1. Human and Rhesus Macaque Immunologic Responses To Anthrax Vaccine Adsorbed CDC Anthrax Vaccine Research Program Conrad P. Quinn Chief, MPIR Laboratory MVPD/DBD/NCIRD FDA Advisory Committee Meeting 16th November 2010 The findings and conclusions in this report are those of the authors and do not necessarily represent the official position of the Centers for Disease Control and Prevention. National Center for Immunization & Respiratory Diseases Division of Bacterial Diseases

2. CDC Anthrax Research  AVRP  Anthrax Vaccine Adsorbed Phase 4 human clinical trial o Double blind, placebo controlled o Route change, dose reduction Non-human primate vaccine correlates of protection o Inhalation anthrax model in Rhesus macaques  AIGIV  Rabbit passive transfer model o Ambulatory NZW infusion model o Human anti-AVA serum o Proof of concept; prophylactic intervention for inhalation anthrax

3. The CDC Anthrax Vaccine Research Program  Document & assure efficacy of AVA  Minimize doses & optimize schedule  Define the AVA correlate of protection  Find an immunologic marker(s) that: Endorses Human Clinical Trial endpoint Confirms human vaccinee protection Identifies when protection is achieved Quantifies how long protection lasts

4. An Integrated Study in Humans & Rhesus macaque NHP AVA Human Clinical Trial AVA Macaque Dose Ranging & Immunogenicity Study Immunocompetence Model Building Survival Against Virulent Challenge NHP Immune Correlates of Protection Human Surrogate Markers of Protection Human Humoral & Cellular Immune Profiles Macaque Humoral & Cellular Immune Profiles Hu-AVA 1/5 – 1/40

5. Challenge vs. Vaccination in Rh. Macaques Study Group IA Label Week 0 Week 2 Week 4 Month 6 Month 12 Month 18 Month 30 Month 42 8SQ 4SQAVA 8IM 4IMAVA 7IM 3IM AVAS 5IM AVAS S S 4IM AVAS SS S IM Placebo PLAC SSSSSS SS SQ Placebo PLAC SSSSSS SS Rh. macaque challenge time points

6. Building Immunological Profiles  Modulation of rh. macaque immune response (3IM)  Variable schedules in humans  Quantification of antibody levels and functional competence (ELISA & TNA)  Th1 – Th2 disposition  Onset and duration of T cell memory & competence (SI)  Onset and duration of B cell memory  Data mining and bridging to humans  Correlate of protection in rh. macaques  Surrogate of protection in human vaccinees

7. Antibody Levels and Functional Competence Are Highly Correlated y = 0.95x + 0.84 R 2 = 0.86 (n=6875) Anti-PA IgG (µg/ml) Lethal Toxin Neutralization Titer (ED50) Regression Human –n=4568, Rsqrd=0.8672 LOGED50=0.7811+0.9774LOGIGG –TNA range ED50 0-40,000 –ELISA range 0-5781.7 (0-4585.6 when a TNA result is present)

8. Diluted AVA Elicits Long-term Circulating Anti-PA IgG 5.32 µg/ml 3.40 µg/ml 1.22 µg/ml 01m6m 5.32 µg/ml 3.40 µg/ml 1.22 µg/ml vaccination

9. Diluted AVA Elicits Potent Anamnestic Responses to infection in Rh. Macaque NHP vaccination challenge 5.32 µg/ml 3.40 µg/ml 4211 µg/ml 3134 µg/ml 37 µg/ml 25 µg/ml 2714 µg/ml 1009 µg/ml 2669 µg/ml 2.63 µg/ml

10. PA Specific NHP T Cell Competence is Maintained for 52m

11. 3-IM Human Dose and Diluted AVA Provides Long Term Protection in Rhesus Macaque NHP 3-IM priming series. No booster vaccinations. Number Survived / Number Challenged (Survival Rate) Challenge Time Undiluted1/51/101/201/40 12 Months-- 8/10 (80.0%) 11/20 (55.0%) 13/20 (65.0%) 30 Months 10/10 (100.0%) 8/8 (100.0%) 6/9 (66.7%) 7/8 (87.5%) - 52 Months 8/10 (80.0%) 9/9 (100.0%) 6/10 (60.0%) --

12. Human 3-IM Priming Establishes Long Term Responses PLAC 433.20 310.02 254.80 320.45 216.83 7IM 47.77 8IM 38.07 8SQ 35.69 5IM 21.58 4IM 6.02 PLAC 1.85 3 yrs 6.02 (5.28-6.87) 1yr 21.58 (18.9-24.7)

13. PA Specific Human T Cell Competence is Maintained for 43m T cell SI were detectable by 0.5m, significantly above control levels by 1m, and sustained for 43m irrespective of schedule.

14. Rabbit Passive Transfer Studies – AIGIV Human anti-AVA IgG fraction Minimum of 4 doses AVA SC Single intravenous infusion 24hr prior to aerosol exposure 3 dose levels human AIGIV 200 LD50 equivalents B. anthracis Ames Flebogamma protein control Ambulatory, minimal invasion, maximum access Facilitates long term, multiple infusions, combination therapies

15. GMC AIGIV Profiles in Rabbits Challenge Infusion 608 µg/ml 333 µg/ml 145 µg/ml 292 µg/ml 161 µg/ml 65 µg/ml

16. AIGIV Passive Transfer Survival 20mg/kg AIGIV 10mg/kg AIGIV 5mg/kg AIGIV Flebogamma Controls Naïve Controls 292 µg/ml 161 µg/ml 65 µg/ml GMTTD 3.9d GMTTD 4.9d GMTTD 5.7d

17. Correlates of Protection Modeling  Approach:  David Madigan, Columbia U, NY Least Squares Regression Analyses LASSO - automatic variable selection and estimation algorithm via constrained maximum likelihood logistic regression o Evaluation of NHP data Bayesian multi-level models o Bridging between genera  Chuck Rose, Lydia Foster, CDC Linear Mixed Effects Model (log-log transformed) Anti-PA IgG longevity and decay in humans and NHPs

18. Active Immunization Antibody Decay in Humans and NHP fits a Conventional Power Law Model Human and NHP1/5_AVA slopes and intercepts are not statistically significantly different Human NHP_1/5_AVA NHP_1/10_AVA NHP_Hu_AVA NHP_1/20_AVA N = 94 vaccinated animals From C.Rose & L.Foster

19. Segregation of NHP Survivors by wk30 Anti-PA IgG Levels and Decay Kinetics

20. Interpretive Value of the GUP Anti-PA IgG Response  Significant NHP survival at 30 and 52 months post- vaccination  IgG and TNA ED50 are correlated with survival  Other assays (SI) only weakly correlate with survival  IgG and TNA ED50 are highly correlated with each other  NHP anti-PA IgG responses to1/5 diluted 3-IM AVA and human responses to a 3-IM schedule fit the same decay model  Peak response to 3-IM vaccination combined with decay kinetics may be useful in predicting protection in NHP

21. 1.Anti-PA IgG antibodies are an adequate, if not perfect, correlate of protection from which to bridge between animal models and humans 2.Bridging from animals to humans may be achieved from active immunization studies 3.Antibody passive transfer studies  comprise only one facet of the protective immune response  are required to overcompensate for the absence of CMI  may set unnecessarily high requirements for measureable circulating antibody from active immunization 4.GUP vaccine efficacy informs PEP  Effects of concomitant abx on human immune response to AVA need to be evaluated Conclusions