DNA Repair Variants Research Articles

Germline DNA damage repair variants and prognosis of patients with high-risk or metastatic prostate cancer.

Deleterious germline variants in certain DNA repair genes are risk factors for developing aggressive prostate cancer. The objective was to quantify their prognostic impact after prostate cancer diagnosis. Men with prostate cancer, predominantly of European ancestry, were included from four cohorts with long-term follow-up. Pathogenic or likely pathogenic germline variants in 26 DNA repair genes were assessed in relation to metastasis-free survival in high-risk, localized prostate cancer and to overall survival in metastatic castration-sensitive prostate cancer (mCSPC) and metastatic castration-resistant prostate cancer (mCRPC). Among 3,525 patients initially diagnosed with non-metastatic prostate cancer, 2,594 had high-risk localized prostate cancer, 429 mCSPC, and 502 mCRPC at inclusion. BRCA2 variant carriers did not have worse metastasis-free survival in high-risk localized prostate cancer (hazard ratio [HR] 1.01, 95% confidence interval [CI] 0.69, 1.46) or overall survival in mCSPC (HR 0.46, 95% CI 0.14, 1.45) or mCRPC (HR 0.60, 95% CI 0.31, 1.17) compared to non-carriers of DNA repair variants. Among 868 additional patients with de novo metastatic (M1) prostate cancer, BRCA2 variant carriers tended to have worse overall survival (HR 1.59, 95% CI 1.01, 2.51). BRCA2 prognostic associations were not explained by radiation, PARP inhibitor, or platinum therapy. Results for other genes were limited in precision because variants were less common. Among patients with high-risk or metastatic prostate cancer who were initially diagnosed with and treated for non-metastatic tumors, germline DNA repair variants in BRCA2 do not confer a substantially worse prognosis.

DNA damage and its association with early-life exposome: Gene-environment analysis in Colombian children under five years old

Environmental exposures and gene-exposure interactions are the major causes of some diseases. Early-life exposome studies are needed to elucidate the role of environmental exposures and their complex interactions with biological mechanisms involved in childhood health. This study aimed to determine the contribution of early-life exposome to DNA damage and the modifying effect of genetic polymorphisms involved in air pollutants metabolism, antioxidant defense, and DNA repair. We conducted a cohort study in 416 Colombian children under five years. Blood samples at baseline were collected to measure DNA damage by the Comet assay and to determine GSTT1, GSTM1, CYP1A1, H2AX, OGG1, and SOD2 genetic polymorphisms. The exposome was estimated using geographic information systems, remote sensing, LUR models, and questionnaires. The association exposome-DNA damage was estimated using the Elastic Net linear regression with log link. Our results suggest that exposure to PM2.5 one year before the blood draw (BBD) (0.83, 95 %CI: 0.76; 0.91), soft drinks consumption (0.94, 0.89; 0.98), and GSTM1 null genotype (0.05, 0.01; 0.36) diminished the DNA damage, whereas exposure to PM2.5 one-week BBD (1.18, 1.06; 1.32), NO2 lag-5 days BBD (1.27, 1.18; 1.36), in-house cockroaches (1.10, 1.00; 1.21) at the recruitment, crowding at home (1.34, 1.08; 1.67) at the recruitment, cereal consumption (1.11, 1.04; 1.19) and H2AX (AG/GG vs. AA) (1.44, 1.11; 1.88) increased the DNA damage. The interactions between H2AX (AG/GG vs. AA) genotypes with crowding and PM2.5 one week BBD, GSTM1 (null vs. present) with humidity at the first year of life, and OGG1 (SC/CC vs. SS) with walkability at the first year of life were significant. The early-life exposome contributes to elucidating the effect of environmental exposures on DNA damage in Colombian children under five years old. The exposome-DNA damage effect appears to be modulated by genetic variants in DNA repair and antioxidant defense enzymes.

Puzzling phenomenon: adult-onset cancer predisposition and pediatric cancer

Pathogenic variants (PVs) in DNA repair–linked adult-onset cancer predisposition genes, including double heterozygosity, are increasingly identified in pediatric patients with cancer. Their role in childhood cancer, however, remains poorly understood. Integrating comprehensive tumor analysis is integral for understanding the contribution of such PVs in cancer development and personalized cancer care.

Prevalence of single-nucleotide variants in twenty-five pharmacogenes from a Cuban sample cohort.

The Cuban population is genetically diverse, and information on the prevalence of genetic variants is still limited. As complex admixture processes have occurred, we hypothesized that the frequency of pharmacogenetic variants and drug responses may vary within the country. The aims of the study were to describe the frequency distribution of 43 single-nucleotide variants (SNVs) from 25 genes of pharmacogenetic interest within the Cuba population and in relation to other populations, while taking into consideration some descriptive variables such as place of birth and skin color. SNVs were analyzed in 357 unrelated healthy Cuban volunteers. Genotype, allele frequencies, and ancestry proportions were determined, and the pairwise fixation index (FST ) was evaluated. Hardy-Weinberg equilibrium (HWE) deviations in six loci (rs11572103, rs2740574, rs776746, rs3025039, rs861539, and rs1762429) were identified. Minor allele frequencies (MAFs) ranged from 0.00 to 0.15 for variants in genes encoding xenobiotic metabolizing enzymes. They also ranged from 0.01 to 0.21 for variants in DNA repair, growth factors, methyltransferase, and methyl-binding proteins, while they ranged from 0.04 to 0.27 for variants in the O-6-methylguanine-DNA methyltransferase enzyme. Moderate genetic divergence was observed upon comparison to Africans (FST = 0.071 and SD 0.079), with 19 markers exhibiting moderate-to-large genetic differentiation. The average European, African, and Amerindian ancestry proportions were 67.8%, 27.2%, and 5.3%, respectively. Ancestry proportions differed by skin color and birthplace for both African and European components, with the exception of the European component, which showed no significant difference between individuals from Western and Eastern regions. Meanwhile, the statistical significance varied in comparisons by skin color and birthplace within the Amerindian component. Low genetic divergence was observed across geographical regions. We identified 12 variants showing moderate-to-large differentiation between White/Black individuals. Altogether, our results may support national strategies for the introduction of pharmacogenetic tools in clinical practice, contributing to the development of precision medicine in Cuba.

GSTP , GSTT1 , XRCC1 and CASP8 Genetic Variations are Associated with Human Papillomavirus in Women with Cervical Cancer From Zimbabwe

Aim: Investigate the role of host genetic variations in high-risk human papillomaviruses (HR-HPVs). Methods: This cross-sectional study recruited 238 cervical cancer patients. Variants in transport (ABCC2), xenobiotic metabolism ( GSTP, GSTT1, GSTM1, NQO1), DNA repair ( ERCC1, XRCC1), immune response ( TLR4) and apoptosis ( CASP8, FASL, p53) genes were characterized. Tumor DNA was genotyped for 14 HR-HPVs. Results: GSTP rs1695GG, XRCC1 rs1799782TT and GSTT1 del/del were associated with HPV51 (odds ratio [OR]: 3.9; 95% confidence interval [CI]: 1.3–11.7; p = 0.02) and HPV58 (OR: 2.4; 95% CI: 1.2–5.8; p = 0.048), respectively. CASP8 rs3834129del/del was associated with HPV16/18 (OR: 2.7; 95% CI: 1.2–6.0; p = 0.017) and HPV monoinfections (OR: 2.3; 95% CI: 1.2–4.4; p = 0.008). Conclusion: GSTP, GSTT1, XRCC1 and CASP8 variants were associated with HPV-positivity. With further research, a genetic-based screening tool can be developed, to use with HPV vaccines toward preventing cervical cancer.

METB-09. GERMLINE PATHOGENIC VARIANTS IN 838 PEDIATRIC BRAIN TUMOR PATIENTS

Abstract The genetic contribution of rare pathogenic germline variation in cancer patients without a family history remains unclear. We sought to characterize the prevalence, spectrum, and clinical significance of pathogenic germline variation in cancer predisposition genes (CPGs) in pediatric brain tumor patients. Paired tumor and normal whole genome or exome sequencing was performed for 838 patients in the Pediatric Brain Tumor Atlas (PBTA). Rare variants in 196 CPGs were annotated as pathogenic (P) or likely pathogenic (LP) in an automated manner consistent with American College of Medical Genetics criteria and reviewed by an interdisciplinary panel. To assess enrichment, the frequency of CPG P-LP variants in cases was compared to the gnomAD v3.1 cancer-free control cohort (n=74,023). Second hit, mutational signature, and gene set enrichment analyses (GSEA) were performed on matched tumor sequencing to characterize differences in patients with and without germline P-LP variants in DNA repair genes. We observed 178 germline CPG P-LP variants in 163 PBTA patients (19.5%). CPG P-LP variants were most prevalent among Neurofibroma plexiform patients (64.3%), and NF1 and TSC2 harbored the most significant P-LP variant burden compared to controls (OR=69.5, p=2.4E-23, CI=34.6-137 and OR=357, p=1.1E-14, CI=71-3669, respectively). DNA repair gene P-LP variants were enriched in cases compared to controls (OR=4.4, p=8.3E-30). Patients harboring mismatch repair (MMR) gene P-LP variants exhibited significantly higher tumor-associated MMR-deficiency mutational signature exposures (p<0.05) and DNA repair GSEA scores relative to other patients (p=0.04). Second hit SNVs in tumors were identified in four patients with P-LP germline variants in NF1 (n=2), APC (n=1), and SUFU (n=1). Germline P-LP variants are enriched in PBTA patients, and DNA repair variants are predictive of repair-deficiency signatures in tumor samples. Our work emphasizes the need for germline sequencing to inform patient care.

Candidate variants in DNA replication and repair genes in early-onset renal cell carcinoma patients referred for germline testing

BackgroundEarly-onset renal cell carcinoma (eoRCC) is typically associated with pathogenic germline variants (PGVs) in RCC familial syndrome genes. However, most eoRCC patients lack PGVs in familial RCC genes and their genetic risk remains undefined.MethodsHere, we analyzed biospecimens from 22 eoRCC patients that were seen at our institution for genetic counseling and tested negative for PGVs in RCC familial syndrome genes.ResultsAnalysis of whole-exome sequencing (WES) data found enrichment of candidate pathogenic germline variants in DNA repair and replication genes, including multiple DNA polymerases. Induction of DNA damage in peripheral blood monocytes (PBMCs) significantly elevated numbers of gammaH2AX foci, a marker of double-stranded breaks, in PBMCs from eoRCC patients versus PBMCs from matched cancer-free controls. Knockdown of candidate variant genes in Caki RCC cells increased gammaH2AX foci. Immortalized patient-derived B cell lines bearing the candidate variants in DNA polymerase genes (POLD1, POLH, POLE, POLK) had DNA replication defects compared to control cells. Renal tumors carrying these DNA polymerase variants were microsatellite stable but had a high mutational burden. Direct biochemical analysis of the variant Pol δ and Pol η polymerases revealed defective enzymatic activities.ConclusionsTogether, these results suggest that constitutional defects in DNA repair underlie a subset of eoRCC cases. Screening patient lymphocytes to identify these defects may provide insight into mechanisms of carcinogenesis in a subset of genetically undefined eoRCCs. Evaluation of DNA repair defects may also provide insight into the cancer initiation mechanisms for subsets of eoRCCs and lay the foundation for targeting DNA repair vulnerabilities in eoRCC.

Older age should not be a barrier to testing for somatic variants in homologous recombination DNA repair-related genes in patients with high-grade serous ovarian carcinoma

BackgroundSomatic pathogenic variants (PVs) in homologous recombination DNA repair (HR)-related genes found in high-grade serous ovarian carcinomas (HGSC) are not well-characterised in older patients (≥70 years). This may reflect low testing rates in older patients. MethodsData from 1210 HGSC patients in AACR Project GENIE and 324 patients in an independent dataset INOVATe were analysed. Cases where somatic variants could be distinguished from germline variants were included, and analysis was restricted to those with a somatic TP53 variant, to ensure cases were HGSC. ResultsOf 1210 patients in GENIE, 27% (n = 325) were aged ≥70 years at testing. Patients with somatic-only PVs in BRCA2 were older compared with BRCA1 (median 71 vs 60 years, p = 0.002). Median age for 21 patients with somatic-only PVs in 11 other HR-related genes ranged from 40 to 67 years. In older patients, 7% (n = 22) had somatic BRCA1/2 PVs, and 1% (n = 2) had PVs other HR-related genes; this rate was not significantly different to younger patients (<70 years), 7% (n = 62) BRCA1/2 and 2% (n = 19) other HR-related genes (p = 0.36). The overall frequency of somatic BRCA1/2 PVs was similar in INOVATe (n = 25; 7.7%) and somatic-only BRCA2 PVs were again found in older patients compared with BRCA1 (median age: at testing, 70 vs 63 years; at diagnosis, 68 vs 60 years). ConclusionsThe overall frequency of somatic-only PVs in HR-related genes was similar in older and younger patients with HGSC, highlighting the importance of somatic testing irrespective of age. Limiting somatic testing by age may exclude patients who could benefit from maintenance poly(ADP-ribose) polymerase (PARP) inhibitors.

RNA-Based Classification of Homologous Recombination Deficiency in Racially Diverse Patients with Breast Cancer.

Aberrant expression of DNA repair pathways such as homologous recombination (HR) can lead to DNA repair imbalance, genomic instability, and altered chemotherapy response. DNA repair imbalance may predict prognosis, but variation in DNA repair in diverse cohorts of breast cancer patients is understudied. To identify RNA-based patterns of DNA repair expression, we performed unsupervised clustering on 51 DNA repair-related genes in the Cancer Genome Atlas Breast Cancer [TCGA BRCA (n = 1,094)] and Carolina Breast Cancer Study [CBCS (n = 1,461)]. Using published DNA-based HR deficiency (HRD) scores (high-HRD ≥ 42) from TCGA, we trained an RNA-based supervised classifier. Unsupervised and supervised HRD classifiers were evaluated in association with demographics, tumor characteristics, and clinical outcomes. : Unsupervised clustering on DNA repair genes identified four clusters of breast tumors, with one group having high expression of HR genes. Approximately 39.7% of CBCS and 29.3% of TCGA breast tumors had this unsupervised high-HRD (U-HRD) profile. A supervised HRD classifier (S-HRD) trained on TCGA had 84% sensitivity and 73% specificity to detect HRD-high samples. Both U-HRD and S-HRD tumors in CBCS had higher frequency of TP53 mutant-like status (45% and 41% enrichment) and basal-like subtype (63% and 58% enrichment). S-HRD high was more common among black patients. Among chemotherapy-treated participants, recurrence was associated with S-HRD high (HR: 2.38, 95% confidence interval = 1.50-3.78). HRD is associated with poor prognosis and enriched in the tumors of black women. RNA-level indicators of HRD are predictive of breast cancer outcomes in diverse populations.

Human Variation in DNA Repair, Immune Function, and Cancer Risk.

DNA damage constantly threatens genome integrity, and DNA repair deficiency is associated with increased cancer risk. An intuitive and widely accepted explanation for this relationship is that unrepaired DNA damage leads to carcinogenesis due to the accumulation of mutations in somatic cells. But DNA repair also plays key roles in the function of immune cells, and immunodeficiency is an important risk factor for many cancers. Thus, it is possible that emerging links between inter-individual variation in DNA repair capacity and cancer risk are driven, at least in part, by variation in immune function, but this idea is underexplored. In this review we present an overview of the current understanding of the links between cancer risk and both inter-individual variation in DNA repair capacity and inter-individual variation in immune function. We discuss factors that play a role in both types of variability, including age, lifestyle, and environmental exposures. In conclusion, we propose a research paradigm that incorporates functional studies of both genome integrity and the immune system to predict cancer risk and lay the groundwork for personalized prevention.

Cyclin-Dependent Kinase 4/6 Inhibitor Outcomes in Patients With Advanced Breast Cancer Carrying Germline Pathogenic Variants in DNA Repair-Related Genes.

In recent years, unprecedented benefits have been observed with the development of cyclin-dependent kinase (CDK) 4 and 6 inhibitors for the treatment of hormone receptor-positive/human epidermal growth factor receptor 2-negative metastatic breast cancer. However, there is scarce evidence of their value in specific populations, such as patients carrying germline pathogenic variants in DNA repair-related genes. We retrospectively studied the efficacy of CDK 4/6 inhibitors plus endocrine therapy in patients with hormone receptor-positive/human epidermal growth factor receptor 2-negative advanced breast cancer. Three cohorts were compared, including patients harboring germline pathogenic variants in DNA repair-related genes (gBRCA1/2-ATM-CHEK2 mutated), those tested without these mutations (wild type [WT]), and the nontested subgroup. Relevant prognostic factors including age, metastatic site (visceral v nonvisceral), Eastern Cooperative Oncology Group, and prior treatment with CDK 4/6 inhibitors were stratified by univariate and multivariate Cox regression models. Among the total population (n = 217), 15 (6.9%) patients carried gBRCA1/2 (n = 10)-ATM (n = 4)-CHEK2 (n = 1) pathogenic variants, 45 (20.7%) were WT, and 157 (72.4%) were nontested. Gene pathogenic variant carriers were younger (P < .001). Most patients (164, 75.6%) had not received prior endocrine therapy in the advanced setting. Median progression-free survival was shorter in patients with evaluated germline pathogenic variants (10.2 months [95% CI, 5.7 to 14.7]), compared with WT and nontested patients (15.6 months [95% CI, 7.8 to 23.4], and (17.6 months [95% CI, 12.9 to 22.2]; P = .002). Consistently, a worse median overall survival was observed in the subgroup with germline pathogenic variants than in the WT group (P = .006). Multivariable analysis showed that mutation status was an independent prognostic factor of progression-free survival (P = .020) and overall survival (P = .012). In this retrospective real-world study, gBRCA1/2-ATM-CHEK2 pathogenic variants were independently associated with poor outcomes in patients with advanced breast cancer treated with CDK4/6 inhibitors.

Histone variants: The unsung guardians of the genome

Histones H2A, H2B, H3, H4 and H1 are highly conserved, positively charged proteins which form a disc-shaped protein core around which genomic DNA is wrapped to form a nucleosome. Immediately following DNA synthesis, replication-dependent canonical histones help package the DNA into nucleosomes to form compact chromatin fibers that can fit within the confines of the cell nucleus. Histone variants, which vary from the canonical histones in their primary amino acid sequence and expression patterns, replace their canonical counterparts throughout the cell cycle in important biological processes such as transcription, replication, DNA repair and heterochromatin formation. DNA damage is a continual threat to genomic stability and cell survival. Unrepaired DNA lesions are either lethal or can promote mutations if the damaged cells escape programmed cell death due to apoptosis. In order to repair DNA damage, cells use multiple DNA repair pathways, all of which require the recruitment of a multiple DNA damage signaling and repair factors. In order for these repair factors to be recruited efficiently and function properly at sites of DNA damage, the local chromatin environment surrounding the DNA lesion is often altered. Cells are able to regulate chromatin structure in the vicinity of DNA lesions through the addition of posttranslational modifications on histones and DNA, as well as through histone variant incorporation or removal. Recruitment or removal of histone variants at sites of DNA damage can alter the local chromatin structure by destabilizing it and making it more accessible to repair factors. Alternatively, some histone variants and their modifications may also provide specific binding sites for the recruitment of various DNA repair factors, thereby influencing repair pathway choice or repair efficiency, or both. This review seeks to provide an overview of our current understanding of the roles played by histone variants in DNA repair, especially in mammalian cells.

In silico assessment of DNA damage response gene variants associated with head and neck cancer

Head and neck cancer (HNC), the sixth most common cancer globally, stands first in India, especially Northeast India, where tobacco usage is predominant, which introduces various carcinogens leading to malignancies by accumulating DNA damages. Consequently, the present work aimed to predict the impact of significant germline variants in DNA repair and Tumour Suppressor genes on HNC development. WES in Ion ProtonTM platform on ‘discovery set’ (n = 15), followed by recurrence assessment of the observed variants on ‘confirmation set’ (n = 40) using Sanger Sequencing was performed on the HNC-prevalent NE Indian populations. Initially, 53 variants were identified, of which seven HNC-linked DNA damage response gene variants were frequent in the studied populations. Different tools ascertained the biological consequences of these variants, of which the non-coding variants viz. EXO1_rs4150018, RAD52_rs6413436, CHD5_rs2746066, HACE1_rs6918700 showed risk, while FLT3_rs2491227 and BMPR1A_rs7074064 conferred protection against HNC by affecting transcriptional regulation and splicing mechanism. Molecular Dynamics Simulation of the full-length p53 model predicted that the observed coding TP53_rs1042522 variant conferred HNC-risk by altering the structural dynamics of the protein, which displayed difficulty in the transition between active and inactive conformations due to high-energy barrier. Subsequent pathway and gene ontology analysis revealed that EXO1, RAD52 and TP53 variants affected the Double-Strand Break Repair pathway, whereas CHD5 and HACE1 variants inactivated DNA repair cascade, facilitating uncontrolled cell proliferation, impaired apoptosis and malignant transformation. Conversely, FLT3 and BMPR1A variants protected against HNC by controlling tumorigenesis, which requires experimental validation. These findings may serve as prognostic markers for developing preventive measures against HNC.

DNA Repair Genes as Drug Candidates for Early Breast Cancer Onset in Latin America: A Systematic Review.

The prevalence of breast cancer in young women (YWBC) has increased alarmingly. Significant efforts are being made to elucidate the biological mechanisms concerning the development, prognosis, and pathological response in early-onset breast cancer (BC) patients. Dysfunctional DNA repair proteins are implied in BC predisposition, progression, and therapy response, underscoring the need for further analyses on DNA repair genes. Public databases of large patient datasets such as METABRIC, TCGA, COSMIC, and cancer cell lines allow the identification of variants in DNA repair genes and possible precision drug candidates. This study aimed at identifying variants and drug candidates that may benefit Latin American (LA) YWBC. We analyzed pathogenic variants in 90 genes involved in DNA repair in public BC datasets from METABRIC, TCGA, COSMIC, CCLE, and COSMIC Cell Lines Project. Results showed that reported DNA repair germline variants in the LA dataset are underrepresented in large databases, in contrast to other populations. Additionally, only six gene repair variants in women under 50 years old from the study population were reported in BC cell lines. Therefore, there is a need for new approaches to study DNA repair variants reported in young women from LA.

An in silico kinetic model of 8-oxo-7,8-dihydro-2-deoxyguanosine and 8-oxo-7,8-dihydroguanosine metabolism from intracellular formation to urinary excretion

Oxidatively generated DNA damage is of paramount importance in a wide range of physiological and pathophysiological processes. Urinary 8-oxo-7,8-dihydro-2-deoxyguanosine (8-oxodG) is often used as an outcome marker in studies on the role of oxidatively generated DNA damage, but its exact relation to intracellular damage levels and variations in DNA repair have been unclear. Using a new approach of quantitative kinetic modeling inspired by pharmacokinetics, we find evidence that in steady state – i.e. when systemic consequences of given change in damage or cellular removal rates have stabilized - the urinary excretion of 8-oxodG is closely correlated to rates of damage and intracellular 8-oxodG levels, but independent of the rate of cellular removal. Steady state was calculated to occur within approximately 12 h. A similar pattern was observed in a model of the corresponding RNA marker 8-oxo-7,8-dihydroguanosine (8-oxoGuo), but with steady-state occurring slower (up to 5 d). These data have significant implications for the planning of studies and interpretation of data involving urinary 8-oxodG/8-oxoGuo excretion as outcome. Highlights The kinetics of 8-oxodG/8-oxoGuo formation, removal and excretion were simulated in silico. The model was based on existing data on 8-oxodG/8-oxoGuo levels and removal/excretion rates. Intracellular 8-oxodG/8-oxoGuo was closely correlated with urinary excretion in steady state. Changes in removal rates did not influence urinary excretion of 8-oxodG/8-oxoGuo.

Abstract 808: Functional evaluation of novel germline DNA repair variants identified in patients with early-onset renal cancer

Abstract Background: Early-onset renal cancer (eoRC) is typically associated with significant family history of cancer, and often associated with germline pathogenic variants (PVs) in ‘RC-specific' genes including VHL, MET, FLCN, TSC1, TSC2, FH, SDH, PTEN and BAP1. However, most eoRC patients lack PVs in RC-specific genes; rather, we recently identified an enrichment of likely PVs in DNA repair genes in eoRC patients. Here, using eoRC patient resources (primary and transformed lymphocytes, lymphocyte DNA, and tumor tissue), we functionally evaluated the DNA repair variants identified in eoRC patients. Methods: We performed whole-exome sequencing (WES) on lymphocyte DNA from 22 patients with genetically undefined eoRC, with extensive family history of RC and/or other cancers, provided by Risk Assessment Program and Genitourinary Group at Fox Chase Cancer Center. Prioritized variants were assessed by structural, biochemical and cellular studies. Results: 70% of patients had rare, heterozygous, missense variants in genes that suppress DNA damage. ~18% of the variants were in DNA polymerase genes (POLD1, POLE, POLH, POLK), which mediate DNA replication-repair. Increased tumor mutation burden (&amp;gt;10 mutations/Mb) was observed in tumors carrying predicted damaging polymerase variants. In functional testing, induction of DNA damage in primary lymphocytes from eoRC patients and matched cancer-free controls (n=20 each) selectively elevated γH2AX foci (indicative of DNA damage) in cells from eoRC patients (P&amp;lt;0.001, pairwise T-test). DNA fiber analysis of eoRC patient-derived lymphocyte lines carrying the polymerase variants, showed significantly slower rate of DNA replication, and replication fork restoration after DNA damage versus the matched control-derived lymphocyte lines (P&amp;lt;0.01, Mann-Whitney test). Biochemical analysis of purified polymerase proteins found that the variants differ from wild type polymerase in their ability to extend DNA substrates and bypass DNA lesion(s), and exhibit dominant negative effects. Conclusion and future work: Our results indicate that germline variants in DNA repair genes impact suppression of DNA damage in cells from patients with eoRC. Biological studies testing the impact of the identified variants on renal carcinogenesis are warranted. Overall, genetic testing for variation in DNA repair genes may provide a more comprehensive risk assessment in eoRC patients, and provide novel opportunities for targeting DNA repair vulnerabilities in RC. Citation Format: Elena V. Demidova, Waleed Iqbal, Ilya G. Serebriiskii, Andrea F. Forman, Tatiana Kent, Richard T. Pomerantz, Erica A. Golemis, Michael J. Hall, David Y. Chen, Mary B. Daly, Sanjeevani Arora. Functional evaluation of novel germline DNA repair variants identified in patients with early-onset renal cancer [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2021; 2021 Apr 10-15 and May 17-21. Philadelphia (PA): AACR; Cancer Res 2021;81(13_Suppl):Abstract nr 808.

Role of DNA Repair Variants and Diagnostic Radiology Exams in Differentiated Thyroid Cancer Risk: A Pooled Analysis of Two Case-Control Studies.

Given the increased use and diversity of diagnostic procedures, it is important to understand genetic susceptibility to radiation-induced thyroid cancer. On the basis of self-declared diagnostic radiology examination records in addition to existing literature, we estimated the radiation dose delivered to the thyroid gland from diagnostic procedures during childhood and adulthood in two case-control studies conducted in France. A total of 1,071 differentiated thyroid cancer (DTC) cases and 1,188 controls from the combined studies were genotyped using a custom-made Illumina OncoArray DNA chip. We focused our analysis on variants in genes involved in DNA damage response and repair pathways, representing a total of 5,817 SNPs in 571 genes. We estimated the OR per milli-Gray (OR/mGy) of the radiation dose delivered to the thyroid gland using conditional logistic regression. We then used an unconditional logistic regression model to assess the association between DNA repair gene variants and DTC risk. We performed a meta-analysis of the two studies. The OR/mGy was 1.02 (95% confidence interval, 1.00-1.03). We found significant associations between DTC and rs7164173 in CHD2 (P = 5.79 × 10-5), rs6067822 in NFATc2 (P = 9.26 × 10-5), rs1059394 and rs699517 both in ENOSF1/THYS, rs12702628 in RPA3, and an interaction between rs7068306 in MGMT and thyroid radiation doses (P = 3.40 × 10-4). Our results suggest a role for variants in CDH2, NFATc2, ENOSF1/THYS, RPA3, and MGMT in DTC risk. CDH2, NFATc2, ENOSF1/THYS, and RPA3 have not previously been shown to be associated with DTC risk.

Investigating the effects of additional truncating variants in DNA-repair genes on breast cancer risk in BRCA1-positive women

BackgroundInherited pathogenic variants in BRCA1 and BRCA2 are the most common causes of hereditary breast and ovarian cancer (HBOC). The risk of developing breast cancer by age 80 in women carrying a BRCA1 pathogenic variant is 72%. The lifetime risk varies between families and even within affected individuals of the same family. The cause of this variability is largely unknown, but it is hypothesized that additional genetic factors contribute to differences in age at onset (AAO). Here we investigated whether truncating and rare missense variants in genes of different DNA-repair pathways contribute to this phenomenon.MethodsWe used extreme phenotype sampling to recruit 133 BRCA1-positive patients with either early breast cancer onset, below 35 (early AAO cohort) or cancer-free by age 60 (controls). Next Generation Sequencing (NGS) was used to screen for variants in 311 genes involved in different DNA-repair pathways.ResultsPatients with an early AAO (73 women) had developed breast cancer at a median age of 27 years (interquartile range (IQR); 25.00–27.00 years). A total of 3703 variants were detected in all patients and 43 of those (1.2%) were truncating variants. The truncating variants were found in 26 women of the early AAO group (35.6%; 95%-CI 24.7 - 47.7%) compared to 16 women of controls (26.7%; 95%-CI 16.1 to 39.7%). When adjusted for environmental factors and family history, the odds ratio indicated an increased breast cancer risk for those carrying an additional truncating DNA-repair variant to BRCA1 mutation (OR: 3.1; 95%-CI 0.92 to 11.5; p-value = 0.07), although it did not reach the conventionally acceptable significance level of 0.05.ConclusionsTo our knowledge this is the first time that the combined effect of truncating variants in DNA-repair genes on AAO in patients with hereditary breast cancer is investigated. Our results indicate that co-occurring truncating variants might be associated with an earlier onset of breast cancer in BRCA1-positive patients. Larger cohorts are needed to confirm these results.

Subsequent neoplasm risk associated with rare variants in DNA repair and clinical radiation sensitivity syndrome genes: A report from the Childhood Cancer Survivor Study.

10028 Background: Radiotherapy for childhood cancer is associated with strikingly elevated risk for developing subsequent neoplasms (SNs). Whether mutations in DNA repair and radiation sensitivity genes modulate SN risks is largely unknown. Methods: We conducted whole-exome sequencing in 5105 long-term childhood cancer survivors of European descent (mean follow-up = 32.7 years). SnpEff and ClinVar identified potentially damaging rare variants in 476 DNA repair or radiation sensitivity genes. Conditional logistic regression assessed SN risk associated with these variants aggregated by gene or pathway (N = 155 with ≥5 carriers). Controls were matched on sex, childhood cancer type and diagnosis age, radiation dose to the SN site, and survival. Exact p-values were calculated by permutation. Analyses used all survivors or subsets stratified on radiation dose. Results: A total of 1108 (21.7%) survivors developed at least one SN type known to be related to ionizing radiation exposure (e.g., breast cancer, basal cell carcinoma, meningioma, thyroid cancer, sarcoma). Radiation-related SN risk was associated with homologous recombination (HR) gene variants for SN sites that received &gt; 0- &lt; 10 Gy (20.9% cases, 11.0% controls; odds ratio [OR] = 2.20, 95% confidence interval [CI] 1.52-3.18; P = 1.41x10-4), most notably for FANCM (3.1% cases, 0.5% controls; OR = 9.91, 95%CI 3.73-26.4; P = 2.85x10-4). For radiation-related SNs at sites with higher doses (≥10 Gy), associations were not observed for the HR pathway (14.4% cases, 12.4% controls, P = 0.17) but were observed for two individual genes implicated in double-strand DNA break repair, EXO1 (1.8% cases, 0.4% controls; OR = 6.50, 95%CI 3.49-12.1; P = 7.43x10-4) and NEIL3 (0% cases, 1.0% controls; P = 3.23x10-4). Conclusions: In this discovery study, we identified dose-specific novel associations between SN risk after radiotherapy for childhood cancer and potentially damaging rare variants in genes involved in double-strand DNA break repair, particularly HR. If replicated, these results could impact long-term screening of childhood cancer survivors and risk-benefit assessments of treatment approaches.

Prevalence of pathogenic germline variants in DNA repair by race, age, and ethnicity in men with prostate cancer.

5062 Background: Patients with advanced prostate cancer (PC) frequently harbor pathogenic or likely pathogenic (P/LP) germline variants (GVs) in mismatch repair (MMR) and homologous repair (HR) enzymes which have clinical and treatment implications. However, whether the prevalence of such GVs differ by race, ethnicity, family history, or age is unknown in men with PC. Methods: This is a retrospective analysis of germline DNA from men with PC in the United States, tested by Invitae. Baseline characteristics including self-identified race, ethnicity, family history (FH), and age were recorded. Race and ethnicity were analyzed by 3 cohorts: non-Ashkenazi Caucasian Americans (CA), non-Ashkenazi African Americans (AA), and Ashkenazi Jewish Americans (AJ). Chi-square testing was performed to identify significant differences across these categories between the three cohorts, with respect to combined and individual pathogenic MMR (MSH2/6, MLH1, PMS2, and MUTYH) and HR genes (BRCA1/2, ATM, CHEK2, RAD51D, and PALB2). Results: 3057 men were included in the final analysis: 2248 (74%) men were CA, 229 (7%) were AA, and 210 (7%) were AJ. Of these, 2665 (87%) men had a FH of PC and 463 (15%) had a P/LP GV. In addition, 1068 (35%) were found to have a variant of uncertain significance, and 35 (1.6%) had the HOXB13 G84E variant. There were no significant differences in the overall prevalence of MMR (CA 1.5% vs AA 0.9% vs AJ 1.4%, p = 0.89) or HR genes (CA 7.8% vs AA 7.9% vs AJ 10.5%, p = 0.37) by race/ethnicity. With respect to individual genes, AJ had a higher prevalence of pathogenic BRCA1 alterations (AJ 3.3% vs CA 0.8% vs AA 1.7%, p = 0.0034) and CHEK2 alterations (AJ 4.3% vs CA 2.8% vs AA 0.4%, p = 0.02). There were no significant differences in the prevalence of individual or specific classes of GVs between those with or without a self-reported FH of prostate or breast/ovarian cancers. There was also no association between prevalence of HR genes and age at germline testing (p = 0.40). Conclusions: This national study found that the overall prevalence of pathogenic GVs in MMR and HR genes do not differ by race, ethnicity, or age at the time of testing, and suggests that all men with advanced prostate cancer should be offered germline testing.