DESCRIPTION (provided by applicant): The long term objective of our laboratory is to determine the mechanism by which the novel c- Myc-interacting tumor suppressor Bin1 renders chemoresistant cancer cells sensitive to DNA- damaging agents such as cisplatin. This project is innovative because it focuses on the 'c-Myc- Bin1-PARP-1' chemoresistant axis, which provides a new and substantially different way of addressing the problem of cisplatin-resistant cancers. We propose to test the hypothesis that the sensitivity of human cancer cells to cisplatin is dependent on the expression of Bin1, a protein that inhibits c-Myc-induced poly(ADP-ribose) polymerase-1 (or PARP-1) activity. Results from our laboratory support our assertion that Bin1 interacts with and inhibits PARP-1, a key nuclear enzyme required for promoting DNA repair. Subsequently, Bin1 decreases chromosome stability and sensitizes cancer cells to cisplatin. Moreover, elevated c-Myc directly represses Bin1 promoter, which subsequently rescues PARP-1 activity and increases cisplatin resistance. We aim to 1) determine the mechanism(s) by which Bin1 suppresses PARP-1-mediated DNA repair, and 2) identify the mechanism(s) by which oncogenic c-Myc alters Bin1 and PARP-1 levels. To achieve the first aim, we will investigate (i) whether the interactions between the automodification domain of PARP-1 and Bin1 are direct and (ii) how Bin1 disrupts the PARP-1-XRCC1 interaction in vivo, an essential step for PARP-1-mediated base excision repair (BER). Given that Bin1 suppresses PARP-1 auto-poly(ADP-ribosyl)ation, we predict that Bin1 inhibits PARP-1-associated XRCC1 recruitment and the subsequent BER pathways. To identify the mechanism by which elevated c- Myc alters Bin1 and PARP-1 expression, we will determine how c-Myc interferes with the transcriptional activities of Miz-1 and E2F1, two transcription factors that separately activate the Bin1 promoter, and conversely, how c-Myc upregulates PARP-1 expression via Brahma-related gene protein 1 (BRG1), a key component of the SWI/SNF chromatin remodeling complex. Cisplatin-induced cell death assays in the presence and absence of small interfering RNAs or chemical inhibitors of Bin1, PARP-1, and c-Myc will determine the manner by which Bin1 disrupts the 'c-Myc-PARP-1' chemoresistant axis. Successful completion of this project will define Bin1-mediated chemosensitizing mechanisms that attenuate PARP-1-mediated DNA repair. This knowledge will be significant, as it may guide the development of therapeutic strategies for the treatment of cisplatin-resistant human cancers that overexpress c-Myc.
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