Allelic heterogeneity in a patient with postzygotic MTOR-related hypomelanosis of Ito with neurodevelopmental abnormalities.

A case of mosaic MTOR-associated hemimegalencephaly and hypomelanosis of Ito, died at 33 probably because of sudden unexpected death in epilepsy. Assessment of the variant allele fraction (VAF) in different tissues postmortem showed high variability not correlated with clinical features, representing the most detailed assessment of VAFs in different tissues to date.

Molecular consequences of PQBP1 deficiency, involved in the X-linked Renpenning syndrome.

Mutations in the PQBP1 gene (polyglutamine-binding protein-1) are responsible for a syndromic X-linked form of neurodevelopmental disorder (XL-NDD) with intellectual disability (ID), named Renpenning syndrome. PQBP1 encodes a protein involved in transcriptional and post-transcriptional regulation of gene expression. To investigate the consequences of PQBP1 loss, we used RNA interference to knock-down (KD) PQBP1 in human neural stem cells (hNSC). We observed a decrease of cell proliferation, as well as the deregulation of the expression of 58 genes, comprising genes encoding proteins associated with neurodegenerative diseases, playing a role in mRNA regulation or involved in innate immunity. We also observed an enrichment of genes involved in other forms of NDD (CELF2, APC2, etc). In particular, we identified an increase of a non-canonical isoform of another XL-NDD gene, UPF3B, an actor of nonsense mRNA mediated decay (NMD). This isoform encodes a shorter protein (UPF3B_S) deprived from the domains binding NMD effectors, however no notable change in NMD was observed after PQBP1-KD in fibroblasts containing a premature termination codon. We showed that short non-canonical and long canonical UPF3B isoforms have different interactomes, suggesting they could play distinct roles. The link between PQBP1 loss and increase of UPF3B_S expression was confirmed in mRNA obtained from patients with pathogenic variants in PQBP1, particularly pronounced for truncating variants and missense variants located in the C-terminal domain. We therefore used it as a molecular marker of Renpenning syndrome, to test the pathogenicity of variants of uncertain clinical significance identified in PQPB1 in individuals with NDD, using patient blood mRNA and HeLa cells expressing wild-type or mutant PQBP1 cDNA. We showed that these different approaches were efficient to prove a functional effect of variants in the C-terminal domain of the protein. In conclusion, our study provided information on the pathological mechanisms involved in Renpenning syndrome, but also allowed the identification of a biomarker of PQBP1 deficiency useful to test variant effect.

Multiple molecular diagnoses in the field of intellectual disability and congenital anomalies: 3.5% of all positive cases.

PURPOSE: Wide access to clinical exome/genome sequencing (ES/GS) enables the identification of multiple molecular diagnoses (MMDs), being a long-standing but underestimated concept, defined by two or more causal loci implicated in the phenotype of an individual with a rare disease. Only few series report MMDs rates (1.8% to 7.1%). This study highlights the increasing role of MMDs in a large cohort of individuals addressed for congenital anomalies/intellectual disability (CA/ID). METHODS: From 2014 to 2021, our diagnostic laboratory rendered 880/2658 positive ES diagnoses for CA/ID aetiology. Exhaustive search on MMDs from ES data was performed prospectively (January 2019 to December 2021) and retrospectively (March 2014 to December 2018). RESULTS: MMDs were identified in 31/880 individuals (3.5%), responsible for distinct (9/31) or overlapping (22/31) phenotypes, and potential MMDs in 39/880 additional individuals (4.4%). CONCLUSION: MMDs are frequent in CA/ID and remain a strong challenge. Reanalysis of positive ES data appears essential when phenotypes are partially explained by the initial diagnosis or atypically enriched overtime. Up-to-date clinical data, clinical expertise from the referring physician, strong interactions between clinicians and biologists, and increasing gene discoveries and improved ES bioinformatics tools appear all the more fundamental to enhance chances of identifying MMDs. It is essential to provide appropriate patient care and genetic counselling.

Lessons from two series by physicians and caregivers' self-reported data in DDX3X-related disorders.

INTRODUCTION AND METHODS: We report two series of individuals with DDX3X variations, one (48 individuals) from physicians and one (44 individuals) from caregivers. RESULTS: These two series include several symptoms in common, with fairly similar distribution, which suggests that caregivers' data are close to physicians' data. For example, both series identified early childhood symptoms that were not previously described: feeding difficulties, mean walking age, and age at first words. DISCUSSION: Each of the two datasets provides complementary knowledge. We confirmed that symptoms are similar to those in the literature and provides more details on feeding difficulties. Caregivers considered that the symptom attention-deficit/hyperactivity disorder were most worrisome. Both series also reported sleep disturbance. Recently, anxiety has been reported in individuals with DDX3X variants. We strongly suggest that attention-deficit/hyperactivity disorder, anxiety, and sleep disorders need to be treated.

Combining globally search for a regular expression and print matching lines with bibliographic monitoring of genomic database improves diagnosis.

Introduction: Exome sequencing has a diagnostic yield ranging from 25% to 70% in rare diseases and regularly implicates genes in novel disorders. Retrospective data reanalysis has demonstrated strong efficacy in improving diagnosis, but poses organizational difficulties for clinical laboratories. Patients and methods: We applied a reanalysis strategy based on intensive prospective bibliographic monitoring along with direct application of the GREP command-line tool (to "globally search for a regular expression and print matching lines") in a large ES database. For 18 months, we submitted the same five keywords of interest [(intellectual disability, (neuro)developmental delay, and (neuro)developmental disorder)] to PubMed on a daily basis to identify recently published novel disease-gene associations or new phenotypes in genes already implicated in human pathology. We used the Linux GREP tool and an in-house script to collect all variants of these genes from our 5,459 exome database. Results: After GREP queries and variant filtration, we identified 128 genes of interest and collected 56 candidate variants from 53 individuals. We confirmed causal diagnosis for 19/128 genes (15%) in 21 individuals and identified variants of unknown significance for 19/128 genes (15%) in 23 individuals. Altogether, GREP queries for only 128 genes over a period of 18 months permitted a causal diagnosis to be established in 21/2875 undiagnosed affected probands (0.7%). Conclusion: The GREP query strategy is efficient and less tedious than complete periodic reanalysis. It is an interesting reanalysis strategy to improve diagnosis.

BRAT1-related disorders: phenotypic spectrum and phenotype-genotype correlations from 97 patients.

BRAT1 biallelic variants are associated with rigidity and multifocal seizure syndrome, lethal neonatal (RMFSL), and neurodevelopmental disorder associating cerebellar atrophy with or without seizures syndrome (NEDCAS). To date, forty individuals have been reported in the literature. We collected clinical and molecular data from 57 additional cases allowing us to study a large cohort of 97 individuals and draw phenotype-genotype correlations. Fifty-nine individuals presented with BRAT1-related RMFSL phenotype. Most of them had no psychomotor acquisition (100%), epilepsy (100%), microcephaly (91%), limb rigidity (93%), and died prematurely (93%). Thirty-eight individuals presented a non-lethal phenotype of BRAT1-related NEDCAS phenotype. Seventy-six percent of the patients in this group were able to walk and 68% were able to say at least a few words. Most of them had cerebellar ataxia (82%), axial hypotonia (79%) and cerebellar atrophy (100%). Genotype-phenotype correlations in our cohort revealed that biallelic nonsense, frameshift or inframe deletion/insertion variants result in the severe BRAT1-related RMFSL phenotype (46/46; 100%). In contrast, genotypes with at least one missense were more likely associated with NEDCAS (28/34; 82%). The phenotype of patients carrying splice variants was variable: 41% presented with RMFSL (7/17) and 59% with NEDCAS (10/17).

Systematic analysis and prediction of genes associated with monogenic disorders on human chromosome X.

Disease gene discovery on chromosome (chr) X is challenging owing to its unique modes of inheritance. We undertook a systematic analysis of human chrX genes. We observe a higher proportion of disorder-associated genes and an enrichment of genes involved in cognition, language, and seizures on chrX compared to autosomes. We analyze gene constraints, exon and promoter conservation, expression, and paralogues, and report 127 genes sharing one or more attributes with known chrX disorder genes. Using machine learning classifiers trained to distinguish disease-associated from dispensable genes, we classify 247 genes, including 115 of the 127, as having high probability of being disease-associated. We provide evidence of an excess of variants in predicted genes in existing databases. Finally, we report damaging variants in CDK16 and TRPC5 in patients with intellectual disability or autism spectrum disorders. This study predicts large-scale gene-disease associations that could be used for prioritization of X-linked pathogenic variants.