BIOEQUIVALENCE STUDY OF ABIRATERONE ACETATE TABLET IN ...

CERTIFICATE OF APPROVAL The foregoing thesis entitled ““BIOEQUIVALENCE STUDY OF ABIRATERONE ACETATE TABLET IN INDIAN HEALTHY ADULT MALE VOLUNTEERS” is hereby approved as a creditable study of research topic and has been presented in satisfactory manner to warrant its acceptance as prerequisite to the degree for which it has been submitted. It is understood that by this approval, the undersigned do not necessarily endorse any conclusion drawn or opinion expressed therein, but approve the thesis for the purpose for which it is submitted.

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Internal Examiner

External Examiner

____________________________ Dr. B. N. Sinha Professor & Head Department of Pharmaceutical Sciences Birla Institute of Technology Mesra, Ranchi, Jharkhand – 835215, India

DECLARATION CERTIFICATE This is to certify that the work presented in the thesis entitled “BIOEQUIVALENCE STUDY OF ABIRATERONE ACETATE TABLET IN INDIAN HEALTHY ADULT MALE VOLUNTEERS” in fulfilment of the requirement for the award of degree of Master of Pharmacy in Clinical Research of Birla Institute of Technology Mesra, Ranchi is an authentic work carried out under our supervision and guidance. To the best of my knowledge, the content of the thesis does not form a basis for the award of any previous degree to anyone else.

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Internal Guide (Dr. Sugato Banerjee) Assistant Professor Department of Pharmaceutical Sciences Birla Institute of Technology Mesra, Ranchi, Jharkhand – 835215, India

External Guide (Dr. Rajeev M. Patil) Head – Biostudies Norwich Clinical Services Pvt. Ltd. 43/2, 3rd floor, Cauvery Medical Center, NH-7, Bellary road, Sahakar Nagar, Bangalore – 560092, Karnataka, India

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Dr. B. N. Sinha Professor & Head Department of Pharmaceutical Sciences Birla Institute of Technology Mesra, Ranchi, Jharkhand – 835215, India

Dean (Post Graduate Studies) Birla Institute of Technology Mesra, Ranchi, Jharkhand – 835215, India

ACKNOWLEDGEMENT Gratitude is one of the least articulate of emotions especially when it is deep. Words are not enough to express my gratitude towards the people who stood behind me during my project work. The highest appreciation is not to utter words but to live by them. I will be indebted throughout my life to my esteemed internal guide, Dr. Sugato Banerjee, Assistant Professor and external guide, Dr. Rajeev M. Patil, Head-Biostudies, Norwich Clinical Services Pvt. Ltd., Bangalore whose guidance, invaluable encouragement, innovative ideas and quest of knowledge beyond present frontiers, enabled me to accomplish this thesis with zest and zeal. I am extremely grateful for his infallible determination, untiring patience and emotional strength that he instilled in me. I am highly grateful to Dr. B.N. Sinha, Head, Department of Pharmaceutical Science & Technology, Birla Institute of Technology, Mesra. I am highly obliged to my respected professors, faculties at BIT Mesra who have a profound influence in shaping my orientation for research. My sincere thanks to staffs of Norwich Clinical Services Mr. Sameer Ranjan, Mr. Joseph and Mr. Suresh for their invaluable words of encouragement constant help. I extend my heartfelt gratitude towards all the faculty members for their support and co-operation. I will be failing in my duty if I fail to express my heartfelt gratitude towards Dr. Arindam Mukhopadhyay, Dean, Lotus Clinical Research Academy, Bangalore for their creative guidance, valuable suggestions and persistent inspiration for the completion of this work. I wish to thank Norwich Clinical Research Pvt. Ltd. for allowing to pusue my project work at their place. I would like to thank my friends Rakesh Kumar, Aishwarya Roy and Pooja Kumari for their overwhelming help, support and co-operation. I find myself lacking in words to express my deepest sense of gratitude towards my beloved parents for their unconditional support, encouragement and motivation. It is all because of their belief and the optimism that they instilled in me that I have been able to complete this work successfully.

Department of Pharmaceutical Sciences,

Abhimanyu Thakur

Birla Institute of Technology

Reg. No. MPH/10066/2013

Mesra, Ranchi, Jharkhand-835215, India

Table of Contents Acknowledgement………………………………………………………………………………………………………………..i A. Introduction….……………………………………………………………………………………………………………….1 1. Abiraterone………………………………………………………………………………………………………………………2 1.1 General Information……………………………………………………………………………………………………..2 1.2 Pharmacology of drug…………………………………………………………………………………………………..3 1.2.1 Pharmacokinetics…………………………………………………………………………………………………3 1.2.2 Pharmacodynamics………………………………………………………………………………………………4 1.2.3 Mechanism of action……………………………………………………………………………………………..4 1.2.4 Therapeutic indications………………………………………………………………………………………...5 1.2.5 Adverse event……………………………………………………………………………………………………….5 1.2.6 Warning and precaution………………………………………………………………………………………..6 1.2.7 Drug interaction…...……………………………………………………………………………………………….6 2.Prostate Cancer…………………………………………………………………………………………………………………..7 2.1 Prostate……………………………………………………………………………………………………………………….7 2.2 Prostate Cancer……………………………………………………………………………………………………….……7 Benign Prostate Hyperplasia…………………………………………………………………………………………7 Castration resistant prostate cancer………………………………………………………………………………8 Risk factor……………………………………………………………………………………………………………………8 Diagnosis…………………………………………………………………………………………………………….............8 Treatment……………………………………………………………………………………………………………………8 3. Bioequivalence study…………………………………………………………………………………………………………9 3.1 Need of bioequivalence study……………………………………………………………………………………….9 3.2 Phases of bioequivalence study…………………………………………………………………………………….9 3.2.1 Clinical phase….…………………………………………………………………………………………………..9 3.2.2 Bio-analytical phase……………………………………………………………………………………….……9 3.2.3 Statistical phase….………………………………………………………………………………………….……9 3.2.4 Clinical phase….……………………………………………………………………………………………….….9 3.2.5 Pharmacokinetic analysis of primary and secondary parameters……………………….…..9 3.2.6 Clinical study report preparation…………………………………………………………………….……9

B. Literature review…………………………………………………………………………………………………..............10 C. Rationale and Objective….………………………………………………………………………………………………12 1. Rationale….……………………………………………………………………………………………………………………....12 2. Objective….………………………………………………………………………………………………………………………12 2.1 Primary Objective.……………………………………………………………………………………………………...12 2.2 Secondary Objective…………………………………………………………………………………………….……..12 D. Plan of work…………………..….…………………………………………………….………………………….………….13 E. Research Methodology…..………………………………………………………………………………….….………..14 1. Study initiation phase…...……………………………………………………………………………………………….14 a. Synopsis…….……….…..………………………………………………………………………………………………..14 b. Study protocol...………………………………………………………………………………………………………..18 c. Inform Consent Form……..…………………………………………………………………………………….…...19 d. Case Report Form………...……………………………………………………………………………………………20 2. Clinical part……………………………………...…………………………………………………………………………....21 3. Bioanalytical Measurements……………………………………………………………………………………….….27 4. Pharmacokinetic analysis.……………………………………………………………………………………………...38 F. Results……….…..………..….………………………………………………………………………………………..….....…..38 G. Discussion and conclusion……………………………………………………………………………………………..70 H. References…..…………..….……………………………………………………………………………………………....….71 I. Appendix………………………………………………………………………………………………………………………….74 1. Demographic data listing………………………………………………………………………………………………..74 2. Abnormal laboratory values…………………………………………………………………………………………...74 3. Source documents (Separately attached)...………………………………………………………………………75 a. Clinical Study Protocol……………………………………………………………………………………………….75 b. Inform Consent Document…………………………………………………………………………………………75 c. Case Report Forms…………………………………………………………………………………………………….75 d. Case Study Report……………………………………………………………………………………………………..75 4. Study Event Chart..………………………………………………………………………………………………………..76

List of Tables Table No.

Title

Page No.

1

Pharmacokinetic parameters

4

2

Scores for PSA level

8

3

Overall Study Plan

21

4

Study count of volunteers who completed the study

22

5

Selection and Timing of Dose in the Study Period I

24

6

Selection and Timing of Dose in the Study Period II

24

7

Selection and Timing of Dose in the Study Period III

25

8

Selection and Timing of Dose in the Study Period IV

25

9

Instruments and equipments

27

10

Details of chemicals used

29

11

Internal standard stock dilutions

30

12

Stock dilutions of Abiraterone acetate

31

13

Stock dilutions for QC

31

14

Spiking preparation

32

15

Spiking preparation for QC

33

16

Various parameters of instrument

34

17

Individual plasma Concentration (ng/ml) for Period I for Abiraterone

35

18

Individual plasma Concentration (ng/ml) for Period II for Abiraterone

36

19

Individual plasma Concentration (ng/ml) for Period III for Abiraterone

36

20

Individual plasma Concentration (ng/ml) for Period IV for Abiraterone

37

21

Calculations of pharmacokinetic parameters of subject 1 in period I

39

22


41

23


43

Table No.

Title

Page No.

24


45

25


47

26


49

27

Calculations of pharmacokinetic parameters of subject 1 in period II

50

28


53

29


55

30


57

31


59

32


61

33

Primary and Secondary parameters that were analysed for the Reference product

63

34

Primary and Secondary parameters that were analysed for the Test product

63

35

ANOVA for Cmax

67

36

ANOVA for AUC 0-t

67

37

ANOVA for AUC 0-∞

67

38

Confidence interval for the ratio of test and reference product

69

39

Demographic Data Listing

74

40

Abnormal laboratory values

74

41

Study events

77

List of Figures Figure No. 1

Title Chemical structure of Abiraterone Acetate

Page No. 2

2

Mechanism of Action of Abiraterone Acetate

5

3

A typical prostate gland

7

4

Steps involved in the study

13

5

Concentration vs. Time plot for subject 1 in period I

39

6

lnC vs. Time plot for subject 1 in period I

40

7


41

8


42

9


43

10


44

11


45

12


46

13


47

14


48

15


49

16


50

17

Concentration vs. Time plot for subject 1 in period II

51

18

lnC vs. Time plot for subject 1 in period II

52

19


53

20


54

21


55

22


56

Figure No.

Title

Page No.

23


57

24


58

25


59

26


60

27


61

28


62

29

Pair wise presentation of individual plasma Abiraterone concentration

64

vs. time curves following a single dose of 250 mg Abiraterone acetate. 30

Disposition of Subject

75

List of Abbreviations µg

Microgram

µL

Microlitre

ADR AE

Adverse Drug Reaction Adverse Event

ALP

Alkaline Phosphate

ANOVA

Analysis of Variance

AUC

Area Under Curve

BA

Bioavailability

BE

Bioequivalence

BLQ

Below Limit of Quantification

BMI

Body Mass Index

CDER CDSCO

Centre for Drug Evaluation and Research Central Drugs Standard Control Organization

CI

Confidence Interval

cm

Centimetre

Cmax

Maximum observed concentration in plasma

COSTART Coding Symbols for Thesaurus of Adverse Reaction Terms CRF CRPC

Case Report Form Castration Resistant Prostate Cancer

CV

Coefficient of Variation

DHEA

Dehydroepiandrosterone

dl

Decilitre

ECG

Electrocardiogram

GCP

Good Clinical Practice

GLP

Good Laboratory Practice

HCV

Hepatitis C Virus

HIV

Human Immunodeficiency Virus

HQC

High Quality Control

hr

Hour

ICD

Informed Consent Document

ICH

International Conference on Harmonization of Technical Requirements for Registration of Pharmaceuticals for Human Use

ICMR IEC IP K2EDTA

Indian Council of Medical Research Independent Ethics Committee Investigational Product Disodium Ethylene Diamine Tetra Acetate

Ke

Elimination Rate Constant

Kg

Kilogram

L

Litre

LC/MS/MS Liquid Chromatography/ Mass Spectroscopy/ Mass Spectroscopy LOQ LOQQC

Limit of Quantification Quality Control at Lower Limit of Quantification

LQC

Low Quality Control

LSM

Least Square Mean

M

Meter

mg

Milligram

ml

Millilitre

mM/L

Millimole per litre

MQC

Middle Quality Control

N

Number

ng

Nanogram

°C

Degree Celsius

PCV

Packed Cell Volume

P-gp

P-glycoprotein

PK

Pharmacokinetic

PSA

Prostate Specific Antigen

QA

Quality Assurance

Rpm

Revolutions Per Minute

RSD

Relative Standard Deviation

SAE

Serious Adverse Event

SAS

Statistical Analysis Software

SD

Standard Deviation

SGOT

Serum Glutamic Oxaloacetic Transaminase

SGPT

Serum Glutamic Pyruvic Transaminase

SOP t1/2

Standard Operating Procedure Elimination half-life

Tmax

Time to observed maximum drug concentration in plasma

W/O

Washout

WBC

White Blood Cell (Leukocyte)

WMA

World Medical Association

Bioequivalence study of Abiraterone acetate tablet in Indian healthy adult male volunteers INTRODUCTION Bioequivalence studies are the commonly accepted method to demonstrate therapeutic equivalence between two medicinal products. Savings with respect to time and cost are substantial when using bioequivalence as an established surrogate marker of therapeutic equivalence. For this reason the design, performance and evaluation of bioequivalence studies have received major attention from academia, pharmaceutical industry and health authorities. Bioequivalence Studies in Drug Development focuses on the planning, conducting, analysing and reporting, covering all aspects required by regulatory authorities. In the present work, bioequivalence study of Abiraterone acetate has been performed. Abiraterone acetate is a prodrug of Abiraterone that specifically inhibits CYP17 and thereby blocks biosynthesis of extragonadal, testicular, tumor androgens. It is used in combination with prednisolone for treatment of metastatic castration resistant prostate cancer. This study was performed to compare the bioavailability of Abiraterone acetate tablet manufactured by X Pharmaceuticals with that of innovator’s (Y Pharmaceuticals’) Abiraetone acetate by using data obtained from plasma Abiraterone concentration for determining bioequivalence between these two products. Eight Indian volunteers participated in this study that was of a crossover design, with seven days washout period. After an overnight fast, a single 250 mg Abiraterone acetate tablet of either the reference product or the test product was orally administered to each subject. Plasma blood sample of 4 ml was drawn prior to dosing and at 0.33, 0.67, 1.00, 2.00, 3.00, 6.00, 8.00, 10.00, 24.00, 48.00, 72.00 hours after dosing. The pre-dose and post-dose blood samples were collected in pre-chilled, pre-labeled vacutainers containing sodium fluoride (anticoagulant). The plasma sample was analyzed for Abiraterone concentration using LC-MS-MS. Subsequently the pharmacokinetic parameters viz., Cmax, Tmax, AUC0-t, AUC0-∞, t1/2, etc. of Abiraterone acetate were calculated. The test and reference formulations were considered to be bioequivalent if at 90% confidence intervals the logtransformed values were within the predetermined equivalence range 8%-125% for AUC and Cmax. The results showed that for Abiraterone acetate, 90% confidence interval ratio for Cmax, AUC0-t, AUC0-∞ were 83.3%, 94.6%, and 88.4% for log-transformed data respectively. It can be concluded that the test product was bioequivalent to the reference product based on the criteria that the percent ratio of test parameters was within the range of 80.00 – 125.00 % with a 90% confidence interval. KEY WORDS: Abiraterone acetate/ Bioequivalence study/ Pharmacokinetic study/ Clinical Research

Birla Institue Technology Mesra, Ranchi, India

Abhimanyu Thakur (MPH/10066/2013)

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Bioequivalence study of Abiraterone acetate tablet in Indian healthy adult male volunteers 1. Abiraterone 1.1.

General information Abiraterone acetate is considered as pro-drug of Abiraterone that specifically inhibits CYP17 and thereby blocks biosynthesis of extragonadal, testicular, tumour androgen1.

Figure 1: Chemical structure of Abiraterone Acetate The IUPAC name of Abiraterone is (3β)-17-(3-Pyridinyl)-16-dien-3-ol-androsta-5 acetate (ester). Its molecular formula2 is C26H33NO2. Brand name3: Zytiga (Janssen Biotech, Inc.) Dosage forms3: Tablet (250 mg) Therapeutic dosage3: Baseline moderate hepatic impairment (Child-Pugh Class B): 250 mg once daily. In case of hepatotoxicity: Hold ZYTIGA until recovery. Reduced dose of ZYTIGA for retreatment should be used. In case of severe hepatotoxicity, ZYTIGA should be discontinued. Uses4: It is used in combination with prednisolone for treating metastatic castration resistant prostate cancer. Patient population5: Adult male suffering from castration resistant prostate cancer. It should not be given to women and children. Physical property6: Off-white solid, molecular formula is C26H33NO2, molecular weight is 391.55 Solubility7: Soluble in organic solvents, mainly in alcohols. Slightly soluble in a 0.1 N HCl solution. Insoluble in aqueous media over a wide range of pH values tested

(2 - 12) at 20°C.

Melting point7: 147°C to 148°



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Bioequivalence study of Abiraterone acetate tablet in Indian healthy adult male volunteers 1.2.

Pharmacology of drug 1.2.1. Pharmacokinetics 8 In fasting state only 5% of Abiraterone acetate is absorbed by the intestine i.e. 50mg of drug is being absorbed per 1000 mg (4 tablets) dose. The absorption of Abiraterone acetate is enhanced tremendously when taken along with food. Thereby 60 mg of drug absorptions occurs per 250 mg of single dose which is far greater than the completed dose administered on an empty stomach. Absorption8 Following oral administration, Abiraterone acetate gets solubilised by bile acids in the stomach. It gets disintegrated in the stomach and the released Abiraterone acetate goes along the small intestine and get absorbed by the wall of small intestine and subsequently enters the blood. As the intestinal blood goes through the liver, the Abiraterone acetate avoids first pass metabolism and it is cleaved by ester to yield Abiraterone. Distribution8 Then the Abiraterone acetate is released by the blood to the various organs including gonads, prostate and adrenal glands. The Abiraterone acetate binds firmly to CYP17 enzyme which is responsible for the production of all androgens in the body. In this way it prevents the enzyme from androgen synthesis and it leads to reduction in testosterone levels. As a result, the prostate tumour becomes starved of testosterone and its multiplicationstops. Metabolism 8 The inactive metabolite of Abiraterone acetate viz. Abiraterone N-Oxide is made by CYP3A4. The pyridyl nitrogen is oxidised to N-oxide Abiraterone which is incapable of blocking

CYP17

and

becomes

inactive.

Excretion 8 The SULT2A1 (steroid sulfactase) enzyme forms water soluble Abiraterone N-Oxide Sulfate from by putting the sulphate group on to the 3-hydroxy moiety of Abiraterone Noxide. The water soluble Abiraterone N-oxide is excreted in urine. This is the fate of 4% of fully metabolised Abiraterone acetate.



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Bioequivalence study of Abiraterone acetate tablet in Indian healthy adult male volunteers

Table 1: Pharmacokinetic parameters 9 AUC

979 ng×h/ml

Cmax

226 ng/ml

Tmax

2 hours

Vd

5630 L

Cl

33%

Plasma protein binding

99.8%

1.2.2. Pharmacodynamics 10 The testosterone and androgen levels are lowered in serum by Abiraterone acetate than that by LHRH agonists alone or by orchiectomy. For prostate cancer patients, Prostate Specific Antigen (PSA) acts as biomarker. In a stage 3 clinical investigation of patients who fizzled former chemotherapy with taxanes, 29% of patients treated with abiraterone, versus 6% of patients treated with placebo, had at any rate a half decay from pattern in PSA levels. Impacts on the QT interim In a cardiovascular security mull over in patients with metastatic progressed prostate malignancy, there were no huge impacts of abiraterone acetic acid derivation on the heart QT/Qtc interim. 1.2.3. Mechanism of action 11 Abiraterone is structurally analogous to pregnenolone and it inhibits enzyme 17αhydroxylase/C17,

20-lyase

(CYP17); required for synthesis of androgen. Thereby the

androgen synthesis is reduced in testes, prostate and adrenal glands which lead to reduction in serum concentration of testosterone.



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Future Oncol, 2010, Future Medicine Ltd. Figure 2: Mechanism of Action of Abiraterone Acetate7 1.2.4. Therapeutic indications9 Abiraterone acetate is indicated for treating metastatic prostate cancer in patients who have taken prior chemotherapy of docetaxel after the androgen deprivation therapy got failed. For geriatric patients (age greater than or equal to 65 years): Overall no difference is found between younger and elderly patients. For paediatric: It has not been studied in this age group of population. 1.2.5. Adverse event 9 Some of the adverse events encountered by the Abiraterone acetate are hypokalemia, edema, increased alanine transferase, urinary tract infection, hematuria, dyspepsia, fracture and hypertension. Elevation in AST and ALT levels are the most common adverse drug reaction.



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Bioequivalence study of Abiraterone acetate tablet in Indian healthy adult male volunteers 1.2.6. Warning and precaution 9 As a matter of fact the blood level of Abiraterone acetate is increased when taken on an empty stomach. Hence, it must be administered on an empty stomach as this may prove to be harmful. No food should be taken two hours prior or at least two hours after the administration of Abiraterone acetate. Prior to use of Abiraterone acetate, the doctor should be consulted in following conditions: 

High blood pressure or low blood potassium.



Any heart related problem viz. Heart failure, heart attack.



Any liver problem.



Any dental problem.

It may be harmful to unborn baby. Condom must be used by male patients during sexual activity with a pregnant woman while taking Abiraterone acetate. Protective gloves should be used by pregnant women while handling Abiraterone acetate. Abiraterone acetate should not be used by under 18 year patients. 1.2.7. Drug interaction9 Drug-Drug interactions CYP1A2 substrate- Theophylline: Systemic exposure is not increased. CYP2D6 substrate- Dextromethorphan: Systemic exposure was enhanced by 200%. CYP2C8 substrate: No clinical evidence. CYP3A4 inducers- Rifampicin: When it is followed by administration of 1000 mg of Abiraterone acetate, the AUC∞ is reduced by 55%. Strong CYP3A4 inducers such as phenytoin, rifampicin, Phenobarbital, carbamazepine and rifabutin should be avoided while treating with Abiraterone acetate. The evaluation of clinical efficacy should be done carefully if at all CYP3A4 must be co-administered with Abiraterone acetate as there is no clinical evidence of dose adjustment. Drug- Food interactions The absorption of Abiraterone acetate is greatly enhanced if taken with food. Drug-Herb interaction St. John’s wort (Hypericum perforatum) when co-administered with Abiraterone acetate, the plasma concentarion of Abiraterone acetate is reduced. Birla Institue Technology Mesra, Ranchi, India


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Bioequivalence study of Abiraterone acetate tablet in Indian healthy adult male volunteers 2. Prostate Cancer 2.1.

Prostate 12 The prostate is a gland in human male which is located in front of the urinary bladder and rectum. Its size varies with age. It is like walnut in younger men and is of much larger size in older men. The prostate gland synthesizes fluid which protects and nourishes sperm cells in semen. Behind the prostate gland, seminal vesicles are located which make major portion of semen fluid. Through the centre of the prostate gland, the urethra goes which is a tubular structure that carries semen and urine through the penis. Prior to birth, the prostate commences to develop. During puberty, its growth is rapid aided by the male androgen hormones. The male androgen hormone viz. testosterone is synthesized in the testicles. The testosterone is converted into dihydrotestosterone (DHT) by the enzyme 5-alpha reductase. The DHT gives signal for the growth of prostate. Normally, the prostate remains of same size or grow gradually till the male hormones are present.

Figure 3: A typical prostate gland12 2.2.

Prostate Cancer 13 When the cells in prostate gland grow abnormally, it forms tumour. It may be benign or malignant. It may spread by the process referred to as metastasis. The cancer that starts in gland cells is referred as adenocarcinoma. Prostate cancer usually occurs in older male. Benign Prostatic Hyperplasia (BPH) 13 This is the medical condition in which the prostate gland overgrows which is non-cancerous.



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Bioequivalence study of Abiraterone acetate tablet in Indian healthy adult male volunteers Castration resistant prostate cancer (CRPC)13 This is the medical condition in which even after the deprivation of male androgen hormone viz. testosterone, the prostate gland tissues keep growing. That means it is no longer dependent on testosterone and get the support from some other body organ. Risk factor 13, 14 Older age: Old age male are more prone to the prostate cancer. Among all prostate cancers, about 65% cases have been found in men having about 65 years of age. Family history: Men having family history of prostate cancer is more likely to develop the prostate cancer. Race: In a study, it has been found that African-American males are more likely to suffer from prostate cancer as compared to Caucasian men. Diagnosis 14, 15 Prostate Specific Antigen (PSA) test: In this test, the level of PSA (a protein produced by prostate) is determined by drawing a small amount of blood sample from the arm. Table 2: Scores for PSA level PSA level

Scoring

Under 4 ng/ml

Normal

Over 10 ng/ml

High

Between 4 and 10 ng/ml

Intermediate

Digital Rectal Exam (DRE): In this technique, the physician examines prostate for any sort of irregularities by inserting the gloved, lubricated finger inside the rectum. Biopsy: Based on the results of PSA test and DRE assessment the biopsy of prostate is recommended. Magnetic Resonance Imaging (MRI): The MRI is performed to check if another biopsy is necessary or not. Treatment 14, 15 Based on the age and stage of prostate cancer treatment options are active surveillance, radical treatment (Prostatectomy), radiation therapy (External beam radiation, bachytherapy), hormone therapy, and chemotherapy. Birla Institue Technology Mesra, Ranchi, India


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Bioequivalence study of Abiraterone acetate tablet in Indian healthy adult male volunteers 3. Bioequivalence study16 According to ICH bioequivalence is “The absence of a significant difference in the rate and extent to which the active ingredient or active moiety in pharmaceutical equivalents or pharmaceutical alternatives becomes available at the site of drug action when administered at the same molar dose under similar conditions in an appropriately designed study”. 3.1.

Need of bioequivalence study To obtain the marketing approval for a generic product, it is necessary that it is bioequivalent to the already established brand (i.e. reference product). Phases of bioequivalence study17

3.2.

3.2.1. Clinical Phase Preparation of clinical trial protocol and study design Determination of sample size and randomization of subjects Designing Case Report Forms (CRFs) Preparation of Informed Consent Forms Regulatory submission to Independent Ethics Committee (IEC) Recruitment of healthy volunteers as per inclusion and exclusion criteria Check-in and examinations such as laboratory tests Bioequivalence phase I- II (a) Reporting adverse events. Processing of Clinical data and statistical analysis 3.2.2. Bio-analytical Phase Development and validation of assay of the drug and its metabolites. Determination and analysis of drug and its metabolites in biological fluid samples. 3.2.3. Statistical Phase Processing of bio-analytical data Calculation of Pharmacokinetic parameters Writing report on clinical, bio-analytical and statistical data Archival of documents 3.2.4. Pharmacokinetic analysis of primary and secondary parameters 3.2.5. Clinical Study Report preparation



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Bioequivalence study of Abiraterone acetate tablet in Indian healthy adult male volunteers LITERATURE REVIEW  Sternberg et al. 2014 analysed an early access protocol trial that was initiated after completion of COUAA-301 to enable preapproval access to abiraterone acetate in patients with casteration resistant prostate cancer progressing after chemotherapy. It was found that median time to PSA progression was 8.5 months and median time to clinical progression was 12.7 months 16.  Matsubara et al. 2014 evaluated the pharmacokinetics, pharmacodynamics, and preliminary efficacy and safety of in Japanese CRPC patients. This study was designed to validate the clinical utility of the globally approved Abiraterone acetate recommended dose (1000 mg) in chemotherapy naive Japanese CRPC patients, and to investigate whether Abiraterone acetate dosing needs are different in the Japanese population. It was found that the PSA response rate of 67% in Japanese CRPC patients is comparable to a 66% response rate in a similar phase I study in a Western population. Overall, the Abiraterone acetate safety profile in Japanese CRPC patients was comparable to that in the Western population17.  Satoh et al. 2014 evaluated the efficacy and safety of oral Abiraterone acetate (1000 mg/once daily) plus prednisolone (5mg/twice daily) in metastatic castration-resistant prostate cancer patients from Japan who had previously received docetaxel-based chemotherapy. It was found that Abiraterone acetate plus prednisolone showed favourable efficacy in metastatic castration-resistant prostate cancer Japanese patients who had received chemotherapy. Abiraterone acetate plus prednisolone had an acceptable safety profile18.  Ryan et al. 2014 studied androgen dynamics and serum PSA level in patients treated with Abiraterone acetate. Abiraterone plus prednisone significantly reduced serum androgens, as measured by ultrasensitive assays and was generally associated with PSA response. However, androgen decline did not uniformly predict PSA decline suggesting ligand independent or other mechanisms for mCRPC progression19.  Goodman et al. 2014 did exploratory analysis of the visceral disease subgroup in a phase III study of Abiraterone acetate in metastatic castration resistant prostate cancer. Abiraterone acetate plus prednisone produced similar absolute improvement in patients with and without visceral disease versus prednisone20.



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Bioequivalence study of Abiraterone acetate tablet in Indian healthy adult male volunteers  Amit et al. 2014 reviewed the Princess Margaret experience of low dose Abiraterone and prior ketoconazole. It was concluded that low dose Abiraterone acetate or prior KT treatment were not associated with poorer outcome in men with mCRPC treated with Abiraterone acetate21.  Basch et al. 2013 analyzed data for patient-reported pain and functional status in a preplanned interim analysis of a phase-III. The results show that median time to progression of mean pain intensity was longer in patients assigned to Abiraterone plus prednisone than in those assigned to placebo plus prednisone22.  Harshman et al. 2013 reviewed for evaluating its efficacy in earlier disease states, exploring its synergy in combination with other therapeutic agents, and assessing the necessity for administration of concurrent steroids and gonadal suppression. It was concluded that patients now have access to more than five therapies that prolong survival, with several more on the horizon23.  Fizazi et al. 2012 analyzed the study (COUAA301) before crossover from placebo to abiraterone acetate. This analysis confirms that Abiraterone acetate significantly prolongs overall survival in patients with metastatic castration resistant prostate cancer who have progressed after docetaxel treatment24.  Acharya et al. 2012 evaluated the pharmacokinetic, safety, tolerability of Abiraterone acetate in healthy men. Two phase-I studies were conducted in healthy men aged 18-55 years. A dose related increase in Abiraterone exposure was observed in both studies25.



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Bioequivalence study of Abiraterone acetate tablet in Indian healthy adult male volunteers RATIONALE AND OBJECTIVE 1. RATIONALE Bioequivalence is a part of regulatory submission which is required for obtaining marketing approval for any generic product in which it is compared with an already established brand (reference product). The present study has been conducted to compare non inferiority of the investigational product, Abiraterone manufactured by X Pharmaceutical with respect to pharmacokinetic profile & bioequivalence as compared to the reference product (Abiraterone acetate) manufactured by Y Pharmaceutical, in normal healthy male adult, human subjects under fasting conditions. 2. OBJECTIVE Primary objective To characterize the rate and extent of Abiraterone absorption after oral administration and to assess the bioequivalence of the sponsor’s test product Abiraterone Acetate Tablets 250 mg relative to that of reference Abiraterone acetate Tablets in healthy, adult, human male subjects under fasting conditions. Secondary objective To assess the safety and tolerability of both the formulations on the basis of clinical and laboratory examination, documentation of the Adverse Event (AE) and or Adverse Drug Reactions (ADRs).



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Bioequivalence study of Abiraterone acetate tablet in Indian healthy adult male volunteers PLAN OF WORK

Figure 4: Steps involved in the study •

Study Initiation - Preparation of Synopsis, Protocol, Informed consent Document, Case Report Form, CRF - Attachments, Ethical Submission.

•

Clinical Phase - Obtaining Voluntary Informed Consent, Screening subjects, Check-in, Dosing, Sample collection, Safety Monitoring, Check-out. Segregation of blood sample and sample shippment for bioanalysis.

•

Bioanalytical Measurement -

Validated LC-MS-MS methods was used for determination of

Abiraterone acetate concentration in blood samples. Data generated was used for Pharmacokinetic analysis. •

Pharmacokinetic Analysis Primary Parameters Cmax, AUC 0-t, Tmax. Secondary Parameters - t1/2, AUC 0-t , AUC 0-α

•

Statistical Analysis - Criteria for Equivalence Cmax, AUC 0-t , AUC 0-α of the Test product for the study should fall within 80% to 125% (both inclusive) .

•

Preparation of Clinical Study Report



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Bioequivalence study of Abiraterone acetate tablet in Indian healthy adult male volunteers RESEARCH METHODOLOGY The present thesis is concerned about the bioequivalence study of Abiraterone acetate in drug development which aims on the planning, conducting, analysing and reporting of bioequivalence studies encompassing all aspects required by regulatory authorities. It is a regulatory requirement for getting marketing approval for a generic product after the expiration of patent of a comparator product. Research methodology is subdivide in following sections: i.

Study Initiation

ii.

Clinical phase

iii.

Bioanalytical measurements

iv.

Pharmacokinetic analysis

STUDY INITIATION Preparation of the study related documents which includes the following and was cross verified and reviewed by the Principle Investigator. 

Synopsis



Protocol



Informed consent form



Case Report Form

SYNOPSIS 1. TITLE:

An open labelled, randomized, balanced, two treatment, two period, two sequence, single dose, four way replicated crossover, oral bioequivalence study of Abiraterone Acetate Tablets 250 mg, manufactured by X Pharmaceutical compared with that of

Abiraterone Acetate Tablets

manufactured by Y Pharmaceutical in healthy adult human male subjects under fasting condition. 2. OBJECTIVE:

a. PRIMARY OBJECTIVE:

To characterize the rate and extent of Abiraterone absorption

after oral administration and to assess the bioequivalence of the sponsor’s test product Abiraterone Acetate Tablets 250 mg relative to that of reference product (Abiraterone acetate tablets) in healthy, adult, human male subjects under fasting conditions.



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Bioequivalence study of Abiraterone acetate tablet in Indian healthy adult male volunteers b. SECONDARY OBJECTIVE: To assess the safety and tolerability of both the formulations on the basis of clinical and laboratory examination, documentation of the Adverse Event (AE) and or Adverse Drug Reactions (ADRs). 3. STUDY DESIGN: An open label, randomized, balanced, two treatment, four period, two sequence,

single dose, four way fully replicated crossover oral bioequivalence study with wasout period of at least 06 days in healthy, adult, human male subjects under fasting conditions. 4. INVESTIGATIONAL PRODUCT:

Test: Abiraterone Acetate Tablet 250 mg Manufactured by X Pharmaceutical Reference: Abiraterone acetate Tablets 250 mg Manufactured by Y Pharmaceutical 5. SAMPLE SIZE: 08 healthy, adult, human male subjects were included in the study. 6. SCREENING PROCEDURE:

During screening procedure, demographic data, medical history, general physical examination including vitals, 12-lead ECG, chest X-ray, hematology, biochemistry, serology (HIV, VDRL, Hepatitis B and C tests), urine drug screening, alcohol breath analysis, urine analysis were be performed for all subjects 7. HOUSING:

Subjects were housed from at least 11 hours prior to drug administration until after 48 hours postdose blood draw in each period. 8. STUDY MEAL:

Food was restricted from at least 10 hours before drug administration until 4 hours post-dose. Predefined standard meals were provided as scheduled. 9. STUDY RESTRICTION:

Water only 240 ml given during dosing. Physical activity: Following drug administration first 4 hours seated upright. Birla Institue Technology Mesra, Ranchi, India


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10. DRUG ADMINISTARTION:

In each period, following an overnight fast of at least 10 hours, subjects received a single oral dose of test or reference product while in sitting posture with 240 ml of ambient water. Drug administration was done as per the randomization schedule generated by the SAS software version 9.2 or higher. Oral cavity and hands were checked by the trained study personnel immediately after drug administration. 11. BLOOD SAMPLING:



In each study period, 12 blood samples (4ml) were collected from each subject.



Pre-dose (0.00 hour) within 1 hour prior to dosing.



Post-dose blood sample collection at 0.33, 0.67, 1.00, 2.00, 3.00, 6.00, 8.00, 10.00, 24.00, 48.00, 72.00 hours.



Blood samples were collected in pre-chilled, pre-labelled vacutainers containing Sodium Fluoride+ Potassium Oxalate (anticoagulant).



Centrifugation of samples at 3500 rpm at 4±2°C for 10 minutes to separate plasma.



Separated plasma were transferred as two aliquots to pre-chilled, pre-labelled polypropylene tubes, packed in black polythene bags and stored upright in deep freezer at a temperature of -70°C.



Blood samples were collected through in dwelling intravenous cannula placed in a forearm vein of the subject.



After each blood draw, 0.5 ml of heparinized saline were injected into the cannula to prevent coagulation.



Before each blood draw, 0.5 ml of blood were discarded to prevent the heparin in cannula from interfering with analysis.

12. INCLUSION/ EXCLUSION CRITERIA:

a. Inclusion criteria  Age: 18 to 45 years.  Weight ≥ 50 Kg  BMI: 18.5 TO 24.99 (Both inclusive).  All the screening parameters should be within normal range.  Volunteers should be capable of giving consent.  Willing to comply with the study requirements. Birla Institue Technology Mesra, Ranchi, India


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Bioequivalence study of Abiraterone acetate tablet in Indian healthy adult male volunteers  Must be a non-smoker, and has no history of smoking or use of nicotine-containing substances within the previous 2 months.  Must have blood pressure between 90 and 140 mmHg systolic, inclusive, and no higher than 90 mmHg diastolic.  Must have a 12-lead electrocardiogram consistent with normal cardiac conduction and function.  Must agree to use an adequate contraception method as deemed appropriate by the investigator and to not donate sperm during the study for 3 months after receiving the last dose of study medication b. Exclusion criteria:  If any type of abnormality.  If hypersensitive to the drug.  If any subject has participated in a trial or blood withdraw recently.  If they are on concomitant medication.  Lactating mother, pregnant female.  History of or current clinically significant medical illness including cardiac arrhythmias or other cardiac disease that could interfere with the interpretation of the study results.  Clinically significant abnormal values for hematology or urinalysis (at screening).  Clinically significant abnormal physical examination vital signs or 12-lead electrocardiogram (at screening).  Usage of any prescription or non prescription medication, except for acetaminophen, and oral contraceptives and hormonal replacement therapy within 14 days before the first dose of the study medication is scheduled.  Unable to swallow solid, oral dosage forms whole with the aid of water (participants may not chew, divide, dissolve, or crush the study medication) 13. SAFETY MONITORING:

a. Vital monitoring: 02 hours prior to dosing, 02, 04, 08, 10 hours post-dose within ± 30 minutes from the scheduled time, prior to check-out and additionally if required. b. Well-being: Enquired twice a day post-dose on all housing days (2-4 hours post-dose and 10-12 hours post dose on day 1; 24-26 hours post-dose and 32-34 hours post-dose on day 2) and prior to check out of each period. Birla Institue Technology Mesra, Ranchi, India


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Bioequivalence study of Abiraterone acetate tablet in Indian healthy adult male volunteers c. ECG: Performed between 2-4 hours post-dose in each period and at the end of study. d. Serum electrolyte test (Sodium, potassium and phosphate) test: Performed at the checkout of period I and Period III. e. Hematology and biochemistry investigations: Performed at screening, repeated during check-out of second period and at the end of study. f. Urine drug screening and alcohol breath analysis for all subjects performed before checkin of each period. 14. BIOANALYTICAL MEASUREMENTS:

Drug samples/plasma samples were analysed using validated LC-MS-MS 15. PHARMACOKINETIC ANALYSIS:

Estimation of Cmax, tmax, AUC, t1/2, Ke 16. BIOEQUIVALENCE CRITERIA: The ratio and 90% confidence interval for pharmacokinetic

parameter of the transformed natural log ratios between the test and reference formulations fall within the range of 80 % to 125%. PROTOCOL: In a clinical trial, the protocol is carefully designed to safeguard the health of the participants as well as answer specific research questions. A protocol describes what types of people may participate in the trial; the schedule of tests, procedures, medications, and dosages; and the length of the study. While in a clinical trial, participants following a protocol are seen regularly by research staff to monitor their health and to determine the safety and effectiveness of their treatment. In the present project, it has been prepared and reviewed by the Principal Investigator. The study protocol was approved by Independent Ethics Committee (IEC). * For Clinical Study Protocol, see APPENDIX (1) Ethical submission and safety considerations: Independent Ethics Committee (IEC) An independent body (a review board or a committee, institutional, regional, national, or

supranational),

constituted of medical professionals and non-medical members, whose responsibility is to ensure the protection of the rights, safety and well-being of human subjects involved in a trial and to provide public assurance of that protection, by, among other things, reviewing and approving / providing favourable



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Bioequivalence study of Abiraterone acetate tablet in Indian healthy adult male volunteers opinion on, the trial protocol, the suitability of the investigator(s), facilities, and the methods and material to be used in obtaining and documenting informed consent of the trial subjects. The legal status, composition, function, operations and regulatory requirements pertaining to Independent Ethics Committees may differ among countries, but should allow the Independent Ethics Committee to act in agreement with GCP as described in this guideline. Ethical principles: It deals with safety, wellbeing and confidentiality of subjects. Grant of approval for study The protocol was approved by the ethical committee and after the grant of permission the study starts with the preparation of Informed consent Document, Case Report Form, CRF - Attachments, Study Essential Documents, Study specific checklists, Randomization schedule, Ethical Submission as per requirement. The following documents were submitted to the IRB/IEC for the review before the conduct of the study:  Trial protocol(s)/amendment(s),  Written informed consent form includes patient information sheet , consent certificate  Subject recruitment procedures (e.g. advertisements),  Investigator's Brochure (IB),  Investigational product related safety information,  Case report form  Payments and compensation information provided to subjects,  Investigator’s current curriculum vitae and other documentations evidencing qualifications and experience of Investigator. Ethical Conduct of the study The study was conducted in compliance with protocol and the requirements of the Schedule Y (Amended Version, 2005) of CDSCO (Central Drugs Standard Control Organization), Ministry of health and family welfare, Government of India, Ethical guidelines for biomedical research on human participant, ICMR (Indian Council of Medical Research, 2006), Drugs and Cosmetics (Amendment) Rules, 2013, ICH (International Conference on Harmonization) E6 “Guideline for Good Clinical Practice”, Declaration of Helsinki (Fortazela 2013) and applicable principles of GLP. INFORMED CONSENT FORM: Informed consent is a process for getting permission before conducting a healthcare intervention on a person. A health care provider may ask a patient to consent to receive therapy before providing it, or Birla Institue Technology Mesra, Ranchi, India


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Bioequivalence study of Abiraterone acetate tablet in Indian healthy adult male volunteers a clinical researcher may ask a research participant before enrolling that person into a clinical trial. Informed consent is collected according to guidelines from the fields of medical ethics and research ethics. A process by which a subject voluntarily confirms his or her willingness to participate in a particular trial, after having been informed of all aspects of the trial that are relevant to the subject's decision to participate. Informed consent is documented by means of a written, signed, and dated informed consent form. Subject Information and Consent Informed Consent Document (English, Kannada and Back translated were approved by IEC) were used for the study. * For informed consent documents (ICD), see APPENDIX (2) CASE REPORT FORM: A case report form (or CRF) is a paper or electronic questionnaire specifically used in clinical trial research. The Case Report Form is the tool used by the sponsor of the clinical trial to collect data from each participating site. All data on each patient participating in a clinical trial are held and/or documented in the CRF, including adverse events. The sponsor of the clinical trial develops the CRF to collect the specific data they need in order to test their hypotheses or answer their research questions. The size of a CRF can range from a handwritten one-time 'snapshot' of a patient's physical condition to hundreds of pages of electronically captured data obtained over a period of weeks or months. (It can also include required check-up visits months after the patient's treatment has stopped.) The sponsor is responsible for designing a CRF that accurately represents the protocol of the clinical trial, as well as managing its production, monitoring the data collection and auditing the content of the filled-in CRFs. Content of Case Report Form i.

Protocol Compliance Form

ii.

Inclusion/ Exclusion Form

iii.

Subject number allotment form

iv.

Subject Number Allotment log & Randomization Code

v.

Check-in form

vi.

Meal Form

vii.

Suitability for drug administration form



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Bioequivalence study of Abiraterone acetate tablet in Indian healthy adult male volunteers viii.

Drug administration form

ix.

Water administration form

x.

Restriction and Prohibition form

xi.

Vitals form

xii.

Cannula Insertion and Removal form

xiii.

Blood Sampling Form

xiv.

Check-out Vitals

xv.

Check-out Form

xvi.

Alcohol Breath Analysis Screening Form

xvii.

Urine Drug Abuse Screening Form

xviii.

Chest X-ray Form

xix.

Electrocardiography Form

xx.

Medical Examination Form

xxi.

Clinical History Form

xxii.

Comment log form

xxiii.

ICD Prestudy Checklist * For Case Report Forms (CRFs), see APPENDIX (3)

CLINICAL PART The clinical study was conducted at Norwich Clinical Services Pvt. Ltd. 43/2, 3rd floor, Cauvery Medical Center, NH-7, Bellary road, Sahakar Nagar, Bangalore – 560092, Karnataka, India under the guidance of Dr. Rajeev M. Patil, MD. The study has been conducted as per protocol, Good Clinical Practice (GCP) and the applicable principles of Good Laboratory Practices (GLPs) and Standard Operating Procedures (SOPs). Table 3: Overall Study Plan Period Number

Check-in

Check-out

I

21 JAN 15

23 JAN 15

II

28 JAN 15

30 JAN 15

III

04 FEB 15

06 FEB 15

IV

11 FEB 15

13 FEB 15



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Bioequivalence study of Abiraterone acetate tablet in Indian healthy adult male volunteers Methodology This was a single-dose, four way fully replicated crossover study design to determine the bioequivalence of test product with reference product following oral administration to healthy adult, human male subjects under fasting condition. During screening in the pre-study period, 08 subjects were selected for the study. Subjects were housed for at least 11 hours prior to drug administration until after 48 hours post-dose blood draw in each period. In each period, following an overnight fast of at least 10 hours, subjects were dosed with a single oral dose of test or reference product in sitting posture with about 240 ml of water at ambient temperature. Drug administration was done as per the randomization schedule generated by the SAS software 9.2. Oral activity and hands were checked immediately after drug administration. 12 blood samples, 4 ml each, were collected at the following times relative to dosing during each treatment period: the pre-dose blood samples (0.00 hour) were collected within 1 hour prior to drug administration, and post-dose blood samples were collected at 0.33, 0.67, 1.00, 2.00, 3.00, 6.00, 8.00, 10.00, 24.00, 48.00 and 72.00. All the in-house post-dose blood samples were collected at the scheduled time with a window period of ± 02 minutes. The 72.00 hours ambulatory blood sample was collected at the scheduled time with a window period of ±01 hour. Subjects were regularly monitored during the study for adverse events (AEs). Data from a total of 08 subjects who completed the study were used for pharmacokinetic anakysis. A washout period of 07 days was given between treatment periods. Table 4: Study count of volunteers who completed the study Number of subjects planned: 08 subjects Period I: 08

Period II: 08

Period III: 07

Period IV: 07

subjects Period I: 08

Period II: 08

Period III: 07

Period IV: 07

Number of subjects dosed Number

of

completed study Number of subjects analyzed on the bioanalytical method: 06 Number of subjects withdrawn: 01 Number of subjects dropped out: NIL Number of subjects included in pharmacokinetic analysis: 06 Number of subjects included in statistical analysis: 06 Inclusion Criteria: Healthy, adult human male subjects aged 18 to 45 years; weighing at least 50 kg and having body mass index of 18.5 to 27.0 Kg/m2 at the time of screening were included in the study. They did not have any significant diseases or clinically significant abnormal conditions during screening which included medical history, physical examination, vital signs, laboratory evaluation, chest X-ray (posteroBirla Institue Technology Mesra, Ranchi, India


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Bioequivalence study of Abiraterone acetate tablet in Indian healthy adult male volunteers interior view) and 12-lead ECG recordings. Volunteers who complied with all the inclusion and exclusion criteria were enrolled into the study. All subjects had signed and dated the EC approved informed consent form prior to participating in the study. Test Product: Abiraterone Acetate Tablets, 250 mg Dose and mode of administration: Single oral dose Manufactured by: X Pharmaceutical Manufacturing Date: 09.2014 Expiry Date: 08.2016 Reference Product: Abiraterone Acetate Tablets, 250 mg Dose and mode of administration: Single oral dose Manufactured by: Y Pharmaceutical Manufacturing Date: 10.2014 Expiry Date: 02.2015 Duration of treatment: The duration of the study starting from check-in of period I to the last subject last visit was 33 days. Withdrawal Criteria: In accordance with the ICH-GCP guidelines, all subjects had the right to withdraw from the study at any time, regardless of their reasons. Over the course of the study, the investigator(s) had the right to withdraw any subject from the study based on the criteria listed in the protocol. Treatment administration: After a supervised overnight fasting of at least 10 hours, the subjects were dosed with the test or reference product (single oral dose of 250 mg tablet) as per randomization schedule with 240 ml of ambient water in sitting posture on dosing day of each period. Drug administration was done under yellow monochromatic light. Investigational Medicinal Products: Reference and test product were supplied by the sponsor and received at the clinical facility of Norwich Clinical Services Pvt. Ltd. The drug accountability and archival were done under yellow monochromatic light. Drug identification was done under normal light. Unused IPs that had been dispensed but not used for the study were returned to the pharmacy and retained along with the remaining IPs of its type.



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Table 5: Selection and Timing of Dose in the Study Period I Date

Dosing Time

Subject No.

22 JAN 15

08:00 AM

01

08:02 AM

02

08:04 AM

03

08:06 AM

04

08:08 AM

05

08:10 AM

06

08:12 AM

07

08:14 AM

08

Table 6: Selection and Timing of Dose in the Study Period II Date

Dosing Time

Subject No.

29 JAN 15

08:00 AM

01

08:02 AM

02

08:04 AM

03

08:06 AM

04

08:08 AM

05

08:10 AM

06

08:12 AM

07

08:14 AM

08



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Table 7: Selection and Timing of Dose in the Study Period III Date

Dosing Time

Subject No.

05 FEB 15

08:00 AM

01

08:02 AM

02

08:04 AM

03

08:06 AM

04

08:08 AM

05

08:10 AM

06

08:12 AM

07

08:14 AM

08

Table 8: Selection and Timing of Dose in the Study Period IV Date

Dosing Time

Subject No.

12 FEB 15

08:00 AM

01

08:02 AM

02

08:04 AM

03

08:06 AM

04

08:08 AM

05

08:10 AM

06

08:12 AM

07

08:14 AM

08

Blinding: This was an open label study. However, personnel involved in bioanalytical and pharmacokinetic analysis were blinded until the completion of the analysis and reporting of results.



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Bioequivalence study of Abiraterone acetate tablet in Indian healthy adult male volunteers Treatment: In each period, one tablet of either test or reference product was administered with 240 ml of ambient water to each subject in a sitting position in accordance with the SAS generated randomization schedule. Immediately after IP administration in each period, the hands and oral cavity of the subjects were examined using a tongue depressor followed by flashlight to confirm complete medication and fluid consumption. Assessment of safety variables: Safety assessment was done for all subjects who received at least one dose of the study drug. Safety evaluation was based on the critical examination prior to start of the study, laboratory (hematology and biochemistry) tests performed prior to start of the study, repeated during checkout of period II (mid-safety laboratory assessment) and at the end of the study. The serum electrolyte test (sodium, potassium and phosphate) were performed at the check-out of period I and period III. Vitals were assessed at the time of admission to clinical unit, within 02 hours prior to dosing (pre-dose), 02, 04, 08, 10 hours post-dose (within ±30 minutes from the scheduled time) and prior to check-out of each period. Well-being of the subjects was enquired twice a day during housing (2-4 hours post-dose and 10-12 hours post-dose on day 1; 24-26 hours post-dose and 32-34 hours post-dose on day 2) and prior to check-out of each period. ECG was performed between 2-4 hours post-dose in each period and at the end of the study. Blood Sampling The time points chosen for PK assessments are based on the literature for the PK of Abiraterone. The design and PK analyses planned for the study are consistent with FDA Guidance to Industry Bioavailability and Bioequivalence Studies for Orally Administered Drug Products-General Considerations, March 2003. Concentration of Abiraterone in plasma was measured for proper characterization of the PK of the drug. As per protocol, a total of 12 blood samples, 4 ml each, were collected at the following times relative to dosing during each treatment period: the pre-dose (0.00 hours) blood samples were collected within 1 hour prior to drug administration, and the post-dose blood samples were collected at 0.33, 0.67, 1.00, 2.00, 3.00, 6.00, 8.00, 10.00, 24.00, 48.00, 72.00 hours. All the in-house post-dose blood samples were collected at the scheduled time with a window period of ±02 minutes. The 72.00 hours ambulatory blood sample was collected at the scheduled time with a window period of ±01 hour. Prior to drug administration, an intravenous cannula was inserted into forearm of subjects for collection of blood samples and retained upto 24 hours after drug administration. To prevent heparin present in the cannula from interfering with the analysis, 0.5 mL of blood was discarded (up until 24.00 hours) before collection of each blood sample. About 350 mL of blood including screening (about 32 mL) and study sample collection (about 288 mL) was drawn from each subject for the study. The pre-dose and post-dose



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Bioequivalence study of Abiraterone acetate tablet in Indian healthy adult male volunteers blood samples will be collected in pre-chilled, pre-labeled vacutainers containing Sodium Fluoride + Potassium Oxalate (anticoagulant) and were kept in wet ice bath. Sample processing: Centrifugation of samples was started within 30 minutes of blood sample collection. The blood samples were centrifuged at 3500 rpm for 10 minutes at 4oC. The resulting plasma was transferred approximately into two aliquots to pre-chilled, pre-labeled polypropylene tubes containing a suitable stabilizing agent, packed in black polythene bags and stored upright in a container containing dry ice (until transferred into the deep freezer) or in a deep freezer at a temperature of about -70 oC until shipped to the bioanalytical facility of Norwich Clinical Services Pvt. Ltd.

MATERIALS AND METHODS BIOANALYTICAL MEASUREMENTS Quantification of Abiraterone in human plasma (Sodium fluoride+Potassium oxalate) using LC-MS/MS. Table 9: Instruments and equipments Instruments and

Equipments

Manufacturer

Food weighing scale

Satwik

Height meter

Bio plus

Weighing machine

Satwik scale

ECG Machine

BPL

Alcohol Breath Analysis

Uniphos

Lab Centrifuge

REMI

BP apparatus

Diamond

Stop watch

DIMENSIONS

Refrigerate centrifuge

Thermo Scientific

Deep freezer

Thermo Scientific

Digital Temperature Display

Delta system

Weighing Machine

Indo systematic

Data Logger

DIMENSIONS

Master Digital Clock

BRIM

Slave Digital Clock

Accuchek (Roche)

Blood Glucose Monitoring

Pennya Industrial

Laminar Air Flow

Denta system



Page 27


Instruments and

Equipments

Manufacturer

Digital temperature humidity display

While- whisting

Dehumidifier

BRIM

Lab refrigerator

Blue star

Freezer (-20 to -30 oC )

Thermolab

o

Deep freezer (-70 C)

Applied Bio-systems

LCMS/MS-API 4000

Thermolab

Cooling cabinet (2-8 o C)

Caliper life sciences

Nitrogen evaporator

Thermo Scientific

Centrifuge

Ultra sonic cleaner

Vibramax

Heidolph

Sonicator

BRIM

Pipette

-

20 - 200 µl

Eppendorf

100 - 1000 µl

Eppendorf

Repeater pipette

Eppendorf

Statistical Analysis

Software SAS version 9.0 Table 10: Details of chemicals used

Chemicals Used

Manufacturer

Methanol (HPLC grade)

Merck

Formic acid (Analytical grade)

Spectrochem

Ammonium Formate (GR grade)

Spectrochem

TBME (HPLC grade)

Bio-oraganics

Abiraterone (Working standard)

Bio-oraganics

Abiraterone D4 (Internal standard)

Millipore

Milli Q water

Pall life sciences

0.45 µm nylon 6,6 membrane filter

Tarson

Micro tips

Chromatopack

HPLC vials RIA vials


Pall life


Page 28

Bioequivalence study of Abiraterone acetate tablet in Indian healthy adult male volunteers Chemicals Used

Manufacturer

Volumetric Flask

Borosil

Reagent bottles

Borosil

Automatic Pippete

Eppendorf

Acetonitrile (Gradient grade)

Merck

Sodium Dihydrogen Orthophosphate Dihydrate

-

(AR grade) Sodium Fluoride + Potassium Oxalate Plasma

-

Potassium Fluoride

-

Oxalic Acid Dihydrate

-

0.45 µm nylon 6,6 membrane filter paper

-

Preparation of the Reagents: 2 mM Ammonium formate (pH 3.5±0.2) 126.12 mg of ammonium formate was weighed and transferred to a 1000 ml volumetric flask and dissolved in Milli Q water. The pH was adjusted to 3.5±0.2 with formic acid. The volume was made up with the Milli Q water. The solution was filtered through 0.45 µm nylon 6, 6 membrane filter paper in a 1000 ml reagent bottle and sonicated. Mobile Phase (2mM Ammonium formate (pH 3.5±0.2): Acetonitrile:: 30:70 v/v) 300 ml of filtered 2mM ammonium formate pH 3.5±0.2 and 700 ml Acetonitrile (filtered through 0.45 µm nylon 6, 6 membrane filter paper in a 1000 ml reagent bottle) were mixed and sonicated.

Methanol: Water:: 50:50 v/v (Diluent) 500 ml methanol was taken and mixed with 500 ml Milli-Q-water in a 1000 ml reagent bottle and sonicated.

Acetonitrile:Water:: 50:50 v/v (Wash Solvent) 500 ml acetonitrile was taken and mixed with 500 ml Milli-Q-water in a 1000 ml reagent bottle and sonicated.



Page 29

Bioequivalence study of Abiraterone acetate tablet in Indian healthy adult male volunteers 0.1 M Sodium dihydrogen orthophosphate dihydrate 1.5601 g of Sodium dihydrogen orthophosphate dehydrate was weighed and transferred to a 100 ml volumetric flask and dissolved in Milli-Q-water and sonicated. 5% (w/v) Oxalic Acid Dihydrate 5.0 g of oxalic acid dihydrate was weighed and transferred to a 100 ml volumetric flask. It was then dissolved in Milli-Q-water and sonicated. 10% (w/v) Potassium Fluoride in 5% (w/v) Oxalic Acid Dihydrate About 10.0 g of Potassium Fluoride was weighed and and dissolved in 5% w/v Oxalic Acid Dihydrate in 100 ml volumetric flask. The volume was made up with 5% w/v Oxalic Acid Dihydrate and sonicated. Potassium Fluoride stabilized sodium fluoride + potassium oxalate plasma 100 µl of 10% w/v Potassium fluoride in 5% w/v Oxalic Acid Dihydrate was added to 900 µl of pooled sodium fluoride + potassium oxalate plasma. This plasma was stored at -70o C. Preparation of stock solution and stock dilutions: Internal standard stock solution preparation About 2 mg of Abiraterone D4 was weighed, transferred to 2ml volumetric flask and was dissolved in methanol. The volume was made up to 2ml with methanol to give 1000 µg/ml solution. The stock solution was stored in cooling cabinet (2-8o C). Internal standard stock dilution preparation Internal standard stock dilution concentration of 100 ng/ml was prepared from 1000 µg/ml solution of Abiraterone D4 using Methanol:water 50:50 v/v as a diluent. Table 11: Internal standard stock dilutions Stock

Stock Aliquot (ml)

Concentration

Diluent Added

Final Volume (ml)

(ml)

Final Concentration

(µg/ml)

(µg/ml)

1000

0.020

1.980

2.000

10

10

0.500

49.500

50.000

100



Page 30

Bioequivalence study of Abiraterone acetate tablet in Indian healthy adult male volunteers Analyte stock solution preparation for Abiraterone 5 mg of Abiraterone was weighed and transferred to 25 ml of volumetric flask dissolved in methanol. Then the volume was made up to 25 ml with methanol to give 200 µg/ml solution. The stock solution was stored in cooling cabinet (2-8o C). Analyte stock dilution preparation for calibration curve standards for Abiraterone The stock dilutions of Abiraterone was prepared in the concentration range of 9.984 ng/ml to 4000.000 ng/ml using methanol:water (50:50 v/v) as diluent. Table 12: Stock dilutions of Abiraterone acetate Stock

Stock Aliquot (ml)

Diluent Added

Concentration

Final Volume (ml)

(ml)

Final Concentration

(µg/ml)

(µg/ml)

200000

0.200

1.800

2.000

20000

20000

0.400

1.600

2.000

4000

4000

1.600

0.400

2.000

3200

3200

1.200

0.800

2.000

1920

1920

1.220

0.780

2.000

1171.20

1171

1.000

1.000

2.000

585.60

585

0.620

1.380

2.000

181.536

181

0.220

1.780

2.000

19.969

19.969

1.000

1.000

2.000

9.984

Analyte stock dilution preparation for Quality Control for Abiraterone The stock dilutions of Abiraterone were prepared in the concentration range of 10.125 ng/ml to 3000.000 ng/ml using Methanol:water (50:50 v/v) as diluent. Table 13: Stock dilutions for QC Stock

Stock Aliquot (ml)

Concentration

Diluent Added

Final Volume (ml)

(ml)

Final Concentration

(µg/ml)

(µg/ml)

200000

0.20

1.80

2

20000

20000

0.30

1.70

2

3000

3000

0.80

1.20

2

1200



Page 31

Bioequivalence study of Abiraterone acetate tablet in Indian healthy adult male volunteers Stock

Stock Aliquot (ml)

Concentration

Diluent Added

Final Volume (ml)

(ml)

Final Concentration

(µg/ml)

(µg/ml)

1200

0.30

1.70

2

180

180

0.30

1.70

2

27

27

0.75

1.25

2

10.125

Spiking preparation for calibration curve standards for Abiraterone The spiked concentration of analyte was prepared in the concentration range of about 0.2 ng/ml to 8 ng/ml for Abiraterone by spiking 20 µl of each calibration curve standard of Abiraterone into 980 µl of Potassium fluoride stabilized Sodium fluoride + potassium oxalate plasma as below. Table 14: Spiking preparation Stock Concentration

Final Spiked Concentration

CCID

Abiraterone

Abiraterone

(µg/ml)

(ng/ml)

4000.000

80.000

H

3200.000

64.000

G

1920.000

38.400

F

1171.200

23.424

E

585.600

11.712

D

181.536

3.631

C

19.969

0.399

B

9.984

0.200

A

* This is just an example. Actual concentration may vary as per potency and stock weight. 0.300 ml of each CC was aliquoted into RIA vials. The bulk spiked samples were stored in deep freezer at about -70 o C. Spiking preparation for quality control for Abiraterone The spiked concentration of analyte was prepared in the concentration range of about 0.203 ng/ml to 60.000 ng/ml for Abiraterone into 980 µl of Potassium fluoride stabilized sodium fluoride + potassium oxalate plasma as below.



Page 32

Bioequivalence study of Abiraterone acetate tablet in Indian healthy adult male volunteers Table 15: Spiking preparation for QC Stock Concentration

Final Spiked Concentration

QCID

Abiraterone

Abiraterone

(µg/ml)

(ng/ml)

3000.000

60.000

HQC

1200.000

24.000

MQC

27.000

0.540

LQC

10.125

0.203

LOQQC

* This is just an example. Actual concentration may vary as per potency and stock weight. 0.300 ml of each CC was aliquoted into RIA vials. The bulk spiked samples were stored in deep freezer at about -70 o C. Preparation of aqueous standard 20 µl of analyte stock solution was taken in 250 µl of Internal standard. Then the volume was made up to 1.500 ml with mobile phase. * Stock weighing, stock dilutions and spiking are done under yellow monochromatic light. Instrument parameters and chromatographic conditions Instrument : API 4000 Mode: Positive API Column: BDS-Hypersil C18 100×4.6 mm, 5µm Injection Volume: 20.0 µl Flow rate: 1.2 ml/min with splitter Run time: 4.0 minutes Column oven temperature: 40 o C Auto Sampler temperature: 10 o C



Page 33

Bioequivalence study of Abiraterone acetate tablet in Indian healthy adult male volunteers Table 16: Various parameters of instrument MS Parameters Abiraterone Abiraterone D4

Temperature

500

Q1

350.3

354.3

GS1

45

Q3

156.1

160.1

GS2

55

DP

90

90

CAD

8

EP

10

10

Dwell time

400 min

CE

65

65

CUR

30

CXP

15

15

IS

550

Sample Preparation i.

Required set of calibration curve standards, quality control samples along with subject samples were withdrawn from deep freezer/ freezer.

ii.

The thawed samples were vortexed for complete mixing.

iii.

50 µl of internal standard (Abiraterone D4) was added to all RIA vials except blank.

iv.

200 µl of sample was added to respectively labelled RIA vials and they were vortexed.

v.

100 µl of 0.1 M Sodium dihydrogen orthophosphate dihydratewas added to respectively labelled RIA vials and were vortexed.

vi.

2 ml of TBME was added to all the samples and vials were capped.

vii.

The samples were kept at Vibramax at 2500 RPM for 10 mins.

viii.

The samples were centrifuged at 4000 RPM for 5 mins in refrigerated centrifuge at about 4oC.

ix.

1.6 ml of supernatant was transferred into repectively labelled RIA vials.

x.

The samples were allowed to dry at 40 oC in nitrogen evaporator.

xi.

The samples were reconstituted with 0.3 ml of mobile phase and vortexed.

xii.

The samples were transferred into respectively labelled HPLC vials and then loaded into autosampler. * Sample processing was done under yellow monochromatic light. Thawing of samples were done in CCB.

Processing of data Chromatograms were acquired by using computer based Analyst software. Sample concentration were calculated by linear regression analysis using the analyst software. Data was processed by peak area ratio. The concentration of unknown was calculated from the following equation using regression analysis of spiked plasma calibration standards with reciprocal of the square of the drug concentration (1/X2). Birla Institue Technology Mesra, Ranchi, India


Page 34

Bioequivalence study of Abiraterone acetate tablet in Indian healthy adult male volunteers y = mx + c Where, x=concentration of analyte m=slope of the calibration curve y=peak area ratio of analyte to internal standard c=y axis intercept of the calibration curve

Table 17: Individual plasma Concentration (ng/ml) for Period I for Abiraterone Calibration Curve Range 0.210 ng/ml to 78.016 ng/ml Time points

0.00

0.33

0.67

1.00

2.00

3.00

6.00

8.00

10.00

24.00

48.00

72.00

Subject 01

0

0.748

19.335

20.634

7.932

4.954

1.543

1.138

1.331

0.712

0

0

Subject 02

0

0

4.010

6.453

37.116

33.782

11.356

5.887

3.678

1.521

0.521

0

Subject 03

0

0

1.264

4.566

10.526

12.986

9.008

6.287

4.126

0.867

211.0

0

Subject 04

0

0

0.409

0.897

24.607

66.540

16.933

9.121

4.586

4.061

0

0

Subject 05

0

0.295

1.968

5.343

22.450

18.710

8.932

4.991

2.850

0.449

0

0

Subject 06

0

0.052

0.569

1.560

4.610

12.384

30.290

13.252

7.132

1.537

0

0

Subject 07

0

0

3.703

14.210

16.804

18.608

4.056

2.441

1.847

0.545

0

0

Subject 08

0

0.102

12.040

29.655

35.326

18.943

9.833

7.894

5.313

1.234

0.301

0



Page 35


Table 18: Individual plasma Concentration (ng/ml) for Period II for Abiraterone Calibration Curve Range 0.210 ng/ml to 78.016 ng/ml Time Points

0.00

0.33

0.67

1.00

2.00

3.00

6.00

8.00

10.00

24.00

48.00

72.00

Subject 01

0

2.456

45.771

58.672

23768.0

12.456

6567.0

5456.0

3.221

1.567

0.467

0

Subject 02

0

0.399

4.765

10.110

37.128

30.278

15.756

8991.0

6.536

2.999

0.416

0

Subject 03

0

0

2.130

4.870

11.770

16.989

11.866

8.976

5.768

1.339

0.435

0

Subject 04

0

0

1.567

4.665

8.561

35.789

18.216

8.656

5.928

0.701

0

0

Subject 05

0

0

1.678

6.372

25.678

34.618

15.719

3.991

2.617

0.678

0.231

0

Subject 06

0

0

0.456

4.129

17.990

22.567

26.872

15.183

7.125

1.456

0.298

0

Subject 07

0

0.789

5738.0

25.790

16.987

13.578

5.008

2.465

1.562

0.593

0

0

Subject 08

0

1.672

19.725

16.218

15.579

9.667

4.790

2.138

1.598

0.999

0.245

0

Table 19: Individual plasma Concentration (ng/ml) for Period III for Abiraterone Calibration Curve Range 0.210 ng/ml to 78.016 ng/ml Time Points

0.00

0.33

0.67

1.00

2.00

3.00

6.00

8.00

10.00

24.00

48.00

72.00

Subject 01

0

0.867

14.557

20.678

9.554

5.009

1.878

1.284

0.853

0.456

0.216

0

Subject 02

0

0

0.253

1.598

6.345

10.234

18.4567

9.475

5.898

2.456

0.388

0

Subject 03

0

0

0.241

1.357

10.198

23.547

16.448

9.556

4.809

1.712

0.391

0

Subject 04

0

0

0.987

3.648

36.853

20.567

14.719

7.781

4.243

1.321

0.452

0

Subject 05

0

0

0.345

1.632

9.22

57.39

12.862

6.011

3.456

0.743

0.289

0

Subject 06

0

0.337

1.647

3.647

22.478

20.8732

15.138

9.3978

5.8374

1.346

0.349

0

Subject 07

0

0.904

7.864

11.534

18.777

16.062

8.6789

6.113

3.634

0.867

0.254

0

Subject 08

0

0.212

8.887

31.098

33.667

18.116

4.998

3.218

2.678

0.798

0

0



Page 36


Table 19: Individual plasma Concentration (ng/ml) for Period IV for Abiraterone Calibration Curve Range 0.210 ng/ml to 78.016 ng/ml Time Points

0.00

0.33

0.67

1.00

2.00

3.00

6.00

8.00

10.00

24.00

48.00

72.00

Subject 01

0

0.584

46.288

49.775

12.764

6.908

2.365

1.898

1.567

0.567

0

0

Subject 02

0

1.004

2.8878

6.745

38.765

26.346

7.387

5.198

2.874

1.2645

0.509

0

Subject 03

0

1.273

4.675

10.129

7.283

5.987

3.345

2.019

1.145

0.290

0

0

Subject 04

0

0.984

7.376

14.389

26.364

50.274

13.283

6.345

3.489

1.384

0.267

0

Subject 05

0

0

3.236

11.384

27.338

19.329

6.273

4.473

2.084

0.456

0

0

Subject 06

0

1.118

3.037

6.287

6.478

5.123

12.384

7.283

3.992

0.982

0.352

0

Subject 07

0

1.127

12.098

18.231

36.128

42.428

5.963

3.387

2.374

0.723

0

0

Subject 08

0

0

3.045

7.265

21.389

23.478

7.472

4.004

2.761

1.283

0.289

0



Page 37


RESULTS PHARMACOKINETIC ANALYSIS Plasma concentration vs. Time graphs were plotted for each subjects (for respective subjects) using Microsoft Excel Version 2007. Pharmacokinetic analysis on the plasma concentrations for Abiraterone was carried out for the test and reference products to determine the following PK parameters in each individual subject. Primary variables: Cmax: Maximum measured plasma concentration following each treatment. AUC0-t: The area under plasma concentration versus time curve from 0 hours to last measurable concentration as calculated by linear trapezoidal method. AUC 0-∞: The area under plasma concentration versus time curve from 0 hours to infinity. Where AUC0-∞ = AUC 0-t + Ct/Kel, Ct is the last measurable concentration and Kel is the terminal elimination rate constant. Secondary Variable: Tmax: Time of the maximum measured plasma concentration. t1/2: The elimination or terminal half-life will be calculated as 0.693/Kel Kel: First order rate constant associated with the terminal portion of the curve. This is estimated via linear regression of time vs log concentration. This parameter has been calculated by using linear least square regression analysis using at least last three or more non zero plasma concentration values.



Page 38

Bioequivalence study of Abiraterone acetate tablet in Indian healthy adult male volunteers Calculation of Pharmacokinetic Parameters using Microsoft Excel Version 2007 PERIOD I Table 21: Calculations of pharmacokinetic parameters of subject 1 in period I

Time (t) 0 0.33 0.67 1 2 3 6 8 10 24 48 72

PERIOD 1

SUB 1

Concentration (c) 0 0.748 19.335 20.634 9.932

c1+c2 0.748 20.083 39.969 30.566 14.886

dt 0.33 0.34 0.33 1 1

AUC- linear 0.12342 3.41411 6.594885 15.283 7.443

lnC -0.290352301 2.961916925 3.026940201 2.295761868

4.954 2.543

7.497 4.131

3 2

11.2455 4.131

1.600195331 0.933344486

1.588 0.712 0.331 0 0

2.3 1.043 0.331 0 0

2 14 24 24 -72 AUC for SUB1 (P1)

2.3 7.301 3.972 0 0 61.807915

0.462475363 -0.339677368 -1.105636904 -

c1 and c2are preceeding and succeeding concentrations of the trapezium, dt is the change in time interval, AUC is area under the concentration vs. time curve, lnC is natural logarithm of the concentrations

Figure 5: Concentration vs. Time plot for subject 1 in period I



Page 39

Bioequivalence study of Abiraterone acetate tablet in Indian healthy adult male volunteers Cmax = 20.634 ng/ml Tmax= 1 hour Kel= 0.6491 h−1 t1/2= 1.067632 h AUC 0-t = 61.807915 ng mL-1h-1 AUC0-∞ = 62.31785184 ng mL-1h-1

Slope of the line = - Elimination rate constant (Ke) Figure 6: ln C vs. Time plot for subject 1 in period I



Page 40

Bioequivalence study of Abiraterone acetate tablet in Indian healthy adult male volunteers Table 22: Calculations of pharmacokinetic parameters of subject 2 in period I PERIOD 1

SUB 2

Time (t) 0 0.33

Concentration (c) 0 0

c1+c2 0 4.01

Dt 0.33 0.34

AUC- linear 0 0.6817

lnC -

0.67 1

4.01 6.453

10.463 44.569

0.33 1

1.726395 22.2845

1.388791241 1.864545139

2 3 6 8 10 24 48 72

38.116 22.782 10.356 5.887 3.678 1.521 0.521 0

60.898 33.138 16.243 9.565 5.199 2.042 0.521 0

1 3 2 2 14 24 24 -72 AUC for SUB2 (P1)

30.449 49.707 16.243 9.565 36.393 24.504 6.252 0 197.805595

3.640634141 3.12597075 2.337566062 1.77274653 1.302369126 0.419368013 -0.652005237 -





Page 41

Bioequivalence study of Abiraterone acetate tablet in Indian healthy adult male volunteers Cmax = 20.634 ng/ml Tmax= 1 hour Kel= 0.6326 h−1 t1/2= 1.067632 h AUC 0-t = 61.807915 ng mL-1 h−1 AUC0-∞ = 62.31785184 ng mL-1 h−1




Page 42


SUB 3

Time (t) 0

Concentration (c) 0

c1+c2 0

dt 0.33

AUC- linear 0

lnC -

0.33 0.67

0 1.264

1.264 5.83

0.34 0.33

0.21488 0.96195

0.234281296

1 2 3 6 8 10 24 48 72

4.566 10.526 12.986 9.008 6.287 4.126 0.867 0.211 0

15.092 23.512 21.994 15.295 10.413 4.993 1.078 0.211 0

1 1 3 2 2 14 24 24 -72 AUC for SUB3 (P1)

7.546 11.756 32.991 15.295 10.413 34.951 12.936 2.532 0 129.59683

1.518637548 2.353848387 2.563871854 2.198113071 1.838484009 1.417308415 -0.142716302 -1.555897146 -





Page 43

Bioequivalence study of Abiraterone acetate tablet in Indian healthy adult male volunteers Cmax = 12.986 ng/ml Tmax= 3 hour Kel = 0.3799 h−1 t1/2 = 1.824164 h AUC 0-t = 129.59683 ng mL-1 h−1 AUC 0-∞ = 130.1522 ng mL-1 h−1




Page 44


Table 24: Calculations of pharmacokinetic parameters of subject 4 in period I

Time (t) 0 0.33 0.67 1 2 3 6 8 10 24 48 72

PERIOD 1

SUB 4

Concentration (c) 0 0

c1+c2 0 0.409

dt 0.33 0.34

AUC- linear 0 0.06953

lnC -

0.409 0.897

1.306 25.504

0.33 1

0.21549 12.752

-0.894040123 -0.108699417

24.607 46.54 24.933 10.121 6.586 2.961 0 0

71.147 71.473 35.054 16.707 9.547 2.961 0 0

1 3 2 2 14 24 24 -72 AUC for SUB4 (P1)

35.5735 107.2095 35.054 16.707 66.829 35.532 0 0 309.94202

3.203030955 3.840312158 3.216192227 2.314612473 1.884946184 1.085527049 -





Page 45


Cmax = 46.54 ng/ml Tmax= 3 hour Kel= 0.6841 h−1 t1/2= 1.01301 h AUC 0-t = 309.94202 ng mL-1 h−1 AUC 0-∞ = 314.2703 ng mL-1 h−1




Page 46


SUB 5

Time (t) 0 0.33 0.67

Concentration (c) 0 0.295 1.968

c1+c2 0.295 2.263 7.311

dt 0.33 0.34 0.33

AUC- linear 0.048675 0.38471 1.206315

lnC -1.220779923 0.677017799

1 2

5.343 22.45

27.793 41.16

1 1

13.8965 20.58

1.675787293 3.111290614

3 6 8 10 24 48 72

18.71 8.932 4.991 2.85 0.449 0 0

27.642 13.923 7.841 3.299 0.449 0 0

3 2 2 14 24 24 -72 AUC for SUB5 (P1)

41.463 13.923 7.841 23.093 5.388 0 0 127.8242

2.92905814 2.189640334 1.60763629 1.047318994 -0.800732391 -





Page 47

Bioequivalence study of Abiraterone acetate tablet in Indian healthy adult male volunteers Cmax = 22.45 ng/ml Tmax= 2 hour Kel= 0.5449 h−1 t1/2= 1.271793 h AUC 0-t = 127.8242 ng mL-1 h−1 AUC 0-∞ = 128.6482 ng mL-1 h−1




Page 48


SUB 6

Time (t) 0 0.33 0.67

Concentration (c) 0 0.052 0.569

c1+c2 0.052 0.621 2.129

dt 0.33 0.34 0.33

AUC- linear 0.00858 0.10557 0.351285

lnC -2.95651156 -0.563874845

1 2

1.56 4.61

6.17 16.994

1 1

3.085 8.497

0.444685821 1.528227857

3 6 8 10 24 48 72

12.384 30.29 13.252 7.132 4.537 0 0

42.674 43.542 20.384 11.669 4.537 0 0

3 2 2 14 24 24 -72 AUC for SUB6 (P1)

64.011 43.542 20.384 81.683 54.444 0 0 276.111435

2.516405317 3.410817625 2.584148484 1.9645917 1.512266001 -





Page 49





Page 50

Bioequivalence study of Abiraterone acetate tablet in Indian healthy adult male volunteers Period II Table 27: Calculations of pharmacokinetic parameters of subject 1 in period II

Time (t) 0 0.33 0.67 1 2 3 6 8 10 24 48 72

PERIOD 2

SUB 1

Concentration (c) 0 2.456 45.771 58.672 28.768

c1+c2 2.456 48.227 104.443 87.44 44.224

dt 0.33 0.34 0.33 1 1

AUC- linear 0.40524 8.19859 17.233095 43.72 22.112

lnC 0.89853401 3.823650703 4.071962611 3.359263658

15.456 7.567 3.456 2.002 1.002 0.234 0

23.023 11.023 5.458 3.004 1.236 0.234 0

3 2 2 14 24 24 -72 AUC for SUB1 (P2)

34.5345 11.023 5.458 21.028 14.832 2.808 0 181.352425

2.737997277 2.023796688 1.240111851 0.694146681 0.001998003 -1.45243416 -


Figure 17: Concentration vs. Time plot for subject 1 in period II



Page 51


Slope of the line = - Elimination rate constant (Ke) Figure 18: ln C vs. Time plot for subject 1 in period II



Page 52

Bioequivalence study of Abiraterone acetate tablet in Indian healthy adult male volunteers Table 28: Calculations of pharmacokinetic parameters of subject 2 in period II PERIOD 2

SUB 2

Time (t) 0 0.33 0.67 1

Concentration (c) 0 0.399 4.765 10.11

c1+c2 0.399 5.164 14.875 47.238

dt 0.33 0.34 0.33 1

2 3 6 8 10 24 48 72

37.128 30.278 15.756 8.991 6.536 2.999 0.416 0

67.406 46.034 24.747 15.527 9.535 3.415 0.416 0

1 3 2 2 14 24 24 -72 AUC for SUB2 (P2)

AUC- linear 0.065835 0.87788 2.454375 23.619 33.703 69.051 24.747 15.527 66.745 40.98 4.992 0 282.76209

lnC -0.91879386 1.561297537 2.313525033 3.614371402 3.410421376 2.757221245 2.196224077 1.877325358 1.0982789 -0.87707002 -





Page 53





Page 54


SUB 3

Time (t) 0 0.33 0.67 1

Concentration (c) 0 0 2.13 4.87

c1+c2 0 2.13 7 16.64

dt 0.33 0.34 0.33 1

AUC- linear 0 0.3621 1.155 8.32

lnC 0.75612198 1.583093937

2 3 6 8 10 24 48 72

11.77 16.989 11.866 8.976 5.768 1.339 0.435 0

28.759 28.855 20.842 14.744 7.107 1.774 0.435 0

1 3 2 2 14 24 24 -72 AUC for SUB3 (P2)

14.3795 43.2825 20.842 14.744 49.749 21.288 5.22 0 179.3421

2.465553921 2.832566076 2.473677168 2.194554349 1.7523254 0.291923067 -0.83240925 -





Page 55





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SUB 4

Time (t) 0 0.33 0.67 1


c1+c2 0 1.567 6.232 13.226

dt 0.33 0.34 0.33 1

AUC- linear 0 0.26639 1.02828 6.613

lnC 0.449162963 1.540087834

2 3 6 8 10 24 48 72

8.561 35.789 18.216 8.656 4.928 0.701 0 0

44.35 54.005 26.872 13.584 5.629 0.701 0 0

1 3 2 2 14 24 24 -72 AUC for SUB4 (P2)

22.175 81.0075 26.872 13.584 39.403 8.412 0 0 199.36117

2.147217006 3.577640584 2.902300329 2.158252722 1.594933226 -0.35524739 -





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Page 58


SUB 5

Time (t) 0 0.33 0.67 1


c1+c2 0 1.678 8.05 32.05

dt 0.33 0.34 0.33 1

AUC- linear 0 0.28526 1.32825 16.025

lnC 0.517602608 1.851913392

2 3 6 8 10 24 48 72

25.678 34.618 16.719 8.991 4.817 0.678 0.231 0

60.296 51.337 25.71 13.808 5.495 0.909 0.231 0

1 3 2 2 14 24 24 -72 AUC for SUB5 (P2)

30.148 77.0055 25.71 13.808 38.465 10.908 2.772 0 216.45501

3.245634594 3.544373778 2.816545797 2.196224077 1.572151328 -0.38860799 -1.46533757 -





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Page 60


SUB 6

Time (t) 0 0.33 0.67 1


c1+c2 0 0.456 4.585 22.119

dt 0.33 0.34 0.33 1

AUC- linear 0 0.07752 0.756525 11.0595

lnC -0.785 1.418

2 3 6 8 10 24 48 72

17.99 22.567 26.872 15.183 8.125 4.456 0.298 0

40.557 49.439 42.055 23.308 12.581 4.754 0.298 0

1 3 2 2 14 24 24 -72 AUC for SUB6 (P2)

20.2785 74.1585 42.055 23.308 88.067 57.048 3.576 0 320.384545

2.889 3.116 3.291 2.720 2.094 1.494 -1.210 -





Page 61





Page 62

Bioequivalence study of Abiraterone acetate tablet in Indian healthy adult male volunteers Table 33: Primary and Secondary parameters that were analysed for the Reference product Primary parameter

Subject No. Cmax(ng/ml)

Secondary parameter

AUC 0-t

AUC 0-inf

(hr*ng/ml)

(hr*ng/ml)

Tmax (hr)

t1/2

Kel

1

58.672

181.352

181.696

1.00

1.019

0.679

2

38.116

197.805

198.629

2.00

1.095

0.632

3

12.986

129.596

130.152

3.00

1.824

0.379

4

35.789

199.361

200.409

3.00

1.035

0.669

5

34.618

216.455

216.808

3.00

1.060

0.653

6

30.290

276.111

283.295

6.00

1.097

0.631

Table 34: Primary and Secondary parameters that were analysed for the Test product Primary parameter

Subject No. Cmax(ng/ml)

Secondary parameter

AUC 0-t

AUC 0-inf

(hr*ng/ml)

(hr*ng/ml)

Tmax (hr)

t1/2

Kel

1

20.634

61.807

62.317

1.00

1.067

0.649

2

37.128

282.762

283.649

2.00

1.478

0.468

3

16.989

179.342

180.577

3.00

1.968

0.352

4

46.540

309.942

314.270

3.00

1.013

0.684

5

22.450

127.824

128.648

2.00

1.271

0.544

6

26.872

320.384

320.879

6.00

1.151

0.601



Page 63

Bioequivalence study of Abiraterone acetate tablet in Indian healthy adult male volunteers Figure 29: Pair wise presentation of individual plasma Abiraterone concentration vs. time curves following a single dose of 250 mg Abiraterone acetate. Subject 1

Subject 2



Page 64


Subject 3

Subject 4



Page 65

Bioequivalence study of Abiraterone acetate tablet in Indian healthy adult male volunteers Subject 5

Subject 6

For statistical analysis purpose, the number of subjects with randomization code T (Test) and R (Reference) should be same. Hence, the last two subjects (subject 7 and subject 8) have not been considered for calculation purpose.



Page 66

Bioequivalence study of Abiraterone acetate tablet in Indian healthy adult male volunteers Analysis of variance (ANOVA) was performed on the untransformed pharmacokinetic parameters AUC 0-t, AUC0-∞, Cmax and Tmax. Additionally, log transformed data was used for analysis of AUC 0-t, AUC0-∞, Cmax and Tmax. The analysis of variance model included sequence, subjects tested within sequence, period and drug formulation as factors. Ratio analysis was reported for untransformed and log-transformed data. For bioequivalence evaluation, in accordance with current FDA guidelines. The products (reference and test) were considered bioequivalent if 90% confidence intervals for Cmax, AUC0-t and AUC0-∞ fell within the range of 80 % to 125 %. Table 35: ANOVA for Cmax Source Between Within Total

SS (Sum of Square)

DF (Degree of freedom)

MS (Mean Square)

F ratio

9.95E-02 1.951497861 2.05E+00

1 10 11

9.95E-02 0.195149786

5.10E-01

Table 36: ANOVA for AUC 0-t Source Between Within Total

SS (Sum of Square)

DF (Degree of freedom)

MS (Mean Square)

F ratio

9.13E-03 2.392 2.40E+00

1 10 11

9.13E-03 0.239

3.82E-02

Table 37: ANOVA for AUC 0-∞ Source Between Within Total

SS (Sum of Square) 4.52E-02 3.526 3.57E+00

DF (Degree of freedom) 1 10 11


MS (Mean Square) 4.52E-02 0.352


F ratio 1.28E-01

Page 67

Bioequivalence study of Abiraterone acetate tablet in Indian healthy adult male volunteers Calculation for analysis of Cmax Difference = Test mean – Reference mean = 3.286 – 3.468 = -0.182 Standard Error (SE) difference = 0.255 Cmax ratio = 100 × e-0.182 = 100 × 0.833 = 83.3 % 90% Confidence interval (C.I.) limits: Lower limit = 100 × e(-0.182 – 1.812 × 0.355) = 100 × 0.525 = 52.5% Upper limit = 100 × e(-0.182 + 1.812 × 0.355)

= 100 × 1.323 = 132.3%

Calculation for analysis of AUC 0-t Difference = Test mean – Reference mean = 5.219 – 5.274 = -0.055 Standard Error (SE) difference = 0.282 AUC 0-t ratio = 100 × e-0.055= 100 × 0.946 = 94.6 % 90% Confidence interval (C.I.) limits: Lower limit = 100 × e(-0.055 – 1.812 × 0.282) = 100 × 0.567 = 56.7% Upper limit = 100 × e(-0.055 + 1.812 × 0.282) = 100 × 1.578 = 157.8%



Page 68

Bioequivalence study of Abiraterone acetate tablet in Indian healthy adult male volunteers Calculation for analysis of AUC 0-inf Difference = Test mean – Reference mean = 5.103– 5.225 = -0.122 Standard Error (SE) difference = 0.342 AUC 0-inf ratio = 100 × e-0.122 = 100 × 0.884 = 88.4 % 90% Confidence interval (C.I.) limits: Lower limit = 100 × e(-0.122– 1.812 × 0.342) = 100 × 0.475 = 47.5% Upper limit = 100 × e(-0.122 + 1.812 × 0.342) = 100 × 1.646 = 164.6% The main objective of the study was to compare the bioavailability and characterize the PK profile of the test formulation of Abiraterone Acetate Tablets 250 mg (manufactured by X Pharmaceutical) relative to that of reference formulation, Abiraterone Acetate Tablets (manufactured by Y Pharmaceutical) in healthy, adult, human male subjects under fasting conditions and to assess the bioequivalence of the two formulations. The ratio and 90% confidence intervals for AUC0-t, AUC 0-∞ and Cmax for test and reference products were found to be in between 80.00% to 125.00%. Table 38: Confidence interval for the ratio of test and reference product PK Parameters

Ratio (%) Test/Reference

90% CI for Ratio (%)

Cmax (ng/mL)

83.3 %

52.5 – 132.3 %

AUC0-t (ng×hr/mL)

94.6 %

56.7 - 157.8 %

AUC0-∞ (ng×hr/mL)

88.4 %

47.5 - 164.6 %



Page 69

Bioequivalence study of Abiraterone acetate tablet in Indian healthy adult male volunteers DISCUSSION AND CONCLUSION The study was planned and conducted as open-label, randomised, four-period, two-treatment, two-sequence, crossover, single dose, bioequivalence under fasting condition. The primary objective of the present study was to assess the bioequivalence of Abiraterone acetate tablet in healthy adult male volunteers under fasting condition. The design of study was adequate to determine the pharmacokinetic end points of the test and reference formulation. The wash out period of 7 days was maintained to avoid any carry over effects. The study was performed in compliance with ICH- GCP guidelines, World Medical Association Declaration of Helsinki Seoul, 2008 and US FDA’s Guidance for industry: Bioavailability and Bioequivalence studies of orally administered drug products-General consideration, March 2003. As per the protocol, samples from all volunteers were analyzed in the Bioanalytical laboratory to determine the plasma drug concentration. The quantification of drug in plasma samples was performed in accordance with GLP requirements using LCMS-MS allowing specific and sensitive determination of Abiraterone acetate in plasma. The bioequivalence evaluation was based on the intra-individual ratios of the log transformed primary pharmacokinetic parameters Cmax, AUC0-t and AUC0-∞. and comparative bioavailability of Abiraterone acetate was assessed. Ratios of Geometric Least Square Means of Cmax, AUC0-t and AUC0- was been found to be 83.3%, 94.6% and 88.4% respectively. Confidence interval (CI) for Cmax, AUC0-t and AUC0- has been found to 52.5 – 132.3%, 56.7 - 157.8 % and 47.5 - 164.6 % respectively. It can be concluded that the Abiraterone acetate tablet of X Pharmaceuticals Limited, India has been found to be bioequivalent compared to the reference product (Abiraterone Acetate Tablets 250 mg) manufactured by Y Pharmaceutical on healthy, adult, males under fasting condition in terms of both rate and extent of absorption after administration of single oral dose.



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Bioequivalence study of Abiraterone acetate tablet in Indian healthy adult male volunteers REFERENCES 1. Rathkopf DE, Smith MR, Bono JS, Logothetis CJ, Shore ND, Sauza P et al. Updated Interim Efficacy Analysis and Long-term Safety of Abiraterone Acetate in Metastatic Castration-resistant Prostate Cancer Patients Without Prior Chemotherapy (COU-AA-302). European Urology, 2014; 66: 815 – 825. 2. http://www.drugs.com/cdi/abiraterone-acetate.html Date accessed: 25.11.2014

3. Prescribing Information of Zytiga. Available http://www.zytiga.com/sites/default/files/pdf/full_product_information.pdf Date 25.11.2014

from: accessed:

4. Summary of Product characteristics. Available http://www.ema.europa.eu/docs/en_GB/document_library/EPAR__Product_Information/human/002321/WC500112858.pdf Date accessed: 26.11.2014

from:

5. http://www.webmd.com/drugs/2/drug-155807/abiraterone-oral/details Date accessed: 23.11.2014

6. Material Safety data sheet, Clearsinth. Available http://www.drugbank.ca/system/msds/DB05812.pdf?1368946350 Date accessed: 02.12.2014

from:

7. Clinical Pharmacology and Biopharmaceutics review. Available http://www.accessdata.fda.gov/drugsatfda_docs/nda/2011/202379Orig1s000ClinPharmR.pdf accessed: 02.12.2014

from: Date

8. Inoue K et al, Pharmacokinetics of abiraterone in healthy Japanese men: dose proportionality and effect of food timing, Cancer chemotherapy and Pharmacology, 2014. Available from: http://link.springer.com/article/10.1007%2Fs00280-014-2616-4 Date accessed: 26.11.2014 9. Product Monograph of Zytiga (Abiraterone acetate). Available http://www.janssen.ca/subcategory_docdownload?id=1874 Date accessed: 23.11.2014 10. A report from American Cancer Society. Available http://www.cancer.org/acs/groups/cid/documents/webcontent/003134-pdf.pdf Date 01.12.2014

from:

from: accessed:

11. http://www.pcf.org/atf/cf/%7B7c77d6a2-5859-4d60-af47 132fd0f85892%7D/INTRO_PROSTATE_CANCER_GUIDE.PDF Date accessed: 23.11.2014 12. Guidelines on Prostate Cancer. European Assciation of Urology. Available http://www.uroweb.org/gls/pdf/09_Prostate_Cancer_LR.pdf Date accessed: 24.11.2014



from:

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Bioequivalence study of Abiraterone acetate tablet in Indian healthy adult male volunteers 13. Prostate Cancer: Diagnosis and Treatment, National Intitute for Health and Care excellance. Available from: http://www.nice.org.uk/guidance/cg175/resources/guidance-prostate-cancerdiagnosis-and-treatment-pdf Date accessed: 25.11.2014 14. http://www.ich.org/fileadmin/Public_Web_Site/ABOUT_ICH/Organisation/GCC/Top ics_under_Harmonisation/Bioequivalence.pdf Date accessed: 25.11.2014

15. https://www.quinta.cz/specialni-populace-subjektu--42/ Date accessed: 07.12.2014

16. Sternberg CN, Castellano D, Daugaard G, Geczi L et al. Abiraterone acetate for patients with mCRPC progression after chemotherapy: final analysis of a multicentre, open label, early access protocol trial, The Lancet Oncology. 2014; 15(11): 1263 – 1268. 17. Matsubara N, Uemura H, Fukui I, Niwakawa M, Yamaguchi A, Iizuka K, Akaza H et al. Phase1study of abiraterone acetate in chemotherapynaïve Japanese patients with Castrationresistant prostate cancer. Cancer Sci. 2014; 44(12): 1313–1320. 18. Satoh T, Uemura H, Tanabe K, Nishiyama T, Terai A, Yokomizo A et al. A Phase 2 Study of Abiraterone Acetate in Japanese Men with Metastatic Castration resistant Prostate Cancer Who Had Received Docetaxel based Chemotherapy. Jpn. J. Clin. Oncol. 2014; 44(12): 1316-1319.

19. Ryan CJ, Peng W, Kheoh T, Welkowsky E, Haqq CM, Chandler DW et al. Androgen dynamics and serum PSA in patients treated with Abiraterone acetate, Prostate Cancer and Prostatic Disease. 2014; 17: 192–198. 20. Goodman OB, Flaig TW, Molina A, Mulders PF, Fizazi K, Suttmann H et al. Exploratory analysis of the visceral disease subgroup in a phase III study of Abiraterone acetate in metastatic castration resistant prostate cancer, Prostate Cancer and Prostatic Disease. 2014; 17: 34–39.

21. Amit RL, Seah J, Atenafu EG, Templeton AJ, Verabadillo FE, Alimohamed N et al. Abiraterone acetate in metastatic castration resistant prostate cancer: A retrospective review of the Princess Margaret experience of (I) low dose Abiraterone and (II) prior ketoconazole. European Journal of Cancer. 2014; 50(14): 2399-2407. 22. Basch E, Autio K, Ryan CJ, Muilders P, Shore N, Kheoh T et al. Abiraterone acetate plus prednisone versus prednisone alone in chemotherapy naive men with metastatic castration resistant prostate cancer: patient reported outcome results of a randomised phase 3 trial. Lancet Oncol. 2013; 14(12): 1193-1199. 23. Harshman L, Taplin M, Abiraterone Acetate: Targeting Persistent Androgen Dependence in Castration-Resistant Prostate Cancer. Adv Ther. 2013; 30:727–747.



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Bioequivalence study of Abiraterone acetate tablet in Indian healthy adult male volunteers 24. Fizazi K, Scher HI, Molina A, Logothetis CJ, Chi KN, Jones RJ et al. Abiraterone acetate for treatment of metastatic castration resistant prostate cancer: final overall survival analysis of the COUAA301 randomised, double blind, placebo controlled phase 3 study. The Lancet Oncology. 2012; 13(10): 983-992. 25. Acharya M, Bernard A, Gonzalez M, Jiao J, Vries R et al. Open label, phase I, pharmacokinetic studies of abiraterone acetate in healthy men, Cancer Chemother Pharmacol. 2012; 69(6):1583–1590. 26. http://www.accessdata.fda.gov/drugsatfda_docs/label/2014/202379s0091b1.pdf 12.02.2015

Date

accessed:

27. http://www.fda.gov/downloads/Drugs/GuidanceComplianceRegulatoryInformation/Guidances/UCM 333001.pdf Date accessed: 12.02.2015 28. Guideline for Bioavailability and Bioequivalence, Central Drugs Standard Organization, Director of General of Health Services, Ministery of Health and Family Welfare, Government of India, New Delhi, Mar 2005 29. Guidance for industry Bioavailability and Bioequivalence studies for orally Administered Drug Products-General considerations U.S. Department of health and Human Services, Food and Drug Administration, Mar 2003 30. U.S. Department of Health and Human Services, Food and Drug Administration Center for Drug Evaluation and Research (CDER), Guidance for Industry, Bio analytical Method Validation, Center for Veterinary Medicine (CVM), May 2001 31. Ministry of Health and Family Welfare (Department of Health), The Drug and Cosmetics Act and Rules, 1945(As amended up to the 30th June, 2005) 32. Ethical Guidelines for Biomedical Research on Human Participants Indian Council of Medical Research, New Delhi 2006, Published by: Director-General Indian Council of Medical Research, New Delhi 110029; Oct, 2006 33. Guidance for Industry. Food-Effect Bioavailability and Fed Bioequivalence Studies, U.S. Department of Health and Human Services, Food and Drug Administration, Center for Drug Evaluation and Research (CDER), Dec 2002, BP



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Bioequivalence study of Abiraterone acetate tablet in Indian healthy adult male volunteers APPENDIX Table 39: Demographic Data Listing Subject No.

Gender

Age (years)

Height (cm)

Weight (kg)

BMI (kg/m2)

01

Male

31

165.60

68.60

25.00

02

Male

24

170.10

63.30

21.90

03

Male

30

167.70

56.60

20.10

04

Male

25

176.80

64.80

20.70

05

Male

27

166.90

53.80

19.30

06

Male

39

170.10

65.50

22.60

07

Male

27

179.10

68.50

21.40

08

Male

41

167.70

61.20

21.80

Table 40: Abnormal laboratory values Period I Check-out (Serum electrolyte) Subject No.

Parameter

Value

Repeat

Conclusion

01

Phosphate

4.8 mg/dL

3.8 mg/dL

Normal

02

Phosphate

5.2 mg/dL

4.1 mg/dL

Normal

04

Phosphate

4.9 mg/dL

4.2 mg/dL

Normal

Period II Check-out mid-safety laboratory assessment 05

WBC

3410 cells/cmm

5210 cells/cmm

Normal

Period III Check-out (Serum electrolyte) 04

Phosphate

4.9 mg/dL

4.2 mg/dL

Normal

Post-study laboratory assessment 03

Bilirubin

2.03 mg/dL

Phosphate

1.9 mg/dL



Page 74

Bioequivalence study of Abiraterone acetate tablet in Indian healthy adult male volunteers Source Documents (Separately Attached) 1. Clinical Study Protocol 2. Inform Consent Documents 3. Case Report Forms 4. Case Study Report Disposition of Subjects

Subjects Enrolled N = 08

Period I Checked-in (N = 08)

Period II Checked-in (N = 08)

Dosed T (N = 05) R (N = 03)

Dosed T (N = 03) R (N = 0R)

Withdrawn (N = 00) Dropout (N = 00)

Completed (N = 08)

Period III Checked-in (N = 07)

Dosed T (N = 04) R (N = 03)



Completed (N = 08)

Completed (N = 07)

Period IV Checked-in (N = 07)

Dosed T (N = 03) R (N = 04)


Completed (N = 07)

Completed (N = 07)

Figure 30: Disposition of subjects



Page 75

Bioequivalence study of Abiraterone acetate tablet in Indian healthy adult male volunteers Event

Screening

Period I D0

Written informed consent

√

MH, DM, PE

√

Vitals Monitoring

√

Well-being of subject

√

ECG recording & chest X-

√

D1

D2

W/O D3

D4

Period II D0

D1

√

√

D2

W/O D3

D4

Priod III D0

D1

√

√

D2

W/O D3

D4

Period IV D0

D1

√

√

D2

D3

D4

√

√

√ √

√ √

√

√

√ √

√

√

√ √

√

√

√ √

√

ray (Screening) Hematology, Biochemistry,

√ Urine

analysis (Screening) HIV,

HBV,

HCV,

√

RPR/VDRL screen Urine drug screening

√

√

√

√

√

Alcohol breath analysis

√

√

√

√

√

√

√

√

√

Check-in Dosing

√

√

Check-out

√ √

PK sampling (post-dose)

√

√

√

√

√

√

√

√

AE monitoring recording

√ safety

√

√

√

√

√

PK sampling (pre-dose)

ECG

√

√

√

√

√

√

√

√

√

√

√

√

√

√

√

√

√

√

√

√

√

√

√

√

√

√

√

√

√

√

√

√

√

√

√

analysis Serum electrolyte test

√

√

Hematology, Biochemistry

√

√

(Safety analysis)



Page 76