title: Cytotoxicity of Tirapazamine (3-Amino-1,2,4-benzotriazine-1,4-dioxide)-Induced DNA Damage in Chicken DT40 Cells.
abstract: Tirapazamine (TPZ) is an anticancer drug with highly selective cytotoxicity toward hypoxic cells. TPZ is converted to a radical intermediate under hypoxic conditions, and this intermediate interacts with intracellular macromolecules, including DNA. TPZ has been reported to indirectly induce DNA double-strand breaks (DSBs) through the formation of various intermediate DNA lesions under hypoxic conditions. Although the topoisomerase II-DNA complex has been identified as one of these intermediates, other lesions have not yet been defined. In order to obtain a deeper understanding of the mechanisms responsible for the selective cytotoxicity of TPZ toward hypoxic cells, its cellular sensitivity was systematically examined with genetically isogenic DNA-repair-deficient mutant DT40 cell lines. Our results showed that tdp1-/-, tdp2-/-, parp1-/-, and aptx1-/-cells displayed hypersensitivity to TPZ only under hypoxic conditions. These results strongly suggest that the accumulation of the topoisomerase I-trapped DNA complex, topoisomerase II-trapped DNA complex, and abortive ligation products with 5'-AMP are the potential causes of TPZ-induced hypoxic cell death. Furthermore, our genetic analysis revealed that under normoxic conditions (as well as hypoxic conditions), TPZ exhibited significant cytotoxicity toward cell lines deficient in homologous recombination, nonhomologous end joining, base excision repair, and translesion synthesis. Ascorbic acid, a radical scavenger, suppressed TPZ-induced cytotoxicity toward normoxic cells. These results suggest the involvement of oxidative DNA damage and DSBs produced by reactive oxygen species generated from superoxide, a byproduct of the oxidation of TPZ radical intermediates in normoxic cells. Collectively, our results demonstrate that TPZ induces oxidative DNA damage under normoxic and hypoxic conditions and selectively introduces abortive topoisomerase-DNA complexes and unligatable DNA ends under hypoxic conditions.
authors.affiliation: Division of Radiation Life Science, Research Reactor Institute, Kyoto University , 2 Asashiro-Nishi, Kumatori-cho, Sennan-gun, Osaka 590-0494, Japan.
authors.forename: Takahito
authors.initials: T
authors.lastname: Moriwaki
authors.affiliation: Division of Radiation Life Science, Research Reactor Institute, Kyoto University , 2 Asashiro-Nishi, Kumatori-cho, Sennan-gun, Osaka 590-0494, Japan.
authors.forename: Saki
authors.initials: S
authors.lastname: Okamoto
authors.affiliation: Department of Radiation Genetics, Graduate School of Medicine, Kyoto University , Yoshida-Konoe-cho, Sakyo-ku, Kyoto 606-8501, Japan.
authors.forename: Hiroyuki
authors.identifier: 0000-0003-2824-4499
authors.initials: H
authors.lastname: Sasanuma
authors.affiliation: Laboratory of Pharmaceutical and Medicinal Chemistry, Gifu Pharmaceutical University , 1-25-4 Daigaku-nishi, Gifu-shi, Gifu 501-1196, Japan.
authors.forename: Hideko
authors.initials: H
authors.lastname: Nagasawa
authors.affiliation: Department of Radiation Genetics, Graduate School of Medicine, Kyoto University , Yoshida-Konoe-cho, Sakyo-ku, Kyoto 606-8501, Japan.
authors.forename: Shunichi
authors.initials: S
authors.lastname: Takeda
authors.affiliation: Division of Radiation Life Science, Research Reactor Institute, Kyoto University , 2 Asashiro-Nishi, Kumatori-cho, Sennan-gun, Osaka 590-0494, Japan.
authors.forename: Shin-Ichiro
authors.initials: SI
authors.lastname: Masunaga
authors.affiliation: Division of Radiation Life Science, Research Reactor Institute, Kyoto University , 2 Asashiro-Nishi, Kumatori-cho, Sennan-gun, Osaka 590-0494, Japan.
authors.forename: Keizo
authors.identifier: 0000-0003-3900-0035
authors.initials: K
authors.lastname: Tano
mesh_terms: D000818:Animals; D000970:Antineoplastic Agents / Q000633:toxicity*; D002460:Cell Line; D002645:Chickens; D020552:Comet Assay; D004247:DNA / Q000187:drug effects*; D004249:DNA Damage; D017382:Reactive Oxygen Species / Q000378:metabolism; D000077704:Tirapazamine; D014227:Triazines / Q000633:toxicity*
chemical_list: D000970:Antineoplastic Agents; D017382:Reactive Oxygen Species; D014227:Triazines; D000077704:Tirapazamine; D004247:DNA