The Lunenfeld-Tanenbaum Research Institute, Mount Sinai Hospital, Toronto, Ontario, Canada.
MRC Human Genetics Unit, Institute of Genetics and Molecular Medicine, University of Edinburgh, Edinburgh, UK.
Nature. 2018 Jul;559(7713):285-289. doi: 10.1038/s41586-018-0291-z. Epub 2018 Jul 4.
The observation that BRCA1- and BRCA2-deficient cells are sensitive to inhibitors of poly(ADP-ribose) polymerase (PARP) has spurred the development of cancer therapies that use these inhibitors to target deficiencies in homologous recombination. The cytotoxicity of PARP inhibitors depends on PARP trapping, the formation of non-covalent protein-DNA adducts composed of inhibited PARP1 bound to DNA lesions of unclear origins. To address the nature of such lesions and the cellular consequences of PARP trapping, we undertook three CRISPR (clustered regularly interspersed palindromic repeats) screens to identify genes and pathways that mediate cellular resistance to olaparib, a clinically approved PARP inhibitor. Here we present a high-confidence set of 73 genes, which when mutated cause increased sensitivity to PARP inhibitors. In addition to an expected enrichment for genes related to homologous recombination, we discovered that mutations in all three genes encoding ribonuclease H2 sensitized cells to PARP inhibition. We establish that the underlying cause of the PARP-inhibitor hypersensitivity of cells deficient in ribonuclease H2 is impaired ribonucleotide excision repair. Embedded ribonucleotides, which are abundant in the genome of cells deficient in ribonucleotide excision repair, are substrates for cleavage by topoisomerase 1, resulting in PARP-trapping lesions that impede DNA replication and endanger genome integrity. We conclude that genomic ribonucleotides are a hitherto unappreciated source of PARP-trapping DNA lesions, and that the frequent deletion of RNASEH2B in metastatic prostate cancer and chronic lymphocytic leukaemia could provide an opportunity to exploit these findings therapeutically.
观察到 BRCA1 和 BRCA2 缺陷细胞对聚（ADP-核糖）聚合酶（PARP）抑制剂敏感，这促使开发了使用这些抑制剂针对同源重组缺陷的癌症治疗方法。PARP 抑制剂的细胞毒性取决于 PARP 捕获，即由结合 DNA 损伤的受抑制 PARP1 形成的非共价蛋白-DNA 加合物的形成，这些 DNA 损伤的来源尚不清楚。为了解决这些损伤的性质以及 PARP 捕获的细胞后果，我们进行了三次 CRISPR（成簇规律间隔短回文重复）筛选，以鉴定介导细胞对奥拉帕利（一种临床批准的 PARP 抑制剂）耐药的基因和途径。在这里，我们提出了一个高可信度的 73 个基因集，当这些基因发生突变时，会导致对 PARP 抑制剂的敏感性增加。除了同源重组相关基因的预期富集外，我们还发现编码核糖核酸酶 H2 的所有三个基因的突变都使细胞对 PARP 抑制敏感。我们确定了核糖核酸酶 H2 缺陷细胞对 PARP 抑制剂敏感性增加的根本原因是核糖核苷酸切除修复受损。在核糖核苷酸切除修复缺陷的细胞的基因组中大量存在的嵌入核糖核苷酸是拓扑异构酶 1 切割的底物，导致 PARP 捕获损伤，从而阻碍 DNA 复制并危及基因组完整性。我们得出结论，基因组核糖核苷酸是 PARP 捕获 DNA 损伤的一个迄今为止尚未被认识到的来源，并且在转移性前列腺癌和慢性淋巴细胞白血病中频繁缺失 RNASEH2B 可能为利用这些发现进行治疗提供机会。
