molecular therapeutic approach for human papillomavirus infections

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We have been investigating the possibility of molecular diagnosis of cancer of the uterine cervix by using HPV infection as a marker. First, we demonstrated that there are cases of advanced cancer of the cervix in which HPV16 type DNA is isolated in which it is possible to isolate antibody to HPV16 E7 protein from the patient's serum (Fujii et al.: Jpn J Cancer Res 86:28-34, 1995). We also collected cells by scraping the uterine cervix of patients in whom precancerous lesions of cervical cancer had been found and showed that the rate of detection of E6/E7 transcription products by RT-PCR rose as the lesions became more advanced (Fujii et al.: Gynecol Oncol 58:210-215, 1995). These findings showed that E6/E7 gene products greatly contribute to the development and maintenance of cancer clinically as well. Since E2 protein is a DNA-binding protein that modulates expression of the E6/E7 genes, it can be concluded to be a very important protein in terms of analyzing the mechanism of viral c … More arcinogenesis. The purpose of the present study was therefore to isolate a peptide that binds to E2 protein and inhibits its transcription function. HPV16 E2 protein was expressed in E. coli, and the protein was purified. The E2 protein was converted to the solid phase on plates, and a phage clone that binds to E2 protein was isolated by using a library in which random amino acid sequences were expressed in phage fiber proteins. A peptide sequence containing tryptophan was isolated as a result, and when the tryptophan was replaced with alanine, comparison of binding to E2 protein showed that binding was decreased with the mutant peptide. We also prepared a synthetic peptide in which the nuclear translocation signal was added to the peptide sequence that was isolated, and assessment of its effect on E2 protein transcription activity by the luciferase assay showed a reduction in transcription activity. These findings demonstrated that the peptide that had been isolated binds to E2 protein and suppresses its transcription activity Less