R-Loop-Mediated ssDNA Breaks Accumulate Following Short-Term Exposure to the HDAC Inhibitor Romidepsin.
R-Loop-Mediated ssDNA Breaks Accumulate Following Short-Term Exposure to the HDAC Inhibitor Romidepsin.
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DOI：
10.1158/1541-7786.mcr-20-0833
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发表时间：
2021-08
期刊：
影响因子：
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通讯作者：
Bates SE
中科院分区：
文献类型：
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作者：
Safari M;Litman T;Robey RW;Aguilera A;Chakraborty AR;Reinhold WC;Basseville A;Petrukhin L;Scotto L;O'Connor OA;Pommier Y;Fojo AT;Bates SE
组蛋白去乙酰酶抑制剂(HDACis)通过阻断组蛋白去乙酰酶的催化位点来诱导组蛋白的超乙酰化。尽管监管部门批准了血液系统恶性肿瘤的治疗，但实体肿瘤有限的临床活动限制了它们的潜力，这表明需要更好地了解其作用机制。HDAC抑制剂的多种活性已被证明，依赖于细胞环境，超出了典型的基因表达诱导。在这里，使用临床相关的暴露时间，我们利用NCI-60细胞系数据库建立了DNA损伤为主要特征，然后重点研究了过乙酰化导致DNA损伤的机制。我们鉴定了罗米非菌素诱导组蛋白超乙酰化后DNA-RNA杂交体(R-环)的积累，并通过单细胞电泳法检测到单链DNA断裂。我们的数据表明，转录偶联碱基切除修复(BER)参与解决单链DNA断裂，当不堪重负时，演变为致命的双链DNA断裂。我们表明，抑制BER蛋白，如PARP，将增加dsDNA在这一背景下的断裂。这些研究证实，R-环的积累是罗米非菌素介导组蛋白超乙酰化的结果。我们相信，所提供的见解将为设计更有效的HDACis联合疗法治疗实体瘤提供参考。
Histone deacetylase inhibitors (HDACis) induce hyperacetylation of histones by blocking HDAC catalytic sites. Despite regulatory approvals in hematological malignancies, limited solid tumor clinical activity has constrained their potential, arguing for better understanding of mechanisms of action. Multiple activities of HDAC inhibitors have been demonstrated, dependent on cell context, beyond the canonical induction of gene expression. Here, using a clinically relevant exposure duration, we established DNA damage as the dominant signature using the NCI-60 cell line database and then focused on the mechanism by which hyperacetylation induces DNA damage. We identified accumulation of DNA-RNA hybrids (R-loops) following romidepsin-induced histone hyperacetylation, with single-stranded DNA breaks detected by single cell electrophoresis. Our data suggest that transcription-coupled base excision repair (BER) is involved in resolving ssDNA breaks that, when overwhelmed, evolve to lethal dsDNA breaks. We show that inhibition of BER proteins such as PARP will increase dsDNA breaks in this context. These studies establish accumulation of R-loops as a consequence of romidepsin-mediated histone hyperacetylation. We believe that the insights provided will inform design of more effective combination therapy with HDACis for treatment of solid tumors.