The USP1 Inhibitor KSQ-4279 Overcomes PARP Inhibitor Resistance in Homologous Recombination-Deficient Tumors.

Defects in DNA repair pathways play a pivotal role in tumor evolution and resistance to therapy. At the same time, they create vulnerabilities that render tumors dependent on the remaining DNA repair processes. This phenomenon is exemplified by the clinical activity of PARP inhibitors in tumors with homologous recombination (HR) repair defects, such as tumors with inactivating mutations in BRCA1 or BRCA2. However, the development of resistance to PARP inhibitors in BRCA-mutant tumors represents a high unmet clinical need. In this study, we identified deubiquitinase ubiquitin-specific peptidase-1 (USP1) as a critical dependency in tumors with BRCA mutations or other forms of HR deficiency and developed KSQ-4279, the first potent and selective USP1 inhibitor to enter clinical testing. The combination of KSQ-4279 with a PARP inhibitor was well tolerated and induced durable tumor regression across several patient-derived PARP-resistant models. These findings indicate that USP1 inhibitors represent a promising therapeutic strategy for overcoming PARP inhibitor resistance in patients with BRCA-mutant/HR-deficient tumors and support continued testing in clinical trials. Significance: KSQ-4279 is a potent and selective inhibitor of USP1 that induces regression of PARP inhibitor-resistant tumors when dosed in combination with PARP inhibitors, addressing an unmet clinical need for BRCA-mutant tumors.

Histone-methyltransferase KMT2D deficiency impairs the Fanconi anemia/BRCA pathway upon glycolytic inhibition in squamous cell carcinoma.

Histone lysine methyltransferase 2D (KMT2D) is the most frequently mutated epigenetic modifier in head and neck squamous cell carcinoma (HNSCC). However, the role of KMT2D in HNSCC tumorigenesis and whether its mutations confer any therapeutic vulnerabilities remain unknown. Here we show that KMT2D deficiency promotes HNSCC growth through increasing glycolysis. Additionally, KMT2D loss decreases the expression of Fanconi Anemia (FA)/BRCA pathway genes under glycolytic inhibition. Mechanistically, glycolytic inhibition facilitates the occupancy of KMT2D to the promoter/enhancer regions of FA genes. KMT2D loss reprograms the epigenomic landscapes of FA genes by transiting their promoter/enhancer states from active to inactive under glycolytic inhibition. Therefore, combining the glycolysis inhibitor 2-DG with DNA crosslinking agents or poly (ADP-ribose) polymerase (PARP) inhibitors preferentially inhibits tumor growth of KMT2D-deficient mouse HNSCC and patient-derived xenografts (PDXs) harboring KMT2D-inactivating mutations. These findings provide an epigenomic basis for developing targeted therapies for HNSCC patients with KMT2D-inactivating mutations.

Nucleolytic processing of abasic sites underlies PARP inhibitor hypersensitivity in ALC1-deficient BRCA mutant cancer cells.

Clinical success with poly (ADP-ribose) polymerase inhibitors (PARPi) is impeded by inevitable resistance and associated cytotoxicity. Depletion of Amplified in Liver Cancer 1 (ALC1), a chromatin-remodeling enzyme, can overcome these limitations by hypersensitizing BReast CAncer genes 1/2 (BRCA1/2) mutant cells to PARPi. Here, we demonstrate that PARPi hypersensitivity upon ALC1 loss is reliant on its role in promoting the repair of chromatin buried abasic sites. We show that ALC1 enhances the ability of the abasic site processing enzyme, Apurinic/Apyrimidinic endonuclease 1 (APE1) to cleave nucleosome-occluded abasic sites. However, unrepaired abasic sites in ALC1-deficient cells are readily accessed by APE1 at the nucleosome-free replication forks. APE1 cleavage leads to fork breakage and trapping of PARP1/2 upon PARPi treatment, resulting in hypersensitivity. Collectively, our studies reveal how cells overcome the chromatin barrier to repair abasic lesions and uncover cleavage of abasic sites as a mechanism to overcome limitations of PARPi.

Targeting DNA Damage Response Deficiency in Thoracic Cancers.

Thoracic cancers comprise non-small cell lung cancers (NSCLCs), small cell lung cancers (SCLCs) and malignant pleural mesotheliomas (MPM). Collectively, they account for the highest rate of death from malignancy worldwide. Genomic instability is a universal feature of cancer, which fuels mutations and tumour evolution. Deficiencies in DNA damage response (DDR) genes amplify genomic instability. Homologous recombination deficiency (HRD), resulting from BRCA1/BRCA2 inactivation, is exploited for therapeutic synthetic lethality with poly-ADP ribose polymerase (PARP) inhibitors in breast and ovarian cancers, as well as in prostate and pancreatic cancers. However, DDR deficiency and its therapeutic implications are less well established in thoracic cancers. Emerging evidence suggests that a subset of thoracic cancers may harbour DDR deficiency and may, thus, be effectively targeted with DDR agents. Here, we review the current evidence surrounding DDR in thoracic cancers and discuss the challenges and promise for achieving clinical benefit with such therapeutics.

Concurrent RB1 Loss and BRCA Deficiency Predicts Enhanced Immunologic Response and Long-term Survival in Tubo-ovarian High-grade Serous Carcinoma.

PURPOSE: The purpose of this study was to evaluate RB1 expression and survival across ovarian carcinoma histotypes and how co-occurrence of BRCA1 or BRCA2 (BRCA) alterations and RB1 loss influences survival in tubo-ovarian high-grade serous carcinoma (HGSC). EXPERIMENTAL DESIGN: RB1 protein expression was classified by immunohistochemistry in ovarian carcinomas of 7,436 patients from the Ovarian Tumor Tissue Analysis consortium. We examined RB1 expression and germline BRCA status in a subset of 1,134 HGSC, and related genotype to overall survival (OS), tumor-infiltrating CD8+ lymphocytes, and transcriptomic subtypes. Using CRISPR-Cas9, we deleted RB1 in HGSC cells with and without BRCA1 alterations to model co-loss with treatment response. We performed whole-genome and transcriptome data analyses on 126 patients with primary HGSC to characterize tumors with concurrent BRCA deficiency and RB1 loss. RESULTS: RB1 loss was associated with longer OS in HGSC but with poorer prognosis in endometrioid ovarian carcinoma. Patients with HGSC harboring both RB1 loss and pathogenic germline BRCA variants had superior OS compared with patients with either alteration alone, and their median OS was three times longer than those without pathogenic BRCA variants and retained RB1 expression (9.3 vs. 3.1 years). Enhanced sensitivity to cisplatin and paclitaxel was seen in BRCA1-altered cells with RB1 knockout. Combined RB1 loss and BRCA deficiency correlated with transcriptional markers of enhanced IFN response, cell-cycle deregulation, and reduced epithelial-mesenchymal transition. CD8+ lymphocytes were most prevalent in BRCA-deficient HGSC with co-loss of RB1. CONCLUSIONS: Co-occurrence of RB1 loss and BRCA deficiency was associated with exceptionally long survival in patients with HGSC, potentially due to better treatment response and immune stimulation.

Homologous Recombination Repair Deficiency in Metastatic Prostate Cancer: New Therapeutic Opportunities.

More than 20% of metastatic prostate cancer carries genomic defects involving DNA damage repair pathways, mainly in homologous recombination repair-related genes. The recent approval of olaparib has paved the way to precision medicine for the treatment of metastatic prostate cancer with PARP inhibitors in this subset of patients, especially in the case of BRCA1 or BRCA2 pathogenic/likely pathogenic variants. In face of this new therapeutic opportunity, many issues remain unsolved. This narrative review aims to describe the relationship between homologous recombination repair deficiency and prostate cancer, the techniques used to determine homologous recombination repair status in prostate cancer, the crosstalk between homologous recombination repair and the androgen receptor pathway, the current evidence on PARP inhibitors activity in metastatic prostate cancer also in homologous recombination repair-proficient tumors, as well as emerging mechanisms of resistance to PARP inhibitors. The possibility of combination therapies including a PARP inhibitor is an attractive option, and more robust data are awaited from ongoing phase II and phase III trials outlined in this manuscript.

H2AX promotes replication fork degradation and chemosensitivity in BRCA-deficient tumours.

Histone H2AX plays a key role in DNA damage signalling in the surrounding regions of DNA double-strand breaks (DSBs). In response to DNA damage, H2AX becomes phosphorylated on serine residue 139 (known as γH2AX), resulting in the recruitment of the DNA repair effectors 53BP1 and BRCA1. Here, by studying resistance to poly(ADP-ribose) polymerase (PARP) inhibitors in BRCA1/2-deficient mammary tumours, we identify a function for γH2AX in orchestrating drug-induced replication fork degradation. Mechanistically, γH2AX-driven replication fork degradation is elicited by suppressing CtIP-mediated fork protection. As a result, H2AX loss restores replication fork stability and increases chemoresistance in BRCA1/2-deficient tumour cells without restoring homology-directed DNA repair, as highlighted by the lack of DNA damage-induced RAD51 foci. Furthermore, in the attempt to discover acquired genetic vulnerabilities, we find that ATM but not ATR inhibition overcomes PARP inhibitor (PARPi) resistance in H2AX-deficient tumours by interfering with CtIP-mediated fork protection. In summary, our results demonstrate a role for H2AX in replication fork biology in BRCA-deficient tumours and establish a function of H2AX separable from its classical role in DNA damage signalling and DSB repair.

Long-molecule scars of backup DNA repair in BRCA1- and BRCA2-deficient cancers.

Homologous recombination (HR) deficiency is associated with DNA rearrangements and cytogenetic aberrations1. Paradoxically, the types of DNA rearrangements that are specifically associated with HR-deficient cancers only minimally affect chromosomal structure2. Here, to address this apparent contradiction, we combined genome-graph analysis of short-read whole-genome sequencing (WGS) profiles across thousands of tumours with deep linked-read WGS of 46 BRCA1- or BRCA2-mutant breast cancers. These data revealed a distinct class of HR-deficiency-enriched rearrangements called reciprocal pairs. Linked-read WGS showed that reciprocal pairs with identical rearrangement orientations gave rise to one of two distinct chromosomal outcomes, distinguishable only with long-molecule data. Whereas one (cis) outcome corresponded to the copying and pasting of a small segment to a distant site, a second (trans) outcome was a quasi-balanced translocation or multi-megabase inversion with substantial (10 kb) duplications at each junction. We propose an HR-independent replication-restart repair mechanism to explain the full spectrum of reciprocal pair outcomes. Linked-read WGS also identified single-strand annealing as a repair pathway that is specific to BRCA2 deficiency in human cancers. Integrating these features in a classifier improved discrimination between BRCA1- and BRCA2-deficient genomes. In conclusion, our data reveal classes of rearrangements that are specific to BRCA1 or BRCA2 deficiency as a source of cytogenetic aberrations in HR-deficient cells.

Cockayne syndrome group B protein regulates fork restart, fork progression and MRE11-dependent fork degradation in BRCA1/2-deficient cells.

Cockayne syndrome group B (CSB) protein has been implicated in the repair of a variety of DNA lesions that induce replication stress. However, little is known about its role at stalled replication forks. Here, we report that CSB is recruited to stalled forks in a manner dependent upon its T1031 phosphorylation by CDK. While dispensable for MRE11 association with stalled forks in wild-type cells, CSB is required for further accumulation of MRE11 at stalled forks in BRCA1/2-deficient cells. CSB promotes MRE11-mediated fork degradation in BRCA1/2-deficient cells. CSB possesses an intrinsic ATP-dependent fork reversal activity in vitro, which is activated upon removal of its N-terminal region that is known to autoinhibit CSB's ATPase domain. CSB functions similarly to fork reversal factors SMARCAL1, ZRANB3 and HLTF to regulate slowdown in fork progression upon exposure to replication stress, indicative of a role of CSB in fork reversal in vivo. Furthermore, CSB not only acts epistatically with MRE11 to facilitate fork restart but also promotes RAD52-mediated break-induced replication repair of double-strand breaks arising from cleavage of stalled forks by MUS81 in BRCA1/2-deficient cells. Loss of CSB exacerbates chemosensitivity in BRCA1/2-deficient cells, underscoring an important role of CSB in the treatment of cancer lacking functional BRCA1/2.

Loss of MED12 activates the TGFß pathway to promote chemoresistance and replication fork stability in BRCA-deficient cells.

Understanding chemoresistance mechanisms in BRCA-deficient cells will allow for identification of biomarkers for predicting tumor response to therapy, as well as the design of novel therapeutic approaches targeting this chemoresistance. Here, we show that the protein MED12, a component of the Mediator transcription regulation complex, plays an unexpected role in regulating chemosensitivity in BRCA-deficient cells. We found that loss of MED12 confers resistance to cisplatin and PARP inhibitors in both BRCA1- and BRCA2-deficient cells, which is associated with restoration of both homologous recombination and replication fork stability. Surprisingly, MED12-controlled chemosensitivity does not involve a function of the Mediator complex, but instead reflects a distinct role of MED12 in suppression of the TGFß pathway. Importantly, we show that ectopic activation of the TGFß pathway is enough to overcome the fork protection and DNA repair defects of BRCA-mutant cells, resulting in chemoresistance. Our work identifies the MED12-TGFß module as an important regulator of genomic stability and chemosensitivity in BRCA-deficient cells.

Sirtuin inhibition is synthetic lethal with BRCA1 or BRCA2 deficiency.

PARP enzymes utilise NAD+ as a co-substrate for their enzymatic activity. Inhibition of PARP1 is synthetic lethal with defects in either BRCA1 or BRCA2. In order to assess whether other genes implicated in NAD+ metabolism were synthetic lethal with BRCA1 or BRCA2 gene defects, we carried out a genetic screen, which identified a synthetic lethality between BRCA1 and genetic inhibition of either of two sirtuin (SIRT) enzymes, SIRT1 or SIRT6. This synthetic lethal interaction was replicated using small-molecule SIRT inhibitors and was associated with replication stress and increased cellular PARylation, in contrast to the decreased PARylation associated with BRCA-gene/PARP inhibitor synthetic lethality. SIRT/BRCA1 synthetic lethality was reversed by genetic ablation of either PARP1 or the histone PARylation factor-coding gene HPF1, implicating PARP1/HPF1-mediated serine ADP-ribosylation as part of the mechanistic basis of this synthetic lethal effect. These observations suggest that PARP1/HPF1-mediated serine ADP-ribosylation, when driven by SIRT inhibition, can inadvertently inhibit the growth of BRCA-gene mutant cells.

A transcription-based mechanism for oncogenic ß-catenin-induced lethality in BRCA1/2-deficient cells.

BRCA1 or BRCA2 germline mutations predispose to breast, ovarian and other cancers. High-throughput sequencing of tumour genomes revealed that oncogene amplification and BRCA1/2 mutations are mutually exclusive in cancer, however the molecular mechanism underlying this incompatibility remains unknown. Here, we report that activation of ß-catenin, an oncogene of the WNT signalling pathway, inhibits proliferation of BRCA1/2-deficient cells. RNA-seq analyses revealed ß-catenin-induced discrete transcriptome alterations in BRCA2-deficient cells, including suppression of CDKN1A gene encoding the CDK inhibitor p21. This accelerates G1/S transition, triggering illegitimate origin firing and DNA damage. In addition, ß-catenin activation accelerates replication fork progression in BRCA2-deficient cells, which is critically dependent on p21 downregulation. Importantly, we find that upregulated p21 expression is essential for the survival of BRCA2-deficient cells and tumours. Thus, our work demonstrates that ß-catenin toxicity in cancer cells with compromised BRCA1/2 function is driven by transcriptional alterations that cause aberrant replication and inflict DNA damage.

BRCA2 deficiency reveals that oxidative stress impairs RNaseH1 function to cripple mitochondrial DNA maintenance.

Oxidative stress is a ubiquitous cellular challenge implicated in aging, neurodegeneration, and cancer. By studying pathogenic mutations in the tumor suppressor BRCA2, we identify a general mechanism by which oxidative stress restricts mitochondrial (mt)DNA replication. BRCA2 inactivation induces R-loop accumulation in the mtDNA regulatory region and diminishes mtDNA replication initiation. In BRCA2-deficient cells, intracellular reactive oxygen species (ROS) are elevated, and ROS scavengers suppress the mtDNA defects. Conversely, wild-type cells exposed to oxidative stress by pharmacologic or genetic manipulation phenocopy these defects. Mechanistically, we find that 8-oxoguanine accumulation in mtDNA caused by oxidative stress suffices to impair recruitment of the mitochondrial enzyme RNaseH1 to sites of R-loop accrual, restricting mtDNA replication initiation. Thus, oxidative stress impairs RNaseH1 function to cripple mtDNA maintenance. Our findings highlight a molecular mechanism that links oxidative stress to mitochondrial dysfunction and is elicited by the inactivation of genes implicated in neurodegeneration and cancer.

The Fanconi anemia ubiquitin E3 ligase complex as an anti-cancer target.

Agents that induce DNA damage can cure some cancers. However, the side effects of chemotherapy are severe because of the indiscriminate action of DNA-damaging agents on both healthy and cancerous cells. DNA repair pathway inhibition provides a less toxic and targeted alternative to chemotherapy. A compelling DNA repair target is the Fanconi anemia (FA) E3 ligase core complex due to its critical-and likely singular-role in the efficient removal of specific DNA lesions. FA pathway inactivation has been demonstrated to specifically kill some types of cancer cells without the addition of exogenous DNA damage, including cells that lack BRCA1, BRCA2, ATM, or functionally related genes. In this perspective, we discuss the genetic and biochemical evidence in support of the FA core complex as a compelling drug target for cancer therapy. In particular, we discuss the genetic, biochemical, and structural data that could rapidly advance our capacity to identify and implement the use of FA core complex inhibitors in the clinic.

Targeting HR Repair as a Synthetic Lethal Approach to Increase DNA Damage Sensitivity by a RAD52 Inhibitor in BRCA2-Deficient Cancer Cells.

BRCA mutation, one of the most common types of mutations in breast and ovarian cancer, has been suggested to be synthetically lethal with depletion of RAD52. Pharmacologically inhibiting RAD52 specifically eradicates BRCA-deficient cancer cells. In this study, we demonstrated that curcumin, a plant polyphenol, sensitizes BRCA2-deficient cells to CPT-11 by impairing RAD52 recombinase in MCF7 cells. More specifically, in MCF7-siBRCA2 cells, curcumin reduced homologous recombination, resulting in tumor growth suppression. Furthermore, a BRCA2-deficient cell line, Capan1, became resistant to CPT-11 when BRCA2 was reintroduced. In vivo, xenograft model studies showed that curcumin combined with CPT-11 reduced the growth of BRCA2-knockout MCF7 tumors but not MCF7 tumors. In conclusion, our data indicate that curcumin, which has RAD52 inhibitor activity, is a promising candidate for sensitizing BRCA2-deficient cells to DNA damage-based cancer therapies.

Homology-directed repair protects the replicating genome from metabolic assaults.

Homology-directed repair (HDR) safeguards DNA integrity under various forms of stress, but how HDR protects replicating genomes under extensive metabolic alterations remains unclear. Here, we report that besides stalling replication forks, inhibition of ribonucleotide reductase (RNR) triggers metabolic imbalance manifested by the accumulation of increased reactive oxygen species (ROS) in cell nuclei. This leads to a redox-sensitive activation of the ATM kinase followed by phosphorylation of the MRE11 nuclease, which in HDR-deficient settings degrades stalled replication forks. Intriguingly, nascent DNA degradation by the ROS-ATM-MRE11 cascade is also triggered by hypoxia, which elevates signaling-competent ROS and attenuates functional HDR without arresting replication forks. Under these conditions, MRE11 degrades daughter-strand DNA gaps, which accumulate behind active replisomes and attract error-prone DNA polymerases to escalate mutation rates. Thus, HDR safeguards replicating genomes against metabolic assaults by restraining mutagenic repair at aberrantly processed nascent DNA. These findings have implications for cancer evolution and tumor therapy.

RNF168 regulates R-loop resolution and genomic stability in BRCA1/2-deficient tumors.

Germline mutations in BRCA1 and BRCA2 (BRCA1/2) genes considerably increase breast and ovarian cancer risk. Given that tumors with these mutations have elevated genomic instability, they exhibit relative vulnerability to certain chemotherapies and targeted treatments based on poly (ADP-ribose) polymerase (PARP) inhibition. However, the molecular mechanisms that influence cancer risk and therapeutic benefit or resistance remain only partially understood. BRCA1 and BRCA2 have also been implicated in the suppression of R-loops, triple-stranded nucleic acid structures composed of a DNA:RNA hybrid and a displaced ssDNA strand. Here, we report that loss of RNF168, an E3 ubiquitin ligase and DNA double-strand break (DSB) responder, remarkably protected Brca1-mutant mice against mammary tumorigenesis. We demonstrate that RNF168 deficiency resulted in accumulation of R-loops in BRCA1/2-mutant breast and ovarian cancer cells, leading to DSBs, senescence, and subsequent cell death. Using interactome assays, we identified RNF168 interaction with DHX9, a helicase involved in the resolution and removal of R-loops. Mechanistically, RNF168 directly ubiquitylated DHX9 to facilitate its recruitment to R-loop-prone genomic loci. Consequently, loss of RNF168 impaired DHX9 recruitment to R-loops, thereby abrogating its ability to resolve R-loops. The data presented in this study highlight a dependence of BRCA1/2-defective tumors on factors that suppress R-loops and reveal a fundamental RNF168-mediated molecular mechanism that governs cancer development and vulnerability.

PCAF-Mediated Histone Acetylation Promotes Replication Fork Degradation by MRE11 and EXO1 in BRCA-Deficient Cells.

Stabilization of stalled replication forks is a prominent mechanism of PARP (Poly(ADP-ribose) Polymerase) inhibitor (PARPi) resistance in BRCA-deficient tumors. Epigenetic mechanisms of replication fork stability are emerging but remain poorly understood. Here, we report the histone acetyltransferase PCAF (p300/CBP-associated) as a fork-associated protein that promotes fork degradation in BRCA-deficient cells by acetylating H4K8 at stalled replication forks, which recruits MRE11 and EXO1. A H4K8ac binding domain within MRE11/EXO1 is required for their recruitment to stalled forks. Low PCAF levels, which we identify in a subset of BRCA2-deficient tumors, stabilize stalled forks, resulting in PARPi resistance in BRCA-deficient cells. Furthermore, PCAF activity is tightly regulated by ATR (ataxia telangiectasia and Rad3-related), which phosphorylates PCAF on serine 264 (S264) to limit its association and activity at stalled forks. Our results reveal PCAF and histone acetylation as critical regulators of fork stability and PARPi responses in BRCA-deficient cells, which provides key insights into targeting BRCA-deficient tumors and identifying epigenetic modulators of chemotherapeutic responses.

Maintenance Therapy for ATM-Deficient Pancreatic Cancer by Multiple DNA Damage Response Interferences after Platinum-Based Chemotherapy.

Personalized medicine in treating pancreatic ductal adenocarcinoma (PDAC) is still in its infancy, albeit PDAC-related deaths are projected to rise over the next decade. Only recently, maintenance therapy with the PARP inhibitor olaparib showed improved progression-free survival in germline BRCA1/2-mutated PDAC patients after platinum-based induction for the first time. Transferability of such a concept to other DNA damage response (DDR) genes remains unclear. Here, we conducted a placebo-controlled, three-armed preclinical trial to evaluate the efficacy of multi-DDR interference (mDDRi) as maintenance therapy vs. continuous FOLFIRINOX treatment, implemented with orthotopically transplanted ATM-deficient PDAC cell lines. Kaplan-Meier analysis, cross-sectional imaging, histology, and in vitro analysis served as analytical readouts. Median overall survival was significantly longer in the mDDRi maintenance arm compared to the maintained FOLFIRINOX treatment. This survival benefit was mirrored in the highest DNA-damage load, accompanied by superior disease control and reduced metastatic burden. In vitro analysis suggests FOLFIRINOX-driven selection of invasive subclones, erased by subsequent mDDRi treatment. Collectively, this preclinical trial substantiates mDDRi in a maintenance setting as a novel therapeutic option and extends the concept to non-germline BRCA1/2-mutant PDAC.

Brca2 deficiency drives gastrointestinal tumor formation and is selectively inhibited by mitomycin C.

BRCA2 is crucial for repairing DNA double-strand breaks with high fidelity, and loss of BRCA2 increases the risks of developing breast and ovarian cancers. Herein, we show that BRCA2 is inactively mutated in 10% of gastric and 7% of colorectal adenocarcinomas, and that this inactivation is significantly correlated with microsatellite instability. Villin-driven Brca2 depletion promotes mouse gastrointestinal tumor formation when genome instability is increased. Whole-genome screening data showed that these BRCA2 monoallelic and biallelic mutant tumors were selectively inhibited by mitomycin C. Mechanistically, mitomycin C provoked double-strand breaks in cancer cells that often recruit wild-type BRCA2 for repair; the failure to repair double-strand breaks caused cell-cycle arrest at the S phase and p53-mediated cell apoptosis of BRCA2 monoallelic and biallelic mutant tumor cells. Our study unveils the role of BRCA2 loss in the development of gastrointestinal tumors and provides a potential therapeutic strategy to eliminate BRCA2 monoallelic and biallelic mutant tumors through mitomycin C.