P05

In recent years, the use of recombinant Adeno-Associated Viruses vectors (rAAV) in gene therapy to treat genetic diseases have proven effectiveness although it raised some unexpected toxicity concerns due to the activation of the immune system in some patients. This immunity against AAV capsids and transgene products compromises the efficacy of the therapy, but can also lead in some cases to important safety issues. The innate immune detection of rAAV vectors was described to involve the TLR9 (Toll Like Receptor 9) pathway through the recognition of unmethylated CpG islands of rAAVs' single-stranded DNAs. For this reason, we assessed rAAV9 genome-engineered viral vectors where TLR9-inhibitory sequences, called DIMS (DNA-based immunomodulating sequences) were inserted, after in vivo high vector doses administration. Sprague Dawley rats were intravenously injected with 10e14viral genomes/kg of rAAV9 vectors carrying a GFP expression cassette, with or without the DIMS sequences. The efficacy and immunogenicity of gene transfer were performed at 3- and 6-months post-injection (p.i.). Although the effect on the adaptive immune response was moderate, DIMS sequences increased the amount of viral genome-derived transcripts in muscle and cardiac tissues at 3 months following vector delivery. Moreover, while 30% of rAAV9 control rats have shown a loss of transgene expression in the liver, engineered rAAV enabled GFP persistence at 3 (n=10/10) and 6 months p.i. (n=4/5). Altogether, these results highlight the potential of rAAV9 DIMS-engineered vectors to improve the efficacy of rAAV-based gene therapies and possibly immunogenicity-related side effects.