Rare ACTN2 Frameshift Variants Resulting in Protein Extension Cause Distal Myopathy and Hypertrophic Cardiomyopathy through Protein Aggregation.

Distal myopathies are a group of rare, inherited muscular disorders characterized by progressive loss of muscle fibers that begins in the distal parts of arms and legs. Recently, variants in a new disease gene, ACTN2 , have been shown to cause distal myopathy. ACTN2 , a gene previously only associated with cardiomyopathies, encodes alpha-actinin-2, a protein expressed in both cardiac and skeletal sarcomeres. The primary function of alpha-actinin-2 is to link actin and titin to the sarcomere Z-disk. New ACTN2 variants are continuously discovered, however, the clinical significance of many variants remains unknown. Thus, lack of clear genotype-phenotype correlations in ACTN2 -related diseases, actininopathies, persists. Objective: The objective of the study is to characterize the pathomechanisms underlying actininopathies. Methods: Functional characterization in C2C12 cell models of several ACTN2 variants is conducted, including frameshift and missense variants associated with dominant actininopathies. We assess the genotype-phenotype correlations of actininopathies using clinical data from several patients carrying these variants. Results: The results show that the missense variants associated with a recessive form of actininopathy do not cause detectable alpha-actinin-2 aggregates in the cell model. Conversely, dominant frameshift variants causing a protein extension do produce alpha-actinin-2 aggregates. Interpretation: The results suggest that alpha-actinin-2 aggregation is the disease mechanism underlying some dominant actininopathies, and thus we recommend that protein-extending frameshift variants in ACTN2 should be classified as pathogenic. However, this mechanism is likely elicited by only a limited number of variants. Alternative functional characterization methods should be explored to further investigate other molecular mechanisms underlying actininopathies.

Idiopathic pulmonary fibrosis with benign SFTPC variant and pathogenic MARS1 mutations: can't see the forest for the trees!

Even in the absence of liver disease, MARS1 screening should be considered in severe lung fibrosis of young individuals. Interpretation of the genetic variants can evolve with improvement of knowledge (databases, bioinformatic tools) over time. https://bit.ly/45OxF5E.

Low risk of embryonic and other cancers in PIK3CA-related overgrowth spectrum: Impact on screening recommendations.

The PIK3CA-related overgrowth spectrum (PROS) encompasses various conditions caused by mosaic activating PIK3CA variants. PIK3CA somatic variants are also involved in various cancer types. Some generalized overgrowth syndromes are associated with an increased risk of Wilms tumor (WT). In PROS, abdominal ultrasound surveillance has been advocated to detect WT. We aimed to determine the risk of embryonic and other types of tumors in patients with PROS in order to evaluate surveillance relevance. We searched the clinical charts from 267 PROS patients for the diagnosis of cancer, and reviewed the medical literature for the risk of cancer. In our cohort, six patients developed a cancer (2.2%), and Kaplan Meier analyses estimated cumulative probabilities of cancer occurrence at 45 years of age was 5.6% (95% CI = 1.35%-21.8%). The presence of the PIK3CA variant was only confirmed in two out of four tumor samples. In the literature and our cohort, six cases of Wilms tumor/nephrogenic rests (0.12%) and four cases of other cancers have been reported out of 483 proven PIK3CA patients, in particular the p.(His1047Leu/Arg) variant. The risk of WT in PROS being lower than 5%, this is insufficient evidence to recommend routine abdominal imaging. Long-term follow-up studies are needed to evaluate the risk of other cancer types, as well as the relationship with the extent of tissue mosaicism and the presence or not of the variant in the tumor samples.

Identification and in vivo functional investigation of a HOMER2 nonstop variant causing hearing loss.

DFNA68 is a rare subtype of autosomal dominant nonsyndromic hearing impairment caused by heterozygous alterations in the HOMER2 gene. To date, only 5 pathogenic or likely pathogenic coding variants, including two missense substitutions (c.188 C > T and c.587 G > C), a single base pair duplication (c.840dupC) and two short deletions (c.592_597delACCACA and c.832_836delCCTCA) have been described in 5 families. In this study, we report a novel HOMER2 variation, identified by massively parallel sequencing, in a Sicilian family suffering from progressive dominant hearing loss over 3 generations. This novel alteration is a nonstop substitution (c.1064 A > G) that converts the translational termination codon (TAG) of the gene into a tryptophan codon (TGG) and is predicted to extend the HOMER2 protein by 10 amino acids. RNA analyses from the proband suggested that HOMER2 transcripts carrying the nonstop variant escaped the non-stop decay pathway. Finally, in vivo studies using a zebrafish animal model and behavioral tests clearly established the deleterious impact of this novel HOMER2 alteration on hearing function. This study identifies the fourth causal variation responsible for DFNA68 and describes a simple in vivo approach to assess the pathogenicity of candidate HOMER2 variants.

French recommendations for the diagnosis and management of lymphangioleiomyomatosis.

BACKGROUND: The present article is an English-language version of the French National Diagnostic and Care Protocol, a pragmatic tool to optimize and harmonize the diagnosis, care pathway, management and follow-up of lymphangioleiomyomatosis in France. METHODS: Practical recommendations were developed in accordance with the method for developing a National Diagnosis and Care Protocol for rare diseases of the Haute Autorité de Santé and following international guidelines and literature on lymphangioleiomyomatosis. It was developed by a multidisciplinary group, with the help of patient representatives and of RespiFIL, the rare disease network on respiratory diseases. RESULTS: Lymphangioleiomyomatosis is a rare lung disease characterised by a proliferation of smooth muscle cells that leads to the formation of multiple lung cysts. It occurs sporadically or as part of a genetic disease called tuberous sclerosis complex (TSC). The document addresses multiple aspects of the disease, to guide the clinicians regarding when to suspect a diagnosis of lymphangioleiomyomatosis, what to do in case of recurrent pneumothorax or angiomyolipomas, what investigations are needed to make the diagnosis of lymphangioleiomyomatosis, what the diagnostic criteria are for lymphangioleiomyomatosis, what the principles of management are, and how follow-up can be organised. Recommendations are made regarding the use of pharmaceutical specialties and treatment other than medications. CONCLUSION: These recommendations are intended to guide the diagnosis and practical management of pulmonary lymphangioleiomyomatosis.

Clinical heterogeneity of NADSYN1-associated VCRL syndrome.

The NADSYN1 gene [MIM*608285] encodes the NAD synthetase 1 enzyme involved in the final step of NAD biosynthesis, crucial for cell metabolism and organ embryogenesis. Perturbating the role of NAD biosynthesis results in the association of vertebral, cardiac, renal, and limb anomalies (VCRL). This condition was initially characterized as severe with perinatal lethality or developmental delay and complex malformations in alive cases. Sixteen NADSYN1-associated patients have been published so far. This study illustrates the wide phenotypic variability in NADSYN1-associated NAD deficiency disorder. We report the clinical and molecular findings in three novel cases, two of them being siblings with the same homozygous variant and presenting with either a very severe prenatal lethal or a mild phenotypic form. In addition to an exhaustive literature, we validate the expansion of the spectrum of NAD deficiency disorder. Our findings indicate that NAD deficiency disorder should be suspected not only in the presence of the full spectrum of VCRL, but even a single of the aforementioned organs is affected. Decreased plasmatic levels of NAD should then strongly encourage the screening for any of the genes responsible for a NAD deficiency disorder.

Congenital hydrocephalus: new Mendelian mutations and evidence for oligogenic inheritance.

BACKGROUND: Congenital hydrocephalus is characterized by ventriculomegaly, defined as a dilatation of cerebral ventricles, and thought to be due to impaired cerebrospinal fluid (CSF) homeostasis. Primary congenital hydrocephalus is a subset of cases with prenatal onset and absence of another primary cause, e.g., brain hemorrhage. Published series report a Mendelian cause in only a minority of cases. In this study, we analyzed exome data of PCH patients in search of novel causal genes and addressed the possibility of an underlying oligogenic mode of inheritance for PCH. MATERIALS AND METHODS: We sequenced the exome in 28 unrelated probands with PCH, 12 of whom from families with at least two affected siblings and 9 of whom consanguineous, thereby increasing the contribution of genetic causes. Patient exome data were first analyzed for rare (MAF < 0.005) transmitted or de novo variants. Population stratification of unrelated PCH patients and controls was determined by principle component analysis, and outliers identified using Mahalanobis distance 5% as cutoff. Patient and control exome data for genes biologically related to cilia (SYScilia database) were analyzed by mutation burden test. RESULTS: In 18% of probands, we identify a causal (pathogenic or likely pathogenic) variant of a known hydrocephalus gene, including genes for postnatal, syndromic hydrocephalus, not previously reported in isolated PCH. In a further 11%, we identify mutations in novel candidate genes. Through mutation burden tests, we demonstrate a significant burden of genetic variants in genes coding for proteins of the primary cilium in PCH patients compared to controls. CONCLUSION: Our study confirms the low contribution of Mendelian mutations in PCH and reports PCH as a phenotypic presentation of some known genes known for syndromic, postnatal hydrocephalus. Furthermore, this study identifies novel Mendelian candidate genes, and provides evidence for oligogenic inheritance implicating primary cilia in PCH.

Exome sequencing as a first-tier test for copy number variant detection: retrospective evaluation and prospective screening in 2418 cases.

BACKGROUND: Despite the availability of whole exome (WES) and genome sequencing (WGS), chromosomal microarray (CMA) remains the first-line diagnostic test in most rare disorders diagnostic workup, looking for copy number variations (CNVs), with a diagnostic yield of 10%-20%. The question of the equivalence of CMA and WES in CNV calling is an organisational and economic question, especially when ordering a WGS after a negative CMA and/or WES. METHODS: This study measures the equivalence between CMA and GATK4 exome sequencing depth of coverage method in detecting coding CNVs on a retrospective cohort of 615 unrelated individuals. A prospective detection of WES-CNV on a cohort of 2418 unrelated individuals, including the 615 individuals from the validation cohort, was performed. RESULTS: On the retrospective validation cohort, every CNV detectable by the method (ie, a CNV with at least one exon not in a dark zone) was accurately called (64/64 events). In the prospective cohort, 32 diagnoses were performed among the 2418 individuals with CNVs ranging from 704 bp to aneuploidy. An incidental finding was reported. The overall increase in diagnostic yield was of 1.7%, varying from 1.2% in individuals with multiple congenital anomalies to 1.9% in individuals with chronic kidney failure. CONCLUSION: Combining single-nucleotide variant (SNV) and CNV detection increases the suitability of exome sequencing as a first-tier diagnostic test for suspected rare Mendelian disorders. Before considering the prescription of a WGS after a negative WES, a careful reanalysis with updated CNV calling and SNV annotation should be considered.

Bi-allelic loss-of-function variants in PPFIBP1 cause a neurodevelopmental disorder with microcephaly, epilepsy, and periventricular calcifications.

PPFIBP1 encodes for the liprin-ß1 protein, which has been shown to play a role in neuronal outgrowth and synapse formation in Drosophila melanogaster. By exome and genome sequencing, we detected nine ultra-rare homozygous loss-of-function variants in 16 individuals from 12 unrelated families. The individuals presented with moderate to profound developmental delay, often refractory early-onset epilepsy, and progressive microcephaly. Further common clinical findings included muscular hyper- and hypotonia, spasticity, failure to thrive and short stature, feeding difficulties, impaired vision, and congenital heart defects. Neuroimaging revealed abnormalities of brain morphology with leukoencephalopathy, ventriculomegaly, cortical abnormalities, and intracranial periventricular calcifications as major features. In a fetus with intracranial calcifications, we identified a rare homozygous missense variant that by structural analysis was predicted to disturb the topology of the SAM domain region that is essential for protein-protein interaction. For further insight into the effects of PPFIBP1 loss of function, we performed automated behavioral phenotyping of a Caenorhabditis elegans PPFIBP1/hlb-1 knockout model, which revealed defects in spontaneous and light-induced behavior and confirmed resistance to the acetylcholinesterase inhibitor aldicarb, suggesting a defect in the neuronal presynaptic zone. In conclusion, we establish bi-allelic loss-of-function variants in PPFIBP1 as a cause of an autosomal recessive severe neurodevelopmental disorder with early-onset epilepsy, microcephaly, and periventricular calcifications.

Epileptic spasms are associated with increased stereo-electroencephalography derived functional connectivity in tuberous sclerosis complex.

OBJECTIVE: Epileptic spasms (ES) are common in tuberous sclerosis complex (TSC). However, the underlying network alterations and relationship with epileptogenic tubers are poorly understood. We examined interictal functional connectivity (FC) using stereo-electroencephalography (SEEG) in patients with TSC to investigate the relationship between tubers, epileptogenicity, and ES. METHODS: We analyzed 18 patients with TSC who underwent SEEG (mean age = 11.5 years). The dominant tuber (DT) was defined as the most epileptogenic tuber using the epileptogenicity index. Epileptogenic zone (EZ) organization was quantitatively separated into focal (isolated DT) and complex (all other patterns). Using a 20-min interictal recording, FC was estimated with nonlinear regression, h2 . We calculated (1) intrazone FC within all sampled tubers and normal-appearing cortical zones, respectively; and (2) interzone FC involving connections between DT, other tubers, and normal cortex. The relationship between FC and (1) presence of ES as a current seizure type at the time of SEEG, (2) EZ organization, and (3) epileptogenicity was analyzed using a mixed generalized linear model. Spike rate and distance between zones were considered in the model as covariates. RESULTS: Six patients had ES as a current seizure type at time of SEEG. ES patients had a greater number of tubers with a fluid-attenuated inversion recovery hypointense center (p < .001), and none had TSC1 mutations. The presence of ES was independently associated with increased FC within both intrazone (p = .033) and interzone (p = .011) networks. Post hoc analyses identified that increased FC was associated with ES across tuber and nontuber networks. EZ organization and epileptogenicity biomarkers were not associated with FC. SIGNIFICANCE: Increased cortical synchrony among both tuber and nontuber networks is characteristic of patients with ES and independent of both EZ organization and tuber epileptogenicity. This further supports the prospect of FC biomarkers aiding treatment paradigms in TSC.

A second individual with rhizomelic spondyloepimetaphyseal dysplasia and homozygous variant in GNPNAT1.

Spondyloepimetaphyseal dysplasias (SEMDs) belong to a clinically and genetically heterogeneous group of inherited skeletal disorders defined by a defect in the growth and shape of vertebrae, epiphyses and metaphyses. Rhizomelic SEMD is characterized by a disproportionate small stature caused by severe shortening and deformation of the limbs' proximal bones, with the cranio-facial sphere unaffected. We report a second individual, an 8-year-old girl, with autosomal recessive rhizomelic SEMD associated with a homozygous exonic missense variant, c.226G > A p.(Glu76Lys), in GNPNAT1 identified by trio genome sequencing. Our data corroborate the recent findings of Ain et al. and further delineate the clinical and radiographic features of this form of SEMD associated with rhizomelic dysplasia while outlining a potential hotspot in this newly described genetic disorder.

GGCX-related congenital combined vitamin K-dependent clotting factors deficiency-1: Description of a fetus with chondrodysplasia punctata.

Congenital combined vitamin K-dependent clotting factors deficiency (VKCFD) is a rare autosomal recessive disease resulting in hemorrhagic symptoms usually associated with developmental disorders and bone abnormalities. Pathogenic variants in two genes encoding enzymes of the vitamin K cycle, GGCX and VKORC1, can lead to this disorder. We present the case of a male fetus with a brachytelephalangic chondrodysplasia punctata (CDP), absence of nasal bone, growth restriction, and bilateral ventriculomegaly at 18 weeks of gestation. Pathological examination showed a Binder phenotype, hypoplastic distal phalanges, stippled epiphyses, and brain abnormalities suggestive of a brain hemorrhage. Two GGCX pathogenic variants inherited respectively from the mother and the father were identified. To our knowledge, this is the first prenatal description of VKCFD. Even if it remains a rare etiology, which is mostly described in children or adult patients, VKCFD should be considered in fetuses with CDP.

Historical Patterns of Diagnosis, Treatments, and Outcome of Epilepsy Associated With Tuberous Sclerosis Complex: Results From TOSCA Registry.

Background: Epilepsy is the most common neurological manifestation in individuals with tuberous sclerosis complex (TSC). However, real-world evidence on diagnosis and treatment patterns is limited. Here, we present data from TuberOus Sclerosis registry to increase disease Awareness (TOSCA) on changes in patterns of epilepsy diagnosis, treatments, and outcomes over time, and detailed epilepsy characteristics from the epilepsy substudy. Methods: TuberOus Sclerosis registry to increase disease Awareness (TOSCA) was a multicentre, international disease registry, consisting of a main study that collected data on overall diagnostic characteristics and associated clinical features, and six substudies focusing on specific TSC manifestations. The epilepsy substudy investigated detailed epilepsy characteristics and their correlation to genotype and intelligence quotient (IQ). Results: Epilepsy was reported in 85% of participants, more commonly in younger individuals (67.8% in 1970s to 91.8% in last decade), while rate of treatments was similar across ages (>93% for both infantile spasms and focal seizures, except prior to 1960). Vigabatrin (VGB) was the most commonly used antiepileptic drugs (AEDs). Individuals with infantile spasms showed a higher treatment response over time with lower usage of steroids. Individuals with focal seizures reported similar rates of drug resistance (32.5-43.3%). Use of vagus nerve stimulation (VNS), ketogenic diet, and surgery remained low. Discussion: The epilepsy substudy included 162 individuals from nine countries. At epilepsy onset, most individuals with infantile spasms (73.2%) and focal seizures (74.5%) received monotherapies. Vigabatrin was first-line treatment in 45% of individuals with infantile spasms. Changes in initial AEDs were commonly reported due to inadequate efficacy. TSC1 mutations were associated with less severe epilepsy phenotypes and more individuals with normal IQ. In individuals with TSC diagnosis before seizure onset, electroencephalogram (EEG) was performed prior to seizures in only 12.5 and 25% of subsequent infantile spasms and focal seizures, respectively. Conclusions: Our study confirms the high prevalence of epilepsy in TSC individuals and less severe phenotypes with TSC1 mutations. Vigabatrin improved the outcome of infantile spasms and should be used as first-line treatment. There is, however, still a need for improving therapies in focal seizures. Electroencephalogram follow-up prior to seizure-onset should be promoted for all infants with TSC in order to facilitate preventive or early treatment.

Care management in a French cohort with Down syndrome from the AnDDI-Rares/CNSA study.

Down syndrome (DS) is a genetic neurodevelopmental disorder. In individuals with DS, a multidisciplinary approach to care is required to prevent multiple medical complications. The aim of this study was to describe the rehabilitation, medical care, and educational and social support provided to school-aged French DS patients with varying neuropsychological profiles. A mixed study was conducted. Quantitative data were obtained from a French multicentre study that included patients aged 4-20 years with diverse genetic syndromes. Qualitative data were collected by semi-structured face-to-face interviews and focus groups. Ninety-five DS subjects with a mean age of 10.9 years were included. Sixty-six per cent had a moderate intellectual disability (ID) and 18.9% had a severe ID. Medical supervision was generally multidisciplinary but access to medical specialists was often difficult. In terms of education, 94% of children under the age of six were in typical classes. After the age of 15, 75% were in medico-social institutions. Analysis of multidisciplinary rehabilitation conducted in the public and private sectors revealed failure to access physiotherapy, psychomotor therapy and occupational therapy, but not speech therapy. The main barrier encountered by patients was the difficulty accessing appropriate facilities due to a lack of space and long waiting lists. In conclusion, children and adolescents with DS generally received appropriate care. Though the management of children with DS has been improved considerably, access to health facilities remains inadequate.

Rare manifestations and malignancies in tuberous sclerosis complex: findings from the TuberOus SClerosis registry to increAse disease awareness (TOSCA).

BACKGROUND: Tuberous sclerosis complex (TSC) is a rare multisystem autosomal dominant disorder caused by pathogenic variants in either the TSC1 or TSC2 gene. Common manifestations of TSC have been grouped into major and minor clinical diagnostic criteria and assessed in clinical routine workup. However, case studies point towards the existence of rare disease manifestations and to the potential association of TSC with malignant tumors. In this study we sought to characterize rare manifestations and malignancies using a large cohort of patients. METHODS: TuberOus SClerosis registry to increAse disease awareness (TOSCA) is a multicenter, international disease registry collecting clinical manifestations and characteristics of patients with TSC, both retrospectively and prospectively. We report rates and characteristics of rare manifestations and malignancies in patients with TSC who had enrolled in the TOSCA registry. We also examined these manifestations by age, sex, and genotype (TSC1 or TSC2). RESULTS: Overall, 2211 patients with TSC were enrolled in the study. Rare manifestations were reported in 382 (17.3%) study participants and malignancies in 65 (2.9%). Of these rare manifestations, the most frequent were bone sclerotic foci (39.5%), scoliosis (23%), thyroid adenoma (5.5%), adrenal angiomyolipoma (4.5%), hemihypertrophy and pancreatic neuroendocrine tumors (pNET; both 3.1%). These rare manifestations were more commonly observed in adults than children (66.2% vs. 22.7%), in females versus males (58.4% vs. 41.6%; except for scoliosis: 48.9% vs. 51.1%), and in those with TSC2 versus TSC1 (67.0% vs. 21.1%; except for thyroid adenoma: 42.9% vs. 57.1%). In the 65 individuals with reported malignancies, the most common were renal cell carcinoma (47.7%), followed by breast (10.8%) and thyroid cancer (9.2%). Although malignancies were more common in adult patients, 26.1% were reported in children and 63.1% in individuals < 40 years. TSC1 mutations were over-represented in individuals with malignancies compared to the overall TOSCA cohort (32.1% vs. 18.5%). CONCLUSION: Rare manifestations were observed in a significant proportion of individuals with TSC. We recommend further examination of rare manifestations in TSC. Collectively, malignancies were infrequent findings in our cohort. However, compared to the general population, malignant tumors occurred earlier in age and some tumor types were more common.

TuberOus SClerosis registry to increAse disease awareness (TOSCA) Post-Authorisation Safety Study of Everolimus in Patients With Tuberous Sclerosis Complex.

This non-interventional post-authorisation safety study (PASS) assessed the long-term safety of everolimus in patients with tuberous sclerosis complex (TSC) who participated in the TuberOus SClerosis registry to increase disease Awareness (TOSCA) clinical study and received everolimus for the licensed indications in the European Union. The rate of adverse events (AEs), AEs that led to dose adjustments or treatment discontinuation, AEs of potential clinical interest, treatment-related AEs (TRAEs), serious AEs (SAEs), and deaths were documented. One hundred seventy-nine patients were included in the first 5 years of observation; 118 of 179 patients had an AE of any grade, with the most common AEs being stomatitis (7.8%) and headache (7.3%). AEs caused dose adjustments in 56 patients (31.3%) and treatment discontinuation in nine patients (5%). AEs appeared to be more frequent and severe in children. On Tanner staging, all patients displayed signs of age-appropriate sexual maturation. Twenty-two of 106 female (20.8%) patients had menstrual cycle disorders. The most frequent TRAEs were stomatitis (6.7%) and aphthous mouth ulcer (5.6%). SAEs were reported in 54 patients (30.2%); the most frequent SAE was pneumonia (>3% patients; grade 2, 1.1%, and grade 3, 2.8%). Three deaths were reported, all in patients who had discontinued everolimus for more than 28 days, and none were thought to be related to everolimus according to the treating physicians. The PASS sub-study reflects the safety and tolerability of everolimus in the management of TSC in real-world routine clinical practice.

Screening of the duplication 24 pb of ARX gene in Moroccan patients with X-linked Intellectual Disability.

OBJECTIVE: Intellectual Disability (ID) represents a neuropsychiatric disorder, which its etiopathogenesis remains insufficiently understood. Mutations in the Aristaless Related Homeobox gene (ARX) have been identified to cause syndromic and nonsyndromic (NS-ID). The most recurrent mutation of this gene is a duplication of 24pb, c.428-451dup. Epidemiological and genetic studies about ID in the Moroccan population remain very scarce, and none study is carried out on the ARX gene. This work aimed to study c.428-451dup (24 bp) mutation in the exon 2 of the ARX gene in 118 males' Moroccan patients with milder NS-ID to evaluate if the gene screening is a good tool for identifying NS-ID. RESULTS: Our mutational analysis did not show any dup(24pb) in our patients. This is because based on findings from previous studies that found ARX mutations in 70% of families with NS-ID, and in most cases, 1.5-6.1% of individuals with NS-ID have this duplication. Since 1/118 = 0.0084 (0.84%) is not much different from 1.5%, then it is reasonable that this could a sample size artifact. A complete screening of the entire ARX gene, including the five exons, should be fulfilled. Further investigations are required to confirm these results.

X-linked partial corpus callosum agenesis with mild intellectual disability: identification of a novel L1CAM pathogenic variant.

Pathogenic variants in L1CAM, the gene encoding the L1 cell adhesion molecule, are responsible for a wide clinical spectrum including X-linked hydrocephalus with stenosis of the Sylvius aqueduct, MASA syndrome (mental retardation, aphasia, shuffling gait, adducted thumbs), and a form of spastic paraplegia (SPG1). A moderate phenotype with mild intellectual disability (ID) and X-linked partial corpus callosum agenesis (CCA) has only been related to L1CAM in one family. We report here a second family, including 5 patients with mild to moderate ID and partial CCA without signs usually associated with L1CAM pathogenic variations (such as hydrocephalus, pyramidal syndrome, thumb adductus, aphasia). We identified a previously unreported c.3226A > C transversion leading to a p.Thr1076Pro amino acid substitution in the fifth fibronectin type III domain (FnIII) of the protein which co-segregates with the phenotype within the family. We performed in vitro assays to assess the pathogenic status of this variation. First, the expression of the novel p.Thr1076Pro mutant in COS7 cells resulted in endoplasmic reticulum (ER) retention and reduced L1CAM cell surface expression, which is expected to affect both L1CAM-mediated cell-cell adhesion and neurite growth. Second, immunoblotting techniques showed that the immature form of the L1CAM protein was increased, indicating that this variation led to a lack of maturation of the protein. ID associated with CCA is not a common clinical presentation of L1CAM pathogenic variants. Genome-wide analyses will identify such variations and it is important to acknowledge this atypical phenotype.

Clinical and neuroimaging findings in 33 patients with MCAP syndrome: A survey to evaluate relevant endpoints for future clinical trials.

Megalencephaly-CApillary malformation-Polymicrogyria (MCAP) syndrome results from somatic mosaic gain-of-function variants in PIK3CA. Main features are macrocephaly, somatic overgrowth, cutaneous vascular malformations, connective tissue dysplasia, neurodevelopmental delay, and brain anomalies. The objectives of this study were to describe the clinical and radiological features of MCAP, to suggest relevant clinical endpoints applicable in future trials of targeted drug therapy. Based on a French collaboration, we collected clinical features of 33 patients (21 females, 12 males, median age of 9.9 years) with MCAP carrying mosaic PIK3CA pathogenic variants. MRI images were reviewed for 21 patients. The main clinical features reported were macrocephaly at birth (20/31), postnatal macrocephaly (31/32), body/facial asymmetry (21/33), cutaneous capillary malformations (naevus flammeus 28/33, cutis marmorata 17/33). Intellectual disability was present in 15 patients. Among the MRI images reviewed, the neuroimaging findings were megalencephaly (20/21), thickening of corpus callosum (16/21), Chiari malformation (12/21), ventriculomegaly/hydrocephaly (10/21), cerebral asymmetry (6/21) and polymicrogyria (2/21). This study confirms the main known clinical features that defines MCAP syndrome. Taking into account the phenotypic heterogeneity in MCAP patients, in the context of emerging clinical trials, we suggest that patients should be evaluated based on the main neurocognitive expression on each patient.

Genotype-first in a cohort of 95 fetuses with multiple congenital abnormalities: when exome sequencing reveals unexpected fetal phenotype-genotype correlations.

PURPOSE: Molecular diagnosis based on singleton exome sequencing (sES) is particularly challenging in fetuses with multiple congenital abnormalities (MCA). Indeed, some studies reveal a diagnostic yield of about 20%, far lower than in live birth individuals showing developmental abnormalities (30%), suggesting that standard analyses, based on the correlation between clinical hallmarks described in postnatal syndromic presentations and genotype, may underestimate the impact of the genetic variants identified in fetal analyses. METHODS: We performed sES in 95 fetuses with MCA. Blind to phenotype, we applied a genotype-first approach consisting of combined analyses based on variants annotation and bioinformatics predictions followed by reverse phenotyping. Initially applied to OMIM-morbid genes, analyses were then extended to all genes. We complemented our approach by using reverse phenotyping, variant segregation analysis, bibliographic search and data sharing in order to establish the clinical significance of the prioritised variants. RESULTS: sES rapidly identified causal variant in 24/95 fetuses (25%), variants of unknown significance in OMIM genes in 8/95 fetuses (8%) and six novel candidate genes in 6/95 fetuses (6%). CONCLUSIONS: This method, based on a genotype-first approach followed by reverse phenotyping, shed light on unexpected fetal phenotype-genotype correlations, emphasising the relevance of prenatal studies to reveal extreme clinical presentations associated with well-known Mendelian disorders.