Deleterious Germline 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.

Germline DNA Damage Repair Gene Alterations in Patients with Metachronous Breast and Colorectal Cancer.

A hereditary component of breast (BC) and colorectal cancer (CRC) has been described in approximately one-third of these tumor types. BC patients have an increased risk of developing CRC as a second primary tumor and vice versa. Germline genomic variants (NextSeq550, Illumina) were investigated in 24 unrelated BC and/or CRC patients and 7 relatives from 3 index patients. Fifty-six pathogenic or likely pathogenic variants were identified in 19 of 24 patients. We detected single-nucleotide variants (SNVs) in CRC predisposition genes (MLH1 and MUTYH) and other promising candidates (CDK5RAP3, MAD1L1, NOS3, and POLM). Eighteen patients presented SNVs or copy number variants (CNVs) in DNA damage repair genes. We also identified SNVs recently associated with BC or CRC predisposition (PABPC1, TYRO3, MAP3K1, SLC15A4, and LAMA1). The PABPC1c.1255C>T variant was detected in nine unrelated patients. Each patient presented at least one SNV/CNV in a candidate gene, and most had alterations in more than one gene, reinforcing a polygenic model for BC/CRC predisposition. A significant fraction of BC/CRC patients with a family history of these tumors harbored deleterious germline variants in DNA repair genes. Our findings can lead to strategies to improve the diagnosis, genetic counseling, and treatment of patients and their relatives.

Genomic ascertainment of CHEK2-related cancer predisposition.

There is clear evidence that deleterious germline variants in CHEK2 increases risk for breast and prostate cancers; there is limited or conflicting evidence for other cancers. Genomic ascertainment was used to quantify cancer risk in CHEK2 germline pathogenic variant heterozygotes. Germline CHEK2 variants were extracted from two exome-sequenced biobanks linked to the electronic health record: UK Biobank (n= 469,765) and Geisinger MyCode (n=170,503). Variants were classified as per American College of Medical Genetics and Genomics (ACMG)/Association for Molecular Pathology (AMP) criteria. Heterozygotes harbored a CHEK2 pathogenic/likely pathogenic (P/LP) variant; controls harbored benign/likely benign CHEK2 variation or wildtype CHEK2. Tumor phenotype and demographic data were retrieved; to adjust for relatedness, association analysis was performed with SAIGE-GENE+ with Bonferroni correction. In CHEK2 heterozygotes in both MyCode and UK Biobank, there was a significant excess risk of all cancers tested, including breast cancer (C50; OR=1.54 and 1.84, respectively), male genital organ cancer (C60-C63; OR=1.61 and 1.77 respectively), urinary tract cancer (C64-C68; OR=1.56 and 1.75, respectively) and lymphoid, hematopoietic, and related tissue cancer (C81-C96; OR=1.42 and 2.11, respectively). Compared to controls, age-dependent cancer penetrance in CHEK2 heterozygotes was significantly younger in both cohorts; no significant difference was observed between the penetrance of truncating and missense variants for cancer in either cohort. Overall survival was significantly decreased in CHEK2 heterozygotes in UK Biobank but there was no statistical difference in MyCode. Using genomic ascertainment in two population-scale cohorts, this investigation quantified the prevalence, penetrance, cancer phenotype and survival in CHEK2 heterozygotes. Tailored treatment options and surveillance strategies to manage those risks are warranted.

Deleterious ZNRF3 germline variants cause neurodevelopmental disorders with mirror brain phenotypes via domain-specific effects on Wnt/β-catenin signaling

Zinc and RING finger 3 (ZNRF3) is a negative-feedback regulator of Wnt/β-catenin signaling, which plays an important role in human brain development. Although somatically frequently mutated in cancer, germline variants in ZNRF3 have not been established as causative for neurodevelopmental disorders (NDDs). We identified 12 individuals with ZNRF3 variants and various phenotypes via GeneMatcher/Decipher and evaluated genotype-phenotype correlation. We performed structural modeling and representative deleterious and control variants were assessed using invitro transcriptional reporter assays with and without Wnt-ligand Wnt3a and/or Wnt-potentiator R-spondin (RSPO). Eight individuals harbored de novo missense variants and presented with NDD. We found missense variants associated with macrocephalic NDD to cluster in the RING ligase domain. Structural modeling predicted disruption of the ubiquitin ligase function likely compromising Wnt receptor turnover. Accordingly, the functional assays showed enhanced Wnt/β-catenin signaling for these variants in a dominant negative manner. Contrarily, an individual with microcephalic NDD harbored a missense variant in the RSPO-binding domain predicted to disrupt binding affinity to RSPO and showed attenuated Wnt/β-catenin signaling in the same assays. Additionally, four individuals harbored de novo truncating or de novo or inherited large in-frame deletion variants with non-NDD phenotypes, including heart, adrenal, or nephrotic problems. In contrast to NDD-associated missense variants, the effects on Wnt/β-catenin signaling were comparable between the truncating variant and the empty vector and between benign variants and the wild type. In summary, we provide evidence for mirror brain size phenotypes caused by distinct pathomechanisms in Wnt/β-catenin signaling through protein domain-specific deleterious ZNRF3 germline missense variants.

Germline Predisposition to Hematopoietic Malignancies: An Overview

Deleterious germline variants are now recognized as common drivers of hematopoietic malignancies (HMs) and bone marrow failure syndromes. With the increasing use of personalized medicine and the application of tumor-based profiling via next-generation sequencing, diagnosis of HM predisposition occurs with increasing frequency. Although deleterious germline variants can be readily identified by comprehensive clinical testing, numerous barriers exist for many clinicians. Observations regarding particular germline predisposition disorders challenge widely held assumptions about these conditions. Here, we review approaches to germline genetic testing, highlighting key points in a typical patient's course that present challenges for testing and interpreting results. Increasing awareness by health care providers of these conditions and improvements in testing platforms are crucial for enabling a proactive approach to tailoring a suitable treatment plan and surveillance program for the patient and their family members.

Germline Predisposition in Hematologic Malignancies: Testing, Management, and Implications.

Although numerous barriers for clinical germline cancer predisposition testing exist, the increasing recognition of deleterious germline DNA variants contributing to myeloid malignancy risk is yielding steady improvements in referrals for testing and testing availability. Many germline predisposition alleles are common in populations, and the increasing number of recognized disorders makes inherited myeloid malignancy risk an entity worthy of consideration for all patients regardless of age at diagnosis. Germline testing is facilitated by obtaining DNA from cultured skin fibroblasts or hair bulbs, and cascade testing is easily performed via buccal swab, saliva, or blood. Increasingly as diagnostic criteria and clinical management guidelines include germline myeloid malignancy predisposition, insurance companies recognize the value of testing and provide coverage. Once an individual is recognized to have a deleterious germline variant that confers risk for myeloid malignancies, a personalized cancer surveillance plan can be developed that incorporates screening for other cancer risk outside of the hematopoietic system and/or other organ pathology. The future may also include monitoring the development of clonal hematopoiesis, which is common for many of these cancer risk disorders and/or inclusion of strategies to delay or prevent progression to overt myeloid malignancy. As research continues to identify new myeloid predisposition disorders, we may soon recommend testing for these conditions for all patients diagnosed with a myeloid predisposition condition.

Abstract GS03-11: Germline-mediated immunoediting sculpts breast cancer subtypes and metastatic proclivity

Abstract Background: Somatic genomic aberrations are acquired within the context of germline genomes which differ across individuals at millions of polymorphic sites. However, the role of germline variation in somatic evolution remains poorly understood. The most compelling example is is that deleterious germline variants in BRCA1 and, to a lesser extent, BRCA2 are preferentially associated with the development of triple-negative breast cancer (BC). The variable frequency of BC subtypes across ancestral populations further suggests a role for germline contributions. On the other hand, various lines of evidence indicate that avoidance of the adaptive immune system is a strong determinant of which somatic mutations persist within a tumor. Whether and how germline differences influence immunoediting has not been studied. Building on these observations, we sought to investigate whether germline variation mediates somatic evolution through immunoediting. Specifically, we hypothesize inherited variation in oncogenes would be subject to varied immunoediting pressures during malignant transformation and progression. A high burden of germline-derived epitopes in recurrently amplified oncogenes is predicted to select against amplification of the cognate gene during malignant transformation because this would increase epitope availability, the likelihood of epitope presentation, and immune-mediated cell death. Instead, immune pressures may select for amplification of an alternate driver gene with a lower germline-mediated epitope burden. We evaluated this hypothesis in a collection of 3,855 BC, spanning ductal carcinoma in situ (DCIS) to invasive BC, and metastatic lesions. Methods: We analyzed paired tumor and normal sequencing data from 1,087 primary and 702 metastatic breast cancer patients as well as somatic genomic profiles from 341 patients with DCIS using a novel algorithm to estimate germline-derived epitope burden (GEB) based on an individual’s genotype and class 1 HLA alleles. The relationship between GEB and subtype commitment, defined by the acquisition of focal oncogenic amplifications in five prognostic subgroups of BC: HER2+ disease and four high-risk of relapse ER+/HER2 integrative subgroups/clusters (ICs) which we previously described (IC1: 17q23, IC2: 11q13, IC6: 8p12, and IC9: 8q24) was evaluated. Specifically, we evaluated the association between the GEB per gene and whether an individual developed the corresponding subtype via logistic regression, correcting for the first six genetic principal components and somatic mutation burden. Outcome associations were evaluated via Cox Proportional Hazards Models. Results: Interrogating 3,855 breast cancer lesions, we demonstrate that germline-derived epitopes in recurrently amplified genes influence somatic evolution by mediating immunoediting. Individuals with a high GEB in ERBB2/HER2 are significantly less likely to develop HER2-positive breast cancer compared to other subtypes. The same holds true for recurrent amplicons that define four aggressive, high-risk of relapse, ER-positive integrative subgroups. Thus, GEB selects against cognate oncogene amplification. Tumors that overcome such immune-mediated negative selection are more aggressive and exhibited microenvironments depleted of lymphocytes, consistent with “immune cold” tumors. Conclusions: We demonstrate that inherited variation sculpts breast cancer subtypes, aggressivity, and immune landscapes by mediating anti-tumor immune responses. The implications of these findings are severalfold. First, GEB is prognostic, complementing other molecular measurements. Second, immunoediting pressures differ across the disease course, with implications for the timing of therapeutic interventions. Third, these data illuminate a broad source of currently under-appreciated immunogenic antigens. Citation Format: Kathleen Houlahan, Aziz Khan, Noah Greenwald, Robert West, Michael Angelo, Christina Curtis. Germline-mediated immunoediting sculpts breast cancer subtypes and metastatic proclivity [abstract]. In: Proceedings of the 2023 San Antonio Breast Cancer Symposium; 2023 Dec 5-9; San Antonio, TX. Philadelphia (PA): AACR; Cancer Res 2024;84(9 Suppl):Abstract nr GS03-11.

Telomere-lengthening germline variants predispose to a syndromic papillary thyroid cancer subtype

Papillary thyroid cancer (PTC) is the most common endocrine malignancy. 10% to 15% of individuals show familial clustering with three or more affected members, but the factors underlying this risk are unknown. In a group of recently studied individuals with POT1 pathogenic variants and ultra-long telomere length, PTC was the second most common solid tumor. We tested whether variants in POT1 and four other telomere-maintenance genes associated with familial cancer underlie PTC susceptibility. Among 470 individuals, we identified pathogenic or likely pathogenic variants in three genes encoding telomere-binding proteins: POT1, TINF2, and ACD. They were found in 4.5% and 1.5% of familial and unselected cases, respectively. Individuals harboring these variants had ultra-long telomere length, and 15 of 18 (83%) developed other cancers, of which melanoma, lymphoma, and sarcoma were most common. Among individuals with PTC and melanoma, 22% carried a deleterious germline variant, suggesting that a long telomere syndrome might be clinically recognizable. Successive generations had longer telomere length than their parents and, at times, developed more cancers at younger ages. Tumor sequencing identified a single oncogenic driver, BRAF p.Val600Glu, in 10 of 10 tumors studied, but no telomere-maintenance mechanism, including at the TERT promoter. These data identify a syndromic subset of PTCs with locusheterogeneity and telomere lengthening as a convergent mechanism. They suggest these germline variants lower the threshold to cancer by obviating the need for an acquired telomere-maintenance mechanism in addition to sustaining the longevity of oncogenic mutations.

Abstract NG01: Germline-mediated immunoediting sculpts breast cancer subtypes and metastatic proclivity

Abstract NG01: Germline-mediated immunoediting sculpts breast cancer subtypes and metastatic proclivity

BARD1 deletion in a patient with suspected hereditary colorectal cancer

Deleterious germline variants in the BRCA1-associated ring domain (BARD1) gene moderately elevate breast cancer risk; however, their potential association with other neoplasms remains unclear. Here, we present the case of a 43-year-old female patient diagnosed with sigmoid colon adenocarcinoma whose maternal family members met the Amsterdam Criteria II for Lynch syndrome. Comprehensive multigene panel testing revealed a heterozygous BARD1 exon 3 deletion.

Insights into the Molecular Mechanisms of Genetic Predisposition to Hematopoietic Malignancies: The Importance of Gene-Environment Interactions.

The recognition of host genetic factors underlying susceptibility to hematopoietic malignancies has increased greatly over the last decade. Historically, germline predisposition was thought to primarily affect the young. However, emerging data indicate that hematopoietic malignancies that develop in people of all ages across the human lifespan can derive from germline predisposing conditions and are not exclusively observed in younger individuals. The age at which hematopoietic malignancies manifest appears to correlate with distinct underlying biological pathways. Progression from having a deleterious germline variant to being diagnosed with overt malignancy involves complex, multistep gene-environment interactions with key external triggers, such as infection and inflammatory stimuli, driving clonal progression. Understanding the mechanisms by which predisposed clones transform under specific pressures may reveal strategies to better treat and even prevent hematopoietic malignancies from occurring.Recent unbiased genome-wide sequencing studies of children and adults with hematopoietic malignancies have revealed novel genes in which disease-causing variants are of germline origin. This paradigm shift is spearheaded by findings in myelodysplastic syndrome/acute myeloid leukemia (MDS/AML) as well as acute lymphoblastic leukemia, but it also encompasses other cancer types. Although not without challenges, the field of genetic cancer predisposition is advancing quickly, and a better understanding of the genetic basis of hematopoietic malignancies risk affects therapeutic decisions as well as genetic counseling and testing of at-risk family members.

Germline mutations in cancer predisposition genes among pediatric patients with cancer and congenital anomalies.

Childhood cancer has a poorly known etiology, and investigating the underlying genetic background may provide novel insights. A recognized association exists between non-chromosomal birth defects and childhood cancer susceptibility. We performed whole-exome sequencing and chromosomal microarray analysis in a cohort of childhood cancer (22 individuals, 50% with congenital anomalies) to unravel deleterious germline variants. A diagnostic yield of 14% was found, encompassing heterozygous variants in bona fide dominant Cancer Predisposition Genes (CPGs). Considering candidate and recessive CPGs harboring monoallelic variants, which were also deemed to play a role in the phenotype, the yield escalated to 45%. Most of the deleterious variants were mapped in genes not conventionally linked to the patient's tumor type. Relevant findings were detected in 55% of the syndromic individuals, mostly variants potentially underlying both phenotypes. We uncovered a remarkable prevalence of germline deleterious CPG variants, highlighting the significance of a comprehensive genetic analysis in pediatric cancer, especially when coupled with additional clinical signs. Moreover, our findings emphasized the potential for oligogenic inheritance, wherein multiple genes synergistically increase cancer risk. Lastly, our investigation unveiled potentially novel genotype-phenotype associations, such as SETD5 in neuroblastoma, KAT6A in gliomas, JAG1 in hepatoblastomas, and TNFRSF13B in Langerhans cell histiocytosis. Novel gene-phenotype associations and candidate genes for pediatric cancer were unraveled, such as KAT6A in gliomas, SETD5 in neuroblastoma, JAG1 in hepatoblastomas, and TNFRSF13B in Langerhans cell histiocytosis. Our analysis revealed a high frequency of deleterious germline variants, particularly in cases accompanied by additional clinical signs, highlighting the importance of a comprehensive genetic evaluation in childhood cancer. Our findings also underscored the potential for oligogenic inheritance in pediatric cancer risk. Understanding the cancer etiology is crucial for genetic counseling, often influencing therapeutic decisions and offering valuable insights into molecular targets for the development of oncological therapies.

Germline Determinants of Toxicity and Efficacy in Patients with Large B-Cell Lymphoma Treated with Anti-CD19 Autologous CAR T-Cell Therapy

Background. Increasing data have demonstrated that acquired genetic aberrations, including TP53 mutations in the tumor cells and clonal hematopoiesis, can impact the efficacy and/or toxicity of anti-CD19 autologous CAR T-cell therapy (CART) in patients with large B-cell lymphoma (LBCL). Recent data have suggested that germline genetic aberrations, including polymorphisms in CD19, ABCB1, MISP and CPVL (Strati P et al, Leukemia 2022) and deleterious germline variants in STXBP2, can also affect outcomes in these patients. However, a comprehensive analysis of germline determinants of response and toxicity after CART in this patient population has not yet been described. Methods. Genome-wide genotyping using the Illumina Genome Screening Array (GSAv2.0) was performed in 170 patients with relapsed or refractory LBCL treated with standard of care axicabtagene ciloleucel in third line and beyond. Response was assessed according to 2014 Lugano criteria. Cytokine release syndrome (CRS) and immune effector cell-associated neurotoxicity syndrome (ICANS) were graded according to ASTCT criteria, and cytopenia according to CTCAE v5. Polygenic risk score (PRS) instruments for blood cell traits and inflammatory markers were obtained from the PGS Catalog and analyzed using PRSice. Risk of poor response and toxicity was estimated using logistic regression models with standardized PRS scores as a continuous variable adjusting for age at diagnosis, gender, and bridging therapy (yes/no). Exploratory genome-wide association study (GWAS) analyses were also performed. Results. The median age was 58 years, 116 (68%) were male, and 91 (54%) received bridging therapy. Interestingly, increasing PRS for baseline IL-6 levels was associated with decreased risk of CRS grade (G) 2-4 (β: -0.15, 95% CI: -0.30 to -0.013, p=0.030) and ICANS G3-4 (β: -0.21, 95% CI: -0.37 to -0.066, p=0.005). Similarly, increased genetically-predicted monocyte levels decreased risk of ICANS G3-4 (β: -0.16, 95% CI: -0.31 to -0.018, p=0.027). Baseline genetically-predicted monocyte levels were also associated with increased probability of a CR at D90 (β: 0.19, 95% CI: 0.041 to 0.34, p=0.011). For prolonged cytopenia (defined as G3-4 cytopenia ongoing at day 30), risk was increased with increasing PRS for reticulocyte levels (β: 0.29, 95% CI: 0.10 to 0.48, p=0.002). No genome-wide significant (p<5x10 -8) variants were identified in the exploratory GWAS analyses, however, several loci at p=10 -5 were located within/near potential biologically-relevant candidate genes. This included an association with prolonged cytopenia for rs111940551 located upstream of IKZF1, a regulator of lymphocyte differentiation, and serving as a putative expression quantitative trait loci (eQTL) for this gene. An intragenic variant identified for ICANS G3-4 (rs1149927) was in high linkage disequilibrium (D'=0.97) with a variant previously shown to be associated with brain morphology. Conclusion. PRS representing baseline levels of several key blood cell traits, particularly monocytes, and inflammatory markers may impact toxicities and efficacy of CART in patients with LBCL. The intriguing finding of an inverse association for IL-6 with both high-grade CRS and ICANS was unexpected and needs further investigation. Larger studies are warranted to confirm these observations and better understand how relevant germline genetic determinants may influence efficacy and toxicity after CART therapy. Elucidating such determinants may help in improving patient selection and in developing strategies to enhance the therapeutic index of CART.

Frequency of CDH1, CTNNA1 and CTNND1 Germline Variants in Families with Diffuse and Mixed Gastric Cancer.

The most well-characterized hereditary form of gastric cancer is hereditary diffuse gastric cancer (HDGC), an autosomal dominant syndrome characterized by an increased risk of diffuse gastric and lobular breast cancer. HDGC is predominantly caused by germline pathogenic variants in the CDH1 gene, and more rarely in the CTNNA1 gene. Furthermore, the International Gastric Cancer Linkage Consortium (IGCLC) guidelines do not clarify whether or not mixed gastric cancer (with a diffuse component) should be considered in the HDGC genetic testing criteria. We aimed to evaluate the contribution of CTNNA1 and CTNND1 germline variants to HDGC. Additionally, we also intended to compare the frequencies of CDH1 and CTNNA1 (and eventually CTNND1) germline variants between patients with diffuse and mixed gastric carcinomas to evaluate if genetic testing for these genes should or should not be considered in patients with the latter. We analyzed the CDH1 gene in 67 cases affected with early-onset/familial mixed gastric carcinomas and the CTNNA1 and CTNND1 genes in 208 cases with diffuse or mixed gastric cancer who had tested negative for CDH1 pathogenic germline variants. A deleterious CTNNA1 germline variant was found in 0.7% (1/141) of diffuse gastric cancer patients meeting the 2020 IGCLC criteria, as compared to the rate of 2.8% of CDH1 deleterious variants found by us in this setting. No deleterious variants were found in CTNND1, but six variants of uncertain significance were identified in this gene. We did not find any pathogenic CDH1, CTNNA1 or CTNND1 variant in index patients with early-onset/familial mixed gastric cancer, so there is no evidence that supports including this tumor type in the testing criteria for germline variants in these genes. The role of the CTNND1 gene in inherited gastric cancer predisposition is still unclear.

Response to the FDA Decision Regarding DPYD Testing Prior to Fluoropyrimidine Chemotherapy.

Fluoropyrimidine (FP) chemotherapy is associated with severe, life-threatening toxicities, particularly among patients who carry deleterious germline variants in the DPYD gene. Pretreatment DPYD testing is standard of care throughout most of Europe; however, it has not been recommended in clinical practice guidelines in the United States. Due to increased risk of severe toxicity, a Citizen's Petition asked the US Food and Drug Administration (FDA) to update language in FP drug labels to recommend DPYD testing as part of a boxed warning and recommend FP dose reduction in patients carrying deleterious germline variants. In response, the FDA updated the capecitabine package insert to inform patients about the toxicity risk and test availability and consider DPYD testing. However, the FDA did not include a testing recommendation or requirement, or a boxed warning. Additionally, the FDA did not recommend FP dose adjustment in DPYD variant carriers. This review provides a critical assessment of the DPYD-FP pharmacogenetic association using the FDA's previously published Pharmacogenetic Pyramid, demonstrating that the evidence is compelling for recommending DPYD testing prior to FP treatment. Additionally, the FDA's stated concerns about recommending DPYD testing and DPYD-guided FP dose adjustment are addressed and discussed in the context of the FDA's other genetic testing and dose adjustment recommendations. We call on the FDA to follow our European counterparts in recommending DPYD testing and genotype-based dose adjustment to ensure patients with cancer receive safe and effective FP chemotherapy.

Abstract 5214: Expanding the spectrum of germline-driven cancers by leveraging population-scale targeted tumor and normal sequencing

Abstract Large case-control and familial studies have established clear cancer-specific risk profiles for several key cancer predisposition genes (CPGs). For example, germline pathogenic variants (PVs) in BRCA1/2 (gBRCA) are associated with increased risk for developing breast, ovarian, pancreatic, and prostate cancers. However, the extent to which gBRCA mutations are involved in mediating the tumorigenesis of other cancer types remains challenging to characterize. We hypothesized that integrating orthogonal features such as selection for biallelic inactivation of the PVs and depletion of canonical somatic drivers among the carriers can enrich the signal for identifying novel gene and cancer type associations. We then extend this framework to identify novel CPGs as well as to understand how tumors arise in patients with PVs in oncogenes. To study this, we leveraged the prospective MSK-IMPACT matched tumor-normal sequencing cohort of 49,291 patients across 77 major cancer types. We study 90 well-known CPGs as well as >300 cancer genes not previously associated with cancer predisposition. Overall, 8% (N=3,964) of patients harbored a PV in high or moderate penetrance CPGs. We identified 90 gene and cancer type associations with enrichment for biallelic inactivation (q<0.05), including 19 novel findings among clinically actionable genes. For example, we find enrichment for biallelic inactivation of BRCA1/2 in unexpected lineages such as hepatobiliary, endometrioid, and ampullary cancers. These tumors were also significantly depleted for somatic gain-of-function driver alterations. Hepatobiliary cancers with gBRCA1 mutations were also enriched for somatic loss of NF1. Among carriers of PVs in oncogenes, we observe two possible mechanisms of first somatic hit towards malignant transformation. We find enrichment for copy number gain or copy neutral loss of heterozygosity of the germline PV in thyroid cancers with a PV in RET. We also find that lung cancers with a germline PV in EGFR frequently developed additional somatic point mutations located in cis with the PV. Investigating genes with no prior association with germline predisposition to cancer, we find evidence for KEAP1 and CIC as likely novel CPGs. Lung (n=8) and thyroid (n=4) cancers with deleterious germline variants in KEAP1 were characterized by loss of the wild-type allele, co-occurring somatic STK11 mutations, and depletion of canonical drivers such as EGFR. We also found biallelic loss of CIC in two patients with Neuroblastoma, each carrying a different germline loss-of-function mutation in CIC. Both tumors were also negative for MYCN and ALK defects. Collectively, our findings expand our understanding of cancer predisposition in cancer, shed new insights into how tumors arise in germline carriers, and provide a framework for identifying new CPGs using population scale tumor-normal paired clinical sequencing data. Citation Format: Miika Mehine, Rebecca Caeser, Yelena Kemel, Daniel Muldoon, Sebastià Franch-Expósito, A. Rose Brannon, Aijazuddin Syed, Ozge Ceyhan-Birsoy, Maksym Misyura, Panieh Terraf, David B. Solit, Marc Ladanyi, Kenneth Offit, Zsofia K. Stadler, Diana L. Mandelker, Yonina R. Murciano-Goroff, Charles M. Rudin, Michael F. Berger, Chaitanya Bandlamudi. Expanding the spectrum of germline-driven cancers by leveraging population-scale targeted tumor and normal sequencing. [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2023; Part 1 (Regular and Invited Abstracts); 2023 Apr 14-19; Orlando, FL. Philadelphia (PA): AACR; Cancer Res 2023;83(7_Suppl):Abstract nr 5214.

Abstract 5238: The functional role of promoter germline variants in breast cancer susceptibility genes

Abstract It is estimated that inherited deleterious or pathogenic germline variants in high- to moderate- penetrance breast cancer (BC) susceptibility genes such as BRCA1, BRCA2, STK11, CDH1, ATM, PALB2, and CHEK2 may explain up to 27% of the BC cases. For the remaining cases, BC develops as a result of the complex interaction between environmental and lifestyle risk factors as well as inherited familial predisposition. While low penetrance genetic variants have limited impact on BC risk when taken individually, their combined effect on polygenic risk scores may provide a risk elevated enough to justify clinical actions. Most of the previous work in hereditary breast cancer research has focused on variants located in the coding region of the genes and as a result, there is an abundance of information on the functional interpretation of such variants, while information on regulatory variants remains limited. The goal of this study was to assess the functional impact of germline variants in the promoter region of BC susceptibility genes on gene expression level. Genes evaluated include CDH1, STK11, and bidirectional promoter genes ATM/NPAT and PALB2/DCTN5. A bioinformatics approach was developed to identify and prioritize variants based on conservation level and predicted binding of transcription regulatory proteins. Selected variants will be introduced into pGL3 constructs harboring the promoter of interest using site-directed mutagenesis followed by transfection into HEK293T cells. Dual-glo luciferase expression assays are used to measure levels of promoter activities. Expression levels from promoters containing genetic variants are compared with those from constructs containing the reference sequence. The reference constructs for STK11 and CDH1 containing the genes complete promoters demonstrated functional activity. For bidirectional promoters, forwards and reverse construct were tested. Forward ATM/NPAT and PALB2/DCTN5 constructs showed luciferase expression under the influence of ATM and DCTN5 respectively. Work is on the way to assess promoter activity of the reverse complement sequence of those promoters and determine the impact of 13 candidate variants on the strength of these promoters. At the clinical level, identifying germline variants that contribute to BC risk could help better define BC risk in women with negative hereditary cancer genetic test results. Citation Format: Nelly Arlene Arroyo, Lenin J. Godoy, Julie Dutil. The functional role of promoter germline variants in breast cancer susceptibility genes. [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2023; Part 1 (Regular and Invited Abstracts); 2023 Apr 14-19; Orlando, FL. Philadelphia (PA): AACR; Cancer Res 2023;83(7_Suppl):Abstract nr 5238.

SPECTRUM OF GERMLINE CANCER SUSCEPTIBILITY GENE MUTATIONS IN BREAST AND OVARIAN CANCER PATIENTS IN THE BLACK SEA REGION OF TURKEY: SINGLE CENTER EXPERIENCE

Introduction: Breast cancer consists huge amount of the cancerrelated death in population. Ovarian cancer is the second most frequent seen type of gynecological cancer and has the highest mortality among gynecological cancers since most cases are detected late. The current study intended to determine the prevalence of oncogene mutations, especially BRCA1 and BRCA2, in high-risk patients diagnosed with ovarian and breast cancer in the Black Sea region of our country. Material and method: Between August 2017 and January 2022, a total of 223 individuals who applied to our center and met the genetic test criteria were included in the study. Next-generation sequencing (NGS) was used to detect germ-line deleterious variants in genes included in the oncogenetic panel of patients (34 genes). Results: Among the 223 patients analyzed within the scope of the study, 195 had breast cancer, and 28 had ovarian cancer, resulting in the detection of 15 different pathogenic variants of BRCA1 (%4,9) and BRCA2 (%6,7) genes in 26 (11.6%) patients. In the analysis of 32 oncogenes other than BRCA1 and BRCA2 genes, 26 different pathogenic (P) or likely pathogenic (LP) variants were detected in a total of 35 patients (15.7%). Based on the analysis of 223 breast/ ovarian cancer patients together, 41 different pathogenic (P) or likely pathogenic (LP) variants were found in 61 patients (27.3%). Furthermore, 65 different VUSs (Variant of Uncertain Significance) were detected in 73 patients (32.7%). Conclusion: This is the first study to be conducted in our region in a single center located in the Black Sea region. The study was conducted in a single center within the Black Sea region and, to our knowledge, provides the first data in this region in terms of cancer genes other than BRCAs. To appreciate of the genetic susceptibility spectrum of hereditary breast and/or ovarian cancer better, it is imperative to clarify the risks associated with genes other than BRCAs, which carry a high risk for other breast and ovarian cancers, as well as BRCA1 and BRCA2. Therefore, patients in the risk group must undergo multigene panel testing in addition to routine BRCA1 and BRCA2 gene testing. We detected two novel variants in the BRCA2 gene and five novel variants other than BRCA oncogenes. Furthermore, the results of this study contributed to the development of our country's specific variant pool.

Abstract P5-12-06: Comprehensive analysis of DNA damage repair gene germline mutations in Chinese breast cancer patients

Abstract Background: Germline DNA damage repair (DDR) mutations has been associated with increased cancer risk, PARP inhibitor therapeutic opportunity for breast cancer (BC) patients. However, the profile of germline mutations in BC covering comprehensive DDR genes remains unclear. Methods: A total of 341 women with breast cancer who tested 102 germline related genes (including 50 DDR genes) between April 2021 to May 2022 in Guangdong Provincial People’s Hospital were identified. Variants were classified into pathogenic, likely pathogenic, variant of uncertain significance (VUS), likely benign and benign groups according to the ACMG/AMP Standards and Guidelines. We defined pathogenic and likely pathogenic variants as deleterious mutations. Results: The median age of 341 breast cancer patients was 48 (range, 20-89) at the first diagnosis of BC. A total of 47 patients (13.78%) carried 53 deleterious germline variants in 21 cancer predisposition genes, 16 of which were DDR genes. DDR deleterious mutations were detected in genes including BRCA2 (n=18), BRCA1(n=7), FANCA (n=4), PMS2 (n=4), PALB2(n=2), RECQL4(n=2), PALB2 (n=2), etc. The younger age at diagnosis (less than 40-year-old) were significantly associated with deleterious mutations in DDR pathway(P=0.02). At least one VUS was identified in 238 (69.79%) patients. The top 5 DDR VUS genes were FANCM (n=21), ATM (n=20), RAD54L (n=17), FANCD2 (n=15) and ATR (n=14). Breast or ovarian cancer family history were significantly correlated with VUS germline mutations in DDR pathway(P=0.039). Interesting, we found that patients with pCR efficacy of neoadjuvant therapy were more likely to have VUS mutations in DDR pathway (table 1). Conclusion: We provided a comprehensive view of germline DDR gene mutations in BC patients and also analyzed the association between clinical characteristics and germline DDR mutation status. DDR mutations are prevalent in Chinese BC patients. Patients with younger and breast or ovarian cancer family history were more likely to carry DDR alterations. Moreover, patients with higher frequency of DDR VUS mutations may benefit from neoadjuvant therapy. Table 1. Clinicopathological characteristics between germline mutation carriers and non-carriers Citation Format: Ning Liao, Li Cao, Guochun Zhang, Junyun Wang, Airong Yang, Yulei Wang, Kai Li, Lingzhu Wen, Chongyang Ren, Minghan Jia, Cheukfai Li, Hsiaopei Mok, Bo Chen, Jianguo Lai, Weikai Xiao. Comprehensive analysis of DNA damage repair gene germline mutations in Chinese breast cancer patients [abstract]. In: Proceedings of the 2022 San Antonio Breast Cancer Symposium; 2022 Dec 6-10; San Antonio, TX. Philadelphia (PA): AACR; Cancer Res 2023;83(5 Suppl):Abstract nr P5-12-06.

Allogeneic hematopoietic stem cell transplant outcomes in adults with inherited myeloid malignancies.

There is increasing recognition that pathogenic germ line variants drive the development of hematopoietic cancers in many individuals. Currently, patients with hereditary hematologic malignancies (HHMs) receive similar standard therapies and hematopoietic stem cell transplant (HSCT) approaches as those with sporadic disease. We hypothesize that patients with myeloid malignancies and deleterious germ line predisposition variants have different posttransplant outcomes than those without such alleles. We studied 472 patients with myeloid neoplasms, of whom 26% had deleterious germ line variants and 34% underwent HSCT. Deleterious germ line variants in CHEK2 and DDX41 were most commonly seen in American and Australian cohorts, respectively. Patients with deleterious germ line DDX41 variants had a higher incidence of severe (stage 3-4) acute graft-versus-host disease (GVHD) (38%) than recipients with deleterious CHEK2 variants (0%), other HHM variants (12%), or patients without such germ line variants (9%) (P= .002). Importantly, the use of posttransplant cyclophosphamide reduced the risk of severe acute GVHD in patients receiving HSCT for deleterious germ line DDX41-associated myeloid neoplasms (0% vs 53%, P= .03). Based on these results, we advocate the use of posttransplant cyclophosphamide when individuals with deleterious germ line DDX41 variants undergo allogeneic HSCT for myeloid malignancies, even when transplantation has been performed using wild-type donors.