Exploring the Synergy between PARP and CHK1 Inhibition in Matched BRCA2 Mutant and Corrected Cells.

Lisa Prendergast,Hannah L Smith,Nicola J Curtin

doi:10.3390/cancers12040878

Lisa Prendergast, Hannah L Smith + Show 1 more

Open Access

https://doi.org/10.3390/cancers12040878

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Journal: Cancers	Publication Date: Apr 4, 2020
Citations: 18	License type: CC BY 4.0

Affiliation: Cancer Research UK, Newcastle University

Abstract

PARP inhibition results in the accumulation of DNA SSBs, causing replication stress (RS) and lesions that can only be resolved by homologous recombination repair (HRR). Defects in HRR, e.g., due to BRCA2 mutation, confer profound sensitivity to PARP inhibitor (PARPi) cytotoxicity. In response to RS, CHK1 is activated to signal to S and G2/M cell cycle checkpoints and also to HRR. To determine the relative contribution of these two functions of CHK1 to survival following PARPi exposure, we investigated the effects of rucaparib (a PARPi) and PF-477736 (a CHK1 inhibitor) alone and in combination in cells with mutated and corrected BRCA2. The BRCA2 mutated V-C8 cells were 1000× more sensitive to rucaparib cytotoxicity than their matched BRCA2 corrected V-C8.B2 cells, but no more sensitive to PF-477736 despite having seven-fold higher levels of RS. PF-477736 caused a five-fold enhancement of rucaparib cytotoxicity in the V-C8.B2 cells, but no enhancement in the V-C8 cells. This differential sensitivity was not due to a difference in PARP1 or CHK1 expression or activity. PF-477736 increased rucaparib-induced RS (γH2AX foci) and completely inhibited RAD51 focus formation, indicating a profound suppression of HRR. Our data suggested that inhibition of HRR was the main mechanism of sensitisation to rucaparib, compounded with an inhibition of cell cycle checkpoints by PF-477736.

Highlights

Replication stress (RS) is a key source of genomic instability, an enabling characteristic of cancer [1]
Three Poly(ADP-ribose) polymerase (PARP) inhibitors (PARPi) are currently approved for the treatment of ovarian cancer, the success being largely due to the high frequency (>50%) of homologous recombination DNA repair (HRR) defects in this cancer type [5,6,7]
No significant difference in cytotoxicity to PF-477736 (Figure 1b) was observed between the cell lines as both V-C8 and V-C8.B2 cells had similar LC50 (100.9 and 87.5 nM, respectively). This suggested that BRCA2/HRR defective (HRD) status was not a determinant of sensitivity to Checkpoint kinase 1 (CHK1) inhibitor PF-477736

Summary

Introduction

Replication stress (RS) is a key source of genomic instability, an enabling characteristic of cancer [1]. RS is increased in cancer due to an almost ubiquitous loss of G1 checkpoint control [2] and unrepaired DNA lesions encountering the replication fork. PARP is the first line of defence against the most common type of endogenous DNA damage, oxidative stress, a major contributor to RS. PARP inhibitors (PARPi) are a significant breakthrough in the treatment of cancer by exploiting cancer-specific defects in homologous recombination DNA repair (HRR), e.g., due to BRCA mutations. Toxicities associated with these drugs are generally mild [4]. Three PARPi are currently approved for the treatment of ovarian cancer, the success being largely due to the high frequency (>50%) of HRR defects in this cancer type [5,6,7]

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