Abstract A92: The role of FOXM1 and NBS1 in DNA damage-induced senescence and epirubicin resistance.

Abstract

Background: The Forkhead box M1 (FOXM1) transcription factor is responsible for the regulation of a wide range of biological processes, including cell cycle, cell proliferation, apoptosis and tumorigenesis. Deregulation of FOXM1 has been suggested to be associated with drug resistance and poor prognosis in breast cancer patients and this could occur through an enhancement of DNA damage repair as well as a reduced propensity to undergo cell death or senescence. However, the exact mechanisms of how FOXM1 modulates drug resistance still remain unclear. The aim of this study is to investigate the new roles of FOXM1 and its downstream targets in DNA damage response and cellular senescence, which are vital for both the tumor progression and the development of chemotherapeutic drug resistance. Materials and Methods: MCF-7 human breast carcinoma cell lines and mouse embryonic fibroblasts (MEFs) were exposed to epirubicin and long-term cell viability was assessed by clonogenic assay. DNA double strand break repair was examined by γH2AX foci and DR-GFP repair assay. Analysis of cellular senescence was based on cellular β-galactosidase activity and changes in cell morphology. Expression levels of key DNA repair genes were assessed by using quantitative real-time PCR and western blot analysis. Results: We previously found that FOXM1 is required for DNA repair via homologous recombination repair. Consistently, reconstitution of FOXM1 in FOXM1-deficient MEFs decreased the accumulation of senescence-associated γH2AX foci after epirubicin treatment. Here, we shows that FOXM1-depletion can sensitize both human breast cancer MCF-7 cells and MEFs into entering epirubicin-induced senescence as indicated by the loss of long-term cell proliferation ability, the accumulation of persistent γH2AX foci, and the induction of senescence-associated β-galactosidase activity. Interestingly, we also demonstrate that FOXM1 regulates NBS1, a key component for the MRN complex-mediated ATM activation in response to DNA double stranded break (DSBs), at the transcriptional level. Using HeLa cell lines harbouring the DR-GFP reporter, overexpression of FOXM1 failed to augment HR activity in the background of NBS1 depletion, suggesting that NBS1 is indispensable for the HR function of FOXM1. Similar to FOXM1, NBS1 is strongly up-regulated in the epirubicin-resistant MCF-7 (MCF-7 EpiR) cells compared with parental MCF-7 cells and the depletion of FOXM1 or NBS1 also renders MCF-7 and MCF-7 EpiR cells more sensitive to epirubicin-induced cellular senescence. Conclusion: Taken together, these results indicate that NBS1 is a key FOXM1 downstream target gene involved in DNA damage response, DNA damage-induced senescence and epirubicin resistance. The FOXM1-NBS1 axis can be a reliable prognostic marker and a potential therapeutic target for overcoming genotoxic agent resistance in cancer. Citation Information: Mol Cancer Ther 2013;12(11 Suppl):A92. Citation Format: Pasarat Khongkow, Upekha Karunarathna, Eric Lam. The role of FOXM1 and NBS1 in DNA damage-induced senescence and epirubicin resistance. [abstract]. In: Proceedings of the AACR-NCI-EORTC International Conference: Molecular Targets and Cancer Therapeutics; 2013 Oct 19-23; Boston, MA. Philadelphia (PA): AACR; Mol Cancer Ther 2013;12(11 Suppl):Abstract nr A92.