Abstract
BackgroundWhile platinum-based chemotherapeutic agents are widely used to treat various solid tumors, the acquired platinum resistance is a major impediment in their successful treatment. Since enhanced DNA repair capacity is a major factor in conferring cisplatin resistance, targeting of DNA repair pathways is an effective stratagem for overcoming cisplatin resistance. This study was designed to delineate the role of nucleotide excision repair (NER), the principal mechanism for the removal of cisplatin-induced DNA intrastrand crosslinks, in cisplatin resistance and reveal the impact of DNA repair interference on cisplatin sensitivity in human ovarian cancer cells.ResultsWe assessed the inherent NER efficiency of multiple matched pairs of cisplatin-sensitive and -resistant ovarian cancer cell lines and their expression of NER-related factors at mRNA and protein levels. Our results showed that only the cisplatin-resistant ovarian cancer cell line PEO4 possessed an increased NER capacity compared to its inherently NER-inefficient parental line PEO1. Several other cisplatin-resistant cell lines, including CP70, CDDP and 2008C13, exhibited a normal and parental cell-comparable NER capacity for removing cisplatin-induced DNA intrastrand cross-links (Pt-GG). Concomitant gene expression analysis revealed discordance in mRNA and protein levels of NER factors in various ovarian cancer cell lines and NER proteins level were unrelated to the cisplatin sensitivity of these cell lines. Although knockdown of NER factors was able to compromise the NER efficiency, it only caused a minimal effect on cisplatin sensitivity. On the contrary, downregulation of BRCA2, a critical protein for homologous recombination repair (HRR), significantly enhanced the efficacy of cisplatin in killing ovarian cancer cell line PEO4.ConclusionOur studies indicate that the level of NER factors in ovarian cancer cell lines is neither a determinant of their NER capacity nor of the sensitivity to cisplatin, and suggest that manipulation of the HRR but not the NER factor expression provides an effective strategy for sensitizing cisplatin-resistant tumors to platinating agents.
Highlights
While platinum-based chemotherapeutic agents are widely used to treat various solid tumors, the acquired platinum resistance is a major impediment in their successful treatment
The amount of Pt-GG produced by 7.5 μM of cisplatin in 2008 cells is equivalent to that produced by 40 μM of cisplatin in 2008C13 cells, and the amount of Pt-GG induced by 15 μM of cisplatin in PEO1 cells is equivalent to that induced by 20 μM of cisplatin in PEO4 cells
In subsequent experiments, we used the different doses of cisplatin to treat different cell lines to ensure that the same initial amount of Pt-GG was produced when assessing the nucleotide excision repair (NER) capacity in this study
Summary
While platinum-based chemotherapeutic agents are widely used to treat various solid tumors, the acquired platinum resistance is a major impediment in their successful treatment. Cisplatin forms primarily 1, 2-intrastrand cross-links between adjacent purines in DNA, e.g. cis-Pt(NH3)2d (GpG) (Pt-GG), with Pt bound to two adjacent guanines, and cis-Pt(NH3)2d(ApG) (Pt-AG), in which the Pt is bound to adenine and an adjacent guanine. These lesions contribute to 90% of total damage introduced by cisplatin. The cisplatin-induced intrastrand cross-links are mainly removed by nucleotide excision repair (NER). Alteration of this DNA repair pathway is believed to confer resistance to platinum-based chemotherapy. The repair of interstrand cross-links induced by cisplatin is more complex, and involves excision repair and homologous recombination (HR) [6]
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