Loss-of-function mutations in MYO15A and OTOF cause non-syndromic hearing loss in two Yemeni families.

BACKGROUND: Hearing loss is a rare hereditary deficit that is rather common among consanguineous populations. Autosomal recessive non-syndromic hearing loss is the predominant form of hearing loss worldwide. Although prevalent, hearing loss is extremely heterogeneous and poses a pitfall in terms of diagnosis and screening. Using next-generation sequencing has enabled a rapid increase in the identification rate of genes and variants in heterogeneous conditions, including hearing loss. We aimed to identify the causative variants in two consanguineous Yemeni families affected with hearing loss using targeted next-generation sequencing (clinical exome sequencing). The proband of each family was presented with sensorineural hearing loss as indicated by pure-tone audiometry results. RESULTS: We explored variants obtained from both families, and our analyses collectively revealed the presence and segregation of two novel loss-of-function variants: a frameshift variant, c.6347delA in MYO15A in Family I, and a splice site variant, c.5292-2A > C, in OTOF in Family II. Sanger sequencing and PCR-RFLP of DNA samples from 130 deaf and 50 control individuals confirmed that neither variant was present in our in-house database. In silico analyses predicted that each variant has a pathogenic effect on the corresponding protein. CONCLUSIONS: We describe two novel loss-of-function variants in MYO15A and OTOF that cause autosomal recessive non-syndromic hearing loss in Yemeni families. Our findings are consistent with previously reported pathogenic variants in the MYO15A and OTOF genes in Middle Eastern individuals and suggest their implication in hearing loss.

An OTOF Frameshift Variant Associated with Auditory Neuropathy Spectrum Disorder.

BACKGROUND: Auditory Neuropathy Spectrum Disorder (ANSD) is manifested as impairment of auditory nerve activity but preservation of the outer hair cell function. OBJECTIVE: This study was to detect the disease-causing gene and variant(s) in a Chinese ANSD family. METHODS: A four-generation consanguineous Chinese ANSD family and 200 unrelated healthy controls were enrolled. Exome sequencing and Sanger sequencing were applied to identify the genetic basis for ANSD in this family. RESULTS: Exome sequencing detected a c.1236delC variant of the otoferlin gene in an apparently homozygous state. Sanger sequencing confirmed that the variant co-segregating with the phenotype of hearing impairments in this family. The variant was not detected in 200 healthy controls. The c.1236delC alteration may result in a truncated otoferlin missing the C2C-C2F domains and the C-terminal transmembrane domain, and thus severely damages Ca2+-dependent synaptic vesicle fusion and targeting function of the otoferlin. CONCLUSION: Our study suggested that the c.1236delC alteration in the otoferlin gene may be the disease-causing variant in this family, and also shed new light on genetic counseling to this ANSD family.

Novel compound heterozygous mutations in the OTOF Gene identified by whole-exome sequencing in auditory neuropathy spectrum disorder.

BACKGROUND: Many hearing-loss diseases are demonstrated to have Mendelian inheritance caused by mutations in single gene. However, many deaf individuals have diseases that remain genetically unexplained. Auditory neuropathy is a sensorineural deafness in which sounds are able to be transferred into the inner ear normally but the transmission of the signals from inner ear to auditory nerve and brain is injured, also known as auditory neuropathy spectrum disorder (ANSD). The pathogenic mutations of the genes responsible for the Chinese ANSD population remain poorly understood. METHODS: A total of 127 patients with non-syndromic hearing loss (NSHL) were enrolled in Guangxi Zhuang Autonomous Region. A hereditary deafness gene mutation screening was performed to identify the mutation sites in four deafness-related genes (GJB2, GJB3, 12S rRNA, and SLC26A4). In addition, whole-exome sequencing (WES) was applied to explore unappreciated mutation sites in the cases with the singularity of its phenotype. RESULTS: Well-characterized mutations were found in only 8.7% (11/127) of the patients. Interestingly, two mutations in the OTOF gene were identified in two affected siblings with ANSD from a Chinese family, including one nonsense mutation c.1273C > T (p.R425X) and one missense mutation c.4994 T > C (p.L1665P). Furthermore, we employed Sanger sequencing to confirm the mutations in each subject. Two compound heterozygous mutations in the OTOF gene were observed in the two affected siblings, whereas the two parents and unaffected sister were heterozygous carriers of c.1273C > T (father and sister) and c.4994 T > C (mother). The nonsense mutation p.R425X, contributes to a premature stop codon, may result in a truncated polypeptide, which strongly suggests its pathogenicity for ANSD. The missense mutation p.L1665P results in a single amino acid substitution in a highly conserved region. CONCLUSIONS: Two mutations in the OTOF gene in the Chinese deaf population were recognized for the first time. These findings not only extend the OTOF gene mutation spectrum for ANSD but also indicate that whole-exome sequencing is an effective approach to clarify the genetic characteristics in non-syndromic ANSD patients.

Further phenotype description, genotype characterization in patients with de novo interstitial deletion on 2p23.2-24.1.

Interstitial deletions of the distal part of chromosome 2p seem to be rarely identified or reported: to date, only nine distinct patients have been published. The last three patients were diagnosed with the use of more recent molecular karyotyping technology (SNP array). We report on the natural history of an 8-year-old boy with dysmorphic features, postnatal overgrowth, microcephaly, generalized hypotonia, and global developmental delay. The diagnosis was accomplished by SNP array investigation that led to the identification of a de novo 7.4 Mb deletion of 2p23.2-p24.1. The present patient also developed a nonsyndromic auditory neuropathy. Since the deletion encompassed the OTOF gene, this haploinsufficiency suggests second allele sequencing as a possible cause (DFNB9). We describe the phenotype of the patient and review reports in patients with del 2p23 subsequent to the advent of the genomic era. At the time of identification of "new" micro- deletion and -duplication syndromes, the present report adds to the description of phenotype in patients with del(2)p(23.2;24.1) and the 2p23.2 region in particular.

A novel mutation in the OTOF gene in a Chinese family with auditory neuropathy.

Gene therapy for monogenic auditory neuropathy (AN) has successfully improved hearing function in target gene-deficient mice. Accurate genetic diagnosis can not only clarify the etiology but also accurately locate the lesion site, providing a basis for gene therapy and guiding patient intervention and management strategies. In this study, we collected data from a family with a pair of sisters with prelingual deafness. According to their auditory tests, subject Ⅱ-1 was diagnosed with profound sensorineural hearing loss (SNHL), Ⅱ-2 was diagnosed with AN, Ⅰ-1 was diagnosed with high-frequency SNHL, and Ⅰ-2 had normal hearing. Using whole-exome sequencing (WES), one nonsense mutation, c.4030C>T (p.R1344X), and one missense mutation, c.5000C>A (p.A1667D), in the OTOF (NM_001287489.1) gene were identified in the two siblings. Their parents were heterozygous carriers of c.5000C>A (father) and c.4030C>T (mother). We hypothesized that c.5000C>A is a novel pathogenic mutation. Thus, subject Ⅱ-1 should also be diagnosed with AN caused by OTOF mutations. These findings not only expand the OTOF gene mutation spectrum for AN but also indicate that WES is an effective approach for accurately diagnosing AN.

AAV-Mediated Gene Therapy Restores Hearing in Patients with DFNB9 Deafness.

Mutations in OTOFERLIN (OTOF) lead to the autosomal recessive deafness 9 (DFNB9). The efficacy of adeno-associated virus (AAV)-mediated OTOF gene replacement therapy is extensively validated in Otof-deficient mice. However, the clinical safety and efficacy of AAV-OTOF is not reported. Here, AAV-OTOF is generated using good manufacturing practice and validated its efficacy and safety in mouse and non-human primates in order to determine the optimal injection dose, volume, and administration route for clinical trials. Subsequently, AAV-OTOF is delivered into one cochlea of a 5-year-old deaf patient and into the bilateral cochleae of an 8-year-old deaf patient with OTOF mutations. Obvious hearing improvement is detected by the auditory brainstem response (ABR) and the pure-tone audiometry (PTA) in these two patients. Hearing in the injected ear of the 5-year-old patient can be restored to the normal range at 1 month after AAV-OTOF injection, while the 8-year-old patient can hear the conversational sounds. Most importantly, the 5-year-old patient can hear and recognize speech only through the AAV-OTOF-injected ear. This study is the first to demonstrate the safety and efficacy of AAV-OTOF in patients, expands and optimizes current OTOF-related gene therapy and provides valuable information for further application of gene therapies for deafness.