Mutational Spectrum of the ABCA12 Gene and Genotype-Phenotype Correlation in a Cohort of 64 Patients with Autosomal Recessive Congenital Ichthyosis.

Autosomal recessive congenital ichthyosis (ARCI) is a non-syndromic congenital disorder of cornification characterized by abnormal scaling of the skin. The three major phenotypes are lamellar ichthyosis, congenital ichthyosiform erythroderma, and harlequin ichthyosis. ARCI is caused by biallelic mutations in ABCA12, ALOX12B, ALOXE3, CERS3, CYP4F22, NIPAL4, PNPLA1, SDR9C7, SULT2B1, and TGM1. The most severe form of ARCI, harlequin ichthyosis, is caused by mutations in ABCA12. Mutations in this gene can also lead to congenital ichthyosiform erythroderma or lamellar ichthyosis. We present a large cohort of 64 patients affected with ARCI carrying biallelic mutations in ABCA12. Our study comprises 34 novel mutations in ABCA12, expanding the mutational spectrum of ABCA12-associated ARCI up to 217 mutations. Within these we found the possible mutational hotspots c.4541G>A, p.(Arg1514His) and c.4139A>G, p.(Asn1380Ser). A correlation of the phenotype with the effect of the genetic mutation on protein function is demonstrated. Loss-of-function mutations on both alleles generally result in harlequin ichthyosis, whereas biallelic missense mutations mainly lead to CIE or LI.

Double-Hit Mutations in Bicuspid Aortic Valve and Blunt Traumatic Acute Aortic Dissection.

We report on a young patient with a bicuspid aortic valve operated on for type A acute aortic dissection because of a blunt thoracic trauma. Aortic root replacement and ascending aortic and total arch repair together with the postoperative course were uneventful. Multigenerational genetic analyses revealed mutations in the NOTCH1 and ACTA2 genes in the patient and his father. The screening of his parents and children revealed no bicuspid aortic valve or aortic root dilation. This exceptionally rare case of double-hit mutations and the presence of blunt trauma reveals the complex etiology of aortic dissection in patients with a bicuspid aortic valve.

Novel Association of a Familial TGFBR1 Mutation in Loeys-Dietz Syndrome with Concomitant Hematologic Malignancy.

Concomitant Loeys-Dietz syndrome (LDS) and hematologic malignancies are exceptionally rare. This is the first report of a patient operated on for aortic root dilation who had been previously diagnosed with LDS and B-cell-lymphoma. After completion of chemotherapy and complete remission, an elective valve-sparing aortic root replacement (using the David-V method) was performed. Due to the positive family history, pre-operative genetic counseling was conducted, and revealed LDS with a TGFBR1 (transforming growth factor beta receptor type I) mutation in 6 probands of the family, albeit in 1 of them posthumously. This missense mutation has been previously described in relation to aortic dissection, but a causative relationship to malignancy has so far neither been proposed nor proven.

Novel MECP2 Mutation c.1162_1172del; p.Pro388* in Two Patients with Symptoms of Atypical Rett Syndrome.

We report 2 cases of girls with MECP2 gene variants who do not have typical clinical features of Rett syndrome except for intellectual disability and seizures. Both patients present with adipositas, macrocephalia, precocious puberty, and seizures. They have prominent eyebrows and a short neck as well as short and plump fingers. Sequencing by NGS revealed a novel variant c.1162_1172del; p.Pro388* in both patients.

A family with limb girdle muscular dystrophy type 1B and multiple exostoses.

INTRODUCTION: Next-generation sequencing in cases of hereditary neuromuscular disorders often yields multiple candidate gene variants. Here, we describe a case with mutations in two genes, lamin A/C (LMNA) and exostosin glycosyltransferase 2 (EXT2), which led to hereditary myopathy combined with multiple exostoses. CASE HISTORY: A 51-year-old German woman with a history of removal of multiple exostoses during childhood presented with proximal limb-girdle muscular dystrophy and a newly diagnosed cardiomyopathy with atrioventricular conduction block. Because her younger son had exostoses and her younger brother had died at age 44 after heart transplantation due to dilated cardiomyopathy, an autosomal dominant inheritance was suspected. RESULTS: Muscle biopsy revealed features of chronic myopathy associated with focal myofibrillar disintegration. Electron microscopy showed myonuclear, myofibrillar, and Z-disc alterations, accumulations of granulofilamentous material, and a large sporadic osmiophilic inclusion body reminiscent of a nemaline body. Mendeliome and Sanger sequencing detected both a c.1129>T LMNA mutation of known pathogenicity and a c.1101_1102delAG (E368Kfs*18) truncating EXT2 mutation in the patient and her affected son. DISCUSSION: The clinical, genetic, and muscle biopsy findings suggest that both mutations are pathogenic. The EXT2 mutation was most likely responsible for the multiple exostoses phenotype in mother and son, whereas the myopathy was probably caused by a combined effect of the LMNA and EXT2 mutations.

Differentiation of MISSLA and Fanconi anaemia by computer-aided image analysis and presentation of two novel MISSLA siblings.

Variants in DONSON were recently identified as the cause of microcephaly, short stature, and limb abnormalities syndrome (MISSLA). The clinical spectra of MISSLA and Fanconi anaemia (FA) strongly overlap. For that reason, some MISSLA patients have been clinically diagnosed with FA. Here, we present the clinical data of siblings with MISSLA featuring a novel DONSON variant and summarize the current literature on MISSLA. Additionally, we perform computer-aided image analysis using the DeepGestalt technology to test how distinct the facial features of MISSLA and FA patients are. We show that MISSLA has a specific facial gestalt. Notably, we find that also FA patients feature facial characteristics recognizable by computer-aided image analysis. We conclude that computer-assisted image analysis improves diagnostic precision in both MISSLA and FA.

Defining and expanding the phenotype of QARS-associated developmental epileptic encephalopathy.

OBJECTIVE: The study is aimed at widening the clinical and genetic spectrum and at assessing genotype-phenotype associations in QARS encephalopathy. METHODS: Through diagnostic gene panel screening in an epilepsy cohort, and recruiting through GeneMatcher and our international network, we collected 10 patients with biallelic QARS variants. In addition, we collected data on 12 patients described in the literature to further delineate the associated phenotype in a total cohort of 22 patients. Computer modeling was used to assess changes on protein folding. RESULTS: Biallelic pathogenic variants in QARS cause a triad of progressive microcephaly, moderate to severe developmental delay, and early-onset epilepsy. Microcephaly was present at birth in 65%, and in all patients at follow-up. Moderate (14%) or severe (73%) developmental delay was characteristic, with no achievement of sitting (85%), walking (86%), or talking (90%). Additional features included irritability (91%), hypertonia/spasticity (75%), hypotonia (83%), stereotypic movements (75%), and short stature (56%). Seventy-nine percent had pharmacoresistant epilepsy with mainly neonatal onset. Characteristic cranial MRI findings include early-onset progressive atrophy of cerebral cortex (89%) and cerebellum (61%), enlargement of ventricles (95%), and age-dependent delayed myelination (88%). A small subset of patients displayed a less severe phenotype. CONCLUSIONS: These data revealed first genotype-phenotype associations and may serve for improved interpretation of new QARS variants and well-founded genetic counseling.

Microcephaly, short stature, and limb abnormality disorder due to novel autosomal biallelic DONSON mutations in two German siblings.

Recently, variants in DONSON have been reported to cause different disorders of the microcephalic primordial dwarfism spectrum. Using whole-exome sequencing, we identified two novel, compound heterozygous DONSON variants in a pair of siblings, one of whom was previously diagnosed with Fanconi anemia. This occurred because the present cases exhibited clinical findings in addition to those of the microcephalic primordial dwarfism disorder, including severe limb malformations. These findings suggest that the DONSON and Fanconi anemia proteins could have supplementary roles in developmental processes as they have in the maintenance of genomic integrity, resulting in related disease phenotypes.

Transforming Growth Factor Beta-2 Mutations in Barlow's Disease and Aortic Dilatation.

We report on a patient operated on for degenerative myxomatous mitral and tricuspid valve disease (Barlow's disease) and aortic root dilatation. A valve repair operation and the postoperative course were uneventful. Multigenerational genetic analyses revealed two different mutations in the transforming growth factor beta-2 gene in the same patient. The two mutations in different exons were inherited from both parents each. None of the parents presented with either valve dysfunction or aortic root dilatation. This rare case illustrates potentially common genetic and signaling pathways of concomitant myxomatous valve disease and aortic root dilatation.

Genetic abnormalities in bicuspid aortic valve root phenotype: preliminary results.

OBJECTIVES: Genetic defects associated with bicuspid aortopathy have been infrequently analysed. Our goal was to examine the prevalence of rare genetic variants in patients with a bicuspid aortic valve (BAV) with a root phenotype using next-generation sequencing technology. METHODS: We investigated a total of 124 patients with BAV with a root dilatation phenotype who underwent aortic valve ± proximal aortic surgery at a single institution (BAV database, n = 812) during a 20-year period (1995-2015). Cross-sectional follow-up revealed 63 (51%) patients who were still alive and willing to participate. Systematic follow-up visits were scheduled from March to December 2015 and included aortic imaging as well as peripheral blood sampling for genetic testing. Next-generation sequencing libraries were prepared using a custom-made HaloPlex HS gene panel and included 20 candidate genes known to be associated with aortopathy and BAV. The primary end-point was the prevalence of genetic defects in our study cohort. RESULTS: A total of 63 patients (mean age 46 ± 10 years, 92% men) with BAV root phenotype and mean post-aortic valve replacement follow-up of 10.3 ± 4.9 years were included. Our genetic analysis yielded a wide spectrum of rare, potentially or likely pathogenic variants in 19 (30%) patients, with NOTCH1 variants being the most common ( n = 6). Moreover, deleterious variants were revealed in AXIN1 ( n = 3), NOS3 ( n = 3), ELN ( n = 2), FBN1 ( n = 2) , FN1 ( n = 2) and rarely in other candidate genes. CONCLUSIONS: Our preliminary study demonstrates a high prevalence and a wide spectrum of rare genetic variants in patients with the BAV root phenotype, indicative of the potentially congenital origin of associated aortopathy in this specific BAV cohort.

[Gastroparesis in Noonan syndrome]. / Gastroparese bei Noonan-Syndrom.

We present a case of a 26-year-old female patient with bloating, postprandial nausea and recurrent vomiting after solid food intake. A gastric emptying scintigraphy showed a delayed gastric emptying, defining gastroparesis. Because of her past medical history of short stature and pulmonary stenosis, we initiated genetic counseling where the diagnosis of Noonan syndrome was made. Dietary therapy and medication with domperidone quickly led to relief of the discomfort due to gastroparesis. However, prokinetics are not indicated for long-term therapy, as cardiac arrhythmia may occur. A risk-benefit assessment should be done. There are several novel approaches which need to be further investigated.

Next-generation sequencing in X-linked intellectual disability.

X-linked intellectual disability (XLID) is a genetically heterogeneous disorder with more than 100 genes known to date. Most genes are responsible for a small proportion of patients only, which has hitherto hampered the systematic screening of large patient cohorts. We performed targeted enrichment and next-generation sequencing of 107 XLID genes in a cohort of 150 male patients. Hundred patients had sporadic intellectual disability, and 50 patients had a family history suggestive of XLID. We also analysed a sporadic female patient with severe ID and epilepsy because she had strongly skewed X-inactivation. Target enrichment and high parallel sequencing allowed a diagnostic coverage of >10 reads for ~96% of all coding bases of the XLID genes at a mean coverage of 124 reads. We found 18 pathogenic variants in 13 XLID genes (AP1S2, ATRX, CUL4B, DLG3, IQSEC2, KDM5C, MED12, OPHN1, SLC9A6, SMC1A, UBE2A, UPF3B and ZDHHC9) among the 150 male patients. Thirteen pathogenic variants were present in the group of 50 familial patients (26%), and 5 pathogenic variants among the 100 sporadic patients (5%). Systematic gene dosage analysis for low coverage exons detected one pathogenic hemizygous deletion. An IQSEC2 nonsense variant was detected in the female ID patient, providing further evidence for a role of this gene in encephalopathy in females. Skewed X-inactivation was more frequently observed in mothers with pathogenic variants compared with those without known X-linked defects. The mutation rate in the cohort of sporadic patients corroborates previous estimates of 5-10% for X-chromosomal defects in male ID patients.

Disturbed neuronal ER-Golgi sorting of unassembled glycine receptors suggests altered subcellular processing is a cause of human hyperekplexia.

Recent studies on the pathogenic mechanisms of recessive hyperekplexia indicate disturbances in glycine receptor (GlyR) α1 biogenesis. Here, we examine the properties of a range of novel glycine receptor mutants identified in human hyperekplexia patients using expression in transfected cell lines and primary neurons. All of the novel mutants localized in the large extracellular domain of the GlyR α1 have reduced cell surface expression with a high proportion of receptors being retained in the ER, although there is forward trafficking of glycosylated subpopulations into the ER-Golgi intermediate compartment and cis-Golgi compartment. CD spectroscopy revealed that the mutant receptors have proportions of secondary structural elements similar to wild-type receptors. Two mutants in loop B (G160R, T162M) were functional, but none of those in loop D/ß2-3 were. One nonfunctional truncated mutant (R316X) could be rescued by coexpression with the lacking C-terminal domain. We conclude that a proportion of GlyR α1 mutants can be transported to the plasma membrane but do not necessarily form functional ion channels. We suggest that loop D/ß2-3 is an important determinant for GlyR trafficking and functionality, whereas alterations to loop B alter agonist potencies, indicating that residues here are critical elements in ligand binding.

Transforming growth factor-beta receptor type II mutation in a patient with bicuspid aortic valve disease and intraoperative aortic dissection.

We report on a patient with familial bicuspid aortic valve disease operated on for proximal aortic aneurysm. The surgery was complicated by intraoperative aortic dissection. Multi-generational genetic analysis demonstrated a mutation in the transforming growth factor-beta receptor type II gene. This case confirms the clinical hypothesis that the proximal aortic disease has a genetic origin in some bicuspid aortic valve patients.

Prenatal diagnosis of Roberts syndrome and detection of an ESCO2 frameshift mutation in a Pakistani family.

OBJECTIVES: We report two siblings with Roberts syndrome (RBS), and an attempt to delineate the underlying molecular mechanism leading to familial recurrence. METHODS: Cytogenetic studies and direct sequencing of the ESCO2 gene were carried out in the second affected fetus and the parents. Fetal DNA was obtained from amniocytes after amniocentesis. Parental DNA was obtained from peripheral blood samples. RESULTS: Cytogenetic analysis of amniocytes revealed a normal male karyotype in 20 analyzed metaphases and chromosomal aneuploidies in 10 metaphases. All metaphases displayed premature separation of centromeres and puffing of heterochromatic regions near the centromere. A homozygous mutation leading to a frameshift in ESCO2 was identified in the fetal DNA sample. Both parents are heterozygous carriers of the same mutation. CONCLUSION: The present case demonstrates the prenatal diagnosis of RBS associated with a frameshift mutation in ESCO2.

Greig cephalopolysyndactyly (GCPS) contiguous gene syndrome in a boy with a 14 Mb deletion in region 7p13-14 caused by a paternal balanced insertion (5; 7).

We report on a six years old boy with several features of Greig cephalopolysyndactyly syndrome (GCPS) including craniofacial dysmorphism, hypertelorism, heart defect, preaxial hexadactyly of toes, partial agenesis of corpus callosum, and severe developmental delay. Greig cephalopolysyndactyly (GCPS) can be caused by GLI3 deletions. In patients with large deletions which include additional genes, it is termed Greig cephalopolysyndactyly-contiguous gene syndrome (GCPS-CGS). It is generally believed that the deletion size correlates with disease severity. Nearly all cases appear to be a result of GLI3 de novo deletions. Chromosome analysis of our patient revealed a large deletion in chromosome 7(p13-p14). Unlike most previously described cases, we found that this deletion resulted from a paternal balanced insertional translocation of 7p13-14 into the long arm of chromosome 5.

Immunocytochemical identification of low-affinity NTS2 neurotensin receptors in parietal cells of human gastric mucosa.

The biological effects of neurotensin (NT) are mediated by two distinct G protein-coupled receptors, NTS(1) and NTS(2). Although it is well established that neurotensin inhibits gastric acid secretion in man, the plasma membrane receptor mediating these effects has not been visualized yet. We developed and characterized a novel antipeptide antibody to the carboxy-terminal region of the human NTS(2) receptor. The cellular and subcellular distribution of NTS(2) receptors was evaluated in various human gastrointestinal tissues. Specificity of the antiserum was demonstrated by (1) detection of a broadband migrating at M(r) 90 000-100 000 in Western blots of membranes from NTS(2)-expressing tissues; (2) cell-surface staining of NTS(2)-transfected cells; (3) translocation of NTS(2) receptor immunostaining after agonist exposure; and (4) abolition of tissue immunostaining by preadsorbtion of the antibody with its immunizing peptide. In the gastrointestinal tract, NTS(2) receptor immunoreactivity was highly abundant in parietal cells of the gastric mucosa, in neuroendocrine cells of the stomach small and large intestine, and in cells of the exocrine pancreas. NTS(2) receptors were clearly located in the plasma membrane and uniformly present on nearly all target cells. The presence of NTS(2) receptors was rarely detected in human tumors. This is the first localization of NTS(2) receptors in human formalin-fixed, paraffin-embedded tissues at the cellular level. The abundant expression of low-affinity NTS(2) receptors on the plasma membrane of human parietal cells provides a morphological substrate for the direct inhibition of gastric acid secretion observed after i.v. administration of neurotensin.