Análisis del genoma en pacientes con enfermedad de Ménière esporádica de inicio precoz

Abstract

Introduction: Meniere disease (MD) is a rare inner disease characterized by attacks of vertigo associated with low and medium frequency sensorineural hearing loss, and tinnitus or ear fullness. Although its etiology is unknown, its prevalence varies from 0.5 to 1 per 1000 individuals, affecting more familial cases (6-9%) than sporadic cases, which suggests a genetic contribution to MD. MD is a complex disease, with a genetic heterogeneity that is accompanied by a great diversity in the phenotype which complicates its diagnosis. Objective: To identify patients with early-onset recessive inheritance MD, as well as to identify new genes that could explain the heterogeneity of the disease. In addition, early-onset sporadic MD will be identified and clinically characterized. Whole genome sequencing will be performed in threesome in which the index will be a sporadic case of MD. Analysis of genomic data will be carried out to demonstrate accumulation of rare variants in genes (homozygous or compound heterozygous). Methods: This study will include a total of 6 threesome of Spanish patients with sporadic MD with early onset (≤35 years) according to the Barany Society’s Hearing and Balance Committee criteria. The DNA of the cases will be extracted from peripheral blood mononuclear cells (PBMCs) by extracting 12ml of venous blood. After the extraction, the DNA samples will be stored at -20ºC. This study will be carried out in accordance with the principles of the Helsinki Declarations of 1975 (revised in 2013) for human experimentation. Patients and controls will be informed about the study and their verbal and written consent will be required. Subsequently, the variant calling will be performed, using GATK for the realignment of the data with the GRCh27/hg19 human reference genome. Variant Call Format (VCF) files will be generated with the variants in each subject, including single nucleotide variants (SNV), insertions/deletions (INDELs), and copy number variations (CNV) and its corresponding annotation, in the population frequency reference databases (ExAC and gnomAD), and clinical characterization (ClinVar) including a pathogenicity prediction through CADD (Combined Annotation Dependent Depletion). With the results of each threesome, various type of analysis will be performed to cover the hypotheses about the appearance of early MD in sporadic cases (appearance of de novo variants, compound heterozygous inheritance, and homozygous inheritance). The gene rare variants aggregation analysis (MAF <0.05) and gene regulation networks will be carried out with the rvtest and GSEA tools. The novel variants of interest will be validated by Sanger sequencing. Results: For the first four threesome, an autosomal recessive inheritance was found, finding homozygous variants with and without change of sense. For the fifth and sixth threesome, dominant inheritance was identified, with one de novo heterozygous variant in the fifth threesome, and one de novo heterozygous copy number variant in the sixth one. Conclusions: Patients with early-onset sporadic MD have a higher prevalence of migraine than late-onset patients. Furthermore, cytokine analysis it was found revealed that migraine is not associated with any cytokine profile in MD patients. Cytokine levels in blood vary between patients with MD, migraine and controls. This could be used as a diagnostic tool. Early-onset sporadic MD could be explained by a homozygous recessive inheritance pattern. This inheritance model for missense SNV has been identified for SH3GL1 and LPCAT2 genes. Non-missense SNV has been identified for RANBP9 and ASH2L genes.