A DNA resequencing array for pathogenic mutation detection in hypertrophic cardiomyopathy

Abstract

Hypertrophic cardiomyopathy (HCM) is a heterogeneous autosomal dominant cardiac disorder with a prevalence of 1 in 500. Over 450 different pathogenic mutations in at least 16 genes have been identified so far. The large allelic and genetic heterogeneity of HCM requires high-throughput, rapid, and affordable mutation detection technologies to efficiently integrate molecular screening into clinical practice. We developed a custom DNA resequencing array that contains both strands of all coding exons (160), splice-site junctions, and 5'UTR regions of 12 genes that have been clearly implicated in HCM (MYH7, MYBPC3, TNNT2, TPM1, TNNI3, MYL3, MYL2, CSRP3, PLN, ACTC, TNNC1, and PRKAG2). We analyzed a first series of 38 unrelated patients with HCM (17 familial, 21 sporadic). A total of 953,306 bp across the 38 patients were sequenced with a mean nucleotide call rate of 96.92% (range: 93-99.9%). Pathogenic mutations (single nucleotide substitutions) in MYH7, MYBPC3, TNNI3, and MYL3 (six known and six novel) were identified in 60% (10/17) of familial HCM and 10% of sporadic cases (2/21). The high-throughput HCM resequencing array is the most rapid and cost-effective tool for molecular testing of HCM to date; it thus has considerable potential in diagnostic and predictive testing, and prognostic stratification.

Hybridisation-based resequencing of 17 X-linked intellectual disability genes in 135 patients reveals novel mutations in ATRX, SLC6A8 and PQBP1

“…Small in-dels cannot be detected because of the nature of hybridisationbased sequencing, 8,9 however, as in-dels in the 17 investigated genes are much less frequent than missense mutations (Human Mutation Database), it is possible, but not very likely, that undetected in-dels are present in these 17 genes in the analysed patient cohort.…”

Recurrent and founder mutations in the Netherlands: cardiac Troponin I (TNNI3) gene mutations as a cause of severe forms of hypertrophic and restrictive cardiomyopathy*

“…An explanation for the presence of founder mutations is that some mutations are better tolerated than others. Indeed, in the literature one large family (Arg145Trp) was described with more than four gene carriers who showed late onset and low-risk disease without progression during follow-up over 3 years [14][15][16]. None of the family members in this large family had any risk factors for sudden death.…”

Next Generation Sequencing for Clinical Diagnostics-Principles and Application to Targeted Resequencing for Hypertrophic Cardiomyopathy

“…59 -62 Genetic testing approaches for HCM have included Sanger sequencing of individual genes and a gene panel approach using resequencing arrays. [62][63][64] Next generation sequencing now offers a new diagnostic approach for HCM.…”

“…62 Primers and reaction conditions were optimized yielding an average amplicon length of 5136 bp with overlaps averaging 550 bases in length. The choice of long-range PCR allowed for the design of fewer amplicons (67 total) and the longer term opportunity to investigate potential deep intronic mutations, which to date have not been extensively studied in HCM.…”