Identification of Genetic Causes of Inherited Peripheral Neuropathies by Targeted Gene Panel Sequencing

Abstract

Inherited peripheral neuropathies (IPN), which are a group of clinically and genetically heterogeneous peripheral nerve disorders including Charcot-Marie-Tooth disease (CMT), exhibit progressive degeneration of muscles in the extremities and loss of sensory function. Over 70 genes have been reported as genetic causatives and the number is still growing. We prepared a targeted gene panel for IPN diagnosis based on next generation sequencing (NGS). The gene panel was designed to detect mutations in 73 genes reported to be genetic causes of IPN or related peripheral neuropathies, and to detect duplication of the chromosome 17p12 region, the major genetic cause of CMT1A. We applied the gene panel to 115 samples from 63 non-CMT1A families, and isolated 15 pathogenic or likely-pathogenic mutations in eight genes from 25 patients (17 families). Of them, eight mutations were unreported variants. Of particular interest, this study revealed several very rare mutations in the SPTLC2, DCTN1, and MARS genes. In addition, the effectiveness of the detection of CMT1A was confirmed by comparing five 17p12-nonduplicated controls and 15 CMT1A cases. In conclusion, we developed a gene panel for one step genetic diagnosis of IPN. It seems that its time- and cost-effectiveness are superior to previous tiered-genetic diagnosis algorithms, and it could be applied as a genetic diagnostic system for inherited peripheral neuropathies.

Keywords: DCTN1; MARS; SPTLC2; charcot-Marie-Tooth disease (CMT); gene panel; inherited peripheral neuropathy.

Fig. 1.

Pedigrees of inherited peripheral neuropathy families with novel pathogenic or likely pathogenic mutations. Arrows indicate the proband in each family. Open and black symbols represent the unaffected and affected individuals, respectively, whereas grey symbols indicate individuals with no clinical information available. Genotypes of the mutations are indicated at the bottom of all examined individuals (mutant allele: bold underlined): (A) FC541 family with c.47T > C in PMP22, (B) FC703 with c.929G > A and c.3272G > T in SH3TC2, (C) FC156 with c.154T > G in MPZ, (D) FC141 with c.262T > C in MPZ, (E) FC180 with c.1019A > G in DCTN1, (F) FC725 with c.286G > C in GJB1, and (G) FC459 with c.435G > T in SPTLC2.

Fig. 2.

Sequencing chromatograms and conservation analysis of novel causative mutations. (A) Confirmation of the causative mutations using the Sanger’s sequencing method. (B) Alignment of the amino acid sequences for the mutation sites and around regions in several vertebrate species: Homo sapiens, Bos taurus, Rattus norvegicus, Mus musculus, Gallus gallus, Xenopus laevis, and Danio rerio.