MYO7A Mutations Research Articles

Focus on the treatment of inherited retinal diseases

Inherited retinal diseases (IRDs) are rare and incurable eye diseases.Gene therapy has become a new method for the treatment of IRDs.At present, at least 26 clinical trials of gene therapy involving at least 16 different IRDs genes are in progress or imminent.Such as autosomal recessive retinitis pigmentosa (RP, MERTK mutation), choroideremia (CHM mutation), Stargardt’s disease (ABCA4 mutation), Usher syndrome 1B subtype (Myo7a mutation), X-linked retinoschisis (RS1 mutation), mitochondrial-related Leber hereditary optic neuropathy (ND4 mutation), panchromatic blindness (CNGA3 mutation and CNNGB3 mutation), sex-linked RP (RPGR mutation). Adeno-associated viral (AAV) vector-mediated gene expression is a conventional method for gene transduction, which has showed efficacies in autosomal recessive IRDs caused by small gene mutations, especially for those IRDs, which have the original lesions in retinal pigment epithelial (RPE) cells and photoreceptor cells.Novel progress has been made in animal experiments by gene editing technologies for autosomal dominant IRDs or autosomal recessive IRDs with large gene mutations, which is going to clinical trial.Oligonucleotide therapy based on RNA has also shown good efficacy for the IRDs caused by large genes, which beyond the AAV capacity.These research achievements are attacking more and more attentions in these fields.Although there are still some efficacy and safety issues on the way to treat IRDs, it is reasonable to expect that more and more patients with IRDs will be treated in the near future. Key words: Inherited retinal diseases; Gene therapy; Clinical trials; Optogenetic gene therapy; Antisense Oliogonucleotide therapy

Retinal findings in pediatric patients with Usher syndrome Type 1 due to mutations in MYO7A gene.

To describe retinal alterations detected by swept-source optical coherence tomography (SS-OCT) in paediatric patients with Usher syndrome type 1 (USH1) and to compare these findings to previously published reports. Thirty-two eyes from 16 patients (11 males and 5 females) with a genetic diagnosis of USH1 because of MYO7A mutations underwent SS-OCT. Patients ranged in age from 4 to 17 years (mean, 11,13 ± 4,29). The subfoveal and macular area were analysed with SS-OCT at 1050 nm using 12 radial scans of 12.0 mm. Structural abnormalities were evaluated and correlated with best-corrected visual acuity (BCVA). The most common qualitative retinal abnormality was external layer damage in macular area. Specific alterations included external limiting membrane loss/disruption (27 eyes; 84.4%), disruption of the Myoid zone (27 eyes; 84.4%); Ellipsoid zone disruption (28 eyes; 87.5%), and loss of the outer segments (29 eyes; 90.6%). The damage of the retinal pigment epithelium was divided according to the loss of the different layers: phagosome zone (30 eyes; 93.8%), melanosome zone (29 eyes; 90.6%) and mitochondria zone (0 eyes; 0%). The presence of cystoid macular oedema (CMO) was significantly correlated with alterations in photoreceptors. Disruption or absence of the myoid and ellipsoid zones of the photoreceptors were the only variables independently associated with decreased BCVA. The findings of this study suggest that the physiopathologic basis of early-stage Usher syndrome (USH) may be changes in the outer retinal layer, particularly the photoreceptors, which in turn may cause alterations-such as CMO-in the inner retinal layers. Accordingly, monitoring the condition of photoreceptors during follow-up may be advisable for the early detection of pathologic changes.

Characterisation of microvascular abnormalities using OCT angiography in patients with biallelic variants in USH2A and MYO7A

AimsUsing optical coherence tomography angiography (OCTA) to characterise microvascular changes in the retinal plexuses and choriocapillaris (CC) of patients with MYO7A and USH2A mutations and correlate with genotype, retinal structure...

Identification of a novel homozygous mutation in the MYO15A gene in a Kazakh family with non-syndromic hearing loss

BackgroundMillions of people around the world are plagued by hearing loss. More than 50% of congenital or pre-lingual deafness is associated with genetic factors and has highly genetic heterogeneity. To date, although hundreds of genes have been found to be implicated in non-syndromic deafness, there are still lots of genes or loci that we need to verify. MethodsIn this study, we performed target sequencing and Sanger sequencing in a Kazakh consanguineous family with autosomal recessive non-syndromic hearing loss. Following that, functional and structural studies predicted the pathogenic effect of novel mutations by use of the online tools. ResultsWe identified a novel homozygous mutation p.R3191C in MYO15A gene causing deafness in this family. The mutation p.R3191C co-segregated with the disease phenotype in this family and was not present in any public databases. Automatic tools predict that the novel mutation makes a great impact on the function and structure of MYO15A protein. ConclusionsThis is a novel mutation of MYO15A causing deafness and also the first report of MYO15A mutations causing deafness in the Kazakh families. This finding expanded the spectrum of MYO15A mutations, making it more precise for future genetic diagnosis in patients with deafness.

Detecting novel mutations and combined Klinefelter syndrome in Usher syndrome cases

Background: Usher syndrome (USH) is an autosomal recessive disease characterized by hearing loss, vision loss, and occasionally vestibular dysfunction. Klinefelter syndrome (KS) is an X chromosome polyploidy characterized by one or more additional X chromosomes in males. To date, there has been no report of USH combined with KS.Objectives: This study examined the causative genes in three Chinese probands with congenital hearing loss.Material and methods: Targeted next-generation sequencing (NGS) was performed to identify mutations in three probands with hearing loss. Low-coverage whole-genome sequencing (WGS) analysis of aneuploidy was used to verify the chromosome aneuploidy.Results: Four novel MYO7A mutations were identified in two USH1 probands who were initially diagnosed with nonsyndromic hearing loss until the onset of vision loss. Another case was initially diagnosed with nonsyndromic hearing loss and USH2 and KS were discovered incidentally after the genetic analysis.Conclusions: Our findings expand the mutation spectrum of MYO7A. This is also the first report of concomitant USH and KS. Genetic testing can help with clinical management, particularly if an unrecognized syndromic disorder is identified before the onset of additional symptoms. A clinical genetic evaluation is recommended as part of the diagnostic work-up in congenital hearing loss.

Identification of four novel mutations in MYO7A gene and their association with nonsyndromic deafness and Usher Syndrome 1B

IntroductionMYO7A gene has been shown to be associated with Usher syndrome 1B and nonsyndromic deafness. Although a lot of mutations have been reported in MYO7A gene, novel MYO7A mutations are continuously to be identified. MethodsTargeted next generation sequencing was performed on the two unrelated patients with Usher syndrome 1B and nonsyndromic deafness respectively. The identified mutations from targeted next generation sequencing were further validated by Sanger sequencing, and analyzed by bioinformatics tools, like SIFT, Polyphen-2, PyMOL, I-Mutant 2.0 and so on. ResultsBy analyzing the sequencing data of these two patients, four novel MYO7A mutations were revealed: (i) MYO7A p.Tyr560Ser and p.Ala2039Pro were associated with Usher syndrome 1B. (ii) MYO7A c.2187 + 2_+8 delTGAGCAC and p.Leu728Pro were related to nonsyndromic hearing loss. These mutations were further proved to be possibly disease-causing by segregation analysis, conservation analysis and bioinformatics tools. ConclusionsFour novel MYO7A mutations were identified in the present study. These findings provided new evidence for the genetic counseling of Usher syndrome 1B and nonsyndromic deafness.

Novel deleterious mutation in MYO7A, TH and EVC2 in two Pakistani brothers with familial deafness

Objective:In Pakistan, 74% of consanguineous marriages are among the first cousins. Continuity of consanguineous marriages over generations increases the risk of recessive diseases such as deafness. The objective of this study was to investigate genetic origin of Pakistani deaf brothers with parents of consanguineous marriage.Methods:DNA was extracted from the blood through Qiagen kit. Paired-end sequencing library was prepared according to protocol of Illumina’s TruSight Rapid Capture kit and TruSight Inherited Disease Panel. Library was normalized and used for Next Generation Sequencing through MiSeq. NGS data were analyzed using various bioinformatics tools.Results:Both brothers were found to have novel deleterious mutation in MYO7A (c.2476G>A) while the younger brother had additional novel deleterious mutation in TH (c.43C>T) and EVC2 (c.2614C>T) genes.Conclusion:It is concluded that in addition to novel mutations in MYO7A, TH and EVC2, the CDH23 and GJB2 can also be responsible for deafness in the family with consanguineous marriages.

Clonal Evolution of Relapsed CBFB/MYH11 Rearranged Acute Myeloid Leukemia (AML)

The leukemia-associated fusion gene CBFB/MYH11 results from a pericentric inversion of chromosome 16, inv(16)(p13.1q22), or less commonly from a t(16;16)(p13.1;q22). Although this cytogenetic aberration is associated with a rather favorable prognosis in acute myeloid leukemia (AML), nearly half of patients eventually relapse after standard chemotherapy.To systematically analyze the clonal evolution in this AML-subgroup, we performed whole exome sequencing (WES) of 13 adult CBFB/MYH11-rearranged AML patients using matched diagnostic, remission and relapse samples (‘triplets‘). Thereby, we found 2-12 (median: 8) somatic sequence variants per patient at diagnosis and 2-13 (median: 4) mutations at relapse. These included mutations in genes known to cooperate with CBFB/MYH11 (e.g. RAS, FLT3, KIT) as well as in genes, which had not been associated with AML previously (MYO15A, EVPL, ROS1, FTCD and ASL). Next, we designed a custom targeted sequencing assay (Haloplex, Agilent), including the candidate genes from exome sequencing, as well as genes known to be recurrently mutated in AML (455 genes, 1.86 Mbp total target sequence) and performed targeted sequencing of 32 CBFB/MYH11-rearranged AML triplet samples (including the 13 triplets initially analyzed by WES) with a median read depth of 500. The results are summarized in Figure 1. Fourteen genes were found mutated in at least two patients at diagnosis and 9 genes at the timepoint of relapse. In all CBFB/MYH11-rearranged patients, more than one additional mutation was identified, each of them at a distinct variant allele-frequency, indicating clonal heterogeneity. All but one FLT3 TKD (D835 or N676) mutation were lost at relapse, whereas FLT3 ITDs were stable in 3 out of 7 patients. One FLT3 ITD was gained at relapse. The majority of RAS,KIT and CBL mutations were lost and none was acquired at relapse. Particularly, the loss of 6 out of 7 KIT exon 8 frameshift mutations was surprising since KIT exon 8 frameshift mutations were negative prognostic markers in a cohort of 162 patients with CBFB/MYH11 rearranged leukemia (OS: HR= 3.12, p= 0.001; Opatz et al. submitted). In contrast, mutations in WT1 and DNMT3A were all stable during relapse evolution and four patients gained mutations in these two genes. Furthermore, aberrations in CSF3R, BCORL1 and ZBTB7A were acquired at relapse. Of note, WT1 mutations causing a frameshift in exon 6 were found in 9% of adult de novo AML with CBFB/MYH11-rearrangement and have recently been characterized by our research group as negative prognostic marker for overall survival (HR: 2.93, p= 0.011) (Opatz et al. submitted). These findings are in line with the observed gain of WT1 mutations in 10% of relapsed cytogenetically normal AML patients (Greif et al., 2018, Clin Cancer Res) suggesting a common mechanism of disease progress across cytogenetic subgroups. Surprisingly, a mutation in ZBTB7A, a gene frequently altered in RUNX1/RUNX1T1 positiv leukemia (23%) but rarely in CBFB/MYH11 positiv leukemia (2%), was gained at relapse in one patient. Mutations in epigenetic modifiers, cohesion complex components and janus kinases are known cooperating events in RUNX1/RUNX1T1 rearranged leukemia and were, except for DNMT3A, not found in our CBFB/MYH11-positive cohort. The new recurring mutations in MYO15A (n=3), ROS1 (n=2), FTCD (n=2) and ASL (n=2) were partially lost at relapse, whereas EVPL (n=2) mutations were stable during the course of disease. In addition, we identified mutations in APC2, TP53 and ZFHX4 (gained at relapse), PTPN11, MECOM, BCOR, NPM1 and IDH2 (stable) as well as in ABL1 (lost at relapse) in individual patients.Taken together, our findings suggest that mutations in signaling pathway genes seem to be unstable during disease progression and may thus not be required for the evolution of relapse. The frequent loss of signaling gene mutations indicates that relapse might evolve from an early ancestral clone carrying the CBFB/MYH11 rearrangement only.Figure 1: Mutation profile of 32 patients with CBFB/MYH11-rearranged AML. The stability of recurrently mutated genes during the evolution of relapse is shown. Diagonal lines indicate two mutations in the respective gene. [Display omitted] DisclosuresThiede:AgenDix: Other: Ownership; Novartis: Honoraria, Research Funding. Middeke:Janssen: Membership on an entity's Board of Directors or advisory committees, Research Funding; Roche: Membership on an entity's Board of Directors or advisory committees; Abbvie: Membership on an entity's Board of Directors or advisory committees. Stoelzel:Neovii: Speakers Bureau. Metzeler:Novartis: Consultancy; Celgene: Consultancy, Research Funding.

Novel mutations in MYTH4-FERM domains of myosin 15 are associated with autosomal recessive nonsyndromic hearing loss

ObjectiveHereditary hearing loss is the most common neurosensory disorder in humans caused by myriad mutations in numerous genes. Autosomal recessive nonsyndromic hearing loss (ARNSHL) accounts for 80% of hearing impairments of genetic origin and is quite prevalent in societies with a high rate of consanguinity. In the current study, we investigated the causes of sensorineural hearing loss in 24 unrelated Iranian families who were mainly consanguineous and had at least two affected children. MethodsAll probands were initially screened for GJB2 mutations, as the most common causes of ARNSHL in Iran. Verified GJB2-negative samples were subsequently subjected to whole exome sequencing (WES) to identify the underlying causes of hearing impairment, and the variants identified in each family were further confirmed by Sanger sequencing. ResultsWES revealed three previously unreported mutations in MYO15A, the gene encoding the unconventional myosin 15 (Myo15). All variants identified, c.C6436T (p.R2146W), c.C9584G (p.P3195R) and c.G10266C (p.Q3422H), reside in the MYTH4 (myosin tail homology) and FERM (4.1 ezrin, radixin, moesin) domains of the protein. ConclusionGlobally, mutations in MYO15A are considered to be among the most prevalent genetic causes of ARNSHL, and they rank as the third leading cause of hearing loss in the Iranian population, below GJB2 and SLC26A4. Yet again, these results endorse the importance of MYO15 screening in hearing impaired populations, particularly in Iran.

A novel nonsense mutation in MYO15A is associated with non-syndromic hearing loss: a case report

BackgroundHearing loss is genetically heterogeneous and is one of the most common human defects. Here we screened the underlying mutations that caused autosomal recessive non-syndromic hearing loss in a Chinese family.Case presentationThe proband with profound hearing loss had received audiometric assessments. We performed target region capture and next generation sequencing of 127 known deafness-related genes because the individual tested negative for hotspot variants in the GJB2, GJB3, SLC26A4, and MTRNR1 genes. We identified a novel c.6892C > T (p.R2298*) nonsense mutation and a c.10251_10253delCTT (p.F3420del) deletion in MYO15A. Sanger sequencing confirmed that both mutations were co-segregated with hearing loss in this family and were absent in 200 ethnically matched controls. Bioinformatics analysis and protein modeling indicated the deleterious effects of both mutations. The p.R2298* mutation leads to a truncated protein and a loss of the functional domains.ConclusionsOur results demonstrated that the hearing loss in this case was caused by novel, compound heterozygous mutations in MYO15A. The p.R2298* mutation in MYO15A was reported for the first time, which has implications for genetic counseling and provides insight into the functional roles of MYO15A mutations.

SD-OCT imaging as a valuable tool to support molecular genetic diagnostics of Usher syndrome type 1

A girl with profound congenital deafness and balance problems was found at 3.5years of age to be a carrier of two novel compound heterozygous mutations in MYO7A that were predicted to be disease-causing. She also carried one known pathogenic mutation and one rare variant in USH2A. Fundus examination performed at 4.75years revealed one small peripheral pigment deposit in the right eye, indicating probable retinal degeneration. Spectral domain optical coherence tomography (SD-OCT) showed a loss of photoreceptors throughout the macular area, except for the foveolar region, clearly confirming a diagnosis of Usher syndrome type 1. This case demonstrates that SD-OCT may be easily used in young children to confirm retinal disease, quantify the extent of retinal damage, and monitor disease progression.

The first case of NSHL by direct impression on EYA1 gene and identification of one novel mutation in MYO7A in the Iranian families.

Objective(s):Targeted next-generation sequencing (NGS) provides a consequential opportunity to elucidate genetic factors in known diseases, particularly in profoundly heterogeneous disorders such as non-syndromic hearing loss (NSHL). Hearing impairments could be classified into syndromic and non-syndromic types. This study intended to assess the significance of mutations in these genes to the autosomal recessive/dominant non-syndromic genetic load among Iranian families.Materials and Methods:Two families were involved in this research and two patients were examined by targeted next-generation sequencing. Here we report two novel mutations in the MYO7A and EYA1 genes in two patients detected by targeted NGS. They were confirmed by Sanger sequencing and quantitative real-time PCR techniques.Results:In this investigation, we identified a novel mutation in MYO7A, c.3751G>C, p.A1251P, along with another previously identified mutation (c.1708C>T) in one of the cases. This mutation is located in the MYTH4 protein domain which is a pivotal domain for the myosin function. Another finding in this research was a novel de-novo deletion which deletes the entire EYA1 coding region (EX1-18 DEL). Mutations in EYA1 gene have been found in branchiootorenal (BOR) syndrome. Interestingly the patient with EYA1 deletion did not show any other additional clinical implications apart from HL. This finding might argue for the sole involvement of EYA1 function in the mechanism of hearing.Conclusion:This investigation exhibited that the novel mutations in MYO7A, c.3751G>C, p.A1251P, and EYA1, EX1-18 DEL, were associated with NSHL. Our research increased the mutation spectrum of hearing loss in the Iranian population.

Utility of whole exome sequencing in the diagnosis of Usher syndrome: Report of novel compound heterozygous MYO7A mutations

Next generation sequencing (NGS), such as targeted panel sequencing, whole-exome sequencing and whole-genome sequencing has led to an exponential increase of elucidated genetic causes in both rare diseases, and common but heterogeneous disorders. NGS is applied in both research and clinical settings, and the clinical exome sequencing (CES), which provides not only the sequence variation data but also clinical interpretation, aids in reaching a final conclusion with regards to a genetic diagnosis.Usher syndrome is a group of disorders, characterized by bilateral sensorineural hearing loss, with or without vestibular dysfunction and retinitis pigmentosa. The index patient, a 2-year-old child was initially diagnosed with nonsyndromic hearing impairment. Homozygosity mapping followed by CES was utilized as a diagnostic tool to identify the genetic basis of his hearing loss. A paternally inherited novel insertion, c.198_199insA (p.Val67Serfs*73) and a maternally inherited novel deletion, c.1219_1226del (p.Phe407Aspfs*33) in gene MYO7A were found in compound heterozygous state in the index patient. The result expands the mutational spectrum of MYO7A. In addition it helped in early diagnosis of the syndrome, for planning and adjustments for the patient, and as well as for future family planning. This study highlights the clinical effectiveness of CES for Usher syndrome diagnosis in a child presented with congenital hearing loss.

Identification of a MYO7A mutation in a large Chinese DFNA11 family and genotype–phenotype review for DFNA11

Background: The molecular and genetic research showed the association between DFNA11 and mutations in MYO7A. This research aimed to identify a MYO7A mutation in a family with nonsyndromic autosomal dominant hearing loss.Methods: We have ascertained one large multigenerational Chinese family (Z029) with autosomal dominant late-onset progressive non-syndromic sensorineural hearing loss. Genome-wide linkage analysis of the family mapped the disease locus to the DFNA11 interval, where the MYO7A was considered as a candidate gene. Sequencing of the PCR products was carried out for each sample. One hundred and fifty one control subjects with normal hearing functions were also evaluated.Results: The pathogenic mutation (c.2011G>A) was identified in the family. This mutation co-segregated with hearing loss in this family. No mutation of MYO7A gene was found in the 151 controls.Conclusions: The missense mutation of MYO7A is identified in the family displaying the pedigree consistent with DFNA11. We not only examined the clinical and genetic characteristics of the family, but also provided a basis for genetic counseling. We also summarized and analyzed the phenotypes and genotypes of all DFNA11 families, four of nine are Chinese families, suggesting that MYO7A mutations are not rare. Therefore, we should pay more attention to Chinese patients.

Identification of Pathogenic Genes of Nonsyndromic Hearing Loss in Uyghur Families Using Massively Parallel DNA Sequencing Technique.

We aim to identify the mutations of deafness genes using massively parallel DNA sequencing in the 12 Uyghur families. SNPscan method was used to screen against the 124 sites in the common deafness genes in probands. Subjects with SNPscan negativity were subject to massively parallel DNA sequencing for the sequencing of 97 genes known to be responsible for hearing loss. Eight families (66.7%) showed biallelic mutations in probands, including MYO15A mutation (6892C>T in J02 family, 9514C>T/7894G>T in J07 family, and 9514C>T in J16 family), MYO7A mutation (1258A>T in J03 family), TMC1 mutation (773G>A in J09 family and 1247T>G/1312G>A in J11 family), and PCDH15 mutation (4658delT in J08 and J13 families). Six novel types of mutation were identified including 6892C>T, 9514C>T/7894G>T, and 9514C>T in MYO15A gene, 1258A>T in MYO7A, 773G>A in TMC1, and 4658delT in PCDH15. The ratio of nonsense mutation and frameshift mutation was comparatively high. All these indicated that the mutation types reported in this study were rare. In conclusion, rare deafness genes were identified in the Uyghur families using massively parallel DNA sequencing, part of which were suggested to be related to the pathogenesis of the disease.

Identification of a novel MYO7A mutation in Usher syndrome type 1.

Usher syndrome (USH) is an autosomal recessive disease characterized by deafness and retinitis pigmentosa. In view of the high phenotypic and genetic heterogeneity in USH, performing genetic screening with traditional methods is impractical. In the present study, we carried out targeted next-generation sequencing (NGS) to uncover the underlying gene in an USH family (2 USH patients and 15 unaffected relatives). One hundred and thirty-five genes associated with inherited retinal degeneration were selected for deep exome sequencing. Subsequently, variant analysis, Sanger validation and segregation tests were utilized to identify the disease-causing mutations in this family. All affected individuals had a classic USH type I (USH1) phenotype which included deafness, vestibular dysfunction and retinitis pigmentosa. Targeted NGS and Sanger sequencing validation suggested that USH1 patients carried an unreported splice site mutation, c.5168+1G>A, as a compound heterozygous mutation with c.6070C>T (p.R2024X) in the MYO7A gene. A functional study revealed decreased expression of the MYO7A gene in the individuals carrying heterozygous mutations. In conclusion, targeted next-generation sequencing provided a comprehensive and efficient diagnosis for USH1. This study revealed the genetic defects in the MYO7A gene and expanded the spectrum of clinical phenotypes associated with USH1 mutations.

Triple Vectors Expand AAV Transfer Capacity in the Retina.

Retinal gene transfer with adeno-associated viral (AAV) vectors holds great promise for the treatment of inherited retinal degenerations (IRDs). One limit of AAV is its transfer capacity of about 5 kb, which can be expanded to about 9 kb, using dual AAV vectors. This strategy would still not suffice for treatment of IRDs such as Usher syndrome type 1D or Alström syndrome type I (ALMS) due to mutations in CDH23 or ALMS1, respectively. To overcome this limitation, we generated triple AAV vectors, with a maximal transfer capacity of about 14 kb. Transcriptomic analysis following triple AAV transduction showed the expected full-length products along a number of aberrant transcripts. However, only the full-length transcripts are efficiently translated in vivo. We additionally showed that approximately 4% of mouse photoreceptors are transduced by triple AAV vectors and showed correct localization of recombinant ALMS1. The low-photoreceptor transduction levels might justify the modest and transient improvement we observe in the retina of a mouse model of ALMS. However, the levels of transduction mediated by triple AAV vectors in pig retina reached 40% of those observed with single vectors, and this bodes well for further improving the efficiency of triple AAV vectors in the retina.

STRC Deletion is a Frequent Cause of Slight to Moderate Congenital Hearing Impairment in the Czech Republic

This study aimed to clarify the molecular epidemiology of hearing loss by identifying the responsible genes in patients without GJB2 mutations. Prospective genetic study. Tertiary referral hospital. Fifty one patients with bilateral sensorineural hearing loss, 20 men, and 31 women, mean age 24.9 years, range 3 to 64 years, from 49 families. GJB2 and deltaGJB6-D13S1830 mutations were excluded previously. Diagnostic. Sixty-nine genes reported to be causative of hearing loss were analyzed. Sequence capture technology, next-generation sequencing, and multiplex ligation-dependent probe amplification (MLPA) were used. Coverage of STRC was screened in Integrative Genomics Viewer software. Identification of causal pathogenic mutations in genes related to deafness. Five families (10%) had recessive STRC deletions or mutations. Five unrelated patients (10%) had recessive mutations in TMPRSS3, USH2A, PCDH15, LOXHD1, and MYO15A. Three families (6%) had autosomal dominant mutations in MYO6A, KCNQ4, and SIX1. One family (2%) had an X-linked POU3F4 mutation. Thus, we identified the cause of hearing loss in 28% of the families studied. Following GJB2, STRC was the second most frequently mutated gene in patients from the Czech Republic with hearing loss. To decrease the cost of testing, we recommend STRC deletion screening with MLPA before next-generation sequencing. The existence of a pseudogene and polymorphic STRC regions can lead to false-positive or false-negative results when copy number variation analysis is based on next-generation sequencing data.

Targeted next generation sequencing in Italian patients with Usher syndrome: phenotype-genotype correlations

We report results of DNA analysis with next generation sequencing (NGS) of 21 consecutive Italian patients from 17 unrelated families with clinical diagnosis of Usher syndrome (4 USH1 and 17 USH2) searching for mutations in 11 genes: MYO7A, CDH23, PCDH15, USH1C, USH1G, USH2A, ADGVR1, DFNB31, CLRN1, PDZD7, HARS. Likely causative mutations were found in all patients: 25 pathogenic variants, 18 previously reported and 7 novel, were identified in three genes (USH2A, MYO7A, ADGRV1). All USH1 presented biallelic MYO7A mutations, one USH2 exhibited ADGRV1 mutations, whereas 16 USH2 displayed USH2A mutations. USH1 patients experienced hearing problems very early in life, followed by visual impairment at 1, 4 and 6 years. Visual symptoms were noticed at age 20 in a patient with homozygous novel MYO7A missense mutation c.849G > A. USH2 patients’ auditory symptoms, instead, arose between 11 months and 14 years, while visual impairment occurred later on. A homozygous c.5933_5940del;5950_5960dup in USH2A was detected in one patient with early deafness. One patient with homozygous deletion from exon 23 to 32 in USH2A suffered early visual symptoms. Therefore, the type of mutation in USH2A and MYO7A genes seems to affect the age at which both auditory and visual impairment occur in patients with USH.

CLINICAL PRESENTATION AND DISEASE COURSE OF USHER SYNDROME BECAUSE OF MUTATIONS IN MYO7A OR USH2A.

To evaluate differences in the visual phenotype and natural history of Usher syndrome caused by mutations in MYO7A or USH2A, the most commonly affected genes of Usher syndrome Type I (USH1) and Type II (USH2), respectively. Eighty-eight patients with a clinical diagnosis of USH1 (26 patients) or USH2 (62 patients) were retrospectively evaluated. Of these, 48 patients had 2 disease-causing mutations in MYO7A (10 USH1 patients), USH2A (33 USH2 patients), and other USH (5 patients) genes. Clinical investigation included best-corrected visual acuity, Goldmann visual field, fundus photography, electroretinography, and audiologic and vestibular assessments. Longitudinal analysis was performed over a median follow-up time of 3.5 years. Patients carrying mutations in MYO7A had a younger age of onset of hearing and visual impairments than those carrying mutations in USH2A, leading to an earlier diagnosis of the disease in the former patients. Longitudinal analysis showed that visual acuity and visual field decreased more rapidly in subjects carrying MYO7A mutations than in those carrying USH2A mutations (mean annual exponential rates of decline of 3.92 vs. 3.44% and of 8.52 vs. 4.97%, respectively), and the former patients reached legal blindness on average 15 years earlier than the latter. The current study confirmed a more severe progression of the retinal disease in USH1 patients rather than in USH2 patients. Furthermore, most visual symptoms (i.e., night blindness, visual acuity worsening) occurred at an earlier age in USH1 patients carrying mutations in MYO7A.