Princess Margaret Hospital (University Health Network) and Departments of Medical Biophysics and Radiation Oncology, University of Toronto, Toronto, Ontario, Canada.
Cancer Res. 2010 Oct 15;70(20):8045-54. doi: 10.1158/0008-5472.CAN-10-2352. Epub 2010 Oct 5.
Acute and chronic hypoxia exists within the three-dimensional microenvironment of solid tumors and drives therapy resistance, genetic instability, and metastasis. Replicating cells exposed to either severe acute hypoxia (16 hours with 0.02% O(2)) followed by reoxygenation or moderate chronic hypoxia (72 hours with 0.2% O(2)) treatments have decreased homologous recombination (HR) protein expression and function. As HR defects are synthetically lethal with poly(ADP-ribose) polymerase 1 (PARP1) inhibition, we evaluated the sensitivity of repair-defective hypoxic cells to PARP inhibition. Although PARP inhibition itself did not affect HR expression or function, we observed increased clonogenic killing in HR-deficient hypoxic cells following chemical inhibition of PARP1. This effect was partially reversible by RAD51 overexpression. PARP1(-/-) murine embryonic fibroblasts (MEF) showed a proliferative disadvantage under hypoxic gassing when compared with PARP1(+/+) MEFs. PARP-inhibited hypoxic cells accumulated γH2AX and 53BP1 foci as a consequence of altered DNA replication firing during S phase-specific cell killing. In support of this proposed mode of action, PARP inhibitor-treated xenografts displayed increased γH2AX and cleaved caspase-3 expression in RAD51-deficient hypoxic subregions in vivo, which was associated with decreased ex vivo clonogenic survival following experimental radiotherapy. This is the first report of selective cell killing of HR-defective hypoxic cells in vivo as a consequence of microenvironment-mediated "contextual synthetic lethality." As all solid tumors contain aggressive hypoxic cells, this may broaden the clinical utility of PARP and DNA repair inhibition, either alone or in combination with radiotherapy and chemotherapy, even in tumor cells lacking synthetically lethal, genetic mutations.
实体瘤的三维微环境中存在急性和慢性缺氧，导致治疗耐药性、遗传不稳定性和转移。暴露于严重急性缺氧（16 小时，氧浓度为 0.02%）后再复氧或中度慢性缺氧（72 小时，氧浓度为 0.2%）处理的复制细胞，同源重组（HR）蛋白表达和功能下降。由于 HR 缺陷与聚（ADP-核糖）聚合酶 1（PARP1）抑制具有合成致死性，我们评估了修复缺陷性缺氧细胞对 PARP 抑制的敏感性。尽管 PARP 抑制本身不会影响 HR 的表达或功能，但我们观察到 HR 缺陷性缺氧细胞在 PARP1 化学抑制后对克隆杀伤的敏感性增加。RAD51 过表达部分逆转了这种效应。与 PARP1(+/+) MEFs 相比，PARP1(-/-) 小鼠胚胎成纤维细胞（MEF）在缺氧通气时表现出生长劣势。PARP 抑制的缺氧细胞在 S 期特异性细胞杀伤过程中由于 DNA 复制起始的改变而积累 γH2AX 和 53BP1 焦点。为了支持这种拟议的作用模式，PARP 抑制剂处理的异种移植物在体内 RAD51 缺陷性缺氧亚区中显示出增加的 γH2AX 和裂解的 caspase-3 表达，这与实验性放射治疗后体外克隆生存能力降低有关。这是首例报道的体内 HR 缺陷性缺氧细胞由于微环境介导的“上下文合成致死性”而发生的选择性细胞杀伤。由于所有实体瘤都含有侵袭性缺氧细胞，这可能扩大 PARP 和 DNA 修复抑制的临床应用，无论是单独使用还是与放射治疗和化疗联合使用，即使在缺乏合成致死性、遗传突变的肿瘤细胞中也是如此。
