两种不同辐射抗性癌症类型中，共同的 Chk1 依赖性 DNA 双链断裂抑制表型。
A common Chk1-dependent phenotype of DNA double-strand break suppression in two distinct radioresistant cancer types.
机构信息
Department of Radiation Oncology, Massachusetts General Hospital, Harvard Medical School, 55 Fruit Street, Boston, MA, 02114, USA.
Laboratory of Radiobiology and Experimental Radiooncology, Clinic of Radiotherapy and Radiooncology, University Medical Center Hamburg-Eppendorf, Hamburg, Germany.
出版信息
Breast Cancer Res Treat. 2019 Apr;174(3):605-613. doi: 10.1007/s10549-018-05079-7. Epub 2019 Jan 3.
PURPOSE
Triple-negative breast cancers (TNBC) are often resistant to treatment with ionizing radiation (IR). We sought to investigate whether pharmacologic inhibition of Chk1 kinase, which is commonly overexpressed in TNBC, preferentially sensitizes TNBC cells to IR.
METHODS
Ten breast cancer cell lines were screened with small molecule inhibitors against Chk1 and other kinases. Chk1 inhibition was also tested in isogenic KRAS mutant or wild-type cancer cells. Cellular radiosensitization was measured by short-term and clonogenic survival assays and by staining for the DNA double-strand break (DSB) marker γ-H2AX. Radiosensitization was also assessed in breast cancer biopsies using an ex vivo assay. Aurora B kinase-dependent mitosis-like chromatin condensation, a marker of radioresistance, was detected using a specific antibody against co-localized phosphorylation of serine 10 and trimethylation of lysine 9 on histone 3 (H3K9me3/S10p). Expression of CHEK1 and associated genes was evaluated in TNBC and lung adenocarcinoma.
RESULTS
Inhibition of Chk1 kinase preferentially radiosensitized TNBC cells in vitro and in patient biopsies. Interestingly, TNBC cells displayed lower numbers of IR-induced DSBs than non-TNBC cells, correlating with their observed radioresistance. We found that Chk1 suppressed IR-induced DSBs in these cells, which was dependent on H3K9me3/S10p-a chromatin mark previously found to indicate radioresistance in KRAS mutant cancers. Accordingly, the effects of Chk1 inhibition in TNBC were reproduced in KRAS mutant but not wild-type cells. We also observed co-expression of genes in this Chk1 chromatin pathway in TNBC and KRAS mutant lung cancers.
CONCLUSIONS
Chk1 promotes an unexpected, common phenotype of chromatin-dependent DSB suppression in radioresistant TNBC and KRAS mutant cancer cells, providing a direction for future investigations into overcoming the treatment resistance of TNBC.
目的
三阴性乳腺癌（TNBC）通常对电离辐射（IR）治疗有抗性。我们试图研究是否抑制在 TNBC 中常过表达的 Chk1 激酶可优先使 TNBC 细胞对 IR 敏感。
方法
用小分子抑制剂筛选了 10 种乳腺癌细胞系，以筛选针对 Chk1 和其他激酶的抑制剂。还在同基因 KRAS 突变或野生型癌细胞中测试了 Chk1 抑制作用。通过短期和克隆存活测定以及针对 DNA 双链断裂（DSB）标记 γ-H2AX 的染色来测量细胞放射增敏性。还使用离体测定法评估了乳腺癌活检中的放射增敏性。通过针对丝氨酸 10 和组蛋白 3（H3）上赖氨酸 9 的三甲基化（H3K9me3/S10p）共定位磷酸化的特异性抗体检测 Aurora B 激酶依赖性有丝分裂样染色质凝聚，这是一种放射抗性的标志物。在 TNBC 和肺腺癌中评估了 CHEK1 和相关基因的表达。
结果
Chk1 激酶的抑制作用在体外和患者活检中优先使 TNBC 细胞对放射敏感。有趣的是，与观察到的放射抗性相关，TNBC 细胞显示出比非 TNBC 细胞更少的 IR 诱导的 DSB。我们发现 Chk1 抑制了这些细胞中的 IR 诱导的 DSB，这依赖于 H3K9me3/S10p-先前在 KRAS 突变型癌症中发现的与放射抗性相关的染色质标记。因此，在 KRAS 突变而非野生型细胞中重现了 Chk1 抑制在 TNBC 中的作用。我们还观察到在 TNBC 和 KRAS 突变型肺癌中，这个 Chk1 染色质途径中的基因表达。
结论
Chk1 促进了在放射抗性 TNBC 和 KRAS 突变型癌细胞中意外的、常见的染色质依赖性 DSB 抑制表型，为未来研究克服 TNBC 的治疗抵抗提供了方向。
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