Cervical cancer accounts for approximately 7% of all female cancers in developed countries and nearly 5,000 deaths annually in the United States. Numerous clinical and epidemiological studies have established a strong link between human papillomavirus type 16 and 18 infections and cervical cancer. Virus infection and gene expression are necessary but not sufficient to induce malignancy. Other factors are required for malignant progression of infected cells. Molecular studies have revealed that the mechanisms of cellular transformation used by HPV oncogenes (E6 and E7) are strikingly similar to those of other DNA tumor viruses such as SV40 (T-Ag) and adenovirus (E1a/E1b). An appealing approach for designing novel therapeutic approaches for the treatment or prevention of HPV-induced cervical cancer is the amelioration of this malignant progression by inhibiting HPV functions. The common, nonpathogenic, defective human parvovirus, Adeno-associated Virus (AAV) has generated much excitement lately because of its potential uses as a gene therapy vector. Another exciting aspect of AAV is that its replication (rep) gene suppresses cellular transformation mediated by the SV40 T-Ag gene, the bovine papillomavirus E5 gene and an adenovirus E1a and activated ras gene pair. To demonstrate the effectiveness of AAV as a therapeutic agent against cervical cancer, we will suppress in vitro HPV18 transformations using focus formation assays in both fibroblast and epithelial cell lines. These experiments will also verify that the rep gene suppresses HPV18 E7-mediated activation of transcription promoters. In order to determine how AAV and the rep gene suppress HPV and E1a/ras mediated transformation, we will investigate the mechanisms by which AAV naturally inhibits adenovirus infections. Using wild type and mutant AAV viruses we will determine how AAV inhibits adenovirus; (i) DNA replication, (ii) gene expression and (iii) induction of cellular DNA synthesis. In these experiments we will also characterize protein- protein interactions mediated by the AAV Rep proteins which play significant roles in their functioning. This proposal will open up an entire field investigation into the development of AAV based gene therapy vectors for the prevention and treatment of cervical cancer.
| Yang, Q; Chen, F; Trempe, J P (1994) Characterization of cell lines that inducibly express the adeno-associated virus Rep proteins. J Virol 68:4847-56 |
