Mutations in CYB561 Causing a Novel Orthostatic Hypotension Syndrome

Rationale: Orthostatic hypotension is a common clinical problem, but the underlying mechanisms have not been fully delineated. Objective: We describe two families, with four patients in total, suffering from severe life-threatening orthostatic hypotension due to a novel cause. Methods and Results: As in dopamine β-hydroxylase deficiency (DβH), concentrations of norepinephrine and epinephrine in the patients were very low. Plasma DβH activity, however, was normal and the DBH gene had no mutations. Molecular genetic analysis was performed to determine the underlying genetic cause. Homozygosity mapping and exome and Sanger sequencing revealed pathogenic homozygous mutations in the gene encoding cytochrome b561 (CYB561); a missense variant c.262G>A, p.Gly88Arg in exon 3 in the Dutch family and a nonsense mutation (c.131G>A, p.Trp44*) in exon 2 in the American family. Expression of CYB561 was investigated using RNA from different human adult and fetal tissues, transcription of RNA into cDNA and real-time quantitative polymerase chain reaction. The CYB561 gene was found to be expressed in many human tissues, in particular the brain. The CYB561 protein defect leads to a shortage of ascorbate inside the catecholamine secretory vesicles leading to a functional DβH deficiency. The concentration of the catecholamines and downstream metabolites was measured in brain and adrenal tissue of six CYB561 knockout mice (reporter-tagged deletion allele (post-Cre), genetic background C57BL/6NTac). The concentration of norepinephrine and normetanephrine was decreased in whole brain homogenates of the CYB561(-/-) mice compared to wild type mice (p<0.01) and the concentration of normetanephrine and metanephrine was decreased in adrenal glands (p<0.01), recapitulating the clinical phenotype. The patients responded favorably to treatment with L-dihydroxyphenylserine, which can be converted directly to norepinephrine. Conclusions: This study is the first to implicate cytochrome b561 in disease by showing that pathogenic mutations in CYB561 cause an as yet unknown disease in neurotransmitter metabolism causing orthostatic hypotension. as yet unknown disease in neurotransmitter metabolism causing orthostatic hypotension

Experiences with array-based sequence capture; toward clinical applications

Although sequencing of a human genome gradually becomes an option, zooming in on the region of interest remains attractive and cost saving. We performed array-based sequence capture using 385K Roche NimbleGen, Inc. arrays to zoom in on the protein-coding and immediate intron-flanking sequences of 112 genes, potentially involved in mental retardation and congenital malformation. Captured material was sequenced using Illumina technology. A data analysis pipeline was built that detects sequence variants, positions them in relation to the gene, checks for presence in databases (eg, db single-nucleotide polymorphism (SNP)) and predicts the potential consequences at the level of RNA splicing and protein translation. In the samples analyzed, all known variants were reliably detected, including pathogenic variants from control cases and SNPs derived from array experiments. Although overall coverage varied considerably, it was reproducible per region and facilitated the detection of large deletions and duplications (copy number variations), including a partial deletion in the B3GALTL gene from a patient sample. For ultimate diagnostic application, overall results need to be improved. Future arrays should contain probes from both DNA strands, and to obtain a more even coverage, one could add fewer probes from densely and more probes from sparsely covered regions

Basal Cell Carcinoma Pathology Requests and Reports Are Lacking Important Information

Introduction. Basal cell carcinoma (BCC) is the most common cancer affecting humans. Luckily it has negligible risk for metastasis; however it can be locally destructive to surrounding tissue. The diagnosis of this tumor relies on clinical and dermoscopic features; however confirmation requires biopsy and histologic examination. Based on clinical and pathologic findings, BCC is classified as low or high risk subtype. The clinician requesting pathology examination for BCC should provide the pathologist with detailed information including patient details, relevant clinical and medical history, site and type of the biopsy, and whether this is a primary or recurrent lesion. The pathologist on the other hand should write an adequate report containing a minimum of core set of parameters including type of BCC, depth of invasion, presence of lymphovascular or perineural invasion, and the excision margins. Objectives. The objective of this study is to evaluate whether requests by clinicians and pathology reports of BCC are adequate. Methods. This is a retrospective analysis done at the dermatology department, faculty of medicine at Jordan University of Science and Technology, Irbid, Jordan. Reports for the period from January 2003 to December 2017 were retrieved and analyzed for data completeness. Results. Most clinical request forms of BCC provided by clinicians are inadequate and lack important relevant information especially in regard to lesion history, patient medical history, and whether BCC is a primary or a recurrent one. Pathology reports for BCC cases also have significant deficiency especially in describing the histologic subtype, depth of invasion, and presence of lymphovascular and perineural invasion. However, the tumor excision margins are adequately described in almost all reports. Conclusions. The study shows that clinicians do not provide adequate clinical information when submitting a request for histopathologic examination of BCC. Similarly, pathologists write incomplete reports that lack important pathologic features. Having pre-set forms (electronic proforma) can help overcome missing information

Yield of peripheral sodium channels gene screening in pure small fibre neuropathy

Background Neuropathic pain is common in peripheral neuropathy. Recent genetic studies have linked pathogenic voltage-gated sodium channel (VGSC) variants to human pain disorders. Our aims are to determine the frequency of SCN9A, SCN10A and SCN11A variants in patients with pure small fibre neuropathy (SFN), analyse their clinical features and provide a rationale for genetic screening. Methods Between September 2009 and January 2017, 1139 patients diagnosed with pure SFN at our reference centre were screened for SCN9A, SCN10A and SCN11A variants. Pathogenicity of variants was classified according to established guidelines of the Association for Clinical Genetic Science and frequencies were determined. Patients with SFN were grouped according to the VGSC variants detected, and clinical features were compared. Results A mong 1139 patients with SFN, 132 (11.6%) patients harboured 73 different (potentially) pathogenic VGSC variants, of which 50 were novel and 22 were found in >= 1 patient. The frequency of (potentially) pathogenic variants was 5.1% (n=58/1139) for SCN9A, 3.7% (n=42/1139) for SCN10A and 2.9% (n=33/1139) for SCN11A. Only erythromelalgia-like symptoms and warmth-induced pain were significantly more common in patients harbouring VGSC variants. Conclusion (Potentially) pathogenic VGSC variants are present in 11.6% of patients with pure SFN. Therefore, genetic screening of SCN9A, SCN10A and SCN11A should be considered in patients with pure SFN, independently of clinical features or underlying conditions

Peripheral Ion Channel Genes Screening in Painful Small Fiber Neuropathy

Neuropathic pain is a characteristic feature of small fiber neuropathy (SFN), which in 18% of the cases is caused by genetic variants in voltage-gated sodium ion channels. In this study, we assessed the role of fifteen other ion channels in neuropathic pain. Patients with SFN (n = 414) were analyzed for ANO1, ANO3, HCN1, KCNA2, KCNA4, KCNK18, KCNN1, KCNQ3, KCNQ5, KCNS1, TRPA1, TRPM8, TRPV1, TRPV3 and TRPV4 variants by single-molecule molecular inversion probes-next-generation sequencing. These patients did not have genetic variants in SCN3A, SCN7A-SCN11A and SCN1B-SCN4B. In twenty patients (20/414, 4.8%), a potentially pathogenic heterozygous variant was identified in an ion-channel gene (ICG). Variants were present in seven genes, for two patients (0.5%) in ANO3, one (0.2%) in KCNK18, two (0.5%) in KCNQ3, seven (1.7%) in TRPA1, three (0.7%) in TRPM8, three (0.7%) in TRPV1 and two (0.5%) in TRPV3. Variants in the TRP genes were the most frequent (n = 15, 3.6%), partly in patients with high mean maximal pain scores VAS = 9.65 ± 0.7 (n = 4). Patients with ICG variants reported more severe pain compared to patients without such variants (VAS = 9.36 ± 0.72 vs. VAS = 7.47 ± 2.37). This cohort study identified ICG variants in neuropathic pain in SFN, complementing previous findings of ICG variants in diabetic neuropathy. These data show that ICG variants are central in neuropathic pain of different etiologies and provides promising gene candidates for future research

Homozygous damaging SOD2 variant causes lethal neonatal dilated cardiomyopathy

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Peripheral Ion Channel Genes Screening in Painful Small Fiber Neuropathy

Neuropathic pain is a characteristic feature of small fiber neuropathy (SFN), which in 18% of the cases is caused by genetic variants in voltage-gated sodium ion channels. In this study, we assessed the role of fifteen other ion channels in neuropathic pain. Patients with SFN (n = 414) were analyzed for ANO1, ANO3, HCN1, KCNA2, KCNA4, KCNK18, KCNN1, KCNQ3, KCNQ5, KCNS1, TRPA1, TRPM8, TRPV1, TRPV3 and TRPV4 variants by single-molecule molecular inversion probes&ndash;next-generation sequencing. These patients did not have genetic variants in SCN3A, SCN7A-SCN11A and SCN1B-SCN4B. In twenty patients (20/414, 4.8%), a potentially pathogenic heterozygous variant was identified in an ion-channel gene (ICG). Variants were present in seven genes, for two patients (0.5%) in ANO3, one (0.2%) in KCNK18, two (0.5%) in KCNQ3, seven (1.7%) in TRPA1, three (0.7%) in TRPM8, three (0.7%) in TRPV1 and two (0.5%) in TRPV3. Variants in the TRP genes were the most frequent (n = 15, 3.6%), partly in patients with high mean maximal pain scores VAS = 9.65 &plusmn; 0.7 (n = 4). Patients with ICG variants reported more severe pain compared to patients without such variants (VAS = 9.36 &plusmn; 0.72 vs. VAS = 7.47 &plusmn; 2.37). This cohort study identified ICG variants in neuropathic pain in SFN, complementing previous findings of ICG variants in diabetic neuropathy. These data show that ICG variants are central in neuropathic pain of different etiologies and provides promising gene candidates for future research