Usher Syndrome Research Articles

Usher syndrome: clinical features, molecular genetics and advancing therapeutics.

Usher syndrome has three subtypes, each being clinically and genetically heterogeneous characterised by sensorineural hearing loss and retinitis pigmentosa (RP), with or without vestibular dysfunction. It is the most common cause of deaf–blindness worldwide with a prevalence of between 4 and 17 in 100 000. To date, 10 causative genes have been identified for Usher syndrome, with MYO7A accounting for >50% of type 1 and USH2A contributing to approximately 80% of type 2 Usher syndrome. Variants in these genes can also cause non-syndromic RP and deafness. Genotype–phenotype correlations have been described for several of the Usher genes. Hearing loss is managed with hearing aids and cochlear implants, which has made a significant improvement in quality of life for patients. While there is currently no available approved treatment for the RP, various therapeutic strategies are in development or in clinical trials for Usher syndrome, including gene replacement, gene editing, antisense oligonucleotides and small molecule drugs.

Structural modeling, mutation analysis, and in vitro expression of usherin, a major protein in inherited retinal degeneration and hearing loss

Usherin is the most common causative protein associated with autosomal recessive retinitis pigmentosa (RP) and Usher syndrome (USH), which are characterized by retinal degeneration alone and in combination with hearing loss, respectively. Usherin is essential for photoreceptor survival and hair cell bundle integrity. However, the molecular mechanism underlying usherin function in normal and disease conditions is unclear. In this study, we investigated structural models of usherin domains and localization of usherin pathogenic small in-frame mutations, mainly homozygous missense mutations. We found that usherin fibronectin III (FN3) domains and most laminin-related domains have a β-sandwich structure. Some FN3 domains are predicted to interact with each other and with laminin-related domains. The usherin protein may bend at some FN3 linker regions. RP- and USH-associated small in-frame mutations are differentially located in usherin domains. Most of them are located at the periphery of β-sandwiches, with some at the interface between interacting domains. The usherin laminin epidermal growth factor repeats adopt a rod-shaped structure, which is maintained by disulfide bonds. Most missense mutations and deletion of exon 13 in this region disrupt the disulfide bonds and may affect local protein folding. Despite low expression of the recombinant entire protein and protein fragments in mammalian cell culture, usherin FN3 fragments are more robustly expressed and secreted than its laminin-related fragments. Our findings provide new insights into the usherin structure and the disease mechanisms caused by pathogenic small in-frame mutations, which will help inform future experimental research on diagnosis, disease mechanisms, and therapeutic approaches.

Retinitis pigmentosa with bilateral irido-fundal coloboma

A 40-year-old male, presented with gradual painless progressive loss of vision in both the eyes more in right since 1 year. Born out of consanguineous marriage, he gave a history of night blindness since childhood. The best-corrected visual acuity was 20/200 and 20/40 in right and left eye, respectively. Refraction showed an axial myopia of -9.00 DS/-1.50 DC at 100 degree in the right eye. The left eye showed refraction of +1.00 DC at 170 degree. Intraocular pressure with noncontact tonometer was 15 and 16 mm of Hg. Axial length was found to be 24.93 mm and 23.60 mm in right and left eye, respectively. Vertical and horizontal corneal diameters were 11 mm in both eyes. Keratometry values in the right eye were K vertical 43.55D, K horizontal 44.06D and in the left eye were K vertical 43.05D, K horizontal 43.49D. Slit-lamp examination revealed a clear cornea and normal anterior chamber depth. Inferonasal iris coloboma was noted in both eyes [Fig. 1]. The lens showed nuclear sclerosis grade 2 with posterior subcapsular opacification in right and nuclear sclerosis grade 2 in the left eye according to the Lens Opacities Classification System II (LOCS II) The posterior segment showed a normal-looking disc with minimal arteriolar attenuation, boney spicules distributed from the mid periphery throughout the retina in both eyes. Chorio-retinal coloboma was type 3 (separated from the optic disc by a clear area of the retina) in the right eye [Fig. 2] and type 7 (extreme peripheral coloboma) in the left eye [Fig. 3] according to Ida Manns classification. B scan ultrasonography showed an excavated lesion in the inferior aspect suggestive of fundus coloboma in the right eye. In the left eye the coloboma was not well demarcated on B scan ultrasonography.Figure 1: Anterior segment photographs with diffuse illumination (a) Right eye showing inferonasal iris coloboma with cataractous lens. (b) Left eye showing inferonasal iris colobomaFigure 2: (a) Fundus photograph of right eye showing coloboma in the inferior quadrant sparing disc and macula with bony spicules in the mid periphery. (b) Fundus photograph right eye, superonasal quadrant showing bony spicules in the mid peripheryFigure 3: (a) Fundus photograph of left eye depicting bony spicules in mid periphery and extreme peripheral coloboma inferiorly. (b) Peripheral coloboma with bony spicules in inferior quadrantPhacoemulsification with intraocular lens implantation under topical anesthesia was done in the right eye. Postoperatively at the end of 1 month best-corrected visual acuity was 20/30 in the right eye with a refraction of -0.75 DC at 80 degrees and near vision of N6 with + 3.00 DS. Discussion Retinitis pigmentosa is most commonly associated with posterior subcapsular cataract 53%[1] glaucoma 2.3%[2] myopia, keratoconus, vitreous detachment, and optic disc drusen. Association of macular colobomas with retinitis pigmentosa, especially autosomal dominant RP and Ushers syndrome has been extensively reported in the literature. In 2004, phenotypically different RP with macular coloboma was reported by Parmeggiani in 3 adult siblings[3], which was stated as an autosomal dominant feature. The association of iris and fundal coloboma with RP has been rarely reported in the literature. Kelly et al.[4] described the incidence, ocular findings, and systemic associations of coloboma in a population-based cohort study in the United States, reported one case with unilateral coloboma in association with RP. Population-based studies in India are scanty. Pallavi Agrawal et al.[5] reported a similar case of unilateral coloboma from a case in Mumbai, India in 2013. Bilateral Irido-fundal coloboma and RP can be separate entities occurring together in this case, or it could be a rare association. Similar cases/case series need to be reported to come to a conclusive result. Declaration of patient consent The authors certify that they have obtained all appropriate patient consent forms. In the form the patient(s) has/have given his/her/their consent for his/her/their images and other clinical information to be reported in the journal. The patients understand that their names and initials will not be published and due efforts will be made to conceal their identity, but anonymity cannot be guaranteed. Financial support and sponsorship Nil. Conflicts of interest There are no conflicts of interest.

Next Generation Sequencing Identifies Five Novel Mutations in Lebanese Patients with Bardet–Biedl and Usher Syndromes

Aim: To identify disease-causing mutations in four Lebanese families: three families with Bardet–Biedl and one family with Usher syndrome (BBS and USH respectively), using next generation sequencing (NGS). Methods: We applied targeted NGS in two families and whole exome sequencing (WES) in two other families. Pathogenicity of candidate mutations was evaluated according to frequency, conservation, in silico prediction tools, segregation with disease, and compatibility with inheritance pattern. The presence of pathogenic variants was confirmed via Sanger sequencing followed by segregation analysis. Results: Most likely disease-causing mutations were identified in all included patients. In BBS patients, we found (M1): c.2258A > T, p. (Glu753Val) in BBS9, (M2): c.68T > C; p. (Leu23Pro) in ARL6, (M3): c.265_266delTT; p. (Leu89Valfs*11) and (M4): c.880T > G; p. (Tyr294Asp) in BBS12. A previously known variant (M5): c.551A > G; p. (Asp184Ser) was also detected in BBS5. In the USH patient, we found (M6): c.188A > C, p. (Tyr63Ser) in CLRN1. M2, M3, M4, and M6 were novel. All of the candidate mutations were shown to be likely disease-causing through our bioinformatic analysis. They also segregated with the corresponding phenotype in available family members. Conclusion: This study expanded the mutational spectrum and showed the genetic diversity of BBS and USH. It also spotlighted the efficiency of NGS techniques in revealing mutations underlying clinically and genetically heterogeneous disorders.

Ataluren for the Treatment of Usher Syndrome 2A Caused by Nonsense Mutations.

The identification of genetic defects that underlie inherited retinal diseases (IRDs) paves the way for the development of therapeutic strategies. Nonsense mutations caused approximately 12% of all IRD cases, resulting in a premature termination codon (PTC). Therefore, an approach that targets nonsense mutations could be a promising pharmacogenetic strategy for the treatment of IRDs. Small molecules (translational read-through inducing drugs; TRIDs) have the potential to mediate the read-through of nonsense mutations by inducing expression of the full-length protein. We provide novel data on the read-through efficacy of Ataluren on a nonsense mutation in the Usher syndrome gene USH2A that causes deaf-blindness in humans. We demonstrate Ataluren´s efficacy in both transiently USH2AG3142*-transfected HEK293T cells and patient-derived fibroblasts by restoring USH2A protein expression. Furthermore, we observed enhanced ciliogenesis in patient-derived fibroblasts after treatment with TRIDs, thereby restoring a phenotype that is similar to that found in healthy donors. In light of recent findings, we validated Ataluren´s efficacy to induce read-through on a nonsense mutation in USH2A-related IRD. In line with published data, our findings support the use of patient-derived fibroblasts as a platform for the validation of preclinical therapies. The excellent biocompatibility combined with sustained read-through efficacy makes Ataluren an ideal TRID for treating nonsense mutations based IRDs.

Clarin-1 expression in adult mouse and human retina highlights a role of Müller glia in Usher syndrome.

Usher syndrome type 3 (USH3) is an autosomal recessively inherited disorder caused by mutations in the gene clarin‐1 (CLRN1), leading to combined progressive hearing loss and retinal degeneration. The cellular distribution of CLRN1 in the retina remains uncertain, either because its expression levels are low or because its epitopes are masked. Indeed, in the adult mouse retina, Clrn1 mRNA is developmentally downregulated, detectable only by RT‐PCR. In this study we used the highly sensitive RNAscope in situ hybridization assay and single‐cell RNA‐sequencing techniques to investigate the distribution of Clrn1 and CLRN1 in mouse and human retina, respectively. We found that Clrn1 transcripts in mouse tissue are localized to the inner retina during postnatal development and in adult stages. The pattern of Clrn1 mRNA cellular expression is similar in both mouse and human adult retina, with CLRN1 transcripts being localized in Müller glia, and not photoreceptors. We generated a novel knock‐in mouse with a hemagglutinin (HA) epitope‐tagged CLRN1 and showed that CLRN1 is expressed continuously at the protein level in the retina. Following enzymatic deglycosylation and immunoblotting analysis, we detected a single CLRN1‐specific protein band in homogenates of mouse and human retina, consistent in size with the main CLRN1 isoform. Taken together, our results implicate Müller glia in USH3 pathology, placing this cell type to the center of future mechanistic and therapeutic studies to prevent vision loss in this disease. © 2019 The Authors. The Journal of Pathology published by John Wiley & Sons Ltd on behalf of Pathological Society of Great Britain and Ireland.

Building, Supporting the Usher Syndrome Community

Building, Supporting the Usher Syndrome Community

Current and future gene‐based therapies for Usher syndromes

AbstractInherited retinal dystrophies are a group of genetically and clinically heterogeneous disorders characterized by photoreceptor degeneration. One common example is retinitis pigmentosa (RP), which is characterized by impaired night vision and progressively decreasing peripheral vision. Moreover, RP can be present as either an isolated visual defect or in association with extra ocular symptoms. The most common syndromic form is Usher syndrome, which is characterized by RP associated with neurosensory deafness of varying severity. The difficulty of this double handicap demands the development of novel therapies. The advantage of the Usher syndrome group is that each disease is monogenic thus lending themselves to gene‐based therapies. The caveat is that the most common forms are due to mutations in large genes, thus rendering gene replacement challenging. This talk will present an overview of current gene‐based therapies for Usher syndrome as a whole and will concentrate on novel genome‐editing advances for the most prevalent causative gene, USH2A.

Introduction and clinical aspects in Usher syndromes

AbstractUsher syndrome (USH, OMIM 276900, OMIM 276905, OMIM 605472, ORPHA: 886) is the most prevalent deaf‐blindness of genetic origin. It is a recessive inherited disease characterized by sensorineural hearing loss (HL), visual loss due to retinitis pigmentosa (RP), and, in some cases, vestibular dysfunction. Prevalence estimates range from 3.2 to 6.2/100,000. Patients with USH are classified into three clinical subtypes (USH1, USH2, or USH3), based on the severity and progression of hearing impairment and the presence or absence of vestibular dysfunction. Usher syndrome type I (USH1) is the most severe type, characterized by severe to profound congenital sensorineural hearing loss, vestibular dysfunction and prepubertal onset of RP eventually leading to legal blindness. USH2 is characterized by moderate to severe hearing impairment, normal vestibular function and later onset of retinal degeneration. USH3 displays progressive hearing loss, RP and variable vestibular phenotype. In the last years, next generation sequencing (NGS) techniques have revolutionized the world of the molecular genetic diagnosis. NGS has allowed to molecularly characterize a high number of patients, thus permitting to establish some phenotype‐genotype correlations that had been elusive to date.

Expanding the clinical and genetic spectrum of Heimler syndrome

BackgroundHeimler syndrome (HS) is a rare hereditary systemic disorder, partial clinically overlapping with Usher syndrome. So far, our knowledge of HS is very limited, many cases are misdiagnosed or may not even be diagnosed at all. This study aimed to analyze the clinical and genetic characteristics of HS, and to evaluate potential phenotype–genotype correlations.ResultsTwo HS cases caused by PEX1 mutations were identified, and a novel likely pathogenic mutation, PEX1 c.895_896insTATA, was found. The main ophthalmic finding of the two patients was consistent with retinitis pigmentosa accompanied by cystoid macular edema, but short axial length and hyperopia were also observed as two previously unreported ocular phenotypes. Analysis of the literature showed that of the 29 HS patients previously reported, 12 had PEX6 mutations, 10 had PEX1 mutations, two had PEX26 mutations, and the remaining patients were not genetically tested. Three novel genotype–phenotype correlations were revealed from analysis of these patients. First, most genotypes of every HS patient include at least one missense variant; second, at least one mutation in PEX1 or PEX6 gene affects the AAA-ATPase region in every HS patient with retinal dystrophy, suggesting AAA-ATPase region is a hypermutable region in patients with a retinal dystrophy; third, there are no significant differences between PEX1-, PEX6-, and PEX26-associated phenotypes.ConclusionNext-generation sequencing is important for the diagnosis of HS. This study expands the clinical and genetic spectrum of HS, and provides additional insights into genotype–phenotype correlations, which is vital for accurate clinical practice, genetic counseling, and pathogenesis studies.

Genetic landscape of Usher syndromes

AbstractUsher syndrome (USH) refers to recessively inherited disorders that associate hearing loss (HL) and retinitis pigmentosa (RP). Three clinical subtypes are defined with respect to the degree of HL, the age of RP onset and the occurrence of vestibular areflexia. Ten genes, that when mutated lead the development of the disease, have been identified: six (MYO7A, USH1C, CDH23, PCDH15, USH1G and CIB2) are involved in Usher type 1, the most disabling form, three (USH2A, ADGRV1 (GPR98) and WHRN) in Usher type 2, the most frequent form, and one, CLRN1, in Usher type 3. For many years, molecular diagnosis of Usher syndrome was laborious as many of these genes consist of numerous exons (e.g. 49 for MYO7A and 72 for USH2A, the most frequently involved genes). In the last years, implementation of MPS (Massively parallel sequencing) in diagnosis laboratories permits simultaneous sequencing of all the USH genes, thus making molecular studies of Usher patients much easier and faster. Yet, additional tools to search for hidden mutations or to classify the numerous variants of unknown clinical significance are needed in order to offer a proper comprehensive diagnosis. When gene panels include additional genes mimicking USH syndrome, it becomes possible to address subtle differential diagnoses. However, since several USH genes are involved in non‐syndromic HL as well, they can now be screened in patients presenting with apparent non‐syndromic form. Then, the challenge is to provide the most accurate genetic counselling as whether a child carrying two USH1 pathogenic variants will develop Usher syndrome or not. Finally, elucidating the molecular causes of Usher syndrome remains the essential step to build patient cohorts and to develop therapeutic approaches.

Antisense oligonucleotide‐induced splice modulation for the treatment of Usher syndrome type 2A

AbstractMutations in USH2A exon13 are the most frequent cause of both syndromic and non‐syndromic retinitis pigmentosa (RP), for which currently no treatment options exist. It is generally believed that RP due to mutations in this gene is caused by a loss‐of‐function mechanism. In total, over 600 different mutations have been identified in USH2A of which at least three are known to be derived from a common ancestor and are therefore seen more commonly: c.2299delG (p.Glu767fs*21), c.2276G>T (p.Cys759Phe) and c.7595‐2144A>G (p.Lys2532Thrfs*56). The size of the coding sequence (15,606 bp) and the presence of multiple alternatively spliced USH2A transcripts with unknown significance, hamper the development of gene augmentation therapy. Another difficulty in the development of a therapy is the lack of a suitable animal model. The currently available Ush2a mouse model displays only mild retina degeneration with a very late age of onset. Zebrafish ush2a mutant models however, show an early onset retinal dysfunction and are as such provide a unique opportunity to evaluate future therapeutic strategies. Antisense oligonucleotide (AON)‐based splice modulation has been proven to hold great promise as a therapeutic strategy for a number of hereditary conditions, including Usher syndrome. Upon pre‐mRNA binding, AONs will prevent or stimulate binding of the spliceosome thereby modulating splicing events. AONs can be designed and applied for different genes and genetic disorders as the specificity depends on their nucleotide sequence. In this study we explored the therapeutic potential of AON‐induced splice modulation as a therapeutic approach for the future treatment of USH2A‐associated retinal degeneration.

The latest on viral gene therapy

AbstractIn this presentation, we will discuss the last developments of ocular gene therapy aiming to restore visual functions and protect the retina from different forms of inherited retinal dystrophies (IRD). The issue of disease models will also be addressed, especially for Usher syndrome, for which the vector should target the sensory cells of the cochlea and the retina. In addition to gene therapy for recessive and dominant forms of IRDs and the new perspectives to treat them with CRISPR/Cas9 technology, the development of optogenetic tools will be described. Different strategies to restore function of large mutated genes using minigenes or triple vectors to expand the capacity of AAV vectors will also be exposed. The state‐of‐the‐art of the new developing technologies should help to choose the best available tool to attempt to treat the disease of interest.

Lack of PDZD7 long isoform disrupts ankle-link complex and causes hearing loss in mice.

Usher syndrome (USH) is the most frequent form of combined hereditary deafness-blindness, characterized by hearing loss and retinitis pigmentosa, with or without vestibular dysfunction. PDZD7 is a PDZ domain-containing scaffold protein that was suggested to be a USH modifier and a contributor to digenic USH. In the inner ear hair cells, PDZD7 localizes at the ankle region of the stereocilia and constitutes the so-called ankle-link complex together with three other USH proteins Usherin, WHRN, and ADGRV1. PDZD7 gene is subjected to alternative splicing, which gives rise to two types of PDZD7 isoforms, namely the long and short isoforms. At present, little is known which specific isoform is involved in ankle-link formation and stereocilia development. In this work, we showed that PDZD7 long isoform, but not short isoforms, localizes at the ankle region of the stereocilia. Moreover, we established Pdzd7 mutant mice by introducing deletions into exon 14 of the Pdzd7 gene, which causes potential premature translational stop in the long isoform but leaves short isoforms unaffected. We found that lack of PDZD7 long isoform affects the localization of other ankle-link complex components in the stereocilia. Consequently, Pdzd7 mutant mice showed stereocilia development deficits and hearing loss as well as reduced mechanotransduction (MET) currents, suggesting that PDZD7 long isoform is indispensable for hair cells. Furthermore, by performing yeast two-hybrid screening, we identified a PDZD7 long isoform-specific binding partner PIP5K1C, which has been shown to play important roles in hearing and might participate in the function and/or transportation of PDZD7.

Genome Editing in Patient iPSCs Corrects the Most Prevalent USH2A Mutations and Reveals Intriguing Mutant mRNA Expression Profiles

Inherited retinal dystrophies (IRDs) are characterized by progressive photoreceptor degeneration and vision loss. Usher syndrome (USH) is a syndromic IRD characterized by retinitis pigmentosa (RP) and hearing loss. USH is clinically and genetically heterogeneous, and the most prevalent causative gene is USH2A. USH2A mutations also account for a large number of isolated autosomal recessive RP (arRP) cases. This high prevalence is due to two recurrent USH2A mutations, c.2276G>T and c.2299delG. Due to the large size of the USH2A cDNA, gene augmentation therapy is inaccessible. However, CRISPR/Cas9-mediated genome editing is a viable alternative. We used enhanced specificity Cas9 of Streptococcus pyogenes (eSpCas9) to successfully achieve seamless correction of the two most prevalent USH2A mutations in induced pluripotent stem cells (iPSCs) of patients with USH or arRP. Our results highlight features that promote high target efficacy and specificity of eSpCas9. Consistently, we did not identify any off-target mutagenesis in the corrected iPSCs, which also retained pluripotency and genetic stability. Furthermore, analysis of USH2A expression unexpectedly identified aberrant mRNA levels associated with the c.2276G>T and c.2299delG mutations that were reverted following correction. Taken together, our efficient CRISPR/Cas9-mediated strategy for USH2A mutation correction brings hope for a potential treatment for USH and arRP patients.

A Novel Mouse Model of MYO7A USH1B Reveals Auditory and Visual System Haploinsufficiencies.

Usher’s syndrome is the most common combined blindness–deafness disorder with USH1B, caused by mutations in MYO7A, resulting in the most severe phenotype. The existence of numerous, naturally occurring shaker1 mice harboring variable MYO7A mutations on different genetic backgrounds has complicated the characterization of MYO7A knockout (KO) and heterozygote mice. We generated a novel MYO7A KO mouse (Myo7a–/–) that is easily genotyped, maintained, and confirmed to be null for MYO7A in both the eye and inner ear. Like USH1B patients, Myo7a–/– mice are profoundly deaf, and display near complete loss of inner and outer cochlear hair cells (HCs). No gross structural changes were observed in vestibular HCs. Myo7a–/– mice exhibited modest declines in retinal function but, unlike patients, no loss of retinal structure. We attribute the latter to differential expression of MYO7A in mouse vs. primate retina. Interestingly, heterozygous Myo7a+/– mice had reduced numbers of cochlear HCs and concomitant reductions in auditory function relative to Myo7a+/+ controls. Notably, this is the first report that loss of a single Myo7a allele significantly alters auditory structure and function and suggests that audiological characterization of USH1B carriers is warranted. Maintenance of vestibular HCs in Myo7a–/– mice suggests that gene replacement could be used to correct the vestibular dysfunction in USH1B patients. While Myo7a–/– mice do not exhibit sufficiently robust retinal phenotypes to be used as a therapeutic outcome measure, they can be used to assess expression of vectored MYO7A on a null background and generate valuable pre-clinical data toward the treatment of USH1B.

Expanding the Genetic Landscape of Usher-Like Phenotypes.

Usher syndrome (USH) is a rare disorder characterized by retinitis pigmentosa (RP) and sensorineural hearing loss. Several genes are responsible for the disease, but not all cases are explained by mutations in any of these, supporting the fact that there remain other unknown genes that have a role in the syndrome. We aimed to find the genetic cause of presumed USH patients lacking pathogenic mutations in the known USH genes. Whole exome sequencing was performed on a priori USH-diagnosed subjects from nine unrelated families, which had shown negative results for an USH-targeted panel in a previous study. We identified possible pathogenic variants in six of the studied families. One patient harbored mutations in REEP6 and TECTA, each gene tentatively causative of one of the two main symptoms of the disease, mimicking the syndrome. In three patients, only the retinal degeneration causative mutations were detected (involving EYS, WDR19, and CNGB1 genes). Another family manifested a dementia-linked retinal dystrophy dependent on an allele dosage in the GRN gene. Last, another case presented a homozygous mutation in ASIC5, a gene not yet associated with USH. Our findings demonstrate that pending cases should be clinically and genetically carefully assessed, since more patients than expected may be either related phenocopies or affected by a more complex disease encompassing additional symptoms rather than classical USH.

Cochlear Implantation in Children with Usher's Syndrome: A South Asian Experience.

Usher's syndrome is an autosomal recessive disorder characterized by dual sensory impairment involving both the ears and eyes. Cochlear implantation paves a way to restore hearing loss in such individuals but poor vision among these patients poses additional challenges for the habilitationists. This study aimed to compare the habilitation outcomes and hearing-related quality of life scores of cochlear implantees having Usher syndrome with age-matched cochlear implantees with no such syndromic association. 27 patients aged 1-6years with Usher syndrome underwent cochlear Implantation over a period of 10years from 2006 to 2016 and were included in this study along with an age-matched cohort of 30 implantees with no additional disabilities. Category of Auditory Performance (CAP) and Speech Intelligibility Rating (SIR) scores were compared at 3, 6, 9 and 12months respectively. Glasgow Benefit Inventory and Health Utility Index (HUI 3.0) questionnaires were used to assess the hearing-related quality of life in both groups at 1 year post implantation. There was significant difference in CAP and SIR scores between children with Usher's syndrome and the control group (p < 0.05). The overall scores in terms of quality of life as well were statistically different (p < 0.05). Though there was improvement in speech and language acquisition after cochlear Implantation this was found to be of lesser extent than the normative cohort. These children with additional visual disabilities required intensive, individualized therapy catering to their complex needs. Their family's perception of expected benefit from cochlear Implantation was guarded and needed to be appraised in detail prior to surgery. This experience helped prepare an institutional protocol for counseling such implantees in future.

Preimplantation genetic testing for a family with usher syndrome through targeted sequencing and haplotype analysis

BackgroundPreimplantation genetic testing for monogenic defects (PGT-M) has been available in clinical practice. This study aimed to validate the applicability of targeted capture sequencing in developing personalized PGT-M assay.MethodsOne couple at risk of transmitting Usher Syndrome to their offspring was recruited to this study. Customized capture probe targeted at USH2A gene and 350 kb flanking region were designed for PGT-M. Eleven blastocysts were biopsied and amplified by using multiple displacement amplification (MDA) and capture sequencing. A hidden Markov model (HMM) assisted haplotype analysis was performed to deduce embryo’s genotype by using single nucleotide polymorphisms (SNPs) identified in each sample. The embryo without paternal rare variant was implanted and validated by conventional prenatal or postnatal diagnostic means.ResultsFour embryos were diagnosed as free of father’s rare variant, two were transferred and one achieved a successful pregnancy. The fetal genotype was confirmed by Sanger sequencing of fetal genomic DNA obtained by amniocentesis. The PGT-M and prenatal diagnosis results were further confirmed by the molecular diagnosis of the baby’s genomic DNA sample. The auditory test showed that the hearing was normal.ConclusionsTargeted capture sequencing is an effective and convenient strategy to develop customized PGT-M assay.

Severe or Profound Sensorineural Hearing Loss Caused by Novel USH2A Variants in Korea: Potential Genotype-Phenotype Correlation.

ObjectivesWe, herein, report two novel USH2A variants from two unrelated Korean families and their clinical phenotypes, with attention to severe or more than severe sensorineural hearing loss (SNHL).MethodsTwo postlingually deafened subjects (SB237-461, M/46 and SB354-692, F/34) with more than severe SNHL and also with suspicion of Usher syndrome type II (USH2) were enrolled. A comprehensive audiological and ophthalmological assessments were evaluated. We conducted the whole exome sequencing and subsequent pathogenicity prediction analysis.ResultsWe identified the following variants of USH2A from the two probands manifesting more than severe SNHL and retinitis pigmentosa (RP): compound heterozygosity for a nonsense (c.8176C>T: p.R2723X) and a missense variant (c.1823G>A: p.C608Y) in SB237, and compound heterozygosity for two frameshift variants (c.14835delT: p.S4945fs & c.13112_13115delAAAT: p.G4371fs) in SB354. Based on the American College of Medical Genetics and Genomics/Association for Molecular Pathology guidelines, two novel variants, c.1823G>A: p.C608Y and c.14835delT: p.Ser4945fs, can be classified as “uncertain significance” and “pathogenic,” respectively. The audiogram exhibited more than severe SNHL and a down-sloping configuration, necessitating cochlear implantation. The ophthalmic examinations revealed typical features of RP. Interestingly, one proband (SB 354-692) carrying two truncating compound heterozygous variants exhibited more severe hearing loss than the other proband (SB 237-461), carrying one truncation with one missense variant.ConclusionOur results provide insight on the expansion of audiological spectrum encompassing more than severe SNHL in Korean subjects harboring USH2A variants, suggesting that USH2A should also be included in the candidate gene of cochlear implantation. A specific combination of USH2A variants causing truncating proteins in both alleles could demonstrate more severe audiological phenotype than that of USH2A variants carrying one truncating mutation and one missense mutation, suggesting a possible genotype-phenotype correlation. The understanding of audiological complexity associated with USH2A will be helpful for genetic counseling and treatment starategy.