Progress in the Development of a Vi-CRM Conjugate Vaccine 9th International Conference on Typhoid and Invasive NTS Disease Akshay Goel, PhD Biological E. Ltd., Hyderabad, India 03 May 2015 1
Vi Polysaccharide EFT • Vi Polysaccharide 50 derived from LotVi004-200L-OD Citrobacter OD 40 LotVi005-200L-OD freundii sensu lato 30 • BSL-I, rapid OD growth, high 20 yields • Vi PS NMR 10 identical to Salmonella Typhi 0 0 2 4 6 8 10 12 14 16 Time (h) 2
Purified Vi Polysaccharide SEC-HPLC Purified Vi-PS meets WHO TRS M av ~200-300kD requirements • NMR • O-Acetyl Content • Size • % Pr • %NA • Endotoxin • Residual reagents 3
rCRM 197 • Developed at BioE using E. coli host • Process demonstrated to be robust, meets yield criteria • rCRM 197 meets all quality criteria 4
Vi-CRM 197 Conjugation Kinetics S. No. Process step 1 CRM 197 Derivatization 2 CRM 197 -ADH Purification 3 Vi activation 4 Conjugation Depth filtration 5 of Conjugation Mixture Vi-CRM 197 Conjugate 6 Purification Buffer Exchange of Vi- 7 CRM 197 Conjugate 8 0.22 µm Filtration 5
Vi-CRM Conjugates : Critical to Quality Bulk Conjugate Formulated Bulk • Identity • Identity • Vi Concentration (25 m g/0.5 mL) • Vi Concentration • Vi:CRM ratio • Vi:CRM ratio • Size • Size • % free PS • % Free PS • O-Acetylation level • Sterility • Residual reagents • Osmolarity, pH • Endotoxin • Stability • Stability 6
ID of Vi and CRM 197 in Conjugate by Dot Blot Identification of CRM 197 in conjugate Identification of Vi in conjugate 7
Balb/c Mice Immunization Plan for TCV Study Plan: 1. Mice: • Inbred Balb/C Female SPF Mice • < 6weeks old • 20 mice/per group 2. Route: Subcutaneous 3. Dose: 2.5µg vaccine/100µl , 3 dose 4. Sera collected by terminal bleeding Samples Evaluated: 1. BE Vi-rCRM 2. BE Vi-rCRM 3. Vi PS negative control (BE Vi) Responses Evaluated: 4. Vi-conjugate positive control • Anti-Vi IgG (Fold increase over Placebo and over PS only) 5. Vi PS negative control (native) • Booster Effect 6. PBS Placebo 8
The Conjugate should induce a response that is at least four times higher than the response induced by Vi ……….. A booster response should occur after the 2nd dose ……….. 9
Mouse Data (Post 3 rd Dose) Fold-increase over Placebo 10000 Fold IgG Increase over Placebo Fold IgG Increase over Placebo 10000 Individual Pooled sera 1000 1000 100 100 10 10 1 1 BE ViPS Vi-CJ Ctrl ViPS Ctrl BE Vi-rCRM (Lot1) BE Vi-rCRM (Lot2) ) ) S l l r r 1 2 P t t t t C C o o i V L L J S ( ( C E P M M B - i i V V R R C C r r - - i i V V E E B B 10
Anti- Vi IgG ELISA: Fold Increase Over PS Baseline 100 Fold Anti-Vi IgG Increase ( v/s Vi-PS Control) Samples 1. BE Vi-rCRM Lot 1 2. BE Vi-rCRM Lot 2 3. BE Vi PS control 4. Vi-CJ positive control 5. Vi PS negative control 10 ~ Conclusions: 3 1. Significant (30X) increase in anti- 0 WHO 4 Vi IgG levels observed for both X TRS Vi-rCRM samples when compared to PS baseline. 2. BE product meets the 4X PS threshold mentioned in WHO 1 Baseline TRS. BE Vi-rCRM (Lot1) BE Vi-rCRM (Lot2) BE ViPS Vi-CJ Ctrl ViPS Ctrl 11
Evidence of Booster Response – Dose I and III Post-DoseI 1000 Post-DoseIII Fold-increase in IgG Fold-increase over placebo baseline 100 10 Vi-CJ ctrl ViPS ctrl BE Vi-rCRM (Lot1) BE Vi-rCRM (Lot2) BE ViPS 12
Initial Immunogenicity Evaluation Vi-rCRM 197 : Conclusions • BE Vi-rCRM is highly immunogenic in mice. BE Vi-rCRM preclinical immunogenicity results meet WHO TRS requirements. • BE Vi-rCRM elicits a booster response in mice. • BE Vi-CRM conjugate have similar immunogenicity to other reported conjugates Vi-CRM by NVGH Vi-rEPA by Szu et al Vi-rCRM by Eubiologics Vi-DT by IVI 13
Next Steps • BioE Vi-CRM targeted to be in clinical trials in 1Q16 • Additional lots under preparation for preclinical immunogenicity evaluation in mice and rabbit models • BioE also working on bivalent TCV candidate vaccine (Vi-CRM and O:2-CRM). Preliminary preclinical immunogenicity evaluation ongoing. 14
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