Progression through mitosis promotes PARP inhibitor-induced cytotoxicity in homologous recombination-deficient cancer cells

Abstract

Mutations in homologous recombination (HR) genes BRCA1 and BRCA2 predispose to tumorigenesis. HR-deficient cancers are hypersensitive to Poly (ADP ribose)-polymerase (PARP) inhibitors, but can acquire resistance and relapse. Mechanistic understanding how PARP inhibition induces cytotoxicity in HR-deficient cancer cells is incomplete. Here we find PARP inhibition to compromise replication fork stability in HR-deficient cancer cells, leading to mitotic DNA damage and consequent chromatin bridges and lagging chromosomes in anaphase, frequently leading to cytokinesis failure, multinucleation and cell death. PARP-inhibitor-induced multinucleated cells fail clonogenic outgrowth, and high percentages of multinucleated cells are found in vivo in remnants of PARP inhibitor-treated Brca2−/−;p53−/− and Brca1−/−;p53−/− mammary mouse tumours, suggesting that mitotic progression promotes PARP-inhibitor-induced cell death. Indeed, enforced mitotic bypass through EMI1 depletion abrogates PARP-inhibitor-induced cytotoxicity. These findings provide insight into the cytotoxic effects of PARP inhibition, and point at combination therapies to potentiate PARP inhibitor treatment of HR-deficient tumours.

Identification of regulators of poly-ADP-ribose polymerase (PARP) inhibitor response through complementary CRISPR knockout and activation screens

“…Previously, PARPi treatment has been shown to induce apoptosis in BRCA2-deficient cells. 19 In line with this, we found that olaparib treatment led to a more than 6-fold increase in cells positive for annexin V, a marker of apoptosis, in BRCA2-knockout cells; however, overexpression of ABCB1 restored olaparib-induced apoptosis to control levels ( Figure 2C). Overall, these findings validate the results of our CRISPR activation screen.…”

“…First, to identify genetic changes which sensitize cells to PARPi treatment, we employed the Brunello human CRISPR knockout pooled library, which targets 19,114 genes with four single-guide RNAs (sgRNAs) per gene. 10 HeLa cells infected with the Brunello library were divided into PARPi (5 µM olaparib)-or vehicle (DMSO)-treated arms, and after 4 days surviving cells were harvested for sgRNA sequencing and bioinformatic analysis ( Figure 1A).…”

Loss of E2F7 confers resistance to poly-ADP-ribose polymerase (PARP) inhibitors in BRCA2-deficient cells

“…1F). Olaparib treatment is known to induce apoptosis in BRCA2-deficient cells (33). E2F7 depletion did not only rescue cellular viability of olaparibtreated BRCA2-deficient cells, but also olaparib-induced apoptosis (Fig.…”

Modulation of Early Mitotic Inhibitor 1 (EMI1) Depletion on the Sensitivity of PARP Inhibitors in BRCA1 Mutated Triple-Negative Breast Cancer Cells

“…Inactivation of EMI1 leads to premature APC/C activation in G2-phase, and consequently blocks cell entry into mitosis (23,24), and as expected, there was a G2 block evident in EMI1-depleted MDA-MB-436 cells (data not shown). PARP-induced cytotoxicity has been attributed to repeated cycles of both replication and mitosis showing that forced mitotic bypass through EMI1 depletion could largely rescue viability of HDR-deficient cells upon PARP inhibition (25). Taken together, we infer that two effects of EMI1 depletion, the increased abundance of RAD51 protein and a cell cycle block in G2, combine to overcome the effects of PARPi, cisplatin, and CHK1i.…”