Creation of immune 'stealth' genes for gene therapy through fusion with the Gly-Ala repeat of EBNA-1.
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 beta-galactosidase, firefly luciferase, and HSV1 thymidine kinase) fused with the GAr retained their function. Moreover, linking GAr with beta-galactosidase successfully prevented recognition of GAr-LacZ-expressing cells by beta-galactosidase-specific CTL. Nonetheless, vaccination with a GAr-LacZ adenovirus or with an allogeneic cell line expressing GAr-LacZ resulted in the induction of beta-gal-specific CTL. This demonstrates that the GAr domain does not inhibit cross presentation 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.
Ossevoort
M
M
Department of Molecular Cell Biology, Leiden University Medical Center, Leiden, The Netherlands.
Visser
B M J
BM
van den Wollenberg
D J M
DJ
van der Voort
E I H
EI
Offringa
R
R
Melief
C J M
CJ
Toes
R E M
RE
Hoeben
R C
RC
D000256:Adenoviridae / Q000235:genetics; D000818:Animals; D017951:Antigen Presentation / Q000235:genetics; D019677:Artificial Gene Fusion; D003602:Cytotoxicity, Immunologic / Q000235:genetics; D004151:Dipeptides / Q000235:genetics; D019309:Epstein-Barr Virus Nuclear Antigens / Q000235:genetics*; D015316:Genetic Therapy / Q000379:methods*; D005822:Genetic Vectors; D007108:Immune Tolerance / Q000235:genetics*; D007763:Lac Operon; D051379:Mice; D008807:Mice, Inbred BALB C; D013602:T-Lymphocytes, Cytotoxic / Q000276:immunology*; D019076:Transgenes / Q000276:immunology; D001616:beta-Galactosidase / Q000378:metabolism
D004151:Dipeptides; D019309:Epstein-Barr Virus Nuclear Antigens; C065992:N-glycylalanine; D001616:beta-Galactosidase; C417029:EBV-encoded nuclear antigen 1