Abstract
Objective
NPRL3
‐related epilepsy (NRE) is an emerging condition set within the wide GATOR‐1 spectrum with a particularly heterogeneous and elusive phenotypic expression. Here, we delineated ...the genotype–phenotype spectrum of NRE, reporting an illustrative familial case and reviewing pertinent literature.
Methods
Through exome sequencing (ES), we investigated a 12‐year‐old girl with recurrent focal motor seizures during sleep, suggestive of sleep‐related hypermotor epilepsy (SHE), and a family history of epilepsy in siblings. Variant segregation analysis was performed by Sanger sequencing. All previously published NRE patients were thoroughly reviewed and their electroclinical features were analyzed and compared with the reported subjects.
Results
In the proband, ES detected the novel
NPRL3
frameshift variant (NM_001077350.3): c.151_152del (p.Thr51Glyfs*5). This variant is predicted to cause a loss of function and segregated in one affected brother. The review of 76 patients from 18 publications revealed the predominance of focal‐onset seizures (67/74–90%), with mainly frontal and frontotemporal (32/67–47.7%), unspecified (19/67–28%), or temporal (9/67–13%) onset. Epileptic syndromes included familial focal epilepsy with variable foci (FFEVF) (29/74–39%) and SHE (11/74–14.9%). Fifteen patients out of 60 (25%) underwent epilepsy surgery, 11 of whom achieved complete seizure remission (11/15–73%). Focal cortical dysplasia (FCD) type 2A was the most frequent histopathological finding.
Significance
We reported an illustrative
NPRL3
‐related epilepsy (NRE) family with incomplete penetrance. This condition consists of a heterogeneous spectrum of clinical and neuroradiological features. Focal‐onset motor seizures are predominant, and almost half of the cases fulfill the criteria for SHE or FFEVF. MRI‐negative cases are prevalent, but the association with malformations of cortical developments (MCDs) is significant, especially FCD type 2a. The beneficial impact of epilepsy surgery in patients with MCD‐related epilepsy further supports the inclusion of brain MRI in the workup of NRE patients.
Congenital central hypoventilation syndrome (CCHS) is a rare neurocristopathy characterized by absence of adequate autonomic control of respiration with decreased sensitivity to hypoxia and ...hypercapnia. Frameshift mutations and polyalanine triplet expansions in the coding region of PHOX2B have been identified in the vast majority of CCHS patients and a correlation between length of the expanded region and severity of CCHS has been reported. In this work, we have undertaken in vitro analyses aimed at identifying the pathogenetic mechanisms which underlie the effects of PHOX2B mutations in CCHS. According to the known role of this gene, a transcription factor expressed during autonomic nervous system development, we have tested the transcriptional activity of WT and mutant PHOX2B expression constructs on the regulatory regions of two target genes, DβH and PHOX2A. We observed that the two sets of mutations play different roles in the transcriptional regulation of these genes, showing a correlation between the length of polyalanine expansions and the severity of reduced transcriptional activity. In particular, although reduced transactivation due to polyalanine expansions may be caused by retention of the mutated protein in the cytoplasm or in the nuclear aggregates, frameshift mutations did not impair the PHOX2B nuclear income, suggesting a different mechanism through which they would exert the observed effects on target promoters. Moreover, the frameshift due to deletion of a cytosine residue seems to cause sequestration of the corresponding mutant PHOX2B in the nucleolar compartment.
Neurofibromatosis type 1 (NF1) is a neurocutaneous genetic disorder with a broad spectrum of associated signs and symptoms, including skeletal anomalies. The association of NF1 with anterior chest ...wall deformities has been recently reported, especially the pectus excavatum (PE). Over the years, several authors have suggested loss of heterozygosity (LOH) as the possible pathogenic mechanism underlying the development of the typical NF1 skeletal features. Here, we report a NF1 patient with severe chest deformity and harboring the germline heterozygous pathogenic NF1 variant NM_001042492.3: c.4271delC p.(Ala1424Glufs∗4). Through next-generation sequencing (NGS), we investigated the affected cartilage from the PE deformity and identified the additional frameshift variant NM_001042492.3: c.2953delC p.(Gln985Lysfs∗7), occurring as a somatic NF1 second hit mutation. Exome sequencing confirmed the absence of additional variants of potential pathogenic relevance. Western blot analysis showed the absence of wild-type NF1 protein in the cartilage of the patient, consistent with a somatic double inactivation (SDI) of NF1. Taken together, our findings support the role of SDI in NF1-related PE, widening the spectrum of the pathophysiological mechanisms involved in NF1-related skeletal features.
Variants in the ACTG2 gene, encoding a protein crucial for correct enteric muscle contraction, have been found in patients affected with chronic intestinal pseudo‐obstruction, either congenital or ...late‐onset visceral myopathy, and megacystis‐microcolon‐intestinal hypoperistalsis syndrome. Here we report about ten pediatric and one adult patients, from nine families, carrying ACTG2 variants: four show novel still unpublished missense variants, including one that is apparently transmitted according to a recessive mode of inheritance. Four of the remaining five probands carry variants affecting arginine residues, that have already been associated with a severe phenotype. A de novo occurrence of the variants could be confirmed in six of these families. Since a genotype–phenotype correlation is affected by extrinsic factors, such as, diagnosis delay, quality of clinical management, and intra‐familial variability, we have undertaken 3D molecular modeling to get further insights into the effects of the variants here described. The present findings and further ACTG2 testing of patients presenting with intestinal pseudo‐obstruction, will improve our understanding of visceral myopathies, including implications in the prognosis and genetic counseling of this set of severe disorders.
Chronic Intestinal Pseudo‐Obstruction (CIPO):
wide phenotypic spectrum of severity
Different ACTG2 variants
AD or AR inheritance
Transmitted or de novo
Heterozygous polyalanine repeat expansions of PHOX2B have been associated with Congenital Central Hypoventilation Syndrome, a rare neurocristopathy characterized by absence of adequate control of ...respiration during sleep. Here we report a PHOX2B mutational screening in 63 CCHS patients, 58 of whom presenting with poly-A expansions or frameshift, missense and nonsense mutations. To assess a somatic or germline occurrence of poly-A length variations, the relative amounts of mutant and wild type alleles have been quantified in 20 selected CCHS patients presenting with an expansion, and in their parents. Somatic mosaicism was shown in four parents, while no mosaic was found among CCHS patients. Moreover, while co-segregation analysis of the PHOX2B poly-A expansions with selected marker alleles in the same 20 CCHS trios has not demonstrated any parent-of-origin effect of the mutations, it has provided further clues to clarify the molecular mechanism underlying the expansion occurrence. Finally, the segregation of PHOX2B poly-A anomalous tracts within family members has allowed us to exclude tendency of polymorphic variations towards expansion. This strengthens the notion that expanded polyalanine tracts, identified as frequent disease-causing mutations also in other human diseases, are mitotically and meiotically stable. Published 2007 Wiley-Liss, Inc.
To evaluate the rate of somatic
mosaicism in an Italian cohort of mutation-negative patients with cryopyrin-associated periodic syndrome (CAPS).
The study enrolled 14 patients with a clinical ...phenotype consistent with CAPS in whom Sanger sequencing of the
gene yielded negative results. Patients' DNA were subjected to amplicon-based
deep sequencing.
Low-level somatic
mosaicism has been detected in 4 patients, 3 affected with chronic infantile neurological cutaneous and articular syndrome and 1 with Muckle-Wells syndrome. Identified nucleotide substitutions encode for 4 different amino acid exchanges, with 2 of them being novel (p.Y563C and p.G564S).
functional studies confirmed the deleterious behavior of the 4 somatic NLRP3 mutations. Among the different neurological manifestations detected, 1 patient displayed mild loss of white matter volume on brain magnetic resonance imaging.
The allele frequency of somatic
mutations occurs generally under 15%, considered the threshold of detectability using the Sanger method of DNA sequencing. Consequently, routine genetic diagnostic of CAPS should be currently performed by next-generation techniques ensuring high coverage to identify also low-level mosaicism, whose actual frequency is yet unknown and probably underestimated.
Neurofibromatosis type 1 (NF1) is a proteiform genetic condition caused by pathogenic variants in
and characterized by a heterogeneous phenotypic presentation. Relevant genotype-phenotype ...correlations have recently emerged, but only few pertinent studies are available. We retrospectively reviewed clinical, instrumental, and genetic data from a cohort of 583 individuals meeting at least 1 diagnostic National Institutes of Health (NIH) criterion for NF1. Of these, 365 subjects fulfilled ≥2 NIH criteria, including 235 pediatric patients. Genetic testing was performed through cDNA-based sequencing, Next Generation Sequencing (NGS), and Multiplex Ligation-dependent Probe Amplification (MLPA). Uni- and multivariate statistical analysis was used to investigate genotype-phenotype correlations. Among patients fulfilling ≥ 2 NIH criteria, causative single nucleotide variants (SNVs) and copy number variations (CNVs) were detected in 267/365 (73.2%) and 20/365 (5.5%) cases. Missense variants negatively correlated with neurofibromas (
= 0.005). Skeletal abnormalities were associated with whole gene deletions (
= 0.05) and frameshift variants (
= 0.006). The c.3721C>T; p.(R1241*) variant positively correlated with structural brain alterations (
= 0.031), whereas Lisch nodules (
= 0.05) and endocrinological disorders (
= 0.043) were associated with the c.6855C>A; p.(Y2285*) variant. We identified novel NF1 genotype-phenotype correlations and provided an overview of known associations, supporting their potential relevance in the implementation of patient management.
Hirschsprung (HSCR) Associated Enterocolitis (HAEC) is a common life-threatening complication in HSCR. HAEC is suggested to be due to a loss of gut homeostasis caused by impairment of immune system, ...barrier defense, and microbiome, likely related to genetic causes. No gene has been claimed to contribute to HAEC occurrence, yet. Genetic investigation of HAEC by Whole-Exome Sequencing (WES) on 24 HSCR patients affected (HAEC) or not affected (HSCR-only) by enterocolitis and replication of results on a larger panel of patients allowed the identification of the HAEC susceptibility variant p.H187Q in the Oncostatin-M receptor (
) gene (14.6% in HAEC and 5.1% in HSCR-only,
= 0.0024). Proteomic analysis on the lymphoblastoid cell lines from one HAEC patient homozygote for this variant and one HAEC patient not carrying the variant revealed two well distinct clusters of proteins significantly up or downregulated upon OSM stimulation. A marked enrichment in immune response pathways (
< 0.0001) was shown in the HAEC H187 cell line, while proteins upregulated in the HAEC Q187 lymphoblasts sustained pathways likely involved in pathogen infection and inflammation. In conclusion,
p.H187Q is an HAEC susceptibility variant and perturbates the downstream signaling cascade necessary for the gut immune response and homeostasis maintenance.
Alexander disease is a leukodystrophy caused by heterozygous mutations of
gene. Recurrence in siblings from healthy parents provides a confirmation to the transmission of variants through germinal ...mosaicism. With the use of DNA isolated from peripheral blood, next-generation sequencing (NGS) of
locus was performed with deep coverage (≥500×) in 11 probands and their parents (trios) with probands heterozygous for apparently
mutations. Indeed, one parent had somatic mosaicism, estimated in the range of 8.9%-16%, for the mutant allele transmitted to the affected sibling. Parental germline mosaicism deserves attention, as it is critical in assessing the risk of recurrence in families with Alexander disease.
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