657. Immune Responses to Transgene Expression Targeted to Keratinocytes

Transgene-specific immune responses impose a major hurdle to effective implementation of clinical gene therapy, especially in situations where neoantigen is expressed. Previously, in our studies of cutaneous gene therapy, we showed that neoantigen expression leads to a potent immune response and rejection of the retroviral-transduced epidermal cells in normal mice. As cutaneous gene therapy will often require prolonged transgene expression, we seek a strategy to avoid this host response. One such strategy utilizes the fact that antigen presentation by non-professional antigen presenting cells (APCs), including keratinocytes, may result in T cell unresponsiveness. To explore this concept, the dorsal skin of immunocompetent mice was transduced in vivo with 2 × 107 transducing units of either standard retroviral vector encoding GFP or retroviral vectors in which GFP expression is targeted to the suprabasal layers of epidermis. Initial GFP expression in both groups of animals was confirmed at 1-week post transduction by fluorescent stereo microscopy. Examination at 3 weeks however, indicated loss of GFP expression in both groups of transduced mice coinciding with the generation of significant levels of anti-GFP antibody. Although expression was directed to epidermal keratinocytes, the acute loss of transduced cells may have resulted from uptake of GFP entrapped in the viral particle by professional APCs, which are abundant in skin. This possibility is currently being examined with an ex vivo model of cutaneous gene transfer in which transgene expression will be confined solely to keratinocytes and transplanted to immunocompetent mice.