OR23

Muscle-specific expression reduces early antigen presentation and promotes CD8 T cell tolerance after rAAV gene transfer

L Jeanpierre(1,2) C Pecquet(1,2) H Saliba(1,2) P Finard(1,2) S Terry(1,2,3) G Tavella(1,2) S Boutin(1,2) B Bertin(1,2) S Benkhelifa-Ziyyat(4) G Ronzitti(1,2) D -A Gross(1,2)

1:Genethon, UMR_S951, Inserm, Univ Evry, Université Paris Saclay, EPHE; 2:Université Paris-Saclay, Univ Evry, Inserm, Genethon, Integrare research unit UMR_S951, 91000 Evry, France; 3:Research Department, Inovarion, 75005 Paris, France; 4:Sorbonne Université, Inserm, Institut de Myologie, Centre de Recherche en Myologie, 75013 Paris, France

Immune response against the transgene product can significantly reduce the efficacy of AAV-based gene transfer. Although numerous parameters controlling this immunogenicity have been identified, the in vivo mechanisms controlling CD8⁺ T cell responses against the transgene product, particularly the initial events driving T-cell activation, remains poorly understood. In this study, we investigated in mice the kinetic of antigen presentation after intramuscular administration of AAV vectors. We found viral genomes in the lymph nodes draining the injection site as early as one hour post AAV administration and activation of capsid-specific CD8⁺ T cells at 24h, suggesting rapid lymphatic drainage and degradation of AAV particles. Unexpectedly, the transgene product was also presented to CD8⁺ T cells early, with an increasing percentage of activated T cells found between day 1 and day 4, and initial T-cell divisions detected by day 4. Moreover, experiments involving the removal of the injection site demonstrated that AAV particles reaching the draining lymph node within the first hour were sufficient to expand transgene-specific CD8⁺ T cells with in vivo cytotoxic function. Finally, we tested different muscle-specific promoters and found that those promoting early transgene presentation efficiently primed CD8⁺ T cells, whereas reduced transgene presentation correlated with an absence of CD8⁺ T cells. Overall, our findings reveal an unexpected early transgene presentation to T cells which critically influences the efficacy of T cell priming. This paves the way for a new approach to assess immunogenicity potential of AAV constructs.