VLX600 Disrupts Homologous Recombination and Synergizes with PARP Inhibitors and Cisplatin by Inhibiting Histone Lysine Demethylases

AbstractPoly (ADP-ribose) polymerase (PARP) inhibitors elicit antitumour activity in homologous recombination-defective cancers by trapping PARP1 in a chromatin-bound state. How cells process trapped PARP1 remains unclear. Using wild-type and a trapping-deficient PARP1 mutant combined with rapid immunoprecipitation mass spectrometry of endogenous proteins and Apex2 proximity labelling, we delineated mass spectrometry-based interactomes of trapped and non-trapped PARP1. These analyses identified an interaction between trapped PARP1 and the ubiquitin-regulated p97 ATPase/segregase. We found that following trapping, PARP1 is SUMOylated by PIAS4 and subsequently ubiquitylated by the SUMO-targeted E3 ubiquitin ligase RNF4, events that promote recruitment of p97 and removal of trapped PARP1 from chromatin. Small-molecule p97-complex inhibitors, including a metabolite of the clinically used drug disulfiram (CuET), prolonged PARP1 trapping and enhanced PARP inhibitor-induced cytotoxicity in homologous recombination-defective tumour cells and patient-derived tumour organoids. Together, these results suggest that p97 ATPase plays a key role in the processing of trapped PARP1 and the response of tumour cells to PARP inhibitors.

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Rucaparib in patients presenting a metastatic breast cancer with Homologous Recombination Deficiency, without germline BRCA1/2 mutation

10.21203/rs.3.rs-78109/v1 ◽

2020 ◽

Author(s):

Anne Patsouris ◽

M'boyba Khadija DIOP ◽

Olivier Tredan ◽

Daniel Nenciu ◽

Anthony Goncalves ◽

...

Keyword(s):

Breast Cancer ◽

Metastatic Breast Cancer ◽

Homologous Recombination ◽

Statistical Significance ◽

Objective Response ◽

Metastatic Breast ◽

Parp Inhibitors ◽

Small Subset ◽

Durable Response ◽

Homologous Recombination Deficiency

Abstract Breast cancer may present genomic alterations leading to homologous recombination deficiency. PARP inhibitors have proved their efficacy in patients with HER2-negative metastatic breast cancer (mBC) harboring germline (g) BRCA1/2 mutations. We conducted the phase 2 RUBY trial to assess the efficacy of rucaparib in HER2-negative mBC with high genomic loss of heterozygosity (LOH) score or somatic, without gBRCA1/2 mutation. 220 of 711 patients with mBC screened for LOH presented high LOH score which was associated with a higher likelihood of death (HR = 1.39, 95% CI: 1.11-1.75, p = 0.005). The primary objective was not reached with a clinical benefit rate (objective response or SD>16 weeks) of 13.5%. Two LOH-high patients, without somatic BRCA1/2 mutation, presented a complete and durable response (14 and 32 months). HRDetect tended to be associated with response to rucaparib, whithout reaching statistical significance (median HRDetect responders versus non responders: 0.465 versus, 0.040, p = 0.2135). Our data suggests that a small subset of patients with high LOH score could derive benefit from PARP inhibitors.

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Laboratory cross-comparison of homologous recombination repair mutation analysis in tumor in a multicenter epithelial ovarian cancer series: The BORNEO GEICO 60-0 study.

Journal of Clinical Oncology ◽

10.1200/jco.2021.39.15_suppl.e17550 ◽

2021 ◽

Vol 39 (15_suppl) ◽

pp. e17550-e17550

Author(s):

Ignacio Romero ◽

Ana Oaknin ◽

Zaida Garcia-Casado ◽

Raul Marquez ◽

Alfonso Yubero Esteban ◽

...

Keyword(s):

Ovarian Cancer ◽

Homologous Recombination ◽

Epithelial Ovarian Cancer ◽

Mutational Analysis ◽

Figo Stage ◽

Parp Inhibitors ◽

Homologous Recombination Repair ◽

Recombination Repair ◽

Class 1 ◽

Uncertain Significance

e17550 Background: In epithelial ovarian cancer (EOC), the identification of mutations in homologous recombination repair (HRR) genes on tumor is prognostic, predictive of response to PARP inhibitors, and a tool to identify individuals at genetic cancer risk. The aim of this study is to compare the concordance between two laboratories in identifying and classifying genetic variants in HRR genes. Methods: In a multicentre ambispective series of unselected, non mucinous EOC of all stages formalin-fixed and paraffin embedded tumors were collected. These samples underwent the same mutational analysis of 15 HRR genes ( ATM, BARD1, BRCA1, BRCA2, BRIP1, CDK12, CHEK1, CHEK2, FANCL, PALB2, PPP2R2A, RAD51B, RAD51C, RAD51D, RAD54L) in two different Laboratories (Lab1, Lab2) that used their own validated multi-gene NGS panels. Variant allele frequency (VAF) threshold was 5% for single nucleotide polymorphism and 10% for indels. Large rearrangements were not analyzed. Variants were classified into three categories based on ACMG criteria: non-mutated (class 1-2), Variants of Uncertain Significance (VUS: class 3) and likely pathogenic/pathogenic (class 4-5). Results: A total of 81 cases were sent for the analysis. One had low DNA quality and therefore 80 cases were finally studied (85% high grade serous and 74% FIGO stage III-IV). Results reported by Lab1 and Lab2 (lab1/Lab2) were the following: 21/19 (26%/24%) cases had BRCA1/2 mutations, 7/8 (8.7%/10%) mutations on other HRR genes including two in ATM and RAD51D, one in CHEK1, CHEK2, and FANCL and one RAD51C reported in Lab2 only while the rest were either VUS 23/27 (29%/34%) or non-mutated 29/26 (36%/33%). Concordance between laboratories in classifying patients was 93.75% (kappa coefficient 0.86). Discrepancies (DC) on variants were classified into arbitrary categories, namely 0= complete concordance, category 1 meaning DC in detection assumed to be due to tumor heterogeneity (VAF nearby the threshold) or technique (1A), or caused by laboratories performance and avoidable (1B) and the category 2 identified DC in interpretation without clinical relevance (2A) or clinically relevant (2B), the results of total number of variants are shown in table. Overall, regarding clinically relevant DC in HRR genes, 9 DC in variants were observed including six 2B, two 1A and one 1B and they affect 5 (6.3%) patients since some were overlapping. Conclusions: In our EOC series the concordance of two Laboratories in the identification of clinically relevant HRR mutations on tumor is high but discrepancies in interpretation remain a challenge that needs further harmonization.[Table: see text]

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GSK3β Inhibition Synergizes with PARP Inhibitors through the Induction of Homologous Recombination Deficiency in Colorectal Cancer

10.21203/rs.3.rs-48785/v1 ◽

2020 ◽

Author(s):

Ning Zhang ◽

Yu-Nan Tian ◽

Li-Na Zhou ◽

Meng-Zhu Li ◽

Shan-Shan Song ◽

...

Keyword(s):

Colorectal Cancer ◽

Homologous Recombination ◽

High Throughput Screening ◽

Glycogen Synthase ◽

Objective Response ◽

Xenograft Model ◽

Parp Inhibitors ◽

High Rate ◽

Parp Inhibition ◽

Mouse Xenograft Model

Abstract Background: Monotherapy with poly ADP-ribose polymerase (PARP) inhibitors results in limited objective response rate (≤ 60% in most cases) in patients with homologous recombination repair (HRR)-deficient cancer, which suggests a high rate of resistance in this subset of patients to PARP inhibitors (PARPi). To overcome resistance to PARPi and to broaden their clinical use, we performed high-throughput screening of 99 anticancer drugs in combination with PARPi to identify potential therapeutic combinations. Methods: The effects of PARPi combined with glycogen synthase kinase 3 (GSK3) inhibitors were investigated in vitro with respect to cell viability, cell cycle and apoptosis. The synergy was assessed by calculation of the combination index (CI). GSK3α null and GSK3β null cells were generated using CRISPR/Cas9 technique. The underlying mechanism was examined by western blotting, flow cytometry, qRT-PCR and fluorescence microscopy. This combination was also evaluated in the mouse xenograft model; tumor growth and tumor lysates were analyzed, and the immunohistochemistry assay was performed. All data are presented as mean ± SD. Comparison between two groups was performed with the Student’s t-test.Result: The data showed that ~25% of oncological drugs and kinase inhibitors that were evaluated displayed synergy with PARPi in HCT-15 cells. Among the tested agents, GSK3 inhibitors (GSK3i) exhibited the strongest synergistic effect with PARPi. Moreover, the synergistic antitumor effect of GSK3 and PARP inhibition was confirmed in a panel of colorectal cancer (CRC) cell lines with diverse genetic backgrounds. Additionally, inhibition or genetic depletion of GSK3β was found to impair HRR of DNA and reduce the mRNA and protein level of BRCA1. Finally, we demonstrated that inhibition or depletion of GSK3β could enhance the in vivo sensitivity to simmiparib without toxicity.Conclusion: Our results provide a mechanistic understanding of combination of PARP and GSK3 inhibition, and support the clinical development of this combination therapy for CRC patients.

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PARP inhibitors alone and in combination with other biological agents in homologous recombination deficient epithelial ovarian cancer: From the basic research to the clinic

Journal of Cancer Science & Therapy ◽

10.4172/1948-5956-c1-110 ◽

2017 ◽

Vol 09 (09) ◽

Author(s):

Angiolo Gadducci

Keyword(s):

Ovarian Cancer ◽

Homologous Recombination ◽

Epithelial Ovarian Cancer ◽

Basic Research ◽

Parp Inhibitors ◽

Biological Agents

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WIP1 Promotes Homologous Recombination and Modulates Sensitivity to PARP Inhibitors

Cells ◽

10.3390/cells8101258 ◽

2019 ◽

Vol 8 (10) ◽

pp. 1258 ◽

Cited By ~ 5

Author(s):

Kamila Burdova ◽

Radka Storchova ◽

Matous Palek ◽

Libor Macurek

Keyword(s):

Dna Damage ◽

Dna Repair ◽

Homologous Recombination ◽

Cancer Cells ◽

Genotoxic Stress ◽

Parp Inhibitors ◽

Cell Cycle Checkpoint ◽

Dna Lesion ◽

Cycle Checkpoint ◽

G2 Cells

Genotoxic stress triggers a combined action of DNA repair and cell cycle checkpoint pathways. Protein phosphatase 2C delta (referred to as WIP1) is involved in timely inactivation of DNA damage response by suppressing function of p53 and other targets at chromatin. Here we show that WIP1 promotes DNA repair through homologous recombination. Loss or inhibition of WIP1 delayed disappearance of the ionizing radiation-induced 53BP1 foci in S/G2 cells and promoted cell death. We identify breast cancer associated protein 1 (BRCA1) as interactor and substrate of WIP1 and demonstrate that WIP1 activity is needed for correct dynamics of BRCA1 recruitment to chromatin flanking the DNA lesion. In addition, WIP1 dephosphorylates 53BP1 at Threonine 543 that was previously implicated in mediating interaction with RIF1. Finally, we report that inhibition of WIP1 allowed accumulation of DNA damage in S/G2 cells and increased sensitivity of cancer cells to a poly-(ADP-ribose) polymerase inhibitor olaparib. We propose that inhibition of WIP1 may increase sensitivity of BRCA1-proficient cancer cells to olaparib.

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PARP Inhibitors as a Therapeutic Agent for Homologous Recombination Deficiency in Breast Cancers

Journal of Clinical Medicine ◽

10.3390/jcm8040435 ◽

2019 ◽

Vol 8 (4) ◽

pp. 435 ◽

Cited By ~ 29

Author(s):

Man Keung ◽

Yanyuan Wu ◽

Jaydutt Vadgama

Keyword(s):

Dna Repair ◽

Homologous Recombination ◽

Cancer Cells ◽

Anticancer Agents ◽

Excision Repair ◽

Parp Inhibitor ◽

Predictive Biomarkers ◽

Parp Inhibitors ◽

Homologous Recombination Deficiency ◽

Repair Pathways

Poly (ADP-ribose) polymerases (PARPs) play an important role in various cellular processes, such as replication, recombination, chromatin remodeling, and DNA repair. Emphasizing PARP’s role in facilitating DNA repair, the PARP pathway has been a target for cancer researchers in developing compounds which selectively target cancer cells and increase sensitivity of cancer cells to other anticancer agents, but which also leave normal cells unaffected. Since certain tumors (BRCA1/2 mutants) have deficient homologous recombination repair pathways, they depend on PARP-mediated base excision repair for survival. Thus, inhibition of PARP is a promising strategy to selectively kill cancer cells by inactivating complementary DNA repair pathways. Although PARP inhibitor therapy has predominantly targeted BRCA-mutated cancers, this review also highlights the growing conversation around PARP inhibitor treatment for non-BRCA-mutant tumors, those which exhibit BRCAness and homologous recombination deficiency. We provide an update on the field’s progress by considering PARP inhibitor mechanisms, predictive biomarkers, and clinical trials of PARP inhibitors in development. Bringing light to these findings would provide a basis for expanding the use of PARP inhibitors beyond BRCA-mutant breast tumors.

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TIP60 K430 SUMOylation attenuates its interaction with DNA-PKcs in S-phase cells: Facilitating homologous recombination and emerging target for cancer therapy

Science Advances ◽

10.1126/sciadv.aba7822 ◽

2020 ◽

Vol 6 (28) ◽

pp. eaba7822 ◽

Cited By ~ 1

Author(s):

Shan-Shan Gao ◽

Hua Guan ◽

Shuang Yan ◽

Sai Hu ◽

Man Song ◽

...

Keyword(s):

Cancer Therapy ◽

Homologous Recombination ◽

Cellular Response ◽

Parp Inhibitors ◽

S Phase ◽

Tumor Growth Inhibition ◽

Dna Double Strand Breaks ◽

End Joining ◽

Pathway Activation ◽

Pathway Choice

Nonhomologous end joining (NHEJ) and homologous recombination (HR) are major repair pathways of DNA double-strand breaks (DSBs). The pathway choice of HR and NHEJ is tightly regulated in cellular response to DNA damage. Here, we demonstrate that the interaction of TIP60 with DNA-PKcs is attenuated specifically in S phase, which facilitates HR pathway activation. SUMO2 modification of TIP60 K430 mediated by PISA4 E3 ligase blocks its interaction with DNA-PKcs, whereas TIP60 K430R mutation recovers its interaction with DNA-PKcs, which results in abnormally increased phosphorylation of DNA-PKcs S2056 in S phase and marked inhibition of HR efficiency, but barely affects NHEJ activity. TIP60 K430R mutant cancer cells are more sensitive to radiation and PARP inhibitors in cancer cell killing and tumor growth inhibition. Collectively, coordinated regulation of TIP60 and DNA-PKcs facilitates HR pathway choice in S-phase cells. TIP60 K430R mutant is a potential target of radiation and PARPi cancer therapy.

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Frequency and prognostic value of mutations associated with the homologous recombination DNA repair pathway in a large pan cancer cohort

Scientific Reports ◽

10.1038/s41598-020-76975-6 ◽

2020 ◽

Vol 10 (1) ◽

Author(s):

Daniel R. Principe ◽

Matthew Narbutis ◽

Regina Koch ◽

Ajay Rana

Keyword(s):

Dna Repair ◽

Homologous Recombination ◽

Synthetic Lethality ◽

Parp Inhibitors ◽

The Cancer Genome Atlas ◽

Parp Inhibition ◽

Repair Pathway ◽

Wide Range ◽

Recombination Pathway ◽

Pan Cancer

AbstractPARP inhibitors have shown remarkable efficacy in the clinical management of several BRCA-mutated tumors. This approach is based on the long-standing hypothesis that PARP inhibition will impair the repair of single stranded breaks, causing synthetic lethality in tumors with loss of high-fidelity double-strand break homologous recombination. While this is now well accepted and has been the basis of several successful clinical trials, emerging evidence strongly suggests that mutation to several additional genes involved in homologous recombination may also have predictive value for PARP inhibitors. While this notion is supported by early clinical evidence, the mutation frequencies of these and other functionally related genes are largely unknown, particularly in cancers not classically associated with homologous recombination deficiency. We therefore evaluated the mutation status of 22 genes associated with the homologous recombination DNA repair pathway or PARP inhibitor sensitivity, first in a pan-cancer cohort of 55,586 patients, followed by a more focused analysis in The Cancer Genome Atlas cohort of 12,153 patients. In both groups we observed high rates of mutations in a variety of HR-associated genes largely unexplored in the setting of PARP inhibition, many of which were associated also with poor clinical outcomes. We then extended our study to determine which mutations have a known oncogenic role, as well as similar to known oncogenic mutations that may have a similar phenotype. Finally, we explored the individual cancer histologies in which these genomic alterations are most frequent. We concluded that the rates of deleterious mutations affecting genes associated with the homologous recombination pathway may be underrepresented in a wide range of human cancers, and several of these genes warrant further and more focused investigation, particularly in the setting of PARP inhibition and HR deficiency.

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