Monoallelic Variant Research Articles

The multifaceted phenotypic and genotypic spectrum of type-IV-collagen-related nephropathy-A human genetics department experience.

Disease-causing variants in COL4A3-5 are associated with type-IV-collagen-related nephropathy, a genetically and phenotypically multifaceted disorder comprising Alport syndrome (AS) and thin basement membrane nephropathy (TBMN) and autosomal, X-linked and a proposed digenic inheritance. Initial symptoms of individuals with AS are microscopic hematuria followed by proteinuria leading to kidney failure (90% on dialysis < age 40 years). In contrast, individuals with TBMN, an outdated histology-derived term, present with microscopic hematuria, only some of them develop kidney failure (>50 years of age). An early diagnosis of type-IV-collagen-related nephropathy is essential for optimized therapy and slowing of the disease. Sixty index cases, in whom exome sequencing had been performed and with disease-causing variant(s) in COL4A3-5, were evaluated concerning their clinical tentative diagnosis and their genotype. Of 60 reevaluated individuals with type-IV-collagen-related nephropathy, 72% had AS, 23% TBMN and 5% focal segmental glomerulosclerosis (FSGS) as clinical tentative diagnosis. The FSGS cases had to be re-classified as having type-IV-collagen-related nephropathy. Twelve percent of cases had AS as clinical tentative diagnosis and a monoallelic disease-causing variant in COL4A3/4 but could not be classified as autosomal dominant AS because of limited or conflicting clinical data. This study illustrates the complex clinical and genetic picture of individuals with a type IV-collagen-related nephropathy indicating the need of a refined nomenclature and the more interdisciplinary teamwork of clinicians and geneticists as the key to optimized patient care.

Monoallelic pathogenic ALG5 variants cause atypical polycystic kidney disease and interstitial fibrosis

Disorders of the autosomal dominant polycystic kidney disease (ADPKD) spectrum are characterized by the development of kidney cysts and progressive kidney function decline. PKD1 and PKD2, encoding polycystin (PC)1 and 2, are the two major genes associated with ADPKD; other genes include IFT140, GANAB, DNAJB11, and ALG9. Genetic testing remains inconclusive in ∼7% of the families. We performed whole-exome sequencing in a large multiplex genetically unresolved (GUR) family affected by ADPKD-like symptoms and identified a monoallelic frameshift variant (c.703_704delCA) in ALG5. ALG5 encodes an endoplasmic-reticulum-resident enzyme required for addition of glucose molecules to the assembling N-glycan precursors. To identify additional families, we screened a cohort of 1,213 families with ADPKD-like and/or autosomal-dominant tubulointerstitial kidney diseases (ADTKD), GUR (n= 137) or naive to genetic testing (n= 1,076), by targeted massively parallel sequencing, and we accessed Genomics England 100,000 Genomes Project data. Four additional families with pathogenic variants in ALG5 were identified. Clinical presentation was consistent in the 23 affected members, with non-enlarged cystic kidneys and few or no liver cysts; 8 subjects reached end-stage kidney disease from 62 to 91 years of age. We demonstrate that ALG5 haploinsufficiency is sufficient to alter the synthesis of the N-glycan chain in renal epithelial cells. We also show that ALG5 is required for PC1 maturation and membrane and ciliary localization and that heterozygous loss of ALG5 affects PC1 maturation. Overall, our results indicate that monoallelic variants of ALG5 lead to a disorder of the ADPKD-spectrum characterized by multiple small kidney cysts, progressive interstitial fibrosis, and kidney function decline.

IRIDA Phenotype in TMPRSS6 Monoallelic-Affected Patients: Toward a Better Understanding of the Pathophysiology.

Iron-refractory iron deficiency anemia (IRIDA) is an autosomal recessive inherited form of iron deficiency anemia characterized by discrepantly high hepcidin levels relative to body iron status. However, patients with monoallelic exonic TMPRSS6 variants have also been reported to express the IRIDA phenotype. The pathogenesis of an IRIDA phenotype in these patients is unknown and causes diagnostic uncertainty. Therefore, we retrospectively summarized the data of 16 patients (4 men, 12 women) who expressed the IRIDA phenotype in the presence of only a monoallelic TMPRSS6 variant. Eight unaffected relatives with identical exonic TMPRSS6 variants were used as controls. Haplotype analysis was performed to assess the (intra)genetic differences between patients and relatives. The expression and severity of the IRIDA phenotype were highly variable. Compared with their relatives, patients showed lower Hb, MCV, and TSAT/hepcidin ratios and inherited a different wild-type allele. We conclude that IRIDA in monoallelic TMPRSS6-affected patients is a phenotypically and genotypically heterogeneous disease that is more common in female patients. We hypothesize that allelic imbalance, polygenetic inheritance, or modulating environmental factors and their complex interplay are possible causes. This explorative study is the first step toward improved insights into the pathophysiology and improved diagnostic accuracy for patients presenting with IRIDA and a monoallelic exonic TMPRSS6 variant.

P1408: MONOALLELIC AND BIALLELIC GERMLINE MUTATIONS AFFECTING THE TRANSCRIPTION FACTOR HELIOS CAUSE PLEIOTROPIC DEFECTS OF IMMUNITY

Background: Helios, encoded by IKZF2, is a member of the Ikaros family of transcription factors with pivotal roles in T-follicular helper, NK- and T-regulatory cell physiology. Somatic IKZF2 mutations are frequently found in lymphoid malignancies. Aims: Although germline mutations in IKZF1 and IKZF3, encoding Ikaros and Aiolos, have recently been identified in patients with phenotypically similar immunodeficiency syndromes, the effect of germline mutations in IKZF2 on human hematopoiesis and immunity remains to be elucidated. Methods: We performed whole exome sequencing in six patients with immune dysregulation and their family members. We then applied Co-immunoprecipitation, electrophoretic mobility shift assay and immunofluorescence microscopy techniques to assess dimerization and DNA binding capabilities of the mutations. Proteomic analysis by proximity-dependent Biotin Identification (BioID) was used to assess affinity of mutant Helios to its interaction partners, while single-cell RNA sequencing was applied to peripheral blood mononuclear cells (PBMCs) of two patients to study the transcriptional effect of the truncating and biallelic IKZF2 mutations. Results: We identified six patients with monoallelic germline IKZF2 mutations (one nonsense (p.R291X)- and 4 distinct missense variants) and one patient with a biallelic germline IKZF2 mutation (p.I325V). While patients with heterozygous variants were affected by immune dysregulation (systemic lupus erythematosus, immune thrombocytopenia or EBV-associated hemophagocytic lymphohistiocytosis), the patient with the homozygous variant suffered from recurrent respiratory tract infections complicated by bronchiectasis. Immunophenotype analysis revealed reduced propensities of NK cells, CD4+ T-cells with an inverted CD4/CD8 ratio, T-follicular helper cells and hypogammaglobulinemia across patients, while the p.I325V patient also had a reduction in relative and absolute B-cells and low relative proportions of Treg, MAIT and iNKT populations. Single-cell RNA sequencing of PBMCs from the patients carrying the monoallelic R291X truncating variant or the biallelic I325V variant revealed upregulation of pro-inflammatory genes associated with T-cell receptor activation and T-cell exhaustion. Our biochemical and proteomic analysis has revealed that only the truncating R291X mutation results in loss of Helios DNA binding and dimerization properties. However, BioID analysis revealed aberrant interaction of 4/6 Helios mutants with core components of the Nucleosome Remodeling histone Deacetylase (NuRD) complex conveying Helios-mediated epigenetic and transcriptional dysregulation. Summary/Conclusion: Thus, germline mutations in IKZF2 cause a novel syndrome associating immunodeficiency and profound immune dysregulation and disrupt interaction with the NuRD complex.

A monoallelic variant in EYA1 is associated with Branchio-Otic syndrome in a Malian family.

BackgroundBranchio‐otic syndrome (BO) is one of the most common types of syndromic hearing impairment (HI) with an incidence of 1/40,000 globally. It is an autosomal dominant disorder typically characterized by the coexistence of branchial cysts or fistulae, malformations of the external, middle, and inner ears with preauricular pits or tags and a variable degree of HI. Most cases of BO have been reported in populations of European ancestry. To date, only few cases have been reported in people from African descent.MethodsAfter a careful clinical examination, a pure tone audiometry was performed. DNA was extracted from peripheral blood and whole exome, and Sanger sequencing were performed for genetic analysis.ResultsEight individuals from a large non‐consanguineous Malian family, with autosomal dominant inheritance were enrolled. The ages at diagnosis ranged from 8 to 54 years. A high phenotypic variability was noted among the affected individuals. Four patients presented with a post‐lingual and mixed type of HI, one individual had conductive HI while three had normal hearing but presented other BO features namely branchial fistulae and preauricular sinus. Serum creatinine level and renal ultrasonography were normal in three affected individuals who performed them. Genetic testing identified a monoallelic pathogenic variant in EYA1 (c.1286A > G; p.Asp429Gly) segregating with BO syndrome in the family.ConclusionThis is the first genetically confirmed case of BO syndrome caused by EYA1 variant in the sub‐Saharan African population, expanding the genetic spectrum of the condition.

Lethal neonatal respiratory failure due to biallelic variants in BBS1 and monoallelic variant in TTC21B.

Bardet-Biedl syndrome (BBS) is a rare, autosomal recessive ciliopathy characterized by early onset retinal dystrophy, renal anomalies, postaxial polydactyly, and cognitive impairment with considerable phenotypic heterogeneity. BBS results from biallelic pathogenic variants in over 20 genes that encode key proteins required for the assembly or primary ciliary functions of the BBSome, a heterooctameric protein complex critical for homeostasis of primary cilia. While variants in BBS1 are most frequently identified in affected individuals, the renal and pulmonary phenotypes associated with BBS1 variants are reportedly less severe than those seen in affected individuals with pathogenic variants in the other BBS-associated genes. We report an infant with severe renal dysplasia and lethal pulmonary hypoplasia who was homozygous for the most common BBS1 pathogenic variant (c.1169T > G; p.M390R) and also carried a predicted pathogenic variant in TTC21B (c.1846C > T; p.R616C), a genetic modifier of disease severity of ciliopathies associated with renal dysplasia and pulmonary hypoplasia. This report expands the phenotypic spectrum of BBS with the first infant with lethal neonatal respiratory failure associated with biallelic, pathogenic variants in BBS1 and a monoallelic, predicted pathogenic variant in TTC21B. BBS should be considered among the ciliopathies in the differential diagnosis of neonates with renal dysplasia and severe respiratory failure.

Phenotype-Based Genetic Analysis Reveals Missing Heritability of ABCA4-Related Retinopathy: Deep Intronic Variants and Copy Number Variations

PurposeTo identify the missing heritability of ABCA4-related retinopathy in a Chinese cohort.MethodsWe recruited 33 unrelated patients with ABCA4-related retinopathy carrying a monoallelic variant in ABCA4. All patients underwent ophthalmic examinations. Next-generation sequencing of the whole ABCA4 sequence, including coding and noncoding regions, was performed to detect deep intronic variants (DIVs) and copy number variations (CNVs).ResultsWe identified eight missing pathogenic ABCA4 variants in 60.6% of the patients (20/33), which comprised five DIVs and three CNVs. The five DIVs, including four novel (c.1555-816T>G, c.2919-169T>G, c.2919-884G>T, and c.5461-1321A>G) and one reported (c.4539+1100A>G), accounted for the missing alleles in 51.5% of the patients. Minigene assays showed that four novel DIVs activated cryptic splice sites leading to the insertions of pseudoexons. The three novel CNVs consisted of one gross deletion of 1273 bp (exon 2) and two gross duplications covering 25.2 kb (exons 28–43) and 9.4 kb (exons 38–44). The microhomology domains were identified at the breakpoints and revealed the potential mechanisms of CNV formation.ConclusionsDIVs and CNVs explained approximately two-thirds of the unresolved Chinese cases with ABCA4-related retinopathy. Combining results from phenotypic-directed screening, targeting the whole ABCA4 sequencing and in silico tools can help to identify the missing heritability.

Mutational screening of the TPO and DUOX2 genes in Argentinian children with congenital hypothyroidism due to thyroid dyshormonogenesis

Primary congenital hypothyroidism (CH) is the most common endocrine disease in children and one of the preventable causes of both cognitive and motor deficits. We present a genetic and bioinformatics investigation of rational clinical design in 17 Argentine patients suspected of CH due to thyroid dyshormonogenesis (TDH). Next-Generation Sequencing approach was used to identify variants in Thyroid Peroxidase (TPO) and Dual Oxidase 2 (DUOX2) genes. A custom panel targeting 7 genes associated with TDH [(TPO), Iodothyrosine Deiodinase I (IYD), Solute Carrier Family 26 Member 4 (SLC26A4), Thyroglobulin (TG), DUOX2, Dual Oxidase Maturation Factor 2 (DUOXA2), Solute Carrier Family 5 Member 5 (SLC5A5)] and 4 associated with thyroid dysembryogenesis [PAX8, FOXE1, NKX2-1, Thyroid Stimulating Hormone Receptor (TSHR)] has been designed. Additionally, bioinformatic analysis and structural modeling were carried out to predict the disease-causing potential variants. Four novel variants have been identified, two in TPO: c.2749-2 A > C and c.2752_2753delAG, [p.Ser918Cysfs*62] and two variants in DUOX2 gene: c.425 C > G [p.Pro142Arg] and c.2695delC [p.Gln899Serfs*21]. Eighteen identified TPO, DUOX2 and IYD variants were previously described. We identified potentially pahogenic biallelic variants in TPO and DUOX2 in 7 and 2 patients, respectively. We also detected a potentially pathogenic monoallelic variant in TPO and DUOX2 in 7 and 1 patients respectively. 22 variants have been identified associated with TDH. All described novel mutations occur in domains important for protein structure and function, predicting the TDH phenotype.

De novo variants in EMC1 lead to neurodevelopmental delay and cerebellar degeneration and affect glial function in Drosophila.

The endoplasmic reticulum (ER)-membrane protein complex (EMC) is a multi-protein transmembrane complex composed of 10 subunits that functions as a membrane-protein chaperone. Variants in EMC1 lead to neurodevelopmental delay and cerebellar degeneration. Multiple families with biallelic variants have been published, yet to date, only a single report of a monoallelic variant has been described, and functional evidence is sparse. Exome sequencing was used to investigate the genetic cause underlying severe developmental delay in three unrelated children. EMC1 variants were modeled in Drosophila, using loss-of-function (LoF) and overexpression studies. Glial-specific and neuronal-specific assays were used to determine whether the dysfunction was specific to one cell type. Exome sequencing identified de novo variants in EMC1 in three individuals affected by global developmental delay, hypotonia, seizures, visual impairment and cerebellar atrophy. All variants were located at Pro582 or Pro584. Drosophila studies indicated that imbalance of EMC1-either overexpression or knockdown-results in pupal lethality and suggest that the tested homologous variants are LoF alleles. In addition, glia-specific gene dosage, overexpression or knockdown, of EMC1 led to lethality, whereas neuron-specific alterations were tolerated. We establish de novo monoallelic EMC1 variants as causative of a neurological disease trait by providing functional evidence in a Drosophila model. The identified variants failed to rescue the lethality of a null allele. Variations in dosage of the wild-type EMC1, specifically in glia, lead to pupal lethality, which we hypothesize results from the altered stoichiometry of the multi-subunit protein complex EMC.

Identification of GJB2 Variants in 75 Unrelated Iranian Autosomal Recessive Non-syndromic Hearing Loss Patients

Background: Hearing loss (HL) is the most prevalent sensory disease in humans. HL is among the most clinically and genetically heterogeneous disorders. Pathogenic variants in GJB2 are the principal cause of HL in many populations. Therefore, GJB2 analysis should be considered as the first step for HL. Objectives: This study aimed to find causative variants in the GJB2 gene in 75 unrelated Iranian patients who suffered from Autosomal recessive non-syndromic hearing loss (ARNSHL). Methods: Peripheral blood samples were used for genomic DNA extraction. PCR-direct sequencing was performed to detect GJB2 mutations. Results: In this study, thirty-four chromosomes (21.33%) carried GJB2 mutations. In general, 10 variants were detected among 19 patients. Seven cases were homozygous for c.35delG; (p.Gly12Valfs*2), two were homozygous for c.71G&gt;A; (p.Trp24*), two were homozygous for c.358-360delGAG (p.Glu120del), and two were homozygous for c.-23+1G&gt;A mutation. One patient was a compound heterozygote for c.35delG and c.-23+1G&gt;A mutations, and another patient with compound heterozygosity for c.290dupA and c.235delC mutations were determined. Four patients carried a mono-allelic variant in the GJB2 including c.126G&gt;T; (p.Glu42Asp), c.23C&gt;T; (p.Thr8Met), c.445G&gt;A; (p.Ala149Thr) and c.269T&gt;C; (p.Leu90Pro). Accordingly, c.35delG mutation was the most common variant in this study. Conclusions: Finding common variants of HL mutations in different populations can elevate the diagnostic value of molecular testing in the screening of affected individuals, and can improve counselling to minimize the risk of having affected offspring for at-risk couples. Besides, early diagnosis can easily lead to speech development and prevents further problems.

The impact of the Eurofever criteria and the new InFevers MEFV classification in real life: Results from a large international FMF cohort

IntroductionNew Eurofever/PRINTO classification criteria (EPCC) for Familial Mediterranean Fever (FMF) and other recurrent fevers have been recently developed, together with the classification of the pathogenicity of MEFV variants. ObjectivesTo evaluate the impact in real life of both the EPCC and INSAID pathogenicity classification of MEFV variants in the large international Eurofever FMF cohort. MethodsBaseline demographic, genetic and clinical data of FMF patients included in the Eurofever registry were evaluated. The EPCC and the 2018 INSAID classification for MEFV variants were applied in all eligible FMF patients. ResultsSince November 2009, clinical information was available for 1012 FMF (532 males/480 females, 827 children/185 adults) from 119 centres. Complete data were available for 887 patients in whom 623 (70.2%) satisfied EPCC (EPCC+), while 264 (29.8%) did not (EPCC−). The majority of the EPCC− patients (172, 65.1%) displayed negative or non-informative genetics (monoallelic or biallelic benign variants, monoallelic variant of unknown significance). At baseline, colchicine was used in most of EPCC+ patients (88%) and in a lower percentage of EPCC− patients (69%, p < 0.0001), who were treated in a higher proportion with steroid or NSAID on demand (p = 0.003 and 0.008, respectively). Four percent of patients received Anti-IL-1 treatment. ConclusionsThe combination of EPCC and the 2018 INSAID classification of MEFV variants is able to identify two distinct groups of patients, which differ in clinical characteristics, therapeutic approach and response to treatment. EPCC+ patients displayed the typical features of FMF, while EPCC− patients had a more variable phenotype with a lower percentage of response to colchicine.

Monoallelic deleterious MUTYH germline variants as a driver for tumorigenesis.

MUTYH encodes a glycosylase involved in the base excision repair of DNA. Biallelic pathogenic germline variants in MUTYH cause an autosomal recessive condition known as MUTYH-associated adenomatous polyposis and consequently increase the risk of colorectal cancer. However, reports of increased cancer risk in individuals carrying only one defective MUTYH allele are controversial and based on studies involving few individuals. Here, we describe a comprehensive investigation of monoallelic pathogenic MUTYH germline variants in 10,389 cancer patients across 33 different tumour types and 117,000 healthy individuals. Our results indicate that monoallelic pathogenic MUTYH germline variants can lead to tumorigenesis through a mechanism of somatic loss of heterozygosity of the functional MUTYH allele in the tumour. We confirmed that the frequency of monoallelic pathogenic MUTYH germline variants is higher in individuals with cancer than in the general population, although this frequency is not homogeneous among tumour types. We also demonstrated that the MUTYH mutational signature is present only in tumours with loss of the functional allele and found that the characteristic MUTYH base substitution (C>A) increases stop-codon generation. We identified key genes that are affected during tumorigenesis. In conclusion, we propose that carriers of the monoallelic pathogenic MUTYH germline variant are at a higher risk of developing tumours, especially those with frequent loss of heterozygosity events, such as adrenal adenocarcinoma, although the overall risk is still low. © 2021 The Pathological Society of Great Britain and Ireland. Published by John Wiley & Sons, Ltd.

A Monoallelic Variant in REST Is Associated with Non-Syndromic Autosomal Dominant Hearing Impairment in a South African Family.

Hearing impairment (HI) is a sensory disorder with a prevalence of 0.0055 live births in South Africa. DNA samples from a South African family presenting with progressive, autosomal dominant non-syndromic HI were subjected to whole-exome sequencing, and a novel monoallelic variant in REST [c.1244GC; p.(C415S)], was identified as the putative causative variant. The co-segregation of the variant was confirmed with Sanger Sequencing. The variant is absent from databases, 103 healthy South African controls, and 52 South African probands with isolated HI. In silico analysis indicates that the p.C415S variant in REST substitutes a conserved cysteine and results in changes to the surrounding secondary structure and the disulphide bonds, culminating in alteration of the tertiary structure of REST. Localization studies using ectopically expressed GFP-tagged Wild type (WT) and mutant REST in HEK-293 cells show that WT REST localizes exclusively to the nucleus; however, the mutant protein localizes throughout the cell. Additionally, mutant REST has an impaired ability to repress its known target AF1q. The data demonstrates that the identified mutation compromises the function of REST and support its implication in HI. This study is the second report, worldwide, to implicate REST in HI and suggests that it should be included in diagnostic HI panels.

Prevalence and Characterization of Biallelic and Monoallelic NTHL1 and MSH3 Variant Carriers From a Pan-Cancer Patient Population.

Patients with pan-cancer (n = 11,081) underwent matched tumor-normal sequencing with consent for germline analysis. Medical records and tumors were reviewed and analyzed. Prevalence of PVs was compared with reference controls (Genome Aggregation Database). NTHL1-PVs were identified in 40 patients including 39 monoallelic carriers (39/11,081 = 0.35%) and one with biallelic variants (1/11,081 = 0.009%) and a diagnosis of isolated early-onset breast cancer. NTHL1-associated mutational signature 30 was identified in the tumors of the biallelic patient and two carriers. Colonic polyposis was not identified in any NTHL1 patient. MSH3-PVs were identified in 13 patients, including 12 monoallelic carriers (12/11,081 = 0.11%) and one with biallelic MSH3 variants (1/11,081 = 0.009%) and diagnoses of later-onset cancers, attenuated polyposis, and abnormal MSH3-protein expression. Of the 12 MSH3 carriers, two had early-onset cancer diagnoses with tumor loss of heterozygosity of the wild-type MSH3 allele. Ancestry-specific burden tests demonstrated that NTHL1 and MSH3 prevalence was not significantly different in this pan-cancer population versus controls. NTHL1 and MSH3 germline alterations were not enriched in this pan-cancer patient population. However, tumor-specific findings, such as mutational signature 30 and loss of heterozygosity of the wild-type allele, suggest the potential contribution of monoallelic variants to tumorigenesis in a subset of patients.

A recurrent familial partial lipodystrophy due to a monoallelic or biallelic LMNA founder variant highlights the multifaceted cardiac manifestations of metabolic laminopathies.

LMNA-linked familial partial lipodystrophy type 2 (FPLD2) leads to insulin resistance-associated metabolic complications and cardiovascular diseases. We aimed to characterise the disease phenotype in a cohort of patients carrying an LMNA founder variant. We collected clinical and biological data from patients carrying the monoallelic or biallelic LMNA p.(Thr655Asnfs*49) variant (n = 65 and 13, respectively) and 19 non-affected relative controls followed-up in Reunion Island Lipodystrophy Competence Centre, France. Two-thirds of patients with FPLD2 (n = 51) and one-third of controls (n = 6) displayed lipodystrophy and/or lean or android morphotype (P = 0.02). Although age and BMI were not statistically different between the two groups, the insulin resistance index (median HOMA-IR: 3.7 vs 1.5, P = 0.001), and the prevalence of diabetes, dyslipidaemia, and non-alcoholic fatty liver disease were much higher in patients with FPLD2 (51.3 vs 15.8%, 83.3 vs 42.1%, and 83.1 vs 33.3% (all P ≤ 0.01), respectively). Atherosclerosis tended to be more frequent in patients with FPLD2 (P = 0.07). Compared to heterozygous, homozygous patients displayed more severe lipoatrophy and metabolic alterations (lower BMI, fat mass, leptin and adiponectin, and higher triglycerides P ≤ 0.03) and tended to develop diabetes more frequently, and earlier (P = 0.09). Dilated cardiomyopathy and/or rhythm/conduction disturbances were the hallmark of the disease in homozygous patients, leading to death in four cases. The level of expression of the LMNA 'Reunionese' variant determines the severity of both lipoatrophy and metabolic complications. It also modulates the cardiac phenotype, from atherosclerosis to severe cardiomyopathy, highlighting the need for careful cardiac follow-up in affected patients.

Novel monoallelic variant in ERLIN2 causes spastic paraplegia converted to amyotrophic lateral sclerosis

Hereditary spastic paraplegias (HSPs) are a group of inherited neurologic disorders characterized by corticospinal tract and dorsal column degeneration. Endoplasmic reticulum lipid raft-associated protein 2 (ERLIN2) has been identified as a causative gene of the autosomal recessive HSP family [1,2]. However, only one report has shown that ERLIN2 also causes amyotrophic lateral sclerosis (ALS) [3]; thus, further investigations are needed for its confirmation. Herein we report a family of HSP converted to ALS caused by a novel monoallelic variant in ERLIN2.

Risk of colorectal adenomas and cancer in monoallelic carriers of MUTYH pathogenic variants: a single-centre experience

PurposeThe carrier frequency of MUTYH pathogenic variants in the population may be as high as one in 45. Some studies have found an increased risk of colorectal cancer (CRC) in monoallelic carriers of MUTYH pathogenic variants, but the role of early surveillance colonoscopy is not conclusive. This study aimed to assess the outcomes of colonoscopy surveillance in MUTYH carriers.MethodsPatients, with a monoallelic pathogenic variant in MUTYH, found at cascade testing, were identified from the St Mark’s Hospital Polyposis Registry database. Findings at surveillance colonoscopy were reviewed.ResultsTwo hundred and forty-nine carriers were identified, of whom 125 had undergone at least one surveillance colonoscopy. Twenty-eight patients (22%) developed at least one adenoma; all adenomas had low-grade dysplasia (LGD). The median age at first colonoscopy was 36 years (range 16–75 years). The median age at first adenoma detection was 43 years (range 22–75 years). The cumulative incidence of adenoma development by age 30, 40, 50, 60 and 70 years was 3.2%, 8.8%, 15.2%, 18.4% and 20.8%, respectively. No CRCs were observed.ConclusionsOur cohort of monoallelic carriers of MUTYH pathogenic variants is a relatively younger group than adults entering population screening colonoscopy, but a high adenoma rate was not observed. No CRCs were detected, suggesting that current guidance that these individuals should be managed in the same way as the general population is reasonable.

PRDX1 gene-related epi-cblC disease is a common type of inborn error of cobalamin metabolism with mono- or bi-allelic MMACHC epimutations

BackgroundThe role of epigenetics in inborn errors of metabolism (IEMs) is poorly investigated. Epigenetic changes can contribute to clinical heterogeneity of affected patients but could also be underestimated determining factors in the occurrence of IEMs. An epigenetic cause of IEMs has been recently described for the autosomal recessive methylmalonic aciduria and homocystinuria, cblC type (cblC disease), and it has been named epi-cblC. Epi-cblC has been reported in association with compound heterozygosity for a genetic variant and an epimutation at the MMACHC locus, which is secondary to a splicing variant (c.515-1G > T or c.515-2A > T) at the adjacent PRDX1 gene. Both these variants cause aberrant antisense transcription and cis-hypermethylation of the MMACHC gene promotor with subsequent silencing. Until now, only nine epi-cblC patients have been reported.MethodsWe report clinical/biochemical assessment, MMACHC/PRDX1 gene sequencing and genome-wide DNA methylation profiling in 11 cblC patients who had an inconclusive MMACHC gene testing. We also compare clinical phenotype of epi-cblC patients with that of canonical cblC patients.ResultsAll patients turned out to have the epi-cblC disease. One patient had a bi-allelic MMACHC epimutation due to the homozygous PRDX1:c.515-1G > T variant transmitted by both parents. We found that the bi-allelic epimutation produces the complete silencing of MMACHC in the patient’s fibroblasts. The remaining ten patients had a mono-allelic MMACHC epimutation, due to the heterozygous PRDX1:c.515-1G > T, in association with a mono-allelic MMACHC genetic variant. Epi-cblC disease has accounted for about 13% of cblC cases diagnosed by newborn screening in the Tuscany and Umbria regions since November 2001. Comparative analysis showed that clinical phenotype of epi-cblC patients is similar to that of canonical cblC patients.ConclusionsWe provide evidence that epi-cblC is an underestimated cause of inborn errors of cobalamin metabolism and describe the first instance of epi-cblC due to a bi-allelic MMACHC epimutation. MMACHC epimutation/PRDX1 mutation analyses should be part of routine genetic testing for all patients presenting with a metabolic phenotype that combines methylmalonic aciduria and homocystinuria.

In-house multiplex ligation-dependent probe amplification assay for citrin deficiency: analytical validation and novel exonic deletions in SLC25A13.

Citrin deficiency is one of the most common inborn errors of metabolism in East Asians, which may manifest as neonatal cholestasis, failure to thrive and dyslipidaemia, or recurrent hyperammonaemic encephalopathy. Its molecular diagnosis requires confirmation of the presence of biallelic pathogenic variants in SLC25A13 gene by sequencing, and analysis for a common insertion IVS16ins3kb. However, patients with compatible biochemical features but only one monoallelic pathogenic variant have remained a diagnostic challenge. Here we report the development, validation and application of a multiplex ligation-dependent probe amplification (MLPA) assay using an in-house oligonucleotide probemix and a customised Coffalyer.NET worksheet for detection of exonic copy number variations in SLC25A13. With this MLPA assay, we successfully identified the presence of a heterozygous exonic deletion in SLC25A13 in three of 15 (20%) unrelated individuals with only one monoallelic pathogenic variant detected using conventional methods. Three exonic deletions, two novel involving exon 14 and one reported involving exon 5, were subsequently confirmed with Sanger sequencing. In summary, we developed, evaluated, and demonstrated the clinical utility of an in-house MLPA assay to look for exonic deletions in SLC25A13 in patients with citrin deficiency. With the discovery of novel deletions, MLPA should be considered a test of choice for molecular diagnosis of citrin deficiency when the sequencing result is inconclusive.

Genetic Study in a Cohort of Children With ROHHAD Syndrome

Abstract Introduction: Rapid-onset obesity, hypoventilation, hypothalamic dysfunction and autonomic dysregulation (ROHHAD) is a rare syndrome beginning at 3-6 years of age with approximately 150 cases described. Additional features include eye abnormalities, neurobehavioral dysfunction and paraneoplastic tumors. The etiology of the complex phenotype remains unknown. Methods: This study aims to investigate the genetic landscape of this complex phenotype by whole exome sequencing (WES) and copy number variation (CNV) analysis. We recruited 33 families (27 trios, 1 duo and 5 singletons) with a proband with ROHHAD syndrome (Ize-Ludlow 2007, Pediatrics). WES of 89 individuals was performed at the Center for Mendelian Genomics, Broad Institute. The Illumina platform with a mean coverage of ~100X (&amp;gt; 90% targets 20x) and Infinium Global Screening Array BeadChip 24v1.0 were used. Results: This report includes 28 probands (female = 18, 64%) with rapid onset obesity (100%), hypoventilation (88%), hypothalamic dysfunction (69%), eye disorders (62%) and neurobehavioral abnormalities (76%). Neuroendocrine tumor, ganglioneuroblastoma, was present in 38% (n=13). No unifying causative single gene or CNV was identified, but a number of sequence variants are prioritized. ARNT2, which encodes for a helix-loop-helix transcription factor, plays a role in the development of the hypothalamic-pituitary axis, postnatal brain growth, and visual and renal function. The de novo monoallelic missense variant was found in a 14-year old white girl (BMIz +3.25) with extreme obesity and a neurobehavioral phenotype. OCRL1, a multi-domain protein involved in cytoskeleton-plasma membrane adhesion, endosomal trafficking and in primary cilium assembly. Mutations in this gene have also been known to cause Lowe syndrome. A hemizygous X-linked frameshift variant in a 5-year old white boy with extreme obesity (BMIz +5.48), central hypoventilation neurobehavioral dysfunction and ganglioneuroblastoma. A monoallelic missense variant in NSD1, a transcriptional intermediary factor acting as a histone methyltransferase, was identified in a 8-year old Hispanic girl with severe obesity (BMIz +2.91), neurobehavioral disorder, pituitary and eye dysfunction and ganglioneuroblastoma. NSD1 is known to cause Sotos and Beckwith-Wiedemann. Compound heterozygous variants in KIF7, a key component of the Hedgehog signaling pathway, were identified in a 14-year old white girl with severe obesity (BMIz +3.00), autistic behavior, pituitary dysfunction and central hypoventilation. This gene is known to cause autosomal recessive hydrolethalis and acroscallosal syndromes with mutations also noted in Bardet-Biedl, Meckel and Joubert syndromes. Conclusion: While no unifying genetic cause has been identified in ROHHAD syndrome, it is possible that the phenotype represents a collection of complex genetic syndromes.