Design and Validation of a Custom NGS Panel Targeting a Set of Lysosomal Storage Diseases Candidate for NBS Applications.

Valentina La Cognata,Maria Guarnaccia,Martino Ruggieri,Sebastiano Cavallaro,Agata Polizzi,Giovanna Morello

doi:10.3390/ijms221810064

Abstract

Lysosomal storage diseases (LSDs) are a heterogeneous group of approximately 70 monogenic metabolic disorders whose diagnosis represents an arduous challenge for clinicians due to their variability in phenotype penetrance, clinical manifestations, and high allelic heterogeneity. In recent years, the approval of disease-specific therapies and the rapid emergence of novel rapid diagnostic methods has opened, for a set of selected LSDs, the possibility for inclusion in extensive national newborn screening (NBS) programs. Herein, we evaluated the clinical utility and diagnostic validity of a targeted next-generation sequencing (tNGS) panel (called NBS_LSDs), designed ad hoc to scan the coding regions of six genes (GBA, GAA, SMPD1, IDUA1, GLA, GALC) relevant for a group of LSDs candidate for inclusion in national NBS programs (MPSI, Pompe, Fabry, Krabbe, Niemann Pick A-B and Gaucher diseases). A standard group of 15 samples with previously known genetic mutations was used to test and validate the entire flowchart. Analytical accuracy, sensitivity, and specificity, as well as turnaround time and costs, were assessed. Results showed that the Ion AmpliSeq and Ion Chef System-based high-throughput NBS_LSDs tNGS panel is a fast, accurate, and cost-effective process. The introduction of this technology into routine NBS procedures as a second-tier test along with primary biochemical assays will allow facilitating the identification and management of selected LSDs and reducing diagnostic delay.

Highlights

Lysosomal storage disorders (LSDs) comprise a heterogeneous group of rare monogenic heritable metabolism defects (~70) occurring mostly in infancy and childhood [1]
We aimed to design and evaluate both the clinical and diagnostic validity of a semi-automated and comprehensive sequencing assay based on a targeted next-generation sequencing (NGS) panel to screen variants in six genes (GBA, GAA, SMPD1, IDUA1, GLA, GALC) whose mutations are responsible for a set of LSDs (MPSI, Pompe, Krabbe, Fabry, Gaucher, and Niemann Pick A-B diseases) that are candidates for inclusion in newborn screening (NBS) programs
To assess the efficiency and accuracy of the panel, we used a reference group of standard DNA samples isolated from clinically diagnosed donor subjects (n = 15, including 4 Gaucher disease, 3 Fabry disease, 3 Pompe, 3 Niemann Pick A-B, 2 MPSI) obtained from the NIGMS Human Genetic Cell Repository at the Coriell Institute for Medical Research

Summary

Introduction

Lysosomal storage disorders (LSDs) comprise a heterogeneous group of rare monogenic heritable (inborn) metabolism defects (~70) occurring mostly in infancy and childhood [1]. The diagnostic work-up currently relies on multiple combined laboratory procedures (marking the infamous note “diagnostic odyssey”), that include either the detection of the single enzyme activity or substrates protein abundance in biological fluids, followed by biomarkers analysis and adjunct confirmatory gene sequencing tests to identify pathogenic mutations [4,5]. In this laborious and time-consuming medical process, the possibility to provide an early diagnosis plays a role of particular relevance for newborns and relative families. A timely diagnosis provides parents with realistic information about their child’s prognosis, enables appropriate genetic counseling about future pregnancies, reduces the psychological burden, and optimizes clinical management [6]

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