Grant: $311,249 - National Institutes of Health - Jun. 2, 2009
No votes have been cast for this award yet
Award Description: Transposase activity was thought to be extinct in humans because DNA movement can be deleterious in higher organisms, resulting in genomic instability and perhaps malignancy. However, we isolated a human transposase protein termed Metnase that had histone methylase and non-homologous end-joining (NHEJ) DNA repair activity. It was found to interact with DNA Ligase IV, consistent with its NHEJ repair activity. Metnase also was an endonuclease preferential for supercoiled DNA. We therefore explored Metnase's role in decatenating replicated chromatids. Metnase interacted with Topoisomerase II (Topo II ), the critical decatenating enzyme, and enhanced its activity in DNA decatenation, both in vitro and intracellularly. The nuclease activity within the transposase domain of Metnase was required for full enhancement of Topo II decatenating activity. Metnase improved the rate at which Topo II decatenated DNA, and increased the ability of Topo II to resist the decatenation inhibitor ICRF-193. The finding that Metnase improved Topo II resistance to ICRF-193 stimulated an investigation into whether it could mediate resistance to the clinically relevant Topo II inhibitor etoposide. We found that Metnase prevented inhibition of Topo II decatenation by etoposide in vitro, and mediated cellular resistance to etoposide, promoting proliferation in the presence of etoposide. Metnase also promoted a more rapid clearance of etoposide-induced DSB. Thus, Metnase appeared to mediate resistance to etoposide-induced DNA damage and cell cycle arrest. This is a novel mechanism of etoposide resistance that is unexplored. This application proposes to define the mechanism by which Metnase mediates resistance to etoposide by asking three questions: 1) What are the upstream signals that regulate the ability of Metnase to reduce etoposide DSBs? 2) What is the downstream pathway by which Metnase reduces etoposide DSBs? 3) Do Metnase levels predic clinical resistance to etoposide in human malignancy? PUBLIC HEALTH RELEVANCE: We have isolated a novel protein termed Metnase that helps chromosomes repair breaks and also untangle, thereby allowing them to separate properly during cell division. Because of these actions, Metnase mediates resistance to the cancer drug etoposide. Understanding the mechanism by which Metnase does this would allow the generation of drugs targeting Metnase, which could help improve the response of cancer patients to etoposide. In addition, the levels of Metnase could predict which patient will respond to etoposide.
Project Description: In previous studies we have shown that Metnase is a novel DNA repair component that mediates resistance to the DNA damaging agent etoposide. In this grant we are studying the mechanism of Metnase's action, and whether inhibiting Metnase could be exploited clinically, since Metnase mediates etoposide resistance in some cancer cells. We virtually screened the recently crystallized Metnase tertiary structure using a docking program. Thirty novel and two known compounds were identified, which all fell into a novel structural family. Nine of these compounds and the two known drugs were found to restore sensitivity to etoposide in a resistant leukemia line and gemcitabine in pancreatic and lung cancer cell lines, indicating that Metnase may be a target for clinical intervention. We have initiated studies repressing Metnase in breast, lung, and pancreatic cancer cells to examine whether it indeed is the target for enhancement of gemcitabine.
Jobs Summary: Postdoctoral Fellow-Performs colony formation studies of the effect of Metnase inhibitors on etoposide-treated cancer cells Postdoctoral Fellow-Performs immunofluorescence of damaged DNA foci after etoposide treatment Research Associate-Generates mutant Metnase cells lines resistant or sensitive to etoposide, performs RT-PCR and western analysis of Metnase expression. Postdoctoral Fellow-Examines the role of Metnase's natural antagonist JHDM1a for its effect on etoposide DNA damage (Total jobs reported: 0)
Project Status: Less Than 50% Completed
This award's data was last updated on Jun. 2, 2009. Help expand these official descriptions using the wiki below.