702. Detection and Isolation of Immune Cells After Intramuscular AAV Injection in Mouse and Human

Abstract

Recombinant adeno-associated viral (rAAV) vectors are promising tools for gene transfer to the skeletal muscle. However, studies on mouse and large animal models as well as humans demonstrate that intramuscular (IM) rAAV delivery can trigger immune responses to AAV capsid and/or transgene. Recently, IM delivery of rAAV1 in humans induced a tolerogenic response to rAAV characterized by the presence of capsid-specific regulatory T cells (Tregs) in the periphery. The aim of our study was to develop an assay to allow detection and characterization of immune cells in situ after AAV gene transfer. Mice were injected intramuscularly with an AAV vector. Seven days after injection, muscles were collected and enzymatic dissociation was performed. Our results show that we are able to detect immune cells (monocytes/macrophages, lymphocytes) in mouse muscle by flow cytometry. This assay was also used on a human muscle biopsy collected 5 years after AAV1-AAT gene transfer and revealed the presence of infiltrated Tregs and CD8+ T-cells at the site of injection. This method appears as a new tool in understanding immune responses in the muscle after AAV gene transfer.