Laboratoire Immunité & Infection INSERM UMR-S 945 IFR 113 Paris, France

1. Laboratoire Immunité & Infection INSERM UMR-S 945 IFR 113 Paris, France Towards an innovative therapeutic vaccine against HIVgp41preventing CD4 cells sensitivity to NK killing P DEBRE

2. PATHOGENESIS INFECTION  CD4 non infected cells Virus  CD4 infected cells NK ? HIV, CD4 & NK cells

3. 40 30 UT 3S 20 10 CD4 2.7% 17.4% 0 NKp44L HIV-1 gp41 Env Fusion Peptide HR1 HR2 TM 517 532 643 678 558 595 3S motif 1- Background : NKp44L is induced during HIV infection by the 3S/gp41 motif Kill % NKp44L CD4 4+ 3- 8+ 8+ 4+ 3- Control PBMC HIV+ PBMC NKp44L MHC-I 40 NKp44 - + p=0.002 30 % NKp44L NK 20 10 0 0 250 500 1000 750 3S : very conserved motif & specific to HIV-1 CD4 / mm3

4. Summary 40 44L+ 30 % NK Lysis 20 44L- Autologous NK cells HIV infected cells 10 0 100/1 50/1 25/1 12.5/1 E/T ratio HIV virion NKp44 NKp44L gp41 Epitope 3S (SWSNKS) CD4+NKp44L+ cell CD4+ T cell CD4+ T cell lysis Anti-3S Ab ex vivo In vitro CD4+ T cell depletion ? HIV1-infected individuals

5. gC1qR is the cell-surface receptor of the 3S motif on CD4+ T cells 3S IgM NA HK C1q 3S C3 3S+ agC1qR NA 0 25 50 75 100 125 150 175 NKp44L expression NKp44L expression (MFI) Listeria monocytogenes Staphylococcus aureus Plasmodium falciparum HCV, rubella virus, HSV and HTNV Deletrious effect of NK cells ? Role of gC1qR ! Fausther-Bovendo et al; PLoS Pathogens 2010

6. 2- Proof of concept in macaques 2-1 : Macaque model of SHIV infection (Vieillard et al, AIDS 2008) 2-2 : Prophylactic vaccination (Vieillard et al, PNAS 2008) 2-3 : Therapeutic vaccination (Vieillard et al, submitted) Collaboration: Roger Le Grand (CEA, Fontenay-aux-roses; France)

7. SHIV162P3 « prophylactic » vaccination by the 3S-gp41 peptide 3S 3S 3S 1500 NS NS 3S 3S 108 106 1000 3S anti-3S (U/mL) Viral load 104 3S 500 102 0 1 -400 200 -200 0 0 50 100 150 200 50 2000 ** * 40 1500 % CD4+NKp44L+ 30 CD4 / mm3 1000 20 500 10 0 0 0 50 100 150 200 0 50 100 150 200 Days post-infection KLH-control macaques 3S-immunized macaques

8. SHIV162P3 50 40 30 CRP (mg/ml) 20 100 ** ** 50 ** ** 10 * 75 40 ** 0 % NK lysis 30 50 * % CD4+AnnexinV+ * 20 25 10 0 0 0 14 21 42 LN UI 14 21 42 LN Days post-infection Days post-infection Effect of 3S vaccination on Apoptosis, NK lysis and inflammation SHIV162P3 80 * 60 ** TNF-(pg/ml) ** 40 20 0 0 113 390 397/7 411/21 0 113 390 397/7 411/21 Days post-immunization Days post-immunization 3S-immunized macaques KLH-control macaques

9. Conclusions • Immunogenicity in cynomolgus (anti 3S response) • BUT Unchanged viral load • Decreased NKp44L expression on CD4+ T cells • Decreased NK cells cytotoxicity on CD4+ T cells • Decreased CD4+ T cells apoptosis • Preservation of CD4+ T cell counts • Decreased immune activation • Decreased inflammation during the acute phase

10. « Therapeutic » vaccination by the 3S-gp41 peptide 1000 800 600 Anti-3S (AU/ml) 400 200 0 UI UI 0 0 30 30 10 20 10 20 Weeks post-immunization Weeks post-immunization 15000 10000 Anti-KLH (AU/ml) p=0.0006 5000 40 60 ** ** ** 30 0 40 UI 0 30 10 20 % NKp44L Weeks post-immunization 20 % NKp44L 20 10 Poor responder macaque KLH-control macaques 3S-immunized macaques 0 0 Lymph nodes Spleen Rectum

11. Effect of 3S vaccination on NK lyis K562 CD4 auto P<0.0001 40 50 40 30 30 20 % specific lysis 10 10 0 0 30 30 20 20 20 10 10 0 0 Weeks post-immunization KLH-control macaques 3S-immunized macaques Poor responder macaque * * * * IgM control % specific lysis Anti-NKp44L 30 W0 W10 W32 20 Weeks post-immunization 10 0

12. Effect of 3S vaccination on apoptosis 60 40 40 * * * * 20 30 Lymph nodes 0 PB Spleen Rectum 20 %CD4+AnnexinV+ %FAS-induced killing 10 0 Lymph nodes Spleen 3S-vaccinated Controls 50 * * 8.7% 28.4% 40 30 % caspase-3 activity 20 10.2% 24.5% 10 0 Spleen Lymph nodes % Caspase-3 activity KLH-control macaques 3S-immunized macaques

13. Effect of 3S vaccination on central memory CD4 T cells p=0.0401 800 600 CD4+CD28+CD95+/mm3 UI 0 30 10 20 400 200 0 Weeks post-immunization KLH-control macaques 3S-immunized macaques Poor responder macaque

14. Conclusions • Immunogenicity in cynomolgus (anti 3S response) • Decrease of NKp44L expression • Decrease of apoptosis in lymphoid tissues • Lower lysis activity of NK cells against autologous CD4+ T cells • Preservation of central memory CD4+ T cells

15. Perspectives Clinical trial Phase I/IIa Animal Model Therapeutic Vaccine Non-infected Asymptomatic phase AIDS HIV-1 infection Antiretroviral Therapy

16. Partners IFR 113 CEA Laboratoire d’ Immuno-Virologie Roger Le Grand Nathalie Deudreude-Bosquet Aurélien Corneau Isabelle Mangeot-Méderlé … Unité Immunité & Infection Team-2 Patrice Debré Hugues Fausther Bovendo Caroline Petitdemange Alexis Sennepin Vincent Vieillard Team-1 Brigitte Autran Assia Samri Unité Epidémiologie Clinique Dominique Costagliola Service de Virologie Henri Agut Vincent Calvez Daniel Candotti Isabelle Malet Service des Maladies Infectieuses & Tropicales Christine Katlama Grants ANRS Sanofi-Pasteur Bill & Melinda Gates Foundation Agence Nationale de Recherche (ANR) InnaVirVax ORVACS Institut Pasteur Olivier Schwartz Nathalie Sol-Foulion Felix Rey