The penetrance of rare variants in cardiomyopathy-associated genes: A cross-sectional approach to estimating penetrance for secondary findings.

Understanding the penetrance of pathogenic variants identified as secondary findings (SFs) is of paramount importance with the growing availability of genetic testing. We estimated penetrance through large-scale analyses of individuals referred for diagnostic sequencing for hypertrophic cardiomyopathy (HCM; 10,400 affected individuals, 1,332 variants) and dilated cardiomyopathy (DCM; 2,564 affected individuals, 663 variants), using a cross-sectional approach comparing allele frequencies against reference populations (293,226 participants from UK Biobank and gnomAD). We generated updated prevalence estimates for HCM (1:543) and DCM (1:220). In aggregate, the penetrance by late adulthood of rare, pathogenic variants (23% for HCM, 35% for DCM) and likely pathogenic variants (7% for HCM, 10% for DCM) was substantial for dominant cardiomyopathy (CM). Penetrance was significantly higher for variant subgroups annotated as loss of function or ultra-rare and for males compared to females for variants in HCM-associated genes. We estimated variant-specific penetrance for 316 recurrent variants most likely to be identified as SFs (found in 51% of HCM- and 17% of DCM-affected individuals). 49 variants were observed at least ten times (14% of affected individuals) in HCM-associated genes. Median penetrance was 14.6% (±14.4% SD). We explore estimates of penetrance by age, sex, and ancestry and simulate the impact of including future cohorts. This dataset reports penetrance of individual variants at scale and will inform the management of individuals undergoing genetic screening for SFs. While most variants had low penetrance and the costs and harms of screening are unclear, some individuals with highly penetrant variants may benefit from SFs.

Facing the Unknown: An Inductive Analysis of the Lived Experience of Medical Residents during the COVID-19 Pandemic.

INTRODUCTION: The COVID-19 pandemic had significant repercussions for the everyday life and public health of society. Healthcare professionals were particularly vulnerable. Here, we interviewed medical residents about their lived experiences during the pandemic to offer a phenomenological analysis. To this end, we discuss their pandemic experiences considering Jaspers' "limit situation" concept - that is, a radical shift from their everyday experiences, to one causing them to question the basis of their very existence. METHODS: We interviewed 33 medical residents from psychiatry and other specialties from the Hospital das Clínicas da Faculdade de Medicina da Universidade de São Paulo (HCFMUSP) who either (a) worked directly with COVID-19 patients or (b) provided psychiatric care to other healthcare professionals. Semi-structured interviews were developed using the Inductive Process to Analyze the Structure of lived Experience (IPSE). RESULTS: The descriptions of the lived experiences of medical residents during the pandemic were organized into four content themes: (a) existential defense, (b) limit situations during the COVID-19 pandemic, (c) changes in lived experience, and (d) new world meanings through lived experience. CONCLUSION: During the COVID-19 pandemic, medical residents experienced what can be thought of as a "limit situation," as they encountered the healthcare delivery challenges coupled with the social isolation imposed by the COVID-19 pandemic. These challenges included fear of infection and potential death, uncertainty about the future, and the emotional overload caused by the sharp increase in patient deaths. That said, after facing such a limit situation, residents reported feeling strengthened by this experience. This is consistent with the notion that when confronted with limit situations, we draw on our resources to overcome adversity and, in turn, reap existential gains. Health care providers might use these experiences to energize their own professional approach.

An assessment of the role of vinculin loss of function variants in inherited cardiomyopathy.

The ACMG/AMP variant classification framework was intended for highly penetrant Mendelian conditions. While it is appreciated that clinically relevant variants exhibit a wide spectrum of penetrance, accurately assessing and expressing the pathogenicity of variants with lower penetrance can be challenging. The vinculin (VCL) gene illustrates these challenges. Model organism data provide evidence that loss of function of VCL may play a role in cardiomyopathy and aggregate case-control studies suggest low penetrance. VCL loss of function variants, however, are rarely identified in affected probands and therefore there is a paucity of family studies clarifying the clinical significance of individual variants. This study, which aggregated data from >18,000 individuals who underwent gene panel or exome testing for inherited cardiomyopathies, identified 32 probands with VCL loss-of-function variants and confirmed enrichment in probands with dilated cardiomyopathy (odds ratio [OR] = 9.01; confidence interval [CI] = 4.93-16.45). Our data revealed that the majority of these individuals (89.5%) had pediatric onset of disease. Family studies demonstrated that heterozygous loss of function of VCL alone is insufficient to cause cardiomyopathy but that these variants do contribute to disease risk. In conclusion, VCL loss-of-function variants should be reported in a diagnostic setting but need to be clearly distinguished as having lower penetrance.

Early somatic mosaicism is a rare cause of long-QT syndrome.

Somatic mosaicism, the occurrence and propagation of genetic variation in cell lineages after fertilization, is increasingly recognized to play a causal role in a variety of human diseases. We investigated the case of life-threatening arrhythmia in a 10-day-old infant with long QT syndrome (LQTS). Rapid genome sequencing suggested a variant in the sodium channel NaV1.5 encoded by SCN5A, NM_000335:c.5284G > T predicting p.(V1762L), but read depth was insufficient to be diagnostic. Exome sequencing of the trio confirmed read ratios inconsistent with Mendelian inheritance only in the proband. Genotyping of single circulating leukocytes demonstrated the mutation in the genomes of 8% of patient cells, and RNA sequencing of cardiac tissue from the infant confirmed the expression of the mutant allele at mosaic ratios. Heterologous expression of the mutant channel revealed significantly delayed sodium current with a dominant negative effect. To investigate the mechanism by which mosaicism might cause arrhythmia, we built a finite element simulation model incorporating Purkinje fiber activation. This model confirmed the pathogenic consequences of cardiac cellular mosaicism and, under the presenting conditions of this case, recapitulated 2:1 AV block and arrhythmia. To investigate the extent to which mosaicism might explain undiagnosed arrhythmia, we studied 7,500 affected probands undergoing commercial gene-panel testing. Four individuals with pathogenic variants arising from early somatic mutation events were found. Here we establish cardiac mosaicism as a causal mechanism for LQTS and present methods by which the general phenomenon, likely to be relevant for all genetic diseases, can be detected through single-cell analysis and next-generation sequencing.

Adaptation and validation of the ACMG/AMP variant classification framework for MYH7-associated inherited cardiomyopathies: recommendations by ClinGen's Inherited Cardiomyopathy Expert Panel.

PurposeIntegrating genomic sequencing in clinical care requires standardization of variant interpretation practices. The Clinical Genome Resource has established expert panels to adapt the American College of Medical Genetics and Genomics/Association for Molecular Pathology classification framework for specific genes and diseases. The Cardiomyopathy Expert Panel selected MYH7, a key contributor to inherited cardiomyopathies, as a pilot gene to develop a broadly applicable approach.MethodsExpert revisions were tested with 60 variants using a structured double review by pairs of clinical and diagnostic laboratory experts. Final consensus rules were established via iterative discussions.ResultsAdjustments represented disease-/gene-informed specifications (12) or strength adjustments of existing rules (5). Nine rules were deemed not applicable. Key specifications included quantitative frameworks for minor allele frequency thresholds, the use of segregation data, and a semiquantitative approach to counting multiple independent variant occurrences where fully controlled case-control studies are lacking. Initial inter-expert classification concordance was 93%. Internal data from participating diagnostic laboratories changed the classification of 20% of the variants (n = 12), highlighting the critical importance of data sharing.ConclusionThese adapted rules provide increased specificity for use in MYH7-associated disorders in combination with expert review and clinical judgment and serve as a stepping stone for genes and disorders with similar genetic and clinical characteristics.

Association of Arrhythmia-Related Genetic Variants With Phenotypes Documented in Electronic Medical Records.

IMPORTANCE: Large-scale DNA sequencing identifies incidental rare variants in established Mendelian disease genes, but the frequency of related clinical phenotypes in unselected patient populations is not well established. Phenotype data from electronic medical records (EMRs) may provide a resource to assess the clinical relevance of rare variants. OBJECTIVE: To determine the clinical phenotypes from EMRs for individuals with variants designated as pathogenic by expert review in arrhythmia susceptibility genes. DESIGN, SETTING, AND PARTICIPANTS: This prospective cohort study included 2022 individuals recruited for nonantiarrhythmic drug exposure phenotypes from October 5, 2012, to September 30, 2013, for the Electronic Medical Records and Genomics Network Pharmacogenomics project from 7 US academic medical centers. Variants in SCN5A and KCNH2, disease genes for long QT and Brugada syndromes, were assessed for potential pathogenicity by 3 laboratories with ion channel expertise and by comparison with the ClinVar database. Relevant phenotypes were determined from EMRs, with data available from 2002 (or earlier for some sites) through September 10, 2014. EXPOSURES: One or more variants designated as pathogenic in SCN5A or KCNH2. MAIN OUTCOMES AND MEASURES: Arrhythmia or electrocardiographic (ECG) phenotypes defined by International Classification of Diseases, Ninth Revision (ICD-9) codes, ECG data, and manual EMR review. RESULTS: Among 2022 study participants (median age, 61 years [interquartile range, 56-65 years]; 1118 [55%] female; 1491 [74%] white), a total of 122 rare (minor allele frequency <0.5%) nonsynonymous and splice-site variants in 2 arrhythmia susceptibility genes were identified in 223 individuals (11% of the study cohort). Forty-two variants in 63 participants were designated potentially pathogenic by at least 1 laboratory or ClinVar, with low concordance across laboratories (Cohen κ = 0.26). An ICD-9 code for arrhythmia was found in 11 of 63 (17%) variant carriers vs 264 of 1959 (13%) of those without variants (difference, +4%; 95% CI, -5% to +13%; P = .35). In the 1270 (63%) with ECGs, corrected QT intervals were not different in variant carriers vs those without (median, 429 vs 439 milliseconds; difference, -10 milliseconds; 95% CI, -16 to +3 milliseconds; P = .17). After manual review, 22 of 63 participants (35%) with designated variants had any ECG or arrhythmia phenotype, and only 2 had corrected QT interval longer than 500 milliseconds. CONCLUSIONS AND RELEVANCE: Among laboratories experienced in genetic testing for cardiac arrhythmia disorders, there was low concordance in designating SCN5A and KCNH2 variants as pathogenic. In an unselected population, the putatively pathogenic genetic variants were not associated with an abnormal phenotype. These findings raise questions about the implications of notifying patients of incidental genetic findings.

Postmortem Genetic Testing Is an Increasingly Utilized Tool in Death Investigation.

Introduction: Postmortem genetic testing (PMGT) can provide valuable information about an individual's cause of death and potentially allow at-risk relatives to discern their risks for inherited cardiac disease. Postmortem genetic testing is most often successful with certain specimens. Methods: Investigators collected data on postmortem referrals to GeneDx, LLC for PMGT. Orders were reviewed and stratified based on provider, specimen type, and tests ordered. Discussion: This cohort included 601 deceased individuals referred for PMGT with a total of 673 genetic tests ordered from 247 different providers. The most common test categories ordered were arrhythmia (33.4%) and cardiomyopathy (29.3%). A likely pathogenic or pathogenic genetic variant was identified in approximately 15% of patients. Blood in EDTA was received for 21.6% of patients with a 95% success rate for completion of all test components. Blood samples in EDTA were most successful in completing PMGT, but sequencing was still successful in the majority of suboptimal specimens. Conclusion: The use of PMGT is increasing. Obtaining optimal samples (blood in EDTA) is important for successful completion of genetic testing. Obstacles may still exist for obtaining and storing ideal specimens. Continued efforts are needed for education and awareness around appropriate specimen types, storage and shipping of specimens, DNA banking, and overall availability of PMGT. In addition, access to resources such as supplies, proper storage conditions, DNA banking, and PMGT will allow for more opportunities to complete testing.

Harmonizing the Collection of Clinical Data on Genetic Testing Requisition Forms to Enhance Variant Interpretation in Hypertrophic Cardiomyopathy (HCM): A Study from the ClinGen Cardiomyopathy Variant Curation Expert Panel.

Diagnostic laboratories gather phenotypic data through requisition forms, but there is no consensus as to which data are essential for variant interpretation. The ClinGen Cardiomyopathy Variant Curation Expert Panel defined a phenotypic data set for hypertrophic cardiomyopathy (HCM) variant interpretation, with the goal of standardizing requisition forms. Phenotypic data elements listed on requisition forms from nine leading cardiomyopathy testing laboratories were compiled to assess divergence in data collection. A pilot of 50 HCM cases was implemented to determine the feasibility of harmonizing data collection. Laboratory directors were surveyed to gauge potential for adoption of a minimal data set. Wide divergence was observed in the phenotypic data fields in requisition forms. The 50-case pilot showed that although demographics and assertion of a clinical diagnosis of HCM had 86% to 98% completion, specific phenotypic features, such as degree of left ventricular hypertrophy, ejection fraction, and suspected syndromic disease, were completed only 24% to 44% of the time. Nine data elements were deemed essential for variant classification by the expert panel. Participating laboratories unanimously expressed a willingness to adopt these data elements in their requisition forms. This study demonstrates the value of comparing and sharing best practices through an expert group, such as the ClinGen Program, to enhance variant interpretation, providing a foundation for leveraging cumulative case-level data in public databases and ultimately improving patient care.

Newborn screening for X-linked adrenoleukodystrophy (X-ALD): validation of a combined liquid chromatography-tandem mass spectrometric (LC-MS/MS) method.

Newborn screening for X-linked adrenoleukodystrophy (X-ALD) has until now been limited in implementation because of the lack of an accepted standard methodology. We have previously reported a technique using LC-MS/MS analysis that could provide the basis for screening of newborns for X-ALD. The target analyte diagnostic for X-ALD and other peroxisomal disorders of peroxisomal beta-oxidation is 1-hexacosanoyl-2-lyso-sn-3-glycero-phosphorylcholine (26:0-lyso-PC). We report here the validation of the analytical method using an authentic standard of the target compound. The method possesses sensitivity of <1.0fmole injected on column with a correlation coefficient (R(2)) of 0.9987. A tetradeuterated analog of 26:0-lyso-PC served as the internal standard. The sensitivity of this clinical method was confirmed using 17 newborn samples of individuals with peroxisomal disorders retrieved from state newborn screening programs. These samples were run masked with over 1000 newborn samples. All affected individuals were identified with one exception. One sample which was retrieved as an affected did not have the biochemical or genetic abnormality of X-ALD and thus is considered an error in sample identity. These studies clearly show that the method is highly sensitive and accurate in identifying individuals with a defect in peroxisomal beta-oxidation such as X-ALD.

The incidence of duplicate genetic testing.

PURPOSE: Duplicate genetic testing (DGT) should give the same results as the initial genetic test. Therefore, DGT is indicated only in the rare instances where the initial results require confirmation. The objective of this study was to determine the incidence of DGT by reviewing TPMT, HFE, and CYP450 2D6 polymorphism testing performed in our institution's laboratories in 2006. A secondary objective was to determine the savings in charges that resulted from a system in place to limit HFE DGT. METHODS: A retrospective records review at an academic medical center. RESULTS: The percentage of patients having the same genetic test more than once in 2006 was 3.3% (253/7710) for TPMT, 0.3% for HFE (24/7851), and 0.9% (4/433) for CYP450 2D6 testing. Retail laboratory charges for the DGT identified in 2006 were $76,728. To estimate the incidence of DGT over a longer period of time than 2006, an all-time records review was performed on a subset of internal patients and found the all-time incidence of DGT for TPMT, HFE, and CYP450 2D6 testing to be 6.9%, 1.9%, and 0.9%, respectively. No case of DGT with an appropriate indication for duplicate testing was found. A system in place to decrease HFE DGT is estimated to have saved $77,479 in charges for 2006 (95% CI, $35,512-184,015). CONCLUSIONS: Indicated DGT is rare and decreasing DGT could result in significant savings. Institutions should consider implementing a systems-based process to limit DGT.

High-Throughput Functional Evaluation of KCNQ1 Decrypts Variants of Unknown Significance.

BACKGROUND: The explosive growth in known human gene variation presents enormous challenges to current approaches for variant classification that have implications for diagnosis and treatment of many genetic diseases. For disorders caused by mutations in cardiac ion channels as in congenital arrhythmia syndromes, in vitro electrophysiological evidence has high value in discriminating pathogenic from benign variants, but these data are often lacking because assays are cost, time, and labor intensive. METHODS: We implemented a strategy for performing high-throughput functional evaluations of ion channel variants that repurposed an automated electrophysiological recording platform developed previously for drug discovery. RESULTS: We demonstrated the success of this approach by evaluating 78 variants in KCNQ1, a major gene involved in genetic disorders of cardiac arrhythmia susceptibility. We benchmarked our results with traditional electrophysiological approaches and observed a high level of concordance. This strategy also enabled studies of dominant-negative behavior of variants exhibiting severe loss-of-function. Overall, our results provided functional data useful for reclassifying >65% of the studied KCNQ1 variants. CONCLUSIONS: Our results illustrate an efficient and high-throughput paradigm linking genotype to function for a human cardiac ion channel that will enable data-driven classification of large numbers of variants and create new opportunities for precision medicine.

Medical errors related to inappropriate genetic testing in liver transplant patients.

Genetic testing of genes that encode proteins expressed by liver hepatocytes (clotting factors, alpha 1-antitrypsin, cytochrome P450 enzymes) is common in clinical practice. These tests use DNA extracted from peripheral blood lymphocytes (PBL) and are based on the assumption that PBL DNA can be used as a surrogate for hepatocyte DNA. However, in individuals who have undergone liver transplantation, hepatocyte DNA is that of the donor while PBL DNA remains that of the recipient. It follows that in liver transplant patients, genetic testing of the recipient's PBL DNA does not provide accurate results for proteins expressed by donor hepatocytes. Therefore, genetic testing of clotting factors, alpha 1-1-antitrypsin, cytochrome P450 enzymes, and other proteins expressed by hepatocytes is unreliable and inappropriate in liver transplant patients (inappropriate genetic testing). A review of the records of 215 consecutive liver transplant patients at our institution identified: one medical error and one near-miss medical error related to inappropriate genetic testing, 14 cases of inappropriate genetic testing, and 21 unnecessary duplicate genetic testing requests. We recommend laboratories performing genetic testing create systems to prevent inappropriate and duplicate genetic testing and that physicians be cognizant of the appropriate indications for genetic testing in liver transplant patients.

Results of genetic testing in 855 consecutive unrelated patients referred for long QT syndrome in a clinical laboratory.

AIM: Our aim was to examine the diagnostic yield of genetic testing in 855 consecutive unrelated cases referred for Long QT syndrome (LQTS). RESULTS: Eight hundred fifty five consecutive patients with a mean age at testing of 27.5±18.6 years, were referred for LQTS genetic testing and had accompanying clinical information. KCNQ1, KCNH2, SCN5A, ANK2, KCNE1, KCNE2, CACNA1C, KCNJ2, CAV3, and SCN4B were analyzed using Next-Generation sequencing in all patients, and 395 patients were also tested for an additional two genes, AKAP9 and SNTA1. We retrospectively analyzed the diagnostic yield of this genetic test and factors that predicted the likelihood of a disease causing mutation using ANOVA, χ2, t-test, and receiver operator curves. At least one mutation was identified in 30.3% of the patients (n=259), and 18 patients (2.1%) had two mutations. Patients with two mutations had a longer QTc interval (p<0.01) than patients with one mutation. A longer QTc duration and family history of LQTS were each associated with a higher yield of positive results on genetic testing (p<0.01 for each). Using a QTc cutoff of 476 msec or greater, the genetic testing had a sensitivity of 72% and a specificity of 49%. Mutations within the transmembrane domain of KCNQ1 were associated with a greater risk of cardiac arrest and syncope relative to mutations in other domains of the gene. Mutations in SCN5A were associated with a higher frequency of cardiac arrest (52.6%). CONCLUSION: Sequencing-based genetic testing has a sensitivity of 72% and has clinical utility.

Sexo sem tabu: relato de um caso de ação comunitária, com foco na promoção de saúde de adolescentes, em uma escola técnica na cidade de São Paulo / Sex without taboo: report of a community action focused on health promotion for teenagers in a technical high school in the city of São Paulo

A sexualidade, apesar de sua relevância no âmbito da promoção da saúde, é pouco abordada durante os cursos de graduação e por profissionais da saúde na prática clínica. O objetivo deste trabalho é descrever o projeto “Sexo sem Tabu”, realizado por alunos do primeiro ano do curso de Medicina da Universidade de São Paulo, dentro da proposta da disciplina “Atenção Primária à Saúde I”. Após delineamento do tema e revisão literária, foram executados dois dias de intervenção. Baseados em dinâmicas propostas pelo Ministério da Saúde, discutimos sexualidade e temas correlatos em saúde, com pequenos grupos de alunos de uma escola técnica na cidade de São Paulo. Dentre os alunos participantes, 115/117 (98,3%) disseram ter dúvidas solucionadas durante a discussão e 114/117 (97,4%) consideraram que as informações que foram trazidas eram relevantes para suas vidas. Os alunos da Faculdade de Medicina relataram a oportunidade de treinar várias habilidades afetivas e cognitivas como habilidades de comunicação, desenvoltura, capacidade de lidar com o imprevisto, além de reconhecer que a abordagem da sexualidade não se restringe aos aspectos relacionados às doenças sexualmente transmissíveis, mas também a questões emocionais, morais e fisiológicas. Em conclusão, esta ação comunitária, com foco em promoção da saúde dos adolescentes e abordando o tema da sexualidade, trouxe resultados positivos para os alunos do ensino médio e para a formação profissional dos estudantes de medicina.

Although sexuality has an evident importance in health promotion, it is rarely discussed during undergraduate courses or assessed by health professionals during clinical practice. The aim of this article is to describe the “Sexo sem Tabu” project, a community action realized by first year medical students from the Faculdade de Medicina da Universidade de São Paulo, during the course “Primary Care I” (“Atenção Primária à Saúde I”). After establishing project design and adequate literature review, we realized two days of community action. Based on dynamics proposed by the Brazilian Ministry of Health, we discussed sexuality and other related health topics within small groups of students from a technical high school in the city of São Paulo. Among high school participants, 115/117 (98.3%) considered that they had their doubts solved during the discussion and 114/117(97.4%) found the information was relevant for their lives. First year medical students documented the opportunity to train several cognitive and affective abilities such as communication, resourcefulness, and how to interact in uncontrolled situations. Furthermore, they recognized that sexuality was not restricted to sexually transmitted diseases, but included emotional, ethical and physiological issues. Concluding, this community action, focused on health promotion for teenagers assessing themes on sexuality, brought positive results for high school students and for the training of medical students.