Highly variable hearing loss due to POU4F3 (c.37del) is revealed by longitudinal, frequency specific analyses.

Genotype-phenotype correlations add value to the management of families with hereditary hearing loss (HL), where age-related typical audiograms (ARTAs) are generated from cross-sectional regression equations and used to predict the audiogram phenotype across the lifespan. A seven-generation kindred with autosomal dominant sensorineural HL (ADSNHL) was recruited and a novel pathogenic variant in POU4F3 (c.37del) was identified by combining linkage analysis with whole exome sequencing (WES). POU4F3 is noted for large intrafamilial variation including the age of onset of HL, audiogram configuration and presence of vestibular impairment. Sequential audiograms and longitudinal analyses reveal highly variable audiogram features among POU4F3 (c.37del) carriers, limiting the utility of ARTAs for clinical prognosis and management of HL. Furthermore, a comparison of ARTAs against three previously published families (1 Israeli Jewish, 2 Dutch) reveals significant interfamilial differences, with earlier onset and slower deterioration. This is the first published report of a North American family with ADSNHL due to POU4F3, the first report of the pathogenic c.37del variant, and the first study to conduct longitudinal analysis, extending the phenotypic spectrum of DFNA15.

Autosomal dominant non-syndromic hearing loss maps to DFNA33 (13q34) and co-segregates with splice and frameshift variants in ATP11A, a phospholipid flippase gene.

Sequencing exomes/genomes have been successful for identifying recessive genes; however, discovery of dominant genes including deafness genes (DFNA) remains challenging. We report a new DFNA gene, ATP11A, in a Newfoundland family with a variable form of bilateral sensorineural hearing loss (SNHL). Genome-wide SNP genotyping linked SNHL to DFNA33 (LOD = 4.77), a locus on 13q34 previously mapped in a German family with variable SNHL. Whole-genome sequencing identified 51 unremarkable positional variants on 13q34. Continuous clinical ascertainment identified several key recombination events and reduced the disease interval to 769 kb, excluding all but one variant. ATP11A (NC_000013.11: chr13:113534963G>A) is a novel variant predicted to be a cryptic donor splice site. RNA studies verified in silico predictions, revealing the retention of 153 bp of intron in the 3' UTR of several ATP11A isoforms. Two unresolved families from Israel were subsequently identified with a similar, variable form of SNHL and a novel duplication (NM_032189.3:c.3322_3327+2dupGTCCAGGT) in exon 28 of ATP11A extended exon 28 by 8 bp, leading to a frameshift and premature stop codon (p.Asn1110Valfs43Ter). ATP11A is a type of P4-ATPase that transports (flip) phospholipids from the outer to inner leaflet of cell membranes to maintain asymmetry. Haploinsufficiency of ATP11A, the phospholipid flippase that specially transports phosphatidylserine (PS) and phosphatidylethanolamine (PE), could leave cells with PS/PE at the extracellular side vulnerable to phagocytic degradation. Given that surface PS can be pharmaceutically targeted, hearing loss due to ATP11A could potentially be treated. It is also likely that ATP11A is the gene underlying DFNA33.

A pathogenic deletion in Forkhead Box L1 (FOXL1) identifies the first otosclerosis (OTSC) gene.

Otosclerosis is a bone disorder of the otic capsule and common form of late-onset hearing impairment. Considered a complex disease, little is known about its pathogenesis. Over the past 20 years, ten autosomal dominant loci (OTSC1-10) have been mapped but no genes identified. Herein, we map a new OTSC locus to a 9.96 Mb region within the FOX gene cluster on 16q24.1 and identify a 15 bp coding deletion in Forkhead Box L1 co-segregating with otosclerosis in a Caucasian family. Pre-operative phenotype ranges from moderate to severe hearing loss to profound sensorineural loss requiring a cochlear implant. Mutant FOXL1 is both transcribed and translated and correctly locates to the cell nucleus. However, the deletion of 5 residues in the C-terminus of mutant FOXL1 causes a complete loss of transcriptional activity due to loss of secondary (alpha helix) structure. FOXL1 (rs764026385) was identified in a second unrelated case on a shared background. We conclude that FOXL1 (rs764026385) is pathogenic and causes autosomal dominant otosclerosis and propose a key inhibitory role for wildtype Foxl1 in bone remodelling in the otic capsule. New insights into the molecular pathology of otosclerosis from this study provide molecular targets for non-invasive therapeutic interventions.

A dominant RAD51C pathogenic splicing variant predisposes to breast and ovarian cancer in the Newfoundland population due to founder effect.

BACKGROUND: RAD51C is important in DNA repair and individuals with pathogenic RAD51C variants have increased risk of hereditary breast and ovarian cancer syndrome (HBOC), an autosomal dominant genetic predisposition to early onset breast and/or ovarian cancer. METHODS: Five female HBOC probands sequenced negative for moderate- and high-risk genes but shared a recurrent variant of uncertain significance in RAD51C (NM_058216.3: c.571 + 4A > G). Participant recruitment was followed by haplotype and case/control analyses, RNA splicing analysis, gene and protein expression assays, and Sanger sequencing of tumors. RESULTS: The RAD51C c.571 + 4A > G variant segregates with HBOC, with heterozygotes sharing a 5.07 Mbp haplotype. RAD51C c.571 + 4A > G is increased ~52-fold in the Newfoundland population compared with the general Caucasian population and positive population controls share disease-associated alleles, providing evidence of a founder effect. Splicing analysis confirmed in silico predictions that RAD51C c.571 + 4A > G causes exon 3 skipping, creating an immediate premature termination codon. Gene and protein expression were significantly reduced in a RAD51C c.571 + 4G > A heterozygote compared with a wild-type relative. Sanger sequencing of tumors from two probands indicates loss-of-heterozygosity, suggesting loss of function. CONCLUSION: The RAD51C c.571 + 4A > G variant affects mRNA splicing and should be re-classified as pathogenic according to American College of Medical Genetics and Genomics guidelines.

A common variant in CLDN14 causes precipitous, prelingual sensorineural hearing loss in multiple families due to founder effect.

Genetic isolates provide unprecedented opportunities to identify pathogenic mutations and explore the full natural history of clinically heterogeneous phenotypes such as hearing loss. We noticed a unique audioprofile, characterized by prelingual and rapid deterioration of hearing thresholds at frequencies >0.5 kHz in several adults from unrelated families from the island population of Newfoundland. Targeted serial Sanger sequencing of probands for deafness alleles (n = 23) that we previously identified in this founder population was negative. Whole exome sequencing in four members of the largest family (R2010) identified a CLDN14 (DFNB29) variant [c.488C>T; p. (Ala163Val)], likely pathogenic, sensorineural hearing loss, autosomal recessive. Although not associated with deafness or disease, CLDN14 p.(Ala163Val) has been previously reported as a variant of uncertain significance (VUS). Targeted sequencing of 169 deafness probands identified one homozygote and one heterozygous carrier. Genealogical studies, cascade sequencing and haplotype analysis across four unrelated families showed all subjects with the unique audioprofile (n = 12) were also homozygous for p.(Ala163Val) and shared a 1.4 Mb DFNB29-associated haplotype on chromosome 21. Most significantly, sequencing 175 population controls revealed 1% of the population are heterozygous for CLDN14 p.(Ala163Val), consistent with a major founder effect in Newfoundland. The youngest CLDN14 [c.488C>T; p.(Ala163Val)] homozygote passed newborn screening and had normal hearing thresholds up to 3 years of age, which then deteriorated to a precipitous loss >1 kHz during the first decade. Our study suggests that genetic testing may be necessary to identify at-risk children in time to prevent speech, language and developmental delay.

Comparison of epicardial potential maps derived from the 12-lead electrocardiograms with scintigraphic images during controlled myocardial ischemia.

Our aim was to cross-validate electrocardiographic (ECG) and scintigraphic imaging of acute myocardial ischemia. The former method was based on inverse calculation of heart-surface potentials from the body-surface ECGs, and the latter, on a single photon emission computed tomography (SPECT). A boundary-element torso model with 352 body-surface and 202 heart-surface nodes was used to perform the ECG inverse solution. Potentials at 352 body-surface nodes were calculated from those acquired at 12-lead ECG measurement sites using regression coefficients developed from a design set (n = 892) of body-surface potential mapping (BSPM) data. The test set (n = 18) consisted of BSPM data from patients who underwent a balloon-inflation angioplasty of either the left anterior descending coronary artery (LAD) (n = 7), left circumflex coronary artery (LCx) (n = 2), or the right coronary artery (RCA) (n = 9). Body-surface potential mapping distributions at J point for 352 nodes were estimated from the 12-lead ECG, and an agreement with those estimated from 120 leads was assessed by a correlation coefficient (CC) (in percent). These estimates yielded very similar BSPM distributions, with a CC of 91.0% ± 8.1% (mean ± SD) for the entire test set and 94.1% ± 1.4%, 96.7% ± 0.8%, and 87.4% ± 10.3% for LAD, LCx, and RCA subgroups, respectively. Corresponding heart-surface potential distributions obtained by inverse solution correlated with a lower CC of 69.3% ± 18.0% overall and 73.7% ± 10.8%, 84.7% ± 1.1%, and 62.6% ± 21.8%, respectively, for subgroups. Bull's-eye displays of heart-surface potentials calculated from estimated BSPM distributions had an area of positive potentials that qualitatively corresponded, in general, with the underperfused territory suggested by SPECT images. For the LAD and LCx groups, all 9 ECG-derived bull's-eye images indicated the expected territory; for the RCA group, 6 of 9 ECG-derived images were as expected; 2 of 3 misclassified cases had very small ECG changes in response to coronary-artery occlusion, and their SPECT images showed indiscernible patterns. In conclusion, our findings demonstrate that noninvasive ECG imaging based on just the 12-lead ECG might provide useful estimates of the regions of myocardial ischemia that agree with those provided by scintigraphic techniques.

Quantitative assessment of myocardial ischemia by electrocardiographic and scintigraphic imaging.

We calculated distributions of epicardial potentials from body-surface electrocardiograms (ECGs) recorded during controlled myocardial ischemia and compared them with scintigraphic estimates of ischemia's extent/severity. The study population consisted of patients suffering from single-vessel coronary artery disease, referred for elective percutaneous transluminal coronary angioplasty of either the left anterior descending (n=7), the right coronary (n=9), or the left circumflex (n=2) artery. After the target vessel had been dilated, a 1960s "study" inflation was performed with a non-perfusion-type balloon catheter; at its commencement, technetium-99m sestamibi was injected via a femoral-vein catheter, and ECGs were recorded throughout the inflation from 120 leads. Single photon emission computed tomographic imaging was performed one hour after the injection of radionuclide to obtain an "occlusion image", and again one hour after a repeat injection 24 hours later to obtain a "control image"; the latter image was subtracted from the former, to derive a scintigraphic difference map (Delta map). The ECGs were signal-averaged over a 10-s window at preinflation and peak-inflation states, the preinflation averaged complexes were subtracted from the peak-inflation ones to produce body-surface Delta maps, and the corresponding Delta maps of epicardial potentials were calculated by applying the electrocardiographic inverse solution; this procedure is referred to as electrocardiographic imaging. The ECG-derived epicardial Delta maps related spatially to the scintigraphic Delta maps in all patients. The percent areas and surface integrals of positive values in ECG-derived Delta maps were found to be very good single-variable predictors of the extent (r=0.73; p=0.0006) and severity (r=0.72; p=0.0008) of the scintigraphically-estimated perfusion defect; a regression equation using two ECG-derived predictors further improved the agreement with scintigraphic estimates (r=0.81; p=0.0004 for estimates of severity). These findings suggest that noninvasive electrocardiographic imaging might provide quantitative estimates of the extent/severity of myocardial ischemia that agree closely with those provided by scintigraphic techniques.