Version 1; 10 March 2009 Page 1 of...

Protocol No.: 323_PREGA_09 Version 1; 10 March 2009

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PROTOCOL No.: 323_PREGA_09

Version 1; 10 March 2009 Supersedes: Not Applicable

DCGI approved protocol No. R1_PREGAa_07_B001

An open label, balanced, randomized, three-treatment, three-period, three-sequence, crossover bioavailability study comparing single dose of two batches of

Pregabalin 600 mg extended release tablet of Ranbaxy Laboratories Limited, with two oral doses of Lyrica 300 mg capsules (each dose containing

Pregabalin 300 mg administered 12 hourly; total dose 600 mg, of Pfizer GmbH) in healthy adult, human,

male subjects under fed condition

Ranbaxy Laboratories Limited Udyog Vihar Industrial Area

Gurgaon - 122 015 Haryana, India


of 28 Table of Contents

Page

1.0 LIST OF ABBREVIATIONS.............................................……… 42.0 INVESTIGATORS' DECLARATION.....................................…….. 63.0 FACILITIES..........................................................................………… 84.0 SYNOPSIS.......................................………………………………… 95.0 INTRODUCTION.................................................................................…… 116.0 BIOEQUIVALENCE CRITERIA............................................… 117.0 OBJECTIVE……………………………………………………………… 118.0 PRODUCTS TO BE EVALUATED…………………………………….. 11 8.1 Reference (R) ...................................................................………….. 11 8.2 Test (A)……………………………………………………………. 11 8.3 Test (B)……………………………………………………………. 129.0 PHARMACOLOGY.......................................................................……… 12 9.1 Absorption, Distribution, Metabolism and Excretion...............…… 12 9.2 Adverse Effects................................................................ 12 9.3 Dosage............................................................................……………. 1310.0 STUDY DESIGN.........................................................................………….. 14 10.1 Summary...........................................................................………….. 14 10.2 Number of Subjects...........................................................………….. 14 10.3 Admission and Stay 14 10.4 Dose.................................................................................………… 14

10.4.1 Reference (R)……………………………………………………… 14 10.4.2 Test (A)…………………………………………………………… 14 10.4.3 Test (B)…………………………………………………………… 15 10.5 Fasting/Meals..................................................................…………… 15 10.6 Sampling Schedule...............................................................……….. 15 10.7 Washout Period ..................................................................………… 1611.0 RESTRICTIONS............................................................................………… 16 11.1 Medications........................................................................…………. 16 11.2 Diet....................................................................................………….. 16 11.3 Activity...............................................................................………… 1712.0 STUDY POPULATION...........................................................………. 17 12.1 Screening Assessments.............................................................…… 17 12.1.1 Laboratory Tests……………………………………………………. 17 12.2 Inclusion Criteria.................................................................………. 18 12.3 Exclusion Criteria................................................................…… 1813.0 STUDY MEDICATION ..................................................................….…… 19 13.1 Handling, Storage and Accountability Procedures.................... 19

Continued


of 28 Table of Contents (Continued)

Page

13.2 Assignment to Treatment Sequences ..................................………. 19 13.3 Assessment of Compliance..................................................………. 1914.0 PHARMACOKINETIC PROCEDURES.....……………………………. 20 14.1 Blood Sampling...................................................................………… 20 14.2 Analytical Procedures........................................................…………. 20 14.3 Pharmacokinetic Parameters................................................………. 2115.0 SAFETY.....................................................................................………….. 22 15.1 Clinical Safety Measurements..............................................………. 22 15.2 Adverse Events.................................................................…………. 2216.0 STATISTICAL ANALYSIS.............................................................……… 24 16.1 Summary Statistics.............................................................………… 24 16.2 Analysis of Variance (ANOVA).............................................……… 24 16.3 90% Confidence Intervals and Ratio Analyses …………………… 2417.0 DEVIATIONS..............................................................................…………. 2418.0 ETHICAL CONSIDERATIONS........................................................…… 25 18.1 Basic Principles...................................................................……….. 25 18.2 Institutional Review Board...................................................……….. 25 18.3 Informed Consent...............................................................………… 25 18.4 Drop-out/Withdrawal of Subjects from Study........................……… 25 18.5 Subject Compensation.....................................................……….. 26 18.6 Medical Treatment for Injury……………………………………… 2619.0 TERMINATION OF THE STUDY ....................................................…… 2620.0 STUDY DOCUMENTATION…………………………………………… 2621.0 QUALITY ASSURANCE AUDITS...................................................…… 2622.0 CONFIDENTIALITY OF DATA.......................................................…… 2723.0 CASE REPORT FORMS………………………………………… 2724.0 ARCHIVES.................................................................................…………. 2725.0 PUBLICATION POLICY.................................................................……… 2726.0 REFERENCES..............................................................................……….. 2727.0 LIST OF APPENDICES.................................................................……….. 28


of 28 1.0 LIST OF ABBREVIATIONS

AE : Adverse event ALP : Alkaline Phosphatase ALT : Alanine aminotransferase ANOVA : Analysis of Variance AST : Aspartate aminotransferase AUC : Area under the curve BLQ : Below Limit of Quantitation BUN : Blood urea nitrogen CFR : Code of Federal Regulations cm : Centimeter Cmax : Maximum measured blood concentration CPP : Clinical Pharmacology and Pharmacokinetics CPU : Clinical Pharmacology Unit CRO : Contract research organization CV : Coefficient of Variation DCGI : Drug Controller General of India ECG : Electrocardiogram gm : Gram Hb : Hemoglobin HCV : Hepatitis C Virus HIV : Human Immunodeficiency Virus HPF : High powered field (microscopic) ICF : Informed Consent Form ICH : International Conference on Harmonisation IEC : Independent Ethics Committee Inc. : Incorporation IRB : Institutional Review Board Kel : Elimination Rate Constant kg : kilogram mg : Milligram mL : Milliliter mm : Millimeter N/AP : Not Applicable Occ. : Occasional OTC : Over the counter RBC : Red Blood Corpuscles


of 28 1.0 LIST OF ABBREVIATIONS (Continued)

SAS : Statistical Analysis Software SD : Standard Deviation SOP : Standard Operating Procedure T1/2 : Terminal elimination half-life Tmax : Time of maximum measured blood concentration USA : United States of America VDRL : Venereal Disease Research Laboratory test WBC : White Blood Corpuscles WMA : World Medical Association


Page 8 of 28 3.0 FACILITIES 3.1 Clinical Services Clinical Pharmacology Unit Ranbaxy Laboratories Limited Majeedia Hospital (2nd Floor) Jamia Hamdard (Hamdard University) Hamdard Nagar New Delhi 110 062 India

Ranbaxy Laboratories Limited Plot No. GP5, Sector-18, HSIDC Old Delhi - Gurgaon Road Gurgaon (Haryana) India.

3.3 Alternate Clinical Laboratory Dr. Lal PathLabs Pvt. Ltd. Eskay House 54, Hanuman Road New Delhi 110 001 India Or

Super Religare Laboratories Regional Reference Laboratory

Plot GP - 26, Maruthi Industrial Estate, Sector – 18, Udyog Vihar Gurgaon - 122015

Clinical Pharmacology & Pharmacokinetics Ranbaxy Laboratories Limited

Plot No. GP-5, Sector 18, HSIDC Old Delhi –Gurgaon Road Gurgaon 122 015, Haryana India

3.2 Clinical Laboratory

3.4 Analytical, Pharmacokinetic & Statistical Services


Page 9 of 28 4.0 SYNOPSIS

Study Number: 323_PREGA_09 Description: An open label, balanced, randomized, three-treatment, three-period, three-sequence, crossover

bioavailability study comparing single dose of two batches of Pregabalin 600 mg extended release tablet of Ranbaxy Laboratories Limited, with two oral doses of Lyrica 300 mg capsules (each dose containing Pregabalin 300 mg administered 12 hourly; total dose 600 mg, of Pfizer GmbH) in healthy adult, human, male subjects under fed condition.

Products to be Evaluated:

Reference (R): Lyrica capsules (containing Pregabalin 300 mg) manufactured by Pfizer GmbH, Germany.

Test (A): Pregabalin 600 mg extended release tablet, manufactured by Ranbaxy Laboratories Limited India. Test (B): Pregabalin 600 mg extended release tablet, manufactured by Ranbaxy Laboratories Limited, India

Dose: Reference: Two oral doses of reference product, each containing Pregabalin 300 mg will be administered, at an interval of 12-hours with 240 mL of drinking water at an ambient temperature 45 minutes after starting of a standard meal in each period under supervision of trained personnel

OR Test: A single oral dose of either test A or test B containing Pregabalin 600 mg will be administered with 240 mL of drinking water at an ambient temperature 45 minutes after starting of a standard meal in each period under supervision of trained personnel.

Number of subjects: 18

Dietary Status: For Reference: After an overnight fast of at least 10 hours, subjects will start the standard meal 45 minutes prior to administration of the morning dose. Study subjects will have to eat this meal in 30 minutes or less. The dose will be administered 45minutes after start of the meal with 240 mL of water. The second dose will be administered at 12.00 hrs after the morning dose. The standard meals will also be served at 45 minutes prior to second dose. Study subjects will have to eat this meal in 30 minutes or less. The dose will be administered 45 minutes after start of the meal with 240 mL of water. No food will be allowed for at least 4 hours post each dose. For Test: After an overnight fast of at least 10 hours, subjects will start the standard meal 45 minutes prior to administration of the study drug. Study subjects will eat this meal in 30 minutes or less. The dose will be administered 45 minutes after start of the meal with 240 mL of water. The standard meals will be served at 11.25 hours post-dose also. No food will be allowed for at least 4 hours post-dose. Apart from the standard meals, subjects will receive Lunch at 4 hours post dose on dosing day, and breakfast, lunch and snacks at 24, 28 and 32 hours post-dose, respectively. Drinking water will not be allowed from 1 hour before dosing until 2 hours post-dose (for Test Product and for first and second dose of Reference Product) except 240 ml of water given during administration of the dose.Thereafter, it will be allowed at all times.

Sampling Schedule:

Test (A & B): Predose (in duplicate) & at 1.000, 2.000, 2.500, 3.000, 3.500, 4.000, 4.500, 5.000, 5.500, 6.000, 6.500, 7.000, 7.500, 8.000, 8.500, 9.000, 10.000, 12.000, 14.000, 18.000, 24.000, 30.000, 36.000 and 48.000 hours post dose in each period. Reference (R): Pre morning dose (in duplicate) & at 0.500, 1.000, 1.333, 1.667, 2.000, 2.333, 2.667, 3.000, 3.333, 3.667, 4.000, 4.500, 5.000, 6.000, 8.000, 10.000, 12.000, 12.500, 13.000, 13.333, 13.667, 14.000, 14.333, 14.667, 15.000, 15.333, 15.667, 16.000, 16.500, 17.000, 18.000, 20.000, 24.000, 30.000, 36.000 and 48.000 hours post morning dose. The sampling at 12.0 h post first dose will be done 2 minutes before scheduled time to enable second dosing on time. Samples will be collected in CPDA vacutainers. A total of 90 blood samples including duplicate (2 X 4 mL) predose blood samples. Plasma samples will be divided into 2 aliquots.


Page 10 of 28 Storage Condition:

Below -15o C.

Housing: At least 10 hours prior to dose until 36 hours post-dose. Visits: After discharge there will be one visit at 48 hours post-dose in each period. Blood Volume:

4-mL/sample, 87 draws, 16 mL for screening, 37.5 mL discarded, 06 mL for safety analysis, total 419.5 mL per subject including duplicate predose samples

Compensation: Rs.8700/- per completed subject Washout Period:

At least 05 days

Clinical Safety Measurements:

Vital signs - oral temperature, sitting BP and radial pulse after admission, within 1.5 hours pre-morning dose, 2, 6, 10, 14, 24, 36 and 48 hours post-dose in each period.

Scan for Drugs of Abuse and alcohol

Scan for drugs of abuse (cannabinoids and opioids) will be carried out prior to admission in each period. Breath test for alcohol will be carried out prior to admission in each period. Fundoscopy of each subject will be done at the time of admission of each period of the study.

Analytical Procedure:

Pregabalin in plasma quantitated using validated chromatographic procedure

Pharmacokinetic Parameters:

AUC0-t, AUC 0-24 , AUC0-∞ , AUC0-t / AUC0-∞, Cmax, Tmax, Kel and T1/2.

Statistical Analysis:

Summary statistics, ANOVA, 90% Confidence Intervals and Ratio Analyses

Bioequivalence criteria

For pregablin: - The 90% confidence interval for the ratio of test and reference product for AUC0-t and AUC0-24 should be between 80% and 125% for the log transformed data.


Page 11 of 28 5.0 INTRODUCTION

Pregabalin binds with high affinity to the alpha2-delta site (an auxiliary subunit of voltage-gated calcium channels) in central nervous system tissues. Although the mechanism of action of pregabalin is unknown, results with genetically modified mice and with compounds structurally related to pregabalin (such as gabapentin) suggest that binding to the alpha2-delta subunit may be involved in pregabalin's antinociceptive and antiseizure effects in animal models. In vitro, pregabalin reduces the calcium-dependent release of several neurotransmitters, possibly by modulation of calcium channel function. Pregabalin is indicated for:

- Management of neuropathic pain associated with diabetic peripheral neuropathy

- Management of postherpetic neuralgia - Adjunctive therapy for adult patients with partial onset seizures - Management of fibromyalgia.

Two new batches of extended release tablet formulations, containing Pregabalin 600 mg, have been developed by Ranbaxy Laboratories Limited, India. For its registration to DCGI, it is mandatory to assess the pharmacokinetics and bioavailability of the new formulations in comparison with the reference formulation, Lyrica capsules (containing Pregabalin 300 mg) of Pfizer GmbH, Germany.

6.0 BIOEQUIVALENCE REQUIREMENTS

For pregabalin: - The 90% confidence interval for the ratio of test and reference product for AUC0-t and AUC0-24 should be between 80% and 125% for the log transformed data.

7.0 OBJECTIVE

To compare the single-dose oral bioavailability of two batches of Pregabalin 600 mg extended release tablet of Ranbaxy Laboratories Limited with two oral doses, administered 12 hourly, of Lyrica capsules (each containing Pregabalin 300 mg) of Pfizer GmbH, Germany in healthy, adult, male human, subjects under fed condition.

8.0 PRODUCTS TO BE EVALUATED

8.1 Reference (R)

Lyrica capsules (containing Pregabalin 300 mg) of Pfizer GmbH, Germany.

8.2 Test (A) Pregabalin 600 mg extended release tablet of Ranbaxy Laboratories Limited, India.


Page 12 of 28 8.3 Test (B)

Pregabalin 600 mg extended release tablet of Ranbaxy Laboratories Limited, India.

9.0 PHARMACOLOGY 9.1 Absorption, Distribution, Metabolism and Elimination1

Absorption Pregabalin is rapidly absorbed when administered in the fasted state, with peak plasma concentrations occurring within 1 hour following both single and multiple dose administration. Pregabalin oral bioavailability is estimated to be 90% and is independent of dose. Following repeated administration, steady state is achieved within 24 to 48 hours.

Food Effect

The rate of pregabalin absorption is decreased when given with food resulting in a decrease in Cmax by approximately 25-30% and a delay in tmax to approximately 2.5 hours. However, administration of pregabalin with food has no clinically significant effect on the extent of pregabalin absorption. Distribution

In preclinical studies, pregabalin has been shown to cross the blood brain barrier in mice, rats, and monkeys. Pregabalin has been shown to cross the placenta in rats and is present in the milk of lactating rats. In humans, the apparent volume of distribution of pregabalin following oral administration is approximately 0.56 l/kg. Pregabalin is not bound to plasma proteins.

Metabolism Pregabalin undergoes negligible metabolism in humans. Following a dose of radiolabelled pregabalin, approximately 98% of the radioactivity recovered in the urine was unchanged pregabalin. The N-methylated derivative of pregabalin, the major metabolite of pregabalin found in urine, accounted for 0.9% of the dose. In preclinical studies, there was no indication of racemisation of pregabalin S-enantiomer to the R-enantiomer. Excretion Pregabalin is eliminated from the systemic circulation primarily by renal excretion as unchanged drug. Pregabalin mean elimination half-life is 6.3 hours. Pregabalin plasma clearance and renal clearance are directly proportional to creatinine clearance.

9.2 Adverse Effects The most commonly observed adverse events (≥ 5%) and twice the rate of that seen in placebo) in pregabalin-treated patients in pre-marketing studies were


Page 13 of 28 dizziness, somnolence, peripheral edema and dry mouth. Adverse events were usually mild to moderate in intensity.1

In a review of seven controlled clinical trials in patients of painful diabetic peripheral neuropathy receiving Pregabalin ranging from 150 mg to 600 mg, most common treatment-emergent adverse events were dizziness, somnolence, and peripheral edema.2

In a Phase I study conducted in 53 healthy subjects, Pregabalin was given in rising, multiple doses (25 to 300 mg). Pregabalin was generally well tolerated. There were no serious AEs. The most frequent AEs were dizziness, headache, stupor, somnolence, rhinitis and amblyopia. Mild transient elevations in hepatic enzymes were observed.3

For details regarding adverse effects of Pregabalin, please refer to Appendix 8.

9.3 Dosage

Neuropathic pain associated with diabetic peripheral neuropathy Dosing should begin at 50 mg three times a day (150 mg/day) and may be increased to 300 mg/day within 1 week based on efficacy and tolerability. Post herpetic neuralgia Dosing should begin at 75 mg two times a day, or 50 mg three times a day (150 mg/day) and may be increased to 300 mg/day within 1 week based on efficacy and tolerability. Patients who do not experience sufficient pain relief following 2 to 4 weeks of treatment with 300 mg/day, and who are able to tolerate LYRICA, may be treated with up to 300 mg two times a day, or 200 mg three times a day (600 mg/day). Adjunctive therapy for adult patients with partial onset seizures LYRICA at doses of 150 to 600 mg/day has been shown to be effective as adjunctive therapy in the treatment of partial onset seizures in adults. The total daily dose should be divided and given either two or three times daily. Based on individual patient response and tolerability, the dose may be increased to a maximum dose of 600 mg/day. Management of Fibromyalgia The recommended dose of LYRICA for fibromyalgia is 300 to 450 mg/day. Dosing should begin at 75 mg two times a day (150 mg/day) and may be increased to 150 mg two times a day (300 mg/day) within 1 week based on efficacy and tolerability.


Page 14 of 28 10.0 STUDY DESIGN 10.1 Summary

An open label, balanced, randomized, three-treatment, three-period, three-sequence, crossover bioavailability study comparing single dose of two batches of Pregabalin 600 mg extended release tablet of Ranbaxy Laboratories Limited, with two oral doses of Lyrica 300 mg capsules (each dose containing Pregabalin 300 mg administered 12 hourly; total dose 600 mg, of Pfizer GmbH) in healthy adult, human, male subjects under fed condition. The study design has been schematically represented in Appendix 1.

10.2 Number of Subjects

Eighteen (18) healthy male adult human subjects will be admitted in the first period. Subsequent dropouts and/or withdrawn subjects will not be replaced. Data will be presented on all completed subjects. If necessary, an unequal number of subjects per sequence will be used.

10.3 Admission and Stay Subjects will be admitted and housed in the Clinical Pharmacology Unit at least

10 hours before dose administration and will be discharged 36 hours after administration of study drug during each period, if the subjects do not suffer from any adverse drug reaction. In case of an adverse event, the subject will be monitored until the event subsides. The subjects will make one visit to the Clinical Pharmacology Unit for vitals and collection of further blood sample at 48 hours post-dose in each period.

10.4 Dose

Either two oral doses of reference product, each containing Pregabalin 300 mg administered at 12-hour intervals or test (A) or test (B) products containing Pregabalin 600mg will be administered with 240 mL of drinking water under fed condition in each period under supervision of trained personnel.

10.4.1 Reference (R)

Two oral doses of Lyrica capsules (each containing Pregabalin 300 mg) manufactured by Pfizer GmbH, Germany, will be administered at an interval of 12-hours with 240 mL of drinking water at an ambient temperature, 45 minutes after starting of a standard meal. The first dose will be administered after an overnight fast of at least 10 hours. The second dose will be administered 12.00 hours after the first dose, 45 minutes after starting of a standard meal.

10.4.2 Test (A)

A single oral dose of Pregabalin 600 mg extended release tablet manufactured by Ranbaxy Laboratories Limited, will be administered with 240 mL of drinking water at an ambient temperature 45 minutes after starting of a standard meal.


Page 15 of 28

10.4.3 Test (B)

A single oral dose of a Pregabalin 600 mg extended release tablet manufactured by Ranbaxy Laboratories Limited, will be administered with 240 mL of drinking water at an ambient temperature 45 minutes after starting of a standard meal.

10.5 Fasting/Meals For Reference: After an overnight fast of at least 10 hours, subjects will start the standard meal 45 minutes prior to administration of the morning dose. Study subjects will have to eat this meal in 30 minutes or less. The dose will be administered 45 minutes after start of the meal with 240 mL of water. The second dose will be administered at 12.00 hrs after the morning dose. The standard meals will be served at 45 minutes prior to second dose. Study subjects will have to eat this meal in 30 minutes or less. The dose will be administered 45 minutes after start of the meal with 240 mL of water. No food will be allowed for at least 4 hours each post-dose.

For Test: After an overnight fast of at least 10 hours, subjects will start the standard meal 45 minutes prior to administration of the study drug. Study subjects will eat this meal in 30 minutes or less. The dose will be administered 45 minutes after start of the meal with 240 mL of water. The standard meals will also be served at 11.25 hrs. No food will be allowed for at least 4 hours post-dose. Apart from the standard meals, subjects will receive lunch, breakfast, lunch and snacks at 4, 24, 28 and 32 hours, respectively, after drug administration. During housing, all meal plans will be identical for all the periods. Information on the amount of meal consumed and the time taken for consuming the meal will be recorded in the appropriate clinical raw data sheets. In case meals and blood sample collection time coincide, samples will be collected before meals are provided. Drinking water will not be allowed from 1 hour before dosing until 2 hours post-dose (for Test Product and for first and second dose of Reference Product) except 240 ml of water given during administration of the dose. Thereafter, it will be allowed at all times. (Refer Appendix I for details).

10.6 Sampling Schedule

A total of ninety (90) 4-mL blood samples {including duplicate predose samples (2 X 4 mL)} will be collected from each subject in CPDA vacutainers during the course of the study through indwelling cannulae placed in forearm veins. The minimum blood sample volume required for analytical purpose is 4 mL in this study. Reference (R): Prior to morning dose (in duplicate) and at 0.500, 1.000, 1.333, 1.667, 2.000, 2.333, 2.667, 3.000, 3.333, 3.667, 4.000, 4.500, 5.000, 6.000, 8.000, 10.000, 12.000, 12.500, 13.000, 13.333, 13.667, 14.000, 14.333, 14.667, 15.000, 15.333, 15.667, 16.000, 16.500, 17.000, 18.000, 20.000, 24.000, 30.000, 36.000 and 48.000 hours after the morning dosing in each period. The sampling at 12.0 h


Page 16 of 28 post first dose will be done 2 minutes before scheduled time to enable second dosing on time. Test (A & B): Predose (in duplicate) and at 1.000, 2.000, 2.500, 3.000, 3.500, 4.000, 4.500, 5.000, 5.500, 6.000, 6.500, 7.000, 7.500, 8.000, 8.500, 9.000, 10.000, 12.000, 14.000, 18.000, 24.000, 30.000, 36.000 and 48.000 hours post dose in each period. The pre-dose (for test product) / Prior to morning dose (for reference product) blood sample in each period will be collected within a period of 1.5 hours before dosing and the post-dose samples will be generally within 2 minutes of the scheduled time. These predose samples will be collected in duplicate. The actual end time of collection of each blood sample will be recorded. For each subject, the total number of blood draws during the study will be 87 and the total volume of blood drawn, including 16 mL for screening, 37.5 mL 'discarded' blood prior to venous cannula collections, 06 mL for safety analysis at the end of study; will not exceed 419.5 mL, including duplicate predose samples.

10.7 Washout Period There will be a washout period of at least 05 days between the administrations of

study drugs in each period. 11.0 RESTRICTIONS

11.1 Medications Subjects should not have received any medication (except vitamins preparations) including over the counter medications (OTC) during the 2 weeks period prior to the onset of the study. They will be instructed during screening not to take any prescription and OTC medications until the completion of the study. If drug therapy other than that specified in the protocol is required during the study or in the washout period, decisions to continue or discontinue the subject will be based on the following:

i) The pharmacology and pharmacokinetics of the non-study medication.

ii) The likelihood of a drug-drug interaction, thereby affecting pharmacokinetic comparison of the study medication.

iii) The time of administration of the non-study medication.

11.2 Diet

All subjects will be instructed to abstain from alcoholic products and grape fruit juice for 48 hours prior to dosing and during in-house stay in each period. They will also abstain from any other xanthine containing food or beverages during in-house stay in each period.


Page 17 of 28 11.3 Activity All subjects will be dosed while seated and will be instructed to remain seated or

ambulatory for the first 2 hours following each drug administration. Thereafter, subjects will be allowed to engage only in normal activities while avoiding severe physical exertion. However, should any adverse medical event occur at any time during housing the subjects will be placed in an appropriate position or will be permitted to lie down on their right side.

12.0 STUDY POPULATION

Adequate number of subjects will be selected randomly from the Volunteer Bank of Clinical Pharmacology Unit and will undergo a standardized screening procedure as described in SOP No. CP-S02. Eighteen (18) healthy human male subjects will be selected on the following inclusion and exclusion criteria and will be documented in the CRITERIA CHECK form (Appendix 2).

12.1 Screening Assessments Medical histories and demographic data, including name, sex, age, body weight

(kg), height (cm) and number of cigarettes smoked per day will be recorded. Each subject will undergo physical examination and the laboratory tests of hematology, hepatic and renal functions as listed below. Only medically healthy male subjects with clinically normal laboratory profiles will be enrolled in the study.

12.1.1. Laboratory Tests

HEMATOLOGY

• Haemoglobin • Total leukocyte

count • Differential

leukocyte count • Platelet count

URINALYSIS

PHYSICAL EXAMINATION • Colour • Appearance • pH • Specific gravity • Protein • Glucose

ADDITIONAL TESTS

• HIV I & II • HBsAg • HCV • VDRL • Urine drug screen

-cannabinoids -opioids

BIO-CHEMISTRY • BUN • Creatinine • Total bilirubin • Alkaline

phosphatase • AST • ALT • Glucose • Cholesterol

MICROSCOPIC EXAMINATION

• RBC • WBC • Epithelial Cells • Crystals • Casts • Others


Page 18 of 28 All the samples during screening will be collected and analyzed at in-house Clinical Laboratory. Dr. Lal Path labs or Super Religare Laboratories will be used as a back-up laboratory for sample analysis whenever in-house clinical laboratory is out of stock of lab kits or whenever there is a malfunction in laboratory instruments. Scan for drugs of abuse (cannabinoids and opioids) by card/cassette method and breath test for alcohol will be carried out prior to admission in each period.

12.2 Inclusion Criteria

- Be in the age range of 18-45 years. - Be neither overweight nor underweight for his height as per the Life Insurance Corporation of India height/weight chart for non-medical cases. - Have voluntarily given written informed consent to participate in this

study. - Be of normal health as determined by medical history and physical

examination of the subjects performed within 21 days prior to the commencement of the study.

- Be a non vegetarian

12.3 Exclusion Criteria

- History of hypersensitivity to Pregabalin or any other drug. - History of dizziness, ataxia, or in-coordination. - History of recurrent headache. - History of excessive somnolence / narcolepsy. - History of drug induced rashes/pruritis. - History of confusion, or abnormal thinking. - History of peripheral edema, or dry mouth. - History of blurred vision. - Abnormal fundoscopic findings at the time of admission of any period

of the study. - History of Myopathy, Myalgia. - PR interval > 200 msec at the time of screening. - Any evidence of organ dysfunction or any clinically significant deviation

from the normal, in physical or clinical determinations. - Presence of disease markers of HIV 1 or 2, Hepatitis B or C viruses or

syphilis infection. - Presence of values, which are significantly different from normal,

reference ranges (as defined in Appendix 4) and/or judged clinically significant for hemoglobin, total white blood cells count, differential WBC count or platelet count.

- Positive for urinary screen testing of drugs of abuse (opiates or cannabinoids).

- Positive for breath alcohol test. - Presence of values, which are significantly different from normal reference

ranges (as defined in Appendix 4) and/or judged clinically significant for serum creatinine, blood urea nitrogen, serum aspartate aminotransferase


Page 19 of 28 (AST), serum alanine aminotransferase (ALT), serum alkaline phosphatase, serum bilirubin, plasma glucose or serum cholesterol.

- Clinically abnormal chemical and microscopic examination of urine defined as presence of RBC, WBC (>4/HPF), epithelial cells (>4/HPF), glucose (positive) or protein (positive).

- Clinically abnormal ECG or Chest X-ray. - History of serious gastrointestinal, hepatic, renal, cardiovascular,

pulmonary, neurological or hematological disease, diabetes or glaucoma. - History of any psychiatric illness, which may impair the ability to provide,

written informed consent. - Regular smokers who smoke more than 10 cigarettes daily or have

difficulty abstaining from smoking for the duration of each study period. - History of drug dependence or excessive alcohol intake on a habitual

basis of more than 2 units of alcoholic beverages per day (1 unit equivalent to half pint of beer or 1 glass of wine or 1 measure of spirit) or have difficulty in abstaining for the duration of each study period.

- Use of any enzyme modifying drugs within 30 days prior to Day 1 of this study.

- Participation in any clinical trial within 12 weeks preceding Day 1 of this study (except for the subjects who dropout/withdrawn from the previous study prior to period I dosing).

- Subject has a hemoglobin concentration of less than 13 gm%.

13.0 STUDY MEDICATION

13.1 Handling, Storage and Accountability Procedures

Investigational products will be supplied in the original manufacturer's packing and the test products will be supplied in an appropriate package deemed to maintain the integrity of the products. At the clinical facility, the investigational products will be logged-in by the Registered Pharmacist and stored under prescribed storage conditions in a controlled access area in the drug store as described in SOP NO. CP-C24. The Investigator and the Registered Pharmacist will be accountable for the study drug products. Study drugs will be dispensed according to the randomization schedule as described in SOP No. CP-C04.

13.2 Assignment to Treatment Sequences The order of receiving the test A or test B or reference product for each subject

during the 3 periods of the study will be determined according to a SAS-generated randomization schedule. The randomization will be balanced and the code will be kept under controlled access in the drug store. A working copy of the same will be provided to study personnel responsible for dosing. The Investigator, Registered Pharmacist, personnel involved in dispensing of study drugs and the dosing will be accountable for ensuring compliance to randomization schedule.

13.3 Assessment of Compliance

Compliance will be assessed by conducting a thorough examination of the oral cavity by trained study personnel after dosing in each period and by measurement of plasma Pregabalin (during the analytical phase of the study).


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14.0 PHARMACOKINETICS PROCEDURES 14.1 Blood Sampling Blood samples will be collected within 2 minutes of the time specified under

STUDY DESIGN. Intravenous indwelling cannula will be kept in situ as long as possible (up to 24 hrs post dose) when multiple samples will be collected. The cannula will be maintained patent by injection of 1 mL of 5 IU/mL of heparin in normal saline solution. In such cases blood samples will be collected after discarding the first 0.5 mL of heparinised blood and heparin solution from the tubing.

Alternatively, blood samples may be collected by a fresh clean venipuncture

using a disposable sterilized syringe and a needle. All blood samples collected earlier or later than 2 minutes of the scheduled collection time as specified in section 10.0 under STUDY DESIGN will be reported as protocol deviations and the actual delay in collection will be adjusted during pharmacokinetic calculations.

After collection of blood samples from all the subjects at each time-point, one of the study personnel or an attendant will transfer all the collection tubes to a sample processing room at the Clinical Pharmacology Unit. Thereafter the blood samples will be centrifuged under refrigeration as soon as possible to separate plasma. All plasma samples will be divided into 2 aliquots and transferred to suitably labeled tubes and re-checked to ensure transfer of plasma to the correct tube. The plasma will be stored at below -15o C, until transfer to the analytical facility for assay.

14.2 Analytical Procedures

14.2.1 Method Validation Bioanalytical method will be validated for the sensitivity, selectivity, linearity,

ruggedness, accuracy and precision (repeatability and reproducibility), percent recovery and stability of samples (freeze-thaw stability, bench-top stability, auto sampler stability, short-term and long-term stability of stock solution and internal standard).

14.2.2 Assay of Samples Samples from all subjects completing the crossover will be assayed for plasma

Pregabalin using chromatographic procedures developed and validated at Ranbaxy. If necessary, an unequal number of subjects per sequence will be used. No additional subjects will be enrolled. Samples from dropouts and /or withdrawals will not be assayed.

Whenever possible, all samples from each subject will be analyzed on the same

standard curve. Quality control samples will be distributed through each batch of study samples assayed. Samples with drug concentrations greater than the upper limit of the validated range of the assay will be diluted with the appropriate drug-


Page 21 of 28 free biological fluid and reassayed; those which are below the lower limit of this range will be reported as being below LOQ.

14.3 Pharmacokinetic Parameters The following pharmacokinetic parameters will be calculated for Pregabalin

using WinNonlin-Node version 5.0.1 or above from Pharsight:

AUC0-t: The area under the plasma concentration versus time curve, from time zero to the last measurable concentration, as calculated by the linear trapezoidal method.

AUC0-24: The area under the plasma concentration versus time curve, from time zero to 24 h. AUC0-∞ : The area under the plasma concentration versus time curve, from time zero to infinity. AUC0-∞ is calculated as the sum of AUC0-t plus the ratio of the last measurable plasma concentration to the elimination rate constant.

AUC0-t / AUC0-∞ : The ratio of AUC0-t to AUC0-∞ . Cmax: Maximum measured plasma concentration over the time span specified. Tmax: Time of the maximum measured plasma concentration. If the maximum value occurs at more than 1 time point, Tmax is defined as the first time point with this value.

Kel: Apparent first-order terminal elimination rate constant calculated from a semi-log plot of the plasma concentration versus time curve. The parameter will be calculated by linear least-square regression analysis using the maximum number of points in the terminal log-linear phase (e.g. three or more non-zero plasma concentrations). T1/2: The apparent first-order terminal elimination half-life will be calculated as 0.693/Kel.

No value of Kel, AUC0-∞ or T1/2 will be reported for cases that do not exhibit a terminal log-linear phase in the concentration versus time profile.


Page 22 of 28 15.0 SAFETY 15.1 Clinical Safety Measurements

Vitals signs recording Vital signs (oral temperature, sitting blood pressure and radial pulse) will be

measured during subject admission, prior to dosing, 2, 6, 10, 14, 24, 36 and 48 hours after administration of investigational products in each period. Vital signs to be measured prior to administration of the dose will be taken within 1.5 hours of the scheduled dosing time. Post-dose, vital signs will be taken within one hour of the scheduled time. In the event of detection of any abnormality during measurement of vital signs, the Investigator must be consulted for necessary action, which will be recorded. Fundoscopy of each subject will be done at the time of admission of each period of the study.

Clinical examination Brief clinical examination of the subject will be conducted by a qualified medical

designate on duty after subject admission, prior to dosing and thereafter approximately every 12 hours until discharge. In the event of detection of any abnormality during clinical examination, the Investigator must be consulted for necessary action, which will be recorded.

Laboratory Evaluations for Safety Laboratory parameters will be repeated at the end of the study as per the SOP No. CP-C43. Any laboratory parameter outside acceptable limits (Appendix 4) will be termed as laboratory abnormality and followed up, if deemed necessary by Investigator.

15.2 Adverse Events

15.2.1 Definition

Adverse event Any untoward medical occurrence in a subject or clinical investigation subjects administered a pharmaceutical product, which does not necessarily have a causal relationship with this treatment.

An adverse Event (AE) can therefore be any unfavorable and unintended sign (including an abnormal laboratory finding, for example), symptom, or disease temporally associated with the use of a medicinal product, whether or not considered related to the medicinal product.

Medical Event

Any event, which occurs in a subject after signing the Informed Consent Form (ICF) and prior to dosing in period I.

Laboratory adverse event A laboratory abnormality, after being evaluated by a physician for clinical significance, if any one of the following conditions is met:

i.When the abnormal lab report is accompanied with associated symptoms,

ii.When medical/surgical intervention is required.


Page 23 of 28 iii.When it is considered by clinical/principal investigator as an adverse

event

15.2.2 Adverse Event Monitoring and Recording The Investigator or a Medical Officer will be available at the site of investigation until 36 hours post-dose during each period. A medically qualified designate will be on call for the remaining period of the study. Subjects will be monitored throughout the study period for adverse events. Subjects will be informed to bring to the notice of the nurse or the physician, any adverse event that may occur during their stay at the site of investigation. Subjects will also be specifically asked about any adverse events at the time of admission, predose and at 4, 8, 12, 16, 20, 24, 36 and 48 hours post dose in each period. Treatment of any adverse events will be done by a physician, either at the site of investigation or at a nearby hospital.

15.2.3 Adverse Event follow-up All adverse events will be followed up at regular intervals at a frequency as decided by the Clinical Research Physician/ Clinical Investigator/ Principal Investigator depending upon the condition. All unresolved adverse events till end of study, shall be followed up for at least 30 days from the start of the adverse event or until resolution of adverse event or determination that no further medical intervention is deemed necessary or the adverse event is otherwise explained or the subject is lost to follow-up. Serious adverse events that are still ongoing at the end of the study period must be followed up to determine the final outcome.

15.2.4 Adverse Event Reporting Adverse events experienced by subjects will be reported as per SOP No. CP-C07. The outcome of all adverse/ medical events/ lab abnormalities will be stated in the final clinical update/summary. The study may be suspended or terminated depending on the seriousness of the adverse effects.

Serious Adverse Event Occurrence of all serious adverse events as defined in the SOP No. CP-C07 will be communicated to the Chairperson, Jamia Hamdard Institutional Review Board and Principal Investigator, Study Director and Vice President, Medical Affairs and Clinical Research, Ranbaxy Research Laboratories and to Drug Controller General of India, and to any other regulatory agencies for necessary action. Detailed information about the serious adverse event(s) will be sent to In-charge, Corporate Pharmacovigilance-Investigational Products within 24 hours of their occurrence. Information will be sent to the following address:

E-mail. [email protected]

Phone: 91-124-2346503

Fax: 91-124-2342018

Corporate Pharmacovigilance Investigational Products will process the information and report to the Drugs Controller General of India.


Page 24 of 28 16.0 STATISTICAL ANALYSIS

Statistical analyses will be performed on plasma Pregabalin using the SAS system for Windows, release 9.1 or above or WinNonlin PK Software, Version 5.0.1 or above. The analyses will include data from subjects 1 to 18 if all these subjects complete the study. In the event of dropouts and/or withdrawals, they will not be replaced. If necessary, an unequal number of subjects per sequence will be used.

16.1 Summary Statistics

Arithmetic means, standard deviations and coefficients of variation will be calculated for the parameters listed in section 14.3. Additionally, geometric means and percentage coefficient of variation of geometric means will be calculated for AUC0-t, AUC0-∞ AUC0-24and Cmax.

16.2 Analysis of Variance (ANOVA)

The log-transformed pharmacokinetic parameters (Cmax, AUC0-t, AUC0-24,and AUC0-∞) for Pregabalin will be analyzed using a mixed effects ANOVA model using Type III sum of squares, with the main effects of sequence, period and formulations as fixed effects and subjects nested within sequence as random effect. A separate ANOVA model will be used to analyze each of the parameters. The sequence effect will be tested at the 10% level of significance using the subjects nested within sequence mean square as the error term. All other main effects will be tested at the 5% level of significance against the residual error (mean square error) from the ANOVA model as the error term. Each analysis of variance will include calculation of least-squares means, the difference between the adjusted formulation means and the standard error associated with the difference. The above analyses will be done using the appropriate SAS®

procedure or the WinNonlin PK Software, Version 5.0.1 or above. 16.3 90% Confidence Intervals and Ratio Analysis

90% confidence interval for the ratio of the test and reference product averages (least square means) for AUC0-24 and AUC0-t will be calculated for pregabalin by first calculating the 90% confidence interval for the differences in the averages (arithmetic means) of the log-transformed data and then taking the antilogs of the obtained confidence limits. The comparison of interest is A vs R & B Vs R, so the ratios will be of the form:- Test/Reference. Ratio of means will be calculated using the LSM for log-transformed Cmax, AUC0-t, AUC0-24,and AUC0-∞ for Pregabalin. Ratio of means will be expressed as a percentage of the LSM for the reference formulations.

17.0 DEVIATIONS All protocol deviations will be appropriately reviewed and documented in the raw

data and those, which will affect the integrity of the study, will be reported. In addition, deviations from the original pharmacokinetic and statistical evaluation plan will be justified in the study report.


Page 25 of 28 18.0 ETHICAL CONSIDERATIONS 18.1 Basic Principles

This research will be carried out in accordance with the Basic Principles defined in US 21 CFR Part 320, the ICH (62FR 25692, 09 May 1997) 'Guidance for Good Clinical Practice', ICMR 'ethical guidelines for biomedical research on human participants (2006)', CDSCO 'guidance on Good Clinical Practices for Clinical Research in India' and the principles enunciated in the Declaration of Helsinki (WMA General Assembly, Seoul 2008) respectively.

18.2 Institutional Review Board This protocol and the corresponding informed consent form (ICF) used to obtain

informed consent of study subjects will be reviewed by the Jamia Hamdard Institutional Review Board and the study subjects will not be dosed until the Board has approved the protocol and the ICF, as submitted or with modifications in subsequent version(s). The Board is constituted and operates in accordance with the Principles and requirements described in the US Code of Federal Regulations (21 CFR Part 56). A copy of the final clinical update/summary will be provided to the Jamia Hamdard Institutional Review Board.

18.3 Informed Consent The Clinical Investigator or his designate will inform the subjects before

initiation of study through an oral presentation regarding the purpose, procedures to be carried out, potential hazards and rights of the subjects. Subjects will be required to understand and sign a consent form summarizing the discussion prior to admission for the study in Period I.

18.4 Drop-out/Withdrawal of Subjects from Study Subjects will be informed that they are free to dropout from the study at any

time without stating any reason. The investigator may withdraw a subject from the study for any of the following reasons:

(i) The subject suffers from significant inter-current illness or undergoes

surgery during the course of the study.

(ii) The subject experiences adverse event, when withdrawal would be in the best interest of the subject.

(iii) The subject fails to comply with the requirements of the protocol. This

would include pre-study directions regarding alcohol and drug use, fasting or if the subject is uncooperative during the study.

(iv) If the subject requires concomitant medications which may interfere with

the pharmacokinetics of study drug. (v) The subject who experience vomiting at any time during sample collection

schedule will be withdrawn from study.


Page 26 of 28

Details of reasons for withdrawal of subjects will be recorded and reported. Every effort will be made to obtain a complete follow-up for any withdrawn subject.

18.5 Subject Compensation The subjects will be adequately compensated on account of their participation in

the study. In case of drop-out/withdrawal of a subject before completion of the study, the guidelines issued by the Jamia Hamdard Institutional Review Board will be final and binding on both Ranbaxy Research Laboratories and the study subjects. The compensation in this study for subjects will be Rs.8700/- per completed subject.

18.6 Medical Treatment for Injury

In case of research related injury, first aid will be available at the Ranbaxy Clinical Pharmacology Unit and treatment of adverse reactions requiring hospitalization will be undertaken at a nearby hospital and the expenses will be borne by Ranbaxy Research Laboratories.

19.0 TERMINATION OF THE STUDY Ranbaxy Research Laboratories reserves the right to discontinue the trial at any

time. Reasons for this termination will be provided to the subjects. The Principal Investigator reserves the right to discontinue the study for safety reasons at any time.

20.0 STUDY DOCUMENTATION All data generated during the conduct of the study will be directly entered in the

raw data recording forms as governed by the SOPs of Department of Clinical Pharmacology & Development, and Clinical Pharmacology & Pharmacokinetics, Ranbaxy Research Laboratories, except the analytical data of clinical laboratory of the Clinical Pharmacology Unit, which will be transcribed into the study related forms and the raw data retained by the laboratory for records. The computer-generated chromatograms will also be treated as raw data. All raw data and transcribed data forms will be completed by the study personnel assisting in the study and will be checked wherever applicable for completeness and logistics by the Clinical Investigator or his designate Research Scientist for clinical data and the Laboratory Supervisor for the bioanalytical data. The Clinical Investigator and the Laboratory Supervisor will supervise compilation of data until ready for archiving.

21.0 QUALITY ASSURANCE AUDITS The raw data generated during the course of the study, including the clinical and

analytical operations and the study reports will be inspected and quality audit for conformance to this protocol and all the governing SOPs by an auditor from the Corporate Quality Assurance Department of Ranbaxy Research Laboratories.


Page 27 of 28 22.0 CONFIDENTIALITY OF DATA The data identifying each study subject by name will be kept confidential and will

be accessible to the study personnel, Quality Assurance Auditor during audits and if necessary, to the Jamia Hamdard Institutional Review Board and various regulatory agencies.

23.0 CASE REPORT FORMS Data generated during the conduct of study will directly be entered in the CRF. 24.0 ARCHIVES Drug products will be handled/ archived as per the in-house SOP. All data generated in connection with this study, together with the original copy

of this protocol and the final report will be archived. 25.0 PUBLICATION POLICY The results of the study may be published by the Principal Investigator. 26.0 REFERENCES

1. SPC of Lyrica capsculrs. Available at:

http://emc.medicines.org.uk/emc/assets/c/html/displaydoc.asp?documentid=14651

2. US Prescribing Information on Lyrica capsules http://media.pfizer.com/files/products/uspi_lyrica.pdf

3. Roy Freeman, Edith Durso-DeCruz et al. Efficacy, Safety, and Tolerability of

Pregabalin Treatment for Painful Diabetic Peripheral Neuropathy. http://care.diabetesjournals.org/cgi/content/abstract/dc07-2105v1

4. J A Busch , J A Strand , E L Posvar et al. Pregabalin (CI-1008) multiple-dose pharmacokinetics and safety/tolerance in healthy volunteers. Available at: http://www.aapsj.org/abstracts/AM_1999/1284.htm


Page 28 of 28

27.0 LIST OF APPENDICES

Appendix Description 1 Schematic Representation of Study Design 2 Criteria check 3 Assessment during the Study 4 Acceptable limits for ‘Out of Range’ laboratory parameter(s) 5 List of Laboratory Reference Range In-House 6 List of Laboratory Reference Range Values – Dr. Lal PathLabs 7 List of Laboratory Reference Range Values –. Super Religare Laboratories 8 Prescribing Information of LYRICA capsules 9 Informed Consent Form 10 Case Report Form 11 Insurance Policy 12 Meal Menu


APPENDIX 1 SCHEMATIC REPRESENTATION OF STUDY DESIGN

Period I, II and III (n=18)Reference:

D V B Dosing V V D Dosing V & B L S V & Di

1500 2100 730 830 900 1100 1300 1500 1900 2030 2100 2300 900 1300 1700 210018 hr 12 hr 1.5 hr 0.5 hr 0 hr 2hr 4hr 6hr 10hr 11.25hr 12hr 14hr 24 hr 28 hr 32 hr 36 hr

Admission & vitals Predose

blood sample Lyrica 300 mg capsule Lyrica 300 mg capsuleBlood samples Prior to morning dose, 0.500, 1.000, 1.333, 1.667, 2.000, 2.333, 2.667, 3.000, 3.333, 3.667, 4.000,

Note: 4.500, 5.000, 6.000, 8.000, 10.000, 12.000, 12.500, 13.000, 13.333, 13.667, 14.000, 14.333, 14.667, 15.000, 15.333, 15.667, 16.000, 16.500, 17.000, 18.000, 20.000, 24.000, 30.000, 36.000 and 48.000 hrs post morning dose in each periodD= Dinner, B= Breakfast, L=Lunch, S=Snacks, V=Vitals, Di=DischargeDosing and subsequent sampling timings will be suitably staggered.

V L V

Test:

D V B Dosing V V D V & B L S V & Di

1500 2100 730 830 900 1100 1300 1500 1900 2030 2300 900 1300 1700 210018 hr 12 hr 1.5 hr 0.5 hr 0 hr 2hr 4hr 6hr 10hr 11.25hr 14hr 24 hr 28 hr 32 hr 36 hr

Admission & vitals Predose

blood sample Pregabalin 600 mg extended release tabletBlood samples: Predose and at 1.000, 2.000, 2.500, 3.000, 3.500, 4.000,4.500, 5.000, 5.500, 6.000,6.500, 7.000, 7.500, 8.000,

Note: 8.500, 9.000, 10.000, 12.000, 14.000, 18.000, 24.000, 30.000, 36.000 and 48.000 hrs post morning dose in each periodD= Dinner, B= Breakfast, L=Lunch, S=Snacks, V=Vitals, Di=Discharge

Post dose Adverse Event monitoring at admission, pre dose, every 4 hours post dose until discharge and at every ambulatory visit during each period. The subjects will make 1 visit to the Clinical Pharmacology Unit for collection of further blood samples and vitals at 48 hours post-dose in each period.

Dosing and subsequent sampling timings will be suitably staggered.

V L V


Inclusion Criteria

Exclusion Criteria Yes No

• History of hypersensitivity to pregabalin or any other drug. • History of dizziness, ataxia or in-coordination. • History of recurrent headache. • History of excessive somnolence / narcolepsy. • History of drug induced rashes/pruritis. • History of confusion, abnormal thinking. • History of peripheral edema, dry mouth. • History of blurred vision. • Abnormal fundoscopic findings at the time of admission of any period of the study. • History of Myopathy, Myalgia • PR interval > 200 msec at the time of screening

• The subject has evidence of organ dysfunction or any clinically significant deviation from the normal, in physical or clinical determinations.

• Investigations with blood samples of the subject shows presence of disease markers of HIV 1 or 2, Hepatitis B or C viruses or syphilis infection.

• Investigations with blood samples of the subject shows the presence of values which are significantly different from normal reference range and/or judged clinically significant for haemoglobin, total white blood cells count, differential WBC count or platelet count.

• Positive for urinary screen testing of drugs of abuse (opiates or cannabinoids).

• Positive for breath alcohol test.

• Investigations with blood samples of the subject shows the presence of values which are significantly different from normal reference range and/or judged clinically significant for serum creatinine, blood urea nitrogen, serum aspartate aminotransferase (AST), serum alanine aminotransferase (ALT), serum alkaline phosphatase, serum bilirubin, plasma glucose or serum cholesterol.

• Investigations with urine samples of the subject show clinically abnormal chemical and microscopic examination of urine defined as presence of RBC, WBC (>4/HPF), epithelial cells (>4/HPF), glucose (positive) or protein (positive).

CLINICAL INVESTIGATOR: DATE:

Reference No. APPENDIX 2

CRITERIA CHECK

Subject No.

• The subject is in the age range of 18-45 years. • The subject is neither overweight nor underweight for his height as per the Life Insurance

Corporation of India height/weight chart for non-medical cases.

• The subject has voluntarily given written informed consent to participate in this study • The subject is of normal health as determined by medical history and physical examination of the

subjects performed within 21days prior to the commencement of the study.

• The subject is a Non vegetarian

Yes No


Reference No. APPENDIX 2

CRITERIA CHECK

Subject No.

Exclusion Criteria (Cont.) Yes No

• Clinically abnormal ECG or Chest X-ray.

• The subject has history of serious gastrointestinal, hepatic, renal, pulmonary, cardiovascular, neurological or haematological disease, diabetes or glaucoma.

• The subject has history of any psychiatric illness, which may impair the ability to provide written informed consent.

• The subject is a regular smoker who smokes more than 10 cigarettes daily or has difficulty abstaining from smoking for the duration of each study period.

• The subject has history of drug dependence or excessive alcohol intake on a habitual basis of more than 2 units of alcoholic beverages per day (1 unit equivalent to half pint of beer or 1 glass of wine or 1 measure of spirit) or has difficulty in abstaining for the duration of each study period.

• Use of any enzyme modifying drugs within 30 days prior to Day 1 of this study. • Participation in any clinical trial within 12 weeks preceding Day 1 of this study (except for

the subjects who dropout/withdrawn from the previous study prior to period I dosing).

• Subject has a hemoglobin concentration of less than 13 gm%.

Based on the above, the subject is SUITABLE / UNSUITABLE for the study.

CLINICAL INVESTIGATOR:

DATE:


APPENDIX 3 ASSESSMENT DURING THE STUDY

Study Day

Days -21 to 1 Screening

Period

Period I

(Treatment R/A or B)

Period II


Period III


Post-study Period

Safety

Demographics x Vital Signs x x x x Allergies and Medication History

x

Medical History x Physical Examination x Clinical Laboratory Tests

x x

Urinary Drug Screen (for opioids and cannabinoids) only

x x x

Breath Test for alcohol x x x ECG x Chest X-ray x Criteria Check x Adverse Events x x x x Written Informed Consent

x

Pharmacokinetics

Pre-dose blood sampling

x x x

Drug Administration x x x Blood sampling for Pregabalin determination

x x x


APPENDIX 4 ACCEPTABLE LIMITS FOR ‘OUT OF RANGE’ LABORATORY

PARAMETER (S)

Laboratory parameter Acceptable limits

Hb (Haemoglobin) > 13 g/dL

Platelets > 20 % of lower limit of reference range

Total Leukocyte count > 20 % of lower limit or < 20 % of upper limit of reference

Neutrophils > 25 % of lower limit or < 25 % of upper limit of reference

Lymphocytes > 25 % of lower limit or < 25 % of upper limit of reference

Monocytes < 50 % of upper limit of reference range

Eosinophils < 50 % of upper limit of reference range

Basophils < 50 % of upper limit of reference range

Total bilirubin < 20 % of upper limit of reference range

ALT < 20 % of upper limit of reference range

AST < 20 % of upper limit of reference range

Alkaline phosphatase < 20 % of upper limit of reference range

BUN < 20 % of upper limit of reference range

Creatinine Upper limit of reference range

Serum cholesterol < 240 mg/dL

Glucose, fasting Reference range

Glucose, random Reference range


APPENDIX 5

LIST OF LABORATORY REFERENCE RANGE VALUES – IN-HOUSE

PARAMETER UNIT REF. RANGE PARAMETER REF.

RANGE HAEMATOLOGY URINE ANALYSIS

Haemoglobin - Male g/dL 14 – 18 Routine

Total WBC Count per mm3

4000- 10000

Colour

Pale straw/straw

Differential WBC Count Appearance Clear Neutrophils % 40-75 pH 5.0 -6.5 Lymphocytes % 20-45 Sp. Gr. 1.010-1.025 Monocytes % 0-8 Glucose Nil Eosinophils % 0-6 Protein Nil Basophils % 0-1 Microscopic

Platelets per mm3

150000- 450000 RBC Nil/HPF

SEROLOGY W.B.C. 0-3/HPF E. Cells 0-3/HPF

HIV 1&2 Non-Reactive Crystals Absent HCV Non-Reactive Casts Absent VDRL Non-reactive Others Absent

BIOCHEMISTRY

Instrument – Dimension R-XI Auto analyser

Instrument – VITROS Auto Analyser

PARAMETER UNIT REF. RANGE UNIT REF.

RANGE Glucose (Fasting) mg/dl 70-110 mg/dl 74-106 Glucose (Random) mg/dl 70-140 mg/dl 70-140 Total Bilirubin mg/dl < 1.00 mg/dl 0.2-1.3 AST IU/L 15-37 U/L 15-46 ALT IU/L 30-65 U/L 13-69 Alk. Phosphatase IU/L 50-136 U/L 38-126 Creatinine - Male mg/dL 0.8 – 1.3 mg/dL 0.8-1.5

Blood Urea Nitrogen mg/dL 7-18 mg/dL 9-20 Cholesterol mg/dL 130-220 mg/dL 130-220


APPENDIX 6

LIST OF LABORATORY REFERENCE RANGE VALUES – DR LAL PATHLABS

PARAMETER UNIT REF. RANGE

PARAMETER REF. RANGE

HAEMATOLOGY URINE ANALYSIS Haemoglobin

- Male

g/dL 13-17 Routine

Total WBC Count thou/ mm3

4-10 Colour Pale yellow

Differential WBC Count Appearance Clear Neutrophils % 40-80 pH 5.0 -6.0 Lymphocytes % 20-40 Sp. Gr. 1.015-1.025 Monocytes % 2-10 Glucose Nil Eosinophils % 1-6 Protein Nil Basophils % < 2.00 Microscopic Platelets thou/

mm3 150.00 - 450.00

RBC Nil W.B.C. < 10 WBC/µL

BIOCHEMISTRY E. Cells Nil Glucose (Fasting) mg/dl 70 - 110 Crystals Nil Glucose (Random) mg/dl 70 - 140 Casts Nil Sr. Total Bilirubin mg/dl < 1.00 Others Nil Sr. AST U/L < 38 SEROLOGY Sr. ALT U/L < 41 HBs Ag Non-Reactive Sr. Alk. Phosphatase U/L 40-129 HIV 1&2 Non-Reactive Sr. Creatinine

- Male

mg/dL

0.5 – 1.20 HCV Non-Reactive

Blood Urea Nitrogen mg/dL 7-20 VDRL Non-reactive Sr. Cholesterol mg/dL < 200


APPENDIX 7

LIST OF LABORATORY REFERENCE RANGE VALUES – Super Religare Laboratories

PARAMETER UNIT REF.

RANGE PARAMETER REF.

RANGE HAEMATOLOGY URINE ANALYSIS Haemoglobin

- Male

g/dL

13 – 17 Routine

Total WBC Count thou/ mm3

4 -10 Colour

Pale yellow

Differential WBC Count Appearance Clear Neutrophils % 40-80 pH 4.7 -7.5 Lymphocytes % 20-40 Sp. Gr. 1.003-1.035 Monocytes % 2-10 Glucose Nil Eosinophils % 1-6 Protein Nil Basophils % < 1- 2 Microscopic

RBC Nil Platelets thou/

mm3 150 - 410

W.B.C. 0 - 5 /HPF

BIOCHEMISTRY E. Cells 0 – 5 /HPF Glucose (Fasting) mg/dl 74 - 100 Crystals Nil Glucose (Random) mg/dl 70 - 140 Casts Nil Sr. Total Bilirubin mg/dl 0.0 - 1.0 Others Nil Sr. AST U/L 15 - 37 SEROLOGY Sr. ALT U/L 30 - 65 HBs Ag Non-Reactive Sr. Alk. Phosphatase - Male

U/L

50 - 136

HIV 1&2

Non-Reactive

Sr. Creatinine - Male

mg/dL

0.6 – 1.3

HCV

Non-Reactive

Blood Urea Nitrogen mg/dL 7 - 18 VDRL Non-reactive Sr. Cholesterol mg/dL < 200


APPENDIX 12 MEAL MENU

Breakfast Menu

S. No.

Food Stuff

Quantity

(gm)

Energy (Kcal)

Carbohydrate

(g)

Proteins

(g)

Fat (g)

01 Chicken 60 146 - 14 10

02 Whole Milk

240 mL 152 12 8 8

03 Hash Brown

Potatoes

120 162 15 3 10

04 Fried Eggs in butter*

2 medium 267 - 15 23

05 a) Bread Slices

2 large 160 35.5 4.5 -

b) Butter 10 90 - - 10

Total: 977 62.5 44.5 61 Kcal. 250 178 549 % 26 18 56

2 eggs to be fried in 3 gm of butter


Dinner Menu

S. No.

Food Stuff

Quantity

(gm)

Energy (Kcal)

Carbohydrate

(g)

Proteins

(g)

Fat (g)

01 Butter Chicken

70 in 70 ml gravy

269 2 17 21

02 Fried Paneer Curry

180 230 4.4 8.7 20.5

03 Tomato onion curd

120 67 4.5 3.3 4

04 Chapati 4*35 273 55.4 9.7 1.4

05 Butter (added in

butter chicken)

9 81 0 0 9

06 Frooti 1 80 20 0 0

07 Salad 50 0 0 0 0 Total: 1000 86.3 38.7 55.9 Kcal. 345.2 154.8 503.1 % 34.5 15.48 50.31

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