Journal Article

Combined biochemical profiling and DNA sequencing in the expanded newborn screening for inherited metabolic diseases: the experience in an Italian reference center

Barretta, Fernando, Albano, Lucia, Ruoppolo, Margherita, Annunziata, Patrizia, Cacchiarelli, Davide, Riccardo, Sara, Fusco, Tonya, Alagia, Marianna, Cesana, Marcella, Brunetti-Pierri, Nicola, Grimaldi, Antonio, Crisci, Daniela, Cacciapuoti, Maria Teresa, Verde, Alessandra, Parenti, Giancarlo, Vaccaro, Lorenzo, Frisso, Giulia, Fecarotta, Simona, Tarallo, Antonietta, Vallone, Fabiana, Rossi, Alessandro, De Santis, Rosa, Colantuono, Chiara

2025

Overview

Background Newborn screening (NBS) programs have significantly improved the health and outcomes of patients with inherited metabolic disorders (IMDs). Methods based on liquid chromatography/mass spectrometry (LC–MS/MS) analysis are viewed worldwide as the gold standard procedure for the expanded NBS programs for these disorders. Advanced molecular technologies point to genomic sequencing as an alternative and feasible strategy for the screening of genetic diseases, including IMDs. However, each of the two approaches has potential limitations when used as a first-tier analysis. In this study, we tested a workflow-based parallel biochemical and sequencing analyses to determine whether this approach could improve the diagnostic outcome. Results For each patient identified by LC–MS/MS as positive, we performed both the biochemical confirmatory tests and next-generation sequencing (NGS) procedures from the same Dried Blood Spot (DBS). NGS analysis was based on applying Exome Sequencing libraries, limiting the analysis to 105 actionable genes involved in IMDs. This allows overtaking the actual limitations of NBS on DBS, enhancing our capacity to identify variants that can drive a genetic disease. Through this approach, we could reach 100% of cases solved, with 37.9% of cases (41/108) for which the combination of the biochemical and NGS analysis was indispensable for a correct diagnosis. In total, we could identify 17 affected, 34 false positives, 12 individuals referred to us for maternal conditions. In 45 newborns the molecular analysis showed heterozygosity for mutations in one or more of the genes analyzed, with results compatible with the biochemical profile indicative of NBS positivity. Conclusions In this study, we validated the performance of the proposed workflow. The advantage of this approach is limiting molecular analysis only to positive newborns and using a restricted panel of 105 genes relevant for the expanded NBS, with a 100% rate of diagnosis and potential reduction of the costs related to NBS procedures and reduced impact on patients and families.

• Publisher: BioMed Central,BioMed Central Ltd,Springer Nature B.V,BMC
• Subject: Chromatography, Liquid / Clinical chemistry / Diagnosis / Disease / DNA fingerprinting / DNA sequencing / Female / Genetic disorders / Genetic screening / Genomics / Heterozygosity / High-Throughput Nucleotide Sequencing - methods / Homocysteine / Hospitals / Human Genetics / Humans / Infant, Newborn / Infants (Newborn) / Inherited metabolic disorders (IMDs) / Italy / Laboratories / Liquid chromatography / Male / Mass spectroscopy / Medical examination / Medical screening / Medicine / Medicine & Public Health / Metabolic Diseases - diagnosis / Metabolic Diseases - genetics / Metabolic disorders / Metabolism, Inborn Errors - diagnosis / Metabolism, Inborn Errors - genetics / Metabolism, Inborn errors of / Metabolites / Methods / Neonatal Screening - methods / Neonates / Newborn babies / Newborn screening (NBS) / Next-generation sequencing / Next-generation sequencing (NGS) / Nucleotide sequencing / Oxidation / Pediatrics / Pharmacology/Toxicology / Sequence Analysis, DNA - methods / Tandem Mass Spectrometry - methods / Whole exome sequencing (WES) / Whole genome sequencing