A Phase II/III, Randomized, Single Blinded, Active Controlled Study to Evaluate the Immunogenicity and Safety of Inactivated Japanese encephalitis Vaccine in healthy volunteers

|Product:  Inactivated Japanese encephalitis Vaccinexml:namespace prefix = o ns = "urn:schemas-microsoft-com:office:office" /

| --- | --- | |Protocol Number

BBIL/JEV/II/III/2011

|Full Title of Project

A Phase II/III, Randomized, Single Blinded, Active Controlled Study to Evaluate the Immunogenicity and Safety of inactivated Japanese encephalitis Vaccine in healthy volunteers.

|Short Title

Randomized, Single Blinded, Active Controlled Immunogenicity and Safety Study of BBIL’s Japanese encephalitis Vaccine.

|Version Number and Date

Version 1.0 and 04/02/2011

|Investigational Product

Inactivated Japanese encephalitis Vaccine

|Clinical Phase

Phase II/III

|Sponsor

Bharat Biotech International Ltd

Genome Valley, Shameerpet,

Hyderabad-500 078,

Andhra Pradesh, India

|Project Technical Advisers

 Dr. Richard Chin, Director,

Dr. Raj Shankar Ghosh, Regional Director, South Asia &

Dr. Biswal

Institute for One World Health

New Delhi -110025

V: +91.11. 40601208/09, http://oneworldhealth.org

Dr. Milind Gore,

National Institute of Virology,

Pune, India.

|  |Study Type

Preventive, Randomized, Single Blinded, Active Controlled Study

| Study Population

 A total of 600 healthy volunteers of age group of 50 to 1 will participate in this study with 450 Test vaccine group and 150 Reference vaccine group. The study will proceed in order of descending age group as mentioned below.

Category

Age in Years

Test

Reference

|1

≤50  to  >18

150

50

|2

≤18  to  > 6

150

50

|3

≤6  to  ≥1

150

50

|Total

450

150

The sample size has been calculated on the following assumptions: allocation ratio of 3:1 (test: reference), 90 % power, a non-inferiority margin of 15%, one sided alpha - 97.5% CI. Based on the above inputs, a total of 600 evaluable subjects are needed (450 in test group and 150 in the Reference group).

|Study Objectives

 The primary objective is to Compare the Immunogenicity of the Test vaccine with reference vaccine in terms of Seroconversion and Geometric Mean Titers of JEV neutralizing antibody four weeks after  two doses.

  |The Secondary objective is to assess and to evaluate the Safety of the cell cultured inactivated Japanese encephalitis Vaccine in healthy volunteers of 50 to 1 years.

|Study Design

Prevention, Randomized, Single Blinded, Active Control Study.

Screening Period

Visit 1:

After obtaining written Informed Consent from the subject/legally acceptable representative of the subject (in case of minor age groups), subjects will be screened for eligibility into the study. Screening period might last for 7days. Screening includes obtaining medical history, physical examination, demographic data (Weight, Height & Gender). Vital signs (Heart rate, Respiratory Rate, Body temperature-oral or auxiliary). After completion of the screening process, if determined to be eligible, subject shall be enrolled into the study.

Treatment & Follow-up period

Visit 2 (Day 0):

Healthy volunteers of 50 to 1 years will be enrolled. 4 ml of blood sample will be collected before the vaccination for antibody estimation. They will receive either Test vaccine or Reference vaccine according to the randomization.

 Prior to vaccination, eligibility and medical history will be reviewed; vital signs and body temperature will be measured. Subjects shall be observed for 30 minutes after vaccine administration for Vital signs, local reactions, and general symptoms for the next 7 days. The information will be recorded in the diary cards.

Visit 3 (Day 28±2):

On the day 28 of the study, a physical examination and medical history performed. 4 ml of blood sample will be collected for JE Anti body estimation, vital signs and body temperature will be recorded, dairy card checked and documented. Subjects will receive Second dose of vaccine. Vital signs and body temperature will be recorded 30-60 minutes post vaccination. Subjects shall be observed for 30 minutes after vaccine administration for local reactions, general symptoms and for the next 7 days. The information will be recorded in the diary cards.

 Visit 4 (Day 56±2):

On day 56±2, the final visit of the study, a physical examination and medical history performed. 4 ml of blood sample will be collected for JE Anti body estimation, vital signs and body temperature will be recorded dairy cards will also be checked and documented.

Unscheduled visits:

Subjects will be advised to return to Hospital as soon as possible during the 56±2 days study period if the subject develops fever or if they are in any way concerned about the health of the subject for treatment. All unscheduled visits will be documented and details of treatments received

and recorded Adverse Events will be documented in the Case Record Form.

               Following immunization, safety will be measured by assessment of Adverse Events (AE) through 28±2 days following dose of vaccine. Also Serious Adverse Events (SAE) and new-onset chronic medical conditions through 2 months after first dose of vaccination (Day 56±2), and reactogenicity to the vaccine for 7 days following vaccination are evaluated.

 Immunogenicity testing will include antibody estimation on Day 0, Day 28±2  and Day 56±2 samples.

|Number of Arm in the Study

Two arm study: One arm is Inactivated Japanese encephalitis vaccine (Test Vaccine) and Second arm will be Live, attenuated Japanese encephalitis vaccine (Reference vaccine).

|Randomization and Blinding

The randomization sheet will be generated by a third party (Asian Clinical Trials, Hyderabad) involvement, it will be sent to the study centers directly and not to the Sponsors. All the test and reference vaccine vials will be coded as per the random numbers. The group assignment will not be known to the Subject/Subject’s legal representative, Sponsor, laboratory staff performing the immunologic and serologic assay. Sufficient care will be taken to blind the sera samples while coding to keep laboratory staff performing serologic assay also blinded. A extra copy of Case Report Forms will be used to blind the codes of serum samples.

A copy of computer generated randomization list of subject numbers and decoded key will be sent to the biostatistician at the end of the study for decoding of the data and statistical analysis.

|Inclusion Criteria

1. Healthy volunteers of 50 to 1 years age group (inclusive) without significant medical history or abnormal findings at screening
2. Willing to give signed written Informed Consent.

3.      Available for all study related visits and procedures for the entire duration of the study, without any known exposure to JE prior to the first screening visit based on previous clinical history.

|Exclusion Criteria

1.      Subjects with the age less than 1 and above 50 years.

2.      Fever of any origin of duration more than 3 days within one month prior to screening or on the day of screening.

3.      History of malaise, head ache, anorexia at the time of screening or during the administration of the vaccine under study.

4.      Past history of JE infection.

5.      Life threatening or serious cardiac (NYHA grades III-IV heart failure), respiratory gastrointestinal, Hepatic, renal, Endocrine, hematological or immune disorders.

6.      Past history of / current   allergic diseases.

7.      Any confirmed or suspected immunosuppressive or immunodeficient condition

8.      Use of any marketed or investigational or herbal medicine or non            registered drug or vaccine for JE or other vaccine in the past 2            months.

9.      Clinically relevant abnormal hematology or biochemistry values in the opinion of the investigator.

10.  Any criteria, which in the opinion of the investigator, suggests that the subject would not be compliant with the study protocol.

11.  Intention to travel out of the area prior to final or follow-up

Visit on day 56±2.

12.  Previous history of hypersensitive reaction to vaccine or

Vaccine component.

|Test Vaccine

  The JE vaccine (BBIL’S) is purified inactivated vaccine, containing the JEV strain isolated from a clinical case of JE infection from an endemic area in India. The strain was adapted to grow in Vero cells. The vaccine was prepared by purification and inactivation process consistent with current Good Manufacturing Practices.

|Reference Vaccine

The SA 14-14-2 is a Live, attenuated Japanese encephalitis vaccine. It was developed and is produced by Chengdu Institute of Biological products (CDIBP), China.

|Dose, Dosage and route of administration

Test vaccine: Liquid 0.5 mL of Test Vaccine will be injected as two doses on day 0 and day 28±2 by   Intramuscular route.

 Reference vaccine: Lyophilized Reference Vaccine 0.5mL will be injected after reconstitution with the diluent supplied as one dose on  day 0 and second dose will be Placebo on day 28±2 by subcutaneous route.

 Subjects will receive either cell culture Inactivated Japanese encephalitis vaccine containing NLT 5µg protein or Reference vaccine as per randomization.

|Endpoints

Primary

·         Proportion of participants achieving 4-fold or greater neutralizing antibody titer in subjects seropositive at baseline (≥1:10) at day 28±2 after a single dose of vaccination.

·         Proportion of participants that are seronegative at baseline (<1:10) will require a PRNT50 titer of ≥1:10 to meet the criteria for seroconversion at day 28±2 after a single dose of vaccination..

• GMT in each group on day 0 and day 28±2.

 Secondary

·          Occurrence of solicited and unsolicited local and systemic AEs within 28 days and 56 days of post vaccination on day 0, 28±2 and 56±2.

·          Occurrence of vaccine-associated SAEs throughout the course of the study.

·          Proportion of participants achieving 4-fold or greater neutralizing antibody titer at day 56±2 after two doses of vaccination.

|Statistical Analysis Plan

The statistical analysis will be performed by Dr. G.S.R. Murthy, Indian Statistical Institute (ISI), Hyderabad. The data from trial center would be collected by representatives of BBIL after reviewing it at the site.

Data analysis would be carried out separately in each of the group based on the size and age as mentioned above.

The test used for evaluation would be as follows:

1. Comparisons of the proportion of subjects with seroconversion in the groups shall be made with the use of the chi-square test with Yates’ correction
2. The differences in the distribution of antibody titers shall be tested with the use of the Kolmogorov-Smirnow nonparametric method
3. The Adverse Reactions between groups will be analyzed by Chi-square or Fisher exact test. The 95% confidence intervals shall be calculated for median values.

 Immunogenicity was analyzed with the endpoints (with 95% Confidence intervals) used by regulatory authorities for evaluation of Japanese encephalitis vaccine. These end points are

·         Seroconversion (Four-fold rise in post-vaccination titers to pre-vaccination titers).

·         Seroprotection (≥1:10, which is deemed to be the seroprotective threshold)

·         Geometric Mean Titer rise (geometric mean of the within subject ratios of post-vaccination reciprocal titter (day 28±2 & 56±2) to the Day 0 reciprocal titter)

|Data Documenting

Case Report Forms and Subject Diary Cards