title: Single-cell microarray enables high-throughput evaluation of DNA double-strand breaks and DNA repair inhibitors.
abstract: 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.
authors: Department of Biological Engineering, Massachusetts Institute of Technology, Cambridge, MA, USA.
authors: David M
authors: DM
authors: Weingeist
authors: Jing
authors: J
authors: Ge
authors: David K
authors: DK
authors: Wood
authors: James T
authors: JT
authors: Mutamba
authors: Qiuying
authors: Q
authors: Huang
authors: Elizabeth A
authors: EA
authors: Rowland
authors: Michael B
authors: MB
authors: Yaffe
authors: Scott
authors: S
authors: Floyd
authors: Bevin P
authors: BP
authors: Engelward
mesh_terms: D000818:Animals; D016466:CHO Cells; D002460:Cell Line; D002867:Chromones / Q000494:pharmacology; D006224:Cricetinae; D053903:DNA Breaks, Double-Stranded / Q000187:drug effects*; D004249:DNA Damage; D004260:DNA Repair / Q000187:drug effects*; D051747:DNA-Activated Protein Kinase / Q000037:antagonists & inhibitors; D019008:Drug Resistance, Neoplasm; D004791:Enzyme Inhibitors / Q000494:pharmacology; D057166:High-Throughput Screening Assays / Q000379:methods*; D006801:Humans; D009025:Morpholines / Q000494:pharmacology; D009369:Neoplasms / Q000188:drug therapy* / Q000235:genetics
chemical_list: C479235:2-(morpholin-4-yl)benzo(h)chromen-4-one; C499693:8-dibenzothiophen-4-yl-2-morpholin-4-yl-chromen-4-one; D002867:Chromones; D004791:Enzyme Inhibitors; D009025:Morpholines; C085911:2-(4-morpholinyl)-8-phenyl-4H-1-benzopyran-4-one; D051747:DNA-Activated Protein Kinase
keywords: DNA double-strand breaks; DNA repair; DNA-PK inhibitors; high throughput; microarray; neutral comet assay; neutral single-cell electrophoresis assay; non-homologous end-joining