Variant Carriers Research Articles

Abstract 4135624: A Hypertrophic Cardiomyopathy Polygenic Score Modifies Penetrance of Pathogenic Hypertrophic and Dilated Cardiomyopathy Variants in Opposite Directions

Background: Hypertrophic (HCM) and dilated (DCM) cardiomyopathy have been postulated to represent morphologically opposing cardiac disease traits, with genetic pathways implicated in HCM demonstrating inverse relationships with DCM. The degree to which polygenic risk for HCM protects from DCM in individuals at genetic risk of DCM remains unexplored. Research Question: Is polygenic risk for HCM protective against developing DCM in carriers of titin truncating variants? Methods: Associations between polygenic risk for HCM, monogenic variants, and cardiomyopathy outcomes were modeled among Penn Medicine BioBank (PMBB) participants genetically similar to European reference populations using logistic regression. Polygenic risk for HCM was determined using a previously published HCM polygenic score (PGS; PGS Catalog PGS000739; Harper et al. 2021); scores were PCA adjusted, and the mean and variance were normalized using pgsc_calc. Pathogenic variants in definitive HCM genes were identified using ACMG criteria. Titin truncating variants in high percentage spliced in exons (TTNtv) were considered pathogenic. HCM and DCM outcomes were assigned based on diagnosis codes +/- clinical notes in the electronic health record. Results: Of 26,728 PMBB participants, 205 (0.77%) had HCM and 589 (2.2%) had DCM. 289 had a TTNtv, and 144 had a pathogenic HCM variant. Pathogenic HCM variant status (n=144) conferred a 76-fold increase in HCM risk (p=1.8 e-87). A one standard deviation change in HCM PGS had a significant positive association with HCM (OR 1.6; p=1.5e-9) and negative association with DCM (OR 0.72; p=1.e-12). TTNtv status (n=289) was associated with an increased risk of DCM (OR 5.0; p=1.4e-12). An interaction term between TTNtv status and HCM PGS had a negative association with DCM (OR 0.61; p=4.5e-2). Among the top HCM PGS quintile, there was no statistically significant difference in DCM risk between those with and without TTNtv (Figure 1). Conclusions: These results provide additional evidence that HCM and DCM are influenced by polygenic background. We observe that polygenic risk of HCM additively increases risk of HCM among pathogenic HCM variant carriers and may synergistically protect from DCM among TTNtv carriers.

Abstract 4144528: The Polygenic Mechanisms of Dilated Cardiomyopathy Contribute to The Development of Tachycardia-Associated Cardiomyopathy in Patients With Atrial Tachyarrhythmias

Background Longstanding tachycardia can lead to reversible left ventricular (LV) systolic dysfunction called tachycardia-associated cardiomyopathy (TAC). It is unknown if there is a genetic predisposition to the development of TAC or whether there is overlap between the genetic pathways of TAC and dilated cardiomyopathy (DCM). Research Questions Is there a genetic predisposition towards the development of TAC in individuals with arrhythmia? If so, do these genetic pathways overlap with those of DCM? Methods/Approach The study is a single-center case control study performed at the Montreal Heart Institute (MHI). Inclusion criteria for TAC cases are shown in Table 1. All cases underwent array genotyping and exome sequencing. Previously genotyped controls with arrhythmia but no documented LV dysfunction were included from the MHI Biobank. Following imputation using the TOPMed imputation server, a polygenic score for DCM (PGS DCM ; PGS000666) was calculated for cases and controls. The rate of (likely) pathogenic variants in known cardiomyopathy genes was reported in cases. Results/Data (descriptive and inferential statistics) The study included 107 cases and 878 controls. Case characteristics are shown in Table 1. The average PGS DCM for cases was significantly higher than that of controls (p-value <10 -10 ; Figure 1). Logistic regression with correction for age, sex, and ancestry (principal components) showed a strong association of PGS DCM with TAC (OR 1.99, 95% CI 1.54-2.58; p-value <10 -6 ). Notably, among patients with atrial tachyarrhythmias, those with a PGS DCM above the 95 th percentile had a 3.53-fold increased risk of TAC (p-value 4.5 -4 ). Exome sequencing in TAC cases identified only 2/107 (2%) carriers of (likely) pathogenic variants (in cardiomyopathy genes TNNT2 and MYH7 ). Conclusions Pathogenic variants account for a small minority of TAC cases. In contrast, a significant association between PGS DCM and TAC is suggestive of a shared polygenic pathway between TAC and DCM.

Abstract 4120375: Penetrance and Phenotype Conversion Rate of Genetic Hypertrophic Cardiomyopathy in an Academic Biobank

Introduction: Hypertrophic cardiomyopathy (HCM) is an autosomal dominant condition with incomplete penetrance and variable expressivity. Given the increasing prevalence of sequencing and current guidelines on secondary finding disclosure, HCM P/LP alleles are increasingly being returned to patients who undergo genetic testing for other reasons. Whether individuals carrying P/LP HCM variants discovered in a genome-first context have progressive endophenotypes indicative of HCM remains poorly understood. Methods: To evaluate the penetrance and phenotype conversion rate of adults with P/LP HCM variants, we leveraged the Massachusetts General Brigham Biobank (MGBB), which consists of 36,417 participants with genotyping array and clinical data. Primary images of the earliest and latest TTE studies were reviewed for measurements of the posterior wall (PW), interventricular septum (IVS), and left ventricular morphology. Phenotype-positive (P+) status was defined by a PW or IVS thickness of at least 13 mm. Results: P/LP HCM variants were identified in 42 (0.1%) individuals with mean (SD) age 50.0 (16.6) years and 21 (50%) female sex. The most common HCM variants were in MYBPC3 (44%) and MYH7 (40%), followed by TNNI3(7%), MYL3 (7%), and TNNT2 (2%). Among the 42 genotype-positive (G+) individuals in MGBB, 17 (40%) had pre-existing HCM diagnoses prior to genomic return of results (gRoR). In contrast, 25 (60%) G+ individuals lacked a prior HCM diagnosis, among whom gRoR prompted TTE studies in 13 patients, revealing evidence of HCM in 9 individuals. Among the 42 G+ patients, 25 (58%) patients had completed at least two TTE studies, with a median [IQR] of 6 [3-9] TTE studies per patient, across a span of 10 [7-15] years follow-up. At the time of first TTE, 16 (64%) patients were P+ while 9 (36%) were P-. No G+, P- individuals converted to P+ over 10 years follow-up, nor was there any significant change in IVS or PW thickness. In contrast, individuals who were P+ at the initial TTE exhibited thickening PW (25% increase, CI 97.5% of 6 to 44) and IVS (26% increase, CI 97.5% of 4 to 49) over 8 years follow-up. Conclusion: In a genome-first approach, we identified 42 individuals in the MGBB with a P/LP HCM variant, among whom 60% lacked a prior clinical diagnosis of HCM. Targeted phenotyping revealed that nearly half of this newly identified group were P+. Over a median of 10 years follow-up, we observed a 0% phenotype conversion rate among G+, P- adult carriers of HCM variants.

Abstract 4115690: COL4A1-related Vascular Disease Displays Strikingly Different Penetrance, Onset, and Expressivity in Literature versus Biobank-Derived Cases

Monogenic cardiovascular diseases present throughout the human lifespan with varying symptoms. For most, our understanding of their penetrance, onset, and expressivity is derived from published case reports. The extent to which these literature-derived cases capture the full allelic and phenotypic spectrum is unknown, particularly since the literature is biased towards severe presentations. To illustrate, we investigated the penetrance (i.e. the presence of at least one symptom), onset, and phenotypic expressivity of a rare genetic disease linked to hemorrhagic stroke and other complications: COL4A1 -related vascular disease (COL4A1VD, or Gould Syndrome). We performed a systematic review of COL4A1VD cases reported in the biomedical literature ( N =459), normalizing their genetic variants and symptoms to controlled terminologies. We then investigated the phenotypic expression among COL4A1VD pathogenic variant carriers isolated from two population-scale biobanks (the UK Biobank and All of Us Research Program; combined N =714,246), identifying 178 subjects (prevalence: 0.025%) that should be at high risk for COL4A1VD-related symptoms. Based on the literature, the penetrance of COL4A1VD is approximately 92% (95% CI: 90%-95%), with each case expressing 4.3 symptoms on average. Neurologic complications were the most common finding, observed in 85% of symptomatic patients. Alternatively, the biobank-derived cohort displayed an overall enrichment for disease-related symptoms (COL4A1 pathogenic carriers are more likely to have increased symptom burden compared to non-carrier controls; P<0.005), but the penetrance of the variants was substantially reduced (34%; 95% CI: 26%-41%). In addition, this cohort expressed fewer symptoms on average (0.8 per carrier), with renal complications being far more common (78% of symptomatic biobank patients vs 22% of literature-derived cases; P < 0.001). Finally, the apparent age-of-onset was much later for the biobank-derived cohort (57-years vs 5-years for the literature cases). Most of these differences reflect the distinct ascertainment of these two COL4A1VD cohorts, but they also highlight our incomplete understanding of the true penetrance, expressivity, and onset for this rare disease. These results have important implications for COL4A1VD-related genetic counseling and surveillance, and they highlight a need for new methods that formally integrate the rare disease knowledge captured from these distinct data sources.

MYH6 Variants Are Associated with Atrial Dysfunction in Neonates with Hypoplastic Left Heart Syndrome

Background: MYH6 variants are the most well-known genetic risk factor (10%) for hypoplastic left heart syndrome (HLHS) and are associated with decreased cardiac transplant-free survival. MYH6 encodes for α-myosin heavy chain (α-MHC), a contractile protein expressed in the neonatal atria. We therefore assessed atrial function in HLHS patients with MYH6 variants. Methods: We performed a retrospective, blinded assessment of pre-stage I atrial function using 2D speckle-tracking echocardiography (2D-STE). Variant carriers were control-matched based on AV valve anatomy, sex, and birth year. Studies were obtained postnatally from awake patients prior to surgical intervention. Right atrial (RA) and right ventricular (RV) strain and strain rate (SR) were measured from the apical four-chamber view. Results: A total of 19 HLHS patients with MYH6 variants had echocardiograms available; 18 were matched to two controls each, and one had a single control. RA active strain (ASct) was decreased in variant carriers (−1.41%, IQR −2.13, −0.25) vs. controls (−3.53%, IQR −5.53, −1.28; p = 0.008). No significant differences were identified in RV strain between the groups. RA reservoir strain (ASr) and conduit strain (AScd) positively correlated with heart rate (HR) in MYH6 variant carriers only (ASr R = 0.499, p = 0.029; AScd R = 0.469, p = 0.043). RV global longitudinal strain (GLS) as well as RV systolic strain (VSs) and strain rate (VSRs) correlated with HR in controls only (GLS R = 0.325, p = 0.050; VSs R = 0.419, p = 0.010; VSRs R = 0.410, p = 0.012). Conclusions: We identified functional consequences associated with MYH6 variants, a known risk factor for poor outcomes in HLHS. MYH6 variant carriers exhibit impaired RA contractility despite there being no differences in RV function between variant carriers and controls. MYH6 variants are also associated with an ineffective RA reservoir and conduit function at high heart rates, despite preserved RV diastolic function. RA dysfunction and reduced atrial “kick” may therefore be a significant contributor to RV failure and worse clinical outcomes in HLHS patients with MYH6 variants.

Clinical and genetic analysis of a case of Triadin knockout syndrome due to variant of TRDN gene and a literature review

To explore the genetic etiology and clinical phenotype of a child with Triadin knockout syndrome (TKOS), and to review the relevant literature of TKOS patients due to variants of TRDN gene. A child who was admitted to the Children's Hospital of Xi'an Jiaotong University on March 19, 2023 due to sudden cardiac arrest 3 days earlier was selected as the study subject. Peripheral blood samples (2 to 3 mL) were collected from the child and her parents for the extraction of genomic DNA and whole exome sequencing (WES). Pathogenic variants were searched from databases such as the Genome Aggregation Database (gnomAD) and Online Mendelian Inheritance in Man (OMIM), and were assessed based on the guidelines from the American College of Medical Genetics and Genomics (ACMG). Sanger sequencing was carried out for family validation of the pathogenic variants. Using keywords such as "arrhythmias" "TRDN" and "Triadin" both in Chinese and English, relevant literature on TKOS patients due to variants of the TRDN gene was retrieved from the CNKI, Wanfang Data Knowledge Service Platform, and PubMed databases, and the time of literature retrieval was set from January 1, 2012 to December 1, 2023. This study has been approved by the Ethics Committee of the Affiliated Children's Hospital of Xi'an Jiaotong University (No. 20230097), and informed consent was obtained from the parents of the child. The child had experienced syncope and cardiac arrest after exercise. Electrocardiographic examination revealed QTc interval prolongation, T-wave inversion in precordial leads V1-V3, polymorphic ventricular premature beat (VPB), and ventricular tachycardia (VT) along with increased heart rate. WES and Sanger sequencing revealed that the child has harbored a homozygous c.463del(p.E155Kfs*20) variant of the TRDN gene, for which both of the parents were heterozygous. Based on the guidelines from the ACMG, the variant was classified as pathogenic (PVS1+PM2+PM3). The child was ultimately diagnosed with TKOS. In total 12 publications on TOKS cases caused by TRDN gene variants were retrieved, which involved 30 patients and 28 carriers of single heterozygous variant of the TRDN gene. Among the 30 TKOS patients, 20 had carried homozygous variants of the TRDN gene, and 10 had carried compound heterozygous variants, and all had exhibited significant clinical phenotype of arrhythmia, with most cases had experienced malignant arrhythmia induced by exercise and/or excitement during infancy or early childhood, leading to recurrent syncope and cardiac arrest. Of note, none of the 28 carriers of single heterozygous variant had abnormal clinical phenotype. The homozygous c.463del(p.E155Kfs20) variant of the TRDN gene probably underlay the pathogenesis of cardiac arrest in this child. Above discovery has enriched the mutational spectrum of the TRDN gene.

DPYD genotype-guided dose personalisation for fluoropyrimidine-based chemotherapy prescribing in solid organ cancer patients in Australia: GeneScreen 5-FU study protocol

BackgroundFluoropyrimidine (FP) chemotherapies are commonly prescribed for upper and lower gastrointestinal, breast and head and neck malignancies. Over 16,000 people with cancer require FP chemotherapies per annum in Australia. Between 10 and 40% patients experience grade 3–4 (≥ G3) toxicities that require hospital-based management ± intensive care admission. Approximately 1% of patients die secondary to FP toxicities. Prospective screening for DPYD gene variants (encoding the key enzyme for FP catabolism) can identify patients at risk of ≥ G3 toxicity and allow for dose adjustment prior to first FP exposure. Evidence supports this as a cost-effective method of improving patient safety and reducing healthcare burden internationally; however, no Australian data confirms its feasibility on a large scale.MethodThis investigator-led, single-arm study will determine large scale feasibility of prospective DPYD genotyping, confirming patient safety and cost-effectiveness within the Australian health care system. 5000 patients aged 18 years and older with solid organ cancers requiring FP chemotherapy will be consented and genotyped prior to commencing treatment, and early toxicity (within 60 days) post-FP exposure will be determined. Toxicity data for DPYD variant carriers who have dose adjustments will be compared to the wild-type cohort and historical cohorts of carriers who did not undergo genotyping prior to FP exposure, and prospective variant carriers who do not undergo dose-adjustment. Prevalence of the four standard DPYD gene variants will be confirmed in an Australian population. Additionally, health economic analysis, implementation research via semi-structured interviews of patients and clinicians, and feasibility of UGT1A1 genotyping will be conducted.DiscussionThis study will determine the prevalence of DPYD gene variant status in Australia and its impact on FP-induced toxicity among Australians with cancer. Feasibility and cost-effectiveness for Australian health care system will be estimated to support national roll-out of prospective DPYD genotyping prior to FP administration. Additionally, feasibility will be confirmed with the intention of including UGT1A1 in future pharmacogenomic panels to aid chemotherapy prescribing.Trial registrationThis trial was registered with the Australian and New Zealand Cancer Trials Registry on 13th Dec 2023, ACTRN12623001301651.

Effects of GBA1 Variants and Prenatal Exposition on the Glucosylsphingosine (Lyso-Gb1) Levels in Gaucher Disease Carriers

Gaucher disease (GD) is a lysosomal lipid storage disorder caused by β-glucocerebrosidase (encoded by GBA1 gene) activity deficiency, resulting in the accumulation of glucosylceramide (Gb1) and its deacylated metabolite glucosylsphingosine (lyso-Gb1). Lyso-Gb1 has been studied previously and proved to be a sensitive biomarker, distinguishing patients with GD from carriers and healthy subjects. It was shown that its level corresponds with β-glucocerebrosidase activity, thus it remains unknown as to why carriers have slightly higher lyso-Gb1 level than healthy population. This is the first report on lyso-Gb1 levels describing representative cohort of GD carriers. Our data of 48 GD carriers, including three newborns, indicated that there are significant differences in lyso-Gb1 levels between carriers having a GD-affected mother and a healthy mother (11.53 and 8.45, respectively, p = 0.00077), and between carriers of the L483P GBA1 variant and carriers of other GBA1 pathogenic variants (9.85 and 7.03, respectively, p = 0.07). Through analysing our unique data of three newborns whose mothers are patients with GD, we also found that lyso-Gb1 is most probably transferred to the foetus via placenta.

The GDF15 3′ UTR Polymorphism rs1054564 Is Associated with Diabetes and Subclinical Atherosclerosis

Growth differentiation factor 15 (GDF15) is a stress-response cytokine related to a wide variety of metabolic diseases. However, the impact of GDF15-specific genetic variants on the abovementioned conditions is poorly known. The aim of this study was to assess the impact of selected GDF15 single-nucleotide polymorphisms (SNPs) on metabolic disturbances and subclinical atherosclerosis. A cross-sectional study involving 153 participants of a metabolic patient-based cohort was performed. Three selected SNPs (rs888663, rs1054564 and rs1059369) in a locus on chromosome 19 including the GDF15 gene were genotyped by Polymerase Chain Reaction (PCR), and its relationship with the serum GDF15 levels, health status and clinical variables were analyzed. Of the three SNPs analyzed, only rs1054564 showed different distributions between the healthy volunteers and patients suffering lipid alterations and associated disorders. Accordingly, just the rs1054564 variant carriers showed a significant increase in GDF15 serum levels compared to the wild-type carriers. The group of variant carriers showed a higher frequency of individuals with diabetes, compared to the wild-type carrier group, without showing differences in other metabolic conditions. Additionally, the frequency of individuals with atherosclerotic carotid plaque was higher in the rs1054564 variant carriers than in the wild-type carriers. Logistic regression models identified that the presence of the rs1054564 variant carriers increase the likelihood for both diabetes and carotid plaque independently of confounding factors. Overall, the findings of this study identify the rs1054564 variant as a potential indicator for the likelihood of diabetes and subclinical atherosclerosis.

The Excess of Carriers in Rare Disorders Suggests a Nonpathogenic Effect for Most Variants of Uncertain Significance.

Functional annotation and interpretation of genetic variants are a critical step in genetic diagnosis, as it may lead to personalized therapeutic options and genetic counseling. While the number of confirmed pathogenic genetic variants in an individual is relatively low, the number of variants of uncertain significance (VOUS) can be considerably higher, increasing the number of potential carriers of genetic disorders. Thus, reducing uncertainty and assessing the real effect of VOUS are crucial for clinical and medical genetics. In this study, we evaluated the efficacy of genetic screening technologies in accurately predicting pathogenic variants and their corresponding disease prevalence in a cohort of over 6000 healthy individuals involved in assisted reproduction programs. Using data from 305 genes associated with recessive disorders, we determined the frequency of carriers of pathogenic variants and VOUS in our dataset and compared the predicted prevalence based on this information with reported population prevalence data. The higher predicted prevalence in some disorders when considering VOUS suggests a mostly benign effect.

Reclassification of Two MLH1 Variants of Uncertain Significance Utilizing Clinical and Functional Data.

Pathogenic variants in the mismatch repair genes are associated with an elevated lifetime risk of colorectal cancer (CRC). We previously identified two variants of uncertain significance (VUS) in the MLH1 gene, c.696_698del, p.(Cys233del) and c.1919C > G, p.(Pro640Arg), in Danish families with numerous occurrences of CRC. To reclassify the variants we collected clinical data, initiated tumor and co-segregation analysis, and performed RNA splicing analysis, subcellular localization, and protein stability studies. The functional analysis revealed that the c.696_698del, p.(Cys233del) variant had an effect at the RNA level, on subcellular localization, and on protein stability, while the c.1919C > G, p.(Pro640Arg) variant showed decreased expression in localization studies and decreased protein stability. These results suggest both variants disrupt DNA mismatch repair. By applying all collected data and functional results we propose to reclassify the c.696_698del, p.(Cys233del) and the c.1919C > G, p.(Pro640Arg) variants as likely pathogenic (class 4) using MMR gene-specific ACMG/AMP guidelines. Consequently, the two MLH1 variants can now be used for risk assessment of variant carriers, while family members without the variants can be excluded from intensified cancer surveillance and follow population recommendations.

Cardiovascular Risk Factors and Genetic Risk in Transthyretin V142I Carriers.

Nearly 3% to 4% of Black individuals in the United States carry the transthyretin V142I variant, which increases their risk of heart failure. However, the role of cardiovascular (CV) risk factors (RFs) in influencing the risk of clinical outcomes among V142I variant carriers is unknown. This study aimed to assess the impact of CV RFs on the risk of heart failure in V142I carriers. This study included self-identified Black individuals without prevalent heart failure from 6 TOPMed (Trans-Omics for Precision Medicine) cohorts, the REGARDS (Reasons for Geographic And Racial Differences in Stroke) study, and the All of Us Research Program. The cohort was stratified based on the V142I genotype and the number of CV RFs (hypertension, diabetes, obesity, and hypercholesterolemia). Adjusted Cox models were used to assess the association of heart failure with the V142I genotype and CV RF profile, taking noncarriers with a favorable CV RF profile as reference. The cross-sectional analysis, including 1,625 V142I carriers among 48,365 Black individuals, found that the prevalence of CV RFs did not vary by V142I carrier status. In the longitudinal analysis, there were 587 (3.2%) V142I carriers among 18,407 Black individuals (median age: 60 years [Q1-Q3: 52-68 years], 63.0% female). Among carriers, the heart failure risk was attenuated with a favorable (0 or 1 RF) CV RF profile (adjusted HR: 2.26; 95%CI: 1.58-3.23) compared with an unfavorable (3 or 4 RFs) CV RF profile (adjusted HR: 4.14; 95%CI: 2.79-6.14). A favorable CV RF profile lowers but does not abrogate V142I variant-associated heart failure risk. This study highlights the importance of having a favorable CV RF profile among V142I carriers for risk reduction of heart failure.

Skeletal muscle from TBC1D4 p.Arg684Ter variant carriers is severely insulin resistant but exhibits normal metabolic responses during exercise.

In the Greenlandic Inuit population, 4% are homozygous carriers of a genetic nonsense TBC1D4 p.Arg684Ter variant leading to loss of the muscle-specific isoform of TBC1D4 and an approximately tenfold increased risk of type 2 diabetes1. Here we show the metabolic consequences of this variant in four female and four male homozygous carriers and matched controls. An extended glucose tolerance test reveals prolonged hyperglycaemia followed by reactive hypoglycaemia in the carriers. Whole-body glucose disposal is impaired during euglycaemic-hyperinsulinaemic clamp conditions and associates with severe insulin resistance in skeletal muscle only. Notably, a marked reduction in muscle glucose transporter GLUT4 and associated proteins is observed. While metabolic regulation during exercise remains normal, the insulin-sensitizing effect of a single exercise bout is compromised. Thus, loss of the muscle-specific isoform of TBC1D4 causes severe skeletal muscle insulin resistance without baseline hyperinsulinaemia. However, physical activity can ameliorate this condition. These observations offer avenues for personalized interventions and targeted preventive strategies.

Exome sequencing data reanalysis of 200 hypertrophic cardiomyopathy patients: the HYPERGEN French cohort 5 years after the initial analysis

BackgroundApproximately half of hypertrophic cardiomyopathy (HCM) patients lack a precise genetic diagnosis. The likelihood of identifying clinically relevant variants increased over time.MethodsIn this study, we conducted a gene-centric reanalysis of exome data of 200 HCM cases 5 years after the initial analysis. This reanalysis prioritized genes with a matched HCM entry in the OMIM database and recently emerging HCM-associated genes gathered using a text mining-based literature review. Further classification of the identified genes and variants was performed using the Clinical Genome Resource (ClinGen) resource and American College of Medical Genetics and Genomics (ACMG) guidelines to assess the robustness of gene–disease association and the clinical actionability of the prioritized variants.ResultsAs expected, the majority of patients carried variants in MYBPC3 and MYH7 genes, 26% (n = 51) and 8% (n = 16), respectively, in accordance with the initial analysis. The vast majority of pathogenic (P) and likely pathogenic (LP) variants were found in MYBPC3 (22 out of 40 variants) and MYH7 (8 out of 16 variants) genes. Three genes—not included in the initial analysis—were identified: SVIL, FHOD3, and TRIM63. Considering only patients with unique variants in the last three genes, there was a 9% enhancement in variant identification. Importantly, SVIL variant carriers presented apical and septal HCM, aortopathies, and severe scoliosis for one patient. Ten patients (5%) carried variants in the FHOD3 gene, six in hotspot regions (exons 12 and 15). We identified seven variants within the TRIM63 gene in 12 patients (6%). Homozygous variants were detected in 2.5% of the cohort in MYBPC3 (n = 1), MYL3 (n = 1), and TRIM63 (n = 3) genes.ConclusionOur study revealed that no variants were found in the ACTC1, TPM1, and TNNI3 genes in the HYPERGEN cohort. However, we identified variants in five out of the eight HCM core genes, with a high prevalence in young patients. We identified variants in three recent HCM-associated genes (SVIL, FHOD3, and TRIM63) in 35 patients, with 18 patients carrying unique variants (9%). Our results further emphasize the usefulness of exome data reanalysis, particularly in genotype-negative patients.

Second Primary Cancer Risks After Breast Cancer in BRCA1 and BRCA2 Pathogenic Variant Carriers.

Second primary cancer (SPC) risks after breast cancer (BC) in BRCA1/BRCA2 pathogenic variant (PV) carriers are uncertain. We estimated relative and absolute risks using a novel linkage of genetic testing data to population-scale National Disease Registration Service and Hospital Episode Statistics electronic health records. We followed 25,811 females and 480 males diagnosed with BC and tested for germline BRCA1/BRCA2 PVs in NHS Clinical Genetics centers in England between 1995 and 2019 until SPC diagnosis, death, migration, contralateral breast/ovarian surgery plus 1 year, or the 31st of December 2020. We estimated standardized incidence ratios (SIRs) using English population incidences, hazard ratios (HRs) comparing carriers to noncarriers using Cox regression, and Kaplan-Meier 10-year cumulative risks. There were 1,840 BRCA1 and 1,750 BRCA2 female PV carriers. Compared with population incidences, BRCA1 carriers had elevated contralateral BC (CBC; SIR, 15.6 [95% CI, 11.8 to 20.2]), ovarian (SIR, 44.0 [95% CI, 31.4 to 59.9]), combined nonbreast/ovarian (SIR, 2.18 [95% CI, 1.59 to 2.92]), colorectal (SIR, 4.80 [95% CI, 2.62 to 8.05]), and endometrial (SIR, 2.92 [95% CI, 1.07 to 6.35]) SPC risks. BRCA2 carriers had elevated CBC (SIR, 7.70 [95% CI, 5.45 to 10.6]), ovarian (SIR, 16.8 [95% CI, 10.3 to 26.0]), pancreatic (SIR, 5.42 [95% CI, 2.09 to 12.5]), and combined nonbreast/ovarian (SIR, 1.68 [95% CI, 1.24 to 2.23]) SPC risks. Compared with females without BRCA1/BRCA2 PVs on testing, BRCA1 carriers had elevated CBC (HR, 3.60 [95% CI, 2.65 to 4.90]), ovarian (HR, 33.0 [95% CI, 19.1 to 57.1]), combined nonbreast/ovarian (HR, 1.45 [95% CI, 1.05 to 2.01]), and colorectal (HR, 2.93 [95% CI, 1.53 to 5.62]) SPC risks. BRCA2 carriers had elevated CBC (HR, 2.40 [95% CI, 1.70 to 3.40]), ovarian (HR, 12.0 [95% CI, 6.70 to 21.5]), and pancreatic (HR, 3.56 [95% CI, 1.34 to 9.48]) SPC risks. Ten-year cumulative CBC, ovarian, and combined nonbreast/ovarian cancer risks were 16%/6.3%/7.8% (BRCA1 carriers), 12%/3.0%/6.2% (BRCA2 carriers), and 3.6%/0.4%/4.9% (noncarriers). Male BRCA2 carriers had higher CBC (HR, 13.1 [95% CI, 1.19 to 146]) and prostate (HR, 5.61 [95% CI, 1.96 to 16.0]) SPC risks than noncarriers. Survivors of BC carrying BRCA1 and BRCA2 PVs are at high SPC risk. They may benefit from enhanced surveillance and risk-reduction measures.

Exploring the Relationship Between NOD2 Risk Variants and First Decompensation Events in Cirrhotic Patients With Varices.

NOD2 mutations are associated with impaired gut mucosal barrier function. According to the systemic inflammation hypothesis, bacterial translocation is central in the development of decompensation. The aim was to evaluate whether the presence of NOD2 variants is associated with the development of first decompensation. Secondary analysis of prospectively collected consecutive patients with compensated cirrhosis, who were screened between 2014 and 2018. Patients with and without NOD2 variants were compared and stratified analysis according to the presence of varices was performed. 360 patients [239 (66%) men, median age 61 (53-69) years, 70 (19%) with NOD2 variants, 90 (25%) with varices] were followed for a median of 9 (4-16) months. Similar baseline characteristics were observed across NOD2 status groups, except for beta-blocker use (45% vs. 32% amongst variant carriers vs. non-carriers, p = 0.05). During follow-up, 34 patients (12%) developed their first decompensation, with no differences according to NOD2 status [HR 1.75 (95% CI 0.84-3.67)]. On multivariate analysis, only MELD remained an independent predictor of decompensation. Amongst patients with varices (n = 90), 18 (24.4%) carried a NOD2 variants, with a higher incidence of first decompensation [HR 3.00 (95% CI 1.08-8.32)], primarily due to ascites [HR 3.32 (95% CI 1.07-10.32)]. In this subgroup, MELD [HR 1.18 (95% CI 1.06-1.32)] and NOD2 variants [HR 2.91 (95% CI 0.95-8.89)] were determined to be independent predictors of decompensation. The presence of NOD2 risk variants leads to a greater incidence of first decompensation only in compensated patients with varices.

Melanoma arising from pre-existing naevus in carriers of a germline CDKN2A pathogenic variant.

In this study we used total body photography (TBP) to determine whether melanoma emerges from a preexisting naevi or normal-appearing skin in individuals with pathogenic CDKN2a variant. In total, 60 melanoma cases occurred in 44 CDKN2a germline carriers. In addition, we observed that 78% of the 60 included melanoma cases developed from preexisting naevi, significantly higher than the 30% reported for sporadic melanoma. This high incidence may be due to the numerous atypical naevi and early bi-allelic loss of CDKN2A. Our findings underscore the importance of routine TBP in these patients for early detection of melanoma. More than previously appreciated, the surgical removal of melanocytic naevi in these high-risk patients may prevent melanoma development.

Exome sequencing in over 63,000 patients in TIMI trials uncovers pathogenic cardiomyopathy variant carriers with high risk for heart failure and cardiovascular death

Abstract Background Rare genetic variants predisposing to cardiomyopathy (CMP) confer increased risk for adverse events. The prevalence of CMP variants in contemporary cardiovascular (CV) trials and their association with prevalent heart failure (HF) and incident HF hospitalizations (HHF) is not established. Purpose We aimed to identify carriers of rare pathogenic/likely pathogenic (PLP) variants for dilated cardiomyopathy (DCM), hypertrophic cardiomyopathy (HCM), and arrhythmogenic right ventricular cardiomyopathy (ARVC) in well-phenotyped clinical trial cohorts and to evaluate their prognostic significance. Methods Rare variants were identified from high-quality exome sequencing data from 5 large clinical trials (ENGAGE AF-TIMI 48, SAVOR-TIMI 53, PEGASUS-TIMI 54, DECLARE-TIMI 58, FOURIER (TIMI 59)). Putatively pathogenic variants for cardiomyopathy were identified using phenotype- and gene-specific curation of protein-truncating variants and high-confidence variant classifications from ClinVar. We conducted logistic regression for prevalent disease and Cox proportional hazard models for incident HHF, both adjusted by age, sex, trial, and first 10 principal components and using those without CMP variants as the reference. Sensitivity analysis by history of HF at baseline was performed for prospective outcomes. Results In 63,700 participants, we identified 866 CMP variant carriers of which 518 identified as DCM carriers (age 65±9 yrs, male 66%), 194 HCM carriers (65.5±9 yrs, male 69%), and 125 ARVC carriers (66±9 yrs, male 77%). Out of the 518 DCM variant carriers, 402 had a PLP variant in TTN. DCM variant carriers had higher odds of history of HF (Odds ratio [OR] 1.94, 1.57-2.39, p<0.0001) whereas this was not observed for HCM variant carriers (OR 1.18, 0.85-1.65, p=0.33) or ARVC variant carriers (OR 1.23, 0.80-1.90, p=0.35). Over a mean follow-up of 2.5 years, the rate of HHF was significantly higher for DCM variant carriers (Hazard ratio [HR] 1.58, 1.14-2.20, p=0.006), HCM variant carriers (HR 2.78, 1.87-4.13, p<0.001), and ARVC variant carriers (HR 1.89, 1.02-3.53, p=0.04). These associations were, if anything, stronger in patients without HF at baseline (HR 2.06, 1.30-3.25) than in patients with HF at baseline (HR 1.48, 1.12-1.95). DCM variant carriers also had a higher rate of CV death (HR 1.50, 1.05-2.12, p=0.02), which was not observed for HCM (HR 1.10, 0.57-2.11, p=0.78) or ARVC (HR 0.35, 0.09-1.41, p=0.14) variant carriers. Conclusion In 5 contemporary CV trials, CMP variant carriers have an increased rate of HHF, while DCM variant carriers also have a higher rate of CV death. These findings motivate better identification of these high-risk individuals and the incorporation of genetic testing in CV trials.

Amplicon-based long-read sequencing is useful for confirming zygosity of DSG2 variants associated with Japanese ARVC

Abstract Background Arrhythmogenic right ventricular cardiomyopathy (ARVC) is an inherited cardiomyopathy mainly caused by desmosomal gene variants. Previously, we reported that DSG2 variants were the most common in 153 Japanese ARVC probands (DSG2 multiple, n=62 and single, n=4; PKP2, n=28; others, n=7). Among the multiple DSG2 variant carriers, 22 probands were compound heterozygous carriers, and 25 were homozygous, which were confirmed by genetic testing for family members (Figure A). On the other hand, 88% of their family members with single DSG2 variant carriers were not diagnosed with ARVC, which meant that recessive DSG2 variants were the major cause of ARVC in Japan. However, zygosities in remaining 15 probands with multiple DSG2 variants were unknown due to the lack of genome data from their parents. Purpose This study aimed to clarify the zygosities of multiple DSG2 variants using amplicon-based long-read sequencing. Methods Genomic DNA was extracted from whole blood using standard protocols. The range containing multiple variants was amplified by long-range PCR. Sequencing libraries were prepared using ONT Ligation Sequencing Kit. Each library was loaded onto a flow cell for sequencing on ONT GridION Mk1 running MinKNOW control software. FASTQ files were aligned to hg38 using minimap2. Variants were called using Clair3 and phased using WhatsHap. Integrative Genomics Viewer (IGV) was used for visualization of the phased variants. Results We successfully phased all the multiple DSG2 variants found in the 15 proband. A typical result of amplicon-based LRS was shown in Figure B. It was IGV screen shot of reads at DSG2 containing c.874C>T (p.R292C) and c.1481A>C (p.D494A). The reads colored in red belonged to one haplotype, while the ones in blue belonged to the other. Each of the two variants was on different color reads, indicating they were compound heterozygous variants. Using LRS, we confirmed all the 15 probands had compound heterozygous variants in DSG2. Overall, the 62 probands (40.5%) with DSG2 variants were recessive variant carriers (Figure A). Conclusions Japanese ARVC was mainly caused by autosomal recessive variants in DSG2. Clarifying the zygosity of causative variants is crucial for the diagnosis in recessive diseases including Japanese ARVC. Amplicon-based LRS was helpful for phasing the multiple DSG2 variants directly without genetic testing of the parents.Figure AFigure B

Pathogenic cardiomyopathy gene variants inform prognosis in atrial fibrillation: results from exome sequencing in over 17,000 patients in TIMI trials

Abstract Background Rare genetic variants in cardiomyopathy genes are associated with risk of atrial fibrillation (AF), even in the absence of overt ventricular dysfunction. Data on clinical outcomes for rare genetic variant carriers among patients with AF remain sparse. Purpose We aimed to study the prognostic implication of rare pathogenic/likely pathogenic (PLP) variants for dilated cardiomyopathy (DCM), hypertrophic cardiomyopathy (HCM) and arrhythmogenic right ventricular cardiomyopathy (ARVC) in patients with AF from large, well-phenotyped randomized clinical trials. Methods and results Rare variants were called using exome sequencing data in 5 large clinical trials from the TIMI study group (ENGAGE-AF, FOURIER, SAVOR, PEGASUS, DECLARE). PLP variants for cardiomyopathy were identified using phenotype- and gene-specific curation of protein-truncating variants and high-confidence variant classifications from the ClinVar database. In 17,190 patients with a history of AF, we identified 421 (2.4%) PLP variant carriers (age 67±10, 35% women) and 16,769 (97.6%) pts without PLP variants (age 70±9, 36% women). Among these individuals, we identified 265 DCM carriers (1.5%), 106 (0.6%) HCM carriers and 54 (0.3%) ARVC carriers. In this cohort, 9,275 (54%) had a history of heart failure (HF) and 2,991 (17.4%) prior ischemic stroke. Over 2.5 years median follow-up, 1,365 pts (7.9%) were hospitalized for HF, 599 (3.5%) had an ischemic stroke, and 1147 (6.7%) died of cardiovascular causes. In logistic regression analyses adjusting for age, sex, trial, and principal components of ancestry, PLP variants were significantly associated with a history of HF (OR 1.66, p<0.0001), most notably for DCM variants (OR 1.97, 1.48-2.62, p<0.0001). PLP variants were independently associated with incident HF admissions (adj HR 1.75, 1.39-2.29, p<0.0001), most notably for HCM (adj HR 2.59, 1.68-3.99, p<0.0001) and ARVC variants (adj HR 2.50, 1.34-4.66, p=0.004). The increased risk of HF admissions remained consistent in patients with (adj HR 1.49, 1.11-2.00, p=0.009) and those without (adj HR 1.88, 1.00-3.54, p=0.049) a history of HF. PLP variants were also associated with higher risk of cardiovascular death (adj HR 1.46, 1.06-2.02, p=0.02), most notably for DCM variants (adj HR 1.77, 1.21-2.58, p=0.003). In contrast, PLP variants were not associated with history of ischemic stroke at baseline (adj OR 0.99, p=0.96) nor with incident ischemic stroke (adj HR 0.95, p=0.84). Conclusion In patients with AF, rare cardiomyopathy gene variants are associated with increased risks of HF and of cardiovascular death, but not stroke. These results, collected from large well-phenotyped randomized clinical trials, demonstrate an important prognostic implication for rare genetic variants for CMP in pts with AF.