Catarina Allegue

Molecular genetics of sudden cardiac death

Sudden cardiac death (SCD) is one of the most common causes of death. An important number of sudden deaths, especially in the young, are due to genetic heart disorders, both with structural and arrhythmogenic abnormalities. In recent years, significant advances have been made in understanding the genetic basis of SCD. Identification of the genetic causes of sudden death is important because close relatives are also at potential risk of having a fatal cardiac condition. A comprehensive post-mortem investigation is vital to determine the cause and manner of death and provides the opportunity to assess the potential risk to the family after appropriate genetic counselling. In this paper, we present an update of the different genetic causes of sudden death, emphasizing their importance for the forensic pathologist due to his relevant role in the diagnosis and prevention of SCD.

Genetics of arrhythmogenic right ventricular cardiomyopathy

Arrhythmogenic right ventricular cardiomyopathy is a rare clinical entity characterised by fibro-fatty replacement of myocardium, mainly involving right ventricular free wall, leading to malignant electrical instability and sudden cardiac death. The disease is inherited in up to 50% of cases, with incomplete penetrance and variable phenotypic expression. To date, more than 300 pathogenic mutations have been identified in 12 genes, mainly with autosomal dominant inheritance. Here, we focus on recent advances in the genetics of arrhythmogenic right ventricular cardiomyopathy. Despite continuous improvements, current genotype–phenotype studies have not contributed yet to establish a genetic risk stratification of the disease.

Negative autopsy and sudden cardiac death.

Forensic medicine defines the unexplained sudden death as a death with a non-conclusive diagnosis after autopsy. Molecular diagnosis is being progressively incorporated in forensics, mainly due to improvement in genetics. New genetic technologies may help to identify the genetic cause of death, despite clinical interpretation of genetic data remains the current challenge. The identification of an inheritable defect responsible for arrhythmogenic syndromes could help to adopt preventive measures in family members, many of them asymptomatic but at risk of sudden death. This multidisciplinary translational research requires a specialized team.

Sarcomeric gene mutations in sudden infant death syndrome (SIDS).

In developed countries, sudden infant death syndrome (SIDS) represents the most prevalent cause of death in children between 1 month and 1 year of age. SIDS is a diagnosis of exclusion, a negative autopsy which requires the absence of structural organ disease. Although investigators have confirmed that a significant percentage of SIDS cases are actually channelopathies, no data have been made available as to whether other sudden cardiac death-associated diseases, such as hypertrophic cardiomyopathy (HCM), could be responsible for some cases of SIDS. The presence of a genetic mutation in the sarcomeric protein usually affects the force of contraction of the myocyte, whose weakness is compensated with progressive hypertrophy and disarray. However, it is unclear whether in the most incipient forms, that is, first years of life, the lack of these phenotypes still confers a risk of arrhythmogenesis. The main goal of the present study is to wonder whether genetic defects in the sarcomeric proteins, previously associated with HCM, could be responsible for SIDS. We have analysed 286 SIDS cases for the most common genes implicated in HCM in adults. A total of 680 mutations localised in 16 genes were analysed by semi-automated matrix-assisted laser desorption/ionisation time-of-flight mass spectrometry (MALDITOF-MS) using the Sequenom MassARRAY(®) System. Ten subjects with completely normal hearts showed mutated alleles at nine of the genetic variants analysed, and one additional novel mutation was detected by conventional sequencing. Therefore, a genetic mutation associated with HCM may cause sudden cardiac death in the absence of an identifiable phenotype.

Determining the Pathogenicity of Genetic Variants Associated with Cardiac Channelopathies

Advancements in genetic screening have generated massive amounts of data on genetic variation; however, a lack of clear pathogenic stratification has left most variants classified as being of unknown significance. This is a critical limitation for translating genetic data into clinical practice. Genetic screening is currently recommended in the guidelines for diagnosis and treatment of cardiac channelopathies, which are major contributors to sudden cardiac death in young people. We propose to characterize the pathogenicity of genetic variants associated with cardiac channelopathies using a stratified scoring system. The development of this system was considered by using all of the tools currently available to define pathogenicity. The use of this scoring system could help clinicians to understand the limitations of genetic associations with a disease, and help them better define the role that genetics can have in their clinical routine.

New technologies in the genetic approach to sudden cardiac death in the young

Sudden cardiac death (SCD) is a major health problem and constitutes one of the most important unsolved challenges in the practice of forensic pathology due to the failure to determine the cause of death. Particularly, an important number of previously healthy young people who have died suddenly and unexpectedly are consequence of genetic heart disorders, either structural cardiomyopathies or arrhythmogenic abnormalities. The technological approach to analyze this type of genetically heterogeneous disorders is far from easy but nowadays the variety of chemistries and methodologies improves choice. This review offers to the reader a state of the art of the available technologies for the study of genetics of sudden cardiac death, including mutation screening approaches, genome wide association studies, and the recently developed next-generation sequencing.

Further evidence of the association between LQT syndrome and epilepsy in a family with KCNQ1 pathogenic variant

PURPOSE Ion channels are expressed both in the heart and in the brain, being advocated as responsible for sudden unexpected death in epilepsy but few pathogenic mutations have been identified. We aim to identify a novel gen associated with channelopathies and epilepsy in a family. METHODS We assessed a family showing epilepsy concomitant with LQTS. Index case showed prolonged QT interval. His father suffers of LQT and epilepsy. We performed a direct sequencing analysis of KCNQ1, KCNH2, KCNE1, KCNE2 and SCN5A genes. RESULTS We identified a non-synonymous heterozygous missense pathogenic mutation (p.L273F) in exon 6 of the KCNQ1 gene. All clinically affected relatives carried the same mutation. CONCLUSION We report, for a first time, a KCNQ1 mutation in a family suffering of both phenotypes, suggesting that KCNQ1 genetic variations may confer susceptibility for recurrent seizure activity increasing the risk or lead to sudden death.

Prevalence of HCM and long QT syndrome mutations in young sudden cardiac death-related cases

Cardiomyopathies and channelopathies are major causes of sudden cardiac death. The genetic study of these diseases is difficult because of their heterogenic nature not only in their genetic traits but also in their phenotypic expression. The purpose of the present study is the analysis of a wide spectrum of previously known genetic mutations in key genes related to hypertrophic cardiomyopathy (HCM), long QT syndrome (LQTS), and Brugada syndrome (BrS) development. The samples studied include cases of sudden cardiac death (SCD) in young adults and their relatives in order to identify the real impact of genetic screening of SCD in forensic cases. Genetic screening of described variation in 16 genes implicated in the development of HCM and three more genes implicated in LQTS and BrS was performed by using MassARRAY™ technology. In addition, direct sequencing of the two most prevalent genes implicated in the development of SQTL type 1 and 2 was also carried out. Genetic screening allowed us to unmask four possibly pathogenic mutation carriers in the 49 SCD cases considered; carriers of mutation represent 9% (2/23) of the probands with structural anomalies found after autopsy and 7% (1/14) of the probands with structurally normal hearts after in depth autopsy protocol. One mutation was found among 12 of the recovered SCD cases considered. In people with direct family history of sudden cardiac death, but not themselves, 11 additional mutation carriers were found. Three different mutations were found in six of the 19 LQTS patients, representing three families and two different mutations were found among six patients with previous syncope. Genetic analysis in sudden cardiac death cases could help to elucidate the cause of death, but it also can help in the prevention of future deaths in families at risk. The study presented here shows the importance and relevance of genetic screening in patients with signs of cardiac hypertrophy and in family cases with more than one relative affected.

Genetic Analysis of Arrhythmogenic Diseases in the Era of NGS: The Complexity of Clinical Decision-Making in Brugada Syndrome

Background The use of next-generation sequencing enables a rapid analysis of many genes associated with sudden cardiac death in diseases like Brugada Syndrome. Genetic variation is identified and associated with 30–35% of cases of Brugada Syndrome, with nearly 20–25% attributable to variants in SCN5A, meaning many cases remain undiagnosed genetically. To evaluate the role of genetic variants in arrhythmogenic diseases and the utility of next-generation sequencing, we applied this technology to resequence 28 main genes associated with arrhythmogenic disorders. Materials and Methods A cohort of 45 clinically diagnosed Brugada Syndrome patients classified as SCN5A-negative was analyzed using next generation sequencing. Twenty-eight genes were resequenced: AKAP9, ANK2, CACNA1C, CACNB2, CASQ2, CAV3, DSC2, DSG2, DSP, GPD1L, HCN4, JUP, KCNE1, KCNE2, KCNE3, KCNH2, KCNJ2, KCNJ5, KCNQ1, NOS1AP, PKP2, RYR2, SCN1B, SCN3B, SCN4B, SCN5A, SNTA1, and TMEM43. A total of 85 clinically evaluated relatives were also genetically analyzed to ascertain familial segregation. Results and Discussion Twenty-two patients carried 30 rare genetic variants in 12 genes, only 4 of which were previously associated with Brugada Syndrome. Neither insertion/deletion nor copy number variation were detected. We identified genetic variants in novel candidate genes potentially associated to Brugada Syndrome. These include: 4 genetic variations in AKAP9 including a de novo genetic variation in 3 positive cases; 5 genetic variations in ANK2 detected in 4 cases; variations in KCNJ2 together with CASQ2 in 1 case; genetic variations in RYR2, including a de novo genetic variation and desmosomal proteins encoding genes including DSG2, DSP and JUP, detected in 3 of the cases. Larger gene panels or whole exome sequencing should be considered to identify novel genes associated to Brugada Syndrome. However, application of approaches such as whole exome sequencing would difficult the interpretation for clinical purposes due to the large amount of data generated. The identification of these genetic variants opens new perspectives on the implications of genetic background in the arrhythmogenic substrate for research purposes. Conclusions As a paradigm for other arrhythmogenic diseases and for unexplained sudden death, our data show that clinical genetic diagnosis is justified in a family perspective for confirmation of genetic causality. In the era of personalized medicine using high-throughput tools, clinical decision-making is increasingly complex.

Natural and Undetermined Sudden Death: Value of Post-Mortem Genetic Investigation

Background Sudden unexplained death may be the first manifestation of an unknown inherited cardiac disease. Current genetic technologies may enable the unraveling of an etiology and the identification of relatives at risk. The aim of our study was to define the etiology of natural deaths, younger than 50 years of age, and to investigate whether genetic defects associated with cardiac diseases could provide a potential etiology for the unexplained cases. Methods and Findings Our cohort included a total of 789 consecutive cases (77.19% males) <50 years old (average 38.6±12.2 years old) who died suddenly from non-violent causes. A comprehensive autopsy was performed according to current forensic guidelines. During autopsy a cause of death was identified in most cases (81.1%), mainly due to cardiac alterations (56.87%). In unexplained cases, genetic analysis of the main genes associated with sudden cardiac death was performed using Next Generation Sequencing technology. Genetic analysis was performed in suspected inherited diseases (cardiomyopathy) and in unexplained death, with identification of potentially pathogenic variants in nearly 50% and 40% of samples, respectively. Conclusions Cardiac disease is the most important cause of sudden death, especially after the age of 40. Close to 10% of cases may remain unexplained after a complete autopsy investigation. Molecular autopsy may provide an explanation for a significant part of these unexplained cases. Identification of genetic variations enables genetic counseling and undertaking of preventive measures in relatives at risk.