53BP1 depletion causes PARP inhibitor resistance in ATM-deficient breast cancer cells.

Ruoxi Hong,Yang Luo,Fei Ma,Changjun Zhu,Binghe Xu,Weimin Zhang,Qing Li,Xiying Yu,Wei Jiang

doi:10.1186/s12885-016-2754-7

Abstract

BackgroundMutations in DNA damage response factors BRCA1 and BRCA2 confer sensitivity to poly(ADP-ribose) polymerase (PARP) inhibitors in breast and ovarian cancers. BRCA1/BRCA2-defective tumors can exhibit resistance to PARP inhibitors via multiple mechanisms, one of which involves loss of 53BP1. Deficiency in the DNA damage response factor ataxia-telangiectasia mutated (ATM) can also sensitize tumors to PARP inhibitors, raising the question of whether the presence or absence of 53BP1 can predict sensitivity of ATM-deficient breast cancer to these inhibitors.MethodsCytotoxicity of PARP inhibitor and ATM inhibitor in breast cancer cell lines was assessed by MTS, colony formation and apoptosis assays. ShRNA lentiviral vectors were used to knockdown 53BP1 expression in breast cancer cell lines. Phospho-ATM and 53BP1 protein expressions were determined in human breast cancer tissues by immunohistochemistry (IHC).ResultsWe show that inhibiting ATM increased cytotoxicity of PARP inhibitor in triple-negative and non-triple-negative breast cancer cell lines, and depleting the cells of 53BP1 reduced this cytotoxicity. Inhibiting ATM abrogated homologous recombination induced by PARP inhibitor, and down-regulating 53BP1 partially reversed this effect. Further, overall survival was significantly better in triple-negative breast cancer patients with lower levels of phospho-ATM and tended to be better in patients with negative 53BP1.ConclusionThese results suggest that 53BP1 may be a predictor of PARP inhibitor resistance in patients with ATM-deficient tumors.Electronic supplementary materialThe online version of this article (doi:10.1186/s12885-016-2754-7) contains supplementary material, which is available to authorized users.

Highlights

Mutations in DNA damage response factors BRCA1 and BRCA2 confer sensitivity to poly(ADP-ribose) polymerase (PARP) inhibitors in breast and ovarian cancers
ataxia-telangiectasia mutated (ATM) inhibition sensitizes breast cancer cells to Olaparib To evaluate whether ATM inhibition increases the cytotoxicity of PARP inhibitors, we treated both triplenegative breast cancer cell lines (CAL-51, MDA-MB-231 and MDA-MB-468) and non-triple-negative breast cancer cell lines (MCF-7, T-47D and SK-BR-3) with increasing concentrations of Olaparib (0–10 μM) alone or in combination with 10 μM ATM inhibitor KU55933
Olaparib inhibited colony formation in CAL-51 and MCF-7 cells in a dose-dependent manner, which KU55933 further enhanced it (Fig. 1d-e). These data suggest that Olaparib suppresses cell proliferation and colony formation in breast cancer cell lines, and that KU55933 reinforces this effect through synthetic lethality

Summary

Introduction

Mutations in DNA damage response factors BRCA1 and BRCA2 confer sensitivity to poly(ADP-ribose) polymerase (PARP) inhibitors in breast and ovarian cancers. Cells with defective BRCA1 or BRCA2 are unable to perform HR, so alternative repair processes kick in, such as nonhomologous DNA end-joining (NHEJ) These alternative processes sometimes fail to repair DSBs, leading to genome instability and cytotoxicity. PARP inhibitor therapy is based on synthetic lethality: it targets two separate molecular pathways that are non-lethal when disrupted independently, but are lethal when inhibited simultaneously [8] Both BRCA1 and BRCA2 function in the homologous recombination (HR) pathway to repair of doublestranded DNA breaks (DSBs), while PARP-1 is a key mediator in the base excision repair (BER) pathway to repair single-stranded DNA breaks (SSBs) [9, 10]

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Conclusion