Genet Med Research Articles

Abstract 7325: Integration of functional data to classify BRCA1/2 missense variants: An ENIGMA project

Abstract The success of precision cancer prevention and treatment hinges on accurate discrimination between benign and pathogenic germline alleles in cancer susceptibility genes. Variants of uncertain clinical significance (VUS) present a challenge for cancer risk assessment and functional data will be essential to resolve many VUS. To address this, we developed a cloud-based environment to collate, curate, integrate and analyze all published functional data related to BRCA1 and BRCA2 missense VUS. Then, functional evidence criteria for pathogenicity were determined for each assay using the American College of Medical Genetics and Genomics/Association for Molecular Pathology (ACMG/AMP) classification guidelines and ClinGen criteria. Our approach involved retrieving published articles reporting one or more functional assays of BRCA1/2 missense variants reporting variant impact on various biochemical and cell biological functions. Functional results were recorded using original authors' thresholds and categories, then converted to ordinal variables to harmonize: [0 = no functional impact], [1 = intermediate impact], and [2 = functional impact]. For BRCA1, we updated our previous published integration (Genet Med. 2021;23:306-315) with 14 additional publications, collating results from 53 instances of functional assays reporting functional data on 3,219 unique missense variants. For BRCA2, 28 publications were identified reporting data from 141 instances of functional assays on 4,759 unique missense variants. Utilizing a panel of 542 and 396 known pathogenic and benign reference variants for BRCA1 and BRCA2, respectively, we determined the sensitivity, specificity, and ACMG/AMP odds of pathogenicity for each instance of the assay. The remaining variants were assigned a relevant functional assay code weight for each assay instance. Our study successfully derived ACMG/AMP evidence strength from functional data for 3,040 BRCA1 (94% of 3,219) and 2,704 BRCA2 (57% of 4,759) missense variants. This comprehensive work underscores the utility of functional data in informing classification of BRCA1 and BRCA2 missense VUS. Citation Format: Paulo Lyra, Lucas Dalcolmo, Michael Parsons, Thales Nepomuceno, Samuel Brito, Nam Phuong N. Nguyen, Geise de Oliveira, Joao Paulo da Siva, Laura Caleca, Tanisha Taneja, Chunling Hu, Marcy Richardson, Maria Rossing, Aditi Hazra, Alexandra Martins, Sandrine Caputo, Gael A. Millot, Joanne NgeowYuen Yie, Marcelo A. Carvalho, Melissa Cline, Paolo Radice, Rachael Carlsen, Romy Mesman, Valentina Zampiga, Rehan Villani, Vijay Josef, Shyam Sharan, Kyriaki Michailidou, Amanda B. Spurdle, Fergus Couch, Maaike P.G. Vreeswijk, Alvaro N. Monteiro. Integration of functional data to classify BRCA1/2 missense variants: An ENIGMA project [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 7325.

Whole Exome Sequencing of Patients with Adult-Onset Evans Syndrome: A Cohort of 80 Patients

Introduction Loss of tolerance toward several self-antigens suggests an underlying genetic predisposition. A previous study (1) conducted in children with Evans syndrome revealed a high prevalence of pathogenic mutations in genes involved in primary immunodeficiencies. Whether these findings are restricted to children is unknown. In this study, we assessed the prevalence of constitutional genetic variants in a cohort of patients with adult onset Evans syndrome. Patients and methods We consecutively included from November 2021 to June 2023 patients that had been diagnosed with adult onset (≥18 years) Evans syndrome (association of immune thrombocytopenia (ITP), warm autoimmune hemolytic anemia (wAIHA) and/or autoimmune neutropenia (AIN) according to international guidelines (2,3)), and identified in France throughout the network of the national reference center for adult' autoimmune cytopenia. We excluded patients with a systemic lupus (according to Systemic Lupus International Collaborating Clinics criteria (4)), and patients with indolent B cell malignancies at Evans syndrome onset. Clinical and biological data were retrospectively collected using a standardized form. Patients underwent whole-exome sequencing after written and informed consent was obtained. Gene variants already published and/or with in vitro confirmed consequences were considered as genetic diagnoses. Clinical interpretation of variants was performed according to the ACMG guidelines (5). A patient was classified as having a genetic diagnosis if carryinghemizygous/heterozygous pathogenic or likely pathogenic variant (for X-linked or autosomal dominant conditions), or 2 pathogenic or likely pathogenic variants in the same gene (for autosomal recessive conditions). Results We identified 80 patients (40% female, median age 31 years at Evans syndrome diagnosis [range 18-76]) with adult-onset Evans syndrome from 19 centers. Median age at first cytopenia was 28 years [range 1-73], and 13 patients (16%) were under 18 years at first cytopenia. 78 patients (96%) had ITP (chronic in 87%), 59 (73%) had wAIHA (with at least one relapse in 58%), and 38 (47%) had AIN (chronic in 74%). Median delay between first and second cytopenia was 18 months (range 0 - 329) and 28 patients (35%) had synchronous onset of cytopenia. Twenty-four patients (30%) had associated immune manifestation (involving endocrine system in 9%, lung in 9%, gastrointestinal tract in 6%, skin in 5%, central nervous system in 4%) and 28% had antinuclear antibody titers ≥ 1/160. Lymphoproliferation was found in 56%, including lymphadenopathy in 41% and splenomegaly in 35%. Nine patients (11%) had lymphoma or cancer history. Twenty patients (25%) had humoral immune defect, including 13 (16%) with a common variable immunodeficiency (CVID)-like phenotype. In the whole cohort, 9 patients (11.25%) had a genetic diagnosis that could be linked to Evans syndrome, with heterozygous variants in CTLA4, STAT3, NFKB1, CYBB, TNFAIP3, DDX41, MYH9 and homozygous variant in PIK3CG. Twenty (25%) patients had genetic testing prior to this study (targeted Sanger sequencing [N=3], or NGS panel [N=17]), resulting in 2 genetic diagnoses ( CYBB and TNFAIP3). A family history of immune manifestations in first-degree relatives was found in 56% of the patients with genetic diagnosis versus 20% of patients without (P=0.0308). However, the proportion of patients with CVID-like phenotype, associated immune disorder, lymphoproliferation, cancer/lymphoma, positive antinuclear antibodies, or pediatric onset of first cytopenia was not significantly increased in patients with genetic diagnosis. Discussion As expected, the proportion of genetic diagnoses in patients with adult onset Evans syndrome was lower that what was previously described in children, but led to a significant impact on clinical management in some patients. We did not find clinical or biological features associated with a genetic diagnosis, suggesting that genetic testing should be considered in every adult patient diagnosed with Evans syndrome in the absence of any obvious underlying cause or condition. References (1) Hadjadj J et al., Blood 2019; 134: 9-21. (2) Provan D et al., Blood Adv 2019; 3: 3780-817. (3) Jäger U et al., Blood Rev 2020; 41: 100648. (4) Petri M et al., Arthritis Rheum 2012; 64: 2677-86. (5) Richards S et al., Genet Med 2015; 17: 405-24.

EDITORIAL COMMENT

Paragangliomas and pheochromocytomas (PGL/PCC) are rare tumors that are derived from neural crest chromaffin cells either from neuroendocrine tissues distributed along the spine or from within the adrenal gland respectively. Approximately 40% of cases are associated with pathogenic germline variants in genes associated to cancer susceptibility syndromes, such as MAX, NF1, RET, SDHA, and SDHAF2, including several gene that associated with inherited kidney cancer, such as VHL, SDHB, SDHC, SDHD, and TMEM127. 1 Shuch B Ricketts CJ Metwalli AR Pacak K Linehan WM. The genetic basis of pheochromocytoma and paraganglioma: implications for management. Urology. 2014; 83: 1225-1232 Google Scholar ,2 Dahia PL. Pheochromocytoma and paraganglioma pathogenesis: learning from genetic heterogeneity. Nat Rev Cancer. 2014; 14: 108-119 Google Scholar A recent addition to this list is germline alteration of the FH gene, known to cause hereditary leiomyomatosis and renal cell cancer (HLRCC). 3 Richter S Gieldon L Pang Y et al. Metabolome-guided genomics to identify pathogenic variants in isocitrate dehydrogenase, fumarate hydratase, and succinate dehydrogenase genes in pheochromocytoma and paraganglioma. Genet Med. 2019; 21: 705-717 Google Scholar , 4 Clark GR Sciacovelli M Gaude E et al. Germline FH mutations presenting with pheochromocytoma. J Clin Endocrinol Metab. 2014; 99: E2046-E2050 Google Scholar , 5 Muller M Ferlicot S Guillaud-Bataille M et al. Reassessing the clinical spectrum associated with hereditary leiomyomatosis and renal cell carcinoma syndrome in French FH mutation carriers. Clin Genet. 2017; 92: 606-615 Google Scholar

Thrombosis in hereditary hemorrhagic telangiectasia

Thrombosis in hereditary hemorrhagic telangiectasia

The Reckoning: The Return of Genomic Results to 1444 Participants Across the eMERGE3 Network

(Abstracted from Genet Med 2022;24:1130–1138) As increased accessibility to genetic testing contributes to finding more individuals with genetic predispositions to a specific conditions or diseases, National Institutes of Health–sponsored networks grow in importance when considering the identification and subsequent management of these conditions. The American College of Genetics and Genomics (ACMG) designated 59 genes (this is now higher with subsequent versions of the guideline) as highly actionable owing to availability of effective medical management guidelines for individuals with detected disease-causing variants in one of these genes.

LIVING WITH EHLERS-DANLOS SYNDROME: FOLLOWING A YOUNG BOY THROUGH HIS UPS AND DOWNS ON HIS JOURNEY INTO ADULTHOOD

LIVING WITH EHLERS-DANLOS SYNDROME: FOLLOWING A YOUNG BOY THROUGH HIS UPS AND DOWNS ON HIS JOURNEY INTO ADULTHOOD

Stewardship of Patient Genomic Data: A Policy Statement of the American College of Medical Genetics and Genomics (ACMG)

(Abstracted from Genet Med 2022;24:509–511) The need for genomic research is a firmly held value of the American College of Medical Genetics and Genomics (ACMG), as this can lead to discovery for the improvement of health outcomes and patient experience. However, an exchange of deidentified genomic data presents a potential opportunity for breach of privacy.

Implementation of Fetal Clinical Exome Sequencing: Comparing Prospective and Retrospective Cohorts

(Abstracted from Genet Med 2022;24:344–363) Approximately 2% to 3% of pregnancies are affected by fetal abnormalities, and many of these are detected by ultrasound. After identification of suspected fetal structural anomaly, invasive procedures such as chorionic villus sampling and amniocentesis are offered as a method of prenatal genetic diagnosis.

Identifying germline genetic alterations or environmental factors associated with bilateral germ cell tumor (GCT).

5035 Background: Germline genomic alterations and/or environmental factors are proposed as critical to GCT development given the significant rates of bilateral and familial cases. A number of low-moderate risk loci for GCT have been identified (Litchfield, Eur Urol 2018 & Pluta, Nat Comm 2021), predominantly by analysing unilateral GCT, with a polygenic heritability model proffered. We postulate bilateral GCT represents an extreme phenotype with higher frequency of loss of function germline genomic alterations and/or pre-disposing environmental exposures. Methods: Bilateral GCT pts who consented to a prospective clinically annotated GCT DNA registry were included for analysis. The registry integrates self-reported questionnaires on environmental exposures, relevant medical history and germline whole exome sequencing (WES). Pathogenicity of variants identified by WES were classified using the algorithm Pathogenicity of Mutation Analyzer (PathoMAN) [Ravichandran, Genet Med 2019]. Aggregated WES results were curated to identify disruptive or deleterious variants affecting the same genes as the most significant individual variants in the Litchfield series. A < 1% minor allele frequency threshold was set for candidate variants, likely benign/benign variants were excluded . Results: Of 43 bilateral cases with germline WES results, median age at first diagnosis was 32.1 years (range 18-73), 9 tumors were synchronous and 34 metachronous with median interval between diagnosis of 76.4 months for metachronous cases. Tumor histology was seminoma in 52 and nonseminoma in 34. 39 of 43 (91%) pts completed questionnaires with 35 (90%) reporting White race, 2 Hispanic/Latino, 1 Asian, and 1 Peruvian. 13 (33%) reported prior smoking, 21 (54%) prior marijuana use, and 6 (15%) other illicit drug use. 6 (15%) pts reported undescended testis and 1 (3%) hypospadias. 24/43 (45%) patients harbored ≥1 variant within 15 Litchfield genes (60%). 43 distinct variants were identified: 33 (77%) nonsynonymous missense alterations, 7 (16%) intron variants, 1 inframe deletion in STH3TC1, and 1 initiator and 1 stop codon variant in DEFB132, which plays a role in binding spermatozoa. Multiple bilateral patients harbored variants in MPDZ (n = 7), EHBP1L1 (n = 6), SKIV2L (n = 5), FAM160A2 (n = 3), ADAMTS18 (n = 3), JMJD4 (n = 3), ABCC4 (n = 3), R3HCC1 (n = 2), MLXIP (n = 2) and VPS16 (n = 2). Conclusions: In this bilateral GCT cohort, we identified several alterations in candidate susceptibility genes from the Litchfield series, whereas no convincing causative recurrent environmental exposures were observed. Our results support a polygenic model of GCT pathogenesis and show that analyzing enriched cohorts such as bilateral GCT may aid understanding of GCT pathogenesis. Additional analysis of the specific variants is required to further assess pathogenicity, elucidate their role and association with bilateral GCT.

Direct-to-Consumer Prenatal Testing for Multigenic or Polygenic Disorders: A Position Statement of the American College of Medical Genetics and Genomics (ACMG)

(Abstracted from Genet Med 2021:23:2027–2028) The American College of Medical Genetics and Genomics (ACMG) recently updated its position statement on direct-to-consumer (DTC) prenatal testing for multigenic or polygenic disorders. The ACMG recommends that prenatal testing for these disorders is not ready for use in the clinic and should not be offered as DTC testing.

Genome-Wide Cell-Free DNA Screening: A Focus on Copy-Number Variants

(Abstracted from Genet Med 2021;23:1847–1853) In 2015, a study found that traditional cell-free DNA (cfDNA) screening had the capability to detect approximately 70% to 80% of the karyotypic abnormalities identified in fetuses and infants. The remaining 20% to 30% that are missed include clinically relevant findings such as subchromosomal deletions and duplications, rare autosomal aneuploidies, and polyploidy, representing an area of growth for prenatal screening tests, and genome-wide cfDNA screening is now available.

Screening for Autosomal Recessive and X-Linked Conditions During Pregnancy and Preconception: A Practice Resource of the American College of Medical Genetics and Genomics (ACMG)

(Abstracted from Genet Med 2021;23:1793–1806) Carrier screening can be used to determine if an individual or couple is at risk to have a child with an autosomal recessive or X-linked genetic disorder. The American College of Medical Genetics and Genomics (ACMG) last published documents specific to carrier screening in 2004 and 2008.

50 Years Ago in TheJournalofPediatrics: From Chromosomes to Clinical Care: Klinefelter Syndrome

Caldwell PD, Smith DW. The XXY (Klinefelter's) syndrome in childhood: detection and treatment. J Pediatr 1972; 80:250-8. Classical dysmorphology was born, like so much of clinical medicine, from painstaking and careful clinical observation of unrelated individuals with shared clinical features. In this report, Caldwell and Smith take the subsequent steps toward broader understanding of the phenotype and treatment. They tackle the challenge of identifying individuals with a common syndrome before obvious presentation of features. They further dive into the now-modern era of treatment for genetic syndromes. Klinefelter syndrome was published in 1942 from observations during Klinefelter's fellowship at Mass General Hospital.1Klinefelter H.F. Reifenstein E.C. Albright F. Syndrome characterized by gynecomastia, aspermatogenesis without A-leydigism, and increased excretion of follicle-stimulating hormone.J Clin Endocrinol Metab. 1942; 2: 615-627Google Scholar Seventeen years later, the cause of Klinefelter syndrome was identified as an additional “X,” for a chromosome compliment of 47,XXY.2Jacobs P.A. Strong D.J.A. A case of human intersexuality having a possible XXY sex-determining mechanism.Nature. 1959; 183: 302-303Google Scholar This made Klinefelter syndrome the first sex chromosome aneuploidy identified—in predoctoral work by Patricia A. Jacobs. Prof Jacobs continues as a genomicist today, directing research at the University of Southampton in the United Kingdom. Despite the definition of Klinefelter syndrome as 47,XXY, the presence of a Barr body (“X chromatin positive”) in a phenotypic male subject often was used as an efficient “biomarker.” We now appreciate that this, in fact, included individuals with alternative diagnoses, including 46,XX sex reversal (eg, with SRY translocation). The challenge of parsing out a screen that merely suggests sex chromosome aneuploidy, and taking appropriate subsequent steps for a complete genetic diagnosis, continues today. In particular, this is an issue with noninvasive prenatal screening, where the positive predictive value for 47,XXY is a mean of 50%.3Cherry A.M. Akkari Y.M. Barr K.M. Kearney H.M. Rose N.C. South S.T. et al.Diagnostic cytogenetic testing following positive noninvasive prenatal screening results: a clinical laboratory practice resource of the American College of Medical Genetics and Genomics (ACMG).Genet Med. 2017; 19: 845-850Google Scholar We also understand now, in part thanks to efforts like that of Caldwell and Smith, that the phenotype of 47,XXY Klinefelter syndrome is broad. Even today, many individuals present initially for fertility evaluation, having experienced minimal learning or pubertal challenges, and yet more are identified now that noninvasive prenatal screening in pregnancy is commonplace regardless of maternal age. Treatment for Klinefelter is also broadening. Hormonal supplementation for those needing pubertal support is commonplace, and investigation is now focused on whether early pediatric hormonal support may benefit learning and neurologic outcomes.4Samango-Sprouse C.A. Tran S.L. Lasutschinkow P.C. Sadeghin T. Powell S. Mitchell F.L. et al.Neurodevelopmental outcome of prenatally diagnosed boys with 47,XXY (Klinefelter syndrome) and the potential influence of early hormonal therapy.Am J Med Genet A. 2020; 182: 1881-1889Google Scholar Where diagnosis using genetics and dysmorphology was once an end point unto itself, Caldwell and Smith's work is an early indication of the future of genomic medicine. Ultimately, prognostication alone is often insufficient. The goal of all pediatrics is improvement of health and outcomes, in this case using the identification of a syndrome as a tool to reach that goal.

Colorectal Cancer Risk in Lynch Syndrome: Of Genes and More

Colorectal Cancer Risk in Lynch Syndrome: Of Genes and More

Beyond Diagnostic Yield: Prenatal Exome Sequencing Results in Maternal, Neonatal, and Familial Clinical Management Changes

(Abstracted from Genet Med 2021;23:909–917) At present, it is recommended that chromosomal microarray (CMA) be offered in cases of fetal congenital anomalies. However, 60% to 70% of these cases do not receive a genetic diagnosis following microarray, and exome sequencing is being used more frequently in these cases with a reported diagnostic yield of approximately 10% to 30%.

Exome Sequencing Analysis on Products of Conception: A Cohort Study to Evaluate Clinical Utility and Genetic Etiology for Pregnancy Loss

(Abstracted from Genet Med 2021;2:435–442) Genetic variants are known to cause pregnancy loss. New data from exome sequencing studies have reported that exome sequencing may be a useful tool in identifying causes of pregnancy loss.

Screening Embryos for Polygenic Conditions and Traits: Ethical Considerations for an Emerging Technology

(Abstracted from Genet Med 2021;23:432–434) Polygenic embryo screening (PES) uses polygenic risk scores (PRSs) to estimate risk for complex disorders and traits as a part of preimplantation genetic testing (PGT). Preimplantation genetic testing has been used for many years in order to avoid transferring embryos that have aneuploidies or single gene Mendelian disorders.

Can we talk about myoblast fusion?

Can we talk about myoblast fusion?

Clinical Outcomes of a Genomic Screening Program for Actionable Genetic Conditions

(Abstracted from Genet Med 2020;22:1874–1882) Hereditary breast and ovarian cancer (HBOC), Lynch syndrome, and familial hypercholesterolemia (FH) are recognized by the Centers of Disease Control and Prevention as tier-1 genetic conditions for which early identification and management could decrease morbidity and mortality. Yet few patients with a personal or familial history of these conditions are screened for genetic risk.

Implementation of Population-Based Newborn Screening Reveals Low Incidence of Spinal Muscular Atrophy

(Abstracted from Genet Med 2020;22:1296–1302) Spinal muscular atrophy (SMA) is a neurodevelopmental disorder that is associated with muscle weakness and atrophy. There are several subtypes that are categorized as SMA type 1 (presenting in early infancy), where infants are unable to sit unassisted; SMA type 2 (presenting between 6 and 18 months of age), where patients are unable to walk independently; SMA type 3 (presenting after 18 months of age), where some patients were able to walk independently, but others lose this ability; and SMA type 4 (presenting in adulthood), which is rare and less severe.