JAK2-mutated acute myeloid leukemia: comparison of next-generation sequencing (NGS) and single nucleotide polymorphism array (SNPa) findings between two cases

JAK2 mutations are rare in de novo acute myeloid leukemia (AML), and JAK2-mutated acute myeloid leukemia (AML) patients usually have a previous history of myeloproliferative neoplasms (MPNs). Current advances in laboratory techniques, such as single nucleotide polymorphism array (SNPa) and next-generation sequencing (NGS), have facilitated new insight into the molecular basis of hematologic diseases. Herein, we present two cases of JAK2-mutated AML in which both SNPa and NGS methods added valuable information. Both cases had leukemogenic collaboration, namely, copy-neutral loss of heterozygosity (CN-LOH), detected on chromosome 9. One of the cases exhibited both JAK2 and IDH2 mutations, most likely having originated as an MPN with leukemic transformation, while the other case was classified as a de novo AML with JAK2, CEBPA, and FLT3 mutations

Targeted Genetic Screen in Amyotrophic Lateral Sclerosis Reveals Novel Genetic Variants with Synergistic Effect on Clinical Phenotype

Amyotrophic lateral sclerosis (ALS) is underpinned by an oligogenic rare variant architecture. Identified genetic variants of ALS include RNA-binding proteins containing prion-like domains (PrLDs). We hypothesized that screening genes encoding additional similar proteins will yield novel genetic causes of ALS. The most common genetic variant of ALS patients is a G4C2-repeat expansion within C9ORF72. We have shown that G4C2-repeat RNA sequesters RNA-binding proteins. A logical consequence of this is that loss-of-function mutations in G4C2-binding partners might contribute to ALS pathogenesis independently of and/or synergistically with C9ORF72 expansions. Targeted sequencing of genomic DNA encoding either RNA-binding proteins or known ALS genes (n = 274 genes) was performed in ALS patients to identify rare deleterious genetic variants and explore genotype-phenotype relationships. Genomic DNA was extracted from 103 ALS patients including 42 familial ALS patients and 61 young-onset (average age of onset 41 years) sporadic ALS patients; patients were chosen to maximize the probability of identifying genetic causes of ALS. Thirteen patients carried a G4C2-repeat expansion of C9ORF72. We identified 42 patients with rare deleterious variants; 6 patients carried more than one variant. Twelve mutations were discovered in known ALS genes which served as a validation of our strategy. Rare deleterious variants in RNA-binding proteins were significantly enriched in ALS patients compared to control frequencies (p = 5.31E-18). Nineteen patients featured at least one variant in a RNA-binding protein containing a PrLD. The number of variants per patient correlated with rate of disease progression (t-test, p = 0.033). We identified eighteen patients with a single variant in a G4C2-repeat binding protein. Patients with a G4C2-binding protein variant in combination with a C9ORF72 expansion had a significantly faster disease course (t-test, p = 0.025). Our data are consistent with an oligogenic model of ALS. We provide evidence for a number of entirely novel genetic variants of ALS caused by mutations in RNA-binding proteins. Moreover we show that these mutations act synergistically with each other and with C9ORF72 expansions to modify the clinical phenotype of ALS. A key finding is that this synergy is present only between functionally interacting variants. This work has significant implications for ALS therapy development

Common and rare variant association analyses in amyotrophic lateral sclerosis identify 15 risk loci with distinct genetic architectures and neuron-specific biology

Amyotrophic lateral sclerosis (ALS) is a fatal neurodegenerative disease with a lifetime risk of one in 350 people and an unmet need for disease-modifying therapies. We conducted a cross-ancestry genome-wide association study (GWAS) including 29,612 patients with ALS and 122,656 controls, which identified 15 risk loci. When combined with 8,953 individuals with whole-genome sequencing (6,538 patients, 2,415 controls) and a large cortex-derived expression quantitative trait locus (eQTL) dataset (MetaBrain), analyses revealed locus-specific genetic architectures in which we prioritized genes either through rare variants, short tandem repeats or regulatory effects. ALS-associated risk loci were shared with multiple traits within the neurodegenerative spectrum but with distinct enrichment patterns across brain regions and cell types. Of the environmental and lifestyle risk factors obtained from the literature, Mendelian randomization analyses indicated a causal role for high cholesterol levels. The combination of all ALS-associated signals reveals a role for perturbations in vesicle-mediated transport and autophagy and provides evidence for cell-autonomous disease initiation in glutamatergic neurons. A cross-ancestry genome-wide association meta-analysis of amyotrophic lateral sclerosis (ALS) including 29,612 patients with ALS and 122,656 controls identifies 15 risk loci with distinct genetic architectures and neuron-specific biology

Common and rare variant association analyses in amyotrophic lateral sclerosis identify 15 risk loci with distinct genetic architectures and neuron-specific biology

Amyotrophic lateral sclerosis (ALS) is a fatal neurodegenerative disease with a lifetime risk of one in 350 people and an unmet need for disease-modifying therapies. We conducted a cross-ancestry genome-wide association study (GWAS) including 29,612 patients with ALS and 122,656 controls, which identified 15 risk loci. When combined with 8,953 individuals with whole-genome sequencing (6,538 patients, 2,415 controls) and a large cortex-derived expression quantitative trait locus (eQTL) dataset (MetaBrain), analyses revealed locus-specific genetic architectures in which we prioritized genes either through rare variants, short tandem repeats or regulatory effects. ALS-associated risk loci were shared with multiple traits within the neurodegenerative spectrum but with distinct enrichment patterns across brain regions and cell types. Of the environmental and lifestyle risk factors obtained from the literature, Mendelian randomization analyses indicated a causal role for high cholesterol levels. The combination of all ALS-associated signals reveals a role for perturbations in vesicle-mediated transport and autophagy and provides evidence for cell-autonomous disease initiation in glutamatergic neurons. A cross-ancestry genome-wide association meta-analysis of amyotrophic lateral sclerosis (ALS) including 29,612 patients with ALS and 122,656 controls identifies 15 risk loci with distinct genetic architectures and neuron-specific biology

Common and rare variant association analyses in amyotrophic lateral sclerosis identify 15 risk loci with distinct genetic architectures and neuron-specific biology

A cross-ancestry genome-wide association meta-analysis of amyotrophic lateral sclerosis (ALS) including 29,612 patients with ALS and 122,656 controls identifies 15 risk loci with distinct genetic architectures and neuron-specific biology. Amyotrophic lateral sclerosis (ALS) is a fatal neurodegenerative disease with a lifetime risk of one in 350 people and an unmet need for disease-modifying therapies. We conducted a cross-ancestry genome-wide association study (GWAS) including 29,612 patients with ALS and 122,656 controls, which identified 15 risk loci. When combined with 8,953 individuals with whole-genome sequencing (6,538 patients, 2,415 controls) and a large cortex-derived expression quantitative trait locus (eQTL) dataset (MetaBrain), analyses revealed locus-specific genetic architectures in which we prioritized genes either through rare variants, short tandem repeats or regulatory effects. ALS-associated risk loci were shared with multiple traits within the neurodegenerative spectrum but with distinct enrichment patterns across brain regions and cell types. Of the environmental and lifestyle risk factors obtained from the literature, Mendelian randomization analyses indicated a causal role for high cholesterol levels. The combination of all ALS-associated signals reveals a role for perturbations in vesicle-mediated transport and autophagy and provides evidence for cell-autonomous disease initiation in glutamatergic neurons

Genome-wide Analyses Identify KIF5A as a Novel ALS Gene

To identify novel genes associated with ALS, we undertook two lines of investigation. We carried out a genome-wide association study comparing 20,806 ALS cases and 59,804 controls. Independently, we performed a rare variant burden analysis comparing 1,138 index familial ALS cases and 19,494 controls. Through both approaches, we identified kinesin family member 5A (KIF5A) as a novel gene associated with ALS. Interestingly, mutations predominantly in the N-terminal motor domain of KIF5A are causative for two neurodegenerative diseases: hereditary spastic paraplegia (SPG10) and Charcot-Marie-Tooth type 2 (CMT2). In contrast, ALS-associated mutations are primarily located at the C-terminal cargo-binding tail domain and patients harboring loss-of-function mutations displayed an extended survival relative to typical ALS cases. Taken together, these results broaden the phenotype spectrum resulting from mutations in KIF5A and strengthen the role of cytoskeletal defects in the pathogenesis of ALS.Peer reviewe

Association of NIPA1 repeat expansions with amyotrophic lateral sclerosis in a large international cohort

NIPA1 (nonimprinted in Prader-Willi/Angelman syndrome 1) mutations are known to cause hereditary spastic paraplegia type 6, a neurodegenerative disease that phenotypically overlaps to some extent with amyotrophic lateral sclerosis (ALS). Previously, a genomewide screen for copy number variants found an association with rare deletions in NIPA1 and ALS, and subsequent genetic analyses revealed that long (or expanded) polyalanine repeats in NIPA1 convey increased ALS susceptibility. We set out to perform a large-scale replication study to further investigate the role of NIPA1 polyalanine expansions with ALS, in which we characterized NIPA1 repeat size in an independent international cohort of 3955 patients with ALS and 2276 unaffected controls and combined our results with previous reports. Meta-analysis on a total of 6245 patients with ALS and 5051 controls showed an overall increased risk of ALS in those with expanded (>8) GCG repeat length (odds ratio = 1.50, p = 3.8×10-5). Together with previous reports, these findings provide evidence for an association of an expanded polyalanine repeat in NIPA1 and ALS

Common and rare variant association analyses in amyotrophic lateral sclerosis identify 15 risk loci with distinct genetic architectures and neuron-specific biology

Amyotrophic lateral sclerosis (ALS) is a fatal neurodegenerative disease with a lifetime risk of one in 350 people and an unmet need for disease-modifying therapies. We conducted a cross-ancestry genome-wide association study (GWAS) including 29,612 patients with ALS and 122,656 controls, which identified 15 risk loci. When combined with 8,953 individuals with whole-genome sequencing (6,538 patients, 2,415 controls) and a large cortex-derived expression quantitative trait locus (eQTL) dataset (MetaBrain), analyses revealed locus-specific genetic architectures in which we prioritized genes either through rare variants, short tandem repeats or regulatory effects. ALS-associated risk loci were shared with multiple traits within the neurodegenerative spectrum but with distinct enrichment patterns across brain regions and cell types. Of the environmental and lifestyle risk factors obtained from the literature, Mendelian randomization analyses indicated a causal role for high cholesterol levels. The combination of all ALS-associated signals reveals a role for perturbations in vesicle-mediated transport and autophagy and provides evidence for cell-autonomous disease initiation in glutamatergic neurons.peer-reviewe

In vitro analysis of amyotrophic lateral sclerosis type 8 and genetic study of spastic paraplegia 4

As doenças do neurônio motor (DNM) apresentam grande variabilidade clínica e genética. A Esclerose Lateral Amiotrófica (ELA) é a forma mais comum de DNM de início tardio. Sua manifestação devastadora e incurável leva a uma profunda perda da qualidade de vida do paciente. A ELA8 é uma forma autossômica dominante de ELA familial causada por mutações no gene VAPB. A proteína VAPB está envolvida com diversos processos celulares. Nossos dados sugerem que a mutação P56S na VAPB diminui a interação dela com outras duas outras proteínas: Tubulina e GAPDH. Por terem sido previamente relacionadas a outras formas de doenças neurodegenerativas, essas proteínas são potenciais pontos-chave para desvendar a patogênese da ELA8 e de outras formas de DNM. Um número significativo de drogas testadas com sucesso em modelos animais de ELA não pôde ser transferido para os pacientes, trazendo à tona a necessidade de novos sistemas de estudo da doença. A tecnologia das iPSC torna possível a reprogramação celular para um estágio de pluripotência, permitindo a modelagem de doenças in vitro. Apresentamos um novo modelo de ELA baseado em células iPSC derivadas de pacientes com ELA8. Este modelo apresenta uma redução dos níveis de VAPB quando comparado com os controles. Contudo, não identificamos os agregados intracitoplasmáticos, característicos dos modelos de super-expressão. Mostramos pela primeira vez que células-tronco embrionárias humanas normais expressam VAPB. Adicionalmente, nossos resultados sugerem que os pacientes com ELA8 teriam uma redução dos níveis de VAPB desde o início de suas vidas e revelam a importância de modelar a ELA num background humano. Dessa forma, sugerimos a hipótese de que quantidades específicas de VAPB sejam cruciais para a viabilidade dos neurônios motores. A busca por alterações no número de cópias levou à identificação do primeiro caso de uma duplicação multi-exônica (exon10_12dup) no gene SPG4, ampliando o espectro de mutações encontradas nesse gene. Esta mutação leva à formação de um códon de parada prematuro, sugerindo que a proteína produzida não é funcional. A análise de 30 indivíduos portadores desta mutação mostrou que os homens possuem, na média, idade de início antecipada e quadro clínico mais severo. Esses dados sugerem que certas doenças neuromusculares podem ser moduladas por fatores relacionados ao background individual e ao gênero. Em suma, apoiamos a idéia de que os microtúbulos estariam envolvidos na patogênese das doenças neuromusculares, visto que tanto a VAPB quanto a Espastina interagem com estes elementos do citoesqueleto. Ainda, apresentamos um novo modelo in vitro de análise da ELA e além de justificar os estudos num background humano.The motor neuron diseases (MND) show a huge clinical and genetic variability. Amyotrophic Lateral Sclerosis (ALS) is the most common late-onset form of MND. ALSs devastating and incurable manifestation leads to a profound loss of life quality. ALS8 is an autosomal dominant form of ALS caused by mutations in the VAPB gene. The VAPB protein is involved with many cellular processes and our data suggest that P56S mutation in this protein reduces its interaction with two other proteins: Tubulin and GAPDH. Since these proteins were previously related to other types of neurodegenerative diseases they potentially are key points to reveal the processes responsible for ALS8 and other MND. A substantial number of successful drug tests in ALS animal models could not be translated to humans, showing the need of novel ALS systems. iPSC technology made possible cellular reprogramming. The iPSC technology brings new hope in this area since it can be used to model diseases in vitro. Here we present a new ALS model based on ALS8-iPSC. Compared to control samples, this model shows a reduction of VAPB levels. However, we could not identify intracytoplasmic aggregates, which characterize overexpression models. We show for the first time that human embryonic stem cells express VAPB. Combined with results showing a VAPB reduction in ALS8 samples, it suggests that ALS8 patients present diminished protein levels since the beginning of their lives and reveal the importance of modeling ALS in a human background. The search for copy number variations has led to the identification of the first multiexonic duplication (exon10_12dup) in SPG4 gene, expanding the mutation spectrum in this gene. This mutation leads to a premature stop codon, which suggests that the expressed protein is not functional. The analysis of 30 individuals who carry the mutation showed that males have on average an earlier AAO and are more severely affected. These data suggest specific neuromuscular diseases can be modulated by factors related to individual background and gender. In sum, we support the idea that microtubules can be involved with neuromuscular disorders pathogenesis, since both VAPB and Spastin interact with these cytoskeleton components. Additionally, we present a new in vitro model to ALS analysis and we justify the studies in a human background