Genomic Studies in a Large Cohort of Hearing Impaired Italian Patients Revealed Several New Alleles, a Rare Case of Uniparental Disomy (UPD) and the Importance to Search for Copy Number Variations

Abstract

Hereditary hearing loss (HHL) is a common disorder characterized by a huge genetic heterogeneity. The definition of a correct molecular diagnosis is essential for proper genetic counseling, recurrence risk estimation, and therapeutic options. From 20 to 40% of patients carry mutations in GJB2 gene, thus, in more than half of cases it is necessary to look for causative variants in the other genes so far identified (~100). In this light, the use of next-generation sequencing technologies has proved to be the best solution for mutational screening, even though it is not always conclusive. Here we describe a combined approach, based on targeted re-sequencing (TRS) of 96 HHL genes followed by high-density SNP arrays, aimed at the identification of the molecular causes of non-syndromic HHL (NSHL). This strategy has been applied to study 103 Italian unrelated cases, negative for mutations in GJB2, and led to the characterization of 31% of them (i.e., 37% of familial and 26.3% of sporadic cases). In particular, TRS revealed TECTA and ACTG1 genes as major players in the Italian population. Furthermore, two de novo missense variants in ACTG1 have been identified and investigated through protein modeling and molecular dynamics simulations, confirming their likely pathogenic effect. Among the selected patients analyzed by SNP arrays (negative to TRS, or with a single variant in a recessive gene) a molecular diagnosis was reached in ~36% of cases, highlighting the importance to look for large insertions/deletions. Moreover, copy number variants analysis led to the identification of the first case of uniparental disomy involving LOXHD1 gene. Overall, taking into account the contribution of GJB2, plus the results from TRS and SNP arrays, it was possible to reach a molecular diagnosis in ~51% of NSHL cases. These data proved the usefulness of a combined approach for the analysis of NSHL and for the definition of the epidemiological picture of HHL in the Italian population.

Lights and Shadows in the Genetics of Syndromic and Non-Syndromic Hearing Loss in the Italian Population

“…On the other hand, no deletions in GJB6 have been detected, and the A1555G mitochondrial mutation. These results, combined with the outcomes of the previous works [ 10 , 52 ] suggest that GJB6 and MT-RNR1 are not a common cause of NSHL in the Italian population, despite their relevant role in other areas of the world.…”

“…The implementation of next-generation sequencing technologies (NGS), together with molecular karyotyping (e.g., Single Nucleotide Polymorphism (SNP) array or Comparative Genomic Hybridization (CGH) array) and other validating assays (e.g., Multiplex Ligation Probe Amplification—MLPA), has dramatically increased the diagnostic rate of HHL, leading to the identification of several mutations and Copy Number Variations (CNVs) in known deafness genes, as well as to the discovery of new disease genes [ 10 , 11 , 12 , 13 ]. The possibility to simultaneously screen large number of genes is essential to address with the genetic heterogeneity of HHL.…”

Mutational Spectrum and Clinical Features of Patients with LOXHD1 Variants Identified in an 8074 Hearing Loss Patient Cohort

“…Few previous reports have discussed the prevalence of the LOXHD1 gene amongst SNHL patients. In relatively large cohorts that evaluated more than 100 patients, the prevalence of this gene was estimated to 0.71% in American SNHL patients (8/1119), 1.5% in the Netherlands (3/200) and 0.97% in Italy (1/103) [19,22,28]. This result is similar with our study in the American patients, but noticeably higher in the other two countries.…”

“…In this study, we identified 28 individuals with LOXHD1 variants including 13 novel variants by MPS screening. A total of 44 LOXHD1 variants have been reported to date from various countries, and this study is the largest analysis of LOXHD1 -related HL yet (Table 3) [5,14,15,16,17,18,19,20,21,22,23,24,25,26,27,28,29,30,31,32,33,34]. Among the identified variants, 11 were non-truncating variations (52.4%) and 10 were truncating variations (47.6%).…”

The Impact of Modern Technologies on Molecular Diagnostic Success Rates, with a Focus on Inherited Retinal Dystrophy and Hearing Loss

“…Moreover, in 2013, Roosing et al described maternal UPD of chromosome 6, which included a pathogenic TULP1 variant responsible for the cone dystrophy phenotype of the proband. For HL, several cases of UPD have been described as well, affecting chromosome 1 ( USH2A ) [ 183 ], chromosome 13 ( GJB2) [ 182 ], and chromosome 18 ( LOXHD1 ) [ 182 , 184 ].…”