Creation of immune ‘stealth’ genes for gene therapy through fusion with the Gly-Ala repeat of EBNA-1

Publisher Website
Google Scholar
Abstract
A major obstacle in gene-therapy protocols is T-cell-mediated destruction of transgene-expressing cells. Therefore new approaches are needed to prevent rapid clearance of transduced cells. We exploited the Gly-Ala repeat (GAr) domain of the Epstein–Barr virus nuclear antigen-1, since the GAr prevents cytotoxic T-lymphocyte-epitope generation. Here we show that three different enzymes (viz. the E. coli LacZ gene encoded -galactosidase, firefly luciferase, and HSV1 thymidine kinase) fused with the GAr retained their function. Moreover, linking GAr with -galactosidase successfully prevented recognition of GAr-LacZ-expressing cells by -galactosidase-specific CTL. Nonetheless, vaccination with a GAr-LacZ adenovirus or with an allogeneic cell line expressing GAr-LacZ resulted in the induction of -gal-specific CTL. This demonstrates that the GAr domain does not inhibit crosspresentation of antigens, but only affects breakdown of endogenously synthesized proteins. These data demonstrate how the GAr domain can be exploited to create immuno'stealth' genes by hiding transgene products from CTL-mediated immune attack.