Development of DNA Confirmatory and High-Risk Diagnostic Testing for Newborns Using Targeted Next-Generation DNA Sequencing

• Published in: Genetics in medicine Vol. 17; no. 5; pp. 337 - 347
• Main Authors: Bhattacharjee, Arindam, Sokolsky, Tanya, Wyman, Stacia K., Reese, Martin G., Puffenberger, Erik, Strauss, Kevin, Morton, Holmes, Parad, Richard B., Naylor, Edwin W.
• Format: Journal Article
• Language: English
• Published: United States Elsevier Inc 01.05.2015; Elsevier Limited
• Subjects: Algorithms; Computational Biology - methods; Genetic Testing - methods; Genetic Variation; Genotype; High-Throughput Nucleotide Sequencing - methods; Humans; Infant, Newborn; Neonatal Screening; neonate; newborn screening; next-generation sequencing; Reproducibility of Results; second-tier testing; Sensitivity and Specificity; Workflow; neonate; next-generation sequencing; newborn screening; second-tier testing
• ISSN: 1098-3600; 1530-0366
• DOI: 10.1038/gim.2014.117

Genetic testing is routinely used for second-tier confirmation of newborn sequencing results to rule out false positives and to confirm diagnoses in newborns undergoing inpatient and outpatient care. We developed a targeted next-generation sequencing panel coupled with a variant processing pipeline and demonstrated utility and performance benchmarks across multiple newborn disease presentations in a retrospective clinical study. The test utilizes an in silico gene filter that focuses directly on 126 genes related to newborn screening diseases and is applied to the exome or a next-generation sequencing panel called NBDx. NBDx targets the 126 genes and additional newborn-specific disorders. It integrates DNA isolation from minimally invasive biological specimens, targeted next-generation screening, and rapid characterization of genetic variation. We report a rapid parallel processing of 8 to 20 cases within 105 hours with high coverage on our NBDx panel. Analytical sensitivity of 99.8% was observed across known mutation hotspots. Concordance calls with or without clinical summaries were 94% and 75%, respectively. Rapid, automated targeted next-generation sequencing and analysis are practical in newborns for second-tier confirmation and neonatal intensive care unit diagnoses, laying a foundation for future primary DNA-based molecular screening of additional disorders and improving existing molecular testing options for newborns. Genet Med17 5, 337–347.