Other Baseline Characteristics ................................................. 35 
8.1.1 Demographics .......................................................................................................... 35 
8.1.2 Medical History ....................................................................................................... 35 
8.1.3 Medication History .................................................................................................. 35 
8.1.4 Clinical Laboratory Assessments ............................................................................ 36 
8.2 
Food and Fluid Intake ............................................................................................. 36 
8.3 
Dosing ..................................................................................................................... 36 


BPSUSA Protocol Number: 2521 
Protocol Version Number: 3.0, Date: Jun 22, 2021 
Tacrolimus Capsule 
Fasting Conditions 
 
Page 8 of 68 
8.4 
Safety Monitoring ................................................................................................... 36 
8.5 
COVID-19 prevention and control .......................................................................... 37 
8.5.1 COVID-19 questionnaire ........................................................................................ 37 
8.5.2 COVID-19 symptom check ..................................................................................... 37 
8.5.3 Temperature check .................................................................................................. 37 
8.5.4 Pulse Oximetry and Respiratory Rate ..................................................................... 37 
8.5.5 SARS-CoV-2 Test ................................................................................................... 37 
8.6 
Vital Signs ............................................................................................................... 38 
8.7 
ECG ......................................................................................................................... 38 
8.8 
Health Monitoring ................................................................................................... 38 
8.9 
Sample Collection and Processing .......................................................................... 38 
8.10 Sample Shipment ..................................................................................................... 39 
9.0 
DRUG PRODUCTS .............................................................................................. 40 
9.1 
Drug Information ..................................................................................................... 40 
9.2 
Labeling, Maintenance, and Retention .................................................................... 40 
9.3 
Drug Inventory ........................................................................................................ 40 
9.4 
Disposition of Unused Tacrolimus .......................................................................... 41 
10.0 POTENTIAL RISKS AND BENEFITS .............................................................. 41 
10.1.1 Potential Risks ........................................................................................................ 41 
10.1.2 Potential Benefits ................................................................................................... 41 
11.0 SAFETY ASSESSMENT ...................................................................................... 42 
11.1 Definitions ............................................................................................................... 42 
11.1.1 Treatment-emergent adverse events (TEAEs)........................................................ 43 
11.2 Characteristics of an Adverse Event ....................................................................... 44 
11.2.1 Relationship to Study Drug .................................................................................... 44 
11.2.2 Expectedness of SAE ............................................................................................. 44 
11.2.3 Severity Assessment ............................................................................................... 45 
11.3 Adverse Events and Serious Adverse Events Reporting ......................................... 45 
11.4 Procedures for Reporting Pregnancy ....................................................................... 47 
12.0 BIOANALYTICAL ANALYSIS ......................................................................... 48 
12.1 Analytical Procedures ............................................................................................. 48 
12.1.1 Samples to be Assayed ........................................................................................... 48 
12.1.2 Analyte(s) in Biological Matrix ............................................................................. 48 
12.1.3 Incurred sample reproducibility ............................................................................. 48 
13.0 PHARMACOKINETIC AND STATISTICAL ANALYSIS ............................. 49 
13.1 Pharmacokinetic and Statistical Analysis Data Set ................................................. 49 
13.2 Analysis of Pharmacokinetic Data .......................................................................... 49 
13.2.1 Pharmacokinetic Analysis ...................................................................................... 50 
13.2.2 Statistical Analysis of Pharmacokinetic Data ........................................................ 51 


BPSUSA Protocol Number: 2521 
Protocol Version Number: 3.0, Date: Jun 22, 2021 
Tacrolimus Capsule 
Fasting Conditions 
 
Page 9 of 68 
13.3 Safety Analysis ........................................................................................................ 55 
13.3.1 Safety Data Set ....................................................................................................... 55 
13.3.2 Safety Listings ........................................................................................................ 55 
13.3.3 Safety Tables .......................................................................................................... 55 
13.4 Bioequivalence Criteria [7, 10] ............................................................................... 56 
14.0 ETHICAL CONSIDERATIONS / PROTECTION OF HUMAN SUBJECTS57 
14.1 Basic Principles ....................................................................................................... 57 
14.2 Institutional Review Board ...................................................................................... 57 
14.3 Informed Consent Form .......................................................................................... 57 
14.4 Confidentiality ......................................................................................................... 57 
14.5 Compensation for Participation ............................................................................... 58 
15.0 ADMINISTRATIVE CONSIDERATIONS ....................................................... 60 
15.1 Revisions and/or Amendments to the Protocol ....................................................... 60 
15.2 Investigator Responsibilities ................................................................................... 60 
15.3 Study Completion/Termination ............................................................................... 60 
15.4 Sponsor Visits ......................................................................................................... 61 
16.0 DATA MANAGEMENT/RECORD KEEPING ................................................ 61 
16.1 Source Data ............................................................................................................. 61 
16.2 Quality of Data ........................................................................................................ 61 
16.3 Retention of Documents .......................................................................................... 61 
17.0 REFERENCES ...................................................................................................... 62 
APPENDIX A – SCHEDULE OF EVENTS ................................................................ 63 
APPENDIX B – CLINICAL LABORATORY ASSESSMENT ................................. 65 
APPENDIX C – LIST OF COMMON ABBREVIATIONS ....................................... 66 
APPENDIX D – PROGRAF® LABEL .......................................................................... 68 
 


BPSUSA Protocol Number: 2521 
Protocol Version Number: 3.0, Date: Jun 22, 2021 
Tacrolimus Capsule 
Fasting Conditions 
 
Page 10 of 68 
SPONSOR, INVESTIGATORS AND FACILITIES 
Principal 
Investigator: 
Dr. Artan Markollari, M.D.  
BioPharma Services USA 
Inc. 
 
 
Pharmacokinetic / 
Statistical Facility: 
 
 
  
 
 
 
 
Clinical 
Laboratory: 
 
 
 
Sponsor: 
OGD/CDER/FDA, 
 
 
 
 
 
 
Clinical Facility 
 
Designated, Local 
Institutional Review 
Board (IRB) Review 
and Oversight: 
 
 
 
 
 
Bioanalytical 
Facility: 
 
 
 
 


BPSUSA Protocol Number: 2521 
Protocol Version Number: 3.0, Date: Jun 22, 2021 
Tacrolimus Capsule 
Fasting Conditions 
 
Page 11 of 68 
 
INVESTIGATOR CREDENTIALS 
Dr. Artan Markollari, Principal Investigator 
 
 
____________________________________ ________________________________ 
Dr. Artan Markollari, M.D.  
BioPharma Services USA Inc. 
 
 
 
Date (Mmm dd, yyyy) 
 


BPSUSA Protocol Number: 2521 
Protocol Version Number: 3.0, Date: Jun 22, 2021 
Tacrolimus Capsule 
Fasting Conditions 
 
Page 12 of 68 
1.0 
SYNOPSIS 
Title: 
A Single Dose, Open-Label, Randomized, Four-Way Crossover, Fully Replicate, 
Bioequivalence Study of Generic Tacrolimus Capsules and Prograf® Capsules in 
Healthy Volunteers under Fasting Conditions. 
Objective: 
The objective of this study is to investigate the bioequivalence of Tacrolimus 1 mg 
capsules 
 and Prograf® 1 mg capsules 
 
in healthy male and non-pregnant, non-lactating female volunteers under fasting 
conditions. 
Experimental Design: 
A single-dose, randomized, open-label, fully replicate crossover, four-period, two-
treatment, two-sequence, bioequivalence study. 
Study Rationale:  
This is an in-vivo study to investigate the bioequivalence of generic tacrolimus and its 
reference listed drug (RLD). The outcome of this study will help further the Agency’s 
understanding about pharmacokinetic (PK) performance of tacrolimus in a healthy 
volunteer population and improve review standards for bioequivalence of narrow 
therapeutic index (NTI) drugs. 
Study Population: 
72 subjects, including 8 standbys, will be enrolled to ensure dosing of approximately 
64 subjects. The subjects may be dosed in more than one cohort. 
 
Healthy male and non-pregnant, non-lactating female volunteers, 18 - 59 years of age, 
inclusive, with a body mass index (BMI) within 18.5-33.0 kg/m2, inclusive. 
Test Drug (T): 
Tacrolimus 1 mg capsules (
 
Reference Drug (R): 
Prograf® 1 mg capsules 
 
Treatment: 
1 × 1 mg capsules in each study period 
Duration of 
Confinement: 
From at least 12 hours prior to dosing until at least 72 hours post-dose, for a total of 
approximately 84 hours for each study period. 
Washout: 
At least 14 days between each dosing. 
Safety Monitoring:  
• 
Vital signs (blood pressure [BP] and heart rate [HR]) will be obtained at pre-
dose and at 1, 2, 4, 6, 12, 24, 48 and 72 hours after dosing in each study 
period.  
• 
All tests in Appendix B at screening, and selected tests at each period check-
in and post-study. 
• 
The PI/Sub-Investigator will be present from approximately 30 minutes 
prior to dosing until at least 4 hours after the last subject is dosed in each 
study period. The PI/Sub-Investigator will remain on-call throughout the 
duration of the study. 
COVID-19 prevention 
and control: 
• Temperature, pulse oximetry and respiratory rate will be measured at screening, at 
each check-in, at pre-dose, at 6- and 12-hours post dose, then at every 12 hours until 
discharge at each study period, at return visits and at end of study (EOS) visit.  


BPSUSA Protocol Number: 2521 
Protocol Version Number: 3.0, Date: Jun 22, 2021 
Tacrolimus Capsule 
Fasting Conditions 
 
Page 13 of 68 
• A COVID-19 symptom check and COVID-19 questionnaire will also be completed 
at screening, at each check-in, at return visits and at EOS visit. 
• If applicable, a locally available test for SARS-CoV-2 will be performed at each 
period check-in. 
Type of Vacutainer  
Pre-chilled K2EDTA Vacutainer®, 4 mL  
Blood Sampling Time 
points: 
28 PK Timepoints: Pre-dose (0 hour) and at 0.25, 0.50, 0.75, 1.00, 1.25, 1.50, 1.75, 
2.00, 2.25, 2.50, 2.75, 3.00, 3.50, 4.00, 5.00, 6.00, 8.00, 10.00, 12.00, 16.00, 24.00, 
36.00, 48.00, 72.00, 96.00, 120.00 and 144.00 hours after study drug dosing in each 
study period. 
Total Blood Volume: 
Approximately 520 mL of blood, including blood samples for PK time points, pre- and 
post- study procedures and the selected laboratory testing at each study period. 
Analyte(s) to be 
measured: 
Whole blood samples will be assayed for tacrolimus using a validated analytical 
method according to the principles of Good Laboratory Practice. 
Statistical Analysis: 
 
 
 
The ln-transformed PK parameters AUC72, AUCt, AUCinf and Cmax will be analyzed 
using both an unscaled average bioequivalence procedure and a reference-scaled 
average bioequivalence procedure.  
 
Determination of the Within-subject Standard Deviation of the Test and 
Reference Products ( 𝑺𝑺𝑾𝑾𝑾𝑾 and 𝑺𝑺𝑾𝑾𝑾𝑾 ) for the PK Parameters AUC72, AUCt, AUCinf 
and Cmax 
 
Reference Scaled Average Bioequivalence Procedure  
The 95% upper confidence bound for
 will be determined for the 
ln-transformed PK parameters AUC72, AUCt, AUCinf and Cmax using a PROC MIXED 
model in SAS.  
 
Unscaled Average Bioequivalence Procedure  
The 90% Confidence Intervals (CI) for the Test/Reference ratios of geometric means 
for the PK parameters AUC72, AUCt, AUCinf and Cmax will be calculated using a PROC 
MIXED model in SAS.  
 
Tmax, Lambda and T1/2 will be analyzed using an un-scaled procedure. 
 
Calculation of the 90% CI for the Ratio of the Within-subject Standard Deviation 
of Test Product to Reference Product (𝝈𝝈𝑾𝑾𝑾𝑾/ 𝝈𝝈𝑾𝑾𝑾𝑾) 
The 90% confidence interval of the ratio of the within subject standard deviation of 
test product to reference product (𝝈𝝈𝑾𝑾𝑾𝑾/ 𝝈𝝈𝑾𝑾𝑾𝑾) will be calculated for tacrolimus.  
 
Bioequivalence 
Criteria: 
 
 
 
To establish bioequivalence, all the following conditions for individual PK 
parameter(s) AUC72and Cmax must be met: 
• 
The 90% CI for the Test/Reference ratios of geometric means are completely 
contained within 80.00%-125.00%. 
2
2)
(
WR
R
T
S
θ
µ
µ
−
−


BPSUSA Protocol Number: 2521 
Protocol Version Number: 3.0, Date: Jun 22, 2021 
Tacrolimus Capsule 
Fasting Conditions 
 
Page 14 of 68 
• 
The 95% upper confidence bound for 
 should be less than 
or equal to 0. 
• 
The upper limit of the 90% confidence interval for the Test/Reference ratio of 
the within-subject standard deviation (𝝈𝝈𝑾𝑾𝑾𝑾/ 𝝈𝝈𝑾𝑾𝑾𝑾) is less than or equal to 2.5. 
 
 
 
2
2)
(
WR
R
T
S
θ
µ
µ
−
−


BPSUSA Protocol Number: 2521 
Protocol Version Number: 3.0, Date: Jun 22, 2021 
Tacrolimus Capsule 
Fasting Conditions 
 
Page 15 of 68 
2.0 
TABLE OF STUDY PROCEDURES 
 
 
Procedure/Activity 
Time points 
Visit 1 
Visit 2 to 17 
Screening 
Each 
Period 
Check-in 
Period 1 
Period 2 
Period 3 
Period 4 
Post-
Study/ 
EOS^ 
Written informed 
consent 
X 
 
 
 
 
 
 
Drugs of Abuse 
X 
X 
 
 
 
 
 
Alcohol Test 
X 
X 
 
 
 
 
 
Pregnancy (hCG) 
Test* 
X 
X 
 
 
 
 
 
Vital Signs (BP & HR) 
X 
X 
Xa 
Xa 
Xa 
Xa 
X 
Safety measures for 
COVID-19# (Symptom 
Check, Temperature, 
Pulse Oximetry and 
RR) 
X 
X 
X 
X 
X 
X 
X  
SARS-CoV-2 Testc 
 
X 
 
 
 
 
 
BMI 
X 
 
 
 
 
 
 
Laboratory Testing⸸ 
(Appendix B) 
X 
X 
 
 
 
 
X 
ECG 
X 
 
 
 
 
 
 
Inclusion/Exclusion 
Assessment 
X 
Xb 
 
 
 
 
 
Restrictions 
Compliance Check 
 
Xd 
Xd 
Xd 
Xd 
Xd 
 
Physical Exam 
X 
  
 
 
 
 
X 
Tacrolimus Capsule 
Administration 
 
 
X 
X 
X 
X 
 
PK Sampling 
 
 
Xe 
Xe 
Xe 
Xe 
 
Adverse Event 
Reporting 
 
Xf 
Xf 
Xf 
Xf 
Xf 
X 


BPSUSA Protocol Number: 2521 
Protocol Version Number: 3.0, Date: Jun 22, 2021 
Tacrolimus Capsule 
Fasting Conditions 
 
Page 16 of 68 
Meals 
 
Xg 
Xg 
Xg 
Xg 
Xg 
 
^ 
EOS with one-week ±3 days window. 
* 
Serum pregnancy test at screening and urine pregnancy test at each period check-in. 
# 
Temperature, pulse oximetry and respiratory rate will be measured at screening, at each check-in, at 
pre-dose, at 6- and 12-hours post dose, then at every 12 hours until discharge at each study period, at 
return visits and at EOS visit. A COVID-19 symptom check, and COVID-19 questionnaire will also 
be completed at screening, at each check-in, at return visits and at EOS visit.  
⸸ 
All tests in Appendix B at screening, and selected tests at each check-in and at EOS (details in 
Appendix A and Appendix B). 
a- 
Vital signs measurements (BP and HR) to be obtained at pre-dose and at 1, 2, 4, 6, 12,24, 48 and 72 
hours after dosing in each study period. 
b 
Only at check-in for the period 1. 
c- 
At each period check-in, if applicable. 
d- 
Confirmed at check-in and each return visits, if applicable. 
e- 
PK sampling: Pre-dose (0 hour) and at 0.25, 0.50, 0.75, 1.00, 1.25, 1.50, 1.75, 2.00, 2.25, 2.50, 2.75, 
3.00, 3.50, 4.00, 5.00, 6.00, 8.00, 10.00, 12.00, 16.00, 24.00, 36.00, 48.00, 72.00, 96.00, 120.00 and 
144.00 hours after study drug dosing in each study period. PK sampling at 96, 120 and 144 hours 
will be collected during outpatient return visits.  
f- 
Pre-dose conditions at Period 1 check-in and each return visit, if applicable. AEs will be monitored 
throughout the study. 
g- 
Meals will be served at the scheduled times during confinement prior to dosing and at approximately 
4.5, 9.5, 13.5, 24.5, 28.5, 33.5, 37.5, 48.5, 52.5, 57.5 and 61.5 hours after dosing. 
 
 


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3.0 
INTRODUCTION: BACKGROUND INFORMATION AND SCIENTIFIC 
RATIONALE 
3.1 
Background Information 
Tacrolimus, a calcineurin inhibitor (CNI), has been widely used in solid organ and 
bone marrow transplants for more than two decades. When an organ is transplanted 
between a donor and a genetically non identical recipient, the recipient’s immune 
system may recognize the transplant “foreign” and can trigger humoral immune 
response by activating immune cells such as T cells, B cells, and macrophages 
causing transplant rejection. Calcineurin, a calcium-dependent phosphatase, is 
instrumental for signal transduction for activation of T cell and B cell, which in turn 
cause production of autoinflammatory cytokines such as such as interleukin 2 (IL-
2), tumor necrosis factor-alpha (TNF-α), and interferon-gamma (IFN-γ). CNIs 
prevents the dephosphorylation and translocation of various factors such as the 
nuclear factor of activated T-cells (NF-AT), and nuclear factor kappa-light-chain 
enhancer of activated B-cells (NF-κB), thereby inhibiting the signal transduction 
responsible for growth and proliferation of activated T cells and expression of 
autoinflammatory 
cytokines. 
This 
mechanism 
of 
action 
results 
in 
immunosuppression and prevents organ rejection. Tacrolimus is one of the widely 
used CNI immunosuppressants in liver and kidney transplant patients [1]. 
 
Oral bioavailability of tacrolimus is low, ranged from 11.2 to 19.1% in previous 
studies in adult kidney recipients. The limited oral bioavailability of tacrolimus 
could be attributed to- 
• Extensive metabolism through CYP3A4 and 3A5 enzymes which are 
abundantly expressed in gut [1],  
• Poor oral absorption of the drug resulting from poor aqueous solubility [1].  
 
Optimum tacrolimus bioavailability is critical for - 
i. Achieving desired clinical endpoint: The trough blood concentrations are 
significantly related to clinical endpoints, i.e., there was a significant 
correlation between increased trough concentration and decreased risk of 
acute rejection and vice versa [2]. 
ii. Minimizing acute toxicity: Long term use of tacrolimus is associated with 
nephrotoxicity. Study showed monitoring trough tacrolimus whole blood 
concentrations below 15 ng/mL decreased the occurrence of nephrotoxicity 
[3]. 
 
To enhance the oral bioavailability intricate formulation strategies have been 
studied. One such strategy in formulating tacrolimus into amorphous solid 
dispersions (ASD). In ASDs, the amorphous form of the drug, which is believed to 
have increased solubility compared to crystalline form, is stabilized with the aid of 
a polymer. Currently, Prograf® and the six approved generic products referencing 
Prograf® are being formulated using ASD strategy. There are different 


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manufacturing processes to produce ASDs such as spray-freeze drying, hot melt 
extrusion etc. However, the interplay of manufacturing, processing, and storage 
conditions on the critical quality attributes of ASDs that impact dissolution, 
precipitation, and ultimately in vivo performances are poorly understood. For 
example, amorphous tacrolimus can be converted to crystalline tacrolimus during 
manufacture and/or storage, which may lead to decreased solubility and thereby 
suboptimal therapeutic level. This is undesirable as blood concentration is tied to 
therapeutic performance [1].  
 
There are concerns in the transplant community that generic ASDs may not perform 
similarly to the brand ASD. To this end, the Office of Generic Drugs aims to 
evaluate BE of one of the generic products (generic from Acord) which showed 
likely inequivalent performance in vitro and in vivo in a preclinical setting [4, 5]. 
3.2 
Rationale 
This is an in-vivo study to investigate the bioequivalence of a generic tacrolimus 
product in comparison with that of its reference listed drug (RLD). The outcome of 
this study will help advance the Agency’s understanding about the generic 
tacrolimus product’s pharmacokinetic (PK) performance in healthy volunteer 
population and improve review standards for bioequivalence of NTI drugs.   
3.3 
Product Information 
PROGRAF® [6] 
 
Prograf® (tacrolimus) capsule is indicated for the prophylaxis of organ rejection in 
adult and pediatric patients receiving allogeneic kidney, liver, and heart transplants, 
in combination with other immunosuppressants. 
 
Mechanism of Action 
 
Tacrolimus binds to an intracellular protein, FKBP-12. A complex of tacrolimus-
FKBP-12, calcium, calmodulin, and calcineurin (a ubiquitous mammalian 
intracellular enzyme) is then formed, after which the phosphatase activity of 
calcineurin is inhibited. Such inhibition prevents the dephosphorylation \ of various 
factors such as the nuclear factor of activated T-cells (NF-AT), and nuclear factor 
kappa-light-chain enhancer of activated B-cells (NF-κB). This dephosphorylation, 
in turn, prevents the translocation of these factors to the nucleus, leading to 
decreased activity and of genes coding for many cytokines.  Tacrolimus inhibits the 
expression and/or production of several cytokines that include interleukin (IL)-1 
beta, IL-2, IL-3, IL-4, IL-5, IL-6, IL-8, IL-10, gamma interferon, tumor necrosis 
factor-alpha, and granulocyte macrophage colony-stimulating factor. Tacrolimus 
also inhibits IL-2 receptor expression and nitric oxide release, induces apoptosis and 
production of transforming growth factor beta that can lead to immunosuppressive 


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activity. The net result is the inhibition of T-lymphocyte activation and proliferation, 
as well as T-helper-cell-dependent B-cell response (i.e., immunosuppression). 
 
Pharmacokinetics 
 
Tacrolimus activity is primarily due to the parent drug. The pharmacokinetic 
parameters (mean ± S.D.) of tacrolimus have been determined following intravenous 
(IV) and/or oral (PO) administration in healthy volunteers, and also in kidney, liver, 
and heart transplant patients. 
 
Population 
N 
Route 
(Dose) 
Parameters 
Cmax 
(ng/mL) 
Tmax 
(hr) 
AUC 
(ng•hr/mL
) 
t1/2 (hr) 
CL 
(L/hr/kg) 
V 
(L/kg) 
Healthy 
Volunteers 
8 
IV (0.025 
mg/kg/4 hr) 
* 
* 
652† ± 156 
34.2 ± 
7.7 
0.040 ± 
0.009 
1.91 ± 
0.31 
30 
PO (5 mg) 
(granules) 
35.6 ± 10.9 
1.3 ± 
0.5 
320† ± 164 
32.1 ± 
5.9 
‡ 
‡ 
PO (5 mg) 
(capsules) 
28.8 ± 8.9 
1.5 ± 
0.7 
266† ± 95 
32.3 ± 
8.8 
‡ 
‡ 
 
Due to inter-subject variability in tacrolimus pharmacokinetics, individualization of 
the dosing regimen is necessary for optimal therapy. Pharmacokinetic data indicate 
that whole blood concentrations rather than plasma concentrations serve as the more 
appropriate sampling compartment to describe tacrolimus pharmacokinetics. 
 
Absorption 
Absorption of tacrolimus from the gastrointestinal tract after oral administration is 
incomplete and variable. The absolute bioavailability of tacrolimus was 17 ± 10% 
in adult kidney transplant patients (N = 26), 22 ± 6% in adult liver transplant patients 
(N = 17), 23 ± 9% in adult heart transplant patients (N = 11) and 18 ± 5% in healthy 
volunteers (N = 16). 
 
A single dose trial conducted in 32 healthy volunteers established the 
bioequivalence of the 1 mg and 5 mg capsules. Another single dose trial in 32 
healthy volunteers established the bioequivalence of the 0.5 mg and 1 mg capsules. 
Tacrolimus maximum blood concentrations (Cmax) and area under the curve (AUC) 
appeared to increase in a dose-proportional fashion in 18 fasted healthy volunteers 
receiving a single oral dose of 3, 7, and 10 mg. 
 
In 18 kidney transplant patients, tacrolimus trough concentrations from 3 to 30 
ng/mL measured at 10-12 hours post-dose (Cmin) correlated well with the AUC 
(correlation coefficient 0.93). In 24 liver transplant patients over a concentration 
range of 10 to 60 ng/mL, the correlation coefficient was 0.94. In 25 heart transplant 
patients over a concentration range of 2 to 24 ng/mL, the correlation coefficient was 
0.89 after an oral dose of 0.075 or 0.15 mg/kg/day at steady state. 
 


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Prograf® granules are indicated for pediatric patients who have difficulty 
swallowing Prograf® capsules. In a healthy volunteer adult study, the systemic 
exposure to tacrolimus (AUC) for Prograf® Granules was approximately 16% 
higher than that for Prograf® capsules when administered as single doses. Hence, if 
pediatric patients are converted between formulations (capsules to granules or 
granules to capsules), therapeutic drug monitoring must be performed, and dose 
adjustments made to ensure that systemic exposure to tacrolimus is maintained. 
 
Food Effects 
The rate and extent of tacrolimus absorption were greatest under fasted conditions. 
The presence and composition of food decreased both the rate and extent of 
tacrolimus absorption when administered to 15 healthy volunteers. 
 
The effect was most pronounced with a high-fat meal (848 kcal, 46% fat): mean 
AUC and Cmax were decreased 37% and 77%, respectively; Tmax was lengthened 5-
fold. A high-carbohydrate meal (668 kcal, 85% carbohydrate) decreased mean AUC 
and mean Cmax by 28% and 65%, respectively. 
 
In healthy volunteers (N = 16), the time of the meal also affected tacrolimus 
bioavailability. When given immediately following the meal, mean Cmax was 
reduced 71%, and mean AUC was reduced 39%, relative to the fasted condition. 
When administered 1.5 hours following the meal, mean Cmax was reduced 63%, and 
mean AUC was reduced 39%, relative to the fasted condition. 
 
In 11 liver transplant patients, Prograf® administered 15 minutes after a high fat (400 
kcal, 34% fat) breakfast, resulted in decreased AUC (27 ± 18%) and Cmax (50 ± 
19%), as compared to a fasted state. 
 
Prograf® capsules should be taken consistently every day either with or without food 
because the presence and composition of food decreases the bioavailability of 
Prograf®. 
 
Distribution 
The plasma protein binding of tacrolimus is approximately 99% and is independent 
of concentration over a range of 5-50 ng/mL. Tacrolimus is bound mainly to albumin 
and alpha-1-acid glycoprotein and has a high level of association with erythrocytes. 
The distribution of tacrolimus between whole blood and plasma depends on several 
factors, such as hematocrit, temperature at the time of plasma separation, drug 
concentration, and plasma protein concentration. In a U.S. trial, the ratio of whole 
blood concentration to plasma concentration averaged 35 (range 12 to 67). 
 
Metabolism 
Tacrolimus is extensively metabolized by the mixed-function oxidase system, 
primarily the cytochrome P-450 system (CYP3A). A metabolic pathway leading to 
the formation of 8 possible metabolites has been proposed. Demethylation and 


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hydroxylation were identified as the primary mechanisms of biotransformation in 
vitro. The major metabolite identified in incubations with human liver microsomes 
is 13-demethyl tacrolimus. In in vitro studies, a 31-demethyl metabolite has been 
reported to have the same activity as tacrolimus. 
 
Excretion 
The mean clearance following IV administration of tacrolimus is 0.040, 0.083, 
0.053, and 0.051 L/hr/kg in healthy volunteers, adult kidney transplant patients, 
adult liver transplant patients, and adult heart transplant patients, respectively. In 
man, less than 1% of the dose administered is excreted unchanged in urine. 
 
In a mass balance study of IV-administered radiolabeled tacrolimus to 6 healthy 
volunteers, the mean recovery of radiolabel was 77.8 ± 12.7%. Fecal elimination 
accounted for 92.4 ± 1.0% and the elimination half-life based on radioactivity was 
48.1 ± 15.9 hours whereas it was 43.5 ± 11.6 hours based on tacrolimus 
concentrations. The mean clearance of radiolabel was 0.029 ± 0.015 L/hr/kg and 
clearance of tacrolimus was 0.029 ± 0.009 L/hr/kg. When administered PO, the 
mean recovery of the radiolabel was 94.9 ± 30.7%. Fecal elimination accounted for 
92.6 ± 30.7%, urinary elimination accounted for 2.3 ± 1.1% and the elimination half-
life based on radioactivity was 31.9 ± 10.5 hours whereas it was 48.4 ± 12.3 hours 
based on tacrolimus concentrations. The mean clearance of radiolabel was 0.226 ± 
0.116 L/hr/kg and clearance of tacrolimus was 0.172 ± 0.088 L/hr/kg. 
 
4.0 
STUDY OBJECTIVE  
4.1 
Study Objective 
The objective of this study is to investigate the bioequivalence of Tacrolimus 1 mg 
capsules 
 and Prograf® 1 mg capsules 
 
in healthy male and non-pregnant, non-lactating female volunteers under fasting 
condition. 
 
5.0 
STUDY DESIGN 
5.1 
Discussion of Study Design 
This is a single-dose, randomized, open-label, four-period, two-sequence, two-
treatment, single-center, crossover, bioequivalence study of tacrolimus 1 mg 
capsules 
 and Prograf® 1 mg capsules 
 
under fasting conditions.  
 
FDA has concluded that tacrolimus is a NTI drug based on the following evidence:  
- 
The range between tacrolimus therapeutic and toxic tacrolimus whole 
blood concentrations is narrow;  


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- 
Some tacrolimus toxicities are serious and/or irreversible;  
- 
Subtherapeutic tacrolimus concentrations may lead to morbidity/mortality 
associated with graft rejection;  
- 
Tacrolimus requires individual dose titration to achieve a satisfactory 
balance between maximizing efficacy and minimizing serious dose-
related toxicity; Therapeutic drug monitoring is routinely employed to 
facilitate tacrolimus dose titration; and  
- 
Tacrolimus has small to medium within subject variability. 
 
This study is designed based on FDA draft guidance on Tacrolimus capsule (revised 
on Dec 2012) [7]. The bioequivalence assessment will be based on the 
recommendation from FDA for NTI drug.  
5.1.1 Screening (Visit 1) 
Subjects will review ICF and document their consent by signing the ICF prior to any 
study related procedures.  
 
Screening procedures will be conducted within thirty (30) days prior to dosing in 
Period 1. The Principal Investigator (PI)/Sub-Investigator will review all screening 
results/data to assess eligibility of each potential subject.  In case of an unforeseeable 
delay, a selected safety screening test may be repeated at the PI’s discretion. 
The Screening procedures will include (but not limited to): COVID-19 prevention 
and control measures, collection of demographic data, medical and medication 
histories, physical examinations, body measurements (height and weight for Body 
Mass Index [BMI] calculation), vital signs (seated blood pressure [BP], heart rate 
[HR], temperature, respiratory rate [RR] and pulse oximetry), electrocardiogram 
(ECG), hematology, biochemistry, serology, urinalysis, and urine screening for 
drugs of abuse, alcohol test and serum pregnancy test (female subjects only). For a 
complete listing of all tests to be performed, please refer to Appendix B - Clinical 
Laboratory Assessment.  
 


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5.1.2 Treatment Phase (Visit 2 to 14) 
5.1.2.1 Study Period Check-in Procedures (Visit 2, 6, 10 and 14) 
At check-in for each study period, subjects will be questioned about whether they 
have complied with the study restrictions and will be asked about the COVID-19 
related questionnaire. Temperature, pulse oximetry and respiratory rate will be 
measured at each check-in and at pre-dose at each study period. Subjects will be 
asked to come to the study site approximately 24 hours prior to dosing and will be 
discharged after 72-hour blood draw.  
If drug therapy other than that specified in the protocol was used, a decision to 
continue or discontinue the subject’s participation will be made by the PI/Sub-
Investigator and will be reported to the Sponsor. 
Urine tests for drugs of abuse and a breath alcohol test will be performed on all 
subjects at each study period check-in. In addition, urine hCG testing will be 
performed on all female subjects at each study period check-in. 
The subject’s hemoglobin and hematocrit levels will be assessed by the PI/Sub-
Investigator regarding the subject’s continued participation. Male subjects having a 
hemoglobin level below 12.0 g/dL and/or hematocrit level below 36.0% or a female 
subjects having hemoglobin level below 12.0 g/dL and/or hematocrit level below 
34.0%, the subjects will not be permitted to continue in the study unless deemed 
non-clinically significant (NCS) by PI/Sub-Investigator. 
Clinical staff reserves the right to conduct random testing (urine drugs of abuse, 
urine hCG [females only], or alcohol) on any subject at any time during the study to 
ensure subject compliance and/or safety. Any subject with a positive test for urine 
drugs of abuse, alcohol or urine hCG (females only) will be dismissed immediately 
from the study. 
 
5.1.2.2 Study Treatment 
Tacrolimus capsules will be studied using a fully replicate crossover design in 72 
healthy male and non-pregnant, non-lactating female volunteers, who will receive a 
single oral dose of 1 × 1 mg capsule of each tacrolimus under fasting conditions. For 
each treatment, safety assessments and monitoring will be performed for the 
duration of confinement and PK assessments for up to 144 hours after each 
tacrolimus administration, as listed Section 2 – Table of Study Procedures. Adverse 
event(s) (AEs) will be monitored throughout the study. There will be at least a 14-
day washout period between each study period. Based on the half-life of tacrolimus 
capsule, this time-period is adequate to avoid drug carry-over from the preceding 
treatments. Blood samples will be collected at pre-dose (0 hour) and at 0.25, 0.50, 
0.75, 1.00, 1.25, 1.50, 1.75, 2.00, 2.25, 2.50, 2.75, 3.00, 3.50, 4.00, 5.00, 6.00, 8.00, 
10.00, 12.00, 16.00, 24.00, 36.00, 48.00, 72.00, 96.00, 120.00 and 144.00 hours after 
study drug dosing in each study period.  


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5.1.3 Post-Study [End-of-Study (EOS)]/Early Termination 
Clinical laboratory tests (hematology, serum chemistry and urinalysis), as per 
Appendix B – Clinical Laboratory Assessment, will be performed at the end of the 
study or after withdrawal/dismissal of a subject from the study.  
At the end of the study or after withdrawal/dismissal of a subject from the study, a 
symptom-directed physical exam with the possibility for a full physical exam if 
deemed necessary by the PI/Sub-Investigator including vital signs (BP, HR, 
temperature, pulse oximetry and RR) will be conducted unless the subject is lost to 
follow up. 
A follow up phone call will be made about a week ±3 days after study participants 
have been discharged from the study i.e., End-of-study. Clinic will make two 
additional attempts to contact study participants. If all three attempts are 
unsuccessful, clinic will close the file and will be deemed lost to follow up. If the 
first attempted phone call is unsuccessful, the second phone call will be attempted 5 
±3 days from the first phone call. If second attempt to contact study participant via 
phone is unsuccessful, a registered letter will be sent 5 ±3 days from the second 
phone call. 
5.2 
Study Duration and Confinement  
Subjects will be confined to the clinic from at least 12 hours prior to dosing until at least 
72-hours post-dose, for a total of approximately 84 hours in each study period. Subjects 
will be required to return to the clinical facility for the 96, 120 and 144-hour blood draws 
in each study period.  
 
This study will consist of four study periods with a washout period of at least 14 
days between each tacrolimus dosing. The typical, total anticipated duration of the 
clinical part of the study for each subject will be approximately 12 weeks, including 
an approximately 30-day screening period and approximately 50 days for the 
treatment periods with a follow-up phone call. 
5.3 
Randomization and Blinding 
In this study, the assignment of treatment groups (simple randomization scheme) 
will be generated by a computer program designed and run in SAS® Version 9.4
 
  
 
Subjects who meet the eligibility criteria will be randomly assigned equally into one 
of the following two sequence groups: 
 


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Table 5-1 Randomization Scheme 
 
Period 1 
Period 2 
Period 3 
Period 4 
Sequence 1 
T 
R 
T 
R 
Sequence 2 
R 
T 
R 
T 
 
Each subject is scheduled to receive each treatment twice by the end of the study. 
Treatment T (Test): 1 × 1mg tacrolimus capsules 
 
Treatment R (Reference): 1 × 1mg Prograf® 1 mg capsules 
 
 
Approximately 32 subjects are planned to receive treatment sequence T-R-T-R and 
approximately 32 subjects are planned to receive treatment sequence R-T-R-T.  
 
The trial will be performed as an open-label study as the pharmacokinetic profile 
should not be expected to be affected by having the knowledge of whether reference 
or generic tacrolimus capsules were administered. Blinding of the subjects as well 
as clinic staff is not considered necessary. Only the bioanalytical laboratory staff 
will not have access to the randomization scheme until the bioanalytical analysis is 
complete. 
 
6.0 
STUDY POPULATION 
A required number of healthy male and non-pregnant, non-lactating female 
volunteers, as intended for general population [9], aged between 18- 59 years of age, 
inclusive will be screened according to the inclusion and exclusion criteria listed in 
Sections 7.2 and 7.3 to be sure that about 64 are randomized.  
To assure subjects’ safety, eligibility will be assessed at the participating clinical 
site at the time of screening and upon entry into the clinic prior to first drug 
administration.   
 
7.0 SUBJECT SELECTION AND WITHDRAWAL/TERMINATION 
7.1 
Sample Size 
Up to a total of 72 subjects will be enrolled in the study, including the targeted 
sample size of approximately 64 subjects and 8 standbys to ensure the completion 
of at least 48 subjects. Subjects who are dosed will not be replaced. The subjects 
may be dosed in more than one cohort to achieve the total required sample size. 


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7.2 
Inclusion Criteria 
Potential subjects meeting all of the following criteria may be included in the study: 
(medical/operative reports may be requested to support assessment, if necessary): 
1. Males and female volunteers, 18 -59 years of age (inclusive). 
2. BMI that is within 18.5-33.0 kg/m², inclusive. 
3. Healthy, according to medical history, ECG, vital signs, laboratory results 
and physical examination as determined by the PI/Sub-Investigator. 
4. Capable of giving written informed consent prior to receiving any study 
medication.  
5. Availability to volunteer for the entire study duration and willing to adhere 
to all protocol requirements. 
6. Smokers: Capable of refraining from smoking for the duration of the 
confinement. 
7. Female subjects must be non-pregnant, non-lactating and fulfill at least one 
of the following: 
• Be surgically sterile for a minimum of 6 months 
• Post-menopausal for a minimum of 12 months; Menopause defined 
as 12 months of amenorrhea without any other obvious pathological 
or physiological cause. 
• Agree to avoid pregnancy and use medically acceptable method of 
contraception from at least 30 days prior to the study until 30 days 
after the study has ended (last study procedure). 
Medically acceptable methods of contraception include non-hormonal 
intrauterine device or double barrier method (foam or vaginal spermicidal 
suppository in conjunction with a male condom, diaphragm with spermicide 
in conjunction with a male condom). Abstinence as a method of 
contraception is acceptable if it is in line with the preferred and usual lifestyle 
of the study participant. 
 
If a female subject becomes pregnant during participation in the study or 
within 30 days after she has completed her last tacrolimus administration 
(whichever was administered last), she must inform BPSUSA staff 
immediately. 
8. Males who are able to father children must agree to use medically acceptable 
methods of contraception and not to donate sperm during the study and for 
30 days after the end of the study. 
Medically acceptable methods of contraception include using a condom with 
a female partner of child-bearing potential who is using: oral contraceptives, 


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hormonal patch, implant or injection, intrauterine device, or diaphragm with 
spermicide.  
If a male subject’s partner becomes pregnant during his participation in the 
study or within 30 days after he has completed his last tacrolimus 
administration (whichever was administered last), he must inform BPSUSA 
staff immediately. 
7.3 
Exclusion Criteria 
Potential subjects meeting any of the following criteria will be excluded: 
1. Known history and/or presence of any clinically significant hepatic (e.g., 
hepatitis, jaundice, hepatic failure, hepatic necrosis, hepatic encephalopathy, 
biliary tract diseases, cirrhosis), renal/genitourinary (e.g., urethral stricture, 
any renal impairment), gastrointestinal, cardiovascular (e.g., hypotension 
including orthostatic hypotension, cor pulmonale, congenital long QT, 
congestive heart failure, brady arrhythmias), cerebrovascular, pulmonary 
(e.g., chronic obstructive pulmonary disease, decreased respiratory reserve, 
hypoxia, pre-existing respiratory depression), endocrine (e.g., myxedema, 
hypothyroidism, 
adrenal 
cortical 
insufficiency), 
immunological, 
musculoskeletal (e.g., kyphoscoliosis), neurological (e.g., CNS depression 
or coma, increased cerebrospinal pressure), psychiatric (e.g., psychosis, 
depression, hallucinations, delirium tremens, suicidal thoughts or behavior), 
dermatological or hematological (e.g., thrombocytopenic purpura) disease or 
condition unless determined as not clinically significant by the PI/Sub-
Investigator. 
2. History or presence of any clinically significant gastrointestinal pathology 
(e.g., chronic diarrhea, inflammatory bowel disease), unresolved 
gastrointestinal symptoms (e.g., diarrhea, vomiting), or other conditions 
known to interfere with the absorption, distribution, metabolism or excretion 
of the drug experienced within 7 days prior to first tacrolimus administration, 
as determined by the PI/Sub-Investigator.  
3. Systolic blood pressure outside 90-130 mmHg, inclusive, and diastolic blood 
pressure outside 55-80 mmHg, inclusive, and heart rate between 50-100 
bpm, inclusive, unless deemed otherwise by the PI/Sub-Investigator. 
4. QTc interval ≥ 440 milliseconds for males and ≥ 460 milliseconds for 
females, unless deemed otherwise by the PI/Sub-Investigator. 
5. Abnormal clinical laboratory values unless values are deemed by the PI/Sub-
Investigator as “Not Clinically Significant”. 
6. Abnormal vital signs (blood pressure [BP], heart rate [HR], respiratory rate 
[RR], pulse oximetry and temperature) measurements, unless deemed 
otherwise by the PI/Sub-Investigator. 


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7. Presence of any clinically significant illness within 30 days prior to first 
dosing, as determined by the PI/Sub-Investigator. 
8. Presence of any significant physical or organ abnormality as determined by 
the PI/Sub-Investigator. 
9. A positive test result for any of the following: HIV, Hepatitis B surface 
antigen, Hepatitis C, drugs of abuse (marijuana, amphetamines, barbiturates, 
cocaine, opiates, phencyclidine and benzodiazepines), and alcohol test. 
Positive serum or urine pregnancy test for female subjects. 
10. Known history or presence of: 
• Alcohol abuse or dependence within one year prior to first study 
drug administration; 
• Drug abuse or dependence; 
• Hypersensitivity or idiosyncratic reaction to tacrolimus, its 
excipients, and/or related substances; 
• Lymphoma and other malignancies (particularly of the skin); 
• Bacterial, viral, fungal and protozoal infections (e.g., polyoma virus-
associated nephropathy [PVAN] due to BK virus infection, JC virus 
associated progressive multifocal leukoencephalopathy [PML], Epstein 
Barr Virus (EBC) associated Post transplant lymphoproliferative 
disorder [PTLD], cytomegalovirus (CMV) infections associated CMV 
viremia and CMV disease); 
• Tuberculosis; 
• Posterior reversible encephalopathy syndrome (PRES); 
• Pure red cell aplasia (PRCA); 
• Gastrointestinal perforation; 
• Hyperkalemia; 
• Hypertension; 
• Lactose intolerance, galactose intolerance, Lapp lactase deficiency or 
glucose-galactose malabsorption;  
• Food allergies and/or presence of any dietary restrictions unless deemed 
by the PI/Sub-I as “Not Clinically Significant”. 
• Severe allergic reactions (e.g. anaphylactic reactions, angioedema). 
11. History of intolerance to and/or difficulty with blood sampling through 
venipuncture. 


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12. Abnormal diet patterns (for any reason) during the four weeks preceding the 
study, including fasting, high protein diets, vegan, etc. 
13. Individuals who have donated, in the days prior to first tacrolimus 
administration: 
• 50-499 mL of blood in the previous 30 days; 
• 500 mL or more or double red blood cell (“Power red”) donation in 
the previous 56 days. 
14. Donation of plasma by plasmapheresis within 7 days prior to first tacrolimus 
administration. 
15. Hemoglobin level of 12.0 g/dL or lower. 
16. Individuals who have participated in another clinical trial or who received 
an investigational drug within 30 days prior to first tacrolimus 
administration.  
17. Not being able to fast for at least 14 hours. 
18. Consumption of food or beverages containing grapefruit and/or pomelo 
within 3 days prior to first study drug administration. 
19. Consumption of food or beverages containing caffeine/methylxanthines, 
poppy seeds and/or alcohol within 48 hours before dosing in each study 
period. 
20. Use of any prescription medication within 14 days prior to first tacrolimus 
administration (except for allowed contraceptive products). 
21. Use of any over-the-counter medications (including low-dose aspirin, oral 
multivitamins, allergy medications, herbal and/or dietary supplements) 
within 14 days prior to first tacrolimus administration (except for 
spermicidal/barrier contraceptive products). 
22. Use of any enzyme-modifying drugs and/or other products, including 
inhibitors of cytochrome P450 (CYP) enzymes such as antifungals (e.g., 
ketoconazole, itraconazole, fluconazole, voriconazole, posaconazole, 
clotrimazole), macrolide antibiotics (e.g., erythromycin, clarithromycin, 
josamycin), protease inhibitors (e.g., ritonavir, telaprevir [IncivekTM], 
boceprevir [VictrelisTM], nelfinavir [Viracept®], saquinavir), calcium 
channel blockers (e.g., verapamil, diltiazem, nifedipine, nicardipine), 
nucleotide reverse transcriptase inhibitors (e.g., tenofovir), gastric acid 
suppressors/neturalizers (e.g., lansoprazole, omeprazole, cimetidine, 
cisapride, magnesium-aluminum hydroxide antacids), bromocriptine, 
cortisone, dapsone, ergotamine, gestodene, lidocaine, mephenytoin, 
miconazole, midazolam, nilvadipine, quinidine, tamoxifen, and triacetyl-
oleandomycin and inducers of CYP enzymes such as anti-myobacterials 
(e.g., 
rifampin, 
rifabutin), 
antifungal 
agent 
(e.g., 
caspofungin), 
anticonvulsants 
(e.g. 
phenytoin, 
carbamazepine, 
phenobarbital), 


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corticosteroids (e.g. prednisolone or methylprednisolone), metamizole, 
isoniazid and products containing St. John’s Wort in the previous 30 days 
before first study drug administration. 
23. Use of medicines such as, mycophenolic acid (MPA) products, 
aminoglycosides, gyrase inhibitors, vancomycin, cotrimoxazole, NSAIDs, 
oral 
anticoagulants, 
or 
oral 
antidiabetics, 
ganciclovir, 
acyclovir, 
amphotericin B, ibuprofen, cisplatin, potassium supplements, potassium-
sparing diuretics (e.g., amiloride, triamterene or spironolactone), ACE 
inhibitors (for example, benazepril, enalapril, quinapril), statins (for 
example, atorvastatin, fluvastatin, simvastatin), angiotensin receptor 
blockers (for example, irbesartan, valsartan, azilsartan), nefazodone, 
metoclopramide, danazol, and herbal products containing Schisandra 
sphenanthera extracts should be avoided during treatment with tacrolimus 
in the previous 30 days before first study drug administration. 
24. Use of any QT prolonging drugs (e.g., citalopram, chlorpromazine, 
haloperidol, 
methadone), 
amiodarone 
[CordaroneTM, 
NexteroneTM, 
PaceroneTM], immunosuppressive or immunomodulating therapies (e.g. 
sirolimus 
[Rapamune®], 
cyclosporins 
[Gengraf®, 
Neoral®, 
and 
Sandimune®], antilymphocytic antibodies [e.g., basiliximab, daclizumab]) 
should be avoided during treatment with tacrolimus in the previous 30 days 
before first study drug administration. 
25. Individuals having undergone any major surgery within 6 months prior to 
the start of the study, unless deemed otherwise by PI/Sub-Investigator. 
26. Does not agree, to refrain from driving or operating heavy machinery if 
feeling dizzy or drowsy following tacrolimus administration until full mental 
alertness is regained. 
27. Does not have an ability to fast for at least 14 hours. 
28. Recent history (within 8 weeks prior to screening) of travel to or emigration 
from any country with high incidence of tuberculosis. 
29. Temperature at visit is ≥ 100.4 F (38.0°C). 
30. Test positive for SARS-CoV-2 by locally available, verified test. 
31. Have had common symptoms of COVID-19 currently or in the past 14 
days as documented in the COVID-19 screening questionnaire and 
symptom checklist 
32. Have had an exposure to suspected or confirmed cases of COVID-19 in the 
past two weeks as documented in the COVID-19 screening questionnaire 


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33. Recent live attenuated vaccine1 recipients in the past month. 
7.4 
Informed Consent Procedure 
The Investigator will be responsible for ensuring that the informed consent is 
obtained before any protocol specific procedures are carried out.  The decision of a 
subject to participate in clinical research is voluntary and will be based on a clear 
understanding of what is involved.   
Subjects will receive adequate oral and written information. The oral explanation to 
the subject will be performed by the Investigator or a designated person and will 
cover all the elements specified in the informed consent form.   
The subject will be given every opportunity to clarify any points they do not 
understand and, if necessary, to ask for more information.  The subject will be given 
sufficient time to consider the provided information.  It will be emphasized that the 
subjects may withdraw their consent to participate at any time without loss of 
benefits to which they otherwise would be entitled. 
The subjects will be informed that all attempts to maintain confidentiality will be 
made. Their names or personally identifiable information will not be disclosed 
outside the site. The Investigator or delegated member of the trial team and the 
subject will sign and date the Informed Consent Form (ICF) to confirm that consent 
has been obtained.  The subject will receive a copy of this document and the original 
will be filed in the subject binder.  In case of a screen failure, it will be filed 
separately in the designated binder.  
7.5 
Study Restrictions 
If at any time prior to or during the study, any subject does not comply with the 
restrictions mentioned below, the subject’s continued participation will be re-
assessed by the PI/Sub-Investigator, Pharmacokinetic staff, and reported to the 
Sponsor. 
1. No food will be allowed from at least 10 hours before dosing until at least 4 hours 
after dosing. Except for the 8 ± 0.1 fl. oz. water given with study medication, no 
fluid will be allowed from 1 hour before dosing until 1-hour post-dose. Water will 
be allowed/provided ad libitum at all other times. 
2. No smoking will be permitted during confinement.  
3. Subjects will be required not to drive or operate heavy machinery as indicated in 
Section 7.3. 
 
1 Adenovirus Type 4 and Type 7 vaccine (Oral), Cholera vaccine, Ebola Zaire vaccine (Live), Influenza vaccine, 
Measles, Mumps, and Rubella Virus Vaccine (Live), Smallpox (Vaccinia) Vaccine (Live), Smallpox and 
Monkeypox vaccine (Live, Non-replicating), Typhoid Vaccine (Live Oral Ty21a), Yellow Fever Vaccine and 
Zoster Vaccine (Live). 


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4. Subjects will be required to limit exposure to sunlight and UV light by wearing 
protective clothing and using a sunscreen with a high protection factor during 
study participation until the study completion. 
5. Subjects will be required not to receive a vaccination (live attenuated vaccine). 
Examples include (but are not limited to): Adenovirus Type 4 and Type 7 vaccine 
(Oral), Cholera vaccine, Ebola Zaire vaccine (Live), Influenza vaccine, Measles, 
Mumps, and Rubella Virus Vaccine (Live), Smallpox (Vaccinia) Vaccine (Live), 
Smallpox and Monkeypox vaccine (Live, Non-replicating), Typhoid Vaccine 
(Live Oral Ty21a), Yellow Fever Vaccine and Zoster Vaccine (Live) during the 
study and until 30 days after the study has ended (last study procedure). 
6. The consumption of alcohol of any kind (e.g., wine, beer, liquor, cocktails), 
products containing caffeine/methylxanthines (e.g., coffee, tea, chocolate, 
caffeine-containing soft drinks [e.g. Coke, Pepsi, Red Bull]) and poppy seeds 
containing products (poppy seed cake, cookies, bagels) will be prohibited for 48 
hours prior to tacrolimus administration until after the last blood draw in each 
study period. 
7. The consumption of foods and/or beverages containing grapefruit and/or pomelo 
(e.g., grapefruit, grapefruit juice, grapefruit candies, pomelo, etc.) will be 
prohibited for 3 days prior to first tacrolimus administration until the last blood 
draw in the final study period. 
8. Any concomitant medication (prescription medications [e.g., prescription pills, 
topical systemic creams, inhalants, sprays] or over-the-counter such as low-dose 
aspirin, allergy medications and oral multivitamins), any herbal and/or dietary 
supplements will not be permitted for 14 days prior to first drug administration. 
9. Any concomitant medication (prescription medications [e.g., prescription pills, 
topical systemic creams, inhalants, sprays] or over-the-counter), any herbal 
and/or dietary supplements from the start of dosing of the study until the last 
blood draw in the final study period unless deemed otherwise by the PI/Sub-
Investigator for treatment of any adverse events (AEs).  
Any use of concomitant medication or herbal/dietary supplement during the 
restriction period for the restriction # 9 will be reported as soon as possible to the 
PI/Sub-Investigator, Pharmacokinetic staff and reported to the Sponsor. In each 
case, the decision whether to continue or discontinue the subject’s participation 
in the study will be made by the PI/Sub-Investigator and/or by the 
Pharmacokinetic staff and reported to the Sponsor. 
10. Use of any enzyme-modifying drugs and/or other products, including inhibitors 
of cytochrome P450 (CYP) enzymes such as antifungals (e.g., ketoconazole, 
itraconazole, fluconazole, voriconazole, posaconazole, clotrimazole), macrolide 
antibiotics (e.g., erythromycin, clarithromycin, josamycin), protease inhibitors 
(e.g., ritonavir, telaprevir [IncivekTM], boceprevir [VictrelisTM], nelfinavir 
[Viracept®], saquinavir), calcium channel blockers (e.g., verapamil, diltiazem, 
nifedipine, nicardipine), nucleotide reverse transcriptase inhibitors (e.g., 


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tenofovir), gastric acid suppressors/neturalizers (e.g., lansoprazole, omeprazole, 
cimetidine, cisapride, magnesium-aluminum hydroxide antacids), bromocriptine, 
cortisone, dapsone, ergotamine, gestodene, lidocaine, mephenytoin, miconazole, 
midazolam, nilvadipine, quinidine, tamoxifen, and triacetyl-oleandomycin and 
inducers of CYP enzymes such as anti-myobacterials (e.g., rifampin, rifabutin), 
antifungal agent (e.g., caspofungin), anticonvulsants (e.g., phenytoin, 
carbamazepine, 
phenobarbital), 
corticosteroids 
(e.g., 
prednisolone 
or 
methylprednisolone), metamizole, isoniazid and products containing St. John’s 
Wort within 30 days prior to first study drug administration until the last blood 
draw in the final study period. 
11. Use of medicines such as, mycophenolic acid (MPA) products, 
aminoglycosides, gyrase inhibitors, vancomycin, cotrimoxazole, NSAIDs, oral 
anticoagulants, or oral antidiabetics, ganciclovir, acyclovir, amphotericin B, 
ibuprofen, cisplatin, potassium supplements, potassium-sparing diuretics (e.g., 
amiloride, triamterene or spironolactone), ACE inhibitors (for example, 
benazepril, enalapril, quinapril), statins (for example, atorvastatin, fluvastatin, 
simvastatin), angiotensin receptor blockers (for example, irbesartan, valsartan, 
azilsartan), nefazodone, metoclopramide, danazol, and herbal products 
containing Schisandra sphenanthera extracts should be avoided during treatment 
with tacrolimus within 30 days prior to first study drug administration until the 
last blood draw in the final study period. 
12. Use of any QT prolonging drugs (e.g., citalopram, chlorpromazine, haloperidol, 
methadone), 
amiodarone 
[CordaroneTM, 
NexteroneTM, 
PaceroneTM], 
immunosuppressive 
or 
immunomodulating 
therapies 
(e.g., 
sirolimus 
[Rapamune®], 
ciclosporins 
[Gengraf®, 
Neoral®, 
and 
Sandimune®], 
antilymphocytic antibodies [e.g., basiliximab, daclizumab]) should be avoided 
during treatment with tacrolimus within 30 days prior to first study drug 
administration until the last blood draw in the final study period. 
13. Physical activity: Subjects will remain awake for the first 4 hours following 
tacrolimus administration. If a medical event (i.e., AE) occurs, subjects may be 
placed in an appropriate position at that time. Subjects will be required to abstain 
from strenuous/vigorous activities for the duration of the study period(s). When 
the subjects leave the clinic, they will be requested to refrain from strenuous 
physical activity until the completion of the 144-hour blood draw in each study 
period. 
14. For 3 hour after each study drug administration, subject visits to the washroom 
will be monitored and recorded by clinic staff. Subjects will be restricted from 
flushing the toilet to allow staff to inspect the content during this time. 
15. Additional study restriction for COVID-19 prevention and control: 
• Subjects must be willing to social distance, wear masks and practice hand 
hygiene while enrolled in the study 


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• Report to study staff in case of exposures to known or possible COVID-19 
positive cases during the study 
7.6 
Withdrawal and Dismissal 
A subject is free to withdraw from participation in the study at any time, for any 
reason. An Investigator may terminate a study subject’s participation in the study if: 
• Any clinical adverse event (AE), laboratory abnormality, or other medical 
condition or situation occurs such that continued participation in the study 
would not be in the best interest of the subject. 
• The subject meets an exclusion criterion (either newly developed or not 
previously recognized) that precludes further study participation. 
The cause, date and time of withdrawal or termination will be documented in the 
source documents and in the final study report. If a subject’s participation is 
terminated prematurely, the cause for the early termination date and time of the 
termination will be documented on the source documents and in the final study 
report. 
Subjects experiencing emesis within 3 hours (2 times median Tmax, 1.5 hour, reported 
in the label [6]) after dosing will be dismissed from the affected period in the study. 
Subjects experiencing 3 or more episodes of loose stool (diarrhea) after dosing will 
be allowed to stay in the study unless the PI/Sub-Investigator recommends the 
subject to be dismissed from the study due to safety reasons. All decisions will be 
made prior to the bioanalytical laboratory commencing bioanalysis. 
 
Subjects who are dismissed or withdrawn from Period 1, may be eligible to return 
for Period 2 and/or Period 3 and/or Period 4, if deemed safe and appropriate by the 
PI/Sub-Investigator. All subjects will be reviewed by the PI and the sponsor prior to 
re-entry. Subjects who are dismissed due to a Treatment-Emergent Adverse Event 
(TEAE) or non-compliance will not be eligible to return for Period 2 and/or Period 
3 and/or Period 4. 
 
Withdrawn and dismissed subjects who are returning for subsequent periods are 
required to adhere to the study specific procedures (e.g., study restrictions). If a 
subject is withdrawn or dismissed following tacrolimus administration and will not 
be returning in subsequent periods (i.e., for Period 2 and/or Period 3 and/or Period 
4), these subjects are still required to adhere to the study restrictions with regards to 
the safety, prescription medication, over-the-counter medication, dietary and/or 
herbal supplements for the expected duration of the restriction relevant to the study 
period the subject withdrew or was dismissed from (where that period becomes the 
subject’s last study period).  
 
Female subjects will be asked to adhere to the requirements of not becoming 
pregnant and using a medically acceptable method of contraception for 30 days after 


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discontinuation from the study (if applicable). Male subjects who are able to father 
children will be asked to adhere to the requirements of using medically acceptable 
methods of contraception for 30 days after discontinuation in the study. 
This study may be suspended or prematurely terminated if there is sufficient 
reasonable cause.  Written notification, documenting the reason for study suspension 
or termination, will be provided by the suspending or terminating party to the 
investigator, sponsor and regulatory authorities. If the study is prematurely 
terminated or suspended, the principal investigator will promptly inform the IRB 
and will provide the reason(s) for the termination or suspension.  
Circumstances that may warrant termination include, but are not limited to: 
• Determination of unexpected, significant, or unacceptable risk to subjects. 
• Insufficient adherence to protocol requirements. 
• Data that are not sufficiently complete and/or evaluable.  
• Determination of futility. 
Subjects whose participation in the study is discontinued (for any reason) will not 
be replaced. If a subject withdraws or is dismissed from the study, a post-study 
symptom-directed physical exam with the possibility for a full physical exam if 
deemed necessary by the PI/Sub-Investigator will be conducted and post-study 
testing will be completed, where possible. 
 
8.0 
STUDY PROCEDURES 
All study assessments will be performed at the visits and time points outlined in the 
Table of Study Procedures (Section 2.0). 
8.1 
 Demographics and Other Baseline Characteristics 
8.1.1 Demographics 
Demographics including age (date of birth and years), sex, race, and ethnicity will 
be recorded at screening. 
8.1.2 Medical History 
The complete medical history will include history of acute, chronic, or infectious 
diseases; surgical or oncologic histories; and any reported conditions affecting major 
body systems. All findings on medical history will be evaluated by the PI/Sub-
Investigator for clinical significance.  
8.1.3 Medication History 
All medications (prescription and non-prescription/OTC, herbal medications/natural 
health products, or investigational drugs) taken by the subjects during the 30 days 
prior to Screening will be recorded in the source documentation as medication 
history. 


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8.1.4 Clinical Laboratory Assessments 
Clinical laboratory assessments will be conducted at Screening, each period check-
in and at the EOS. All laboratory safety data will be reviewed by the PI/Sub-
Investigator for clinical significance. Additional laboratory samples may be taken at 
the discretion of the PI/Sub-Investigator (e.g., if the results of any tests fall outside 
reference ranges or clinical symptoms necessitate testing to ensure safety). 
 
Blood and urine samples collected for clinical laboratory assessments will be 
processed and shipped according to the research site’s SOPs and instructions from 
the clinical laboratory.  
 
For a complete listing of all tests to be performed, please refer to Appendix B - 
Clinical Laboratory Assessment.  
8.2 
Food and Fluid Intake 
Following at least a 10-hour overnight fasting period, subjects will be dosed with 
tacrolimus and will be required to continue to fast for at least 4 hours after dosing. 
 
Following the fasting period of at least 4-hours after dosing, subjects will be given 
standardized meals and caffeine/methylxanthine-free beverages at approximately 
4.5, 9.5, 13.5, 24.5, 28.5, 33.5, 37.5, 48.5, 52.5, 57.5 and 61.5 hours after dosing. 
Meals and beverages during confinement will be identical between each study 
period. 
Except for water given with study medication, no fluid will be allowed from 1 hour 
before dosing until 1-hour post-dose. Water will be allowed/provided ad libitum at 
all other times. 
8.3 
Dosing 
After a fast of at least 10 hours, subjects will take their assigned formulation with 8 
± 0.1 fl. oz. of ambient temperature water at their scheduled time point in each study 
period 
Subjects will be instructed not to touch, chew, bite, or break tacrolimus capsules. If 
a subject chews, bites, or breaks the capsules, that subject will be removed from the 
study. If the subject touches the capsules a replacement dose will be administered. 
Subjects must swallow the capsules and finish the water within 60 seconds.  
 
Clinical staff will conduct a hand and mouth check immediately following dosing. 
8.4 
Safety Monitoring 
In the interest of subjects’ safety, the following safety parameters will be assessed 
in all subjects. 


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8.5 
COVID-19 prevention and control 
Additional procedures will be performed to prevent, and control spread of COVID-
19 among the study subjects 
8.5.1 COVID-19 questionnaire 
The COVID-19 questionnaire for each subject will be completed by a clinic staff 
to document all history of exposure and history of symptoms related to COVID-19. 
The COVID-19 questionnaire is provided in the site’s COVID-19 related 
procedural SOP. 
8.5.2 COVID-19 symptom check 
The medical staff will check for any common symptoms for COVID-19 at screening, 
at each check-in, discharge at each study period, at return visits, and at EOS. 
Additional symptom checks might be performed during the study as deemed 
necessary by the PI or study staff. The COVID-19 symptom checklist is provided in 
the site’s COVID-19 related procedural SOP. 
8.5.3 Temperature check 
Temperature will be measured using an oral thermometer at screening, each check-
in, at 6- and 12-hours post dose, then at every 12 hours until discharge at each study 
period, at return visits and at EOS. Additional temperature checks might be 
performed during the study as deemed necessary by the PI or study staff. 
8.5.4 Pulse Oximetry and Respiratory Rate 
Pulse oximetry and respiratory rate will be performed at screening, each check-in, 
at pre-dose, at 6- and 12-hours post dose, then at every 12 hours until discharge at 
each study period, at return visits and at EOS. Additional pulse oximetry and 
respirator rate measurements might be taken during the study as deemed necessary 
by the PI or study staff. The normal ranges of pulse oximetry and respiratory rate 
are: 
- 
a pulse oximeter reading of:  ≥ 95% 
- 
a range of resting respiratory rate as 12 to 20 per minute 
 
8.5.5 SARS-CoV-2 Test 
If applicable, a nasal, oral swab or blood might be collected from each subject at 
each check-in for detection of SARS-CoV-2. Additional tests might be performed 
during the study as deemed necessary by the PI or study staff. The swabs or blood 
will be sent to a local CLIA certified laboratory for analysis using a verified, locally 
acceptable test. The sample collection and shipment will follow a specific procedure 


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provided by the laboratory.as described. The subjects who test positive for SARS-
CoV-2 will be excluded from any subsequent study procedures. 
8.6 
Vital Signs 
Vital signs (BP and HR) will be monitored at pre-dose and at 1, 2, 4, 6, 12-,24-, 48- 
and 72-hours post-dose in each study period. 
Pre-Dose and Post-Dose (all periods): 
Acceptable Range - systolic blood pressure between 90-130 mmHg, diastolic blood 
pressure between 55-80 mmHg, inclusive, and heart rate between 50-100 bpm, 
inclusive. 
 
If the Investigator is not immediately available on site to assess out of range vital 
signs, the vital signs will be repeated up to two times. If vital signs are still outside 
of the acceptable range, the PI/Sub-Investigator will be informed, and PI/Sub-
Investigator will determine the appropriate course of action. 
 
Additional vital signs measurements will be taken if deemed necessary by the 
PI/Sub-Investigator. Blood draws will take precedence over vital signs 
measurements and other scheduled activities if the scheduled time of several 
activities is overlapped to each other unless deemed necessary by the PI/Sub-
Investigator or due to subject’s safety. 
8.7 
ECG 
A 12-lead ECG with rhythm strip will be performed at screening, Additional 
measurements are not required during this study unless deemed necessary by the 
PI/Sub-Investigator. 
8.8 
Health Monitoring 
The PI/Sub-Investigator will be present approximately 30 minutes prior to dosing 
until at least 4 hours after the last subject is dosed in each study period. The PI/Sub-
Investigator will remain on-call throughout the duration of the study. 
Health monitoring will be conducted throughout the study or as needed. Adverse 
events will be monitored throughout the study. 
8.9 
Sample Collection and Processing 
Blood will be obtained by direct venipuncture in the arm. Blood sample collection 
times will be recorded on the appropriate source documents and reported for each 
subject.  
 


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Number of Samples 
28 samples from 28 time points in each study 
period. 
Total Volume of Blood including blood 
samples for PK time points, pre- and post- study 
procedures and the selected laboratory testing 
at each study period. 
Approximately 520 mL of blood 
Type of Vacutainer 
Pre-chilled K2EDTA Vacutainer®, 4 mL 
Blood Sampling Time Points2,3 
Pre-dose (0 hour) and at 0.25, 0.50, 0.75, 1.00, 
1.25, 1.50, 1.75, 2.00, 2.25, 2.50, 2.75, 3.00, 
3.50, 4.00, 5.00, 6.00, 8.00, 10.00, 12.00, 
16.000, 24.000, 36.00, 48.00, 72.00, 96.00, 
120.00 and 144.00 hours post study in each 
study period. 
The actual blood sample collection time will be 
documented. For all blood sample time points, 
any sample collected outside of the scheduled 
time will not be considered as protocol 
deviation. 
 
Whole blood will be aspirated and aliquoted into 2 pre-chilled clear polypropylene 
tubes. A minimum of 2 mL whole blood will be transferred to the first tube and the 
remaining whole blood (if any) will be aliquoted into a second tube. Polypropylene 
tubes will be pre-chilled and pre-labeled with at least the following information: 
Time-point, Protocol Number, Study Period, Aliquot Number, Matrix and Subject 
Number. The samples will be stored at -20ºC (-15ºC or colder) in a freezer pending 
shipment. 
Throughout sample collection, the samples will be maintained in an ice-bath until 
stored in the freezer. 
8.10 Sample Shipment 
One set of aliquots from all subjects will be delivered to the bioanalytical facility 
packed on dry ice. Once the initial set has been confirmed to have been received by 
the bioanalytical facility, the second set of aliquots will then be shipped/delivered. 
All shipments will be accompanied by an inventory list and delivered to the 
bioanalytical facility:  
                  
 
2 - Pre-dose samples to be obtained within 2 hours prior to study drug administration. 
3 - The blood sample will be collected within ± 1-hour window for the 96, 120 and 144-hours time 
points.  


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Clinic personnel will notify the bioanalytical laboratory prior to shipment by phone, 
fax or e-mail. The bioanalytical laboratory will notify BPSUSA by either e-mail or 
fax of confirmation of receipt of each aliquot. 
 
9.0 
DRUG PRODUCTS 
9.1 
Drug Information  
Table 9-1 Study Drugs Information 
 
T (Test) 
R (Reference) 
Drug Name: 
Tacrolimus 
Prograf® 
Strength: 
1 mg 
1 mg 
Dosage Form: 
Capsules 
Capsules  
Manufactured 
by/for: 
 
 
Dose: 
1 × 1 mg 
1 × 1 mg 
 
9.2 
Labeling, Maintenance, and Retention  
Each investigational study drug will be labeled (in English) with a statement 
indicating that the drug is an investigational drug to be used only by a Qualified 
Investigator and will include but is not limited to: Drug Name, Strength, Protocol 
Number, Sponsor’s Name and Address, the recommended storage conditions for the 
drug, Expiry/Retest Date (when available), and Lot/Batch Number. 
9.3 
Drug Inventory 
The BPSUSA will supply sufficient quantities of the study drugs for the following: 
(1) completion of this study and (2) retention, as per applicable regulations. All drug 
supplies provided for this study will be stored in a secure area with restricted access, 
under controlled storage conditions described in the product package labelling, 
unless otherwise instructed per protocol. 
Records will be made of receipt and dispensing of tacrolimus supplied.  


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9.4 
Disposition of Unused Tacrolimus 
Upon completion or termination of the study, all remaining study supplies will be 
retained according to applicable regulations. Once the retention period has elapsed, 
any remaining unused drug will be returned to the Sponsor in the original containers, 
or destroyed, as directed in writing by the Sponsor. 
 
10.0 POTENTIAL RISKS AND BENEFITS  
10.1.1 Potential Risks   
In clinical studies, the most common adverse reactions (≥ 15%) were abnormal renal 
function, hypertension, diabetes mellitus, fever, CMV infection, tremor, 
hyperglycemia, leukopenia, infection, anemia, bronchitis, pericardial effusion, 
urinary tract infection, constipation, diarrhea, headache, abdominal pain, insomnia, 
paresthesia, peripheral edema, nausea, hyperkalemia, hypomagnesemia, and 
hyperlipemia. [6]. 
 
The following serious adverse reactions were also reported in the patients who were 
tacrolimus users [6]: 
 
• Lymphoma and Other Malignancies 
• Serious Infections 
• New Onset Diabetes After Transplant 
• Nephrotoxicity 
• Neurotoxicity 
• Hyperkalemia 
• Hypertension 
• QT Prolongation 
• Myocardial Hypertrophy 
• Immunizations 
• Pure Red Cell Aplasia 
 
COVID-19 related risks 
BPSUSA is taking significant steps to minimize subject exposure to COVID-19. 
However, risk of exposure of the subjects to COVID-19 due to close contact with 
study staff or other study subjects does exist during the conduct of the study. 
10.1.2 Potential Benefits 
While it may not provide direct benefit to subjects, the knowledge that may result 
from the study may help with the development of generic formulations of tacrolimus 
and may enhance the regulatory review standards for equivalence of this category.  


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Protocol Version Number: 3.0, Date: Jun 22, 2021 
Tacrolimus Capsule 
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Page 42 of 68 
11.0 SAFETY ASSESSMENT 
BioPharma Services USA Inc. (BPSUSA) ensure the timely, accurate and complete 
reporting of safety information in conformity with regulatory requirements. 
PI/Sub-Investigators at BPSUSA are responsible for monitoring the safety and for 
providing appropriate medical care for subjects who have entered this trial (i.e., from 
the signing of ICF onwards). In addition, the PI/Sub-Investigator remains 
responsible for following-up all AEs.  
Each subject will be carefully questioned and/or examined by the PI/Sub-
Investigator or a medically qualified delegate (i.e., authorized by the Investigator, in 
a separate form, to record AEs) to obtain information regarding AEs. All AEs will 
be reported and documented as stated below. 
 
Any symptoms or positive tests related to COVID-19 will be documented as part of 
AEs reporting. This documentation applies from the time that volunteers receive the 
study drugs until the completion of the end-of-study procedures. 
11.1 Definitions 
Adverse Event (AE) – is any untoward medical occurrence in clinical trial subject 
who are administered with any of study drugs and which does not necessarily have 
a causal relationship with this treatment. An adverse event can therefore be any 
unfavorable and unintended sign (e.g. an abnormal laboratory finding), symptom, 
or disease temporally associated with the use of a study drug, whether or not 
considered related to the study drug.  
 
Adverse Drug Reaction (ADR) of study drug any of study drugs is any untoward 
and unintended response to any of study drugs related to any dose administered. 
 
Unexpected Adverse (Drug) Reaction – is an adverse reaction, the nature or 
severity of which is not consistent with the applicable product information (e.g., 
Prescribing Information for an authorized product). 
 
Serious Adverse Event (SAE) / Reaction – a serious adverse event (experience) or 
reaction is any untoward medical occurrence or effect that at any dose can: 


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Page 43 of 68 
• result in death, 
• be life-threatening, 
• require hospitalization or prolongation of existing inpatient hospitalization, 
• result in persistent or significant disability or incapacity, 
• cause a congenital anomaly/birth defect, 
• be other medically important event that may not be immediately life-
threatening or result in death or hospitalization but may jeopardize the subject 
or may require intervention to prevent one of the above-listed outcomes. 
Suspected Unexpected Serious Adverse Reaction (SUSAR) – is all suspected 
adverse reactions related to Prograf® and generic tacrolimus capsules which can 
occur in the concerned trial, and that are both unexpected and serious. 
Unanticipated Problems – The unanticipated problems involving risks to subjects 
or others to include, in general, any incident, experience, or outcome that meets all 
of the following criteria: 
• unexpected in terms of nature, severity, or frequency given (a) the research 
procedures that are described in the protocol-related documents, such as the 
IRB-approved research protocol and informed consent document; and (b) the 
characteristics of the subject population being studied; 
• related or possibly related to participation in the research (“possibly related” 
means there is a reasonable possibility that the incident, experience, or 
outcome may have been caused by the procedures involved in the research); 
and 
• suggests that the research places subjects or others at a greater risk of harm 
(including physical, psychological, economic, or social harm) than was 
previously known or recognized. 
Subjects will be instructed to inform clinic personnel of any untoward medical 
symptoms and/or events that may arise during the study. 
11.1.1 Treatment-emergent adverse events (TEAEs) 
Treatment emergent adverse events (TEAEs) are undesirable events, not present 
prior to medical treatment, or an already present event that worsens either in 
intensity or frequency following the treatment. A TEAE is an adverse event that 
occurs only once treatment has started. All TEAEs and all treatment emergent SAEs 
will be summarized. 
 
The PI/sub-I will be responsible for determining whether a TEAE is possibly related 
to the SARS-CoV-2 and document accordingly. 


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Protocol Version Number: 3.0, Date: Jun 22, 2021 
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Page 44 of 68 
11.2 Characteristics of an Adverse Event 
11.2.1 Relationship to Study Drug 
The PI/Sub-Investigator will assess the relationship of all adverse reactions to the 
study drug, using the following scale: 
 
Probable 
A clinical event, including laboratory test abnormality, with a 
reasonable time sequence to drug administration, unlikely to be 
attributed to concurrent disease or other drugs or chemicals, and 
which follows a clinically reasonable response on withdrawal. 
Possible 
A clinical event, including laboratory test abnormality, with a 
reasonable time sequence to drug administration, but which could 
also be explained by concurrent disease or other drugs or chemicals. 
Information on drug withdrawal may be lacking or unclear. 
Unlikely 
A clinical event, including laboratory test abnormality, with a 
temporal relationship to drug administration which makes a causal 
relationship improbable, and which other drugs, chemicals or 
underlying disease provide plausible explanation. 
Unrelated 
This category is applicable to AEs which are judged to be clearly and 
incontrovertibly due to extraneous causes (diseases, environment, 
etc.) and do not meet the criteria for drug relationship listed for the 
above-mentioned conditions. 
 
All AEs will be evaluated by the PI/Sub-Investigator, who must approve the subject 
for subsequent dosing. 
Any AEs, whether serious or non-serious, will be monitored throughout the study 
and followed to resolution, when possible, regardless of whether the subject is still 
participating in the study. 
 
Adverse Event monitoring and reporting will be followed-up until resolution.  If 
resolution cannot be achieved within 1 week of study completion, the PI/Sub-
investigator will decide the course of action. 
11.2.2 Expectedness of SAE 
The Study PI will be responsible for determining whether an SAE is expected or 
unexpected as per drug labels. An adverse event will be considered unexpected if 
the nature, severity, or frequency of the event is not consistent with the risk 
information previously described in the Investigator’s Brochure, Product 
Monograph, Product Label, or on the label of the drug.   


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11.2.3 Severity Assessment 
The term "severe" describes the intensity (severity) of a specific event; the event 
itself, however, may be of relatively minor medical significance (such as severe 
headache). This means it is not the same as "serious", which is based on 
patient/event outcome or action criteria usually associated with events that pose a 
threat to a patient's life or functioning 
The severity of all AEs will be graded by the PI/Sub-Investigator or a medical 
qualified delegate according to the following definitions: 
Every effort will be made to obtain an adequate evaluation of the severity. 
11.3 Adverse Events and Serious Adverse Events Reporting  
An Adverse Event as defined above generally includes any condition that was: 
1) Not present prior to administration of study drugs but appeared after 
initiation of administration of study drug; or 
2) Present prior to administration of study drugs but worsened after 
administration of study drug; or   
3) Reported as part of the subject's history, and while not present immediately 
prior to initiating administration of study drugs, reappeared after 
administration of study drug.  
 
During the study, an AE can also occur outside the time that the investigational 
product(s) was given (e.g., during a washout period).  
 
Pre-existing conditions, diseases, or disorders are not considered AEs unless there 
is a change in intensity, frequency, or quality. Planned hospital admissions and/or 
surgical operations/procedures for an illness or disease that existed before the 
investigational product was given or the subject was enrolled in a clinical trial are 
not to be considered AEs. 
 
Clinical laboratory data collected during the course of the study, which exceeds or 
drops below the acceptable limits for the subject population and which, based on 
baseline values, are considered by the Investigator to be clinically significant, will 
be reported as an AE. If clinically significant abnormal ECGs and/or laboratory 
Mild 
Adverse event resulting in discomfort, but not sufficient to cause 
interference in normal daily activities. 
Moderate 
Adverse event resulting in discomfort that is sufficient to cause 
interference in daily activities.  
Severe 
Adverse event resulting in discomfort causing an inability to carry out 
normal daily activities.  


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Protocol Version Number: 3.0, Date: Jun 22, 2021 
Tacrolimus Capsule 
Fasting Conditions 
 
Page 46 of 68 
values lead to, or are associated with clinical symptom(s), the diagnosis should be 
reported as an AE. 
If a subject discontinues from the study because of an AE, study site personnel must 
clearly document the circumstances and data leading to the reason for 
discontinuation. 
The investigator/designee is responsible for recording all AEs which have occurred 
during the study (including clinically important deviations of laboratory values from 
normal ranges), regardless of their relationship to the study drugs. This includes AEs 
spontaneously reported by the patient, observed by the investigator/designee or 
elicited by general questioning. All AEs will be determined as Serious or Non-
Serious.  
 
Subjects will be instructed to inform clinic personnel of AEs that may arise during 
the study.  
 
Treatment of any AEs will be administered under the direction of a physician, either 
at BPSUSA or at a nearby hospital emergency room. First-aid treatment will be 
provided on site, but if additional treatment is required, subjects will be transferred 
to a hospital. 
 
All symptoms will be recorded by clinic staff and will be reviewed by the PI/Sub-
Investigator prior to any subsequent dosing. When appropriate, medical tests and 
examinations will be performed to document resolution of the event(s).  
All reported adverse events will be documented in the Clinical Report. 
The Investigator will monitor the subject's condition until recovery to a satisfactory 
state or stabilization. Thus, follow-up visits may be required even after the 
administration of the study drugs has been discontinued.   
 
All SAE(s), whether or not the event is deemed study drug-related, will be reported 
to the Sponsor by telephone and email within 24 hours of BPSUSA being aware of 
SAE, followed by a written report within five business days.  
Reports of all SAEs must be communicated as soon as possible to the appropriate 
Institutional Review Board (IRB) or Independent Ethics Committee (IEC) and/or 
reported in accordance with local laws and regulations. 
Adverse events will be coded into the Preferred Term (PT), classified according to 
the Medical Dictionary for Regulatory Activities (MedDRA) with System Organ 
Classification (SOC) and reported with severity, duration, onset time and 
relationship to study drugs and action taken. 
BPSUSA will be responsible for notifying the local IRB. The Sponsor will be 
responsible for notifying the regulatory agencies, if applicable. BPSUSA will also 
report SAEs to FDA Adverse Event Reporting System (FAERS) per their 
procedures. The Principal Investigator will report to the local IRB and to the FDA 


BPSUSA Protocol Number: 2521 
Protocol Version Number: 3.0, Date: Jun 22, 2021 
Tacrolimus Capsule 
Fasting Conditions 
 
Page 47 of 68 
Sponsor any unanticipated problems involving risk to human subjects or others 45 
CFR 46.103(b)(5)(i). Certain adverse events must be reported to the IRB within a 
specified period of time after the discovery of the event. The Office for Human 
Research Protections guidance defines these adverse events as including, but not 
limited to the following:  
− An adverse event that is not expected, i.e., not listed in the informed consent 
document or the investigator’s brochure;  
− An expected adverse event that occurs at a greater frequency or duration than 
expected;  
− Any adverse event that would require modification of the protocol and/or 
informed consent document. 
All SAEs need to be emailed with SAE in a subject line to: 
 
 
  
 
  
 
11.4 Procedures for Reporting Pregnancy 
A positive urine pregnancy test or any hCG level ≥ 5 IU/L for females of child-
bearing potential and ≥ 8 IU/L for post-menopausal females will be assessed as 
clinically significant and will result in the immediate dismissal of the subject from 
the study, unless deemed otherwise by the Investigator. The elevated hCG will be 
investigated as a potential pregnancy and will be followed up in the same manner as 
an AE. A confirmatory hCG blood test will be required for any elevated values. If 
pregnancy is confirmed, any out of range hCG levels will not be considered an 
adverse event. 
Once notified that pregnancy has occurred during the course of the study or within 
30 days of study completion, clinic personnel will report the pregnancy within 24 
hours of the confirmatory hCG blood test, to the PI/Sub-Investigator, the IRB, and 
appropriate Sponsor representative. A Pregnancy Notification Form will be 
completed as per BPSUSA internal Standard Operating Procedures and will be 
included alongside any pertinent information at the time of pregnancy reporting. The 
confirmed pregnancy will be followed to birth of the child or elective/spontaneous 
termination of the pregnancy. Follow up will occur at minimum once every three 
months (i.e., once per trimester) until the end of the pregnancy (birth, termination, 
etc.).  
Elective termination will not be considered an adverse event, however supporting 
documentation detailing the outcome of the procedure will be required. Spontaneous 


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Protocol Version Number: 3.0, Date: Jun 22, 2021 
Tacrolimus Capsule 
Fasting Conditions 
 
Page 48 of 68 
termination of the pregnancy will be considered a Serious Adverse Event and will 
be followed up accordingly (as per Section 11.3). Pregnancy will be treated as AEs 
for this study. 
 
12.0 BIOANALYTICAL ANALYSIS 
12.1 Analytical Procedures 
Data management, quality review and reporting of study data pertaining to 
laboratory analysis of study data will be the responsibility of the bioanalytical 
facility.  
12.1.1 Samples to be Assayed 
Sample from all subjects who complete at least 1 study period will be analyzed for 
drug concentration. 
12.1.2 Analyte(s) in Biological Matrix 
Whole blood samples will be assayed for tacrolimus using a validated analytical 
method according to the principles of Good Laboratory Practice.  
12.1.3 Incurred sample reproducibility 
Incurred Sample Reanalysis (ISR) shows repeatability and stability of an analytical 
method in real sample matrix.  Approximately 10% of the samples will be 
reanalyzed in case the number of samples is less than 1000 samples.  If the number 
of samples is greater than 1000, 10% of the first 1000 samples and 5% of the 
remaining samples will be reanalyzed. The ISR test will contain samples from all 
analyzed subjects.  For each subject, at least two samples should be chosen, one at 
or near Cmax and the other one in the elimination phase.  
 
If an ISR sample result is not valid for analytical reasons (e.g. low area of internal 
standard response, sample processing error, instrument failure etc.), this sample will 
be excluded from the statistical evaluation of the ISR test and will not be reanalyzed.  
Acceptance criterion of the ISR test: the concentration obtained for the initial 
analysis and the concentration obtained by reanalysis should be within 20% of their 
mean for at least 67% of the repeats.  If acceptance criterion of the ISR test is not 
met, an investigation will be conducted to determine the reason for differences, and 
adequate steps will be taken to minimize inaccuracy (and imprecision).  
 
The ISR test results will be presented in the Bioanalytical Report; they will not be 
used for any correction of the concentration results for pharmacokinetic and 
statistical evaluation of the study. 
 


BPSUSA Protocol Number: 2521 
Protocol Version Number: 3.0, Date: Jun 22, 2021 
Tacrolimus Capsule 
Fasting Conditions 
 
Page 49 of 68 
13.0 PHARMACOKINETIC AND STATISTICAL ANALYSIS 
13.1 Pharmacokinetic and Statistical Analysis Data Set 
The data from at least one of the following subjects will be included in the PK and 
statistical analysis: 
1. Subjects who complete all study periods. 
2. Subjects who have missed samples but for whom it has been predicted prior 
to the start of bioanalytical analysis that reliable estimates of the PK 
parameters should be possible. 
3. Subjects who drop out from the study but have completed at least 2 periods. 
If a subject’s pre-dose concentration is less than or equal to 5% of the Cmax value for 
that subject in the given period, then the subject’s data in that period without any 
adjustments can be included in all PK measurements and calculations. If the pre-
dose value is greater than 5% of the Cmax, data from that subject in that period will 
be dropped from the PK and statistical analysis. Data for subjects dropped due to 
higher than 5% of Cmax pre-dose concentrations will be included in a separate 
appendix in the final study report. 
For subjects who are dismissed from the study due to non-compliance or any reason 
other than AEs,  other data of non-impacted period(s) for that subject, which fulfills 
the PK analysis data set requirements, as above, will be included in PK stats analysis. 
Data does not fulfill the requirements will be presented separately.  
If a subject is dismissed/withdrawn due to an AE from a study period, the data from 
that period will not be included in the PK stats analysis and will be presented 
separately. All other data for that subject, which fulfills the PK analysis data set 
requirement, as above, will be included in PK stats analysis. Data does not fulfill the 
requirements will be presented separately. 
Data from other subjects who were dismissed or withdrew due to elevated hCG 
levels or are confirmed to be pregnant will not be included in the PK and statistical 
analysis. The data for these subjects will be presented separately. 
 
Any decision for excluding data prior to start of sample bioanalysis (e.g Major 
protocol deviation) will be provided with a detailed explanation and will be properly 
recorded and dated. 
13.2 Analysis of Pharmacokinetic Data  
The PK and statistical analysis will be performed at BPSI using SAS® Version 9.4. 
The PK and/or statistical analyses outlined in this protocol may be altered with 
appropriate justification. 


BPSUSA Protocol Number: 2521 
Protocol Version Number: 3.0, Date: Jun 22, 2021 
Tacrolimus Capsule 
Fasting Conditions 
 
Page 50 of 68 
13.2.1 Pharmacokinetic Analysis  
Pharmacokinetic parameters will be calculated using non-compartmental analysis 
(NCA) method. The following PK parameters will be estimated (where possible) for 
tacrolimus and included in the PK and statistical analysis: 
Primary endpoint 
AUC72:  
Area under the concentration-time curve from time zero 
until time point 72 hours post dose. AUC72 is estimated 
using the linear trapezoidal method. 
Cmax: 
The maximal observed whole blood concentration. 
 
Secondary endpoint 
AUCinf: 
Area under the concentration-time curve from time zero to 
infinity, calculated as AUCt + Clast/λ, where Clast is the last 
measurable concentration. 
AUCt: 
Area under the concentration-time curve from time zero until the 
last measurable concentration or last sampling time t, whichever 
occurs first. AUCt is estimated using the linear trapezoidal 
method. 
T1/2: 
Terminal elimination half-life, estimated as ln(2)/λ. 
Tmax: 
Time when the maximal whole blood concentration is observed. 
λ: 
Terminal elimination rate constant, estimated by linear regression 
analysis of the terminal portion of the ln-concentration vs. time 
plot. 
 
During PK and statistical analyses, drug concentrations below the lower limit of 
quantitation (BLQ) of an assay will be considered as zero except when they occur 
between two non-BLQ concentrations where they will be considered as missing 
during PK calculations and estimations. 
The actual time of blood samples collection will be used for pharmacokinetic and 
statistical analysis. 
Missed samples and non-reportable concentrations (e.g. quantity not sufficient) 
from the analytical laboratory will be treated in the pharmacokinetic analysis as if 
they have not been scheduled for collection. 
For subjects with missing or non-reportable tacrolimus concentration at time point 
72-hour or the sampling time deviation at 72-hours is greater than 2 hours, the 
AUC72 will not be calculated. If concentration(s) at 72 hours or at the time point(s) 
immediately preceding 72 hours is/are BLQ(s), this/these BLQ value(s) will be set 
as zero and the calculated AUC will be treated as AUC72. 


BPSUSA Protocol Number: 2521 
Protocol Version Number: 3.0, Date: Jun 22, 2021 
Tacrolimus Capsule 
Fasting Conditions 
 
Page 51 of 68 
The λ, T1/2, and AUCinf parameters will not be estimated for whole blood 
concentration-time profiles where the terminal linear phase is not clearly defined. 
For subjects with missing or non-reportable tacrolimus concentrations for three or 
more of the last samples, only the Cmax and Tmax will be presented and included in 
the statistical analysis. Other PK parameters will not be reported. The AUC72 could 
also be estimated for these subjects if data allows.  
13.2.2 Statistical Analysis of Pharmacokinetic Data    
Step 1: Determine the Within-subject Standard Deviation of the Test and 
Reference Products ( 𝑺𝑺𝑾𝑾𝑾𝑾 and 𝑺𝑺𝑾𝑾𝑾𝑾 ) for the PK Parameters AUC72, AUCt, 
AUCinf and Cmax:  
The within-subject standard deviation of the test and reference products will be 
determined for the ln-transformed PK parameters AUC72, AUCt, AUCinf and Cmax for 
tacrolimus.  
For a fully replicated 4-way design with 2 sequences TRTR, RTRT, the within 
subject variance of the reference product,           can be determined as follows: 
 
Where, 
 Number of sequences (2 used in the study), 
Number of subjects within each sequence, 
  (where
 represents the first administered (early) reference and 
 represents the second administered (late) reference.) 
 (Mean within sequence .) 
 (total number of subjects used in the study, while 
 is the number of 
subjects used in sequence .) 
 
 will be calculated as follows: 
 
 
The
 calculation will include subjects with estimated PK parameters for both 
reference administrations. 
)
2
(
2
)
(
2
1
2
1
.
2
−
−
=
∑∑
=
=
n
D
D
S
i
n
j
i
ij
WR
i
=
i
=
j
2
1
ij
ij
ij
R
R
D
−
=
1
ij
R
2
ij
R
i
n
j
ij
i
n
D
D
i∑
=
=
1
.
i
∑
=
=
2
1
i
in
n
in
i
WR
S
2
WR
WR
S
S
=
WR
S
2
WR
S


BPSUSA Protocol Number: 2521 
Protocol Version Number: 3.0, Date: Jun 22, 2021 
Tacrolimus Capsule 
Fasting Conditions 
 
Page 52 of 68 
For a fully replicated 4-way design with 2 sequences TRTR, RTRT, the above 
method can be applied for the calculation of the within subject variance of the test 
product 𝑆𝑆𝑊𝑊𝑊𝑊 as well. 
Step 2: Reference Scaled Average Bioequivalence Procedure 
The 95% upper confidence bound for
 will be determined for the 
ln-transformed PK parameters AUC72, AUCt, AUCinf and Cmax using a PROC 
MIXED model in SAS®, where 
 and 
are the means of the ln-transformed PK 
parameters obtained from the BE study for the test and reference products, 
respectively. 
 is the scaled average bioequivalence limit defined as 𝜃𝜃=
ቀ
ln (1 0.9
⁄
)
𝜎𝜎𝑤𝑤0
ቁ
2
, where 
 is a regulatory constant which is set at 0.10. 
 
The method of obtaining the upper confidence bound is based on Howe’s 
Approximation [8]. The following steps will be followed to construct the 95% upper 
confidence bound. 
1. Let 
, 
, 
 and 
be the log transformed observations on early test, 
late test, early reference and late reference for subject 
 within sequence i. 
Then we will calculate the following two terms:  
i. 
, and 
ii. 
. 
 
2. Fit the Generalized linear mixed model (GLMM) with 
values as dependent 
variable.  Sequence is the explanatory variable of the model.  The confidence 
interval for the least squares mean of 
  will be estimated at 10% significance 
level. Let 
 and 
be the lower and upper bounds of the confidence interval. 
Also the least squares mean of 
 will be calculated by determining the average 
of the 
 sequence means, using the ESTIMATE function of GLMM. Suppose 
 is the estimated least squares mean of 
 and 
 is the standard error of 
this estimate.  
 
Calculate the following estimates: 
 
 
3. Fit the GLMM with 
values as the dependent variable.  Sequence is the 
explanatory variable of the model.  The within subject variance of the reference 
product,
, will be calculated as, 
2
2)
(
WR
R
T
S
θ
µ
µ
−
−
T
µ
R
µ
θ
0
w
σ
1
ij
T
2
ij
T
1
ij
R
2
ij
R
j
2
2
2
1
2
1
ij
ij
ij
ij
ij
R
R
T
T
I
+
−
+
=
2
1
ij
ij
ij
R
R
D
−
=
ijI
I
L
U
I
2
Φ
I
)
(Φ
SE
2
2
1
)
(Φ
−
Φ
=
∆
SE
ij
D
2
WR
S
2
1
(abs(U))))
L)),
(max((abs(
=
B


BPSUSA Protocol Number: 2521 
Protocol Version Number: 3.0, Date: Jun 22, 2021 
Tacrolimus Capsule 
Fasting Conditions 
 
Page 53 of 68 
 
Where MSE is the mean square error from the model and let 
be the 
corresponding degrees of freedom. 
4. Now the 95% upper confidence bound for 
 will be calculated 
as follows: 
 
 
Where, 
 
 
 is the chi-square tabulated values at 5% significance level. 
 
Step 3: Unscaled Average Bioequivalence Procedure 
The unscaled average bioequivalence analysis was performed using PROC MIXED 
in SAS@, with SEQUENCE, PERIOD, and TREATMENT as fixed effects, 
TREATMENT and SUBJECT as random effects on the logarithm of the 
pharmacokinetic parameters (AUC72, AUCt, AUCinf or Cmax).  
 
The following illustrates the program statements to run the ANOVA using PROC 
MIXED in SAS, with SEQ, SUBJ, PER, and TRT identifying sequence, subject, 
period, and treatment variables, respectively, and Y denoting the response measure 
(e.g.,ln(AUC72), ln(AUCt), ln( AUCinf), ln(Cmax)) being analyzed: 
PROC MIXED; 
 CLASSES SEQ SUBJ PER TRT; 
 MODEL Y = SEQ PERTRT / DDFM=SATTERTH; 
 RANDOM TRT/TYPE=FA0(2) SUB=SUBJ G; 
 REPEATED/GRP=TRT SUB=SUBJ; 
 ESTIMATE 'T vs. R' TRT 1 -1/CL ALPHA=0.1; 
 
All the fixed effects will be tested using the Type 3 Tests of Fixed Effects in the 
ANOVA to detect statistically significant differences (α=0.05). 
The REPEATED statement in PROC MIXED is used to specify covariance 
structures for repeated measurements on subjects. Least squares means for the 
treatments and the adjusted difference between treatment means together with the 
associated standard error (σD) will be estimated in this procedure. 
2
2
MSE
S WR =
κ
2
2)
(
WR
R
T
S
θ
µ
µ
−
−
2
2
2
2
1
1
2
1
)
(
)
(
)
(
∆
−
+
∆
−
+
∆
+
∆
B
B
2
2
WR
S
θ
−
=
∆
2
95
.0
,
2
2
κ
χ
κ
∆
=
B
2
95
.0
,
κ
χ


BPSUSA Protocol Number: 2521 
Protocol Version Number: 3.0, Date: Jun 22, 2021 
Tacrolimus Capsule 
Fasting Conditions 
 
Page 54 of 68 
The two one-sided hypotheses [9] at the alpha level (α=0.05) of significance will be 
tested by constructing the 90% confidence interval for the ratio between the test and 
reference means. 
The same ANOVA using the linear mixed-effects model will be performed on the 
raw data for Tmax, Lambda (or λ), and T1/2 parameters. 
 
Step 4: Calculate the 90% CI of the Ratio of the Within-subject Standard 
Deviation of Test Product to Reference Product (𝝈𝝈𝑾𝑾𝑾𝑾𝝈𝝈𝑾𝑾𝑾𝑾
⁄
). 
 
The 90% CI for 𝜎𝜎𝑊𝑊𝑊𝑊𝜎𝜎𝑊𝑊𝑊𝑊
⁄
 is given by 
 
ቆ𝑆𝑆𝑊𝑊𝑊𝑊𝑆𝑆𝑊𝑊𝑊𝑊
⁄
𝐹𝐹𝛼𝛼2
ൗ(𝑣𝑣1, 𝑣𝑣2) ,
𝑆𝑆𝑊𝑊𝑊𝑊𝑆𝑆𝑊𝑊𝑊𝑊
⁄
𝐹𝐹1−𝛼𝛼2
ൗ(𝑣𝑣1, 𝑣𝑣2) ቇ 
where 
• 𝑆𝑆𝑊𝑊𝑊𝑊 is the estimate of 𝜎𝜎𝑊𝑊𝑊𝑊 with 𝑣𝑣1 as the degree of freedom; 
• 𝑆𝑆𝑊𝑊𝑊𝑊 is the estimate of 𝜎𝜎𝑊𝑊𝑊𝑊 with 𝑣𝑣2 as the degree of freedom; 
• 𝐹𝐹𝛼𝛼2
ൗ(𝑣𝑣1, 𝑣𝑣2)  is the value of the F-distribution with 𝑣𝑣1 (numerator) and 
𝑣𝑣2 (denominator) degrees of freedom that has the probability of 𝛼𝛼2
ൗ to its right 
• 𝐹𝐹1−𝛼𝛼2
ൗ(𝑣𝑣1, 𝑣𝑣2)  is the value of the F-distribution with 𝑣𝑣1 (numerator) and 
𝑣𝑣2 (denominator) degrees of freedom that has the probability of 1 −𝛼𝛼2
ൗ to its 
right 
• here 𝛼𝛼= 0.1. 
 
Individual concentration at each scheduled time point and individual PK parameters 
will be listed by treatment. Two SAS transport files including xconc.xpt for 
individual concentration at each scheduled time point and xpara.xpt for individual 
PK parameters, as well as a Define.pdf file to describe each SAS transport file will 
be provided. 
 
Descriptive statistics (including number of subjects (N), arithmetic mean, standard 
deviation (SD), minimum, maximum, median and coefficient of variation (CV%)) 
of concentrations will be presented by treatment at each scheduled time point. 
 
Descriptive statistics (including N, arithmetic mean, SD, minimum, maximum, 
median and CV%) of all PK parameters will be presented by treatment. 
 
The mean and individual concentration-time profiles will be plotted in linear and 
semi-logarithmic scales. 
 
If the study is dosed in two or more groups of subjects, the statistical model will be 
modified to reflect the multi-group nature of the study. 
Additional statistical and alternate tests will be performed if necessary. 


BPSUSA Protocol Number: 2521 
Protocol Version Number: 3.0, Date: Jun 22, 2021 
Tacrolimus Capsule 
Fasting Conditions 
 
Page 55 of 68 
 
Further details for statistical analysis of PK data will be described in the Statistical 
Analysis Plan (SAP). 
13.3 Safety Analysis 
13.3.1 Safety Data Set 
Safety data set is defined as all subjects dosed in this study. In a case where SARS-
CoV-2 related TEAEs lead to the withdrawal or dismissal of approximately 5 % or 
more of the dosed subjects, a subgroup analysis related to Adverse Events (AE) may 
be performed. And the safety listings and tables for AE with subgroup and the total 
safety population will both be presented.  
 
13.3.2 Safety Listings 
The safety analysis data set for Adverse Events (AE), demographic characteristics, 
vital signs and clinical laboratory will be listed by subject or by subject and treatment. 
 
A complete listing of AE or (Serious Adverse Events) SAE (if applicable) including 
period, actual treatment received, primary System Organ Class (SOC) and Preferred 
Term (PT), reported term, AE severity, AE causality, start/end date and time, 
duration, action taken etc., will be presented by subject. 
 
A complete listing of demographic characteristics including age, race, gender, 
ethnicity, BMI, weight and height will be presented by subject. 
 
Vital signs parameters including Systolic Blood Pressure (SBP), Diastolic Blood 
Pressure (DBP), Heart Rate (HR), Respiratory Rate (RR), temperature and oxygen 
saturation (if applicable) will be presented by subject. 
 
A complete listing of clinical laboratory measurements (including hematology, 
serum chemistry, urinalysis, serology, breath alcohol test, urine tests for drugs of 
abuse, urine HCG test (for females), serum HCG test (for females), magnesium, 
creatinine kinase and lipid panel will be presented by subject. 
13.3.3 Safety Tables 
The safety data set for AE, demographic characteristics and vital signs will be 
summarized in frequency or using descriptive statistics (including N, arithmetic 
mean, SD, minimum, maximum and median) of quantitative measures by treatment, 
as appropriate.  
 


BPSUSA Protocol Number: 2521 
Protocol Version Number: 3.0, Date: Jun 22, 2021 
Tacrolimus Capsule 
Fasting Conditions 
 
Page 56 of 68 
A brief overall summary of AEs including number of AEs/TEAEs (Treatment 
Emergent Adverse Event) /SAEs and number and percentage of subjects experienced 
at least one AEs/TEAEs/SAEs will be presented by treatment and the overall 
population. TEAEs/SAEs will also be summarized using the number and percentage 
of subjects experiencing at least one TEAE/SAE (with the number of subjects who 
received study treatment as the denominator of percentage) by primary SOC, PT, AE 
severity and AE causality, for each treatment and the overall population.  
 
Descriptive statistics of age, BMI, weight and height and frequencies for race, gender 
and ethnicity will be provided by treatment and the overall population. 
 
Descriptive statistics of each vital sign’s parameters will be presented by treatment 
at each visit. 
 
Descriptive statistics of all quantitative measurements from all clinical laboratory 
parameters will be presented by the type of laboratory test, laboratory parameter, 
and visit. 
 
Further details for safety analysis will be described in the Statistical Analysis Plan 
(SAP). 
13.4 Bioequivalence Criteria [7, 10] 
To establish bioequivalence, all the following conditions for individual PK 
parameter(s) AUC72, and Cmax must be met: 
• The 90% CI for the Test/Reference ratios of geometric means are completely 
contained within 80.00%-125.00%.  
• The 95% upper confidence bound for 
 should be less than 
or equal to 0. 
• The upper limit of the 90% confidence interval for the Test/Reference ratio of 
the within-subject standard deviation (𝝈𝝈𝑾𝑾𝑾𝑾𝝈𝝈𝑾𝑾𝑾𝑾
⁄
) is less than or equal to 2.5. 
 
2
2)
(
WR
R
T
S
θ
µ
µ
−
−


BPSUSA Protocol Number: 2521 
Protocol Version Number: 3.0, Date: Jun 22, 2021 
Tacrolimus Capsule 
Fasting Conditions 
 
Page 57 of 68 
14.0 ETHICAL CONSIDERATIONS / PROTECTION OF HUMAN SUBJECTS 
14.1 Basic Principles 
This research will be carried out in accordance with Good Clinical Practice (GCP) 
as set out by the International Conference on Harmonization (ICH), the basic 
principles defined in the U.S. Code of Federal Regulations (21 CFR Part 312), the 
Belmont Report, Directive 2001/20/EC (Europe), and the principles enunciated in 
the most recent version of the World Medical Association Declaration of Helsinki. 
14.2 Institutional Review Board 
The local IRB will be the Institutional Review Board (IRB) for this study. This 
Protocol and the ICF will be reviewed and approved by the IRB prior to the initiation 
of the study. Termination / suspension of any prior approval / favorable opinion can 
be done by IEC (Institutional Ethics Committee)/IRB. 
The board is constituted and operates in accordance with Division 5 of the Food and 
Drug Regulations, ICH Harmonized Tripartite Guideline (GCP Consolidated 
Guideline), and 21 CFR Parts 56 and 50. 
14.3 Informed Consent Form 
Subjects will sign a study ICF prior to start of any study related procedures. Each 
subject will be provided with verbal and written information, in non-technical terms, 
which will describe the nature of the screening procedures as well as conduct of the 
study. Prior to signing the ICF, subjects will be allowed adequate time to consider 
the potential benefits and risks associated with their participation in the study. 
Signed and dated ICF will be retained with the study records and a copy will be 
provided to the subject. 
14.4 Confidentiality 
The information in this study protocol is confidential. This document contains trade 
secrets and commercial information that is confidential and may not be disclosed to 
third parties (except to the IRB, FDA and/or relevant regulatory agencies). These 
restrictions will apply as well to all future communications if deemed privileged or 
confidential. 
Sponsor, study monitors, auditors, the IRB, FDA and applicable regulatory 
authorities will be granted direct access to the subject’s original medical records for 
verification of clinical trial procedure and/or data, without violating the 
confidentiality of the subject, to the extent permitted by applicable laws and 
regulations and that, by signing a written informed consent form, the subject or the 
subjects legally acceptable representative is authorizing such access. 
All documentation collected by BPSI/BPSUSA Inc. will be kept confidential. The 
name and identity of the subjects will remain confidential. If documents containing 


BPSUSA Protocol Number: 2521 
Protocol Version Number: 3.0, Date: Jun 22, 2021 
Tacrolimus Capsule 
Fasting Conditions 
 
Page 58 of 68 
the subjects’ names are photocopied, the name will be omitted from the photocopied 
version. 
The Certificate of Confidentiality (CoC), issued by the FDA in accordance with new 
requirements under the 21st Century Cures Act, will be filed and maintained in the 
regulatory binder. As per the CoC, researchers cannot disclose a subject’s name or 
other information that could identify them in any civil, criminal, administrative, 
legislative or other proceedings (such as a court trial), without their consent. Also, 
information collected for this research that could identify a subject cannot be used 
as evidence in a legal proceeding without their consent.  
In addition, with the CoC, researchers involved in this study generally may not 
provide a subject’s name, or any other information that could identify them, to 
anyone who is not connected with the research. However, in the following situations, 
the CoC does not prevent the researchers involved in this study from disclosing 
study information that could identify a subject: 
• If the subject consents to someone receiving their information from this study, 
including situations where the information is necessary for their medical 
treatment 
• When the subject’s study information is used for other scientific research, as 
allowed by federal regulations protecting research subjects 
• When information is needed by the FDA, which is funding this study, in order 
to audit or evaluate federally-funded studies 
• When a law otherwise requires disclosure [such as requirements to 1) make 
certain reports to FDA, 2) report threats of harm to self/others, 3) report 
incidents of child abuse, or 4) report cases of contagious disease, such as HIV, 
to the state as mandated by state law]. This does not apply to disclosure in a 
legal proceeding.  
The CoC does not prevent a subject from voluntarily providing information about 
themselves or their involvement in this research study to others.  
The CoC will not be used to prevent disclosure for any purpose the subject has 
consented to in this ICF, such as sharing their participation and research data with 
their family doctor (if the subject has agreed to share this information). 
14.5 Compensation for Participation 
 
 
 
 
 
 
 
: 


BPSUSA Protocol Number: 2521 
Protocol Version Number: 3.0, Date: Jun 22, 2021 
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Fasting Conditions 
 
Page 59 of 68 
 
 
 
 
 
 
 
 
 
 
 
 
 
 
 
 
 
 
 
 
 
 
 


BPSUSA Protocol Number: 2521 
Protocol Version Number: 3.0, Date: Jun 22, 2021 
Tacrolimus Capsule 
Fasting Conditions 
 
Page 60 of 68 
 
 
 
 
 
 
 
 
 
 
 
 
 
  
 
15.0 ADMINISTRATIVE CONSIDERATIONS 
15.1 Revisions and/or Amendments to the Protocol 
Revisions and/or amendments to the protocol must be documented and approved by 
the Principal Investigator and discussed with sponsor prior to finalization. If the 
revision/amendment will affect subject safety and/or study design, the amendment 
will be re-submitted to the IRB for approval. Administrative changes (i.e., change 
of analytical facility, typographical errors, discrepancies, clarifications) will also be 
submitted to the IRB, but may not require approval. A copy of the IRB’s approval 
or acknowledgement documents will be included in the final report. 
 
For revisions or amendments to the protocol that substantially alter the study design 
after initiation of the study, a signed revised informed consent form will be needed 
for continued participation by the subject. 
15.2 Investigator Responsibilities 
The Principal Investigator is responsible for ensuring that the clinical study is 
performed in accordance with the protocol, the current revision of the Declaration 
of Helsinki, ICH - GCP guidelines and applicable regulatory requirements. 
15.3 Study Completion/Termination 
BioPharma Services USA Inc. and/or the Sponsor reserve the right to terminate the 
study at any time for any reason related to the safety of subjects or due to any other 
non-study specific reason. 


BPSUSA Protocol Number: 2521 
Protocol Version Number: 3.0, Date: Jun 22, 2021 
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Fasting Conditions 
 
Page 61 of 68 
15.4 Sponsor Visits 
The Sponsor is encouraged to visit BioPharma Services USA Inc., if desired, and at 
their convenience. The PI and staff will provide, if requested, all source documents 
and/or other study-related documents. The PI will maintain regular written and 
telephone communication with the Sponsor. 
 
16.0 DATA MANAGEMENT/RECORD KEEPING 
16.1 Source Data 
All data will be recorded in accordance with GCP to ensure accuracy, completeness, 
legibility, and timeliness of the data reported. 
 
All data will be recorded directly on the source documents and will be considered 
source data. Subjects will be identified using an assigned ID number and/or 
randomization number on source documents. 
16.2 Quality of Data 
All source documents and laboratory reports will be Quality Control reviewed to 
ensure accuracy and completeness. Adverse events will be reviewed and assessed 
for severity and causality by the PI/Sub-Investigator. Specific processes of the study, 
its source documentation and any reports (if applicable) will be audited by the 
Quality Assurance (QA) unit of BPSI/BPSUSA. 
16.3 Retention of Documents 
All records and documents pertaining to the study will be retained by BioPharma 
Services USA Inc. for at least 25 years from the completion of the study and will be 
available for inspection by the Sponsor and/or Regulatory Agencies. 
 


BPSUSA Protocol Number: 2521 
Protocol Version Number: 3.0, Date: Jun 22, 2021 
Tacrolimus Capsule 
Fasting Conditions 
 
Page 62 of 68 
17.0 REFERENCES 
1. Zhang W et al., Recent Topics on The Mechanisms of Immunosuppressive 
Therapy-Related, Int J Mol Sci. 2019, 20(13):3210.  
2. Gaynor J et al., Lower tacrolimus trough levels are associated with 
subsequently higher acute rejection risk during the first 12 months after kidney 
transplantation. Transplant International, 2016, 29: 216  
3. Sikma M et al., Association of Whole Blood Tacrolimus Concentrations with 
Kidney Injury in Heart Transplantation Patients, Eur J Drug Metab 
Pharmacokinet. 2018; 43(3): 311. 
4. Hitesh S. Purohit et al., Investigating the Impact of Drug Crystallinity in 
Amorphous Tacrolimus Capsules on Pharmacokinetics and Bioequivalence 
Using Discriminatory In Vitro Dissolution Testing and Physiologically Based 
Pharmacokinetic Modeling and Simulation, J Pharm Sci. 2018, 107(5):1330 
5. Hitesh S. Purohit et al., Assessing the Impact of Endogenously Derived 
Crystalline Drug on the in Vivo Performance of Amorphous Formulations. Mol. 
Pharmaceutics 2019, 16 (8): 3617 
6. PROGRAF
® (tacrolimus) capsules, for oral use, [label]. 
 
 
7. FDA draft guidance on Tacrolimus capsule (revised on Dec 2012) 
https://www.accessdata.fda.gov/drugsatfda_docs/psg/Tacrolimus_cap_50708_
RV12-12.pdf 
8. W. G. Howe (1974), Approximate Confidence Limits on the Mean of X+Y 
Where X and Y are Two Tabled Independent Random Variables, Journal of 
the American Statistical Association, 69 (347): 789 – 794. 
9. Schuirmann DJ.  A comparison of the two one-sided tests procedure and the 
power approach for assessing the equivalence of average bioavailability.  J. 
Pharmacokinet BioPharm 1987; 15:657-80. 
10. FDA draft guidance on Warfarin Sodium tablet (revised on Dec 2012) 
https://www.accessdata.fda.gov/drugsatfda_docs/psg/Warfarin_Sodium_tab_0
9218_RC12-12.pdf 
 


BPSUSA Protocol Number: 2521 
Protocol Version Number: 3.0, Date: Jun 22, 2021 
Tacrolimus Capsule 
Fasting Conditions 
 
Page 63 of 68 
APPENDIX A – SCHEDULE OF EVENTS 
 
Screening 
Check
-in 
Period 1, 2, 3 and 4 
Return Visit 
EOS 
Study Day 
-30 to 0 
 
-1 
 
1 
 
2 
 
3  
 
4 
 
5 
 
6 
 
7  
Last visit of 
Period 4 
Confinement 
 
X 
X 
X 
X 
X# 
 
 
 
 
ICF 
X 
 
 
 
 
 
 
 
 
 
Demographics 
X 
 
 
 
 
 
 
 
 
 
Medical/Medication History 
X 
 
 
 
 
 
 
 
 
 
Physical Exam 
X 
 
 
 
 
 
 
 
 
X 
Body weight 
X 
 
 
 
 
 
 
 
 
 
Height 
X 
 
 
 
 
 
 
 
 
 
BMI 
X 
 
 
 
 
 
 
 
 
 
Vital Signs (BP & HR) 
X 
X 
Xa 
Xb 
Xc 
Xd 
 
 
 
X 
Safety measures for 
COVID-19# (Symptom 
Check, Temperature, Pulse 
Oximetry and RR) 
 
X 
 
 
Xe 
 
Xe 
 
  Xe 
 
 
Xe 
 
 
Xe 
 
 
 
X 
 
 
X 
 
 
X 
 
SARS-CoV-2 Test (If 
applicable) 
 
X 
 
 
 
 
 
 
 
 
Urine Drug Tests 
X 
X 
 
 
 
 
 
 
 
 
Alcohol Test 
X 
X 
 
 
 
 
 
 
 
 
Pregnancy Test 
X* 
X* 
 
 
 
 
 
 
 
 
12-Lead ECG 
X 
 
 
 
 
 
 
 
 
 
Hematology 
X 
X 
 
 
 
 
 
 
 
X 
Serum Chemistry 
X 
Xf 
 
 
 
 
 
 
 
X 
Urinalysis 
X 
X 
 
 
 
 
 
 
 
X 
Serology  
(HIV, Hepatitis B and C) 
Magnesium 
Creatinine Kinase 
Lipid panel  
X 
 
 
 
 
 
 
 
 
 
Inclusion/Exclusion criteria 
X 
Xg 
 
 
 
 
 
 
 
 
Compliance check 
 
X 
 
 
 
 
X 
X 
X 
 
Randomization 
 
 
X 
 
 
 
 
 
 
 
Tacrolimus Capsule 
Administration 
 
 
X 
 
 
 
 
 
 
 
PK Sampling 
 
 
Xh 
Xh 
Xh 
Xh 
Xh 
Xh 
Xh 
 
Questioning subjects if use of 
any concomitant medication 
X 
X 
 
 
 
 
X 
X 
X 
 
Adverse Events 
 
 
X 
X 
X 
X 
X 
X 
X 
 
Meals 
 
X 
X 
X 
X 
X 
 
 
 
 
* Serum pregnancy test at screening and urine pregnancy test at each period check-in. 
# Study exit at 72 hours post-dose tacrolimus 
a: 1, 2, 4, 6- and 12-hours post-dose  
b: 24hours post-dose 
c: 48 hours post-dose 
d: 72 hours post-dose 
e: At pre-dose, at 6- and 12-hours post-dose and then at every 12 hours until discharge at each study period. 
f: At check-in period 1 and 3. 


BPSUSA Protocol Number: 2521 
Protocol Version Number: 3.0, Date: Jun 22, 2021 
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Fasting Conditions 
 
Page 64 of 68 
g: Only at check-in for the period 1 
h: Pre-dose (0 hour) and at 0.25, 0.50, 0.75, 1.00, 1.25, 1.50, 1.75, 2.00, 2.25, 2.50, 2.75, 3.00, 3.50, 4.00, 5.00, 6.00, 8.00, 10.00, 
12.00, 16.000, 24.000, 36.00, 48.00, 72.00, 96.00, 120.00 and 144.00 hours after study drug dosing in each study period 


BPSUSA Protocol Number: 2521 
Protocol Version Number: 3.0, Date: Jun 22, 2021 
Tacrolimus Capsule 
Fasting Conditions 
 
Page 65 of 68 
APPENDIX B – CLINICAL LABORATORY ASSESSMENT 
TYPE OF 
TEST 
COMPONENTS 
Hematology4 
• Hemoglobin 
• Hematocrit 
• RBC 
• Platelet count  
• WBC and automated differential  
Serum5 
Chemistry 
• Glucose 
• Calcium 
• Sodium 
• Chloride 
• Bicarbonate 
(Total CO2) 
 
• Albumin 
• Protein 
• Bilirubin 
 
 
• AST 
• ALT 
• Potassium 
• Alkaline 
Phosphatase 
• BUN 
• Urea 
• Uric Acid 
• Creatinine 
Urinalysis 
• Bilirubin 
• Blood  
• Glucose 
• pH  
• Ketones 
• Leukocytes 
• Nitrites 
• Protein 
• Specific Gravity 
Additional 
Tests 
Serology (HIV, Hepatitis B surface 
antigen, Hepatitis C antibody)6 
Magnesium6 
Creatine Kinase6 
Lipid panel6 
Alcohol test 
Serum hCG (only females, at Screening) 
Urine hCG (only females, at each period 
check-in) 
Urine Tests 
for Drugs of 
Abuse  
Marijuana, Amphetamines, Phencyclidine, Barbiturates, Cocaine, Opiates, 
Benzodiazepines 
 
 
 
 
 
 
4 At Screening, at each check-in and at EOS 
5 At screening, at check-in for period 1 and 3 and at EOS 
6 At screening only 


BPSUSA Protocol Number: 2521 
Protocol Version Number: 3.0, Date: Jun 22, 2021 
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Fasting Conditions 
 
Page 66 of 68 
APPENDIX C – LIST OF COMMON ABBREVIATIONS 
AE: 
Adverse Event 
LSMEANS: 
Least Square Means 
ALT: 
Alanine Aminotransferase 
MedDRA: 
Medical Dictionary for Regulatory Activities 
ANOVA 
Analysis of Variance 
mg: 
Milligram 
AST: 
Aspartate Aminotransferase 
mL: 
Milliliters  
AUC72 
Area under the concentration-time curve 
from time zero until time point 72 hours 
post dose. AUC72 is estimated using the 
linear trapezoidal method. 
mmHg: 
Millimeter of Mercury 
AUCt: 
Area under the concentration-time curve 
from time zero until the last measurable 
concentration or last sampling time t, 
whichever occurs first. AUCt is 
estimated using the trapezoidal method 
N/A or NA: 
Not applicable, Not Available 
AUCinf: 
Area under the concentration-time curve 
from time zero to infinity, calculated as 
AUCt + Clast/λ, where Clast is the last 
measurable concentration 
PI: 
Principal Investigator 
AUCτ,ss: 
Area under the concentration-time curve 
from time 0 to τ (the length of the dosing 
interval) at steady state. AUCτ is 
estimated using the trapezoidal method. 
PK: 
Pharmacokinetic 
BLQ  
Below the Limit of Quantitation / Below 
Quantitation Limit 
PT: 
Preferred Term 
BMI: 
Body Mass Index (kg/m2) 
QA: 
Quality Assurance 
BP: 
Blood Pressure 
RBC: 
Red Blood Cell 
BPM: 
Beats Per Minute 
RPM: 
Revolutions Per Minute 
BPSI: 
BioPharma Services Inc. 
RR: 
Respiratory Rate 
BPSUSA: 
BioPharma Services USA Inc. 
RLD: 
Reference Listed Drug 
cGMP 
Current Good Manufacturing Practice 
SAE: 
Serious Adverse Event 
CI: 
Confidence Interval 
SAS®: 
Statistical Analysis System 
Cmax: 
The maximal observed-whole blood 
concentration. 
SD or STD: 
Standard Deviation 
CoC 
Certificate of Confidentiality 
SOC: 
System Organ Classification 
CYP: 
Cytochrome P450 
SOP: 
Standard Operating Procedure 
ECG: 
Electrocardiogram 
Sub-I 
Sub-Investigator 
FDA: 
Food and Drug Administration 
T1/2: 
Terminal elimination half-life, estimated as 
ln(2)/λ 
FSH: 
Follicle-Stimulating Hormone 
Tmax / tmax: 
Time when the maximal-whole blood 
concentration is observed 
GCP: 
Good Clinical Practice 
WBC: 
White Blood Cell 
GLP: 
Good Laboratory Practice 
 
 
hCG: 
Human Chorionic Gonadotropin 
 
 
HIV: 
Human Immunodeficiency Virus 
 
 
ICF: 
Informed Consent Form 
 
 
ICH: 
International Conference on 
Harmonization 
 
 
IEC 
Institutional Ethic Committee 
 
 
IRB: 
Institutional Review Board 
 
 


BPSUSA Protocol Number: 2521 
Protocol Version Number: 3.0, Date: Jun 22, 2021 
Tacrolimus Capsule 
Fasting Conditions 
 
Page 67 of 68 
Lambda / λ: 
Terminal elimination rate constant, 
estimated by linear regression analysis of 
the terminal portion of the ln-
concentration vs. time plot 
 
 
kg/m2: 
Kilogram/Meter Squared 
 
 
 
 
 
 
 
 
 
 
 
 
 
 
 
 
 
 
 
 
 
 
 
 
 
 
 
 
 
 
 
 
 
 
 
 
 
 
 
 
 
 


BPSUSA Protocol Number: 2521 
Protocol Version Number: 3.0, Date: Jun 22, 2021 
Tacrolimus Capsule 
Fasting Conditions 
 
Page 68 of 68 
APPENDIX D – PROGRAF® LABEL 
(See attached)