Identification of small molecule inhibitors of ERCC1-XPF that inhibit DNA repair and potentiate cisplatin efficacy in cancer cells.

Sanjeevani Arora,Ashley M Floyd,Vivian Kalman-Maltese,Kristin Tillison,Joshua Heyza,Hao Zhang,Steve M Patrick,Elaine M Chalfin,Gerold Bepler

doi:10.18632/oncotarget.12072

Sanjeevani Arora, Ashley M Floyd + Show 7 more

Open Access

https://doi.org/10.18632/oncotarget.12072

Copy DOI

Journal: Oncotarget	Publication Date: Sep 16, 2016
Citations: 49	License type: cc-by

Affiliation: University of Toledo, Karmanos Cancer Institute

Abstract

ERCC1-XPF heterodimer is a 5′-3′ structure-specific endonuclease which is essential in multiple DNA repair pathways in mammalian cells. ERCC1-XPF (ERCC1-ERCC4) repairs cisplatin-DNA intrastrand adducts and interstrand crosslinks and its specific inhibition has been shown to enhance cisplatin cytotoxicity in cancer cells. In this study, we describe a high throughput screen (HTS) used to identify small molecules that inhibit the endonuclease activity of ERCC1-XPF. Primary screens identified two compounds that inhibit ERCC1-XPF activity in the nanomolar range. These compounds were validated in secondary screens against two other non-related endonucleases to ensure specificity. Results from these screens were validated using an in vitro gel-based nuclease assay. Electrophoretic mobility shift assays (EMSAs) further show that these compounds do not inhibit the binding of purified ERCC1-XPF to DNA. Next, in lung cancer cells these compounds potentiated cisplatin cytotoxicity and inhibited DNA repair. Structure activity relationship (SAR) studies identified related compounds for one of the original Hits, which also potentiated cisplatin cytotoxicity in cancer cells. Excitingly, dosing with NSC16168 compound potentiated cisplatin antitumor activity in a lung cancer xenograft model. Further development of ERCC1-XPF DNA repair inhibitors is expected to sensitize cancer cells to DNA damage-based chemotherapy.

Highlights

Platinum based chemotherapy is used to treat a variety of cancers including testicular, ovarian, non-small cell lung, cervical and head and neck cancers
Using the DNA substrate and the high throughput screen (HTS) assay as described in the Material and Methods we screened for the ability to inhibit the endonuclease activity of excision repair crosscomplementation group 1 (ERCC1)-xeroderma pigmentation group F (XPF)
Cleavage of the DNA substrate by HhaI is unaffected by these compounds and minimal to no effect on XPG cleavage demonstrating excellent specificity for ERCC1-XPF

Summary

Introduction

Platinum based chemotherapy is used to treat a variety of cancers including testicular, ovarian, non-small cell lung, cervical and head and neck cancers. A major limitation is either intrinsic resistance to the chemotherapy or acquired resistance during the course of treatment These factors limit clinical response and make it important to identify factors that could increase its efficacy [1, 2]. Unrepaired lesions result in inhibition of DNA replication and transcription, which can lead to the induction of apoptosis, and cancer cell killing and tumor regression. It has been well documented in the literature that increased DNA repair capacity of cancer cells is an important mechanism of cisplatin resistance and it is well accepted that therapies that target DNA repair could be important in potentiating sensitivity to cisplatin and its analogues [5, 6]

Methods

Results

Conclusion