The human papillomavirus oncoproteins E6 and E7 promote cell proliferation and contribute to carcinogenesis by interfering with the activities of cellular tumor suppressors. We used a small interfering RNA molecule targeting the E7 region of the bicistronic E6 and E7 mRNA to induce RNA interference, thereby reducing expression of E6 and E7 in HeLa cells. RNA interference of E6 and E7 also inhibited cellular DNA synthesis and induced morphological and biochemical changes characteristic of cellular senescence. These results demonstrate that reducing E6 and E7 expression is sufficient to cause HeLa cells to become senescent.Human papillomavirus (HPV) infection contributes to the development of over 90% of anogenital cancers (2). HPVs are oncogenic because they lack the DNA and RNA polymerases required for the viral life cycle and therefore must induce production of host cell replication proteins by driving the host cell into a proliferative state. The HPV oncoproteins E6 and E7 direct the progression of the cell cycle and play a major role in HPV-induced carcinogenesis by interfering with host cell regulatory proteins (14). The E7 protein inhibits the function of the retinoblastoma (Rb) family proteins, leading to cell cycle progression (13). In response to abnormal E7-driven proliferation, the host cell triggers apoptosis or senescence by activating the p53 tumor suppressor. To defeat this antiproliferative host cell response, HPVs produce the E6 protein, which targets p53 for degradation (12).While E6 and E7 mask the activities of p53 and Rb proteins, HPV-induced malignancies generally maintain wild-type copies of the p53 and Rb genes. This suggests that reducing E6 and E7 expression in cells transformed with HPV would restore the activity of endogenous tumor suppressors and, thus, prevent proliferation of cells transformed with E6 and E7. Several laboratories have shown that the introduction of the HPV transcriptional regulator E2 into cells transformed with HPV induces either apoptosis or senescence, at least in part by inhibiting E6 and E7 expression (4,7,8,9,17). These studies raise the question of whether E2 expression is necessary to inhibit growth or whether a reduction in E6 and E7 expression alone would be sufficient. In order to determine the effects of reducing E6 and E7 levels in cells without the introduction of additional proteins, we used RNA interference (RNAi).RNAi is a process in which double-stranded RNA (dsRNA) homologous to a specific mRNA dramatically inhibits the expression of the protein made from that mRNA (3, 16). It has been shown that small interfering dsRNA of approximately 21 bases (siRNA) potently induces RNAi in mammalian cells without provoking a nonspecific interferon response (6). Using RNAi to specifically reduce expression of E6 and E7, we found that inhibiting E6 and E7 expression alone is sufficient to induce senescence in HeLa cells.siRNA targeting E7 reduces expression of the E6 and E7 viral oncogenes in HeLa cells. To inhibit E6 and E7 expression in HeLa cells (a cell lin...

The incidence of infection immediately after transplantation did not differ significantly by type of transplant in this pediatric population. Type of transplant predicted increased incidence of infection 30 days posttransplantation and increased incidence of infection with several organisms traditionally associated with a high mortality rate in the transplant population.

In RNA interference (RNAi), double-stranded short interfering RNA (ds-siRNA) inhibits expression from complementary mRNAs. Recently, it was demonstrated that short, single-stranded antisense RNA (ss-siRNA) can also induce RNAi. While ss-siRNA may offer several advantages in both clinical and research applications, its overall poor activity compared with ds-siRNA has prevented its widespread use. In contrast to the poor gene silencing activity of native ss-siRNA, we found that the silencing activity of boranophosphate-modified ss-siRNA is comparable with that of unmodified ds-siRNA. Boranophosphate ss-siRNA has excellent maximum silencing activity and is highly effective at low concentrations. The silencing activity of boranophosphate ss-siRNA is also durable, with significant silencing up to 1 week after transfection. Thus, we have demonstrated that boranophosphate-modified ss-siRNA can silence gene expression as well as native ds-siRNA, suggesting that boranophosphate-modified ss-siRNAs should be investigated as a potential new class of therapeutic agents.