Homologous recombination is the principal pathway for the repair of DNA damage induced by tirapazamine in mammalian cells.
Tirapazamine (3-amino-1,2,4-benzotriazine-1,4-dioxide) is a promising hypoxia-selective cytotoxin that has shown significant activity in advanced clinical trials in combination with radiotherapy and cisplatin. The current study aimed to advance our understanding of tirapazamine-induced lesions and the pathways involved in their repair. We show that homologous recombination plays a critical role in repair of tirapazamine-induced damage because cells defective in homologous recombination proteins XRCC2, XRCC3, Rad51D, BRCA1, or BRCA2 are particularly sensitive to tirapazamine. Consistent with the involvement of homologous recombination repair, we observed extensive sister chromatid exchanges after treatment with tirapazamine. We also show that the nonhomologous end-joining pathway, which predominantly deals with frank double-strand breaks (DSB), is not involved in the repair of tirapazamine-induced DSBs. In addition, we show that tirapazamine preferentially kills mutants both with defects in XPF/ERCC1 (but not in other nucleotide excision repair factors) and with defects in base excision repair. Tirapazamine also induces DNA-protein cross-links, which include stable DNA-topoisomerase I cleavable complexes. We further show that gamma H2AX, an indicator of DNA DSBs, is induced preferentially in cells in the S phase of the cell cycle. These observations lead us to an overall model of tirapazamine damage in which DNA single-strand breaks, base damage, and DNA-protein cross-links (including topoisomerase I and II cleavable complexes) produce stalling and collapse of replication forks, the resolution of which results in DSB intermediates, requiring homologous recombination and XPF/ERCC1 for their repair.
Evans
James W
JW
Department of Radiation Oncology, Division of Radiation and Cancer Biology, Stanford University, Stanford, California 94305-5152, USA.
Chernikova
Sophia B
SB
Kachnic
Lisa A
LA
Banath
Judit P
JP
Sordet
Olivier
O
Delahoussaye
Yvette M
YM
Treszezamsky
Alejandro
A
Chon
Brian H
BH
Feng
Zhihui
Z
Gu
Yongchuan
Y
Wilson
William R
WR
Pommier
Yves
Y
Olive
Peggy L
PL
Powell
Simon N
SN
Brown
J Martin
JM
D000818:Animals; D000970:Antineoplastic Agents / Q000494:pharmacology*; D045744:Cell Line, Tumor; D006224:Cricetinae; D004247:DNA / Q000187:drug effects*; D053903:DNA Breaks, Double-Stranded; D004249:DNA Damage; D004260:DNA Repair / Q000235:genetics*; D004268:DNA-Binding Proteins / Q000235:genetics / Q000502:physiology; D019008:Drug Resistance, Neoplasm / Q000235:genetics*; D004720:Endonucleases / Q000235:genetics / Q000502:physiology; D006657:Histones / Q000378:metabolism; D006801:Humans; D009154:Mutation; D010766:Phosphorylation; D011995:Recombination, Genetic / Q000235:genetics*; D000077704:Tirapazamine; D014227:Triazines / Q000494:pharmacology*
D000970:Antineoplastic Agents; D004268:DNA-Binding Proteins; C468783:H2AX protein, human; D006657:Histones; D014227:Triazines; D000077704:Tirapazamine; D004247:DNA; D004720:Endonucleases