Mutations In DNA Repair Genes Research Articles

Abstract 324: SIK2 inhibitors regulate DNA repair pathway and sensitize ovarian cancer to PARP1 inhibitors

Abstract Genomic instability is a recognized hallmark of cancer. Germline mutations in critical DNA-repair and DNA-damage response genes, including BRCA1and BRCA2, predispose to cancer development, but also create vulnerabilities that can be exploited for cancer therapy. The successful development of PARP inhibitors for cancers with BRCA1/2 mutations and a deficiency in homologous recombination DNA repair has provided proof of concept. Inhibition of kinases that promote transcription of DNA repair genes might provide targets that enhance the activity of PARP-inhibitors. One novel candidate, salt-inducible kinase 2 (SIK2), is overexpressed in approximately 30% of high grade serous ovarian cancers. Inhibition of SIK2 induces tetraploidy, triggers apoptotic cell death, prevents metastasis, reduces AKT/survivin signaling and decreases phosphorylation of class-IIa histone deacetylases (HDACs) enhancing inhibition of gene expression. In contrast to other HDACs, class-IIa HDACs function as signal-responsive transcriptional corepressors for the myocyte enhancer factor-2 (MEF2) family of transcription factors. MEF2s play important roles in regulating growth factor responses, neuronal survival, T-cell apoptosis and, importantly, B-cell DNA double strand break (DSB) repair. Here we ask whether inhibition of SIK2 can enhance HDAC class-IIa inhibition of MEF activity, decrease DNA repair and enhance sensitivity to the PARP inhibitor Olaparib in ovarian cancer cells. Treatment with SIK2 inhibitors (ARN3236 or ARN3261) enhanced sensitivity to Olaparib in each of 8 ovarian cancer cell lines without BRCA1/2 mutations (OC316, OVCAR8, IGROV1, A2780, HCC5030, HCC5032, SKOv3 and OVCAR5) and significantly reduced the IC50 of Olaparib in two PARP inhibitor-resistant ovarian cancer cell lines (A2780CP-R and UWB1-289-10R). Synergistic cytotoxicity, judged with a Chou-Talaylay combination index (CI), was observed in all 8 cell lines. Treatment with a SIK2 inhibitor decreased the phosphorylation of class-IIa HDAC4/5/7, abolished class-IIa HDAC4/5/7-associated transcriptional activity of MEF2 and decreased MEF2 binding to regulatory regions with high-chromatin accessibility in DNA repair genes, resulting in repression of critical gene expression in DNA repair pathways. DNA damage, judged by rH2AX expression was enhanced by SIK2 inhibition. These observations not only provide new insights into the transcriptional regulation of DNA repair gene expression, but also have important implications for enhancing sensitivity of high grade serous ovarian cancers to PARP inhibition, with or without BRCA1 or BRCA2 germline mutations. Citation Format: Zhen Lu, Wequn Mao, Lan Pang, Janice M. Santiago-O'Farrill, Haling Yang, Ahmed Ahmed, Hariprasad Vankayalapati, Robert C. Bast. SIK2 inhibitors regulate DNA repair pathway and sensitize ovarian cancer to PARP1 inhibitors [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2018; 2018 Apr 14-18; Chicago, IL. Philadelphia (PA): AACR; Cancer Res 2018;78(13 Suppl):Abstract nr 324.

Telomere shortening associated with increased levels of oxidative stress in sulfur mustard-exposed Iranian veterans

Sulfur Mustard (SM) is the most widely used chemical weapon. It was used in World War 1 and in the more recent Iran-Iraq conflict. Genetic toxicity and DNA alkylation effects of SM in molecular and animal experiments are well documented. In this study, lymphocytic telomere lengths and serum levels of isoprostane F2α were measured using q-PCR and enzyme immunoassay-based methods in 40 Iranian veterans who had been exposed to SM between 1983-88 and 40 non-exposed healthy volunteers. The relative telomere length in SM-exposed individuals was found to be significantly shorter than the non-exposed individuals. In addition, the level of 8-isoprostane F2α was significantly higher in the SM-exposed group compared to controls. Oxidative stress can be caused by defective antioxidant responses following gene mutations or altered activities of antioxidant enzymes. Chronic respiratory diseases and infections may also increaseoxidative stress. The novel finding of this study was a the identification of ‘premature ageing phenotype’. More specifically, telomere shortening which occurs naturally with aging is accelerated in SM-exposed individuals. Oxidative stress, mutations in DNA repair genes and epimutaions may be among the major mechanisms of telomere attrition. These findings may help for a novel therapeutic strategy by telomere elongation or for validation of an exposure biomarker for SM toxicity.

Germline mutations in DNA repair genes are associated with bladder cancer risk and unfavourable prognosis.

To perform a systematic evaluation of whether germline DNA repair gene mutations in bladder cancer (BCa) are associated with increased risk of BCa and aggressive disease. Germline DNA from 98 patients with BCa was analysed for 54 DNA repair genes using a customized targeted sequencing panel. Population control data were obtained from the public databases the Exome Aggregation Consortium database and the Genome Aggregation Database. Mutation pathogenicity was annotated based on American College of Medical Genetics criteria, mutation frequencies in the general population and the ClinVar database. Mutation frequencies were compared based on case-control and case-case designs for disease risks, disease aggressiveness and outcomes. The frequency of pathogenic/likely pathogenic germlineDNA repair gene mutations was 10.2% among patients with BCa. Within the subset of patients withcarcinoma invading the bladder muscle, the frequency was 15.8%, ~2.4-fold higher than in patients with non-muscle invasive BCa (6.67%). The mutation frequencyamong patients with early-onset disease (at age <45years) was ~3-fold higher than among those diagnosedafter age 45years (28.57% vs 8.79%). Mutation carriers had a significantly higher frequency of unfavourable clinical outcomes (disease recurrence or progression to metastatic BCa) than non-carriers (50.0% vs 13.64%; P = 0.013). Pathogenic and likely pathogenic mutations in DNA repair genes were associated with unfavourable prognosis of BCa.

Inherited predisposition to malignant mesothelioma (MM) due to mutations in DNA repair genes.

8504Background: Identifying the profile of DNA repair genes predisposing to MM will enable treatment options for patients (pts) and risk assessment for their families. Methods: In this prospective study of the natural history of MM (NCT01950572) we enrolled 239 consecutive pts independent of site of disease, family history of cancer, age at diagnosis, ethnicity, or asbestos exposure. Germline DNA was sequenced for all 239 pts, identifying mutations of all classes in 73 DNA repair genes. Tumor DNA from 12 pts with germline BAP1 mutations was evaluated by whole exome sequencing. Results: Of the 239 pts, 29 (12%) carried a pathogenic germline mutation in a DNA repair gene: BAP1 (N = 17 pts), CHEK2 (N = 5), PALB2 (N = 2), and BRCA2, MLH1, POT1, TP53, and MRE11A (N = 1 each). Pts with mutations were more likely to be female (P = 0.02) and to have been diagnosed with another cancer (P = 0.009). Pts with germline mutations were more likely to have a 1o relative with a diagnosis of MM (P < 0.0001), melanoma (P = ...

Phase II trial of rucaparib (Without ADT) in patients with metastatic hormone-sensitive prostate cancer harboring germline DNA repair gene mutations (TRIUMPH).

TPS5095Background: The clinical activity of PARPi in patients with homologous recombination DNA-repair mutations and metastatic prostate cancer has now been established. Focusing specifically on pa...

The GENTleMEN study: Genetic testing for men with metastatic prostate cancer in Washington state and beyond.

TPS5098Background: Germline DNA repair gene mutations are present in ~10% of patients with metastatic prostate cancer (mPC) irrespective of age at diagnosis or family history, and have potential pr...

Whole genome sequencing data of 1110 Mycobacterium tuberculosis isolates identifies insertions and deletions associated with drug resistance

BackgroundDrug resistance in Mycobacterium tuberculosis (MTB) is one of the major challenges in tuberculosis (TB) treatment. However, known mutations cannot explain all of the cases of resistance and little research has focused on the relationship between insertions / deletions (indels) and drug resistance.ResultsHere, we retrieved whole genome sequencing data of 743 drug-resistant MTB strains and 367 pan-susceptible strains from TB patients from the public domain to identify novel genomic markers of drug resistance. A total of 20 region markers containing genes and intergenic regions (IGRs) with significant statistical correlation with antibiotic resistance were revealed, four of which have been previously reported to be associated with drug resistance. In addition, 83 point markers containing frameshift (FS) mutations and IGR indels were also identified independently based on differences in their incidence rates between drug-sensitive and -resistant strains. Among the 83 point markers, eight indels were detected in known drug-associated genes or IGRs. Furthermore, the overlap between 20 region markers and 83 point markers further indicated their associations with drug resistance. The markers identified were involved in essential bacterial metabolic functions, including cell wall and transmembrane transporter functions. A strong correlation between FS mutations and mutations in DNA repair genes including I21V in alkA, R48G in mutT4 and P2R in nth was also found.ConclusionsThis study identified a set of novel genetic markers with FS mutations and IGR indels associated with MTB drug resistance, which greatly broadens the pool of mutations related to MTB drug resistance. This insight may be important in identifying novel mechanisms of drug resistance in MTB.

Primary Mutational Landscape Linked with Pre-Docetaxel Lactate Dehydrogenase Levels Predicts Docetaxel Response in Metastatic Castrate-Resistant Prostate Cancer

BackgroundDocetaxel chemotherapy is a standard of care for metastatic castrate-resistant prostate cancer (mCRPC): 40–50% of patients achieve a biochemical response. However, there is a lack of response predictive biomarkers. ObjectiveTo assess lactate dehydrogenase (LDH) as a docetaxel response biomarker in mCRPC and to examine the association of LDH with genomic alterations in primary diagnostic biopsies. Design, setting, and participantsClinical and associated primary tumour-targeted next-generation sequencing data from matched training (n=150) and test (n=120) cohorts of progressive mCRPC patients receiving docetaxel therapy were analysed. Data were correlated with large-scale prostate cancer genomic datasets. Outcome measurements and statistical analysisProstate-specific antigen (PSA) response, radiographic response, biochemical progression-free survival (PFS), overall survival (OS), genomic analysis of primary biopsies, and genomic datasets (Memorial Sloan Kettering Cancer Center [MSKCC] and SU2C/PCF). Results and limitationsSerum LDH ≥450U/l is a reliable prognostic biomarker (area under the curve: 0.757 [standard deviation 0.054, 95% confidence interval [CI] 0.650–0.864, p<0.001]) in progressive mCRPC, predicting PFS at 3 mo. Patients with LDH ≥450U/l were poorer PSA responders, with shorter PFS (213 vs 372 d, hazard ratio [HR] 1.876, 95% CI 1.289–2.7300) and OS (362 vs 563 d, HR 1.630, 95% CI 1.127–2.357). High LDH is an independent surrogate marker for survival following docetaxel and predicts a poor radiological response (p=0.043). Of the 14 patients with LDH ≥450U/l available for next-generation sequencing, nine (64.3%) were more likely to have DNA repair gene mutation(s) (BRCA1/2, ATM, CHEK2, Fanconi anaemia gene) in their primary biopsy. Cross correlation with MSKCC and SU2C/PCF databases revealed a positive correlation between LDHA, PARP1 (r=0.667, p<0.01), and other DNA repair genes. ConclusionsGenomic abnormalities of LDHA and DNA repair in primary biopsies link to high pretreatment LDH and poor response to docetaxel in mCRPC. Patient summaryThe presence of mutations of the lactate dehydrogenase and DNA repair pathways are associated with aggressive prostate cancer and poor response to chemotherapy later in the disease.

Rare DNA repair gene mutations predispose to young onset and lethal prostate cancer in the UK

Background: Prostate cancer (PrCa) is the most common solid tumour in men in the Western world. There is evidence that PrCa predisposition is due both to common and rare germline variation.

Inherited DNA-Repair Defects in Colorectal Cancer

Colorectal cancer (CRC) heritability has been estimated to be around 30%. However, mutations in the known CRC-susceptibility genes explain CRC risk in fewer than 10% of affected individuals. Germline mutations in DNA-repair genes (DRGs) have recently been reported in CRC, but their contribution to CRC risk is largely unknown. We evaluated the gene-level germline mutation enrichment of 40 DRGs in 680 unselected CRC individuals and 27,728 ancestry-matched cancer-free adults. Significant findings were then examined in independent cohorts of 1,661 unselected CRC individuals and 1,456 individuals with early-onset CRC. Of the 680 individuals in the discovery set, 31 (4.56%) individuals harbored germline pathogenic mutations in known CRC-susceptibility genes, and another 33 (4.85%) individuals had DRG mutations that have not been previously associated with CRC risk. Germline pathogenic mutations in ATM and PALB2 were enriched in both the discovery (OR = 2.81 and p = 0.035 for ATM and OR = 4.91 and p = 0.024 for PALB2) and validation (OR = 2.97 and adjusted p = 0.0013 for ATM and OR = 3.42 and adjusted p = 0.034 for PALB2) sets. Biallelic loss of ATM was evident in all individuals with matched tumor profiling. CRC individuals also had higher rates of actionable mutations in the HR pathway, which can substantially increase the risk of developing cancers other than CRC. Our analysis provides evidence for ATM and PALB2 as CRC-risk genes, underscoring the importance of the homologous recombination pathway in CRC. In addition, we identified frequent complete homologous recombination deficiency in CRC tumors, representing a unique opportunity to explore targeted therapeutic interventions such as poly-ADP ribose polymerase inhibitor (PARPi).

Comparative assessment of abiraterone or enzalutamide activity in the PROREPAIR-B study.

164 Background: Germline mutations in DNA repair genes have been associated with poor prostate cancer outcomes in retrospectives studies. Such defects have been identified in 12% of mCRPC patients. Several studies are ongoing to assess the benefit of these patients from platinum-based chemotherapy and PARP inhibitors, but no conclusive data are available with regards to currently approved therapies for mCRPC, as Abiraterone or Enzalutamide. Methods: PROREPAIR-B (NCT03075735) is a prospective multicentre observational cohort study. Patients diagnosed with mCRPC, with unknown mutational status at study entry and who were going to start a first-line treatment for mCRPC were eligible. For this sub-analysis patients who received Abiraterone or Enzalutamide as first androgen receptor targeted therapy (ART) were selected. The endpoints of this sub-analysis included to assess the impact of BRCA1, BRCA2, ATM, PALB2 and other germline mutations in DNA repair genes on cause-specific survival (CSS), progression-free survival (PFS), time to PSA progression (bPFS) and response to the first ART received as 1st or 2nd line therapy. Results: 337 patients were eligible for this analysis. CSS from mCRPC was not significantly different between gDDR carriers and non-carriers. However, CSS from mCRPC in BRCA2 carriers was significantly shorter than in non-carriers (23.3 Vs 34.6 months, p = 0.02). CSS from first ART, PFS and response-rates were not significantly different between both groups. However, the bPFS was significantly shorter in patients harbouring gDDR mutations (7.3 Vs 3.8 months, p = 0.04), especially in BRCA2 carriers (7.3 Vs 3.0 months, p = 0.03). Conclusions: This is the first study to prospectively follow-up DNA repair germline mutations to determine the outcome on standard treatment for mCRPC. The results suggest that different gDDR defects may have different impact on mCRPC outcomes. Clinical trial information: NCT03075735.

Correlation of primary biopsy mutational landscape to pre-docetaxel LDH for prediction of docetaxel therapeutic response.

220 Background: There is a lack of biomarkers to predict docetaxel response. Here we assess the potential of pre-treatment LDH, associated to the mutational background of the primary biopsy taken prior to 1st line therapy with ADT, to predict docetaxel response in the castrate resistant setting. Methods: Clinical and associated primary biopsy targeted next-generation sequencing data from matched training (n = 150) and test (n = 120) sets of retrospective progressive mCRPC patients (diagnosed 1996-2006) treated with docetaxel therapy were analyzed. LDH thresholds were determined and validated on the training and test sets respectively. Clinico-pathological associations with LDH were assessed using student t-test, Wilcoxon test and Chi-square test and predictive value of LDH determined by linear regression analysis for PSA and radiologic response. Time to event outcomes (PFS &amp; OS) was evaluated by Kaplan-Meier. Meta-analysis of LDH association with DNA repair genes in the MSKCC and SU2C/PCF was conducted using cBioPortal Results: Using pre-docetaxel serum LDH of ≥450U/L as a prognostic threshold (AUC:0.75 (SD 0.054, 95% CI, 0.650–0.864, P &lt; 0.001)) patients with LDH ≥450U/L were poorer PSA responders, with shorter PFS (213 vs 372 days, HR 1.876 (95% CI, 1.289-2.730)) and OS (362 vs 563 days, HR 1.630 (95% CI, 1.127-2.357)). LDH was also an independent surrogate marker for prediction of poor radiologic response and survival following docetaxel chemotherapy (P = 0.043). Genomic analysis showed that patients with LDH ≥450U/L (9/14 (64.3%)) have a higher tumour mutational burden (TMB) at the point of primary pre-ADT biopsy (mean, 46.2 vs 6.2 mutations/MB; p = 0.129) and that they were more likely to have DNA repair gene mutation(s) (BRCA1/2, ATM, CHEK2 and Fanconi’s anemia gene). Meta-analysis of the large MSKCC and the SU2C-PCF patient genomic databases confirmed the positive correlation between the LDH gene LDHA and PARP1 gene (r = 0.667, p &lt; 0.01) expression and expression of other DNA repair genes. Conclusions: High pre-treatment serum LDH coupled with mutations in the DNA repair pathway in primary pre-ADT biopsy correlates with poor docetaxel response and overall survival in the castrate resistant state.

Effect of germline DNA repair gene mutations on outcomes in men with metastatic castration-resistant prostate cancer receiving first-line abiraterone and enzalutamide.

221 Background: Inherited DNA repair gene mutations are more prevalent in men with advanced prostate cancer than previously thought, but their clinical implications are not fully understood. Here we investigated the clinical significance of germline DNA repair gene alterations in men with metastatic castration-resistant prostate cancer (mCRPC) receiving first-line novel hormonal therapy (NHT), with a particular emphasis on BRCA1/2 and ATM mutations. Methods: We interrogated 50 DNA repair genes for pathogenic or likely pathogenic germline mutations using leukocyte DNA from 172 mCRPC patients beginning treatment with first-line NHT: abiraterone or enzalutamide. We assessed the impact of germline DNA repair gene mutation status on ≥50% and ≥90% PSA response rates, PSA progression-free survival (PSA-PFS), clinical/radiologic progression-free survival (PFS), and overall survival (OS). Outcomes were adjusted using propensity score-weighted multivariable Cox regression analyses. Results: Among 172 mCRPC patients, germline mutations (in any DNA repair gene) were found in 12.8% (22/172) of men, and germline BRCA/ATM mutations were found in 5.2% (9/172) of men. In unadjusted analyses, outcomes to first-line NHT were better in men with germline BRCA/ATM mutations (vs. no mutations) with respect to ≥90% PSA responses (78% vs. 28%, P = 0.004), PSA-PFS (HR 0.47, P = 0.061), PFS (HR 0.50, P = 0.090) and OS (HR 0.28, P = 0.059). In propensity score-weighted multivariable analyses, outcomes remained superior in men with germline BRCA/ATM mutations with respect to PSA-PFS (HR 0.48, 95%CI 0.25–0.92, P = 0.027), PFS (HR 0.52, 95%CI 0.28–0.98, P = 0.044) and OS (HR 0.34, 95%CI 0.12–0.99, P = 0.048), but this was not true for men with non- BRCA/ATM germline mutations (all endpoints, P &gt; 0.10). Conclusions: Outcomes to first line NHT appeared better in mCRPC patients harboring germline BRCA/ATM mutations (vs. no mutations), but not for patients with other non- BRCA/ATM germline mutations. These results support the hypothesis that AR may promote DNA repair, and that inhibiting AR in the context of homologous recombination deficiency may lead to synthetic lethality.

New Insights into the Molecular Characteristics of Pulmonary Carcinoids and Large Cell Neuroendocrine Carcinomas, and the Impact on Their Clinical Management.

Carcinoids and large cell neuroendocrine carcinomas (LCNECs) are rare neuroendocrine lung tumors. Here we provide an overview of the most updated data on the molecular characteristics of these diseases. Recent genomic studies showed that carcinoids generally contain a low mutational burden and few recurrently mutated genes. Most of the reported mutations occur in chromatin-remodeling genes (e.g., menin 1 gene [MEN1]), and few affect genes of the phosphoinositide 3-kinase (PI3K)-AKT-mechanistic target of rapamycin gene pathway. Aggressive disease has been related to chromothripsis, DNA-repair gene mutations, loss of orthopedia homeobox/CD44, and upregulation of ret proto-oncogene gene (RET) gene expression. In the case of LCNECs, which present with a high mutation burden, two major molecular subtypes have been identified: one with biallelic inactivation of tumor protein p53 gene (TP53) and retinoblastoma gene (RB1), a hallmark of SCLC; and the other one with biallelic inactivation of TP53 and serine/threonine kinase 11 gene (STK11)/kelch like ECH associated protein 1 gene (KEAP1), genes that are frequently mutated in NSCLC. These data, together with the identification of common mutations in the different components of combined LCNEC tumors, provide further evidence of the close molecular relation of LCNEC with other lung tumor types. In terms of therapeutic options, future studies should explore the association between mechanistic target of rapamycin pathway mutations and response to mechanistic target of rapamycin inhibitors in carcinoids. For LCNEC, preliminary data suggest that the two molecular subtypes might have a predictive value for chemotherapy response, but this observation needs to be validated in randomized prospective clinical trials. Finally, delta like Notch canonical ligand 3 inhibitors and immunotherapy may provide alternative options for patient-tailored therapy in LCNEC.

Germline DNA-repair Gene Mutations and Outcomes in Men with Metastatic Castration-resistant Prostate Cancer Receiving First-line Abiraterone and Enzalutamide

BackgroundInherited DNA-repair gene mutations are more prevalent in men with advanced prostate cancer than previously thought, but their clinical implications are not fully understood. ObjectiveTo investigate the clinical significance of germline DNA-repair gene alterations in men with metastatic castration-resistant prostate cancer (mCRPC) receiving next-generation hormonal therapy (NHT), with a particular emphasis on BRCA/ATM mutations. Design, setting, and participantsWe interrogated 50 genes for pathogenic or likely pathogenic germline mutations using leukocyte DNA from 172 mCRPC patients beginning treatment with first-line NHT with abiraterone or enzalutamide. Outcome measurements and statistical analysisWe assessed the impact of germline DNA-repair gene mutation status on ≥50% and ≥90% PSA responses, PSA progression-free survival (PSA-PFS), clinical/radiologic progression-free survival (PFS), and overall survival (OS). Survival outcomes were adjusted using propensity score-weighted multivariable Cox regression analyses. Results and limitationsAmong 172 mCRPC patients included, germline mutations (in any DNA-repair gene) were found in 12% (22/172) of men, and germline BRCA/ATM mutations specifically in 5% (9/172) of men. In unadjusted analyses, outcomes to first-line NHT were better in men with germline BRCA/ATM mutations (vs no mutations) with respect to PSA-PFS (hazard ratio [HR] 0.47; p=0.061), PFS (HR 0.50; p=0.090), and OS (HR 0.28; p=0.059). In propensity score-weighted multivariable analyses, outcomes were superior in men with germline BRCA/ATM mutations with respect to PSA-PFS (HR 0.48, 95% confidence interval [CI] 0.25–0.92; p=0.027), PFS (HR 0.52, 95% CI 0.28–0.98; p=0.044), and OS (HR 0.34, 95% CI 0.12–0.99; p=0.048), but not in men with non-BRCA/ATM germline mutations (all p>0.10). These results require prospective validation, and our conclusions are limited by the small number of patients (n=9) with BRCA/ATM mutations. ConclusionsOutcomes to first-line NHT appear better in mCRPC patients harboring germline BRCA/ATM mutations (vs no mutations), but not for patients with other non-BRCA/ATM germline mutations. Patient summaryPatients with metastatic castration-resistant prostate cancer and harboring germline mutations in BRCA1/2 and ATM benefit from treatment with abiraterone and enzalutamide.

Association of DNA repair gene-mutation, mutation burden and, neoantigen load in breast cancer.

9 Background: Anti-cancer therapy with immune checkpoint inhibitors demonstrated high efficacy in hypermutated tumors; Food and Drug Administration recently proved PD1/PDL-1 blockade therapy in solid tumors characterized with deficiency of mismatch repair (MMR). However, several malignancies contain less feature of MMR deficiency. Except MMR, it remains uncertain whether mutations of other DNA repair genes are associated with hypermutated status. We aim to analyze the relationship between DNA repair gene-mutation statuses, mutation burden and neoantigen load in breast cancer. Methods: Whole exome sequencing was performed on germline and tumor DNA in 82 patients with early-onset breast cancer (≤ 40 year-old). Calculation of mutation burden includes tumor-specific insertion/deletion (Indel) and single nucleotide variants. Missense mutations, frameshift mutations and uncorrected splicing were considered to generate new peptides (neopeptides). Binding affinity of neopeptide-major histocompatibility complex class I and their immunogenicity were predicted by NetMHC 4.0 and POPI 2.0. Results: A total of 18 pathogenic mutations were found, 4 in ATM (2 germline and 2 somatic), 3 in BRCA1 (germline), 4 in BRCA2 (germline), 1 in BRIP1 (germline), 1 in CHK2 (germline), 1 in ERCC8 (germline), 2 in MSH2 (1 germline and 1 somatic), 1 in RAD51C (somatic) and 1 in OGG1 (germline). The median number of exomic mutations (missense mutations and Indels) were 744 (min. 355; max. 3587) variants per sample. Tumors with mutations of homologous recombination (HR) and MMR genes contained a higher trend of exomic mutations than those without mutation. However, tumors with mutations of HR and MMR genes contained significantly higher numbers of neopeptides with high binding affinity and immunogenicity than those without mutation (p = 0.04). Conclusions: Tumors with mutations of HR and MMR genes contained higher numbers of neoantigens. This study showed that HR and MMR mutation may be a biomarker predicted clinical benefit of immune checkpoint blockade therapy in breast cancer.

High throughput gene sequencing reveals altered landscape in DNA damage responses and chromatin remodeling in sporadic pancreatic neuroendocrine tumors

PurposeThe main objectives of this retrospective study were to survey the genetic landscape of PNETs in a clinical cohort by using the high throughput gene sequencing method and to determine cellular signaling networks affected by the uncovered gene mutations. Materials and methodsWe retrieved the demographics and tumor characteristics of 13 patients. Cellular DNA was extracted from fresh snap frozen tumor tissues and was subject to high throughput gene sequencing analysis using the Illumina NextSeq500 System. Furthermore, the interaction network was constructed from the input gene set by Reactome and performed gene set enrichment analysis was performed with a cutoff FDR of ≤0.01. ResultsTotally 74 mutated genes and 93 mutations were identified. The median number of mutations was 7 (range 1–20) and that of mutated genes was 6 (range 1–17). Among these mutations, 48 (51.6%) were substitution mutations, nine (9.7%) were duplication mutations, 28 (30.1%) were deletion mutations and eight (8.6%) were deletion/insertion mutations. Gene set enrichment analysis generated a network of 21 interactions, 10 of which were associated with DNA repair like the Fanconi anemia pathway, nucleotide excision repair, and homologous recombination repair, or chromosome maintenance. Moreover, 9 patients had one or more mutations in DNA repair genes including the mismatch repair genes MSH2/MSH6. ConclusionsThe study has uncovered genetic alterations of genes implicated in DNA damage responses and chromatin remodeling. Our findings will prompt further studies into the role of these mutated genes in the oncogenesis and molecular stratification of PNETs.

Intraductal/ductal histology and lymphovascular invasion are associated with germline DNA-repair gene mutations in prostate cancer.

Germline mutations in genes mediating DNA repair are common in men with recurrent and advanced prostate cancer, and their presence may alter prognosis and management. We aimed to define pathological and clinical characteristics associated with germline DNA-repair gene mutations, to facilitate selection of patients for germline testing. We retrospectively evaluated 150 unselected patients with recurrent or metastatic prostate cancer who were offered germline genetic testing by a single oncologist using a clinical-grade assay (Color Genomics). This platform utilizes next-generation sequencing from saliva to interrogate 30 cancer-susceptibility genes. Presence or absence of a deleterious germline mutation was correlated with histological and clinical characteristics, and with family history of cancer. All patients with DNA-sequence alterations (pathogenic or variants) were offered genetic counseling. Between July 2016 and July 2017, 150 consecutive patients underwent germline testing; pathogenic mutations were identified in 21 men (14%). Among those with germline mutations, 9 (43%) were in BRCA2, 3 (14%) were in ATM, 3 (14%) were in CHEK2, and 2 (9%) were in BRCA1. While there were no associations between germline mutations and age, tumor stage, Gleason sum or family history; mutation-positive patients had lower median PSA levels at diagnosis (5.5 vs 8.6 ng/mL, P = 0.01) and unique pathologic features. Namely, men with germline mutations were more likely to harbor intraductal/ductal histology (48% vs 12%, P < 0.01) and lymphovascular invasion (52% vs 14%, P < 0.01). Finally, 44% of patients with a positive germline test would not have been offered genetic screening according to current National Comprehensive Cancer Network (NCCN) guidelines. Presence of intraductal/ductal histology and lymphovascular invasion appear to be associated with pathogenic germline DNA-repair gene mutations in men with prostate cancer, and identification of these features may help to select patients for germline testing. NCCN guidelines may be inadequate in predicting which prostate cancer patients should undergo genetic screening.

Mutations in DNA repair genes are associated with increased neo-antigen load and activated T cell infiltration in lung adenocarcinoma.

Mutations in DNA repair genes lead to increased genomic instability and mutation frequency. These mutations represent potential biomarkers for cancer immunotherapy efficacy, as high tumor mutational burden has been associated with increased neo-antigens and tumor infiltrating lymphocytes. While mismatch repair mutations have successfully predicted response to anti-PD-1 therapy in colorectal and other cancers, they have not yet been tested for lung cancer, and few have investigated genes from other DNA repair pathways. We utilized TCGA samples to comprehensively immunophenotype lung tumors and analyze the links between DNA repair mutations, neo-antigen and total mutational burden, and tumor immune infiltration. Overall, 73% of lung tumors contained infiltration by at least one T cell subset, with high mutational burden tumors containing significantly increased infiltration by activated CD4 and CD8 T cells. Further, mutations in mismatch repair genes, homologous recombination genes, or POLE accurately predicted increased tumor mutational burden, neo-antigen load, and T cell infiltration. Finally, neo-antigen load correlated with expression of M1-polarized macrophage genes, PD-1, PD-L1, IFNγ, GZMB, and FASLG, among other immune-related genes. Overall, after defining the immune infiltrate in lung tumors, we demonstrate the potential value of utilizing gene mutations from multiple DNA repair pathways as biomarkers for lung cancer immunotherapy.

Clinical characteristics and outcomes of patients with BRCA1 or RAD51C methylated versus mutated ovarian carcinoma

ObjectiveIn ovarian carcinoma, mutations in homologous recombination DNA repair (HRR) genes, including BRCA1 and RAD51C, are associated with increased survival and specific clinical features. Promoter hypermethylation is another mechanism of reducing gene expression. We assessed whether BRCA1 and RAD51C promoter hypermethylation is associated with similar survival and clinical characteristics. MethodsPromoter methylation of BRCA1 and RAD51C was evaluated using methylation-sensitive PCR in 332 primary ovarian carcinomas. Damaging germline and somatic mutations in 16 HRR genes were identified using BROCA sequencing. ResultsBRCA1 methylation was detected in 22 carcinomas (6.6%) and RAD51C methylation in 9 carcinomas (2.7%). These small numbers limited the power to detect differences in survival and platinum sensitivity. Mutations in one or more HRR genes were found in 95 carcinomas (29%). Methylation of BRCA1 or RAD51C was mutually exclusive with mutations in these genes (P=0.001). Patients whose carcinomas had BRCA1 methylation (57.7years±2.5) or BRCA1 mutations (54.1years±1.4) were younger than those without (63.3years±0.8; P=0.029, P<0.0001). BRCA1 methylation and germline BRCA1 mutation were associated with high grade serous (HGS) histology (P=0.045, P=0.001). BRCA1 mutations were associated with increased sensitivity to platinum chemotherapy while BRCA1 methylation was not (P=0.034, P=0.803). Unlike HRR mutations, methylation was not associated with improved overall survival compared to cases without methylation or mutation. ConclusionsPatients with BRCA1-methylated carcinomas share clinical characteristics with patients with BRCA1-mutated carcinomas including younger age and predominantly HGS histology. However, unlike mutation, RAD51C and BRCA1 methylation were not associated with improved survival or greater sensitivity to platinum chemotherapy.