Novel Role of Base Excision Repair in Mediating Cisplatin Cytotoxicity

Anbarasi Kothandapani,Venkata Srinivas Mohan Nimai Dangeti,Ashley R Brown,Lauren A Banze,Xiao-Hong Wang,Robert W Sobol,Steve M Patrick

doi:10.1074/jbc.m111.225375

Abstract

Using isogenic mouse embryonic fibroblasts and human cancer cell lines, we show that cells defective in base excision repair (BER) display a cisplatin-specific resistant phenotype. This was accompanied by enhanced repair of cisplatin interstrand cross-links (ICLs) and ICL-induced DNA double strand breaks, but not intrastrand adducts. Cisplatin induces abasic sites with a reduced accumulation in uracil DNA glycosylase (UNG) null cells. We show that cytosines that flank the cisplatin ICLs undergo preferential oxidative deamination in vitro, and AP endonuclease 1 (APE1) can cleave the resulting ICL DNA substrate following removal of the flanking uracil. We also show that DNA polymerase β has low fidelity at the cisplatin ICL site after APE1 incision. Down-regulating ERCC1-XPF in BER-deficient cells restored cisplatin sensitivity. Based on our results, we propose a novel model in which BER plays a positive role in maintaining cisplatin cytotoxicity by competing with the productive cisplatin ICL DNA repair pathways.

Highlights

Intrastrand adducts is primarily via the nucleotide excision repair (NER) pathway, which is a versatile pathway for the removal of a variety of bulky lesions, whereas repair of the cisplatin interstrand crosslinks (ICLs) is less understood [3, 4]
Based on the recent studies targeting base excision repair (BER) proteins to sensitize cells to antitumor agents and due to the existing conflicting observations [11,12,13], we evaluated the effect of defective BER on cisplatin cytotoxicity using isogenic mouse embryonic fibroblasts and human cancer cells (MDA-MB-231, HeLaS3 and A2780)
We report that defective BER does not affect the processing of cisplatin intrastrand adducts but is involved in modulating cisplatin ICL DNA repair

Summary

Introduction

Intrastrand adducts is primarily via the nucleotide excision repair (NER) pathway, which is a versatile pathway for the removal of a variety of bulky lesions, whereas repair of the cisplatin ICLs is less understood [3, 4]. Utilizing synthetic oligonucleotides containing a site-specific cisplatin ICL, we demonstrate that preferential deamination occurs on the flipped out cytosine bases adjacent to the cross-linked guanines when compared with undamaged and intrastrand adduct containing DNA substrates. To assess whether the resistant phenotype in BER-deficient cells is general to DNA damage or is cisplatin-specific, we utilized transplatin, a trans isomer of cisplatin, mitomycin C, which induces ICLs, and an intrastrand-specific agent, UV light (supplemental Fig. S2).

Results

Conclusion