Rapid Targeted Next-Generation Sequencing Platform for Molecular Screening and Clinical Genotyping in Subjects with Hemoglobinopathies

TLDR

Hemoglobinopathies are among the most common autosomal‑recessive disorders worldwide. The authors aim to develop a comprehensive NGS test that facilitates screening and diagnosis of hemoglobinopathies and offers advantages over traditional methods. They designed an NGS panel targeting coding regions of hemoglobin genes and four modifier genes, validated it on 2,522 affected individuals, and applied it to carrier testing in 10,111 couples previously screened by conventional methods. The assay identified additional pathogenic variants in 12.1% of cases, detected 4,840 mutant alleles in 4,180 carriers, identified 186 at‑risk couples (including 35 additional at‑risk couples beyond traditional methods), and represents one of the first large‑scale population studies evaluating NGS for hemoglobinopathy screening and diagnosis.

Abstract

Hemoglobinopathies are among the most common autosomal-recessive disorders worldwide. A comprehensive next-generation sequencing (NGS) test would greatly facilitate screening and diagnosis of these disorders. An NGS panel targeting the coding regions of hemoglobin genes and four modifier genes was designed. We validated the assay by using 2522 subjects affected with hemoglobinopathies and applied it to carrier testing in a cohort of 10,111 couples who were also screened through traditional methods. In the clinical genotyping analysis of 1182 β-thalassemia subjects, we identified a group of additional variants that can be used for accurate diagnosis. In the molecular screening analysis of the 10,111 couples, we detected 4180 individuals in total who carried 4840 mutant alleles, and identified 186 couples at risk of having affected offspring. 12.1% of the pathogenic or likely pathogenic variants identified by our NGS assay, which were undetectable by traditional methods. Compared with the traditional methods, our assay identified an additional at-risk 35 couples. We describe a comprehensive NGS-based test that offers advantages over the traditional screening/molecular testing methods. To our knowledge, this is among the first large-scale population study to systematically evaluate the application of an NGS technique in carrier screening and molecular diagnosis of hemoglobinopathies.