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A couple of Ashkenazi Jewish descent was referred for an early anatomy scan at 14 + 2 weeks of gestation following a previous pregnancy termination due to posterior encephalocele and enlarged kidneys. The index pregnancy was also positive for several fetal abnormalities, including enlarged kidneys with cystic dysplasia and abnormal cerebellar morphology highly suggestive of Joubert syndrome.Trio exome sequencing revealed compound heterozygosity for variants in the TMEM67 gene: a known pathogenic maternally inherited variant found in trans with a paternal intronic variant of unknown significance. RNA analysis revealed that the intronic variant creates a cryptic acceptor splice site in intron 12, leading to the insertion of 22 bp and causing a frameshift with a premature stop codon. This analysis enabled the reclassification of the intronic variant to likely pathogenic.This information empowered the couple to make informed reproductive choices and opt for preimplantation genetic testing (PGT) for future pregnancies.

AOA2 is a rare progressive adolescent-onset disease characterised by cerebellar vermis atrophy, peripheral neuropathy and elevated serum alpha-fetoprotein (AFP) caused by pathogenic bi-allelic variants in SETX, encoding senataxin, involved in DNA repair and RNA maturation. Sanger sequencing of genomic DNA, co-segregation and oxidative stress functional studies were performed in Family 1. Trio whole-exome sequencing (WES), followed by SETX RNA and qRT-PCR analysis, were performed in Family 2. Sanger sequencing in Family 1 revealed two novel in-frame SETX deletion and duplication variants in trans (c.7009_7011del; p.Val2337del and c.7369_7371dup; p.His2457dup). Patients had increased induced chromosomal aberrations at baseline and following exposure to higher mitomycin-C concentration and increased sensitivity to oxidative stress at the lower mitomycin-C concentration in cell viability test. Trio WES in Family 2 revealed two novel SETX variants in trans, a nonsense variant (c.568C T; p.Gln190*), and a deep intronic variant (c.5549-107A G). Intronic variant analysis and SETX mRNA expression revealed activation of a cryptic exon introducing a premature stop codon (p.Met1850Lysfs*18) and resulting in aberrant splicing, as shown by qRT-PCR analysis, thus leading to higher levels of cryptic exon activation. Along with a second deleterious allele, this variant leads to low levels of SETX mRNA and disease manifestations. Our report expands the phenotypic spectrum of AOA2. Results provide initial support for the hypomorphic nature of the novel in-frame deletion and duplication variants in Family 1. Deep-intronic variant analysis of Family 2 variants potentially reveals a previously undescribed poison exon in the SETX gene, which may contribute to tailored therapy development.

The complement system regulator CD55 was initially found to carry the Cromer blood group system antigens, and its complete loss of function was subsequently revealed to cause a severe monogenic gastrointestinal syndrome characterized by protein-losing enteropathy and susceptibility to venous thrombosis. Here we present homozygosity to the CD55 c.596CT; p.Ser199Leu variant, which was previously described as the Cromer Dr(a-) genotype, in two Bukharan Jewish CD55-deficiency patients with variable disease severity. We confirm that this missense variant causes aberrant splicing and deletion of 44 bp in exon 5, leading to premature termination and low expression of the CD55 protein. Furthermore, Patient 1 exhibited a mildly abnormal B cell immunophenotyping profile. By population screening we established that this variant is highly prevalent in the Bukharan Jewish population, with a carrier frequency of 1:17, suggesting that many similar patients are un- or mis-diagnosed. The phenotypic variability, ranging from abdominal pain when eating a high-fat diet to the full CD55-deficiency phenotype, is likely related to modifiers affecting the proportion of the variant that is able to escape aberrant splicing. Establishing the diagnosis of CD55-deficiency in a timely manner, even in patients with milder symptoms, may have a critical effect on their management and quality-of-life since treatment with the complement inhibitor eculizumab is highly effective in ameliorating disease manifestations. Awareness of founder mutations within certain populations can further guide genetic testing and prevent a diagnostic odyssey, by placing this CD55 variant high on the differential diagnosis.

BACKGROUND & AIMS: Constitutional mismatch repair deficiency (CMMRD) is a rare recessive childhood cancer predisposition syndrome caused by germline mismatch repair variants. Constitutional microsatellite instability (cMSI) is a CMMRD diagnostic hallmark and may associate with cancer risk. We quantified cMSI in a large CMMRD patient cohort to explore genotype-phenotype correlations using novel MSI markers selected for instability in blood.METHODS: Three CMMRD, 1 Lynch syndrome, and 2 control blood samples were genome sequenced to >120× depth. A pilot cohort of 8 CMMRD and 38 control blood samples and a blinded cohort of 56 CMMRD, 8 suspected CMMRD, 40 Lynch syndrome, and 43 control blood samples were amplicon sequenced to 5000× depth. Sample cMSI score was calculated using a published method comparing microsatellite reference allele frequencies with 80 controls.RESULTS: Thirty-two mononucleotide repeats were selected from blood genome and pilot amplicon sequencing data. cMSI scoring using these MSI markers achieved 100% sensitivity (95% CI, 93.6%-100.0%) and specificity (95% CI 97.9%-100.0%), was reproducible, and was superior to an established tumor MSI marker panel. Lower cMSI scores were found in patients with CMMRD with MSH6 deficiency and patients with at least 1 mismatch repair missense variant, and patients with biallelic truncating/copy number variants had higher scores. cMSI score did not correlate with age at first tumor.CONCLUSIONS: We present an inexpensive and scalable cMSI assay that enhances CMMRD detection relative to existing methods. cMSI score is associated with mismatch repair genotype but not phenotype, suggesting it is not a useful predictor of cancer risk.

BackgroundWe previously reported inborn errors of TLR3- and TLR7-dependent type I interferon (IFN) immunity in 1-5% of unvaccinated patients with life-threatening COVID-19, and auto-antibodies against type I IFN in another 15-20% of cases.MethodsWe report here a genome-wide rare variant burden association analysis in 3,269 unvaccinated patients with life-threatening COVID-19 (1,301 previously reported and 1,968 new patients), and 1,373 unvaccinated SARS-CoV-2-infected individuals without pneumonia. A quarter of the patients tested had antibodies against type I IFN (234 of 928) and were excluded from the analysis.ResultsNo gene reached genome-wide significance. Under a recessive model, the most significant gene with at-risk variants wasTLR7, with an OR of 27.68 (95%CI:1.5-528.7,P=1.1×10−4), in analyses restricted to biochemically loss-of-function (bLOF) variants. We replicated the enrichment in rare predicted LOF (pLOF) variants at 13 influenza susceptibility loci involved in TLR3-dependent type I IFN immunity (OR=3.70 [95%CI:1.3-8.2],P=2.1×10−4). Adding the recently reportedTYK2COVID-19 locus strengthened this enrichment, particularly under a recessive model (OR=19.65 [95%CI:2.1-2635.4];P=3.4×10−3). When these 14 loci andTLR7were considered, all individuals hemizygous (n=20) or homozygous (n=5) for pLOF or bLOF variants were patients (OR=39.19 [95%CI:5.2-5037.0],P=4.7×10−7), who also showed an enrichment in heterozygous variants (OR=2.36 [95%CI:1.0-5.9],P=0.02). Finally, the patients with pLOF or bLOF variants at these 15 loci were significantly younger (mean age [SD]=43.3 [20.3] years) than the other patients (56.0 [17.3] years;P=1.68×10−5).ConclusionsRare variants of TLR3- and TLR7-dependent type I IFN immunity genes can underlie life-threatening COVID-19, particularly with recessive inheritance, in patients under 60 years old.

Since the arrival of severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) in December 2019, its characterization as a novel human pathogen, and the resulting coronavirus disease 2019 (COVID-19) pandemic, over 6.5 million people have died worldwide—a stark and sobering reminder of the fundamental and nonredundant roles of the innate and adaptive immune systems in host defense against emerging pathogens. Inborn errors of immunity (IEI) are caused by germline variants, typically in single genes. IEI are characterized by defects in development and/or function of cells involved in immunity and host defense, rendering individuals highly susceptible to severe, recurrent, and sometimes fatal infections, as well as immune dysregulatory conditions such as autoinflammation, autoimmunity, and allergy. The study of IEI has revealed key insights into the molecular and cellular requirements for immune-mediated protection against infectious diseases. Indeed, this has been exemplified by assessing the impact of SARS-CoV-2 infection in individuals with previously diagnosed IEI, as well as analyzing rare cases of severe COVID-19 in otherwise healthy individuals. This approach has defined fundamental aspects of mechanisms of disease pathogenesis, immunopathology in the context of infection with a novel pathogen, and therapeutic options to mitigate severe disease. This review summarizes these findings and illustrates how the study of these rare experiments of nature can inform key features of human immunology, which can then be leveraged to improve therapies for treating emerging and established infectious diseases.

Eliglustat, an oral substrate reduction therapy, is approved for eligible adults with Gaucher disease type 1. In the Phase 3 ENGAGE trial of previously untreated adults with Gaucher disease type 1, eliglustat‐treated patients had statistically significant improvements in organ volumes and hematologic parameters compared with placebo in the 9‐month primary analysis. We report final outcomes by time on eliglustat among all patients who participated in the ENGAGE trial and extension. No patient deteriorated clinically or withdrew due to adverse events; 39/40 patients entered the open‐label extension period and 34/40 (85%) remained in the trial until completion or switching to commercial eliglustat after its approval (2.3–6 years). Clinically meaningful improvements in Gaucher disease manifestations were seen in all patients concomitant with reductions in pathological lipid substrate levels (glucosylceramide and glucosylsphingosine). Among patients with 4.5 years of eliglustat exposure, mean spleen volume decreased by 66% (from 17.1 to 5.8 multiples of normal [MN], n = 13), mean liver volume decreased by 23% (from 1.5 to 1.1 MN, n = 13), mean hemoglobin increased 1.4 g/dl (from 11.9 to 13.4 g/dl, n = 12), mean platelet count increased by 87% (from 67.6 to 122.6 × 109/L, n = 12), median chitotriosidase decreased by 82% (from 13 394 to 2312 nmol/h/ml, n = 11), median glucosylceramide decreased by 79% (from 11.5 to 2.4 μg/ml, n = 11), median glucosylsphingosine decreased by 84% (from 518.5 to 72.1 ng/ml, n = 10), and mean spine T‐score increased from −1.07 (osteopenia) to −0.53 (normal) (n = 9). The magnitude of improvement in Gaucher disease manifestations and biomarkers over time was similar among the full trial cohort. Eliglustat was well‐tolerated and led to clinically significant improvements in previously untreated patients with Gaucher disease type 1 during 4.5 years of treatment.

Tel Hashomer camptodactyly syndrome is a long-known entity characterized by camptodactyly with muscular hypoplasia, skeletal dysplasia, and abnormal palmar creases. Currently, the genetic basis for this disorder is unknown, thus there is a possibility that this clinical presentation may be contained within another genetic diagnosis. Here, we present a multiplex family with a previous clinical diagnosis of Tel Hashomer camptodactyly syndrome. Whole exome sequencing and pedigree-based analysis revealed a novel hemizygous truncating variant c.269_270dup (p.Phe91Alafs*34) in the FGD1 gene (NM_004463.3) in all three symptomatic patients, congruous with a diagnosis of Aarskog-Scott syndrome. Our report adds to the limited data on Aarskog-Scott syndrome, and emphasizes the importance of unbiased comprehensive molecular testing toward establishing a diagnosis for genetic syndromes with unknown genetic basis.

Germline pathogenic variants (PVs) in BRCA1/BRCA2 are well-established risk factors for breast cancer (BC) and/or ovarian cancer (OC). Founder PVs have been described in BRCA1/ BRCA2 in several genetic isolates. The Christian Arab population in the Middle East is a relatively isolated ethnic group, yet founder, or recurrent BRCA1/BRCA2 PVs have not been reported in this population. In this study we describe PVs detected in cancer susceptibility genes among a cohort of Christian Arabs from Israel. We reviewed patient records from the Oncogenetic clinic at Rambam Health Care Campus during the years 2013- mid 2020. Thirty-five unrelated Christian Arab patients, with personal or family history of BC and/or OC underwent BRCA1/BRCA2 (14/35) testing or cancer gene panel testing (21/35) as part of their diagnostic workup. Three clinically significant variants in BRCA2, CHEK2 and RAD51C were found in 7/35 patients (20%). A recurrent duplication of the BRCA2 genomic region, encompassing exons 5-10 and the 5' portion of exon 11, was found in 5/33 (15.2%) patients for whom copy number variants (CNVs) analysis was performed. We identified a recurrent pathogenic BRCA2 duplication in Christian Arab patients with a personal/ family history of BC and/or OC. Our findings emphasize the importance of inclusion of CNVs analysis in BRCA1/BRCA2 genetic testing, and specifically for Christian Arab patients suspected of hereditary BC and/or OC.

RBL2/p130, a member of the retinoblastoma family of proteins, is a key regulator of cell division and propagates irreversible senescence. RBL2/p130 is also involved in neuronal differentiation and survival, and eliminating Rbl2 in certain mouse strains leads to embryonic lethality accompanied by an abnormal central nervous system (CNS) phenotype. Conflicting reports exist regarding a role of RBL2/p130 in transcriptional regulation of DNA methyltransferases (DNMTs), as well as the control of telomere length. Here we describe the phenotype of three patients carrying bi-allelic RBL2-truncating variants. All presented with infantile hypotonia, severe developmental delay and microcephaly. Malignancies were not reported in carriers or patients. Previous studies carried out on mice and human cultured cells, associated RBL2 loss to DNA methylation and telomere length dysregulation. Here, we investigated whether patient cells lacking RBL2 display related abnormalities. The study of primary patient fibroblasts did not detect abnormalities in expression of DNMTs. Furthermore, methylation levels of whole genome DNA, and specifically of pericentromeric repeats and subtelomeric regions, were unperturbed. RBL2-null fibroblasts show no evidence for abnormal elongation by telomeric recombination. Finally, gradual telomere shortening, and normal onset of senescence were observed following continuous culturing of RBL2-mutated fibroblasts. Thus, this study resolves uncertainties regarding a potential non-redundant role for RBL2 in DNA methylation and telomere length regulation, and indicates that loss of function variants in RBL2 cause a severe autosomal recessive neurodevelopmental disorder in humans.

Feingold syndrome 1 (FGLDS1) is an autosomal dominant malformation syndrome, characterized by skeletal anomalies, microcephaly, facial dysmorphism, gastrointestinal atresias and learning disabilities. Mutations in the MYCN gene are known to be the cause of this syndrome. Congenital absence of the flexor pollicis longus (CAFPL) tendon is a rare hand anomaly. Most cases are sporadic and no genetic variants have been described associated with this abnormality. We describe here a pedigree combining familial CAFPL tendon as a feature of FGLDS1. Molecular analyses of whole exome sequence data in five affected family members spanning three generations of this family revealed a novel mutation in the MYCN gene (c.1171C>T; p.Arg391Cys). Variants in MYCN have not been published in association with isolated or syndromic CAFPL tendon, nor has this been described as a skeletal feature of Feingold syndrome. This report expands on the clinical and molecular spectrum of MYCN-related disorders and highlights the importance of MYCN protein in normal human thumb and foramen development.

Mutations in more than 150 genes are responsible for inherited hearing loss, with thousands of different, severe causal alleles that vary among populations. The Israeli Jewish population includes communities of diverse geographic origins, revealing a wide range of deafness-associated variants and enabling clinical characterization of the associated phenotypes. Our goal was to identify the genetic causes of inherited hearing loss in this population, and to determine relationships among genotype, phenotype, and ethnicity. Genomic DNA samples from informative relatives of 88 multiplex families, all of self-identified Jewish ancestry, with either non-syndromic or syndromic hearing loss, were sequenced for known and candidate deafness genes using the HEar-Seq gene panel. The genetic causes of hearing loss were identified for 60% of the families. One gene was encountered for the first time in human hearing loss: ATOH1 (Atonal), a basic helix-loop-helix transcription factor responsible for autosomal dominant progressive hearing loss in a five-generation family. Our results demonstrate that genomic sequencing with a gene panel dedicated to hearing loss is effective for genetic diagnoses in a diverse population. Comprehensive sequencing enables well-informed genetic counseling and clinical management by medical geneticists, otolaryngologists, audiologists, and speech therapists and can be integrated into newborn screening for deafness.

Agenesis of the corpus callosum (ACC) is a common prenatally-detected brain anomaly. Recently, an association between mutations in the DCC Netrin 1 receptor (DCC) gene and ACC, with or without mirror movements, has been demonstrated. In this manuscript, we present a family with a novel heterozygous frameshift mutation in DCC, review the available literature, and discuss the challenges involved in the genetic counseling for recently discovered disorders with paucity of medical information. We performed whole exome sequencing in a healthy nonconsanguineous couple that underwent two pregnancy terminations due to prenatal diagnosis of ACC. A heterozygous variant c.2774dupA (p.Asn925Lysfs*17) in the DCC gene was demonstrated in fetal and paternal DNA samples, as well as in a healthy 4-year-old offspring. When directly questioned, both father and child reported having mirror movements not affecting quality of life. Segregation analysis demonstrated the variant in three paternal siblings, two of them having mirror movements. Brain imaging revealed normal corpus callosum. Summary of literature data describing heterozygous loss-of-function variants in DCC (n = 61) revealed 63.9% penetrance for mirror movements, 9.8% for ACC, and 5% for both. No significant neurodevelopmental abnormalities were reported among the seven published patients with DCC loss-of-function variants and ACC. Prenatal diagnosis of ACC should prompt a specific anamnesis regarding any neurological disorder, as well as intentional physical examination of both parents aimed to detect mirror movements. In suspicious cases, detection of DCC pathogenic variants might markedly improve the predicted prognosis, alleviate the parental anxiety, and possibly prevent pregnancy termination.

Gaucher disease (GD) is the most prevalent lysosomal disorder caused by GBA mutations and abnormal glucocerebrosidase function, leading to glucocerebrosideaccumulation mainly in the liver, spleen, bone marrow, lungs, and occasionally in the central nervous system. Gaucher disease type 3c (GD3c) is a rare subtype of the subacute/chronic neuronopathic GD3, caused by homozygosity for the GBA p.Asp448His (D409H) mutation. GD3c is characterized mainly by cardiovascular and neuro-ophthalmological findings. In this paper, we describe four new GD3c patients exhibiting rare cardiovascular, pulmonary and psychiatric findings, as well as atypical disease courses. Review of the GD3c-related literature revealed clinical descriptions of 36 patients, presenting predominantly with cardiovascular calcifications; 15%, including Patient 1b in this study, had non-calcified lesions - fibrosis and atherosclerosis. Only 7.5% of patients have been described without heart disease, including Patient 3; however, Patient 2 had a fulminant coronary disease. Neurological findings in GD3c consist mainly of oculomotor apraxia (80%), which is absent in Patient 3, while other neurological findings are common (65%) but diverse. Patient 1b developed a psychiatric behavioral disorder, which has not been previously described in GD3c. Patient 1b also had interstitial lung disease, which was only described in one GD3c patient as pulmonary fibrosis. In view of these unique features, we recommend a revised surveillance protocol; however, further studies are required to establish the management of these patients and the role of GBA in the described pathologies.

This study aimed to characterize the clinical phenotype, genetic basis, and consequent immunological phenotype of a boy with severe infantile-onset colitis and eosinophilic gastrointestinal disease, and no evidence of recurrent or severe infections. Trio whole-exome sequencing (WES) was utilized for pathogenic variant discovery. Western blot (WB) and immunohistochemical (IHC) staining were used for protein expression analyses. Immunological workup included in vitro T cell studies, flow cytometry, and CyTOF analysis. WES revealed a homozygous variant in the capping protein regulator and myosin 1 linker 2 (CARMIL2) gene: c.1590C>A; p.Asn530Lys which co-segregated with the disease in the nuclear family. WB and IHC analyses demonstrated reduced protein levels in patient’s cells compared with controls. Moreover, comprehensive immunological workup revealed severely diminished blood-borne regulatory T cell (Treg) frequency and impaired in vitro CD4+ T cell proliferation and Treg generation. CyTOF analysis showed significant shifts in the patient’s innate and adaptive immune cells compared with healthy controls and ulcerative colitis patients. Pathogenic variants in CARMIL2 have been implicated in an immunodeficiency syndrome characterized by recurrent infections, occasionally with concurrent chronic diarrhea. We show that CARMIL2-immunodeficiency is associated with significant alterations in the landscape of immune populations in a patient with prominent gastrointestinal disease. This case provides evidence that CARMIL2 should be a candidate gene when diagnosing children with very early onset inflammatory and eosinophilic gastrointestinal disorders, even when signs of immunodeficiency are not observed.

Translation of mitochondrial-specific DNA is required for proper mitochondrial function and energy production. For this purpose, an elaborate network of dedicated molecular machinery including initiation, elongation and termination factors exists. We describe a patient with an unusual phenotype and a novel homozygous missense variant in TUFM (c.344A>C; p.His115Pro), encoding mtDNA translation elongating factor Tu (EFTu). To date, only four patients have been reported with bi-allelic mutations in TUFM, leading to combined oxidative phosphorylation deficiency 4 (COXPD4) characterized by severe early-onset lactic acidosis and progressive fatal infantile encephalopathy. The patient presented here expands the phenotypic features of TUFM-related disease, exhibiting lactic acidosis and dilated cardiomyopathy without progressive encephalopathy. This warrants the inclusion of TUFM in differential diagnosis of metabolic cardiomyopathy. Cases that further refine genotype-phenotype associations and characterize the molecular basis of mitochondrial disorders allow clinicians to predict disease prognosis, greatly impacting patient care, as well as provide families with reproductive planning options.

BackgroundChromosomal instability, as reflected by structural or copy-number changes, is a known cancer characteristic but are rarely observed in healthy tissue. Mutations in DNA repair genes disrupt the maintenance of DNA integrity and predispose to hereditary cancer syndromes.ObjectiveTo clinically characterise and genetically diagnose two reportedly unrelated patients with unique cancer syndromes, including multiorgan tumourogenesis (patient 1) and early-onset acute myeloid leukaemia (patient 2), both displaying unique peripheral blood karyotypes.MethodsGenetic analysis in patient 1 included TruSight One panel and whole-exome sequencing, while patient 2 was diagnosed by FoundationOne Heme genomic analysis; Sanger sequencing was used for mutation confirmation in both patients. Karyotype analysis was performed on peripheral blood, bone marrow and other available tissues.ResultsBoth patients were found homozygous for CHEK2 c.499G>A; p.Gly167Arg and exhibited multiple different chromosomal translocations in 30%–60% peripheral blood lymphocytes. This karyotype phenotype was not observed in other tested tissues or in an ovarian cancer patient with a different homozygous missense mutation in CHEK2 (c.1283C>T; p.Ser428Phe).ConclusionsThe multiple chromosomal translocations in patient lymphocytes highlight the role of CHK2 in DNA repair. We suggest that homozygosity for p.Gly167Arg increases patients' susceptibility to non-accurate correction of DNA breaks and possibly explains their increased susceptibility to either multiple primary tumours during their lifetime or early-onset tumourigenesis.

Loss of the complement inhibitor CD55 leads to a syndrome of early-onset protein-losing enteropathy (PLE), associated with intestinal lymphangiectasia and susceptibility to large-vein thrombosis. The in vitro and short-term treatment benefits of eculizumab (C5-inhibitor) therapy for CD55-deficiency have been previously demonstrated. Here we present the 18-months treatment outcomes for 3 CD55-deficiency patients with sustained therapeutic response.Three CD55-deficiency patients received off-label eculizumab treatment. Clinical and laboratory treatment outcomes included frequency and consistency of bowl movements, weight, patient/parent reports of overall well-being, and serum albumin and total protein levels. Membrane attack complex deposition on leukocytes was tested by flow cytometry, before and during eculizumab treatment.Marked clinical improvement was noted in all 3 patients with resolution of PLE manifestations, that is, diarrhea, edema, malabsorption, overall well-being, growth, and quality of life. In correlation with the clinical observations, we observed progress in all laboratory outcome parameters, including increase in albumin and total protein levels, and up to 80% reduction in membrane attack complex deposition on leukocytes (P0.001). The progress persisted over 18 months of treatment without any severe adverse events.CD55-deficiency patients present with early-onset diarrhea, edema, severe hypoalbuminemia, abdominal pain, and malnutrition. Targeted therapy with the terminal complement inhibitor eculizumab has positive clinical and laboratory outcomes in PLE related to CD55 loss-of-function mutations, previously a life-threatening condition. Our results demonstrate the potential of genetic diagnosis to guide tailored treatment, and underscore the significant role of the complement system in the intestine.

Gaucher disease (GD) is an autosomal recessive disease characterized by the buildup of glucocerebrosides in macrophages, resulting in the formation of “Gaucher cells.” These cells predominantly infiltrate the liver, spleen, and bone marrow leading to hepatosplenomegaly, cytopenia, and bone pain. Anemia in GD is typically considered to result from non-hemolytic processes. Although rare, a higher rate of hemolytic anemia of the autoimmune type has been reported in GD than in the general population. The literature on non-immune hemolytic anemia in GD is scarce. We review the literature on hemolytic anemia in GD and report on a case of non-immune hemolytic anemia secondary to GD. We believe this is the first description of a patient with confirmed GD and symptomatic non-immune hemolytic anemia that responded to GD-specific treatment.

Circulating autoantibodies (auto-Abs) neutralizing high concentrations (10 ng/mL, in plasma diluted 1 to 10) of IFN-α and/or -ω are found in about 10% of patients with critical COVID-19 pneumonia, but not in subjects with asymptomatic infections. We detect auto-Abs neutralizing 100-fold lower, more physiological, concentrations of IFN-α and/or -ω (100 pg/mL, in 1/10 dilutions of plasma) in 13.6% of 3,595 patients with critical COVID-19, including 21% of 374 patients80 years, and 6.5% of 522 patients with severe COVID-19. These antibodies are also detected in 18% of the 1,124 deceased patients (aged 20 days-99 years; mean: 70 years). Moreover, another 1.3% of patients with critical COVID-19 and 0.9% of the deceased patients have auto-Abs neutralizing high concentrations of IFN-β. We also show, in a sample of 34,159 uninfected subjects from the general population, that auto-Abs neutralizing high concentrations of IFN-α and/or -ω are present in 0.18% of individuals between 18 and 69 years, 1.1% between 70 and 79 years, and 3.4%80 years. Moreover, the proportion of subjects carrying auto-Abs neutralizing lower concentrations is greater in a subsample of 10,778 uninfected individuals: 1% of individuals70 years, 2.3% between 70 and 80 years, and 6.3%80 years. By contrast, auto-Abs neutralizing IFN-β do not become more frequent with age. Auto-Abs neutralizing type I IFNs predate SARS-CoV-2 infection and sharply increase in prevalence after the age of 70 years. They account for about 20% of both critical COVID-19 cases in the over-80s, and total fatal COVID-19 cases.

Cell adhesion molecules are membrane-bound proteins predominantly expressed in the central nervous system along principal axonal pathways with key roles in nervous system development, neural cell differentiation and migration, axonal growth and guidance, myelination, and synapse formation. Here, we describe ten affected individuals with bi-allelic variants in the neuronal cell adhesion molecule NRCAM that lead to a neurodevelopmental syndrome of varying severity; the individuals are from eight families. This syndrome is characterized by developmental delay/intellectual disability, hypotonia, peripheral neuropathy, and/or spasticity. Computational analyses of NRCAM variants, many of which cluster in the third fibronectin type III (Fn-III) domain, strongly suggest a deleterious effect on NRCAM structure and function, including possible disruption of its interactions with other proteins. These findings are corroborated by previous in vitro studies of murine Nrcam-deficient cells, revealing abnormal neurite outgrowth, synaptogenesis, and formation of nodes of Ranvier on myelinated axons. Our studies on zebrafish nrcama(Delta) mutants lacking the third Fn-III domain revealed that mutant larvae displayed significantly altered swimming behavior compared to wild-type larvae (p < 0.03). Moreover, nrcama(Delta) mutants displayed a trend toward increased amounts of alpha-tubulin fibers in the dorsal telencephalon, demonstrating an alteration in white matter tracts and projections. Taken together, our study provides evidence that NRCAM disruption causes a variable form of a neurodevelopmental disorder and broadens the knowledge on the growing role of the cell adhesion molecule family in the nervous system.

Exome sequencing and Drosophila modeling revealed variants in SMARCA5 as a novel cause of a neurodevelopmental disorder., Intellectual disability encompasses a wide spectrum of neurodevelopmental disorders, with many linked genetic loci. However, the underlying molecular mechanism for more than 50% of the patients remains elusive. We describe pathogenic variants in SMARCA5, encoding the ATPase motor of the ISWI chromatin remodeler, as a cause of a previously unidentified neurodevelopmental disorder, identifying 12 individuals with de novo or dominantly segregating rare heterozygous variants. Accompanying phenotypes include mild developmental delay, frequent postnatal short stature and microcephaly, and recurrent dysmorphic features. Loss of function of the SMARCA5 Drosophila ortholog Iswi led to smaller body size, reduced sensory dendrite complexity, and tiling defects in larvae. In adult flies, Iswi neural knockdown caused decreased brain size, aberrant mushroom body morphology, and abnormal locomotor function. Iswi loss of function was rescued by wild-type but not mutant SMARCA5. Our results demonstrate that SMARCA5 pathogenic variants cause a neurodevelopmental syndrome with mild facial dysmorphia.

RBL2/p130, a member of the retinoblastoma family of proteins, is a key regulator of cell division and propagates irreversible senescence. RBL2/p130 is also involved in neuronal differentiation and survival, and eliminating Rbl2 in certain mouse strains leads to embryonic lethality accompanied by an abnormal central nervous system (CNS) phenotype. Conflicting reports exist regarding a role of RBL2/p130 in transcriptional regulation of DNA methyltransferases (DNMTs), as well as the control of telomere length. Here we describe the phenotype of three patients carrying bi-allelic RBL2-truncating variants. All presented with infantile hypotonia, severe developmental delay and microcephaly. Malignancies were not reported in carriers or patients. Previous studies carried out on mice and human cultured cells, associated RBL2 loss to DNA methylation and telomere length dysregulation. Here, we investigated whether patient cells lacking RBL2 display related abnormalities. The study of primary patient fibroblasts did not detect abnormalities in expression of DNMTs. Furthermore, methylation levels of whole genome DNA, and specifically of pericentromeric repeats and subtelomeric regions, were unperturbed. RBL2-null fibroblasts show no evidence for abnormal elongation by telomeric recombination. Finally, gradual telomere shortening, and normal onset of senescence were observed following continuous culturing of RBL2-mutated fibroblasts. Thus, this study resolves uncertainties regarding a potential non-redundant role for RBL2 in DNA methylation and telomere length regulation, and indicates that loss of function variants in RBL2 cause a severe autosomal recessive neurodevelopmental disorder in humans.

Iron‑sulfur clusters (FeSCs) are vital components of a variety of essential proteins, most prominently within mitochondrial respiratory chain complexes I-III; Fe—S assembly and distribution is performed via multi-step pathways. Variants affecting several proteins in these pathways have been described in genetic disorders, including severe mitochondrial disease. Here we describe a Christian Arab kindred with two infants that died due to mitochondrial disorder involving Fe—S containing respiratory chain complexes and a third sibling who survived the initial crisis. A homozygous missense variant in NFS1: c.215G>A; p.Arg72Gln was detected by whole exome sequencing. The NFS1 gene encodes a cysteine desulfurase, which, in complex with ISD11 and ACP, initiates the first step of Fe—S formation. Arginine at position 72 plays a role in NFS1-ISD11 complex formation; therefore, its substitution with glutamine is expected to affect complex stability and function. Interestingly, this is the only pathogenic variant ever reported in the NFS1 gene, previously described once in an Old Order Mennonite family presenting a similar phenotype with intra-familial variability in patient outcomes. Analysis of datasets from both populations did not show a common haplotype, suggesting this variant is a recurrent de novo variant. Our report of the second case of NFS1-related mitochondrial disease corroborates the pathogenicity of this recurring variant and implicates it as a hot-spot variant. While the genetic resolution allows for prenatal diagnosis for the family, it also raises critical clinical questions regarding follow-up and possible treatment options of severely affected and healthy homozygous individuals with mitochondrial co-factor therapy or cysteine supplementation.

BackgroundConstitutional mismatch repair deficiency syndrome (CMMRD) is the most aggressive cancer predisposition syndrome associated with multiorgan cancers, often presenting in childhood. There is variability in age and presentation of cancers and benign manifestations mimicking neurofibromatosis type 1. Genetic testing may not be informative and is complicated by pseudogenes associated with the most commonly associated gene, PMS2. To date, no diagnostic criteria exist. Since surveillance and immune-based therapies are available, establishing a CMMRD diagnosis is key to improve survival.MethodsIn order to establish a robust diagnostic path, a multidisciplinary international working group, with representation from the two largest consortia (International Replication Repair Deficiency (IRRD) consortium and European Consortium Care for CMMRD (C4CMMRD)), was formed to establish diagnostic criteria based on expertise, literature review and consensus.ResultsThe working group established seven diagnostic criteria for the diagnosis of CMMRD, including four definitive criteria (strong evidence) and three likely diagnostic criteria (moderate evidence). All criteria warrant CMMRD surveillance. The criteria incorporate germline mismatch repair results, ancillary tests and clinical manifestation to determine a diagnosis. Hallmark cancers for CMMRD were defined by the working group after extensive literature review and consultation with the IRRD and C4CMMRD consortia.ConclusionsThis position paper summarises the evidence and rationale to provide specific guidelines for CMMRD diagnosis, which necessitates appropriate surveillance and treatment.

The genetic causes of multiple congenital anomalies are incompletely understood. Here, we report novel heterozygous predicted loss-of-function (LoF) and predicted damaging missense variants in the WW domain binding protein 11 (WBP11) gene in seven unrelated families with a variety of overlapping congenital malformations, including cardiac, vertebral, tracheo-esophageal, renal and limb defects. WBP11 encodes a component of the spliceosome with the ability to activate pre-messenger RNA splicing. We generated a Wbp11 null allele in mouse using CRISPR-Cas9 targeting. Wbp11 homozygous null embryos die prior to E8.5, indicating that Wbp11 is essential for development. Fewer Wbp11 heterozygous null mice are found than expected due to embryonic and postnatal death. Importantly, Wbp11 heterozygous null mice are small and exhibit defects in axial skeleton, kidneys and esophagus, similar to the affected individuals, supporting the role of WBP11 haploinsufficiency in the development of congenital malformations in humans. LoF WBP11 variants should be considered as a possible cause of VACTERL association as well as isolated Klippel-Feil syndrome, renal agenesis or esophageal atresia.

Germline pathogenic variants (PVs) in BRCA1/BRCA2 are well-established risk factors for breast cancer (BC) and/or ovarian cancer (OC). Founder PVs have been described in BRCA1/ BRCA2 in several genetic isolates. The Christian Arab population in the Middle East is a relatively isolated ethnic group, yet founder, or recurrent BRCA1/BRCA2 PVs have not been reported in this population. In this study we describe PVs detected in cancer susceptibility genes among a cohort of Christian Arabs from Israel. We reviewed patient records from the Oncogenetic clinic at Rambam Health Care Campus during the years 2013- mid 2020. Thirty-five unrelated Christian Arab patients, with personal or family history of BC and/or OC underwent BRCA1/BRCA2 (14/35) testing or cancer gene panel testing (21/35) as part of their diagnostic workup. Three clinically significant variants in BRCA2, CHEK2 and RAD51C were found in 7/35 patients (20%). A recurrent duplication of the BRCA2 genomic region, encompassing exons 5-10 and the 5' portion of exon 11, was found in 5/33 (15.2%) patients for whom copy number variants (CNVs) analysis was performed. We identified a recurrent pathogenic BRCA2 duplication in Christian Arab patients with a personal/ family history of BC and/or OC. Our findings emphasize the importance of inclusion of CNVs analysis in BRCA1/BRCA2 genetic testing, and specifically for Christian Arab patients suspected of hereditary BC and/or OC.

Intellectual disability (ID) encompasses a wide spectrum of neurodevelopmental disorders, with many linked genetic loci. However, the underlying molecular mechanism for over 50% of the patients remains elusive. We describe mutations inSMARCA5, encoding the ATPase motor of the ISWI chromatin remodeler, as a cause of a novel neurodevelopmental disorder, identifying twelve individuals withde novoor dominantly segregating rare heterozygous variants. Accompanying phenotypes include mild developmental delay, frequent postnatal short stature, and microcephaly, and recurrent dysmorphic features. Loss of function of the SMARCA5DrosophilaorthologIswiled to smaller body size, reduced dendrite complexity, and tiling defects in larvae. In adult flies, Iswi neural knockdown caused decreased brain size, aberrant mushroom body morphology and abnormal locomotor function.Iswiloss of function was rescued by wild-type but not mutant SMARCA5. Our results demonstrate thatSMARCA5pathogenic variants cause a neurodevelopmental syndrome with mild facial dysmorphia.

Mutations in more than 150 genes are responsible for inherited hearing loss, with thousands of different, severe causal alleles that vary among populations. The Israeli Jewish population includes communities of diverse geographic origins, revealing a wide range of deafness-associated variants and enabling clinical characterization of the associated phenotypes. Our goal was to identify the genetic causes of inherited hearing loss in this population, and to determine relationships among genotype, phenotype, and ethnicity. Genomic DNA samples from informative relatives of 88 multiplex families, all of self-identified Jewish ancestry, with either non-syndromic or syndromic hearing loss, were sequenced for known and candidate deafness genes using the HEar-Seq gene panel. The genetic causes of hearing loss were identified for 60% of the families. One gene was encountered for the first time in human hearing loss:ATOH1(Atonal), a basic helix-loop-helix transcription factor responsible for autosomal dominant progressive hearing loss in a five-generation family. Our results demonstrate that genomic sequencing with a gene panel dedicated to hearing loss is effective for genetic diagnoses in a diverse population. Comprehensive sequencing enables well-informed genetic counseling and clinical management by medical geneticists, otolaryngologists, audiologists, and speech therapists and can be integrated into newborn screening for deafness.

Sifrim–Hitz–Weiss syndrome (SIHIWES) is a recently described multisystemic neurodevelopmental disorder caused by de novo variants in CHD4. In this study, we investigated the clinical spectrum of the disorder, genotype–phenotype correlations, and the effect of different missense variants on CHD4 function. We collected clinical and molecular data from 32 individuals with mostly de novo variants in CHD4, identified through next-generation sequencing. We performed adenosine triphosphate (ATP) hydrolysis and nucleosome remodeling assays on variants from five different CHD4 domains. The majority of participants had global developmental delay, mild to moderate intellectual disability, brain anomalies, congenital heart defects, and dysmorphic features. Macrocephaly was a frequent but not universal finding. Additional common abnormalities included hypogonadism in males, skeletal and limb anomalies, hearing impairment, and ophthalmic abnormalities. The majority of variants were nontruncating and affected the SNF2-like region of the protein. We did not identify genotype–phenotype correlations based on the type or location of variants. Alterations in ATP hydrolysis and chromatin remodeling activities were observed in variants from different domains. The CHD4-related syndrome is a multisystemic neurodevelopmental disorder. Missense substitutions in different protein domains alter CHD4 function in a variant-specific manner, but result in a similar phenotype in humans.

MYH7-related disease (MRD) is the most common hereditary primary cardiomyopathy (CM), with pathogenic MYH7 variants accounting for approximately 40% of familial hypertrophic CMs. MRDs may also present as skeletal myopathies, with or without CM. Since pathogenic MYH7 variants result in highly variable clinical phenotypes, from mild to fatal forms of cardiac and skeletal myopathies, genotype-phenotype correlations are not always apparent, and translation of the genetic findings to clinical practice can be complicated. Data on genotype-phenotype correlations can help facilitate more specific and personalized decisions on treatment strategies, surveillance, and genetic counseling. We present a series of six MRD pedigrees with rare genotypes, encompassing various clinical presentations and inheritance patterns. This study provides new insights into the spectrum of MRD that is directly translatable to clinical practice.