D.P.7 Whole exome sequencing as genetic diagnostic tool in myofibrillar myopathies

Abstract Myofibrillar myopathies (MFM) represents a group of neuromuscular disorders either familial or sporadic with clinical and genetic heterogeneity. To date, mutations in six genes are known to cause MFM, but a large number of these diseases are caused by so far unidentified gene defects. Within the NMD-chip project, we had previously studied by CGH arrays 21 sporadic patients with a clinical and immunohistochemical diagnosis of myofibrillar myopathy negative at conventional sequencing for small mutations in desmin, α B-crystallin, myotilin, Z-band alternatively spliced PDZ motif containing protein (ZASP) Bcl2-associated athanogene 3 (BAG3). The CGH analysis did not reveal any rearrangement in myofibrillar myopathy known genes. We have therefore run whole exome sequencing (WES) by Illumina GAIIe platform in seven out our 21 negative patients. This approach has allowed us to identified a mean of 60,000 SNPs or DIPs passing standard quality filters. We applied a further bioinformatics filter consisting in 880 candidate genes selected using the MedScan interactome pathway developed by Ariadne Genomics. We looked at SNPs or DIPs occurring in heterozygosity (according with a dominant model of inheritance) within all these genes. All SNPs already reported in the dbSNP database were excluded. This analysis led to the identification of a mean of 10 variations/patient. We detected (in heterozygosis) a nonsense and a missense variations in Filamin C, and 3 missense variations in three unrelated patients, within the Titin gene. Mutations in this huge gene, are known to cause a recessive form of LGMD (LGMD2J), but have not been previously related to MFMs. We have also identified stop mutations in other three novel possibly causative genes. All variations are in course of validation. We demonstrate here that WES, following facilitation of both molecular and bioinformatics procedures, may represent a “second step” powerful diagnostic tool in NMDs with high genetic heterogeneity.