Deleterious Germline Variants Research Articles

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.

Germline heterozygous exons 8–11 pathogenic BARD1 gene deletion reported for the first time in a family with suspicion of a hereditary colorectal cancer syndrome: more than an incidental finding?

BackgroundColorectal cancer (CRC) is a highly prevalent disease in developed countries. Inherited Mendelian causes account for approximately 5% of CRC cases, with Lynch syndrome and familial adenomatous polyposis being the most prevalent forms. Scientific efforts are focused on the discovery of new candidate genes associated with CRC and new associations of phenotypes with well-established cancer-related genes. BRCA1-associated ring domain (BARD1) gene deleterious germline variants are associated with a moderate increase in the relative risk of breast cancer, but their association with other neoplasms, such as CRC, remains unclear.Case presentationWe present the case of a 49-year-old male diagnosed with rectal adenocarcinoma whose maternal family fulfilled Amsterdam clinical criteria for Lynch syndrome. Genetic test confirmed the presence in heterozygosis of a germline pathogenic deletion of exons 8–11 in BARD1 gene. The predictive genetic study of the family revealed the presence of this pathogenic variant in his deceased cancer affected relatives, confirming co-segregation of the deletion with the disease.ConclusionsTo the best of our knowledge, this is the first published work in which this BARD1 deletion is detected in a family with familial colorectal cancer type X (FCCTX) syndrome, in which the clinical criteria for Lynch syndrome without alteration of the DNA mismatch repair (MMR) system are fulfilled. Whether this incidental germline finding is the cause of familial colorectal aggregation remains to be elucidated in scientific forums. Patients should be carefully assessed in specific cancer genetic counseling units to account for hypothetical casual findings in other genes, in principle unrelated to the initial clinical suspicion, but with potential impact on their health.

Exome sequencing of affected duos and trios uncovers PRUNE2 as a novel prostate cancer predisposition gene.

Prostate cancer (PrCa) is one of the most hereditable human cancers, however, only a small fraction of patients has been shown to carry deleterious variants in known cancer predisposition genes. Whole-exome sequencing was performed in multiple affected members of 45 PrCa families to select the best candidate genes behind part of the PrCa missing hereditability. Recurrently mutated genes were prioritised, and further investigated by targeted next-generation sequencing in the whole early-onset and/or familial PrCa series of 462 patients. PRUNE2 stood out from our analysis when also considering the available data on its association with PrCa development. Ten germline pathogenic/likely pathogenic variants in the PRUNE2 gene were identified in 13 patients. The most frequent variant was found in three unrelated patients and identical-by-descent analysis revealed that the haplotype associated with the variant is shared by all the variant carriers, supporting the existence of a common ancestor. This is the first report of pathogenic/likely pathogenic germline variants in PRUNE2 in PrCa patients, namely in those with early-onset/familial disease. Importantly, PRUNE2 was the most frequently mutated gene in the whole series, with a deleterious germline variant identified in 2.8% of the patients, representing a novel prostate cancer predisposition gene.

Rare Deleterious TCF3 Germline Variants and Predisposition to Acute Lymphoblastic Leukemia in Children

Introduction TCF3 is a hematopoietic transcription factor essential for lymphoid differentiation. The role of TCF3 in the pathogenesis of B-cell acute lymphoblastic leukemia (B-ALL) is well documented because of recurrent chromosomal rearrangements leading to TCF3 fusion with PBX1 or HLF1. In contrast, less is known about the occurrence and significance of germline TCF3 variants in this blood cancer. Here we present a comprehensive screen and characterization of TCF3 germline variants in children with B-ALL. Methods First, we performed targeted sequencing of TCF3 coding regions in 4,183 children with newly diagnosed B-ALL enrolled in Children's Oncology Group (COG) clinical trials AALL0232, P9904/5/6, and AALL0331 as well as St. Jude Total Therapy XIII and XV clinical trials. Rare variant analysis was performed using the CoCoRV pipeline (Chen W et al., Nat Commun. 2022, PMID: 35545612), where the GnomAD v2 genome-based summary counts (n=15,708) were used as the non-ALL control cohort. Rare deleterious TCF3 variants were defined as follows: allele frequency ≤5x10-4 in GnomAD, protein-truncating variants (frameshift and nonsense) or missense variants with a REVEL score ≥0.65. For transcription factor activity measurement, we used a dual-luciferase reporter assay in HEK 293T cells. Finally, we compared the clinical characteristics of B-ALL patients with rare deleterious TCF3 germline variants to B-ALL patients without these variants enrolled in COG AALL0232 (n=2,180) and COG P9904/5/6 (n=1,619) clinical trials. Results TCF3 alternative splicing gives rise to two distinct isoforms, E47 and E12, of which we found TCF3-E12 to be the predominant transcript expressed in B-ALL. Focusing our analysis on the E12 isoform (NM_003200), we identified 12 rare deleterious TCF3 variants in 12 B-ALL patients, significantly and strikingly more frequent than what is seen in non-ALL controls (p=1.24×10-5, OR=19.9) (Figure 1A). Nine TCF3 variants lead to protein truncation with the loss of the basic helix-loop-helix (b-HLH) domain and three missense variants cluster within TCF3 exon 18a, which encodes the b-HLH domain that is responsible for TCF3 dimerization and DNA binding. Functional characterization by luciferase reporter assay revealed a severe reduction of TCF3 transcriptional activity for two B-ALL-related missense variants. TCF3 E12-p.N554S and E12-p.A568D showed a 6.2- (p<0.0001) and 6.9-fold (p<0.0001) reduction in transcription factor activity compared to wildtype TCF3 E12 (Figure 1B). TCF3 E47-p.E555K is a known loss of function variant and therefore was included as a positive control. As expected, its transcription factor activity was reduced by 11.8-fold compared to wildtype TCF3 E47 (p<0.0001). These results are consistent with the notion that single nucleotide variations within the b-HLH domain impair TCF3 dimerization and DNA binding. Nonsense and frameshift variants result in the loss of the b-HLH domain and were presumed to be deleterious. Finally, we compared clinical features of 10 TCF3 germline variant carriers to 3,799 B-ALL patients without TCF3 variation enrolled in COG clinical trials. We did not observe any difference in age, sex, ancestry, or white blood count at diagnosis. Further, none of the 10 patients with TCF3 germline variant had somatic fusion genes involving TCF3 (i.e., TCF3-PBX1 or TCF3-HLF1). Conclusion We systematically examined germline genetic variants in the TCF3 gene in B-ALL patients, reporting 12 rare deleterious TCF3 variants with a cumulative frequency of 0.29%. These variants directly affect the TCF3 b-HLH domain, which is critical for TCF3 transcription factor activity. Taken together, our findings provide new insights into genetic predisposition to pediatric B-ALL. Figure 1View largeDownload PPTFigure 1View largeDownload PPT Close modal

Allogeneic Hematopoietic Stem Cell Transplant Outcomes in Adults with Inherited Myeloid Malignancies

Recognition of hereditary hematopoietic malignancies (HHMs) has accelerated in recent years, with deleterious germline variants in more than 20 genes recognized as drivers of myeloid malignancies. Individuals with HHMs generally receive similar treatment plans as those with de novo disease. Hematopoietic stem cell transplantation (HSCT) is often employed as a curative approach for myeloid malignancies, and in those with germline pathogenic (P)/likely pathogenic (LP) predisposition variants, there is a preference for use of related donors who lack the familial variant. However, there are limited data on the outcomes of patients with HHMs who are treated in this manner. In this study, we used comprehensive clinical and genomic data to investigate the allogeneic transplant-related outcomes in the largest international myeloid HHM cohort reported to date. We performed a retrospective analysis of all patients with myeloid malignancies who underwent germline predisposition testing at The University of Chicago (UofC) or the Central Adelaide Health Network between 2010 and 2021; or were known to have deleterious germline DDX41 variants from the University of Pennsylvania and Moffitt Cancer Center. We studied 472 patients, of whom 28% (80/289) in the UofC cohort and 20% (35/175) in the Adelaide cohort had a deleterious germline variant causative of HHMs. In the UofC cohort, P/LP CHEK2 variants were the most common, whereas P/LP DDX41 variants were most common in the Adelaide cohort (Fig. 1). A total of 161 patients (34% of the entire cohort) underwent allogeneic HSCT for their myeloid neoplasm (Table 1). There were no differences in the distribution of diagnoses, donor types, disease status at the time of HSCT (remission vs active disease), conditioning regimens, or conditioning intensity. All donor cells were mobilized from peripheral blood. Almost all donors lacked the deleterious germline causative variant, with one donor sharing a deleterious familial DDX41 variant with the relative/HSCT recipient. The median time to engraftment was similar between those who had vs lacked germline HHM variants. Patients with P/LP germline DDX41 variants had higher rates of severe (stage 3-4) acute graft versus host disease (GVHD) when compared to patients with P/LP CHEK2, those with other germline variants, or those without P/LP germline variants (38% vs 0, 12%, and 9%, respectively; p=0.002). Notably, severe gastrointestinal GVHD was seen in a third of patients with P/LP germline DDX41 variants as opposed to <10% incidence in other groups (p<0.001, Table 1). Similarly, 33% of patients with P/LP germline DDX41 variants experienced moderate to severe chronic GVHD, whereas this incidence was 29% for patients with P/LP germline CHEK2 variants, 4% for patients with other P/LP germline variants, and 10% for patients without a P/LP inherited variant (p=0.008). Patients with P/LP germline DDX41 variants who received post-HSCT cyclophosphamide had lower rates of severe acute GVHD compared to those who received other forms of GVHD prophylaxis (0% vs 53%, p=0.03). To adjust for the effects of known GVHD risk factors, we performed a multivariable logistic regression analysis. Germline DDX41 status remained an independent predictor of both acute GVHD (OR: 4.76, range 1.27-17.9; p=0.01) and chronic GVHD (OR: 3.39, range 1.09-10.5; p=0.03). Importantly, post-HSCT cyclophosphamide use was associated with a reduced risk of acute GHVD when adjusted for other variables (OR: 0.10, range 0.01-0.71; p=0.008). Our data establish severe GVHD as a clinically relevant post-transplant phenomenon in germline DDX41-associated HHMs and suggest that donor-derived wild-type T-cells may become activated by a pro-inflammatory milieu conferred by P/LP germline DDX41 variants. Moreover, our observations suggest that post-HSCT cyclophosphamide is an effective strategy for GVHD prevention in these individuals. Broader confirmation of our results in other heterogeneous patient cohorts will serve to expand the spectrum of known pathogenic variants implicated in HHMs and support the development of targeted interventions for those with deleterious germline HHM alleles. Finally, these results argue for germline genetic testing in patients prior to allogeneic HSCT. Figure 1View largeDownload PPTFigure 1View largeDownload PPT Close modal

Patients with Deleterious Germline Variants in STXBP2 Develop Toxicity after CAR-T Cell Therapy with Axicabtagene Ciloleucel

Patients with Deleterious Germline Variants in STXBP2 Develop Toxicity after CAR-T Cell Therapy with Axicabtagene Ciloleucel