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.

Unsuspected consequences of synonymous and missense variants in OCA2 can be detected in blood cell RNA samples of patients with albinism.

Oculocutaneous albinism type 2 (OCA2) is the second most frequent form of albinism and represents about 30% of OCA worldwide. As with all types of OCA, patients present with hypopigmentation of hair and skin, as well as severe visual abnormalities. We focused on a subgroup of 29 patients for whom genetic diagnosis was pending because at least one of their identified variants in or around exon 10 of OCA2 is of uncertain significance (VUS). By minigene assay, we investigated the effect of these VUS on exon 10 skipping and showed that not only intronic but also some synonymous variants can result in enhanced exon skipping. We further found that excessive skipping of exon 10 could be detected directly on blood samples of patients and of their one parent with the causal variant, avoiding invasive skin biopsies. Moreover, we show that variants, which result in lack of detectable OCA2 mRNA can be identified from blood samples as well, as shown for the most common OCA2 pathogenic missense variant c.1327G>A/p.(Val443Ile). In conclusion, blood cell RNA analysis allows testing the potential effect of any OCA2 VUS on transcription products. This should help to elucidate yet unsolved OCA2 patients and improve genetic counseling.

2p25.3 microduplications involving MYT1L: further phenotypic characterization through an assessment of 16 new cases and a literature review.

Microduplications involving the MYT1L gene have mostly been described in series of patients with isolated schizophrenia. However, few reports have been published, and the phenotype has still not been well characterized. We sought to further characterize the phenotypic spectrum of this condition by describing the clinical features of patients with a pure 2p25.3 microduplication that includes all or part of MYT1L. We assessed 16 new patients with pure 2p25.3 microduplications recruited through a French national collaboration (n = 15) and the DECIPHER database (n = 1). We also reviewed 27 patients reported in the literature. For each case, we recorded clinical data, the microduplication size, and the inheritance pattern. The clinical features were variable and included developmental and speech delays (33%), autism spectrum disorder (ASD, 23%), mild-to-moderate intellectual disability (ID, 21%), schizophrenia (23%), or behavioral disorders (16%). Eleven patients did not have an obvious neuropsychiatric disorder. The microduplications ranged from 62.4 kb to 3.8 Mb in size and led to duplication of all or part of MYT1L; seven of these duplications were intragenic. The inheritance pattern was available for 18 patients: the microduplication was inherited in 13 cases, and all parents but one had normal phenotype. Our comprehensive review and expansion of the phenotypic spectrum associated with 2p25.3 microduplications involving MYT1L should help clinicians to better assess, counsel and manage affected individuals. MYT1L microduplications are characterized by a spectrum of neuropsychiatric phenotypes with incomplete penetrance and variable expressivity, which are probably due to as-yet unknown genetic and nongenetic modifiers.

Contribution of DNA methylation profiling to the reclassification of a variant of uncertain significance in the KDM5C gene.

KDM5C encodes a demethylase of the histone H3 lysine 4 residue, involved in chromatin regulation and gene expression. Hemizygous KDM5C pathogenic variants cause X-linked intellectual disability of Claes-Jensen type. Because of its mode of inheritance and the low specificity of the clinical phenotype, interpretation of variants can be difficult, hence the need for functional studies and biomarkers specific to this disorder. We present the case of a male patient with intellectual disability, behavioral abnormalities and subtle dysmorphic features, in which genetic investigation identified a hemizygous novel missense KDM5C variant of uncertain significance (VUS), inherited from his asymptomatic mother and present in his paucisymptomatic sister. We assessed the global genomic DNA methylation status from a whole blood sample of the proband. Global DNA methylation profiling specifically identified the recently discovered epi-signature of Claes-Jensen syndrome. This result served as a biomarker which independently highlighted KDM5C as the cause of the disorder in this patient. Because of the X-linked mode of inheritance, variant reclassification had a high impact on genetic counseling in this family. This example highlights the value of global methylome profiling in situations of variants of uncertain significance in genes with a known specific epi-signature.

Deep intronic NIPBL de novo mutations and differential diagnoses revealed by whole genome and RNA sequencing in Cornelia de Lange syndrome patients.

Cornelia de Lange syndrome (CdLS; MIM# 122470) is a rare developmental disorder. Pathogenic variants in 5 genes explain approximately 50% cases, leaving the other 50% unsolved. We performed whole genome sequencing (WGS) ± RNA sequencing (RNA-seq) in 5 unsolved trios fulfilling the following criteria: (i) clinical diagnosis of classic CdLS, (ii) negative gene panel sequencing from blood and saliva-isolated DNA, (iii) unaffected parents' DNA samples available and (iv) proband's blood-isolated RNA available. A pathogenic de novo mutation (DNM) was observed in a CdLS differential diagnosis gene in 3/5 patients, namely POU3F3, SPEN, and TAF1. In the other two, we identified two distinct deep intronic DNM in NIPBL predicted to create a novel splice site. RT-PCRs and RNA-Seq showed aberrant transcripts leading to the creation of a novel frameshift exon. Our findings suggest the relevance of WGS in unsolved suspected CdLS cases and that deep intronic variants may account for a proportion of them.

uORF-introducing variants in the 5'UTR of the NIPBL gene as a cause of Cornelia de Lange syndrome.

Cornelia de Lange syndrome (CdLS) is a clinically-recognizable rare developmental disorder. About 70% of patients carry a missense or loss-of-function pathogenic variant in the NIPBL gene. We hypothesized that some variants in the 5'-untranslated region (UTR) of NIPBL may create an upstream open reading frame (uORF), putatively leading to a loss of function. We searched for NIPBL 5'-UTR variants potentially introducing uORF by (i) reannotating NGS data of 102 unsolved CdLS patients and (ii) literature and variant databases search. We set up a green fluorescent protein (GFP) reporter assay and studied NIPBL expression in a lymphoblastoid cell line (LCL). We identified two variants introducing a novel ATG codon sequence in the 5'-UTR of NIPBL, both predicted to introduce uORF: a novel c.-457_-456delinsAT de novo mutation in a 15-year-old male with classic CdLS, and a c.-94C>T variant in a published family. Our reporter assay showed a significant decrease of GFP levels in both mutant contexts, with similar levels of messenger RNA (mRNA) as compared to wt constructs. Assessment of LCL of one patient showed consistent results with decreased NIPBL protein and unchanged mRNA levels. 5'-UTR uORF-introducing NIPBL variants may represent a rare source of pathogenic variants in unsolved CdLS patients.

MYT1L-associated neurodevelopmental disorder: description of 40 new cases and literature review of clinical and molecular aspects.

Pathogenic variants of the myelin transcription factor-1 like (MYT1L) gene include heterozygous missense, truncating variants and 2p25.3 microdeletions and cause a syndromic neurodevelopmental disorder (OMIM#616,521). Despite enrichment in de novo mutations in several developmental disorders and autism studies, the data on clinical characteristics and genotype-phenotype correlations are scarce, with only 22 patients with single nucleotide pathogenic variants reported. We aimed to further characterize this disorder at both the clinical and molecular levels by gathering a large series of patients with MYT1L-associated neurodevelopmental disorder. We collected genetic information on 40 unreported patients with likely pathogenic/pathogenic MYT1L variants and performed a comprehensive review of published data (total = 62 patients). We confirm that the main phenotypic features of the MYT1L-related disorder are developmental delay with language delay (95%), intellectual disability (ID, 70%), overweight or obesity (58%), behavioral disorders (98%) and epilepsy (23%). We highlight novel clinical characteristics, such as learning disabilities without ID (30%) and feeding difficulties during infancy (18%). We further describe the varied dysmorphic features (67%) and present the changes in weight over time of 27 patients. We show that patients harboring highly clustered missense variants in the 2-3-ZNF domains are not clinically distinguishable from patients with truncating variants. We provide an updated overview of clinical and genetic data of the MYT1L-associated neurodevelopmental disorder, hence improving diagnosis and clinical management of these patients.

Hypersociability associated with developmental delay, macrocephaly and facial dysmorphism points to CHD3 mutations.

CHD3-related syndrome, also known as Snijders Blok-Campeau syndrome, is a rare developmental disorder described in 2018, caused by de novo pathogenic variants in the CHD3 gene. This syndrome is characterized by global developmental delay, speech delay, intellectual disability, hypotonia and behavioral disorders including autism spectrum disorder (ASD). Typical dysmorphic features include macrocephaly, hypertelorism, enophthalmia, sparse eyebrows, bulging forehead, midface hypoplasia, prominent nose and pointed chin. To our knowledge, there have been no other clinical descriptions of patients since the initial publication. We report the clinical description of a 21-year-old patient harboring a pathogenic de novo variant in CHD3. We reviewed the clinical features of the 35 previously reported patients. Main features were severe intellectual disability, dysmorphic facies, macrocephaly, cryptorchidism, pectus carinatum, severe ophthalmologic abnormalities and behavioral disorders including ASD, and a frank happy demeanor. Hypersociability, which was a noticeable clinical feature in our case, despite ASD, is an uncommon behavioral feature in syndromic intellectual disabilities. Our report supports hypersociability as a suggestive feature of CHD3-related syndrome along with developmental delay, macrocephaly and a dysmorphic facies.

Exome sequencing identifies the first genetic determinants of sirenomelia in humans.

Sirenomelia is a rare severe malformation sequence of unknown cause characterized by fused legs and severe visceral abnormalities. We present a series of nine families including two rare familial aggregations of sirenomelia investigated by a trio-based exome sequencing strategy. This approach identified CDX2 variants in the two familial aggregations, both fitting an autosomal dominant pattern of inheritance with variable expressivity. CDX2 is a major regulator of caudal development in vertebrate and mouse heterozygotes are a previously described model of sirenomelia. Remarkably, the p.(Arg237His) variant has already been reported in a patient with persistent cloaca. Analysis of the sporadic cases revealed six additional candidate variants including a de novo frameshift variant in the genetically constrained NKD1 gene, encoding a known interactor of CDX2. We provide the first insights for a genetic contribution in human sirenomelia and highlight the role of Cdx and Wnt signaling pathways in the development of this disorder.