Discovering the ANK2-related autism phenotype.

Interestingly, disease-causing mutations in the ANK2 gene have been identified in patients with autism since 2012, though with no full clinical description. In this Research Letter, for the first time, we describe the detailed characteristics of a patient with autism caused by a new mutation in this gene. Our report is a first step to better understanding ANK2-related autism and will contribute to facilitating its further diagnosis.

Evaluation of CSF1R-related adult onset leukoencephalopathy with axonal spheroids and pigmented glia diagnostic criteria.

BACKGROUND AND PURPOSE: Diagnostic criteria for adult onset leukoencephalopathy with axonal spheroids and pigmented glia (ALSP) due to colony-stimulating factor 1 receptor (CSF1R) mutation have recently been proposed. Our objective was to assess their accuracy in an independent multicenter cohort. METHODS: We evaluated the sensitivity and specificity of the diagnostic criteria for ALSP (including the "probable" and "possible" definitions) in a national cohort of 22 patients with CSF1R mutation, and 59 patients with an alternative diagnosis of adult onset inherited leukoencephalopathy. RESULTS: Overall, the sensitivity of the diagnostic criteria for ALSP was 82%, including nine of 22 patients diagnosed as probable and nine of 22 diagnosed as possible. Twenty of the 59 CSF1R mutation-negative leukoencephalopathies fulfilled the diagnostic criteria, leading to a specificity of 66%. CONCLUSIONS: Diagnostic criteria for ALSP have an overall limited sensitivity along with a modest specificity. We suggest that in patients suspected of genetic leukoencephalopathy, a comprehensive magnetic resonance imaging pattern-based approach is warranted, together with white matter gene panel or whole exome sequencing.

Impact of a frequent nearsplice SOD1 variant in amyotrophic lateral sclerosis: optimising SOD1 genetic screening for gene therapy opportunities.

OBJECTIVE: Mutations in superoxide dismutase 1 gene (SOD1), encoding copper/zinc superoxide dismutase protein, are the second most frequent high penetrant genetic cause for amyotrophic lateral sclerosis (ALS) motor neuron disease in populations of European descent. More than 200 missense variants are reported along the SOD1 protein. To limit the production of these aberrant and deleterious SOD1 species, antisense oligonucleotide approaches have recently emerged and showed promising effects in clinical trials. To offer the possibility to any patient with SOD1-ALS to benefit of such a gene therapy, it is necessary to ascertain whether any variant of unknown significance (VUS), detected for example in SOD1 non-coding sequences, is pathogenic. METHODS: We analysed SOD1 mutation distribution after SOD1 sequencing in a large cohort of 470 French familial ALS (fALS) index cases. RESULTS: We identified a total of 27 SOD1 variants in 38 families including two SOD1 variants located in nearsplice or intronic regions of the gene. The pathogenicity of the c.358-10T>G nearsplice SOD1 variant was corroborated based on its high frequency (as the second most frequent SOD1 variant) in French fALS, the segregation analysis confirmed in eight affected members of a large pedigree, the typical SOD1-related phenotype observed (with lower limb onset and prominent lower motor neuron involvement), and findings on postmortem tissues showing SOD1 misaccumulation. CONCLUSIONS: Our results highlighted nearsplice/intronic mutations in SOD1 are responsible for a significant portion of French fALS and suggested the systematic analysis of the SOD1 mRNA sequence could become the method of choice for SOD1 screening, not to miss these specific cases.

Clinical and Molecular Landscape of ALS Patients with SOD1 Mutations: Novel Pathogenic Variants and Novel Phenotypes. A Single ALS Center Study.

Mutations in the copper zinc superoxide dismutase 1 (SOD1) gene are the second most frequent cause of familial amyotrophic lateral sclerosis (ALS). Nearly 200 mutations of this gene have been described so far. We report all SOD1 pathogenic variants identified in patients followed in the single ALS center of Lyon, France, between 2010 and 2020. Twelve patients from 11 unrelated families are described, including two families with the not yet described H81Y and D126N mutations. Splice site mutations were detected in two families. We discuss implications concerning genetic screening of SOD1 gene in familial and sporadic ALS.

Electrophysiological Characterization of C9ORF72-Associated Amyotrophic Lateral Sclerosis: A Retrospective Study.

OBJECTIVE: C9ORF72 is the most common genetic cause of amyotrophic lateral sclerosis (ALS). The aim of the present study was to determine whether C9ORF72-associated ALS (C9-ALS) patients present distinctive electrophysiological characteristics that could differentiate them from non C9ORF72-associated ALS (nonC9-ALS) patients. METHODS: Clinical and electrodiagnostic data from C9-ALS patients and nonC9-ALS patients were collected retrospectively. For electroneuromyography, the mean values of motor conduction, myography, and the mean values of sensory conduction were considered. Furthermore, the proportion of ALS patients with electrophysiological sensory neuropathy was determined. RESULTS: No significant difference was observed between 31 C9-ALS patients and 22 nonC9-ALS patients for mean motor conduction and myography. For sensory conduction analyses, mean sensory conduction was not significantly different between both groups. In total, 38% of -C9-ALS patient and 21% of nonC9-ALS patients presented electrophysiological sensory neuropathy (p = 0.33). In -C9-ALS patients with electrophysiological sensory neuropathy, 80% (8/10) were male and 67% (6/9) presented spinal onset compare to 25% (4/16, p = 0.014) male and 25% (4/16, p = 0.087) with spinal onset in those without electrophysiological sensory neuropathy. CONCLUSION: Although not different from nonC9-ALS, these results suggest that sensory involvement is a frequent feature of C9-ALS patients, expanding the phenotype of the disease beyond the motor and cognitive domains.

Regulation of Brain Cholesterol: What Role Do Liver X Receptors Play in Neurodegenerative Diseases?

Liver X Receptors (LXR) alpha and beta are two members of nuclear receptor superfamily documented as endogenous cholesterol sensors. Following conversion of cholesterol in oxysterol, both LXR isoforms detect intracellular concentrations and act as transcription factors to promote expression of target genes. Among their numerous physiological roles, they act as central cholesterol-lowering factors. In the central nervous system (CNS), cholesterol has been shown to be an essential determinant of brain function, particularly as a major constituent of myelin and membranes. In the brain, LXRs act as cholesterol central regulators, and, beyond this metabolic function, LXRs have additional roles such as providing neuroprotective effects and lowering neuroinflammation. In many neurodegenerative disorders, including amyotrophic lateral sclerosis (ALS), Alzheimer's disease (AD), and multiple sclerosis (MS), dysregulations of cholesterol and oxysterol have been reported. In this paper, we propose to focus on recent advances in the knowledge of the LXRs roles on brain cholesterol and oxysterol homeostasis, neuroinflammation, neuroprotection, and their putative involvement in neurodegenerative disorders. We will discuss their potential use as candidates for both molecular diagnosis and as promising pharmacological targets in the treatment of ALS, AD, or MS patients.

Liver X receptors: from cholesterol regulation to neuroprotection-a new barrier against neurodegeneration in amyotrophic lateral sclerosis?

Cholesterol plays a central role in numerous nervous system functions. Cholesterol is the major constituent of myelin sheaths, is essential for synapse and dendrite formation, axon guidance as well as neurotransmission. Among regulators of cholesterol homeostasis, liver X receptors (LXRs), two members of the nuclear receptor superfamily, play a determinant role. LXRs act as cholesterol sensors and respond to high intracellular cholesterol concentration by decreasing plasmatic and intracellular cholesterol content. Beyond their cholesterol-lowering role, LXRs have been proposed as regulators of immunity and anti-inflammatory factors. Dysregulation of cholesterol metabolism combined to neuroinflammatory context have been described in neurodegenerative diseases, including amyotrophic lateral sclerosis (ALS). ALS is characterized by the progressive loss of motoneurons in the brain and spinal cord, leading to severe paralytic condition and death of patients in a median time of 3 years. Motoneuron degeneration is accompanied by chronic neuroinflammatory response, involving microglial and astrocytic activation, infiltration of blood-derived immune cells and release of pro-inflammatory factors. We propose to discuss here the role of LXRs as a molecular link between the central nervous system cholesterol metabolism, neuroinflammation, motoneuron survival and their potential as promising therapeutic candidates for ALS therapy.

Meta-analysis of SHANK Mutations in Autism Spectrum Disorders: a gradient of severity in cognitive impairments.

SHANK genes code for scaffold proteins located at the post-synaptic density of glutamatergic synapses. In neurons, SHANK2 and SHANK3 have a positive effect on the induction and maturation of dendritic spines, whereas SHANK1 induces the enlargement of spine heads. Mutations in SHANK genes have been associated with autism spectrum disorders (ASD), but their prevalence and clinical relevance remain to be determined. Here, we performed a new screen and a meta-analysis of SHANK copy-number and coding-sequence variants in ASD. Copy-number variants were analyzed in 5,657 patients and 19,163 controls, coding-sequence variants were ascertained in 760 to 2,147 patients and 492 to 1,090 controls (depending on the gene), and, individuals carrying de novo or truncating SHANK mutations underwent an extensive clinical investigation. Copy-number variants and truncating mutations in SHANK genes were present in ∼1% of patients with ASD: mutations in SHANK1 were rare (0.04%) and present in males with normal IQ and autism; mutations in SHANK2 were present in 0.17% of patients with ASD and mild intellectual disability; mutations in SHANK3 were present in 0.69% of patients with ASD and up to 2.12% of the cases with moderate to profound intellectual disability. In summary, mutations of the SHANK genes were detected in the whole spectrum of autism with a gradient of severity in cognitive impairment. Given the rare frequency of SHANK1 and SHANK2 deleterious mutations, the clinical relevance of these genes remains to be ascertained. In contrast, the frequency and the penetrance of SHANK3 mutations in individuals with ASD and intellectual disability-more than 1 in 50-warrant its consideration for mutation screening in clinical practice.

A blinded international study on the reliability of genetic testing for GGGGCC-repeat expansions in C9orf72 reveals marked differences in results among 14 laboratories.

BACKGROUND: The GGGGCC-repeat expansion in C9orf72 is the most frequent mutation found in patients with amyotrophic lateral sclerosis (ALS) and frontotemporal dementia (FTD). Most of the studies on C9orf72 have relied on repeat-primed PCR (RP-PCR) methods for detection of the expansions. To investigate the inherent limitations of this technique, we compared methods and results of 14 laboratories. METHODS: The 14 laboratories genotyped DNA from 78 individuals (diagnosed with ALS or FTD) in a blinded fashion. Eleven laboratories used a combination of amplicon-length analysis and RP-PCR, whereas three laboratories used RP-PCR alone; Southern blotting techniques were used as a reference. RESULTS: Using PCR-based techniques, 5 of the 14 laboratories got results in full accordance with the Southern blotting results. Only 50 of the 78 DNA samples got the same genotype result in all 14 laboratories. There was a high degree of false positive and false negative results, and at least one sample could not be genotyped at all in 9 of the 14 laboratories. The mean sensitivity of a combination of amplicon-length analysis and RP-PCR was 95.0% (73.9-100%), and the mean specificity was 98.0% (87.5-100%). Overall, a sensitivity and specificity of more than 95% was observed in only seven laboratories. CONCLUSIONS: Because of the wide range seen in genotyping results, we recommend using a combination of amplicon-length analysis and RP-PCR as a minimum in a research setting. We propose that Southern blotting techniques should be the gold standard, and be made obligatory in a clinical diagnostic setting.

A rare motor neuron deleterious missense mutation in the DPYSL3 (CRMP4) gene is associated with ALS.

The dihydropyrimidinase-like 3 (DPYSL3) or Collapsin Response Mediator Protein 4a (CRMP4a) expression is modified in neurodegeneration and is involved in several ALS-associated pathways including axonal transport, glutamate excitotoxicity, and oxidative stress. The objective of the study was to analyze CRMP4 as a risk factor for ALS. We analyzed the DPYSL3/CRMP4 gene in French ALS patients (n = 468) and matched-controls (n = 394). We subsequently examined a variant in a Swedish population (184 SALS, 186 controls), and evaluated its functional effects on axonal growth and survival in motor neuron cell culture. The rs147541241:A>G missense mutation occurred in higher frequency among French ALS patients (odds ratio = 2.99) but the association was not confirmed in the Swedish population. In vitro expression of mutated DPYSL3 in motor neurons reduced axonal growth and accelerated cell death compared with wild type protein. Thus, the association between the rs147541241 variant and ALS was limited to the French population, highlighting the geographic particularities of genetic influences (risks, contributors). The identified variant appears to shorten motor neuron survival through a detrimental effect on axonal growth and CRMP4 could act as a key unifier in transduction pathways leading to neurodegeneration through effects on early axon development.

Genetic variations related to inflammation in suicidal ideation and behavior: A systematic review.

Background/objectives: Immune-inflammatory changes have been found in all types of suicidal ideation and behavior (SIB), independently of associated mental disorders. Since several Single Nucleotide Polymorphisms (SNPs) affect the function of inflammation-related genes, we searched the literature for genetic variations potentially altering inflammatory processes in SIB. Methods: We included studies that looked for associations between SIB and SNPs in genes related to inflammatory processes. Case reports, literature reviews, and animal studies were excluded. Articles were retrieved from PubMed and PsycINFO databases, Google Scholar and GreySource Index until September 17th, 2022. Quality was assessed using Q-Genie. Results: We analyzed 32 studies. SIB has been associated with eighteen SNPs located in genes encoding for interleukin-8 (rs4073), C-reactive protein (rs1130864), tumor necrosis factor α (rs1800629, rs361525, and rs1099724), tumor necrosis factor receptor 2 (rs1061622), transforming growth factor ß-1 (rs1982073), acid phosphatase 1 (rs7419262, rs300774), interleukin-10 (rs1800896), interferon γ (rs2430561), amino-carboxy muconate semialdehyde decarboxylase (rs2121337), interleukin 7 (rs10448044, rs10448042), macrophage migration inhibitory factor (rs755622), interleukin 1-α (rs1800587), and interleukin 1-ß (rs1143634 and rs16944. A genome-wide association study reported one association at the threshold of significance with the rs300774 SNP, located in the 2p25 region containing ACP1 gene. Discussion: The studies included were methodologically and clinically diverse and of moderate quality. Their findings suggest that some inflammation-related SNPs could increase the likelihood of SIB but the evidence to date is insufficient. Further research using gene-gene (GxG) and gene-environment (GxE) approaches is warranted. Systematic review registration: [https://www.crd.york.ac.uk], identifier [CRD42022296310].

Juvenile amyotrophic lateral sclerosis associated with biallelic c.757delG mutation of sorbitol dehydrogenase gene.

Mutation in the sorbitol dehydrogenase gene (SORD) has been recently described to cause axonal Charcot-Marie-Tooth disease (CMT), intermediate CMT, and distal hereditary motor neuropathy (dHMN). We herein report the case of a 24-year-old patient diagnosed with juvenile amyotrophic lateral sclerosis (JALS) who carried the homozygous c.757delG mutation in SORD. No other pathogenic variant in frequent JALS-causative genes was found. Our findings expand the phenotype related to SORD mutation, a new and potentially treatable genetic disease.

Amyotrophic lateral sclerosis associated with a pathological expansion in the ATXN7 gene.

Spinocerebellar ataxia type 7 (SCA7) is an autosomal dominant hereditary neurodegenerative disease caused by the expansion of a CAG-repeat in the ataxin-7 (ATXN7) gene, usually characterized by progressive cerebellar ataxia and retinal dystrophy. We report the case of a 45-year-old woman presenting with a rapid-onset amyotrophic lateral sclerosis (ALS) phenotype associated with a 39-CAG-repeat expansion in ATXN7. This patient had neither ataxia nor retinal dystrophy, but she had an oculomotor cerebellar syndrome and a family history suggestive of SCA7. In SCA7, shorter expansions may be associated with less severe and incomplete clinical phenotypes, which could explain the patient's phenotype. Unknown genetic and environmental factors may also influence the patient's phenotype. We suggest that a pathological expansion in ATXN7 should be considered in cases of ALS-like phenotype, particularly when associated with oculomotor abnormalities or a family history of ataxia or blindness.

Compound heterozygous P67S/D91A SOD1 mutations in an ALS family with apparently sporadic case.

Objectives: To describe a family with heterozygous P67S and D91A SOD1 mutations. Methods: The ALS profile of the proband was described. SOD1 gene sequencing was performed in the proband and his children. Results: The affected individual presented with progressive left peripheral facial palsy and slow progression with late limb involvement. Unequivocal upper and lower motor neuron signs were present, together with diffuse denervation at myography. The absence of trigeminal involvement excluded a FOSMN syndrome. Pedigree analysis did not show any other ALS case in the family. Genetic analysis of this patient showed P67S and D91A SOD1 mutations. The genetic analysis of the children showed that the mutations were each one carried by a different chromosome. Conclusions: P67S SOD1 mutation has been described in several ALS cases, either with familial or apparently sporadic ALS. The mutation is located in a mutational hotspot and was predicted pathogenic by in silico prediction software. The study of phylogenetic data show that at this codon, the proline is highly conserved throughout species reinforcing causality. Conversely, the D91A variant is known to have a recessive influence. Unilateral motor facial involvement, even after several years, in an ALS patient is unusual. The present case with compound heterozygosity and unusual onset in a patient with apparently sporadic ALS, widens the clinical spectrum of the disease and adds further arguments to support the systematic genetic screening of all ALS cases in referral ALS clinics.

A common polymorphism in NR1H2 (LXRbeta) is associated with preeclampsia.

BACKGROUND: Preeclampsia is a frequent complication of pregnancy and a leading cause of perinatal mortality. Both genetic and environmental risk factors have been identified. Lipid metabolism, particularly cholesterol metabolism, is associated with this disease. Liver X receptors alpha (NR1H3, also known as LXRalpha) and beta (NR1H2, also known as LXRbeta) play a key role in lipid metabolism. They belong to the nuclear receptor superfamily and are activated by cholesterol derivatives. They have been implicated in preeclampsia because they modulate trophoblast invasion and regulate the expression of the endoglin (CD105) gene, a marker of preeclampsia. The aim of this study was to investigate associations between the NR1H3 and NR1H2 genes and preeclampsia. METHODS: We assessed associations between single nucleotide polymorphisms of NR1H3 (rs2279238 and rs7120118) and NR1H2 (rs35463555 and rs2695121) and the disease in 155 individuals with preeclampsia and 305 controls. Genotypes were determined by high-resolution melting analysis. We then used a logistic regression model to analyze the different alleles and genotypes for those polymorphisms as a function of case/control status. RESULTS: We found no association between NR1H3 SNPs and the disease, but the NR1H2 polymorphism rs2695121 was found to be strongly associated with preeclampsia (genotype C/C: adjusted odds ratio, 2.05; 95% CI, 1.04-4.05; p = 0.039 and genotype T/C: adjusted odds ratio, 1.85; 95% CI, 1.01-3.42; p = 0.049). CONCLUSIONS: This study provides the first evidence of an association between the NR1H2 gene and preeclampsia, adding to our understanding of the links between cholesterol metabolism and this disease.

Genes containing hexanucleotide repeats resembling C9ORF72 and expressed in the central nervous system are frequent in the human genome.

More than 40 human diseases, mainly diseases affecting the central nervous system, are caused by the expansion of unstable nucleotide repeats. Repeats of sequences like (CAG)n present in different genes can be responsible for various diseases of the central nervous system. An expanded hexanucleotide repeat (GGGGCC)n in the C9ORF72 gene has been characterized as the most frequent genetic cause of amyotrophic lateral sclerosis and frontotemporal lobar dementia. In this study, we performed a genome-wide analysis in the human genome and identified 74 genes containing this precise hexanucleotide repeat, with a preference for a location in exon 1 or intron 1, similar to the C9ORF72 gene. A total of 36 of these 74 genes may be of interest as candidates in neurodevelopmental or neurodegenerative diseases, based on their function.

High-resolution melting analysis of sequence variations in the cytidine deaminase gene (CDA) in patients with cancer treated with gemcitabine.

Gemcitabine (2',2'-difluorodeoxycytidine) is a major antimetabolite cytotoxic drug with a wide spectrum of activity against solid tumors. Hepatic elimination of gemcitabine depends on a catabolic pathway through a deamination step driven by the enzyme cytidine deaminase (CDA). Severe hematologic toxicity to gemcitabine was reported in patients harboring genetic polymorphisms in CDA gene. High-resolution melting (HRM) analysis of polymerase chain reaction amplicon emerges today as a powerful technique for both genotyping and gene scanning strategies. In this study, 46 DNA samples from gemcitabine-treated patients were subjected to HRM analysis on a LightCycler 480 platform. Residual serum CDA activity was assayed as a surrogate marker for the overall functionality of this enzyme. Genotyping of three well-described single nucleotide polymorphisms in coding region (c.79A>C, c.208G>A and c.435C>T) was successfully achieved by HRM analysis of small polymerase chain reaction fragments, whereas unknown single nucleotide polymorphisms were searched by a gene scanning strategy with longer amplicons (up to 622 bp). The gene scanning strategy allowed us to find a new intronic mutation c.246+37G>A in a female patient displaying marked CDA deficiency and who had an extreme toxic reaction with a fatal outcome to gemcitabine treatment. Our work demonstrates that HRM-based methods, owing to their simplicity, reliability, and speed, are useful tools for diagnosis of CDA deficiency and could be of interest for personalized medicine.

Premature termination codons in SOD1 causing Amyotrophic Lateral Sclerosis are predicted to escape the nonsense-mediated mRNA decay.

Amyotrophic lateral sclerosis (ALS) is the most common and severe adult-onset motoneuron disease and has currently no effective therapy. Approximately 20% of familial ALS cases are caused by dominantly-inherited mutations in the gene encoding Cu/Zn superoxide dismutase (SOD1), which represents one of the most frequent genetic cause of ALS. Despite the overwhelming majority of ALS-causing missense mutations in SOD1, a minority of premature termination codons (PTCs) have been identified. mRNA harboring PTCs are known to be rapidly degraded by nonsense-mediated mRNA decay (NMD), which limits the production of truncated proteins. The rules of NMD surveillance varying with PTC location in mRNA, we analyzed the localization of PTCs in SOD1 mRNA to evaluate whether or not those PTCs can be triggered to degradation by the NMD pathway. Our study shows that all pathogenic PTCs described in SOD1 so far can theoretically escape the NMD, resulting in the production of truncated protein. This finding supports the hypothesis that haploinsufficiency is not an underlying mechanism of SOD1 mutant-associated ALS and suggests that PTCs found in the regions that trigger NMD are not pathogenic. Such a consideration is particularly important since the availability of SOD1 antisense strategies, in view of variant treatment assignment.