Performant Mutation Identification Using Targeted Next-Generation Sequencing of 14 Thoracic Aortic Aneurysm Genes

Abstract

At least 14 causative genes have been identified for both syndromic and nonsyndromic forms of thoracic aortic aneurysm/dissection (TAA), an important cause of death in the industrialized world. Molecular confirmation of the diagnosis is increasingly important for gene-tailored patient management but consecutive, conventional molecular TAA gene screening is expensive and labor-intensive. To circumvent these problems, we developed a TAA gene panel for next-generation sequencing of 14 TAA genes. After validation, we applied the assay to 100 Marfan patients. We identified 90 FBN1 mutations, 44 of which were novel. In addition, Multiplex ligation-dependent probe amplification identified large deletions in six of the remaining samples, whereas false-negative results were excluded by Sanger sequencing of FBN1, TGFBR1, and TGFBR2 in the last four samples. Subsequently, we screened 55 syndromic and nonsyndromic TAA patients. We identified causal mutations in 15 patients (27%), one in each of the six following genes: ACTA2, COL3A1, TGFBR1, MYLK, SMAD3, SLC2A10 (homozygous), two in NOTCH1, and seven in FBN1. We conclude that our approach for TAA genetic testing overcomes the intrinsic hurdles of consecutive Sanger sequencing of all candidate genes and provides a powerful tool for the elaboration of clinical phenotypes assigned to different genes.

Loss-of-function mutations in the X-linked biglycan gene cause a severe syndromic form of thoracic aortic aneurysms and dissections

“…Samples were paired-end sequenced using 2x100 bp on a HiSeq1500 in high output mode (Illumina, San Diego, CA). Enrichment, data analysis, variant annotation and confirmation of the variants were performed as previously described 10,11. GRCh37 was used as a reference build.…”

Notch1 haploinsufficiency causes ascending aortic aneurysms in mice

“…Subsequently, there have been multiple reports of NOTCH1 mutations in patients with BAV and associated aortopathy (28)(29)(30). NOTCH1 encodes one of the 4 transmembrane receptors (NOTCH1-4) that functions in the highly conserved Notch signaling pathway, which is a critical player in cardiovascular development and disease.…”

Findings made in gene panel to whole genome sequencing: data, knowledge, ethics – and consequences?

“…The latter is less sensitive [34] and has lower throughput coupled with higher cost when applied for the analysis of many and/or large genes [35]. A wide spectrum of instruments has entered the market, now covering the full range from low throughput to genome-scale sequencing.…”