CDK5RAP3, a New BRCA2 Partner That Regulates DNA Repair, Is Associated with Breast Cancer Survival

BRCA2 is essential for homologous recombination DNA repair. BRCA2 mutations lead to genome instability and increased risk of breast and ovarian cancer. Similarly, mutations in BRCA2-interacting proteins are also known to modulate sensitivity to DNA damage agents and are established cancer risk factors. Here we identify the tumor suppressor CDK5RAP3 as a novel BRCA2 helical domain-interacting protein. CDK5RAP3 depletion induced DNA damage resistance, homologous recombination and single-strand annealing upregulation, and reduced spontaneous and DNA damage-induced genomic instability, suggesting that CDK5RAP3 negatively regulates double-strand break repair in the S-phase. Consistent with this cellular phenotype, analysis of transcriptomic data revealed an association between low CDK5RAP3 tumor expression and poor survival of breast cancer patients. Finally, we identified common genetic variations in the CDK5RAP3 locus as potentially associated with breast and ovarian cancer risk in BRCA1 and BRCA2 mutation carriers. Our results uncover CDK5RAP3 as a critical player in DNA repair and breast cancer outcomes.

Beyond Brca1/2: Homologous Recombination Repair Genetic Profile in a Large Cohort of Apulian Ovarian Cancers

Background: Pathogenic variants in homologous recombination repair (HRR) genes other than BRCA1/2 have been associated with a high risk of ovarian cancer (OC). In current clinical practice, genetic testing is generally limited to BRCA1/2. Herein, we investigated the mutational status of both BRCA1/2 and 5 HRR genes in 69 unselected OC, evaluating the advantage of multigene panel testing in everyday clinical practice. Methods: We analyzed 69 epithelial OC samples using an NGS custom multigene panel of the 5 HRR pathways genes, beyond the genetic screening routine of BRCA1/2 testing. Results: Overall, 19 pathogenic variants (27.5%) were detected. The majority (21.7%) of patients displayed a deleterious mutation in BRCA1/2, whereas 5.8% harbored a pathogenic variant in one of the HRR genes. Additionally, there were 14 (20.3%) uncertain significant variants (VUS). The assessment of germline mutational status showed that a small number of variants (five) were not detected in the corresponding blood sample. Notably, we detected one BRIP1 and four BRCA1/2 deleterious variants in the low-grade serous and endometrioid histology OC, respectively. Conclusion: We demonstrate that using a multigene panel beyond BRCA1/2 improves the diagnostic yield in OC testing, and it could produce clinically relevant results.

The ubiquitin-dependent ATPase p97 removes cytotoxic trapped PARP1 from chromatin

Poly (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.

Homologous recombination between tandem paralogues drives evolution of Type VII secretion system immunity genes in firmicute bacteria

The Type VII secretion system (T7SS) is found in many Gram-positive firmicutes and secretes protein toxins that mediate bacterial antagonism. Two T7SS toxins have been identified in Staphylococcus aureus, EsaD a nuclease toxin that is counteracted by the EsaG immunity protein, and TspA, which has membrane depolarising activity and is neutralised by TsaI. Both toxins are polymorphic, and strings of non-identical esaG and tsaI immunity genes are encoded in all S. aureus strains. During genome sequence analysis of closely related S. aureus strains we noted that there had been a deletion of six consecutive esaG copies in one lineage. To investigate this further, we analysed the sequences of the tandem esaG genes and their encoded proteins. We identified three blocks of high sequence homology shared by all esaG genes, and identified evidence of extensive recombination events between esaG paralogues facilitated through these conserved sequence blocks. Recombination between these blocks accounts for loss of esaG genes from S. aureus genomes. TipC, an immunity protein for the TelC lipid II phosphatase toxin secreted by the streptococcal T7SS, is also encoded by multiple gene paralogues. Two blocks of high sequence homology locate to the 5-prime and 3-prime end of tipC genes, and we found strong evidence for recombination between tipC paralogues encoded by Streptococcus mitis BCC08. By contrast, we found only a single block of homology across tsaI genes, and little evidence for intergenic recombination within this gene family. We conclude that homologous recombination is one of the drivers for the evolution of T7SS immunity gene clusters.

Homologous Recombination Deficiency Associated With Response to Poly (ADP-Ribose) Polymerase Inhibitors in Ovarian Cancer Patients: The First Real-Word Evidence From China

Homologous recombination deficiency (HRD) is an approved predictive biomarker for Poly (ADP-ribose) polymerase inhibitors (PARPi) in ovarian cancer. However, the proportion of positive HRD in the real world and the relationship between HRD status and PARPi in Chinese ovarian cancer patients remain unknown. A total of 67 ovarian cancer patients who underwent PARPi, either olaparib or niraparib, were enrolled and passed inclusion criteria from August 2018 to January 2021 in the Affiliated Cancer Hospital of Nanjing Medical University. HRD status correlation with Progression-free survival (PFS) was analyzed and summarized with a log-rank test. Univariate and multiple cox-regression analyses were conducted to investigate all correlated clinical factors. Approximately 68.7% (46/67) patients were HRD positive and the rest 31.3% (21/67) were HRD negative. The PFS among HRD-positive patients was significantly longer than those HRD-negative patients (medium PFS 9.4 m vs 4.1 m, hazard ratio [HR]: 0.52, 95% CI: [0.38–0.71], p <0.001). Univariate cox-regression found that HRD status, Eastern Cooperative Oncology Group (ECOG) status, BRCA status, previous treatment lines, secondary cytoreductive surgery and R0 resection were significantly associated with PFS after PARPi treatment. After multiple regression correction, HRD status and ECOG were the independent factors to predict PFS (HR: 0.67, 95% CI: [0.49–0.92], p = 0.01; HR: 2.20, 95% CI: [1.14–4.23], p = 0.02, respectively). In platinum sensitivity evaluable subgroup (N = 49), HRD status and platinum sensitivity status remain significant to predict PFS after multiple regression correction (HR: 0.71, 95% CI: [0.51–0.98], p = 0.04; HR: 0.49, 95% CI: [0.24–1.0], p = 0.05, respectively). This is the first real-world study of HRD status in ovarian cancer patients in China, and we demonstrate that HRD is an independent predictive biomarker for PARP inhibitors treatment in Chinese ovarian cancer patients.

Platinum-Based Neoadjuvant Chemotherapy Before Radical Prostatectomy for Locally Advanced Prostate Cancer With Homologous Recombination Deficiency: A Case Report

More emerging evidence showed that homologous recombination (HR) defect (HRD) may predict sensitivity to platinum agents in metastatic prostate cancer (PCa). Platinum-based neoadjuvant chemotherapy for PCa with HRD has not been reported. Here, we reported a man diagnosed as locally advanced PCa with high Gleason Score (5 + 5) and low PSA level (5.2 ng/ml). Next-generation sequencing (NGS) demonstrated HRD. He received six cycles of platinum-based neoadjuvant chemotherapy before radical prostatectomy (RP). Fifteen months after RP, his PSA level was still undetectable, and no imaging progression was found, indicating a potential role for platinum-based neoadjuvant chemotherapy in locally advanced PCa with HRD.

Arterial Administration of DNA Crosslinking Agents with Restraint of Homologous Recombination Repair by Intravenous Low-Dose Gemcitabine Is Effective for Locally Advanced Pancreatic Cancer

(1) Background: Pretreatment by Rad51-inhibitory substances such as gemcitabine followed by arterial chemotherapy using antineoplastic agents causing DNA crosslink might be more beneficial for patients with locally advanced pancreatic cancers than conventional treatments. The efficacy of arterial administration of DNA crosslinking agents with pretreatment of intravenous low-dose gemcitabine for patients with unresectable locally advanced or metastatic pancreatic cancer (LAPC or MPC) is evaluated. (2) Methods: A single-arm, single-center, institutional review board-approved prospective study was conducted between 2005 and 2015. Forty-five patients (23 LAPC, 22 MPC) were included. Patients received a weekly low dose of gemcitabine intravenously for three weeks followed by arterial administration of mitomycin C and epirubicin hydrochloride at tumor-supplying arteries on the fifth or sixth week. This treatment course was repeated at 1.5-to-2-month intervals. Overall survival (OS), local progression-free survival (LPFS), and therapeutic response were evaluated. LAPC or MPC were divided according to treatment compliance, excellent or poor (1 or 2), to subgroups L1, L2, M1, and M2. (3) Results: OS of LAPC and MPC were 23 months and 13 months, respectively. The OS of LAPC with excellent treatment compliance (subgroup L1, 10 patients) was 33 months with 31 months of LPFS, and four patients (40%) had a complete response (CR). The OS of the L1 subgroup was significantly longer than those of other subgroups L2, M1, and M2, which were 17 months, 17 months, and 8 months, respectively. As Grade 3 adverse effects, severe bone marrow suppression, interstitial pneumonitis, and hemolytic uremic syndrome were observed in six (13.0%), three (6.5%), and three (6.5%) patients, respectively. (4) Conclusions: Arterial DNA crosslinking with the systemic restraint of homologous recombination repair can be a new treatment option for LAPC.

Landscape of homologous recombination deficiencies in solid tumours: analyses of two independent genomic datasets

Background DNA repair deficiencies are characteristic of cancer and homologous recombination deficiency (HRD) is the most common. HRD sensitizes tumour cells to PARP inhibitors so it is important to understand the landscape of HRD across different solid tumour types. Methods Germline and somatic BRCA mutations in breast and ovarian cancers were evaluated using sequencing data from The Cancer Genome Atlas (TCGA) database. Secondly, a larger independent genomic dataset was analysed to validate the TCGA results and determine the frequency of germline and somatic mutations across 15 different candidate homologous recombination repair (HRR) genes, and their relationship with the genetic events of bi-allelic loss, loss of heterozygosity (LOH) and tumour mutation burden (TMB). Results Approximately one-third of breast and ovarian cancer BRCA mutations were somatic. These showed a similar degree of bi-allelic loss and clinical outcomes to germline mutations, identifying potentially 50% more patients that may benefit from precision treatments. HRR mutations were present in sizable proportions in all tumour types analysed and were associated with high TMB and LOH scores. We also identified numerous BRCA reversion mutations across all tumour types. Conclusions Our results will facilitate future research into the efficacy of precision oncology treatments, including PARP and immune checkpoint inhibitors.