Contribution of x-linked genes to the origin and development of intellectual disability in patients from the spanish Basque Country

167 p.La Discapacidad Intelectual (DI) es un desorden del neurodesarrollo que afecta aproximadamente al 1-3 % de la población y que supone un serio problema, tanto médico como social, en los países desarrollados. La DI ligada al cromosoma X (XLID) representa un grupo importante, por haber más varones que mujeres con DI y en muchos casos se segrega además vía materna. En los últimos años, las tecnologías de secuenciación masiva (NGS), han contribuido de forma importante al diagnóstico de la XLID, lo que nos hizo plantear la HIPÓTESIS de que en nuestros pacientes sin filiar podríamos aplicar la NGS encontrando nuevos genes/variantes causantes de DI y aplicando estos resultados al diagnóstico y al Consejo genético en las familias.- OBJETIVOS: Se marcaron 3 objetivos, cuyos resultados siguen el mismo orden:- RESULTADOS:1.- Se revisó la base de datos del laboratorio de los 1909 pacientes varones que llegaron para el estudio del gen FMR1 durante 1991-2015. Se seleccionaron 230 que tenían historia familiar (el 12%) y se cuantificaron y clasificaron los 59 diagnósticos moleculares ya obtenidos, la mayoría de los cualesestaban ligados al X (52/59), siendo el Síndrome X Frágil el más prevalente (43 casos) y demostrando ya la importancia de los genes del X en el origen de la DI (52/230=22,6%).2.- Se diseñó un panel de 82 genes del X para aplicarlo por NGS a los casos sin filiar. En total se estudiaron con el panel 61 pacientes varones con posible DI ligada al X. Se identificaron 17 variantes candidatas en 16 pacientes (16/61=26.23%) y se pudieron recontactar 6 familias con 7 variantes para estudios de cosegregación. Esto hizo posible la interpretación de algunas variantes como patogénicas, probablemente patogénicas o benignas.3.-Se validó el panel de genes del X comparándolo con el exoma completo en trio en 13 parejas de hermanos varones con DI. Aunque el panel de genes resultó en una tasa más baja de diagnóstico (1/13) que el exoma (4/13), el panel ofreció una mayor profundidad de cobertura de las regiones cubiertas y por lo tanto mejor detección de variantes. Finalmente, juntando los datos de estas 13 familias con las parejas de hermanos de la base de datos que ya tenían diagnóstico molecular (en total 45), se comprobó que la mayoría tenían variantes en el cromosoma X (15/45) por lo que la contribución del cromosoma X es del 33.33% en nuestra cohorte de parejas de hermanos con DI.- CONCLUSIONES: Este estudio demuestra la importante contribución de los genes del X en pacientes varones con DI e historia familiar, del orden del 22 al 33% en nuestra cohorte. Además, este trabajo ha demostrado la utilidad del panel de genes del X en el desciframiento de la DI ligada al X; ha puesto de manifiesto las limitaciones de los análisis de segregación y la necesidad de los estudios funcionales; y ha demostrado la necesidad de estudiar retrospectivamente los pacientes para aplicarles las nuevas tecnologías con el fin de lograr más y mejores diagnósticos genéticos moleculare

Contribution of x-linked genes to the origin and development of intellectual disability in patients from the spanish Basque Country

167 p.La Discapacidad Intelectual (DI) es un desorden del neurodesarrollo que afecta aproximadamente al 1-3 % de la población y que supone un serio problema, tanto médico como social, en los países desarrollados. La DI ligada al cromosoma X (XLID) representa un grupo importante, por haber más varones que mujeres con DI y en muchos casos se segrega además vía materna. En los últimos años, las tecnologías de secuenciación masiva (NGS), han contribuido de forma importante al diagnóstico de la XLID, lo que nos hizo plantear la HIPÓTESIS de que en nuestros pacientes sin filiar podríamos aplicar la NGS encontrando nuevos genes/variantes causantes de DI y aplicando estos resultados al diagnóstico y al Consejo genético en las familias.- OBJETIVOS: Se marcaron 3 objetivos, cuyos resultados siguen el mismo orden:- RESULTADOS:1.- Se revisó la base de datos del laboratorio de los 1909 pacientes varones que llegaron para el estudio del gen FMR1 durante 1991-2015. Se seleccionaron 230 que tenían historia familiar (el 12%) y se cuantificaron y clasificaron los 59 diagnósticos moleculares ya obtenidos, la mayoría de los cualesestaban ligados al X (52/59), siendo el Síndrome X Frágil el más prevalente (43 casos) y demostrando ya la importancia de los genes del X en el origen de la DI (52/230=22,6%).2.- Se diseñó un panel de 82 genes del X para aplicarlo por NGS a los casos sin filiar. En total se estudiaron con el panel 61 pacientes varones con posible DI ligada al X. Se identificaron 17 variantes candidatas en 16 pacientes (16/61=26.23%) y se pudieron recontactar 6 familias con 7 variantes para estudios de cosegregación. Esto hizo posible la interpretación de algunas variantes como patogénicas, probablemente patogénicas o benignas.3.-Se validó el panel de genes del X comparándolo con el exoma completo en trio en 13 parejas de hermanos varones con DI. Aunque el panel de genes resultó en una tasa más baja de diagnóstico (1/13) que el exoma (4/13), el panel ofreció una mayor profundidad de cobertura de las regiones cubiertas y por lo tanto mejor detección de variantes. Finalmente, juntando los datos de estas 13 familias con las parejas de hermanos de la base de datos que ya tenían diagnóstico molecular (en total 45), se comprobó que la mayoría tenían variantes en el cromosoma X (15/45) por lo que la contribución del cromosoma X es del 33.33% en nuestra cohorte de parejas de hermanos con DI.- CONCLUSIONES: Este estudio demuestra la importante contribución de los genes del X en pacientes varones con DI e historia familiar, del orden del 22 al 33% en nuestra cohorte. Además, este trabajo ha demostrado la utilidad del panel de genes del X en el desciframiento de la DI ligada al X; ha puesto de manifiesto las limitaciones de los análisis de segregación y la necesidad de los estudios funcionales; y ha demostrado la necesidad de estudiar retrospectivamente los pacientes para aplicarles las nuevas tecnologías con el fin de lograr más y mejores diagnósticos genéticos moleculare

Missense MED12 variants in 22 males with intellectual disability: From nonspecific symptoms to complete syndromes

Genotype; Intellectual disability; PhenotypeGenotipo; Discapacidad intelectual; FenotipoGenotip; Discapacitat intel·lectual; FenotipWe describe the phenotype of 22 male patients (20 probands) carrying a hemizygous missense variant in MED12. The phenotypic spectrum is very broad ranging from nonspecific intellectual disability (ID) to the three well-known syndromes: Opitz–Kaveggia syndrome, Lujan–Fryns syndrome, or Ohdo syndrome. The identified variants were randomly distributed throughout the gene (p = 0.993, χ2 test), but mostly outside the functional domains (p = 0.004; χ2 test). Statistical analyses did not show a correlation between the MED12-related phenotypes and the locations of the variants (p = 0.295; Pearson correlation), nor the protein domain involved (p = 0.422; Pearson correlation). In conclusion, establishing a genotype–phenotype correlation in MED12-related diseases remains challenging. Therefore, we think that patients with a causative MED12 variant are currently underdiagnosed due to the broad patients' clinical presentations.Foundation for Science and Technology (FCT), Grant/Award Number: UIDB/00215/2020 UIDP/00215/2020 LA/P/0064/2020; Broad Institute; National Eye Institute; National Heart, Lung and Blood Institute, Grant/Award Number: UM1 HG008900; National Human Genome Research Institute, Grant/Award Number: R01 HG009141

Functional analyses of a novel splice variant in the CHD7 gene, found by next generation sequencing, Confirm Its pathogenicity in a Spanish patient and diagnose him with CHARGE syndrome

Mutations in CHD7 have been shown to be a major cause of CHARGE syndrome, which presents many symptoms and features common to other syndromes making its diagnosis difficult. Next generation sequencing (NGS) of a panel of intellectual disability related genes was performed in an adult patient without molecular diagnosis. A splice donor variant in CHD7 (c.5665 + 1G > T) was identified. To study its potential pathogenicity, exons and flanking intronic sequences were amplified from patient DNA and cloned into the pSAD® splicing vector. HeLa cells were transfected with this construct and a wild-type minigene and functional analysis were performed. The construct with the c.5665 + 1G > T variant produced an aberrant transcript with an insert of 63 nucleotides of intron 28 creating a premature termination codon (TAG) 25 nucleotides downstream. This would lead to the insertion of 8 new amino acids and therefore a truncated 1896 amino acid protein. As a result of this, the patient was diagnosed with CHARGE syndrome. Functional analyses underline their usefulness for studying the pathogenicity of variants found by NGS and therefore its application to accurately diagnose patients.This work was funded by Jesús de Gangoiti Barrera Foundation (FJGB15/005). The EAV laboratory is funded by projects of the Spanish Ministry of Economy and Competitiveness, National Plan for R & D 2013–2016, ISCIII (FIS: PI13/01749) co-financed by FEDER from Regional Development European Funds (European Union) and the project CSI090U14 of the Regional ministry of Education (ORDER EDU/122/2014) (Castilla y León, Spain). This study made use of data generated by the UK10K Project. Funding for the UK10K Project was provided by the Wellcome Trust under award WT091310.Peer reviewe

Targeted Next-Generation Sequencing in Patients with Suggestive X-Linked Intellectual Disability

X-linked intellectual disability (XLID) is known to contribute up to 10% of intellectual disability (ID) in males and could explain the increased ratio of affected males observed in patients with ID. Over the past decade, next-generation sequencing has clearly stimulated the gene discovery process and has become part of the diagnostic procedure. We have performed targeted next-generation sequencing of 82 XLID genes on 61 non-related male patients with suggestive non-syndromic XLID. These patients were initially referred to the molecular genetics laboratory to exclude Fragile X Syndrome. The cohort includes 47 male patients with suggestive X-linked family history of ID meaning that they had half-brothers or maternal cousins or uncles affected; and 14 male patients with ID and affected brothers whose mothers show skewed X-inactivation. Sequencing data analysis identified 17 candidate variants in 16 patients. Seven families could be re-contacted and variant segregation analysis of the respective eight candidate variants was performed: HUWE1, IQSEC2, MAOA, MED12, PHF8, SLC6A8, SLC9A6, and SYN1. Our results show the utility of targeted next-generation sequencing in unravelling the genetic origin of XLID, especially in retrospective cases. Variant segregation and additional studies like RNA sequencing and biochemical assays also helped in re-evaluating and further classifying the genetic variants found

Екатеринбургская неделя. 1890. № 13

Background: Androgen steroid hormones are key drivers of prostate cancer. Previous work has shown that androgens can drive the expression of alternative mRNA isoforms as well as transcriptional changes in prostate cancer cells. Yet to what extent androgens control alternative mRNA isoforms and how these are expressed and differentially regulated in prostate tumours is unknown. Methods: Here we have used RNA-Seq data to globally identify alternative mRNA isoform expression under androgen control in prostate cancer cells, and profiled the expression of these mRNA isoforms in clinical tissue. Results: Our data indicate androgens primarily switch mRNA isoforms through alternative promoter selection. We detected 73 androgen regulated alternative transcription events, including utilisation of 56 androgen-dependent alternative promoters, 13 androgen-regulated alternative splicing events, and selection of 4 androgen-regulated alternative 3′ mRNA ends. 64 of these events are novel to this study, and 26 involve previously unannotated isoforms. We validated androgen dependent regulation of 17 alternative isoforms by quantitative PCR in an independent sample set. Some of the identified mRNA isoforms are in genes already implicated in prostate cancer (including LIG4, FDFT1 and RELAXIN), or in genes important in other cancers (e.g. NUP93 and MAT2A). Importantly, analysis of transcriptome data from 497 tumour samples in the TGCA prostate adenocarcinoma (PRAD) cohort identified 13 mRNA isoforms (including TPD52, TACC2 and NDUFV3) that are differentially regulated in localised prostate cancer relative to normal tissue, and 3 (OSBPL1A, CLK3 and TSC22D3) which change significantly with Gleason grade and  tumour stage. Conclusions: Our findings dramatically increase the number of known androgen regulated isoforms in prostate cancer, and indicate a highly complex response to androgens in prostate cancer cells that could be clinically important

Functional Analyses of a Novel Splice Variant in the CHD7 Gene, Found by Next Generation Sequencing, Confirm Its Pathogenicity in a Spanish Patient and Diagnose Him with CHARGE Syndrome

Mutations in CHD7 have been shown to be a major cause of CHARGE syndrome, which presents many symptoms and features common to other syndromes making its diagnosis difficult. Next generation sequencing (NGS) of a panel of intellectual disability related genes was performed in an adult patient without molecular diagnosis. A splice donor variant in CHD7 (c.5665 + 1G > T) was identified. To study its potential pathogenicity, exons and flanking intronic sequences were amplified from patient DNA and cloned into the pSAD® splicing vector. HeLa cells were transfected with this construct and a wild-type minigene and functional analysis were performed. The construct with the c.5665 + 1G > T variant produced an aberrant transcript with an insert of 63 nucleotides of intron 28 creating a premature termination codon (TAG) 25 nucleotides downstream. This would lead to the insertion of 8 new amino acids and therefore a truncated 1896 amino acid protein. As a result of this, the patient was diagnosed with CHARGE syndrome. Functional analyses underline their usefulness for studying the pathogenicity of variants found by NGS and therefore its application to accurately diagnose patients

Dietary inflammatory index of mothers during pregnancy and Attention Deficit-Hyperactivity Disorder symptoms in the child at preschool age: a prospective investigation in the INMA and RHEA cohorts

Inflammation provides a substrate for mechanisms that underlie the association of maternal diet during pregnancy with Attention Deficit-Hyperactivity Disorder (ADHD) symptoms in childhood. However, no previous study has quantified the proinflammatory potential of maternal diet as a risk factor for ADHD. Thus, we evaluated the association of maternal dietary inflammatory index (DII®) scores during pregnancy with ADHD symptoms in 4-year-old children born in two Mediterranean regions. We analyzed data from two population-based birth cohort studies-INMA (Environment and Childhood) four subcohorts in Spain (N = 2097), and RHEA study in Crete (Greece) (N = 444). The DII score of maternal diet was calculated based on validated food frequency questionnaires completed during pregnancy (12th and/or 32nd week of gestation). ADHD symptoms were assessed by ADHD-DSM-IV in INMA cohort and by ADHDT test in RHEA cohort, with questionnaires filled-out by teachers and parents, respectively. The associations between maternal DII and ADHD symptoms were analysed using multivariable-adjusted zero-inflated negative binomial regression models in each cohort study separately. Meta-analysis was conducted to combine data across the cohorts for fitting within one model. The DII was significantly higher in RHEA (RHEA = 2.09 [1.94, 2.24]) in comparison to INMA subcohorts (Asturias = - 1.52 [- 1.67, - 1.38]; Gipuzkoa = - 1.48 [- 1.64, - 1.33]; Sabadell = - 0.95 [- 1.07, - 0.83]; Valencia = - 0.76 [- 0.90, - 0.62]). Statistically significant reduced risk of inattention symptomatology (OR = 0.86; CI 95% = 0.77-0.96), hyperactivity symptomatology (OR = 0.82; CI 95% = 0.72-0.92) and total ADHD symptomatology (OR = 0.82; CI 95% = - 0.72 to 0.93) were observed with increased maternal DII in boys. No statistically significant associations were observed in girls between maternal DII and inattention, hyperactivity and total ADHD symptomatology. We found reduced risk of ADHD symptomatology with increased DII only in boys. This relationship requires further exploration in other settings