New Mutations in NEB Gene Discovered by Targeted Next-Generation Sequencing in Nemaline Myopathy Italian Patients

Abstract Nemaline myopathy represents a group of clinically and genetically heterogeneous neuromuscular disorders. Different clinical-genetic entities have been characterized in thelastfewyears,withimplicationsfordiagnosticsandgenetic counseling. Fifty percent of nemaline myopathy forms are due to NEB mutations, but genetic analysis of this large and complexgeneby Sanger sequencingistimeconsumingand expensive. We selected 10 Italian patients with clinical and biopsy features suggestive for nemaline myopathy and negative for ACTA1, TPM2 and TPM3 mutations. We applied a targeted next-generation sequencing strategy designed to analyse NEB coding regions, the relative full introns and the promoter. We also evaluated copy number variations (by CGH array) and transcriptional changes by RNA Sanger sequencing, whenever possible. This combined strategy revealed 11 likely pathogenic variants in 8 of 10 patients. The molecular diagnosis was fully achieved in 3 of 8 patients, while only one heterozygous muta tionwas observed in5subjects. This approachrevealed tobea fast and cost-effective way to analyse the large NEB gene in a small group of patients and might be promising for the detection of pathological variants of othergenes featuring large coding regions and lacking mutational hotspots.