Characterization of an immuno 'stealth' derivative of the herpes simplex virus thymidine-kinase gene.
The cellular immune response against transgene-encoded neoantigens is a potential hurdle in gene therapy applications where long-term expression of transgenes is desired. Here a new optimized derivative of the herpes simplex virus 1-thymidine-kinase (HSV1-TK) gene is described. The HSV-TK gene is frequently used in experimental studies on gene-directed enzyme prodrug therapy. In the optimized gene, the HSV-TK coding region is fused with the codons for the Gly-Ala repeat of the Epstein-Barr virus nuclear-antigen 1 to prevent proteasomal degradation of the HSV-TK. To measure the protective effect in vitro, a model cytotoxic T lymphocyte epitope derived from the ovalbumin was inserted in the TK. Cells expressing the GAr-modified TK do not present TK-derived peptides in the major histocompatibility complex. Furthermore, conservative nucleotide substitutions were introduced, which prevent splicing, as well as mutations that render the TK-expressing cells more sensitive to ganciclovir (GCV). The GAr HSV-TK fusion protein is fully functional in vitro. This HSV-TK gene may be especially useful in those gene therapy applications where an immune response against the transgene-encoded product would frustrate the treatment.
Ossevoort
M
M
Department of Molecular Cell Biology, Leiden University Medical Center, Leiden, The Netherlands.
Zaldumbide
A
A
Cramer
S J
SJ
van der Voort
E I H
EI
Toes
R E M
RE
Hoeben
R C
RC
D017951:Antigen Presentation; D001483:Base Sequence; D003001:Cloning, Molecular; D019309:Epstein-Barr Virus Nuclear Antigens / Q000235:genetics; D015316:Genetic Therapy / Q000379:methods*; D005822:Genetic Vectors; D006801:Humans; D008969:Molecular Sequence Data; D011971:Receptors, Immunologic / Q000378:metabolism; D011993:Recombinant Fusion Proteins / Q000235:genetics; D018139:Simplexvirus / Q000201:enzymology*; D013602:T-Lymphocytes, Cytotoxic / Q000378:metabolism; D013937:Thymidine Kinase / Q000235:genetics* / Q000276:immunology*
D019309:Epstein-Barr Virus Nuclear Antigens; D011971:Receptors, Immunologic; D011993:Recombinant Fusion Proteins; D013937:Thymidine Kinase; C417029:EBV-encoded nuclear antigen 1