Single-cell microarray enables high-throughput evaluation of DNA double-strand breaks and DNA repair inhibitors
Single-cell microarray enables high-throughput evaluation of DNA double-strand breaks and DNA repair inhibitors
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DOI：
10.4161/cc.23880
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发表时间：
2013-03-15
期刊：
影响因子：
4.3
通讯作者：
Engelward, Bevin P.
中科院分区：
文献类型：
--
作者：
Weingeist, David M.;Ge, Jing;Engelward, Bevin P.
非手术治疗癌症的一种关键方式是DNA损伤治疗，这种治疗导致DNA双链断裂，对快速分裂的癌细胞具有优先毒性。双链断裂修复能力被认为是耐药的重要机制，因此是辅助化疗的潜在靶点。此外，已知自发和环境诱导的DSB可促进癌症，因此DSB评价作为流行病学、临床评价和新药物开发的重要工具。目前可用的检测双链断裂的方法在通量和特异性上是有限的，并且提供的关于修复动力学的信息很少。在这里，我们展示了CometChip，这是一个96孔的平台，可以并行评估双链断裂水平和多种细胞类型和条件的修复能力，并集成了标准的高通量筛选和分析技术。我们展示了检测双链断裂修复中多种遗传缺陷的能力，并评估了一组临床相关的双链断裂修复途径之一的非同源末端连接的化学抑制剂。当其他高通量修复检测测量残留损伤或间接损伤标记时，CometChip检测物理双链断裂，提供损伤诱导和修复能力的直接测量，这可能有助于开发和实施减少副作用的治疗策略。
A key modality of non-surgical cancer management is DNA damaging therapy that causes DNA double-strand breaks that are preferentially toxic to rapidly dividing cancer cells. Double-strand break repair capacity is recognized as an important mechanism in drug resistance and is therefore a potential target for adjuvant chemotherapy. Additionally, spontaneous and environmentally induced DSBs are known to promote cancer, making DSB evaluation important as a tool in epidemiology, clinical evaluation and in the development of novel pharmaceuticals. Currently available assays to detect double-strand breaks are limited in throughput and specificity and offer minimal information concerning the kinetics of repair. Here, we present the CometChip, a 96-well platform that enables assessment of double-strand break levels and repair capacity of multiple cell types and conditions in parallel and integrates with standard high-throughput screening and analysis technologies. We demonstrate the ability to detect multiple genetic deficiencies in double-strand break repair and evaluate a set of clinically relevant chemical inhibitors of one of the major double-strand break repair pathways, non-homologous end-joining. While other high-throughput repair assays measure residual damage or indirect markers of damage, the CometChip detects physical double-strand breaks, providing direct measurement of damage induction and repair capacity, which may be useful in developing and implementing treatment strategies with reduced side effects.