1. Improving Rev.1 Vaccine Stability Produced in Iran
بسم الله الرحمن الرحیم
Improving Rev.1 Vaccine Stability Produced in Iran
Dr. Ramin Bagheri Nejad
Head of Brucellosis Department
Razi Vaccine & Serum Research Institute

2. Rev.1 Vaccine Importance
B. melitensis as the main cause of human brucellosis
Primary cause of Small ruminants brucellosis
Sheep & goats: Preferred natural hosts
Rev.1 vaccine the most effective and the only approved vaccine by the OIE

3. Role of Animal Vaccination
Necessary measure for the control of animal brucellosis but not all required
Effective on decreasing human disease incidence proved by several studies around the globe (Greece, Mongolia and…)
Vaccination efficacy depends on:
Vaccination coverage
Prevalence of infection
Animal production practices
Sanitary measures
Control of animal movements

4. Quality of Rev.1 Vaccine Quality: fitness for the intended use
Safety & potency
Stability: no specified duration
Determined based on standard stability studies
Done on 3 successive batches
The product should retain its potency during shelf life
Important factors: safety, identity, purity & potency

5. Standards for Rev.1 Vaccine Production & Quality Control
International standards:
OIE Manual of diagnostic test & vaccines (2012)
European Pharmacopeia (2015)
WHO: Requirements for Rev.1 vaccine production (1970)

6. Standard Quality Control Tests
Identity:
Biochemical and microbiological tests
Purity: free from contaminating agents
Safety in target animal species
Potency:
Smoothness
Enumeration of live bacteria per dose:
EP:0.5- 4×109, OIE: 0.5-2×109
Immunogenicity in mouse model
No requirement for testing on target animal species

7. Standard Quality Control Tests

8. Standard Quality Control Tests

9. Rev.1 vaccine in Iran Rev.1 production since 1341
Collaboration with the WHO
Field trials among first ones in the world
First supplied as liquid
Then lyophilized
Shelf life of 4 months

10. Rev.1 Vaccine Quality Control in Razi Institute
Production of each batch
Sampling by QC department and sending production documents
QC tests
Report of QC tests to Release Office
Final decision of release or fail by Release Office

12. Materials & Methods Focus on the stabilizer composition Stabilizer:
The most important factor to assure vaccine strain viability
Protects the vaccine strain during lyophilization
Protects final products against environmental stresses during storage
Conventional stabilizer composition as recommended by the WHO and OIE:
Pancreatic digest of casein
Saccharose
Sodium L-glutamate
Different studies by adding cryoprotectants and lyophilization exepients

13. Materials & Methods Evaluation of stability by accelerated testing
Production of a lab-scale batch for extended studies1
Real-time real-temperature stability study
Safety evaluation
Determination of immunogenicity in mice model as recommended by the OIE and European Pharmacopeia

14. Number of live bacteria (cfu/dose)
Results
Identification of 3 more substances
Accelerated Stability study:
Day post-incubation
Number of live bacteria (cfu/dose)
1.66 × 109 1
1.96 × 109 3
1.28 × 109 4
1.33 × 109 5
1.16 × 109 7
1.02 × 109 8
1.11 × 109

15. Results
Real-time stability study:

16. Results Safety: According to European pharmacopeia (2012)
Injection to 2 sheep
Observation of animals for 21 days
No local and systemic adverse effects

17. Results Immunogenicity in mice OIE (2012) & EP protocol 3 mice groups
Challenge with B. abortus strain 544 one month after vaccination
Euthanizing all mice 2 weeks post-challenge
Enumeration on challenge strain in spleens

18. Log of challenge strain in spleen
Results
8 months after production
Mouse Group
Log of challenge strain in spleen
(M±SD)
Y=log(X/logX)
Original seed
2.55±0.95 a
2.1±0.7 a
New Rev.1
3.04±0.57 a
2.5±0.4 a
Non-vaccinated Control
5.32± 0.34 b
4.6± 0.3 b

19. Future Plan Production of 3 successive industrial batches
Performing stability studies on all 3 batches to show consistency of production
Identity
Purity
Enumeration of live bacteria
Safety in target animal species
Immunogenicity in mice
Application to Razi Quality Assurance Deputy and IVO to acquire permission for introduction of changes in industrial production

20. Thanks for your Attention