Diagnostic Application of Targeted Next-Generation Sequencing of 80 Genes Associated with Disorders of Sexual Development

Abstract

Disorders of sexual development (DSD) are estimated to occur in 1 of 4500 births. Since the genetic etiology of DSD is highly heterogeneous, obtaining a definitive molecular diagnosis by single gene test is challenging. Utilizing a high-throughput sequencing upfront is proposed as an efficient approach to aid in the diagnosis. This study aimed to examine the diagnostic yield of next-generation sequencing in DSD. 32 DSD patients that previously received clinical examinations and single gene tests were selected, with or without a diagnosis. Prior single gene tests were masked, and then samples went through targeted next-generation sequencing of 80 genes from which the diagnostic yield was assessed. A likely diagnosis, with pathogenic or likely pathogenic variants identified, was obtained from nine of the 32 patients (i.e., 28.1%, versus 10% by single gene tests). In another five patients (15.6%), variants of uncertain significance were found. Among 18 variants identified (i.e., 17 single nucleotide variants and one small deletion), eight had not been previously reported. This study supports the notion that next-generation sequencing can be an efficient tool in the clinical diagnosis and variant discovery in DSD.

Multiplex targeted high‐throughput sequencing in a series of 352 patients with congenital limb malformations

“…Overall, in this series of 352 patients, 138 pathogenic or likely pathogenic variants were identified, among which 26 CNVs, and 22 variants of uncertain significance. Our approach revealed to be a useful strategy, providing a molecular diagnosis in 35.2% of patients, similar to other heterogeneous conditions (50% for inborn errors of metabolism (Yubero et al, ), 47.3% for myopathies and muscular dystrophies (Stehlikova et al, ), 39% for intellectual disability (Martinez et al, ), 28.5% for epileptic encephalopathy (Kothur et al, ), 28.1% for disorders of sexual development (Fan et al, ), 24.5% for developmental eye disorders (Patel et al, ), 20% for primary arrhythmia syndrome and cardiomyopathy (Robyns et al, ), and 18% for inherited polyneuropathy (Wang et al, ). A 7.7% increase was observed in our diagnostic performance, compared to our previous workflow (diagnostic yield 27.5% before 2014, data not shown).…”

A rare ANOS1 variant in siblings with Kallmann syndrome identified by whole exome sequencing

“…The presence of this hemizygous variant was confirmed in both brothers and heterozygous in their mother by Sanger sequencing at GENEWIZ (http://www.genecards.org) (Figure ). This variant is not reported in the three genomic databases studied, but has been found associated with Kallmann syndrome twice in the literature (Jiang et al ., ; Fan et al ., ). Therefore, we conclude that this is a pathogenic variant according to American College of Medical Genetics guidelines and the cause of Kallmann syndrome in this family.…”

“…It was subsequently reported in a study performing next‐generation sequencing of genes associated with disorders of sexual development (DSD) to identify novel disease‐causing variants. In this study, there were four patients with clinical hypogonadotropic hypogonadism and one was identified to have the c.1267C>T variant in ANOS1 (Fan et al ., ). Although our patients may fit into the spectrum of DSD, none of them fulfilled diagnostic criteria for autism on evaluation.…”

The molecular pathways underlying early gonadal development

“…While we know the genetic aetiology of a small number of DSDs, up to as many as 75% of individuals with a DSD will remain without a genetic diagnosis (Arboleda et al 2014). Even with whole genome sequencing, it is often difficult to identify functional variants and causal mutations, rendering many sequencing approaches somewhat ineffective (Fan et al 2017). Gene expression levels, epigenetic modifiers and genetic background, along with the type of mutation and its functional consequence can all influence the resulting phenotype for both humans and mice.…”