Pathogenic neurofibromatosis type 1 (NF1) RNA splicing resolved by targeted RNAseq.

Neurofibromatosis type 1 (NF1) is caused by loss-of-function variants in the NF1 gene. Approximately 10% of these variants affect RNA splicing and are either missed by conventional DNA diagnostics or are misinterpreted by in silico splicing predictions. Therefore, a targeted RNAseq-based approach was designed to detect pathogenic RNA splicing and associated pathogenic DNA variants. For this method RNA was extracted from lymphocytes, followed by targeted RNAseq. Next, an in-house developed tool (QURNAs) was used to calculate the enrichment score (ERS) for each splicing event. This method was thoroughly tested using two different patient cohorts with known pathogenic splice-variants in NF1. In both cohorts all 56 normal reference transcript exon splice junctions, 24 previously described and 45 novel non-reference splicing events were detected. Additionally, all expected pathogenic splice-variants were detected. Eleven patients with NF1 symptoms were subsequently tested, three of which have a known NF1 DNA variant with a putative effect on RNA splicing. This effect could be confirmed for all 3. The other eight patients were previously without any molecular confirmation of their NF1-diagnosis. A deep-intronic pathogenic splice variant could now be identified for two of them (25%). These results suggest that targeted RNAseq can be successfully used to detect pathogenic RNA splicing variants in NF1.

Surgical management for isolated macrodactyly in an adult PIK3CA mutant.

Isolated macrodactyly in adults caused by mosaic pattern PIK3CA mutation can result in significant functional impairment and psychological burden. Due to the rarity of this condition there are no clear treatment guidelines, and those few available are focused on paediatric cases. Reports on surgical management of isolated macrodactyly in adults are lacking. We present here the surgical management through partial amputation of enlarged rays of the right hand in an individual affected by low-grade mosaic PIK3CA mutation.

Schnitzler's syndrome - a novel hypothesis of a shared pathophysiologic mechanism with Waldenström's disease.

Schnitzler's syndrome is an auto-inflammatory disorder which is characterized by two mandatory features: an urticarial rash and a monoclonal gammopathy. Although the pathophysiology of this syndrome is not yet fully understood, a role for interleukin-1 seems apparent. While this presumed link between interleukin-1 and the monoclonal gammopathy is not yet elucidated, a mutual factor in pathophysiology however seems likely. Here we present a novel hypothesis of a shared pathophysiologic mechanism between Schitzler's syndrome and monoclonal gammopathy.

Genetic profiling of basal cell carcinomas detects postzygotic mosaicism in basal cell naevus syndrome.

Basal cell naevus syndrome (BCNS) is associated with germline mutations in the PTCH1 gene. Postzygotic mosaicism can also cause BCNS. Here we describe two patients, one with multiple basal cell carcinomas (BCCs) and one with clinical BCNS, who had no PTCH1 mutation in DNA extracted from blood. In both patients, we performed genetic analysis on different BCCs, revealing the presence of a shared PTCH1 mutation in all tumours. Our findings show that in patients with symptoms of BCNS and initial absence of a PTCH1 mutation in blood, genetic profiling of BCCs can detect postzygotic mosaicism. What's already known about this topic? Basal cell naevus syndrome (BCNS) is associated with germline mutations in the PTCH1 gene, but it can also be caused by low-grade postzygotic mosaicism in PTCH1. What does this study add? In patients suspected of having BCNS or patients with multiple basal cell carcinomas (BCCs) with a special distribution on the body and no mutation detected in blood, it is worthwhile to search for a shared PTCH1 mutation in their BCCs as this can detect postzygotic mosaicism. This information is important to ensure proper surveillance programmes, choose the right therapy and provide adequate genetic counselling.

Okur-Chung neurodevelopmental syndrome: Eight additional cases with implications on phenotype and genotype expansion.

Okur-Chung syndrome is a neurodevelopmental condition attributed to germline CSNK2A1 pathogenic missense variants. We present 8 unreported subjects with the above syndrome, who have recognizable dysmorphism, varying degrees of developmental delay and multisystem involvement. Together with 6 previously reported cases, we present a case series of 7 female and 7 male subjects, highlighting the recognizable facial features of the syndrome (microcephaly, hypertelorism, epicanthic fold, ptosis, arched eyebrows, low set ears, ear fold abnormality, broad nasal bridge and round face) as well as frequently occurring clinical features including neurodevelopmental delay (93%), gastrointestinal (57%), musculoskeletal (57%) and immunological (43%) abnormalities. The variants reported in this study are evolutionary conserved and absent in the normal population. We observed that the CSNK2A1 gene is relatively intolerant to missense genetic changes, and most variants are within the protein kinase domain. All except 1 variant reported in this cohort are spatially located on the binding pocket of the holoenzyme. We further provide key recommendations on the management of Okur-Chung syndrome. To conclude, this is the second case series on Okur-Chung syndrome, and an in-depth review of the phenotypic features and genomic findings of the condition with suggestions on clinical management.

Postzygotic mosaicism in basal cell naevus syndrome.

Basal cell naevus syndrome (BCNS) is an autosomal dominant disorder most commonly caused by a germline mutation in the Drosophila homologue of patched-1 gene (PTCH1). Here we describe a patient with clinical signs of BCNS, caused by postzygotic mosaicism of a PTCH1 mutation. We performed restriction fragment length polymorphism analysis and Droplet Digital polymerase chain reaction to determine the degree of mosaicism in different tissues of this patient. Our case shows that a relatively low-grade mosaicism can lead to clinical signs reminiscent of those caused by a germline mutation. This finding has important implications for genetic counselling and therefore is pivotal to recognize for dermatologists, as well as for clinical geneticists and clinical laboratory geneticists.

What are we waiting for? Factors influencing completion times in an academic and peripheral emergency department.

BACKGROUND: A long completion time in the Emergency Department (ED) is associated with higher morbidity and in-hospital mortality. A completion time of more than four hours is a frequently used cut-off point. Mostly, older and sicker patients exceed a completion time of four hours on the ED. The primary aim was to examine which factors currently contribute to overcrowding and a time to completion of more than four hours on the EDs of two different hospitals, namely: the VU Medical Center (VUmc), an academic level 1 trauma centre and the St. Antonius Hospital, a large community hospital in Nieuwegein. In addition, we compared the differences between these hospitals. METHODS: In this observational study, the time steps in the process of diagnosing and treatment of all patients visiting the EDs of the two hospitals were measured for four weeks. Patients triaged as Emergency Severity Index (ESI) category 2/3 or Manchester Triage System (MTS) orange/yellow were followed more closely and prospectively by researchers for detailed information in the same period from 12.00-23.00 hrs. RESULTS: In the VUmc, 89% of the patients had a completion time of less than four hours. The average completion time (n = 2262) was 2:10 hours, (median 1:51 hours, range: 0:05-12:08). In the St. Antonius Hospital, 77% of patients had a completion time shorter than four hours (n = 1656). The average completion time in hours was 2:49 (n = 1655, median 2:34, range: 0:08-11:04). In the VUmc, a larger percentage of ESI 1, 2 and 3 patients did not achieve the 4-hour target (14%, 20% and 19%) compared with ESI 4 and 5 patients (2.7% and 0%), p < 0.001. At the St. Antonius Hospital, a greater percentage of orange and yellow categorised patients exceeded four hours on the ED (32% and 28%) compared with red (8%) and green/blue (13%), p < 0.001. For both hospitals there was a significant dependency between exceeding four hours on the ED and the following: whether a consultation was performed (p < 0.001), the number of radiology tests performed (p < 0.001), and an age above 65 years. CONCLUSION: Factors leading to ED stagnation were similar in both hospitals, namely old age, treatment by more than one speciality and undergoing radiological tests. Uniform remedial measures should be taken on a nationwide level to deal with these factors to reduce stagnation in the EDs.

Cutaneous clues for diagnosing X-chromosomal disorders.

In a multidisciplinary outpatient clinic for hereditary skin diseases and/or syndromes involving the skin, 7% (30 of 409) of patients were found to have an abnormality involving the X chromosome, a mutation in a gene located on the X chromosome or a clinical diagnosis of an X-linked monogenetic condition. The collaboration of a dermatologist and a clinical geneticist proves to be very valuable in recognizing and diagnosing these conditions. By combining their specific expertize in counselling an individual patient, X-linked diagnoses were recognized and could be confirmed by molecular and/or cytogenetic studies in 24 of 30 cases. Mosaicism plays an important role in many X-linked hereditary skin disorders. From our experience, we extracted clinical clues for specialists working in the field of genetics and/or dermatology for considering X-linked disorders involving the skin.

Genodermatoses caused by genetic mosaicism.

Genetic mosaicism is defined as the existence of at least two genetically distinct cell populations within one individual. Mosaic presentation of genetic disorders is common and is often particularly obvious in the skin, because there it will generate recognizable patterns. Recognizing those can frequently assist in establishing a diagnosis. In this review, we discuss the mechanisms that give rise to genetic mosaicism. We describe its most frequent cutaneous manifestations that are relevant to paediatric practice. While most mosaic genetic diseases are rare, it is important to recognize them so that patients and parents may receive appropriate genetic counselling. Moreover, recent developments are now resulting in novel, targeted treatments for such disorders that promise to considerably improve patients' lives.

Focal dermal hypoplasia in a male patient due to mosaicism for a novel PORCN single nucleotide deletion.

BACKGROUND: Focal dermal hypoplasia (FDH) is an X-linked dominant disorder caused by nonsense mutations and deletions in the PORCN gene coding for a transmembrane endoplasmic reticulum protein required for Wingless signalling. Symptoms consist mainly of linear atrophic skin defects, skeletal deformities and, in many cases, mental retardation. Osteopathia striata is a nearly constant feature. Approximately 90% of patients are women. A few instances of father-to-daughter transmission and a number of sporadic male cases presumably as a result of somatic mosaicism have been recorded. OBJECTIVES: The aim of this study was to demonstrate the presence of somatic mosaicism for PORCN mutations in a male patient. METHODS: We sequenced the PORCN gene in different tissues from a boy with symptoms of FDH. RESULTS: We demonstrate post-zygotic mosaicism for a novel deletion in the PORCN gene. CONCLUSIONS: A novel PORCN deletion, present in a post-zygotic mosaic, causes focal dermal hyplasia in a male patient.

Contiguous gene syndrome due to a maternally inherited 8.41 Mb distal deletion of chromosome band Xp22.3 in a boy with short stature, ichthyosis, epilepsy, mental retardation, cerebral cortical heterotopias and Dandy-Walker malformation.

Microdeletions of Xp22.3 are associated with contiguous gene syndromes, the extent and nature of which depend on the genes encompassed by the deletion. Common symptoms include ichthyosis, mental retardation and hypogonadism. We report on a boy with short stature, ichthyosis, severe mental retardation, cortical heterotopias and Dandy-Walker malformation. The latter two abnormalities have so far not been reported in terminal Xp deletions. MLPA showed deletion of SHOX and subsequent analysis using FISH and SNP-arrays revealed that the patient had an 8.41 Mb distal deletion of chromosome region Xp22.31 --> Xpter. This interval contains several genes whose deletion can partly explain our patient's phenotype. His cortical heterotopias and DWM suggest that a gene involved in brain development may be in the deleted interval, but we found no immediately obvious candidates. Interestingly, further analysis of the family revealed that the patient had inherited his deletion from his mother, who has a mos 46,X,del(X)(p22)/45,X/46, XX karyotype.

Granulomatous rosacea and Crohn's disease in a patient homozygous for the Crohn-associated NOD2/CARD15 polymorphism R702W.

NOD2/CARD15 belongs to the N-terminal caspase recruitment domain family of proteins involved in regulating NF-kB activation in response to inflammatory stimuli transduced through Toll-like receptors. Mutations and polymorphisms in the NOD2/CARD15 gene reduce antibacterial responses and are associated with granulomatous inflammatory conditions such as Blau syndrome and early-onset sarcoidosis. The polymorphism R702W (arginine to tryptophan) is strongly associated with susceptibility to Crohn's disease in Caucasian populations. Skin abnormalities (other than cutaneous manifestations of Crohn's disease) have not been previously associated with R702W. We report on a female patient homozygous for R702W who developed granulomatous rosacea at the age of 12 years old. From the occurrence in the context of Crohn associated with R702W, we speculate that granulomatous rosacea may be an entity distinct from other forms of rosacea, which are associated with increased production of antibacterial proteins such as cathelicidin.

Novel EBP gene mutations in Conradi-Hünermann-Happle syndrome.

BACKGROUND: Conradi-Hünermann-Happle syndrome [X-linked dominant chondrodysplasia punctata type 2 (CDPX2); MIM no. 302960] is an X-linked dominant disorder of cholesterol metabolism that causes a wide spectrum of skeletal abnormalities and linear ichthyosiform skin lesions. Mosaicism is probably responsible for the variability of the phenotype. OBJECTIVES: To describe new mutations in patients with variable manifestations of the disease. METHODS: We studied three patients with CDPX2. We performed mutation analysis of the EBP (formerly known as CDPX2) gene and gas chromatography-mass spectroscopy on serum of two patients. RESULTS: We found two novel (3G-->T and 419-422delTTCT) and one known mutation in the EBP gene. We demonstrated the presence of increased levels of dehydrocholesterol and 8(9)-cholestenol in the two patients with new mutations, confirming the diagnosis of CDPX2 and strongly suggesting that the mutations are indeed pathogenic. One patient had a very mild phenotype, presenting with linear alopecia and a mild symmetrical epiphyseal dysplasia. X-inactivation studies in peripheral blood of all patients showed skewing in only the most severely affected patient. CONCLUSIONS: The strong phenotypic variability in our patients suggests that there is no clear genotype-phenotype correlation.

Skin changes in oculo-dento-digital dysplasia are correlated with C-terminal truncations of connexin 43.

Oculo-dento-digital dysplasia (ODDD, OMIM no.164210) is a pleiotropic disorder caused by mutations in the GJA1 gene that codes for the gap junction protein connexin 43. While the gene is highly expressed in skin, ODDD is usually not associated with skin symptoms. We recently described a family with ODDD and palmoplantar keratoderma. Interestingly, mutation carriers had a novel dinucleotide deletion in the GJA1 gene that resulted in truncation of part of the C-terminus. We speculated, that truncation of the C-terminus may be uniquely associated with skin disease in ODDD. Here, we describe a patient with ODDD and palmar hyperkeratosis caused by a novel dinucleotide deletion that truncates most of the connexin 43 C-terminus. Thus, our findings support the notion that such mutations are associated with the occurrence of skin symptoms in ODDD and provide the first evidence for the existence of a genotype-phenotype correlation.

Recurring HRAS mutation G12S in Dutch patients with Costello syndrome.

Costello syndrome (CS) is a rare multiple congenital anomaly/mental retardation syndrome characterized by coarse face, loose skin and cardiomyopathy. It is often associated with benign and malignant tumors. Several groups have now demonstrated that CS is caused by recurring mutations in the HRAS gene in different ethnic groups. Here, we describe three unrelated Dutch patients and show that they all have the same mutation, G12S, in HRAS. To our knowledge, our patients are the first Dutch to be analysed. The syndrome seems to be genetically homogeneous. We discuss the pertinent nosology of the syndrome.

Myhre syndrome in a female with previously undescribed symptoms: further delineation of the phenotype.

Myhre syndrome is a rare connective tissue disease characterized by nonprogressive stiffness of the large joints, short stature with a peculiar build, and a distinctive facial phenotype. Developmental delay is common. Three female patients have so far been described. Here, we report on a 16-year-old female with Myhre syndrome. She has vertebral defects, hypertrophic scar formation, and a stiff skin in addition to the features that have previously been reported in association with Myhre syndrome.