Post-mortem genetic analysis in juvenile cases of sudden cardiac death

doi:10.1016/j.forsciint.2014.10.004

Forensic Science International

Volume 245, December 2014, Pages 30-37

https://doi.org/10.1016/j.forsciint.2014.10.004 Get rights and content

Highlights

•
Molecular autopsy should be implemented in forensic protocols.
•
Nearly 40% of sudden death young cases carry a cardiac potentially pathogenic variant.
•
It is crucial to undertake a careful genetic analysis in a clinical context.
•
Genetic analyses help to identify relatives at risk of sudden death.

Abstract

Background

The reason behind a sudden death of a young individual remains unknown in up to 50% of postmortem cases. Pathogenic mutations in genes encoding heart proteins are known to cause sudden cardiac death.

Objective

The aim of our study was to ascertain whether genetic alterations could provide an explanation for sudden cardiac death in a juvenile cohort with no-conclusive cause of death after comprehensive autopsy.

Methods

Twenty-nine cases <15 years showing no-conclusive cause of death after a complete autopsy were studied. Genetic analysis of 7 main genes associated with sudden cardiac death was performed using Sanger technology in low quality DNA cases, while in good quality cases the analysis of 55 genes associated with sudden cardiac death was performed using Next Generation Sequencing technology.

Results

Thirty-five genetic variants were identified in 12 cases (41.37%). Ten genetic/variants in genes encoding cardiac ion channels were identified in 8 cases (27.58%). We also identified 9 cases (31.03%) carrying 25 genetic variants in genes encoding structural cardiac proteins. Nine cases carried more than one genetic variation, 5 of them combining structural and non-structural genes.

Conclusions

Our study supports the inclusion of molecular autopsy in forensic routine protocols when no conclusive cause of death is identified. Around 40% of sudden cardiac death young cases carry a genetic variant that could provide an explanation for the cause of death. Because relatives could be at risk of sudden cardiac death, our data reinforce their need of clinical assessment and, if indicated, of genetic analysis.

Introduction

Sudden death in people younger than 15 years old is a rare event, with an incidence between 1–5/100,000 individuals each year in developed countries [1]. Despite this low prevalence, when a death occurs in this juvenile population, it carries a tremendous impact in both the family and community. Sudden death constitutes one of the most important unsolved challenges in the practice of forensic pathology. Several studies have reported that most part of sudden deaths in the young (<40 years) is of cardiac origin (sudden cardiac death -SCD-), mainly caused by structural heart abnormalities identifiable at autopsy (cardiomyopathies) [2]. However, in 10–35% of these deaths, no structural alterations can be identified. In these cases a channelopathy, a genetic disease of the cardiac ion channels, is suspected [3], [4], [5]. Both groups of cardiac alterations are due to inherited genetic defects, thus family members of the deceased individual are at risk of sudden death [6]. This fact carries important implications in diagnosis and counselling of relatives. Though, the application of genetic testing in routine forensic investigation, to benefit diagnosis and possible family prevention, remains still very limited [7].

Currently, numerous genes have been associated with SCD but most part in low frequency [8], [9]. However, in these last years, genetic research has focused on the identification of pathogenic mutations in seven main genes (SCN5A, KCNQ1, KCNH2, KCNE1, KCNE2, KCNE3, and RyR2) associated with channelopathies (like Brugada Syndrome-BrS-, Long QT Syndrome -LQTS-, Short QT Syndrome -SQTS-, and Catecholaminergic Polymorphic Ventricular Tachycardia -CPVT-) and 7 main genes (MYBPC3, MYH7, PKP2, DSC2, DSP, DSC2, and LMNA) associated with cardiomyopathies (like Hypertrophic Cardiomyopathy -HCM-, Arrhythmogenic Right Ventricular Cardiomyopathy -ARVC-, and Dilated Cardiomyopathy -DCM-).

Genetic analysis of these genes can help in the identification of the cause of death, even using mRNA [10], improving the evaluation of relatives at potential risk. Traditional Sanger sequencing is expensive to undertake this extensive analysis. However, new genetic technologies (Next generation Sequencing -NGS-) have emerged as a cost-effective technology for broad genetic studies [11], [12], [13]. The ability to perform analysis of large amount of genes at once has been brought to the clinical arena of several medical specialities, including cardiology. It is no secret though, that the large amount of data generated is causing difficulties in clinical interpretation, especially when dealing with genetic variants of unknown significance (GVUS) or genetic variants in less common genes. In our study we analyzed a cohort of post-mortem cases, aged less than 15 years old, in order to investigate the role of genetics in death causality.

Section snippets

Forensics

A complete autopsy examination was performed according to current international regulations [14]. Our inclusion criteria was: (a) age <15 years, (b) non-conclusive cause of death after complete autopsy, (c) no signs of congenital heart alterations, cardiac infarct or other macroscopic anomalies, (d) blood obtained <48 h after death. The study was approved by the ethics committee of our Hospital, and follows the Helsinki II declaration.

DNA sample

Genomic DNA was extracted with Chemagic MSM I from

Results

A total of 29 cases collected at Institut de Medicina Legal de Catalunya (IMLC), from April 2012 until June 2013, were included in our study. All cases included in our study were <15 years old (mean age 3.29 years old, with a wide range of death from 21 days to 14 years old) and complete autopsy concluded an undetermined cause of death. Toxicological results were negative in all cases. Macroscopic analysis did not showed any anomaly. Microscopic/histological analyses were also negative in all

Discussion

Our study identifies genetic alterations associated with SCD that provide a cause of death in a juvenile post-mortem cohort. Our cohort showed a global gender ratio 1:1, in divergence with published data about higher incidence of sudden death in male gender, mainly at young ages [15]. Specifically, in individuals younger than 1 year old, the number of females studied was higher (3:1). This fact could be a spurious result due to reduced number of cases. Regarding the cause of death, it has been

Conclusion

We have identified a potentially pathogenic genetic variation in 41% of SCD young cases (of which nearly 30% in arrhythmogenic genes), supporting the implementation of the molecular autopsy in forensic protocols. Despite present lack of knowledge in pathogenicity classification of ambiguous genetic variants, identification of pathogenic or potentially pathogenic genetic variations in cases of unexplained sudden death enables the undertaking of clinical assessment, genetic counselling and

Conflict of interest

Dr. Ramon Brugada is consultant of Ferrer-inCode. The other authors declare no conflicts of interest to disclose.

Acknowledgements

This study has been funded by Societat Catalana de Cardiologia (SCC), Fundacion Eugenio Rodriguez Pascual, Academia de Ciències Mèdiques de Catalunya i Balears (ACMCB), and Fundació “Obra social La Caixa”.

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The data demonstrate that increased myofilament Ca²⁺ sensitivity is not sufficient to induce arrhythmias in the Mybpc3-KI mouse model and suggest that reduced K⁺ currents can be a pro-arrhythmic trigger in Hom-KI mice, probably already in early disease stages. However, neither data from EHTs nor from left ventricular samples indicate relevant reduction of K⁺ currents in human HCM. Therefore, our study highlights the species difference between mouse and human and emphasizes the importance of research in human samples and human-like models.
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Sudden cardiac death (SCD) in young people often has a genetic cause. Consequently, the results of “molecular autopsy” may have important implications for their relatives. Our objective was to evaluate the diagnostic yield of a molecular autopsy program using next-generation sequencing.
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¹: These authors equally contributed to this work.

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Post-mortem genetic analysis in juvenile cases of sudden cardiac death

Highlights

Abstract

Background

Objective

Methods

Results

Conclusions

Introduction

Section snippets

Forensics

DNA sample

Results

Discussion

Conclusion

Conflict of interest

Acknowledgements

Forensic Sci. Int.

Cell

Heart Rhythm

Heart Rhythm

Heart Rhythm

Mayo Clin. Proc. Mayo Clin.

Heart Rhythm

Heart Rhythm

J. Am. Coll. Cardiol.

Mayo Clin. Proc. Mayo Clin.

Int. J. Cardiol.

Lancet

Forensic Sci. Int.

Heart Rhythm

J. Am. Coll. Cardiol.

Molecular autopsy of sudden unexplained death in the young

Am. J. Forensic Med. Pathol.

Sudden death in persons younger than 40 years of age: incidence and causes

Eur. J. Cardiovasc. Prev. Rehabil.

Prevalence of long-QT syndrome gene variants in sudden infant death syndrome

Circulation