DNA vaccine against human papillomavirus type 16: modifications of the E6 oncogene.
Since its discovery, DNA vaccination has become an effective strategy for the development of vaccines against cancer including cervical carcinoma (CC). The formation of CC is associated with human papillomavirus (HPV) infection. Viral E6 and E7 oncoproteins are suitable targets for therapeutic vaccination. To adapt the HPV16 E6 oncogene for DNA immunisation, we performed several modifications. First we fused the E6 gene with the 5' or 3'-terminus of the Escherichia coli beta-glucuronidase (GUS) gene and showed enhanced immunogenicity of the 3' fusion (GUS.E6). Then, as the E6 oncogene contains two alternative introns that result in the production of truncated forms of the E6 protein, we abolished the 5' splice site in the E6 gene. This modification completely eliminated the expression of the truncated E6 transcripts and thus increased the production of the full-length E6 protein. At the same time, it moderately reduced the immunogenicity of the modified non-fused (E6cc) or fused (GUS.E6cc) genes, probably as a consequence of the substitution in the immunodominant E6 epitope following the abolishment of the splice site. Furthermore, we reduced the oncogenicity of the E6 protein by two point mutations (E6GT) that, together, prevented E6-mediated p53 degradation. Finally, we constructed the GUS.E6GT gene characterized by enhanced safety and immunogenicity when compared with the wild-type E6 gene.
Poláková
Ingrid
I
Department of Experimental Virology, Institute of Hematology and Blood Transfusion, U Nemocnice 1, 12820 Prague 2, Czech Republic. ingrid.polakova@uhkt.cz
Pokorná
Dana
D
Dusková
Martina
M
Smahel
Michal
M
D000818:Animals; D000914:Antibodies, Viral / Q000097:blood; D002460:Cell Line; D016207:Cytokines / Q000096:biosynthesis; D029968:Escherichia coli Proteins / Q000235:genetics / Q000276:immunology; D005260:Female; D015870:Gene Expression; D005966:Glucuronidase / Q000235:genetics / Q000276:immunology; D052162:Human papillomavirus 16 / Q000235:genetics / Q000276:immunology*; D006801:Humans; D007438:Introns; D007963:Leukocytes, Mononuclear / Q000276:immunology; D051379:Mice; D008810:Mice, Inbred C57BL; D009369:Neoplasms / Q000517:prevention & control; D009856:Oncogene Proteins, Viral / Q000235:genetics / Q000276:immunology*; D053918:Papillomavirus Vaccines / Q000235:genetics / Q000276:immunology*; D017354:Point Mutation; D011993:Recombinant Fusion Proteins / Q000235:genetics / Q000276:immunology; D012097:Repressor Proteins / Q000235:genetics / Q000276:immunology*; D019444:Vaccines, DNA / Q000235:genetics / Q000276:immunology*
D000914:Antibodies, Viral; D016207:Cytokines; C068651:E6 protein, Human papillomavirus type 16; D029968:Escherichia coli Proteins; D009856:Oncogene Proteins, Viral; D053918:Papillomavirus Vaccines; D011993:Recombinant Fusion Proteins; D012097:Repressor Proteins; D019444:Vaccines, DNA; D005966:Glucuronidase