Pathogenic Intronic Splice-Affecting Variants in MYBPC3 in Three Patients with Hypertrophic Cardiomyopathy

Katherine A Wood,Raymond T O’Keefe,James Eden,Claire Hopton,William G Newman,Huw B Thomas,Jamie M Ellingford

doi:10.3390/cardiogenetics11020009

Abstract

Genetic variants in MYBPC3 are one of the most common causes of hypertrophic cardiomyopathy (HCM). While variants in MYBPC3 affecting canonical splice site dinucleotides are a well-characterised cause of HCM, only recently has work begun to investigate the pathogenicity of more deeply intronic variants. Here, we present three patients with HCM and intronic splice-affecting MYBPC3 variants and analyse the impact of variants on splicing using in vitro minigene assays. We show that the three variants, a novel c.927-8G>A variant and the previously reported c.1624+4A>T and c.3815-10T>G variants, result in MYBPC3 splicing errors. Analysis of blood-derived patient RNA for the c.3815-10T>G variant revealed only wild type spliced product, indicating that mis-spliced transcripts from the mutant allele are degraded. These data indicate that the c.927-8G>A variant of uncertain significance and likely benign c.3815-10T>G should be reclassified as likely pathogenic. Furthermore, we find shortcomings in commonly applied bioinformatics strategies to prioritise variants impacting MYBPC3 splicing and re-emphasise the need for functional assessment of variants of uncertain significance in diagnostic testing.

Highlights

Clinical Characteristics Associated with Three Intronic myosin-binding protein C3 (MYBPC3) Variants
The affected nucleotide is 8 base pairs upstream of the canonical 30 splice site in intron 11, and potentially creates a new cryptic 30 splice site AG. This c.927-8G>A variant has not been described previously in hypertrophic cardiomyopathy (HCM) patients in the literature, is not present in the gnomAD population genomic sequence database and is classified as a variant of uncertain significance. This variant has been identified in seven additional unrelated individuals with HCM in diagnostic laboratories performing cardiomyopathy testing in the United Kingdom
A variant in the adjacent residue (c.927-9G>A) has been reported as likely pathogenic, analysis of patient RNA derived from blood for this adjacent variant did not confirm any splicing errors caused by the variant [37,44]

Summary

Introduction

Received: 30 March 2021Accepted: 25 May 2021Published: 2 June 2021Publisher’s Note: MDPI stays neutral with regard to jurisdictional claims in published maps and institutional affiliations.Licensee MDPI, Basel, Switzerland.Attribution (CC BY) license (https://creativecommons.org/licenses/by/ 4.0/).Hypertrophic cardiomyopathy (HCM) is a relatively common genetic disorder (with a prevalence of 1:500) characterised by left ventricular cardiomyopathy, a non-dilated left ventricle and normal, or increased, ejection fraction, and is associated with myocardial fibre disarray [1,2,3]. HCM is usually asymmetrical and develops in the absence of an identifiable secondary cause such as hypertension or aortic valvular stenosis. Patients with

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Conclusion