APOBEC3 enzymes mediate efficacy of cisplatin and are epistatic with base excision repair and mismatch repair in platinum response.

Identifying the mechanisms mediating cisplatin response is essential for improving patient response. Previous research has identified base excision repair (BER) and mismatch repair (MMR) activity in sensitizing cells to cisplatin. Cisplatin forms DNA adducts including interstrand cross-links (ICLs) that distort the DNA helix, forcing adjacent cytosines to become extrahelical. These extrahelical cytosines provide a substrate for cytosine deaminases. Herein, we show that APOBEC3 (A3) enzymes are capable of deaminating the extrahelical cytosines to uracils and sensitizing breast cancer cells to cisplatin. Knockdown of A3s results in resistance to cisplatin and induction of A3 expression in cells with low A3 expression increases sensitivity to cisplatin. We show that the actions of A3s are epistatic with BER and MMR. We propose that A3-induced cytosine deamination to uracil at cisplatin ICLs results in repair of uracils by BER, which blocks ICL DNA repair and enhances cisplatin efficacy and improves breast cancer outcomes.

HPV induction of APOBEC3 enzymes mediate overall survival and response to cisplatin in head and neck cancer.

Human papillomavirus (HPV) is associated with the development of head and neck squamous cell carcinomas (HNSC). Cisplatin is used to treat HNSC and induces DNA adducts including interstrand crosslinks (ICLs). Previous reports have shown that HPV positive HNSC patients respond better to cisplatin therapy. Our previous reports highlight that loss of base excision repair (BER) and mismatch repair (MMR) results in cisplatin resistance. Of importance, uracil DNA glycosylase (UNG) is required to initiate the BER response to cisplatin treatment and maintain drug sensitivity. These previous results highlight that specific cytidine deaminases could play an important role in the cisplatin response by activating the BER pathway to mediate drug sensitivity. The APOBEC3 (A3) family of cytidine deaminases are enzymes that restrict HPV as part of the immune defense to viral infection. In this study, the Cancer Genome Atlas (TCGA) HNSC data were used to assess the association between the expression of the seven proteins in the A3 cytidine deaminase family, HPV-status and survival outcomes. Higher A3 G expression in HPV-positive tumors corresponds with better overall survival (OS) (HR 0.33, 95 % CI 0.11-0.93, p = 0.04). FaDu and Scc-25 HNSC cell lines were used to assess alterations in A3, BER and MMR expression in response to cisplatin. We demonstrate that A3, Polß, and MSH6 knockdown in HNSC cells results in resistance to cisplatin and carboplatin as well as an increase in the rate of ICL removal in FaDu and Scc-25 HNSC cells. Our results suggest that A3s activate BER in HNSC, mediate repair of cisplatin ICLs and thereby, sensitize cells to cisplatin which likely contributes to the improved patient responses observed in HPV infected patients.

Targeting the DNA Repair Endonuclease ERCC1-XPF with Green Tea Polyphenol Epigallocatechin-3-Gallate (EGCG) and Its Prodrug to Enhance Cisplatin Efficacy in Human Cancer Cells.

The 5'-3' structure-specific endonuclease ERCC1/XPF (Excision Repair Cross-Complementation Group 1/Xeroderma Pigmentosum group F) plays critical roles in the repair of cisplatin-induced DNA damage. As such, it has been identified as a potential pharmacological target for enhancing clinical response to platinum-based chemotherapy. The goal of this study was to follow up on our previous identification of the compound NSC143099 as a potent inhibitor of ERCC1/XPF activity by performing an in silico screen to identify structural analogues that could inhibit ERCC1/XPF activity in vitro and in vivo. Using a fluorescence-based DNA-endonuclease incision assay, we identified the green tea polyphenol (-)-epigallocatechin-3-gallate (EGCG) as a potent inhibitor of ERCC1/XPF activity with an IC50 (half maximal inhibitory concentration) in the nanomolar range in biochemical assays. Using DNA repair assays and clonogenic survival assays, we show that EGCG can inhibit DNA repair and enhance cisplatin sensitivity in human cancer cells. Finally, we show that a prodrug of EGCG, Pro-EGCG (EGCG octaacetate), can enhance response to platinum-based chemotherapy in vivo. Together these data support a novel target of EGCG in cancer cells, namely ERCC1/XPF. Our studies also corroborate previous observations that EGCG enhances sensitivity to cisplatin in multiple cancer types. Thus, EGCG or its prodrug makes an ideal candidate for further pharmacological development with the goal of enhancing cisplatin response in human tumors.