SUFU haploinsufficiency causes a recognisable neurodevelopmental phenotype at the mild end of the Joubert syndrome spectrum.

BACKGROUND: Joubert syndrome (JS) is a recessively inherited ciliopathy characterised by congenital ocular motor apraxia (COMA), developmental delay (DD), intellectual disability, ataxia, multiorgan involvement, and a unique cerebellar and brainstem malformation. Over 40 JS-associated genes are known with a diagnostic yield of 60%-75%.In 2018, we reported homozygous hypomorphic missense variants of the SUFU gene in two families with mild JS. Recently, heterozygous truncating SUFU variants were identified in families with dominantly inherited COMA, occasionally associated with mild DD and subtle cerebellar anomalies. METHODS: We reanalysed next generation sequencing (NGS) data in two cohorts comprising 1097 probands referred for genetic testing of JS genes. RESULTS: Heterozygous truncating and splice-site SUFU variants were detected in 22 patients from 17 families (1.5%) with strong male prevalence (86%), and in 8 asymptomatic parents. Patients presented with COMA, hypotonia, ataxia and mild DD, and only a third manifested intellectual disability of variable severity. Brain MRI showed consistent findings characterised by vermis hypoplasia, superior cerebellar dysplasia and subtle-to-mild abnormalities of the superior cerebellar peduncles. The same pattern was observed in two out of three tested asymptomatic parents. CONCLUSION: Heterozygous truncating or splice-site SUFU variants cause a novel neurodevelopmental syndrome encompassing COMA and mild JS, which likely represent overlapping entities. Variants can arise de novo or be inherited from a healthy parent, representing the first cause of JS with dominant inheritance and reduced penetrance. Awareness of this condition will increase the diagnostic yield of JS genetic testing, and allow appropriate counselling about prognosis, medical monitoring and recurrence risk.

Expanding the phenotype of Wiedemann-Steiner syndrome: Craniovertebral junction anomalies.

Wiedemann-Steiner syndrome (WDSTS) is a rare autosomal dominant condition caused by heterozygous loss of function variants in the KMT2A (MLL) gene, encoding a lysine N-methyltransferase that mediates a histone methylation pattern specific for epigenetic transcriptional activation. WDSTS is characterized by a distinctive facial phenotype, hypertrichosis, short stature, developmental delay, intellectual disability, congenital malformations, and skeletal anomalies. Recently, a few patients have been reported having abnormal skeletal development of the cervical spine. Here we describe 11 such individuals, all with KMT2A de novo loss-of-function variants: 10 showed craniovertebral junction anomalies, while an 11th patient had a cervical abnormality in C7. By evaluating clinical and diagnostic imaging data we characterized these anomalies, which consist primarily of fused cervical vertebrae, C1 and C2 abnormalities, small foramen magnum and Chiari malformation type I. Craniovertebral anomalies in WDSTS patients have been largely disregarded so far, but the increasing number of reports suggests that they may be an intrinsic feature of this syndrome. Specific investigation strategies should be considered for early identification and prevention of craniovertebral junction complications in WDSTS patients.

Expanding the clinical phenotype of individuals with a 3-bp in-frame deletion of the NF1 gene (c.2970_2972del): an update of genotype-phenotype correlation.

PURPOSE: Neurofibromatosis type 1 (NF1) is characterized by a highly variable clinical presentation, but almost all NF1-affected adults present with cutaneous and/or subcutaneous neurofibromas. Exceptions are individuals heterozygous for the NF1 in-frame deletion, c.2970_2972del (p.Met992del), associated with a mild phenotype without any externally visible tumors. METHODS: A total of 135 individuals from 103 unrelated families, all carrying the constitutional NF1 p.Met992del pathogenic variant and clinically assessed using the same standardized phenotypic checklist form, were included in this study. RESULTS: None of the individuals had externally visible plexiform or histopathologically confirmed cutaneous or subcutaneous neurofibromas. We did not identify any complications, such as symptomatic optic pathway gliomas (OPGs) or symptomatic spinal neurofibromas; however, 4.8% of individuals had nonoptic brain tumors, mostly low-grade and asymptomatic, and 38.8% had cognitive impairment/learning disabilities. In an individual with the NF1 constitutional c.2970_2972del and three astrocytomas, we provided proof that all were NF1-associated tumors given loss of heterozygosity at three intragenic NF1 microsatellite markers and c.2970_2972del. CONCLUSION: We demonstrate that individuals with the NF1 p.Met992del pathogenic variant have a mild NF1 phenotype lacking clinically suspected plexiform, cutaneous, or subcutaneous neurofibromas. However, learning difficulties are clearly part of the phenotypic presentation in these individuals and will require specialized care.

Refining the phenotype associated with GNB1 mutations: Clinical data on 18 newly identified patients and review of the literature.

De novo germline mutations in GNB1 have been associated with a neurodevelopmental phenotype. To date, 28 patients with variants classified as pathogenic have been reported. We add 18 patients with de novo mutations to this cohort, including a patient with mosaicism for a GNB1 mutation who presented with a milder phenotype. Consistent with previous reports, developmental delay in these patients was moderate to severe, and more than half of the patients were non-ambulatory and nonverbal. The most observed substitution affects the p.Ile80 residue encoded in exon 6, with 28% of patients carrying a variant at this residue. Dystonia and growth delay were observed more frequently in patients carrying variants in this residue, suggesting a potential genotype-phenotype correlation. In the new cohort of 18 patients, 50% of males had genitourinary anomalies and 61% of patients had gastrointestinal anomalies, suggesting a possible association of these findings with variants in GNB1. In addition, cutaneous mastocytosis, reported once before in a patient with a GNB1 variant, was observed in three additional patients, providing further evidence for an association to GNB1. We will review clinical and molecular data of these new cases and all previously reported cases to further define the phenotype and establish possible genotype-phenotype correlations.

Genotype-Phenotype Correlation in NF1: Evidence for a More Severe Phenotype Associated with Missense Mutations Affecting NF1 Codons 844-848.

Neurofibromatosis type 1 (NF1), a common genetic disorder with a birth incidence of 1:2,000-3,000, is characterized by a highly variable clinical presentation. To date, only two clinically relevant intragenic genotype-phenotype correlations have been reported for NF1 missense mutations affecting p.Arg1809 and a single amino acid deletion p.Met922del. Both variants predispose to a distinct mild NF1 phenotype with neither externally visible cutaneous/plexiform neurofibromas nor other tumors. Here, we report 162 individuals (129 unrelated probands and 33 affected relatives) heterozygous for a constitutional missense mutation affecting one of five neighboring NF1 codons-Leu844, Cys845, Ala846, Leu847, and Gly848-located in the cysteine-serine-rich domain (CSRD). Collectively, these recurrent missense mutations affect ∼0.8% of unrelated NF1 mutation-positive probands in the University of Alabama at Birmingham (UAB) cohort. Major superficial plexiform neurofibromas and symptomatic spinal neurofibromas were more prevalent in these individuals compared with classic NF1-affected cohorts (both p < 0.0001). Nearly half of the individuals had symptomatic or asymptomatic optic pathway gliomas and/or skeletal abnormalities. Additionally, variants in this region seem to confer a high predisposition to develop malignancies compared with the general NF1-affected population (p = 0.0061). Our results demonstrate that these NF1 missense mutations, although located outside the GAP-related domain, may be an important risk factor for a severe presentation. A genotype-phenotype correlation at the NF1 region 844-848 exists and will be valuable in the management and genetic counseling of a significant number of individuals.

A patient with germ-line gain-of-function PDGFRB p.N666H mutation and marked clinical response to imatinib.

PurposeHeterozygous germ-line activating mutations in PDGFRB cause Kosaki and Penttinen syndromes and myofibromatosis. We describe a 10-year-old child with a germ-line PDGFRB p.N666H mutation who responded to the tyrosine kinase inhibitor imatinib by inhibition of PDGFRB.MethodsThe impact of p.N666H on PDGFRB function and sensitivity to imatinib was studied in cell culture.ResultsCells expressing the p.N666H mutation showed constitutive PDGFRB tyrosine phosphorylation. PDGF-independent proliferation was abolished by imatinib at 1 µM concentration. Patient fibroblasts showed constitutive receptor tyrosine phosphorylation that was also abrogated by imatinib with reduced proliferation of treated cells.This led to patient treatment with imatinib at 400 mg daily (340 mg/m2) for a year with objective improvement of debilitating hand and foot contractures, reduced facial coarseness, and significant improvement in quality of life. New small subcutaneous nodules developed, but remained stable. Transient leukopenia, neutropenia, and fatigue resolved without intervention; however, mildly decreased growth velocity resulted in reducing imatinib dose to 200 mg daily (170 mg/m2). The patient continues treatment with ongoing clinical response.ConclusionTo our knowledge, this is one of the first personalized treatments of a congenital disorder caused by a germ-line PDGF receptor mutation with a PDGFRB inhibitor.

Retinal dystrophy in two boys with Costello syndrome due to the HRAS p.Gly13Cys mutation.

Features of Costello Syndrome, a systemic disorder caused by germline mutations in the proto-oncogene HRAS from the RAS/MAPK pathway, include failure-to-thrive, short stature, coarse facial features, cardiac defects including hypertrophic cardiomyopathy, intellectual disability, and predisposition to neoplasia. Two unrelated boys with Costello syndrome and an HRAS mutation (p.Gly13Cys) are presented with their ophthalmologic findings. Both had early symptoms of nystagmus, photophobia, and vision abnormalities. Fundus examination findings of retinal dystrophy were present at age 3 years. Both boys have abnormal electroretinograms with reduced or undetectable rod responses along with reduced cone responses consistent with rod-cone dystrophy. Our observations suggest that early ophthalmic examination and re-evaluations are indicated in children with Costello syndrome.

MKS1 regulates ciliary INPP5E levels in Joubert syndrome.

BACKGROUND: Joubert syndrome (JS) is a recessive ciliopathy characterised by a distinctive brain malformation 'the molar tooth sign'. Mutations in >27 genes cause JS, and mutations in 12 of these genes also cause Meckel-Gruber syndrome (MKS). The goals of this work are to describe the clinical features of MKS1-related JS and determine whether disease causing MKS1 mutations affect cellular phenotypes such as cilium number, length and protein content as potential mechanisms underlying JS. METHODS: We measured cilium number, length and protein content (ARL13B and INPP5E) by immunofluorescence in fibroblasts from individuals with MKS1-related JS and in a three-dimensional (3D) spheroid rescue assay to test the effects of disease-related MKS1 mutations. RESULTS: We report MKS1 mutations (eight of them previously unreported) in nine individuals with JS. A minority of the individuals with MKS1-related JS have MKS features. In contrast to the truncating mutations associated with MKS, all of the individuals with MKS1-related JS carry ≥ 1 non-truncating mutation. Fibroblasts from individuals with MKS1-related JS make normal or fewer cilia than control fibroblasts, their cilia are more variable in length than controls, and show decreased ciliary ARL13B and INPP5E. Additionally, MKS1 mutant alleles have similar effects in 3D spheroids. CONCLUSIONS: MKS1 functions in the transition zone at the base of the cilium to regulate ciliary INPP5E content, through an ARL13B-dependent mechanism. Mutations in INPP5E also cause JS, so our findings in patient fibroblasts support the notion that loss of INPP5E function, due to either mutation or mislocalisation, is a key mechanism underlying JS, downstream of MKS1 and ARL13B.

Overcoming the barriers to diagnosis of Morquio A syndrome.

BACKGROUND: Morquio A syndrome is an autosomal recessive lysosomal storage disease often resulting in life-threatening complications. Early recognition and proficient diagnosis is imperative to facilitate prompt treatment and prevention of clinical complications. METHODS: Experts in Asia Pacific reviewed medical records focusing on presenting signs and symptoms leading to a diagnosis of Morquio A syndrome. RESULTS: Eighteen patients (77% female) had a mean (median; min, max) age of 77.1 (42.0; 0.0, 540.0) months at symptom onset, 78.9 (42.0; 4.5, 540.0) months at presentation and 113.8 (60.0; 7.0, 540.0) months at diagnosis. Orthopedic surgeons and pediatricians were most frequently consulted pre-diagnosis while clinical geneticists/metabolic specialists most frequently made the diagnosis. Delayed diagnoses were due to atypical symptoms for 5 patients (28%), while 4 patients (22%) experienced each of subtle symptoms, symptoms commonly associated with other diseases, or false-negative urine glycosaminoglycan analysis. Two patients (11%) each experienced overgrowth within the first year of life. Two patients with Morquio A syndrome (11%) were diagnosed with craniosynostosis and 1 (6%) for each of Legg-Calvé-Perthes disease, Leri-Weill syndrome, and pseudoachondroplasia. Early radiographic features of Morquio A syndrome led to more efficient diagnosis. CONCLUSIONS: Increased awareness of clinical symptomology overlapping with Morquio A syndrome is essential. Clinicians encountering patients with certain skeletal dysplasia should consider Morquio A syndrome in their differential diagnosis. Atypical or subtle symptoms should not eliminate Morquio A syndrome from the differential diagnosis, especially for patients who may have non-classical phenotype of Morquio A syndrome.

Mucopolysaccharidosis IVA (Morquio A syndrome) and VI (Maroteaux-Lamy syndrome): under-recognized and challenging to diagnose.

OBJECTIVE: Mucopolysaccharidosis IVA (MPS IVA, or Morquio A syndrome) and VI (MPS VI, or Maroteaux-Lamy syndrome) are autosomal recessive lysosomal storage disorders. Skeletal abnormalities are common initial presenting symptoms and, when recognized early, may facilitate timely diagnosis and intervention, leading to improved patient outcomes. Patients with slowly progressing disease and nonclassic phenotypes can be particularly challenging to diagnose. The objective was to describe the radiographic features of patients with a delayed diagnosis of MPS IVA or VI. MATERIALS AND METHODS: This was a retrospective study. The records of 5 MPS IVA and 3 MPS VI patients with delayed diagnosis were reviewed. Radiographs were evaluated by a radiologist with special expertise in skeletal dysplasias. RESULTS: An important common theme in these cases was the appearance of multiple epiphyseal dysplasia (MED) with epiphyseal changes seemingly confined to the capital (proximal) femoral epiphyses. Very few patients had the skeletal features of classical dysostosis multiplex. CONCLUSIONS: Radiologists should appreciate the wide phenotypic variability of MPS IVA and VI. The cases presented here illustrate the importance of considering MPS in the differential diagnosis of certain skeletal dysplasias/disorders, including MED, some forms of spondylo-epiphyseal dysplasia (SED), and bilateral Perthes-like disease. It is important to combine radiographic findings with clinical information to facilitate early testing and accurate diagnosis.

Spondyloepiphyseal dysplasias and bilateral legg-calvé-perthes disease: diagnostic considerations for mucopolysaccharidoses.

Mucopolysaccharidosis type VI (MPS VI, Maroteaux-Lamy syndrome, MIM 253200 ) is an autosomal recessive lysosomal storage disease (LSD) caused by decreased activity of arylsulfatase B (N-acetylgalactosamine 4-sulfatase) enzyme resulting in dermatan sulfate accumulation; mucopolysaccharidosis type IVA (MPS IVA, Morquio syndrome A, MIM 253000 ) by decreased activity of N-acetylgalactosamine 6-sulfatase enzyme resulting in accumulation of keratan sulfate. Clinical symptoms include coarse facial features, joint stiffness, hepatosplenomegaly, hip osteonecrosis, and dysostosis multiplex. MPS IVA symptoms are similar but with joint hypermobility.With suspicion of MPS disease, clinicians request urine studies for quantitative and qualitative glycosaminoglycans (GAGs). Diagnosis is confirmed by decreased enzyme activity in leukocytes or cultured skin fibroblasts. Further confirmation is obtained with identification of two mutations in the ARSB gene for MPS VI or mutations in the GALNS gene for MPS IVA.We report slowly progressing patients, one with MPS VI and two with MPS IVA, who presented with skeletal changes and hip findings resembling Legg-Calvé-Perthes disease or spondyloepiphyseal dysplasia and normal/near normal urine GAG levels. The urine analysis data presented suggest that present screening techniques for MPS are inadequate in milder patients and result in delayed or missed diagnoses. The patients presented in this paper emphasize the importance of enzymatic and molecular testing.

WNT1 mutations in families affected by moderately severe and progressive recessive osteogenesis imperfecta.

Osteogenesis imperfecta (OI) is a heritable disorder that ranges in severity from death in the perinatal period to an increased lifetime risk of fracture. Mutations in COL1A1 and COL1A2, which encode the chains of type I procollagen, result in dominant forms of OI, and mutations in several other genes result in recessive forms of OI. Here, we describe four recessive-OI-affected families in which we identified causative mutations in wingless-type MMTV integration site family 1 (WNT1). In family 1, we identified a homozygous missense mutation by exome sequencing. In family 2, we identified a homozygous nonsense mutation predicted to produce truncated WNT1. In family 3, we found a nonsense mutation and a single-nucleotide duplication on different alleles, and in family 4, we found a homozygous 14 bp deletion. The mutations in families 3 and 4 are predicted to result in nonsense-mediated mRNA decay and the absence of WNT1. WNT1 is a secreted signaling protein that binds the frizzled receptor (FZD) and the coreceptor low-density lipoprotein-receptor-related protein 5 (LRP5). Biallelic loss-of-function mutations in LRP5 result in recessive osteoporosis-pseudoglioma syndrome with low bone mass, whereas heterozygous gain-of-function mutations result in van Buchem disease with elevated bone density. Biallelic loss-of-function mutations in WNT1 result in a recessive clinical picture that includes bone fragility with a moderately severe and progressive presentation that is not easily distinguished from dominant OI type III.

Genotype-phenotype analysis of the branchio-oculo-facial syndrome.

Branchio-oculo-facial syndrome (BOFS; OMIM#113620) is a rare autosomal dominant craniofacial disorder with variable expression. Major features include cutaneous and ocular abnormalities, characteristic facies, renal, ectodermal, and temporal bone anomalies. Having determined that mutations involving TFAP2A result in BOFS, we studied a total of 30 families (41 affected individuals); 26/30 (87%) fulfilled our cardinal diagnostic criteria. The original family with the 3.2 Mb deletion including the TFAP2A gene remains the only BOFS family without the typical CL/P and the only family with a deletion. We have identified a hotspot region in the highly conserved exons 4 and 5 of TFAP2A that harbors missense mutations in 27/30 (90%) families. Several of these mutations are recurrent. Mosaicism was detected in one family. To date, genetic heterogeneity has not been observed. Although the cardinal criteria for BOFS have been based on the presence of each of the core defects, an affected family member or thymic remnant, we documented TFAP2A mutations in three (10%) probands in our series without a classic cervical cutaneous defect or ectopic thymus. Temporal bone anomalies were identified in 3/5 patients investigated. The occurrence of CL/P, premature graying, coloboma, heterochromia irides, and ectopic thymus, are evidence for BOFS as a neurocristopathy. Intrafamilial clinical variability can be marked. Although there does not appear to be mutation-specific genotype-phenotype correlations at this time, more patients need to be studied. Clinical testing for TFAP2A mutations is now available and will assist geneticists in confirming the typical cases or excluding the diagnosis in atypical cases.

Importance of surgical history in diagnosing mucopolysaccharidosis type II (Hunter syndrome): data from the Hunter Outcome Survey.

PURPOSE: To characterize surgical histories typical of patients with mucopolysaccharidosis type II, thereby broadening understanding of the natural history of these patients and helping physicians recognize the disease. METHODS: Data on surgical interventions from the Hunter Outcome Survey--a multinational, observational database of patients with mucopolysaccharidosis type II-were analyzed. The study population comprised 527 patients for whom surgical data were reported on/before July 23, 2009. RESULTS: Surgical interventions were performed in 83.7% of the study population. Patients underwent their first operation at a median age of 2.6 years. Tympanostomies, repairs of inguinal hernias, and operations for carpal tunnel syndrome were performed in a greater proportion of the study population than the general population. A median of 3.0 operations was performed per patient; repeat operations for hernia or carpal tunnel syndrome were common. The majority of patients (221/389) underwent at least one surgical intervention before diagnosis of mucopolysaccharidosis type II. CONCLUSION: Patients with mucopolysaccharidosis type II typically undergo surgical intervention at a young age, often before diagnosis. Repeated early surgical interventions, particularly for hernias or carpal tunnel syndrome, are characteristic of patients with mucopolysaccharidosis type II. We recommend that such patients are carefully examined for manifestations of mucopolysaccharidosis disorders and referred for diagnostic testing.

Effects of germline mutations in the Ras/MAPK signaling pathway on adaptive behavior: cardiofaciocutaneous syndrome and Noonan syndrome.

Cardiofaciocutaneous syndrome (CFC) and Noonan syndrome (NS) are two phenotypically overlapping genetic disorders whose underlying molecular etiologies affect a common signaling pathway. Mutations in the BRAF, MEK1, and MEK2 genes cause most cases of CFC and mutations in PTPN11, SOS1, KRAS, and RAF1 typically cause NS. Although both syndromes are associated with developmental delays of varying severity, the extent to which the behavioral profiles differ may shed light on the different roles these respective genes play in development of skills necessary for everyday functioning. In this study, profiles of adaptive behavior of individuals with CFC and NS who had confirmed pathogenic mutations in Ras/mitogen-activated protein kinase (MAPK) pathway genes were investigated. Patterns of strengths and weaknesses, age-related differences, and risk factors for difficulties in adaptive skills were assessed. Although genes acting more downstream in the Ras/MAPK pathway were associated with more difficulties in adaptive functioning than genes more upstream in the pathway, several inconsistencies highlight the wide spectrum of possible developmental courses in CFC and NS. Along with clinical and genetic factors, variables such as chronological age, gestational age at birth, and parental education levels accounted for significant variance in adaptive skills. Results indicate that there is wide heterogeneity in adaptive functioning in CFC and NS, but that these abilities are correlated to some extent with the specific disease-causing genes.

Sectoral iris heterochromia and retinal pigment variation in 13q-syndrome.

Chromosome 13q deletion syndrome is characterized by growth retardation, cognitive delays, and organ and musculoskeletal deformities. Typical ocular associations include retinoblastoma, microphthalmia, and colobomas. We report a case of bilateral iris heterochromia and retinal pigment abnormalities in a child with 13q-syndrome.

Molecular and clinical analyses of Greig cephalopolysyndactyly and Pallister-Hall syndromes: robust phenotype prediction from the type and position of GLI3 mutations.

Mutations in the GLI3 zinc-finger transcription factor gene cause Greig cephalopolysyndactyly syndrome (GCPS) and Pallister-Hall syndrome (PHS), which are variable but distinct clinical entities. We hypothesized that GLI3 mutations that predict a truncated functional repressor protein cause PHS and that functional haploinsufficiency of GLI3 causes GCPS. To test these hypotheses, we screened patients with PHS and GCPS for GLI3 mutations. The patient group consisted of 135 individuals: 89 patients with GCPS and 46 patients with PHS. We detected 47 pathological mutations (among 60 probands); when these were combined with previously published mutations, two genotype-phenotype correlations were evident. First, GCPS was caused by many types of alterations, including translocations, large deletions, exonic deletions and duplications, small in-frame deletions, and missense, frameshift/nonsense, and splicing mutations. In contrast, PHS was caused only by frameshift/nonsense and splicing mutations. Second, among the frameshift/nonsense mutations, there was a clear genotype-phenotype correlation. Mutations in the first third of the gene (from open reading frame [ORF] nucleotides [nt] 1-1997) caused GCPS, and mutations in the second third of the gene (from ORF nt 1998-3481) caused primarily PHS. Surprisingly, there were 12 mutations in patients with GCPS in the 3' third of the gene (after ORF nt 3481), and no patients with PHS had mutations in this region. These results demonstrate a robust correlation of genotype and phenotype for GLI3 mutations and strongly support the hypothesis that these two allelic disorders have distinct modes of pathogenesis.