title: Downregulation of XPF-ERCC1 enhances cisplatin efficacy in cancer cells.
abstract: Bulky cisplatin lesions are repaired primarily by nucleotide excision repair (NER), in which the structure specific endonuclease XPF-ERCC1 is a critical component. It is now known that the XPF-ERCC1 complex has repair functions beyond NER and plays a role in homologous recombination (HR). It has been suggested that expression of ERCC1 correlates with cisplatin drug resistance in non-small cell lung cancer (NSCLC). In our study, using NSCLC, ovarian, and breast cancer cells, we show that the XPF-ERCC1 complex is a valid target to increase cisplatin cytotoxicity and efficacy. We targeted XPF-ERCC1 complex by RNA interference and assessed the repair capacity of cisplatin intrastrand and interstrand crosslinks by ELISA and alkaline comet assay, respectively. We also assessed the repair of cisplatin-ICL-induced double-strand breaks (DSBs) by monitoring gamma-H2AX focus formation. Interestingly, XPF protein levels were significantly reduced following ERCC1 downregulation, but the converse was not observed. The transcript levels were unaffected suggesting that XPF protein stability is likely affected. The repair of both types of cisplatin-DNA lesions was decreased with downregulation of XPF, ERCC1 or both XPF-ERCC1. The ICL-induced DSBs persist in the absence of XPF-ERCC1. The suppression of the XPF-ERCC1 complex significantly decreases the cellular viability which correlates well with the decrease in DNA repair capacity. A double knockdown of XPF-ERCC1 displays the greatest level of cellular cytotoxicity when compared with XPF or ERCC1 alone. The difference in cytotoxicity observed is likely due to the level of total protein complex remaining. These data demonstrate that XPF-ERCC1 is a valid target to enhance cisplatin efficacy in cancer cells by affecting cisplatin-DNA repair pathways.
authors.affiliation: Department of Biochemistry & Cancer Biology, University of Toledo Health Science Campus, OH, United States.
authors.forename: Sanjeevani
authors.initials: S
authors.lastname: Arora
authors.forename: Anbarasi
authors.initials: A
authors.lastname: Kothandapani
authors.forename: Kristin
authors.initials: K
authors.lastname: Tillison
authors.forename: Vivian
authors.initials: V
authors.lastname: Kalman-Maltese
authors.forename: Steve M
authors.initials: SM
authors.lastname: Patrick
mesh_terms: D000970:Antineoplastic Agents / Q000494:pharmacology*; D045744:Cell Line, Tumor; D002945:Cisplatin / Q000494:pharmacology*; D004249:DNA Damage; D004260:DNA Repair / Q000235:genetics; D004268:DNA-Binding Proteins / Q000037:antagonists & inhibitors* / Q000235:genetics; D015536:Down-Regulation; D019008:Drug Resistance, Neoplasm; D004720:Endonucleases / Q000037:antagonists & inhibitors* / Q000235:genetics; D055785:Gene Knockdown Techniques; D006657:Histones / Q000378:metabolism; D006801:Humans; D009369:Neoplasms / Q000378:metabolism*; D034741:RNA, Small Interfering / Q000235:genetics
chemical_list: D000970:Antineoplastic Agents; D004268:DNA-Binding Proteins; C468783:H2AX protein, human; D006657:Histones; D034741:RNA, Small Interfering; C095888:xeroderma pigmentosum group F protein; C071447:ERCC1 protein, human; D004720:Endonucleases; D002945:Cisplatin