Targeting the Sam68-stimulated PARP1 activation for cancer treatment

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Project Summary The inhibition of poly(ADP-ribose) polymerase 1 (PARP1) protein has emerged as a promising therapeutic treatment for several types of cancers. However, the current classes of PARP1 inhibitors are all derivatives of nicotinamide, based on the strategy of competitive binding against nicotinamide. Because the PARP1-catalyzed PARylation is critical for many cellular processes, therapeutic targeting of PARP1 in cancerous cells with current classes of PARP1 inhibitors could negatively affect many cell functions in normal cells. There is an urgent clinical need for improving the inhibitor specificity for PARP1 and lowering the off- target effects and toxicity for better therapeutic value and modalities. However, the rapid recruitment of PARP1 to sites of DNA lesions (within milliseconds) and the vigorously synthesized PAR chains make it difficult to elucidate the precise stimulatory and regulatory mechanisms on PARP1 activation. We recently discovered that Src-associated-substrate-during-mitosis-of-68kDa (Sam68) plays an important role in stimulating the DNA damage-specific PARP1 activation, which suggests that targeting the Sam68-stimulated PARP1 activation could be a novel strategy to develop a new category of PARP1 inhibitors. The objective of our research is to identify specific molecule inhibitors of the Sam68-stimulated PARP1 activation via a high throughput screening, and our hypothesis is that these inhibitors would alleviate DNA damage-triggered PARP1 activation thus being potential pharmacological agents for treating cancers. We have developed a novel ELISA-based assay suitable for screening of small molecule libraries. In this proposed research, we will conduct high throughput screening to identify small molecule compounds inhibiting the Sam68-stimulated PARP1 activation. We will employ interdisciplinary approach combining biochemical, biophysical, and molecular experiments, cellular assays, and imaging methods to validate the screening hits and to identify high quality small molecule inhibitors of Sam68-PARP1 interaction. Activity of the most potent compounds will be evaluated in a panel of cell-based and mouse model experiments to assess their capability to inhibit the survival and development of colon cancer cells. Our project represents a very innovative strategy to inhibit PARP1 activity through blocking the Sam68-conferred stimulatory mechanism. At the conclusion of these studies, we expect to identify highly valuable compounds that can serve as chemical probes suitable for further development into potent PARP1 inhibitors for cancer therapeutics. Additionally, discovery of these inhibitors will greatly facilitate our research on the regulation and function of the Sam68-stimulated PARP1 activation under normal and disease conditions.