Mondal, Gourish

Abstract Purpose: Recent studies suggest that SIRT1-activating compounds (STAC) are a promising class of anticancer drugs, although their mechanism of action remains elusive. The main goal of this study is to determine the role of STACs as a potential therapy for pancreatic cancer. In addition, we also explored the mechanism by which these compounds affect pancreatic cancer. Experimental design: Using in vitro (cell culture experiments) and in vivo (xenograft experiments) approaches, we studied the role of SIRT1 agonists (STAC) in human pancreatic cancer cell viability and growth. Results: We show that SIRT1 is highly expressed in pancreatic cancer cells and that the STACs SRT1720, SRT1460, and SRT3025 inhibited cell growth and survival of pancreatic cancer cells. STACs enhanced the sensitivity of pancreatic cells to gemcitabine and paclitaxel, indicating that these drugs could be used in combination with other chemotherapy drugs. We also show that STACs were very effective in inhibiting tumor xenograft growth. In mechanistic studies, we observed that STACs activated a SIRT1 lysosomal–dependent cell death. Furthermore, the effect of STACs on cell viability was also dependent on the expression of the endogenous SIRT1 inhibitor DBC1. Conclusions: Taken together, our results reveal an essential role for SIRT1 and lysosomes in the death pathway regulated by STACs in pancreatic cancer cells. Clin Cancer Res; 22(10); 2496–507. ©2015 AACR.

One of the mechanisms of tumorigenesis is that the failure of cell division results in genetically unstable, multinucleated cells. Here we show that pVHL, a tumor suppressor protein that has been implicated in the pathogenesis of renal cell carcinoma (RCC), plays an important role in regulation of cytokinesis. We found that pVHL-deficient RCC 786-O cells were multinucleated and polyploid. Reintroduction of wild-type pVHL into these cells rescued the diploid cell population, whereas the mutant pVHL-K171G failed to do so. We demonstrate that lysine 171 of pVHL is important for the final step of cytokinesis: the midbody abscission. The pVHL-K171G caused failure to localize the ESCRT-1 interacting protein Alix and the v-SNARE complex component Endobrevin to the midbody in 786-O cells, leading to defective cytokinesis. Moreover, SUMOylation of pVHL at lysine 171 might modulate its function as a cytokinesis regulator. pVHL tumor suppressor function was also disrupted by the K171G mutation, as evidenced by the xenograft tumor formation when 786-O clones expressing pVHL-K171G were injected into mice. Most RCC cell lines show a polyploid chromosome complement and consistent heterogeneity in chromosome number. Thus, this study offers a way to explain the chromosome instability in RCC and reveals a new direction for the tumor suppressor function of pVHL, which is independent of its E3 ubiquitin ligase activity.

Abstract Microcephalin (MCPH1) is a BRCA1 COOH terminal (BRCT) domain containing protein involved in the cellular response to DNA damage that has been implicated in autosomal recessive primary microcephaly. MCPH1 is recruited to sites of DNA double-strand breaks by phosphorylated histone H2AX (γH2AX), but the mechanism by which MCPH1 contributes to the repair process remains to be determined. Here, we show that MCPH1 binds to BRCA2 and regulates the localization of BRCA2 and Rad51 at sites of DNA damage. The interaction occurs through the NH2 terminus of BRCA2 and the COOH terminal BRCT domains of MCPH1. Disruption of the interaction between MCPH1 and BRCA2 has no effect on the ability of BRCA2 to form a complex with Rad51 but is associated with substantially reduced levels of both BRCA2 and Rad51 at sites of DNA double-strand breaks. Uncoupling of MCPH1 from BRCA2 also interferes with Rad51-dependent and BRCA2-dependent homologous recombination repair activity. These results suggest that the role of MCPH1 in the DNA damage response is in part associated with the ability to localize BRCA2 to sites of DNA double-stand breaks. [Cancer Res 2009;69(13):5531–6]