Germline Sequencing Research Articles

Abstract 6109: Determinants of generalized self-efficacy and genetic knowledge among Hispanic/Latino colorectal cancer patients participating in ENLACE: A Cancer Moonshot Study

Abstract Background: Colorectal cancer (CRC) is the third most common cancer, and second cause of cancer death in the United States (US). Among US Hispanic/Latino/a/x (H/L) individuals, CRC represents the second and the third most common cancer and the most common cause of cancer death for men and women, respectively. Despite this, the tumor landscape, and key determinants of outcomes in CRC H/L patients are understudied. Methods: To address this need, we launched the ENLACE study, to engage H/L CRC patients in germline and somatic sequencing and identify the optimal approaches for patient participation. Through the Center for Patient Engagement in Cancer Characterization Studies (COPECC) patients were recruited from two healthcare facilities: a safety-net hospital (Los Angeles General Medical Center, LA Gen) and the USC Norris Comprehensive Cancer Center (Norris) a university medical center. Surveys to assess demographics, acculturation, Latino values, health literacy, discrimination, health system distrust, numeracy, and generalized self-efficacy (GSE) were evaluated upon recruitment, and after return-of-results (ROR) for somatic and germline testing. Results: After the first year of the project, a total of 100 H/L CRC patients >18 years of age were included in the ENLACE study, 89 participants completed baseline surveys (49 female, median age 53 (IQR = 16) and 21 completed questionnaires after ROR. A majority self-identified as White (61%), Mexico born (51%), catholic (74%), only spoke Spanish at home (35%) and married (49%). Higher levels of GSE were reported in patients with higher scores for numeracy (p = 0.005) and in males when compared to females (p = 0.016). This association was driven by participants from Norris (n=33) as reported by stratified analyses. In contrast, among participants from LA Gen (n=66) we observed an association with higher median values for the Mexican American acculturation scale and mainstream values and a marginally significant increase in the GSE (p = 0.072 & 0.083, respectively). Additionally, LA Gen participants with a greater than the median score for GSE had a marginally significant higher mean score in the KnowGene scale (p = 0.056) with a reduction in the number of times the “Don’t Know” option was selected (p = 0.045). No statistically significant difference in the generalized self-efficacy score and KnowGene scale were reported after reception of results (p = 0.2386 & 0.1337, respectively) Conclusion: We report differences in GSE and cancer genetic knowledge amongst CRC H/L patients in the ENLACE study that may inform understanding of disparities and social determinants in the populations treated at two different healthcare facilities. Citation Format: Joel Sanchez Mendez, Julie O. Culver, Charité N. Ricker, Natalia Gutierrez, Sandra Algaze, John D. Carpten, Heinz-Josef Lenz, Mariana C. Stern. Determinants of generalized self-efficacy and genetic knowledge among Hispanic/Latino colorectal cancer patients participating in ENLACE: A Cancer Moonshot Study [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2024; Part 1 (Regular Abstracts); 2024 Apr 5-10; San Diego, CA. Philadelphia (PA): AACR; Cancer Res 2024;84(6_Suppl):Abstract nr 6109.

Improving somatic exome sequencing performance by biological replicates

BackgroundNext-generation sequencing (NGS) technologies offer fast and inexpensive identification of DNA sequences. Somatic sequencing is among the primary applications of NGS, where acquired (non-inherited) variants are based on comparing diseased and healthy tissues from the same individual. Somatic mutations in genetic diseases such as cancer are tightly associated with genomic instability. Genomic instability increases heterogenity, complicating sequencing efforts further, a task already challenged by the presence of short reads and repetitions in human DNA. This leads to low concordance among studies and limits reproducibility. This limitation is a significant problem since identified mutations in somatic sequencing are major biomarkers for diagnosis and the primary input of targeted therapies. Benchmarking studies were conducted to assess the error rates and increase reproducibility. Unfortunately, the number of somatic benchmarking sets is very limited due to difficulties in validating true somatic variants. Moreover, most NGS benchmarking studies are based on relatively simpler germline (inherited) sequencing. Recently, a comprehensive somatic sequencing benchmarking set was published by Sequencing Quality Control Phase 2 (SEQC2). We chose this dataset for our experiments because it is a well-validated, cancer-focused dataset that includes many tumor/normal biological replicates. Our study has two primary goals. First goal is to determine how replicate-based consensus approaches can improve the accuracy of somatic variant detection systems. Second goal is to develop highly predictive machine learning (ML) models by employing replicate-based consensus variants as labels during the training phase.ResultsEnsemble approaches that combine alternative algorithms are relatively common; here, as an alternative, we study the performance enhancement potential of biological replicates. We first developed replicate-based consensus approaches that utilize the biological replicates available in this study to improve variant calling performance. Subsequently, we trained ML models using these biological replicates and achieved performance comparable to optimal ML models, those trained using high-confidence variants identified in advance.ConclusionsOur replicate-based consensus approach can be used to improve variant calling performance and develop efficient ML models. Given the relative ease of obtaining biological replicates, this strategy allows for the development of efficient ML models tailored to specific datasets or scenarios.

Abstract 6372: Germline pathogenic variants are enriched in pediatric central nervous system tumor patients and associate with somatic functional consequences

Abstract The genetic contribution of rare pathogenic germline variation to central nervous system (CNS) tumor formation in pediatric patients without a family history of cancer remains unclear. We sought to characterize the prevalence, spectrum, and clinical significance of pathogenic germline variation in cancer predisposition genes (CPGs) in 838 patients profiled in the Pediatric Brain Tumor Atlas (PBTA) by whole genome sequencing (n=821) or whole exome sequencing (n=17). Rare variants in 214 CPGs were annotated as pathogenic (P) or likely pathogenic (LP) consistent with American College of Medical Genetics criteria using AutoGVP, our open-source automated pathogenicity assessment tool. To assess enrichment of P/LP variants in the PBTA, we compared the frequency of CPG P/LP variants in cases to a tumor-free control cohort from Penn Medicine BioBank (PMBB; n=6,295). Somatic alterations, mutational signatures, and gene set enrichment analyses from matched tumor sequencing were integrated to characterize functional consequences associated with germline pathogenic variation. We observed 215 germline P/LP variants within 76 distinct CPGs in 189 (22.6%) PBTA patients. CPG P/LP variants were most prevalent among patients with a plexiform neurofibroma diagnosis (9/14, 64.3%), and least prevalent among patients with craniopharyngioma (3/39, 7.8%). We detected syndrome-associated P/LP variants in 45/58 (77.6%) patients with a clinically reported cancer predisposition syndrome and incidentally in another 37 patients. CPG P/LP variants were significantly enriched in the PBTA cohort relative to the PMBB control cohort (OR=2.0, p=1.0E-12, CI=1.6-2.4), and NF1 harbored the most significant excess P/LP variant burden (OR=30.7, p=4.57E-13, CI=11.2-105.1). We observed a significant enrichment of mismatch repair (MMR) gene P/LP variants in patients with high-grade gliomas (HGG) and tumors from MMR P/LP carriers exhibited significantly higher MMR-deficiency mutational signature exposures relative to tumors from non-carrier HGG patients (SBS14: W=478, p=1.71E-05). Overall, 90/215 (41.9%) of identified germline P/LP variants co-occurred with at least one corresponding somatic alteration in matched tumors, including oncogenic SNV (n=5), copy number variation (n=28), loss of heterozygosity (n=54), differential gene expression (n=23), and proximal alternative splicing (n=19). In summary, rare germline P/LP variants are enriched in PBTA patients and are associated with somatic events in the same gene, resulting in gene- and pathway-specific functional consequences. Our work emphasizes a role for pathogenic germline variation in pediatric CNS tumor development and frequent under-identification of cancer predisposition syndromes, underscoring the importance of germline sequencing in personalized patient care strategies. Citation Format: Ryan J. Corbett, Rebecca Kaufman, Shelly W. McQuaid, Zalman Vaksman, Saksham Phul, Miguel A. Brown, Jennifer L. Mason, Heena Desai, Ryan Hausler, Ammar S. Naqvi, Antonia Chroni, Zhuangzhuang Geng, Bo Zhang, Chuwei Zhong, Yuankun Zhu, Allison P. Heath, Marilyn Li, Penn Medicine BioBank, Phillip B. Storm, Adam C. Resnick, Kara N. Maxwell, Miriam Bornhorst, Kristina A. Cole, Angela J. Waanders, Suzanne P. MacFarland, Jo Lynne Rokita, Sharon J. Diskin. Germline pathogenic variants are enriched in pediatric central nervous system tumor patients and associate with somatic functional consequences [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2024; Part 1 (Regular Abstracts); 2024 Apr 5-10; San Diego, CA. Philadelphia (PA): AACR; Cancer Res 2024;84(6_Suppl):Abstract nr 6372.

Abstract 7334: Germline mutations in cancer susceptibility genes in patients with mucosal melanoma

Abstract Background: Mucosal melanoma (MM) is the rarest subtype of melanoma with markedly worse outcomes compared to cutaneous melanoma (CM). Unlike CM which is driven by somatic mutations due to UV exposure, risk factors for MM are less defined. While germline mutations in cancer susceptibility genes are well-studied in CM, they are less understood in MM. Herein, we aimed to identify the prevalence of germline mutations in cancer susceptibility genes in a large MM cohort. Methods: A cohort of 247 MM patients (pts) from MD Anderson Cancer Center with available blood for germline sequencing was identified. Pt characteristics were reflective of U.S. MM population with median age of 63, 60% females and 84.6% white. Clinicodemographic features of pts including tumor site, key somatic mutations (BRAF, NRAS, KIT) and personal history of other malignancies were abstracted, and their frequencies were compared in pts with and w/o germline mutation. Frequency of MITF and CHEK2 alleles were compared to expected population control rate based on TOPMED by Fisher’s exact test. Results: Among 247 pts, 10.9% had germline mutations, primarily in MITF (c.G952A:p.E318K) (2.4%) and CHEK2 (c.1100delC:p.T367fs) (1.6%) at OR of 13.9 (95%CI 5.0, 30.9) and 16.4 (4.4, 42.9) compared to population control rate (Table 1). MM sites of origin were gastrointestinal (36%), genitourinary (34%) and head & neck (30%). In terms of somatic mutations, KIT (26%) was most common, followed by NRAS (15%), non-V600E BRAF (6.5%) and BRAF V600E (2.4%). These variables did not differ between those with and w/o germline mutation. Pts with germline mutation had a higher incidence of non-melanoma malignancies (48% vs 29%; p=0.044). Table 1. Summary of identified germline mutations in the mucosal melanoma cohort. Germline mutation Protein MITF (N= 6) p.E318K CHEK2 (N=4) p.T367fs MUYTH p.G368D ATM splice ATM p.L263fs ATM p.R1875X ATM p.L1327X ATM p.A2067D ATM p.R2034X BRCA2 p.A1689fs CHEK2 p.S400T ERCC3 p.R109X MITF p.C29Y NF1 p.L1183R PALB2 p.V221X POLE p.D287E TP53 p.R306X TP53 p.V73Wfs*50 TSC1 p.Q844X Conclusions: This study reports a high prevalence of germline cancer susceptibility mutations in the largest cohort of MM to date, including mutations which are actionable. There is overlap with known MITF risk alleles for CM. We are currently performing a GWAS to comprehensively understand the genetic architecture of risk in MM. Citation Format: Afsaneh Amouzegar, Xiaogang Wu, James Long, Sabitha Prabhakaran, Latasha Little, Curtis Gumbs, Jared Malke, Julie Simon, Jianhua Zhang, Jennifer Wargo, Paul Scheet, Justin W. Wong, Priyadharsini Nagarajan, Jennifer L. McQuade, Andrew Futreal. Germline mutations in cancer susceptibility genes in patients with mucosal melanoma [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2024; Part 1 (Regular Abstracts); 2024 Apr 5-10; San Diego, CA. Philadelphia (PA): AACR; Cancer Res 2024;84(6_Suppl):Abstract nr 7334.

Diagnostic yield and clinical impact of germline sequencing in children with CNS and extracranial solid tumors—a nationwide, prospective Swedish study

Childhood cancer predisposition (ChiCaP) syndromes are increasingly recognized as contributing factors to childhood cancer development. Yet, due to variable availability of germline testing, many children with ChiCaP might go undetected today. We report results from the nationwide and prospective ChiCaP study that investigated diagnostic yield and clinical impact of integrating germline whole-genome sequencing (gWGS) with tumor sequencing and systematic phenotyping in children with solid tumors. gWGS was performed in 309 children at diagnosis of CNS (n=123, 40%) or extracranial (n=186, 60%) solid tumors and analyzed for disease-causing variants in 189 known cancer predisposing genes. Tumor sequencing data were available for 74% (227/309) of patients. In addition, a standardized clinical assessment for underlying predisposition was performed in 95% (293/309) of patients. The prevalence of ChiCaP diagnoses was 11% (35/309), of which 69% (24/35) were unknown at inclusion (diagnostic yield 8%, 24/298). A second-hit and/or relevant mutational signature was observed in 19/21 (90%) tumors with informative data. ChiCaP diagnoses were more prevalent among patients with retinoblastomas (50%, 6/12) and high-grade astrocytomas (37%, 6/16), and in those with non-cancer related features (23%, 20/88), and ≥2 positive ChiCaP criteria (28%, 22/79). ChiCaP diagnoses were autosomal dominant in 80% (28/35) of patients, yet confirmed de novo in 64% (18/28). The 35 ChiCaP findings resulted in tailored surveillance (86%, 30/35) and treatment recommendations (31%, 11/35). Overall, our results demonstrate that systematic phenotyping, combined with genomics-based diagnostics of ChiCaP in children with solid tumors is feasible in large-scale clinical practice and critically guides personalized care in a sizable proportion of patients. The study was supported by the Swedish Childhood Cancer Fund and the Ministry of Health and Social Affairs.

AutoGVP: a dockerized workflow integrating ClinVar and InterVar germline sequence variant classification.

With the increasing rates of exome and whole genome sequencing, the ability to classify large sets of germline sequencing variants using up-to-date American College of Medical Genetics-Association for Molecular Pathology (ACMG-AMP) criteria is crucial. Here, we present Automated Germline Variant Pathogenicity (AutoGVP), a tool that integrates germline variant pathogenicity annotations from ClinVar and sequence variant classifications from a modified version of InterVar (PVS1 strength adjustments, removal of PP5/BP6). This tool facilitates large-scale, clinically focused classification of germline sequence variants in a research setting. AutoGVP is an open source dockerized workflow implemented in R and freely available on GitHub at https://github.com/diskin-lab-chop/AutoGVP.

Experiences of pediatric cancer patients (age 12–18 years) with extensive germline sequencing for cancer predisposition: a qualitative study

This study explored the experiences and needs of adolescents, ranging from 12 to 18 years old, who have recently been diagnosed with cancer and participated in a nationwide germline genetic sequencing study within the context of pediatric oncology. The 21 adolescents in this qualitative interview study viewed genetic sequencing as an integral part of their cancer journey. They often characterized germline sequencing as “good-to-know” without specifying immediate utility. While the adolescents comprehended the significance of germline genetic sequencing, they were less focused on its potential long-term implications. Adolescents expressed a strong desire to be actively engaged in decisions related to genetics. They advocated for a participatory role in genetic decision-making from a young age onwards. They recommended that re-consent should be sought before re-analysis of their genetic data is performed and believe that patients should have the opportunity to provide (re-)consent once they reach adulthood. Moreover, the adolescents emphasized the importance of developing counseling materials that are not only concise but also visually attractive. In conclusion, this study underscores the positive perception that adolescents diagnosed with cancer hold regarding germline genetic sequencing. They articulate a strong interest in being actively involved in genetic decision-making. To address these articulated needs and preferences, we recommend the development of visually engaging counseling materials. These materials should effectively convey both the immediate and long-term implications of genetic sequencing, enabling adolescents with cancer to make informed decisions about genetic sequencing.

Germline sequencing in men with metastatic castration-resistant prostate cancer from the BARCODE2 study reveals a wide range of pathogenic variants in DNA repair genes

BackgroundThe presence of germline mutations plays an increasingly important role in risk assessment and treatment of prostate cancer (PrCa). Screening for high-risk mutations in subsets of patients is becoming routine. We explore the prevalence of germline genetic mutations in men with metastatic castration-resistant prostate cancer (mCRPC) recruited to the BARCODE2 trial.MethodsThe BARCODE2 trial is a two-part study investigating the response to carboplatin chemotherapy in mCRPC patients carrying a germline variant in a DNA repair gene (DRG). We report interim data from Part 1, in which participants are recruited for germline genetic testing using a customised next-generation sequencing panel consisting of 115 genes.ResultsThese interim results (N = 220) demonstrate a similar frequency of germline DRG variants in mCRPC patients compared with previously published data (15% detection rate). No significant clinical differences were identified between all carriers and non-carriers, though BRCA2/ATM carriers were found to have a shorter time to mCRPC diagnosis.ConclusionsGermline pathogenic/likely pathogenic (P/LP) variants in BRCA2 and ATM genes are associated with a shorter time to progression and rarer P/LP variants in other DRG genes may play a role in mCRPC. This justifies the use of routine screening of men with advanced PrCa for germline variants and supports the need for an expanded panel test.

The survival outcomes for men with metastatic castration-resistant prostate cancer (mCRPC) with and without homologous recombination deficiencies (HRD) treated with radium-223: Princess Margaret Cancer Centre (PMCC) experience.

179 Background: Radium-223 (223Ra) is a radiopharmaceutical that emits alpha particles and specifically targets bone metastases in mCRPC, where it has been shown to improve overall survival (OS). Since 223Ra induces double-strand DNA breaks, we hypothesized that patients with HRD may exhibit heightened sensitivity to 223Ra, resulting in improved survival outcomes compared to patients without HRD. Methods: This retrospective analysis was performed in men with mCRPC and bone metastases, with and without HRD, treated with 223Ra at PMCC. Demographics and disease characteristics were collected. Germline and/or somatic DNA sequencing data were identified. OS and progression free survival (PFS) were calculated using the Kaplan–Meier method; differences in outcomes were assessed using the log-rank test. Alkaline phosphatase (ALP) and prostate specific antigen (PSA) responses were calculated at 12 weeks post 223Ra treatment. Results: We identified 40 mCRPC patients who had germline and/or somatic DNA sequencing and received 223Ra between December 2015 and May 2022. The median age at the start of 223Ra was 76.5 (range: 66.5-80.8), and ECOG was 0/1 (75%). A total of 29 (72.5%) patients received 233 Ra following abiraterone or enzalutamide treatment while 10 (25%) received 233 Ra post docetaxel. Overall, 22 (54%) received ≥4 cycles of ²²³Ra. Median baseline PSA was 58.6 (range: 25.2-143) and median baseline ALP was 109 (range: 71.8-200). Germline/somatic HRD mutations were found in 9/40 (22.5%) patients (BRCA2 [n=6], CHEK2 [n=2], CDK12 [n=1]). Baseline characteristics were well balanced between HRD and non-HRD groups. With median follow up of 13.7 months, the median OS of HRD group vs. non-HRD group was 24 months (95%CI: 14-not evaluable [NE]) vs. 12 months (95%CI: 7-22); p=0.038). The median PFS of HRD group vs. non-HRD group was 5.7 months (95%CI: 3-NE) vs. 3.3 months (95%CI: 2.5-13.4); p=0.74). The median time to the next treatment of HRD group vs. non-HRD group was 4.2 months (95%CI: 3.1-NE) vs. 3.75 months (95%CI: 3.5-13.5); p=0.89. ALP response was 66.7% of HRD group vs. 58.1% of non-HRD group; p=0.72. PSA response was 33.3% of HRD group vs. 9.7% of non-HRD group; p=0.11. For all patients with ALP response, the three years survival probability of HRD group vs. non-HRD group was 33% vs. 11%; p=0.03. Conclusions: While the number of patients included in our review was small, our analysis suggested that patients with HRD may have a slight improvement in OS after 223Ra treatment. Validation in a prospective dataset is required, and whether HRD status has implications for other radiopharmaceuticals such as lutetium-177 remains to be seen.

Feasibility and Clinical Utility of Reporting Hereditary Cancer Predisposition Pathogenic Variants Identified in Research Germline Sequencing: A Prospective Interventional Study.

Patients with cancer frequently undergo research-grade germline sequencing but clinically actionable results are not routinely disclosed. The objective of this study is to evaluate the feasibility of reporting clinically relevant secondary findings (SF) identified in germline research sequencing using the institutional molecular tumor board (MTB) and the treating oncology physician. This prospective, interventional cohort study enrolled Total Cancer Care participants with any cancer diagnosis at a single institution. Patients underwent research-grade germline whole-exome sequencing, with bioinformatic analysis in a Clinical Laboratory Improvement Amendments-certified laboratory to verify pathogenic/likely pathogenic germline variants (PGVs) in any American College of Medical Genomics and Genetics SF v2.0 genes. After a protocol modification in consenting patients, the MTB reported PGVs to treating oncology physicians with recommendations for referral to a licensed genetic counselor and clinical confirmatory testing. Of the 781 enrolled participants, 32 (4.1%) harbored cancer predisposition PGVs, 24 (3.1%) were heterozygous carriers of an autosomal recessive cancer predisposition syndrome, and 14 (1.8%) had other hereditary disease PGVs. Guideline-directed testing would have missed 37.5% (12/32) of the inherited cancer predisposition PGVs, which included BRCA1, BRCA2, MSH6, SDHAF2, SDHB, and TP53 variants. Three hundred fifteen participants consented to reporting results; results for all living patients were reported to the clinical team with half referred to a licensed genetic counselor. There was concordance between all research variants identified in patients (n = 9) who underwent clinical confirmatory sequencing. MTB reporting of research-grade germline sequencing to the clinical oncology team is feasible. Over a third of PGVs identified using a universal testing strategy would have been missed by guideline-based approach, suggesting a role for expanding germline testing.

Abstract C104: Dual primary pancreas cancers – Related or independent lesions?

Abstract Background: Not infrequently, an apparent second pancreas ductal adenocarcinoma (PDAC) is identified either synchronously or metachronously. To date, it remains unclear whether these individual PDACs represent intrapancreatic metastases or a new primary PDAC. Here, we sought to understand the clinicogenomic features of this patient population to provide insights on the underlying biology and management implications. Methods: Memorial Sloan Kettering Cancer Center (MSK) institutional databases were searched for patients with two PDAC diagnoses and at least one pancreatic resection or patients who had undergone completion/total pancreatectomy and then manually curated. Detailed demographic and clinical information were abstracted from the medical record. Metachronous patients excluded if previous surgical margin positive for invasive or pre-invasive carcinoma. All patients were either consented to MSK IMPACT for somatic and germline sequencing (N=16), were approved for de-identified genomic analyses by IRB (N=3), or were excluded from genomic analysis (N=2). KRAS wild-type (WT) tumors were interrogated for targetable fusions with Archer FusionPlex solid tumor panel. Results: N=21 patients (pts) identified, N=6 synchronous, N=15 metachronous. Mean age at diagnosis of first PDAC (PDAC1): 67.1y (51.2–79.1y). PDAC risk factors: 52% (10/19) smoking history, 15% (2/13) with first degree relative with PDAC, 85% (11/13) with first degree relative with any malignancy, and 40% (8/20) with history of premalignant pancreas lesion. N=2 pts with pathogenic germline mutation in ATM. By morphology, 80% (4/5) of synchronous and 65% (11/17) of asynchronous lesions indistinguishable. Two lesions, both metachronous, demonstrated divergent histology while remaining cases demonstrated focal differences. Genomically, 40% (10/25) samples with mKRAS G12V, G12R (16%), G12D (12%), Q61H (8%), G12C (4%); 20% KRAS WT. Of available paired tumor samples, 71% (5/7) with identical KRAS alleles and additional driver mutations. N= 1 pt with different KRAS alleles also had divergent TP53 alleles and CDKN2A/B loss occur in metachronous lesions 14 m apart. N=1 pt with KRAS WT tumor had an actionable fusion involving FGFR2. For metachronous lesions, median interval PDAC1 to PDAC2: 5.1 y (1.1 – 15.2 y). At median follow-up of 7.2 y (0.2 – 15.4 y) from PDAC1, 90% (19/21) have undergone completion/total pancreatectomy and N=16 (76%) alive with 5-year survival rate of 93% (14/15). Conclusions: The clinical, pathologic, and genomic data strongly suggest that two PDAC, independent of disease interval, most often represent the same phylogenetic entity. KRAS allele variant adjudication was the single most important discriminator and was in concordance with other driver oncogenes. Pathogenic germline variants were rare and limited to gATM; notably no gBRCA/PALB2. Acknowledging the highly selected nature of this cohort, solitary intrapancreatic metastases demonstrate a more favorable biology and these patients may benefit from personalized management approaches beyond traditional paradigms. Citation Format: Joshua D. Schoenfeld, Hulya Sahan-Ozkan, Nadeem Bilani, Michael I. D'Angelica, William R. Jarnagin, Jeffrey Drebin, Diane Reidy-Lagunes, Alice C. Wei, Sree B. Chalasani, Anna M. Varghese, Fiyinfolu Balogun, Wungki Park, Kenneth H. Yu, Olca Basturk, Eileen M. O'Reilly. Dual primary pancreas cancers – Related or independent lesions? [abstract]. In: Proceedings of the AACR Special Conference in Cancer Research: Pancreatic Cancer; 2023 Sep 27-30; Boston, Massachusetts. Philadelphia (PA): AACR; Cancer Res 2024;84(2 Suppl):Abstract nr C104.

Abstract IA15: What the surgeon has learned from pancreas cancer genomics, and vice versa

Abstract Recent advances in molecular profiling and fundamental science discoveries are impacting care of PDAC patients, including surgical decision making. Our multidisciplinary group at the Ontario Institute for Cancer Research and the Princess Margaret in Toronto has focused on elucidating both germline and somatic genomic changes in PDAC, with aims to understand genetic predisposition to the disease, foster early detection and conduct trials for improved selection of standard and novel therapies. Using laser capture microdissection overseen by expert pathologists, seamless tumor banking and aggressive biopsy of chemo-naïve and progressive metastases, we have generated high epithelial tumor cell content whole genomes and transcriptomes (WGTS) on over 600 PDAC patients with all stages of the disease. Germline sequencing of all PDAC patients should be considered standard of care to detect patients with uniquely actionable genomic changes due to less common mutational pathways. The genetic basis of familial pancreas cancer remains elusive and we continue to collaborate with the PACGENE Network spearheaded by colleagues at Mayo, Hopkins and McGill to discover novel genes in high risk cases. Our genomic work and others explains the clinical conundrum of late presentation of PDAC due to rapid punctuated evolution which challenges the ‘holy grail’ of early detection. Thus, screening high risk groups is still within the realm of investigational studies such as the PRECEDE consortium, of which we are an active member. Focused clinical trials (eg COMPASS and PASS-01) using WGTS has revealed both large scale changes in genomes (eg. HRD, tandem duplicators, etc) and transcriptomes (the Moffit ‘continuum’) and discreet mutations (eg BRAF mutations in Kras wild type tumors) that are important to identify as early as possible in the patient’s clinical course. Subspecialty PDAC surgeons need to keep abreast of recent developments of both prognostic and predictive transcriptomic biomarkers that will soon be important in selecting appropriate patients for appropriate neoadjuvant therapies. A few clinical vignettes will be shown to provide evidence for the importance of WGTS and other profiling approaches for almost all PDAC patients. Citation Format: Steven Gallinger. What the surgeon has learned from pancreas cancer genomics, and vice versa [abstract]. In: Proceedings of the AACR Special Conference in Cancer Research: Pancreatic Cancer; 2023 Sep 27-30; Boston, Massachusetts. Philadelphia (PA): AACR; Cancer Res 2024;84(2 Suppl):Abstract nr IA15.

Comparison of immediate germline sequencing and multi-step screening for Lynch syndrome detection in high-risk endometrial and colorectal cancer patients.

Lynch syndrome (LS) is a hereditary cancer predisposition syndrome with a significantly increased risk of colorectal and endometrial cancers. Current standard practice involves universal screening for LS in patients with newly diagnosed colorectal or endometrial cancer using a multi-step screening protocol (MSP). However, MSP may not always accurately identify LS cases. To address this limitation, we compared the diagnostic performance of immediate germline sequencing (IGS) with MSP in a high-risk group. A total of 31 Taiwanese women with synchronous or metachronous endometrial and colorectal malignancies underwent MSP which included immunohistochemical staining of DNA mismatch repair (MMR) proteins, MLH1 promoter hypermethylation analysis, and germline sequencing to identify pathogenic variants. All patients who were excluded during MSP received germline sequencing for MMR genes to simulate IGS for the detection of LS. Our findings indicate that IGS surpassed MSP in terms of diagnostic yield (29.0% vs. 19.4%, respectively) and sensitivity (90% vs. 60%, respectively). Specifically, IGS successfully identified nine LS cases, which is 50% more than the number detected through MSP. Additionally, germline methylation analysis revealed one more LS case with constitutional MLH1 promoter hypermethylation, bringing the total LS cases to ten (32.3%). Intriguingly, we observed no significant differences in clinical characteristics or overall survival between patients with and without LS in our cohort. Our study suggests that IGS may potentially offer a more effective approach compared to MSP in identifying LS among high-risk patients. This advantage is evident when patients have been pre-selected utilizing specific clinical criteria.

Genomic ascertainment of primary central nervous system cancers in adolescents and young adults.

Genomic ascertainment is the inversion of the traditional phenotype-first approach; with a "genome-first" approach, a cohort linked to electronic health records (EHR) undergoes germline sequencing (array, panel, exome, and genome) and deleterious variation of interest in a gene (or set of genes) are identified. Phenotype is then queried from the linked EHR and from call-back investigation and estimates of variant prevalence, disease penetrance, and phenotype can be determined. This should permit a better estimate of the full phenotypic spectrum, severity, and penetrance linked to a deleterious variant. For now, given the modest size, limited EHR, and age of participants in sequenced cohorts, genomic ascertainment approaches to investigate cancer in children and young adults will likely be restricted to descriptive studies and complement traditional phenotype-first work. Another issue is the ascertainment of the cohort itself: Participants need to survive long enough to enroll. Not accounting for this may lead to bias and incorrect estimates of variant prevalence. Adult-focused cohorts with EHR extending back into childhood, linked to cancer registries, and/or studies that permit recontact with participants may facilitate genomic ascertainment in pediatric cancer research. In summary, genomic ascertainment in pediatric primary brain cancer research remains largely untapped and merits further investigation.

Medulloblastomas with ELP1 pathogenic variants: A weakly penetrant syndrome with a restricted spectrum in a limited age window.

ELP1 pathogenic variants (PV) have been recently identified as the most frequent variants predisposing to Sonic Hedgehog (SHH) medulloblastomas (MB); however, guidelines are still lacking for genetic counseling in this new syndrome. We retrospectively reviewed clinical and genetic data of a French series of 29 ELP1-mutated MB. All patients developed SHH-MB, with a biallelic inactivation of PTCH1 found in 24 tumors. Other recurrent alterations encompassed the TP53 pathway and activation of MYCN/MYCL signaling. The median age at diagnosis was 7.3 years (range: 3-14). ELP1-mutated MB behave as sporadic cases, with similar distribution within clinical and molecular risk groups and similar outcomes (5 y - OS = 86%); no unusual side effect of treatments was noticed. Remarkably, a germline ELP1 PV was identified in all patients with available constitutional DNA (n = 26); moreover, all tested familial trio (n = 11) revealed that the PVs were inherited. Two of the 26 index cases from the French series had a family history of MB; pedigrees from these patients and from 1 additional Dutch family suggested a weak penetrance. Apart from MB, no cancer was associated with ELP1 PVs; second tumors reported in 4 patients occurred within the irradiation fields, in the usual time-lapse for expected radiotherapy-induced neoplasms. The low penetrance, the "at risk' age window limited to childhood and the narrow tumor spectrum, question the actual benefit of genetic screening in these patients and their family. Our results suggest restricting ELP1 germline sequencing to patients with SHH-MB, depending on the parents" request.

BIOM-12. ULTRA-DEEP WHOLE GENOME SEQUENCING UNCOVERS A HIGH PREVALENCE OF GLIOBLASTOMA-DERIVED CELL-FREE DNA IN PLASMA

Abstract The detection of glioblastoma-derived circulating cell-free DNA (ccfDNA) has proven formidable because tumor-specific mutations are hidden among the abundance of ccfDNA from healthy cells. While capture-enrichment panels provide the necessary read depth to detect low-frequency variants, searches are constrained to a narrow range of targets and largely limited to single nucleotide variants (SNVs). Here, we use ultra-deep whole genome sequencing (WGS) to broaden the search for somatic mutations across the entire genome. In addition, a kmer-based reference-free variant caller, which mitigates the bias and errors that accrue during alignment, is used to extend the mutation search beyond SNVs to include insertions/deletions (INDELS) and structural variants (SVs) such as large INDELS ( >20 bp) and tandem duplications. In tumor DNA and ccfDNA, we achieve a WGS (GRCh38) mean read depth of 263X and 225X, respectively. WGS of matched buffy coat DNA and sequencing reads from the 1000 Genome Database were used to eliminate germline polymorphisms, sequencing artifacts, and common variants. We detected 20,488 somatic mutations in tumor DNA of which 17.3% affected a coding sequence, 31.9% were INDELS, 91 were large INDELs, and 18 were tandem duplications. A tumor-informed search identified 489 of these somatic mutations in ccfDNA which included a large INDEL and a similar portion of mutations affecting a coding sequence. Using an unguided search for glioblastoma-derived ccfDNA uncovered an additional 8,794 somatic mutations of which 16.6% involved a coding sequence, 60.7% were INDELS, 30 were large INDELS, and 3 were tandem duplications. The high prevalence of somatic mutations discovered in ccfDNA using an unbiased approach highlights the low yield of tumor-informed searches in spatially genetically heterogeneous cancers like glioblastoma. Our findings demonstrate that ultra-deep WGS detects glioblastoma-derived ccfDNA by broadening the search for variants across the entire genome and through the inclusion of INDELS and SVs.

Investigating the Influence of Germline ATM Variants on Susceptibility to Chronic Lymphocytic Leukemia and Other Malignancies

Introduction: Chronic lymphocytic leukemia (CLL) patients (pts) face an increased susceptibility to secondary cancers. Furthermore, the hereditary predisposition to CLL is influenced by factors that have yet to be fully elucidated by GWAS-identified common variants. Among the hereditary cancer risk mutations observed, germline Ataxia-telangiectasia mutated ( ATM) aberrations are among the most prevalent. Rare germline ATM variants are detected in 24% of CLL pts, surpassing the frequency observed in other hematologic neoplasms and the general population. In CLL, loss of ATM via 11q23 deletion is linked to an unfavorable prognosis and a shortened progression-free survival. The extent to which germline ATM variants predispose CLL pts to other malignancies remains unclear; to examine this potential risk we conducted a retrospective study to assess the influence of germline ATM variants on the predisposition to secondary neoplasms in CLL pts and their relatives. Methods: Five hundred and eighty-five pts seen at Dana-Farber Cancer Institute and who had NGS performed to evaluate germline ATM status, either through direct germline sequencing of saliva or by inference according to the hierarchical algorithm we have previously published (Lampson, 2022), were mailed a questionnaire that investigated: demographics; personal and family history of any cancer; non-medical radiation and Agent Orange exposure; and Ataxia-Telangiectasia syndrome (AT). Of these, 333 replied (57%). Information collected from our CLL database included FISH, IGHV status and treatment status. European ancestry was categorized based on the United Nations geoscheme (UNSD, 2019). Pt characteristics are in Table 1. Pts were stratified into 4 groups, based on ATM mutational status and del(11q): Group-1, ATM germline variants alone (N=63); Group-2, ATM germline with somatic ATM mutation and/or del(11q)(N=22); Group-3, ATM somatic aberration alone (including del(11q))(N=41); and Group-4, no ATM aberration (N=207). Results: Of the 333 pts in our cohort, baseline demographic and clinical characteristics among the 4 groups were balanced, except for unmutated IGHV status. Nineteen (86%) and 36 (86%) pts of groups 2 and 3, carrying somatic ATM aberrations, had unmutated IGHV, compared to 21 (33%) and 76 (37%) of groups 1 and 4, without somatic ATM aberrations(p<0.01). Regarding treatment, 60% of our cohort had received at least one line of therapy (median: 1 (1-7)). Among the 164 pts (49%) with history of an additional non-CLL neoplasm, 221 cancers were reported. Excluding pts with only non-melanoma skin cancers, the incidence is reduced to 31% (104 pts). The median number of second cancers per pt is 1 (1-4). Of the 134 pts with available age of onset data for the secondary malignancy, 80 (60%) were diagnosed with the secondary cancer after CLL. Comparing pts with germline ATM variants (N=85) to the ones without germline ATM variants (N=248), no significant difference in the incidence of secondary cancer was found (p=0.73). The median age of secondary cancer onset was also similar between the groups (median: 63 and 64 years in germline ATM present vs absent, p=0.28). Of the whole cohort, 283 pts (85%) reported at least one relative with a cancer history (median 2 relatives affected (1-17)). The incidence of B-cell lymphoproliferative disorders was significantly higher (p=0.02) in pts with germline ATM variants (32%) compared to those without germline ATM variants (21%), including familial CLL (25% vs18%) (p=0.04). Familial CLL was also strongly associated with Ashkenazi Jewish ethnicity (39% incidence in Ashkenazi pts vs 16% in non-Ashkenazi, p=0.004). No difference in the incidence of other hematologic or solid malignancies was found between the relatives in the two groups. No pts reported a familial or personal history of AT. Compared to pts without any ATM aberration, time to first treatment (TTFT) was shorter in pts harboring somatic ATM events, while no difference was observed between pts with germline ATM variants (median TTFT: 82, 59, 52, 90 months for group 1, 2, 3 and 4 respectively, p=0.01) (Figure 1). Conclusions: Our results suggest a higher incidence of B-cell lymphoproliferative disorders, including familial CLL, in the relatives of CLL pts carrying germline ATM variants. The presence of these germline variants did not impact TTFT, compared to pts harboring somatic ATM mutations.

MO10-5 Germline and somatic whole exome sequencing identifies a novel candidate gene in double primary BCRA-associated cancers

MO10-5 Germline and somatic whole exome sequencing identifies a novel candidate gene in double primary BCRA-associated cancers

Only 32.3% of Breast Cancer Families with Pathogenic Variants in Cancer Genes Utilized Cascade Genetic Testing.

Hereditary cancer predisposition syndromes are responsible for approximately 5-10% of all diagnosed cancer cases. In order to identify individuals at risk in a cost-efficient manner, family members of individuals carrying pathogenic alterations are tested only for the specific variant that was identified in their carrier relative. The purpose of this study was to investigate the clinical use and implementation of cascade family testing (CFT) in families of breast cancer patients with pathogenic/likely pathogenic variants (PVs/LPVs) in cancer-related predisposition genes. Germline sequencing was carried out with NGS technology using a 52-gene panel, and cascade testing was performed by Sanger sequencing or MLPA. In a cohort of 1785 breast cancer patients (families), 20.3% were found to have PVs/LPVs. Specifically, 52.2%, 25.1%, and 22.7% of patients had positive findings in high-, intermediate-, and low-penetrance breast cancer susceptibility genes, respectively. Although CFT was recommended to all families, only 117 families (32.3%) agreed to proceed with genetic testing. Among the first-degree relatives who underwent CFT, 70.3% were female, and 108 of 121 (89.3%) were cancer free. Additionally, 42.7%, 36.7%, and 20.6% were offspring, siblings, and parents of the subject, respectively. Our data suggest that CFT was mostly undertaken (104/117, 88.8%) in families with positive findings in high-risk genes. Cascade family testing can be a powerful tool for primary cancer prevention by identifying at-risk family members. It is of utmost importance to implement genetic counseling approaches leading to increased awareness and communication of genetic testing results.

Whole exome germline sequencing in early-onset prostate cancer patients: Genomic findings and clinical outcomes.

Whole exome sequencing (WES) furthered our understanding of various tumors. We assessed the occurrence of germline likely pathogenic/pathogenic (LP/P) variants, disease features, and clinical outcomes in early-onset prostate cancer. This retrospective study (N = 134) included consecutive prostate cancer patients who donated blood samples for research purposes to the Kahn-Sagol-Maccabi biobank. Patients diagnosed at >65 years were excluded. Clinical characteristics were extracted from the medical records. Germline WES was performed with analysis reporting on oncogenetic, two immunogenic, and a secondary minimum list panels (121, 468, 76, and 59 genes, respectively). Median age at diagnosis was 61 (range 46-65) years; 131 (98%) were diagnosed with local disease. The median follow-up time from diagnosis was 14 (range <1-25) years. Of the patients with local disease, 32 (24%) and 10 (8%) had biochemical and distant recurrences, respectively. Twenty-five patients (19%) had ≥1 additional cancer (excluding non-melanoma skin cancer), most frequently bladder (6), colorectal (5), and lymphoma (5). Seven (5%) deaths were reported, with only one related to prostate cancer. LP/P variants were identified in 8 patients (6%), all in genes from the oncogenetic panel: ATM, BRCA1 (in two patients), BRCA2 (in two patients), HOXB13, MUTYH, and MYH7. Of these eight patients, with a median follow-up of 7 years (range <1-15), two (25%) had biochemical recurrences, one had (12.5%) distant recurrence, and no deaths were reported. In this cohort of 134 early-onset prostate cancer patients, we identified germline LP/P variants in an oncogenetic panel in 6% of participants, with no unique clinical outcome.