Set up and validation of a targeted NGS approach for the diagnosis of lysosomal storage disorders

In this study, we evaluated an LSD targeted sequencing panel as a tool capable to potentially reverse this 'classic' diagnostic route. The panel includes 50 LSD genes and 230 Conserved Intronic Fragments (CIF), obtained comparing the intronic sequences of 33 placental mammals.For the validation phase we analyzed 56 positive controls, 13 biochemically diagnosed and 9 undiagnosed patients. Disease-causing variants were identified in 66% of the positive control alleles and in 62% of the biochemically diagnosed patients. Three undiagnosed patients were diagnosed. Eight patients undiagnosed by the panel were analyzed by whole exome sequencing: for two of them the disease-causing variants were identified. Five patients, undiagnosed by both panel and exome analyses, were investigated through array CGH: one of them was diagnosed. CIF analysis performed in cases unresolved by the first-level analysis, evidenced no candidate intronic variants.Targeted sequencing has low sequencing costs and short sequencing time. However a coverage above 60-80X must be assured and/or Sanger validation should be performed. Moreover, it must be supported by a thorough clinical phenotyping.

Source: The Journal of Molecular Diagnostics - February 8, 2020 Category: Pathology Source Type: research