Hypersensitivity of nonhomologous DNA end-joining mutants to VP-16 and ICRF-193: implications for the repair of topoisomerase II-mediated DNA damage.
A number of clinically useful anticancer drugs, including etoposide (VP-16), target DNA topoisomerase (topo) II. These drugs, referred to as topo II poisons, stabilize cleavable complexes, thereby generating DNA double-strand breaks. Bis-2,6-dioxopiperazines such as ICRF-193 also inhibit topo II by inducing a distinct type of DNA damage, termed topo II clamps, which has been believed to be devoid of double-strand breaks. Despite the biological and clinical importance, the molecular mechanisms for the repair of topo II-mediated DNA damage remain largely unknown. Here, we perform genetic analyses using the chicken DT40 cell line to investigate how DNA lesions caused by topo II inhibitors are repaired. Notably, we show that LIG4-/- and KU70-/- cells, which are defective in nonhomologous DNA end-joining (NHEJ), are extremely sensitive to both VP-16 and ICRF-193. In contrast, RAD54-/- cells (defective in homologous recombination) are much less hypersensitive to VP-16 than the NHEJ mutants and, more importantly, are not hypersensitive to ICRF-193. Our results provide the first evidence that NHEJ is the predominant pathway for the repair of topo II-mediated DNA damage; that is, cleavable complexes and topo II clamps. The outstandingly increased cytotoxicity of topo II inhibitors in the absence of NHEJ suggests that simultaneous inhibition of topo II and NHEJ would provide a powerful protocol in cancer chemotherapy involving topo II inhibitors.
Adachi
Noritaka
N
Kihara Institute for Biological Research, Graduate School of Integrated Science, Yokohama City University, Totsuka-ku, Yokohama 244-0813, Japan. nadachi@yokohama-cu.ac.jp
Suzuki
Hiromi
H
Iiizumi
Susumu
S
Koyama
Hideki
H
D000818:Animals; D000970:Antineoplastic Agents / Q000494:pharmacology; D000972:Antineoplastic Agents, Phytogenic / Q000494:pharmacology; D002460:Cell Line; D002645:Chickens; D004249:DNA Damage / Q000187:drug effects*; D004260:DNA Repair / Q000187:drug effects*; D004250:DNA Topoisomerases, Type II / Q000378:metabolism*; D054659:Diketopiperazines; D004305:Dose-Response Relationship, Drug; D019008:Drug Resistance, Neoplasm; D005047:Etoposide / Q000494:pharmacology*; D005822:Genetic Vectors; D006579:Heterozygote; D008954:Models, Biological; D008957:Models, Genetic; D009154:Mutation; D010879:Piperazines / Q000494:pharmacology*; D011995:Recombination, Genetic / Q000187:drug effects*; D014162:Transfection; D019076:Transgenes
D000970:Antineoplastic Agents; D000972:Antineoplastic Agents, Phytogenic; D054659:Diketopiperazines; D010879:Piperazines; C070899:4,4'-(1,2-dimethyl-1,2-ethanediyl)bis-2,6-piperazinedione; D005047:Etoposide; D004250:DNA Topoisomerases, Type II