Silencing of Integrated Human Papillomavirus-16 Oncogenes by Small Interfering RNA-Mediated Heterochromatization

Double-stranded RNAs or small interfering RNAs (siRNA) targeting the promoters of genes are known to cause gene knockdown by a process known as transcriptional gene silencing (TGS). We screened multiple siRNAs homologous to one of the NF-1 binding sites in the human papillomavirus-16 (HPV-16) enhancer and identified one siRNA which causes specific TGS of the HPV-16 oncogenes E6 and E7 when transfected into two HPV-16-positive cell lines siHa and CaSki. This phenomenon was specific to the HPV-16 enhancer with no effect on the HPV-18 enhancer. TGS was associated with heterochromatization of the targeted region of the enhancer but no DNA methylation was noted during the time period studied. The choice of target in the enhancer was important as siRNAs differing by one or two bases showed no suppression of downstream gene expression. A low copy number enhancer-associated transcript was detected in the cell lines studied and its level decreased significantly after treatment with the siRNA that caused TGS. This supports the RNA: RNA model described previously for TGS. This siRNA which causes simultaneous silencing of E6 as well as E7 oncogenes by an epigenetic mechanism might be useful as a therapeutic modality for HPV-16-positive cervical and other epithelial cancers. Mol Cancer Ther; 9(7); 2114-22. (C) 2010 AACR.