Abstract
BackgroundHuman papillomavirus (HPV) infection is known to be the most important etiologic factor of cervical cancer. There is no HPV specific therapy available for treatment of invasive squamous cell carcinoma of the cervix and its precursor lesions. The present study elucidates the potential to use herpes simplex virus (HSV) derived vectors for expression of antisense RNA to HPV -16 E7 oncogene.ResultsWe have constructed replication competent, nonneuroinvasive HSV-1 vectors, deleted of the γ134.5 gene. The vectors express RNA antisense to the first 100 nucleotides of the HPV-16 E7 gene. We assayed the ability of the antisense E7 vectors R5225 (tk-) and R5226 (tk+), to produce antisense RNA, as well as the consequent effects on E7 mRNA and protein levels in HPV-16 positive CaSki cells. Anti-E7 RNA was expressed by both constructs in a dose-dependent manner. Expression of HPV-16 E7 mRNA was downregulated effectively in CaSki cells infected with the tk- recombinant R5225 or with R5226. The tk+ recombinant R5226 was effective in downregulating E7 protein expression.ConclusionWe have shown that anti-E7 RNA expressed from an HSV vector could efficiently downregulate HPV-16 E7 mRNA and E7 protein expression in CaSki cells. We conclude that HSV vectors may become a useful tool for gene therapy of HPV infections.
Highlights
Human papillomavirus (HPV) infection is known to be the most important etiologic factor of cervical cancer
The first 100 nucleotides of the E7 gene of HPV-16 were cloned in antisense orientation in to a plasmid pRB 4878 [29] under the egr-1 promoter, flanked by sequences derived from the γ134.5 gene of herpes simplex virus (HSV)-1 (F)
In separate experiments we observed that the expression of the viral transgenes was highest at 16–20 h.p.i. in our cell culture setting, and the effects on the HPV-16 E7 mRNA were most evident at 16 h.p.i
Summary
Human papillomavirus (HPV) infection is known to be the most important etiologic factor of cervical cancer. There is no HPV specific therapy available for treatment of invasive squamous cell carcinoma of the cervix and its precursor lesions. E7 can interact with the pRb tumor suppressor protein [8,9,10,11], which results in release of transcription factor E2F, leading to increased cell cycle progression. CaSki cells, originally derived from a human cervical cancer [13], contain approximately 600 copies of HPV-16 DNA, and the E7 gene is continuously expressed in these cells
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