Joana Chora

DFNB1-associated deafness in Portuguese cochlear implant users: prevalence and impact on oral outcome.

OBJECTIVES Hearing loss is a condition that interferes with the development of the child at a cognitive and language level. Therefore, early diagnosis of deafness is important for (re)habilitation, namely through the use of cochlear implant (CI). The present study aimed at screening CI Portuguese individuals for the presence of mutations in the genes GJB2 and GJB6 (DFNB1 locus), and searching a possible correlation between the genotype and the oral habilitation outcome following implantation. METHODS Our sample included 117 CI individuals implanted longer than 5 years. Sequencing of GJB2 entire coding region was first performed. The presence of deletions del(GJB6-D13S1830) and del(GJB6-D13S1854) was subsequently tested by multiplex PCR. To assess the oral outcome of these individuals, a global score is calculated through a formula that integrates the results of a battery of speech and audiological tests routinely used in ORL services. This global oral performance score was used to test whether individuals with DFNB1-associated deafness perform significantly better than individuals without DFNB1-associated deafness. RESULTS In 35% of the cases, deafness was clearly associated to DFNB1. The most common mutated allele was c.35delG (85%). Other variants have also been found, namely p.Gly130Ala, p.Asn206Ser, p.Val37Ile, p.Glu47X, p.Arg184Trp, p.Trp24X and the two common GJB6 deletions, del(GJB6-D13S1854) and del(GJB6-D13S1830), the last one identified for the first time in our population. Regarding the oral outcome, after testing the homogeneity of the two groups it could be observed that, in mean, the individuals with DFNB1-associated deafness perform significantly better (p=0.012) than the individuals without DFNB1-associated deafness. DISCUSSION AND CONCLUSION This first screening of DFNB1 genes in the Portuguese CI population provides clear evidence of the high proportion of DFNB1-associated deafness amongst the Portuguese implanted individuals. DFNB1 status is significantly associated to higher oral performance scores, with DFNB1 individuals performing, on average, 6% better than the individuals without DFNB1-associated deafness.

Spectrum and frequency of GJB2 mutations in a cohort of 264 Portuguese nonsyndromic sensorineural hearing loss patients

Abstract Objective: To assess the spectrum and prevalence of mutations in the GJB2 gene in Portuguese nonsyndromic sensorineural hearing loss (NSSHL) patients. Design: Sequencing of the coding region, basal promoter, exon 1, and donor splice site of the GJB2 gene; screening for the presence of the two common GJB6 deletions. Study sample: A cohort of 264 Portuguese NSSHL patients. Results: At least one out of 21 different GJB2 variants was identified in 80 (30.2%) of the 264 patients analysed. Two mutant alleles were found in 53 (20%) of these probands, of which 83% (44/53) harboured at least one c.35delG allele. Twenty-seven (10.2%) of the probands harboured only one mutant allele. Subsequent analysis revealed that the GJB6 deletion del(GJB6-D13S1854) was present in at least 7.4% (2/27) of the patients carrying only one mutant GJB2 allele. Overall, one in five (55/264) of the patients were diagnosed as having DFNB1-related NSSHL, of which the vast majority (53/55) harboured only GJB2 mutations. Conclusions: This study provides clear demonstration that mutations in the GJB2 gene are an important cause of NSSHL in Portugal, thus representing a valuable indicator as regards therapeutical and rehabilitation options, as well as genetic counseling of these patients and their families.

Analysis of publicly available LDLR , APOB , and PCSK9 variants associated with familial hypercholesterolemia: application of ACMG guidelines and implications for familial hypercholesterolemia diagnosis

PurposeFamilial hypercholesterolemia (FH) is an autosomal disorder of lipid metabolism presenting with increased cardiovascular risk. Although more than 1,700 variants have been associated with FH, the great majority have not been functionally proved to affect the low-density lipoprotein receptor cycle. We aimed to classify all described variants associated with FH and to establish the proportion of variants that lack evidence to support their pathogenicity.MethodsWe followed American College of Medical Genetics and Genomics (ACMG) guidelines for the classification, and collected information from a variety of databases and individual reports. A worldwide overview of publicly available FH variants was also performed.ResultsA total of 2,104 unique variants were identified as being associated with FH, but only 166 variants have been proven by complete in vitro functional studies to be causative of disease. Additionally, applying the ACMG guidelines, 1,097 variants were considered pathogenic or likely pathogenic. Only seven variants were found in all five continents.ConclusionThe lack of functional evidence for about 85% of all variants found in FH patients can compromise FH diagnosis and patient prognosis. ACMG classification improves variant interpretation, but functional studies are necessary to understand the effect of about 40% of all variants reported. Nevertheless, ACMG guidelines need to be adapted to FH for a better diagnosis.

A novel p.Leu213X mutation in GJB2 gene in a Portuguese family

INTRODUCTION Hearing loss is the most common sensory disability and is present in about 1.9 per 1000 infants at birth. The DFNB1 locus (13q11-q12) includes the genes GJB2, coding for connexin 26, and GJB6, encoding connexin 30. More than 100 mutations have been identified associated with autosomal dominant and recessive hearing loss in the GJB2 gene. OBJECTIVES The aim of the present study was to identify the genetic aetiology of deafness in two Portuguese individuals, presenting nonsyndromic sensorineural moderate and severe hearing loss, respectively. PATIENTS AND METHODS The individuals were evaluated in both ears by pure tone audiometry and blood samples were collected after written informed consent was signed. DNA extraction and PCR amplification of GJB2 coding region followed standard methodologies. PCR products were automatically sequenced in both directions. RESULTS We identified a novel mutation, c.638T>A (p.Leu213X), in GJB2 gene. This nonsense mutation was found in both siblings, and was inherited from their hearing father. Molecular analysis showed that the two siblings were also heterozygous for c.333-334delAA, a previously described GJB2 deletion. This novel mutation was not found in a random control sample of 480 individuals that were screened for coding region of GJB2 gene. p.Leu213X mutation identified in this study for the first time changes the codon 213, coding for a highly conserved and slowly evolving residue of connexin 26, localised to the C-terminus domain of the protein, to a STOP codon, leading to the deletion of the last 14 amino acids of the protein. CONCLUSION We can conclude that the aetiology of deafness in these individuals is due to the GJB2 genotype involving the c.333-334delAA deletion and the novel p.Leu213X mutation in compound heterozygosity.

ClinVar database of global familial hypercholesterolemia-associated DNA variants

Accurate and consistent variant classification is imperative for incorporation of rapidly developing sequencing technologies into genomic medicine for improved patient care. An essential requirement for achieving standardized and reliable variant interpretation is data sharing, facilitated by a centralized open‐source database. Familial hypercholesterolemia (FH) is an exemplar of the utility of such a resource: it has a high incidence, a favorable prognosis with early intervention and treatment, and cascade screening can be offered to families if a causative variant is identified. ClinVar, an NCBI‐funded resource, has become the primary repository for clinically relevant variants in Mendelian disease, including FH. Here, we present the concerted efforts made by the Clinical Genome Resource, through the FH Variant Curation Expert Panel and global FH community, to increase submission of FH‐associated variants into ClinVar. Variant‐level data was categorized by submitter, variant characteristics, classification method, and available supporting data. To further reform interpretation of FH‐associated variants, areas for improvement in variant submissions were identified; these include a need for more detailed submissions and submission of supporting variant‐level data, both retrospectively and prospectively. Collaborating to provide thorough, reliable evidence‐based variant interpretation will ultimately improve the care of FH patients.