TRANSGENE SA, FRANCE 2. INSTITUT PASTEUR, FRANCE On behalf. AFEF 2013, Lille 02 nd October 2013

Transcription

1 A MULTIVALENT ADENOVIRUS-BASED IMMUNOTHERAPEUTIC FOR TREATMENT OF CHRONIC HEPATITIS B INDUCES BROAD, ROBUST AND POLYFUNCTIONAL T CELLS IN NAIVE MICE AND EXERT AN EARLY ANTIVIRAL EFFECT IN HBV TOLERANT MICE Perrine Martin 1, Clarisse Dubois 1, Emilie Jacquier 1, Alexei Evlachev 1, Houda Boukhebza 1, Sarah Dion 2, Maryline Mancini-Bourgine 2, Ophélie Godon 2, Annie Findeli 1, Yasmin Schlesinger 1, Renée Brandely 1, Jean-Baptiste Marchand 1, Thierry Menguy 1, Nathalie Silvestre 1, Marie-Louise Michel 2, Geneviève Inchauspé 1 1 TRANSGENE SA, FRANCE 2 INSTITUT PASTEUR, FRANCE On behalf AFEF 2013, Lille 02 nd October

2 NO CONFLICT OF INTEREST TO DECLARE 2

3 Rationale for an immunotherapeutic product to treat Chronic Hepatitis B (CHB) Current therapies based on nucleoside analogs (NA) or peg-ifnα achieve the control of HBV replication but rarely lead to complete cure of HBV patients Current small molecules aiming at treating CHB do not recruit the host immune system yet: A strong inverse correlation exists between HBV specific functional T cells and control/eradication of viremia (many publications contrast with HIV/HCV) HBV DNA HBV infection Cytokine mediated mechanisms Innate immunity (NK and NKT cells) CD8 + T cells Resolved HBV infection Clearance of HBV DNA Cytolytic mechanisms Major role of CD8+ T cells (and CD4+ T cells to a less extent), Broad and multispecific (Core, Pol and HBsAg) 3

4 Objective and product description OBJECTIVE Develop a viral vector-based HBV-specific immunotherapeutic inducing potent, multispecific, sustained and cross reactive T cell responses displaying properties of immune responses detected in HBV resolving patients Based on non-replicative E1 and E3 deleted human adenovirus serotype 5 encoding a fusion protein comprising truncated HBV Core fused to a deleted and mutated HBV polymerase and 2 selected HBsAg domains (genotype D sequence) 1 Core t Pol1 Env 1 Pol2 Env 2 Pol3* (37 aa) (29 aa) 832 4

5 Key pre-clinical assessments for Immunogenicity in three naive mouse models (HLA-A2, BALB/c and C57BL/6J) following single subcutaneous injection (10 8 iu = 2x10 9 vp /mouse) using IFNγ Elispot, in vivo CTL assays Induction of functional T cells without liver inflammation in a HBV tolerant mouse model AAV-HBV model expressing full-length HBV genome (ML Michel lab, Dion et al 2013 J Virol) 5

6 induces robust and multispecific IFNγ-producing T cells in naive mice IFNγ ELISPOT assay, 2 weeks post-immunisation (pi) HLA-A2 mice BALB/c mice Cumulative number of IFNγproducing cells/10 6 spleen cells Empty Ad C57BL/6J mice Empty Ad Pol Env Core individual mouse median Empty Ad 6

7 induces in vivo functional T cells displaying cytolytic activity in naive mice HLA-A2 transgenic (HLA-A2) and BALB/C (H2 d ) mice, in vivo CTL assays, 2 weeks p.i. CORE ENV % in vivo specific lysis FLP* (HLA-A2) Empty Ad Individual mouse Mean Cut-off value Empty Ad SYV (H2 d ) *FLP epitope (Core ) : associated with spontaneous resolution in patients % in vivo specific lysis VLQ (HLA-A2) Empty Ad IPQ (H2 d ) Empty Ad POL SLY (HLA-A2) HYF (H2 d ) Empty Ad Empty Ad 7

8 evaluation in a AAV-HBV mouse model Model description: Set up and published by Institut Pasteur (Dion et al., J Virol 2013) Performed protocols Based on recombinant AAV coding for 1.2 copies of HBV genome Presence of HBV Ag and HBV replication intermediates in liver Persistence of HBsAg, HBeAg and HBV DNA in sera at least for 1 year Mimic chronic infection by HBV No HBV specific T-cell response and no antibody against HBcAg Presence of regulatory T cells (Tregs) and T-cells producing IL10 in the liver HLA-A2/HLA- DR1 mice Short term (N=2) D0 D21 D28 D32 D38 D42 D46 AAV-HBV( g.e., iv) or PBS or empty Ad + ICS IFNγ/ TNFα/IL2 Blood sample to measure HBV DNA, HBsAg and ALT 8

9 induces functional HBV-specific T cells in the spleens and livers of AAV-HBV injected mice 9 IFNγ/TNFα ICS assay (2 weeks pi) SPLEEN LIVER %of CD8+ T cells producing at least 1 cytokine %of CD8+ T cells producing at least 1 cytokine PBS + AAV-HBV + AAV-HBV + Empty Ad IFN+ TNF+ IL2+ IFN+ TNF+ IL IFN+ TNF+ IL cytokines IFN+ TNF+ IL IFN+ TNF+ IL2+ IFN+ TNF+ IL2+ % responding mice % responding mice

10 induces an early antiviral effect on HBV DNA and HBsAg level in the AAV-HBV model, without ALAT elevation Exp1 Exp2 Fold change (mean +/-SEM) HBV DNA Average HBV DNA titers at D21 : IU/mL * Average HBV DNA titers at D28 : IU/mL Fold change (mean +/-SEM) HBsAg Average HBsAg titers at D28 : 65µg/mL * * * Average HBsAg titers at D28 : 118µg/mL Mean of fold change +/- SEM TG0150 Empty Ad PBS immunization * p< 0.05 (mixed models / repeated measures ANOVA) ALAT (IU/L) (mean +/- SEM) ALAT days days Empty Ad PBS 10

11 Conclusions sc injection Transduction of APC Priming of naive T cells and/or preexisiting HBV T cells CHB patient Migration to the liver + contribution of other cells of immune system (including innate immunity/ad vector) HBV-specific T cells HBV-Infected Hepatocytes HBV-specific T cells LIVER CTL killing Cytokines Hepatocytes killed by T-cell lysis Cleared Hepatocytes Pre-clinical data for in naive and HBV tolerant mouse models show that following ONE single injection of : Induction of HBV-multispecific and long-lasting T cells Induction of IFNγ/TNFα-producing and in vivo functional CTL HBV-specific T cells Migration of induced T cells to the liver where functionality is displayed Early antiviral effect (additional data being collected) No liver inflammation 11

12 Conclusions currently in manufacturing of GMP clinical lot First in Man study planned for second half of 2014 To be combined with current therapies (in particular NA) Foreseen clinical end-points : No exarcerbation of inflammation, safety HBeAg seroconversion HBsAg drop/ HBsAg seroconversion Increase of cure rate 12

13 Acknowledgements TRANSGENE HBV Research Team (Strasbourg and Lyon, France) Nathalie Accart-Gris Emilie Jacquier Houda Boukhebza Catherine Ledoux Renée Brandely Karine Lélu-Santolaria Sandrine Cochin Jean-Baptiste Marchand Clarisse Dubois Bérangère Marie-Bastien Alexei Evlachev Perrine Martin Annie Findeli Thierry Menguy Michel Geist Yasmin Schlesinger Murielle Klein Doris Schmitt Geneviève Inchauspe Nathalie Silvestre AAV-HBV model and experiments INSTITUT PASTEUR, Paris, France, (Pathogénèse des virus de l hépatite B, INSERM U845) Sarah Dion Ophélie Godon Maryline Bourgine Marie-Louise Michel HBV DNA quantification HOPITAL COCHIN, Paris, France (Virology department) Jean François Meritet 13