Japanese Amyotrophic Lateral Sclerosis Patients Research Articles

Updated Genetic Analysis of Japanese Familial ALS Patients Carrying SOD1 Variants Revealed Phenotypic Differences for Common Variants.

Amyotrophic lateral sclerosis (ALS) is an adult-onset progressive neurodegenerative disease. Approximately 10% of ALS cases are familial, and more than 20 causative genes have been identified. As we have previously reported, SOD1 variants are the most common causes of familial ALS in Japan. Because antisense oligonucleotides for SOD1-linked ALS are being used in practical applications, the types of variants and the clinical features of patients need to be updated. We consecutively recruited 160 families with familial ALS in Japan. We performed genetic analyses, focusing on SOD1-linked ALS as the most common in our cohort, updated their genotypes, and characterized clinical phenotypes. A total of 26 SOD1 variants in 56 patients and 49 families (30.6%) were collected, with the 3 most common (p.His47Arg [the conventional numbering; H46R], p.Leu127Ser [L126S], p.Asn87Ser [N86S]) accounting for 38.8% of all families. We also identified 2 novel variants (p.Ile36Phe [I35F] and p.Asn132Argfs*3 [N131Rfs*3]). The mean age at onset was 48.9 ± 12.2 (mean ± SD) years for all patients with SOD1-linked ALS. Lower limb onset comprised 70% of cases. The mean disease duration was 64.7 ± 82 months, and the median survival was 71.5 months. Some variants led to a relatively homogeneous phenotype, although clinical characteristics differed among types of variants and families. Patients with p.His47Arg (H46R) showed slower progression with lower limb onset and a predominance of lower motor neuron involvement. The p.Leu127Ser (L126S) variant led to varying degrees of progression in heterozygous or homozygous states and presented incomplete penetrance. Intrafamilial phenotypic differences were observed in families carrying p.Asn87Ser (N86S). Four variants (p.Cys7Gly [C6G], p.His44Arg [H43R], p.Leu85Val [L84V], and p.Cys147Arg [C146R]) were found to be associated with rapid disease progression. The genetic basis of familial ALS, at least for SOD1 variants, still differed by geographic and ethnic background. Understanding these clinical profiles will help optimize evaluation in targeted gene therapy worldwide and benefit efficient diagnosis, leading to precise application in clinical practice.

Endogenous human retrovirus-K is not increased in the affected tissues of Japanese ALS patients

Activation of human endogenous retrovirus-K (HERV-K) is one of the proposed risk factors for amyotrophic lateral sclerosis (ALS). The HERV-K envelope protein has been reported to show neurotoxicity, and development of therapy with reverse transcriptase inhibitors is being investigated. On the other hand, some reports have failed to show HERV-K activation in ALS. In this study, we analyzed the expression of HERV-K mRNA in the motor cortex and spinal cord of 15 Japanese patients with sporadic ALS and 19 controls using reverse transcriptase droplet digital PCR. This revealed no significant increase of HERV-K expression in ALS-affected tissues, suggesting that the association between ALS and HERV-K remains questionable.

Mutation screening of the DNAJC7 gene in Japanese patients with sporadic amyotrophic lateral sclerosis

DNAJC7 has recently been identified as an amyotrophic lateral sclerosis (ALS) gene via large-scale exome analysis, and its involvement in ALS is still unclear in various populations. This study aimed to determine the frequencies and characteristics of the DNAJC7 variants in a Japanese ALS cohort. A total of 807 unrelated Japanese patients with sporadic ALS were screened via exome analysis. In total, we detected six rare missense variants and one splice-site variant of the DNAJC7 gene, which are not reported in the Japanese public database. Furthermore, the missense variants are located around the TPR domain, which is important for the function of DNAJC7. The total frequency of the DNAJC7 variants in Japanese ALS patients was estimated at 0.87%. Collectively, these results suggest that variants of DNAJC7 are rare cause of Japanese patients with sporadic ALS.

SQSTM1L341V variant that is linked to sporadic ALS exhibits impaired association with MAP1LC3 in cultured cells.

Amyotrophic lateral sclerosis (ALS) and frontotemporal dementia (FTD) are genetically, pathologically and clinically-related progressive neurodegenerative diseases. Thus far, several SQSTM1 variations have been identified in patients with ALS and FTD. However, it remains unclear how SQSTM1 variations lead to neurodegeneration. To address this issue, we investigated the effects of ectopic expression of SQSTM1 variants, which were originally identified in Japanese and Chinese sporadic ALS patients, on the cellular viability, their intracellular distributions and the autophagic activity in cultured cells. Expression of SQSTM1 variants in PC12 cells exerted no observable effects on viabilities under both normal and oxidative-stressed conditions. Further, although expression of SQSTM1 variants in PC12 cells and Sqstm1-deficient mouse embryonic fibroblasts resulted in the formation of numerous granular SQSTM1-positive structures, called SQSTM1-bodies, their intracellular distributions were indistinguishable from those of wild-type SQSTM1. Nonetheless, quantitative colocalization analysis of SQSTM1-bodies with MAP1LC3 demonstrated that among ALS-linked SQSTM1 variants, L341V variant showed the significantly lower level of colocalization. However, there were no consistent effects on the autophagic activities among the variants examined. These results suggest that although some ALS-linked SQSTM1 variations have a discernible effect on the intracellular distribution of SQSTM1-bodies, the impacts of other variations on the cellular homeostasis are rather limited at least under transiently-expressed conditions.

The Edinburgh Cognitive and Behavioural ALS Screen (ECAS) for Japanese ALS and FTD patients

Objective Amyotrophic lateral sclerosis (ALS) patients might present with cognitive and behavioural abnormalities resembling frontotemporal dementia (FTD). The Edinburgh Cognitive and Behavioural ALS Screen (ECAS) was developed as an easy to administer cognitive screen for detecting these symptoms. The aim of the present study was to develop and validate a Japanese version of the ECAS. Methods In this single centre observational study, 35 ALS patients and 28 healthy controls were enrolled. Three patients in the ALS group fulfilled the criteria for behavioural variant FTD (ALS-FTD) and the rest were grouped as ALS without FTD. Participants were subjected to the Japanese version of the ECAS. ALS patients were also subjected to the Montreal Cognitive Assessment, Frontal Assessment Battery, ALS Functional Rating Scale-Revised, and respiratory function testing. Demographic and disease characteristics (e.g., sex, age at examination, and years of education) were also recorded. Results Internal consistency and correlations with general cognitive screenings were sufficient in the Japanese adaptation. Executive functions were the most commonly affected ECAS domain, followed by fluency and language. Compared to control subjects, ALS patients without FTD had low scores in the ECAS ALS-specific functions but not in ALS-nonspecific functions. Meanwhile ALS-FTD patients markedly underperformed both in the ECAS ALS-specific and ALS-nonspecific functions. Furthermore, the Japanese ECAS score correlated positively with years of education and negatively with age at onset. Conclusion The Japanese version of the ECAS is a valid and useful screening tool to identify multiple types of cognitive impairment in ALS patients.

Prognosis of amyotrophic lateral sclerosis patients undergoing tracheostomy invasive ventilation therapy in Japan

ObjectiveThe aim of this study is to describe and clarify the factors affecting the prognosis of Japanese patients with amyotrophic lateral sclerosis (ALS) undergoing tracheostomy invasive ventilation (TIV) therapy.MethodsWe conducted...

Cognitive and behavioral status in Japanese ALS patients: a multicenter study

Amyotrophic lateral sclerosis (ALS) patients may present with cognitive and behavioral abnormalities similar to frontotemporal dementia (FTD). In this multicenter study we examined Japanese ALS patients with and without FTD in order to characterize the full extent of cognitive and behavioral abnormalities, including associations with functional motor status, anxiety and depression. Patients were evaluated using the Montreal Cognitive Assessment (MoCA), Frontal Assessment Battery (FAB), Hospital Anxiety and Depression Scale, ALS Functional Rating Scale-Revised, spirometry, and verbal fluency tests. Caregivers were asked to complete the ALS-FTD-Questionnaire (ALS-FTD-Q), a behavioral screen. We defined severe cognitive impairment (MoCA < 21 or FAB < 11), mild impairment (11 ≤ MoCA ≤ 25 or 11 ≤ FAB ≤ 15), and normal cognition (MoCA > 25 or FAB > 15). Severe and mild behavioral impairments and normal behavior were defined by the ALS-FTD-Q scores. In 145 ALS patients, better cognitive scores were correlated with earlier age at onset, whereas a worse behavioral score was associated with a longer disease duration and higher level of anxiety and depression. Around seventy percent of all ALS patients showed mild (40-45%) or severe cognitive impairment with cognitive impairment outnumbering behavioral impairment fivefold. Cognitive functions were more impaired in patients with age of onset over 65years, while behavioral scores were not related to age. Considering the high prevalence of in particular cognitive impairment, and the diversity of impairments, the cognitive and behavioral aspects of Japanese ALS patients should be given more attention clinically.

Diaphragm Pacing Could Have Efficacy for Sleep Condition in Patients with Amyotrophic Lateral Sclerosis

Background: To investigate the efficacy of electrical diaphragm pacing (DP) for sleep condition in patients with amyotrophic lateral sclerosis (ALS). Method: We investigated 5 Japanese ALS patients without positive-pressure mechanical ventilation (3 men and 2 women, aged 59.6 ± 9.6 years). All of them were implanted NeuRx RA/4 Diaphragm Pacing System R (NeuRx) in the diaphragm laparoscopically. We assessed physical status and polysomnographic recordings before implantation and after 6 months of conditioning with DP turned on. Results: 4 patients completed the evaluation. Sleep condition showed the tendency to improve far from getting worse despite of deterioration of physical status and respiratory function. Conclusion: DP with NeuRx might assist the sleep condition in ALS patients at least in 6 months of follow up.

Difficulty in determining the association of a single nucleotide polymorphism in the ZNF512B gene with the risk and prognosis of amyotrophic lateral sclerosis.

The advent of next-generation sequencing technology is expected to accelerate the identification of novel genes, and this technology will likely supersede Sanger sequencing. Thus, genome-wide association studies (GWASs) are performed more routinely in an effort to identify disease-susceptibility genes for sporadic amyotrophic lateral sclerosis (ALS). Previously, a Japanese team conducted a large-scale GWAS with 1,305 Japanese ALS patients and discovered a new single nucleotide polymorphism (SNP) rs2275294 associated with susceptibility to sporadic ALS (sALS) in the ZNF512B gene on chromosome 20q13.33. Ju et al. recently performed a case-control study to examine the possible association of rs2275294 with the risk of sALS. Their results, however, indicated that the SNP in ZNF512B is not associated with sALS susceptibility in the Chinese population. A precise diagnosis of neurodegenerative diseases, especially ALS, is highly challenging. For GWASs and other clinical research studies that require a large sample size, if true ALS patients are not selected initially, then all subsequent research is futile. Here, I evaluate the factors that are likely responsible for the inconsistent results obtained by GWASs and propose the development of a new classification system and diagnostic criteria for ALS as the first step towards conducting better clinical studies on ALS. I have attempted to explain the reasons for the inconsistent association between rs2275294 and ALS progression by listing the gene-gene/gene-environment interactions, age of onset, sample size, odds ratio, and inappropriate ALS diagnosis criteria for stratifying this heterogeneous disease in this review.

Next-generation sequencing of 28 ALS-related genes in a Japanese ALS cohort

We investigated the frequency and contribution of variants of the 28 known amyotrophic lateral sclerosis (ALS)-related genes in Japanese ALS patients. We designed a multiplex, polymerase chain reaction–based primer panel to amplify the coding regions of the 28 ALS-related genes and sequenced DNA samples from 257 Japanese ALS patients using an Ion Torrent PGM sequencer. We also performed exome sequencing and identified variants of the 28 genes in an additional 251 ALS patients using an Illumina HiSeq 2000 platform. We identified the known ALS pathogenic variants and predicted the functional properties of novel nonsynonymous variants in silico. These variants were confirmed by Sanger sequencing. Known pathogenic variants were identified in 19 (48.7%) of the 39 familial ALS patients and 14 (3.0%) of the 469 sporadic ALS patients. Thirty-two sporadic ALS patients (6.8%) harbored 1 or 2 novel nonsynonymous variants of ALS-related genes that might be deleterious. This study reports the first extensive genetic screening of Japanese ALS patients. These findings are useful for developing genetic screening and counseling strategies for such patients.

Recurrent K3E mutation in Cu/Zn superoxide dismutase gene associated with amyotrophic lateral sclerosis

Cu/Zn superoxide dismutase (SOD1) gene mutations are the most frequently reported genetic causes of amyotrophic lateral sclerosis (ALS). The objective of the study was to describe a clinical phenotype and haplotype background of Polish and Japanese ALS patients harbouring the K3E SOD1 mutation. The K3E mutation was identified by direct sequencing, high resolution melting analysis or high-throughput microarray-based resequencing system. Microsatellite polymorphic markers flanking SOD1 were genotyped in members of six kindreds and two SALS patients. Results demonstrated that the K3E mutation was responsible for classic ALS. The median age of onset was 54 years. The clinical phenotype did not substantially differ between SALS and FALS cases of either ethnic origin, with some intrafamiliar variabilities. There was a limb onset in 92% of patients. In patients with bulbar syndrome, dysphagia predominated over dysarthria. Respiratory insufficiency was found in 61.1% of patients (19–84 months after the first symptoms onset). Median survival was 101 months with age of death ranging from 45 to 77 years. K3E was the most frequent SOD1 mutation among Polish FALS patients. It originated independently, on different haplotype background in the Polish and Japanese populations. In conclusion, recurrent K3E mutation results in a relatively slowly progressing limb onset ALS with classic phenotype.

Reduction rate of body mass index predicts prognosis for survival in amyotrophic lateral sclerosis: A multicenter study in Japan

Malnutrition in the early stage has been reported as an independent predictor of survival in amyotrophic lateral sclerosis (ALS). We analyzed retrospectively the effect of variation of body mass index (BMI) on survival in ALS patients. In total, 77 consecutive ALS patients were enrolled from nine hospitals in Japan. Reduction rate of BMI was calculated from BMI before the disease onset and at the time of the first visit to each hospital. We analyzed the correlation between BMI reduction rate and total disease duration. Results showed that the median BMI reduction rate was 2.5 per year (interquartile range 1.3-3.8). The BMI reduction rate was significantly correlated with survival length (p <0.0001). There was also a significant difference in survival between ALS patients with a BMI reduction rate ≥ and < 2.5 (Kaplan-Meier survival analysis and the log-rank test, p < 0.0001; hazard ratio by the Cox model, 2.9816). In conclusion, faster reduction of BMI at the initial stage before the first visit to hospital predicts shorter survival length also in Japanese ALS patients.

JaCALS: a prospective multicenter ALS cohort study

To investigate the longitudinal course of Japanese patients with Amyotrophic Lateral Sclerosis (ALS), we constructed a multicenter registration and follow-up system called Japanese Consortium for Amyotrophic Lateral Sclerosis research (JaCALS). Genomic DNA samples of ALS patients were stored and linked to the clinical information. We designed a telephone survey system using a clinical research coordinator (CRC) to check the score of the ALS Functional Rating Scale-R (ALSFRS-R) and the prognosis every 3 months. In January 2006, we began registering ALS patients, and, at present, 22 neurology facilities are participating in the JaCALS. Currently, 571 Japanese ALS patients are registered. From the longitudinal data of the 279 patients who were registered before September 2009, the older age at onset was a significant risk factor for not only earlier death or introduction of mechanical ventilation, but also earlier loss of speech, loss of swallowing function and loss of upper limb function. In collaboration with the RIKEN Center for Genomic Medicine, genome-wide association studies (GWAS) using 1,305 ALS samples from the JaCALS and BioBank Japan were conducted, which showed that ZNF512B gene was associated with susceptibility to ALS. The JaCALS has established an efficient registration and follow-up system with genomic DNA resources of ALS patients, and will contribute to identify ALS-associated genes and to promote clinical researches.

Multiple system degeneration with basophilic inclusions in Japanese ALS patients with FUS mutation

Mutations in the fused in sarcoma gene (FUS) were recently found in patients with familial amyotrophic lateral sclerosis (ALS). The present study aimed to clarify unique features of familial ALS caused by FUS mutation in the Japanese population. We carried out clinical, neuropathological, and genetic studies on a large Japanese pedigree with familial ALS. In six successive generations of this family, 16 individuals of both sexes were affected by progressive muscle atrophy and weakness, indicating an autosomal dominant trait. Neurological examination of six patients revealed an age at onset of 48.2+/-8.1 years in fourth generation patients, while it was 31 and 20 years in fifth and sixth generation patients, respectively. Motor paralysis progressed rapidly in these patients, culminating in respiratory failure within 1 year. The missense mutation c.1561 C>T (p.R521C) was found in exon 15 of FUS in the four patients examined. Neuropathological study of one autopsied case with the FUS mutation revealed multiple system degeneration in addition to upper and lower motor neuron involvement: the globus pallidus, thalamus, substantia nigra, cerebellum, inferior olivary nucleus, solitary nucleus, intermediolateral horn, Clarke's column, Onuf's nucleus, central tegmental tract, medial lemniscus, medial longitudinal fasciculus, superior cerebellar peduncle, posterior column, and spinocerebellar tract were all degenerated. Argyrophilic and basophilic neuronal or glial cytoplasmic inclusions immunoreactive for FUS, GRP78/BiP, p62, and ubiquitin were detected in affected lesions. The FUS R521C mutation in this Japanese family caused familial ALS with pathological features of multiple system degeneration and neuronal basophilic inclusions.

Immunohistochemical expression of IGF‐I and GSK in the spinal cord of Kii and Guamanian ALS patients

Insulin-like growth factor-I (IGF-I) is a potent survival factor for motor neurons in animals, and glycogen synthase kinase-3beta (GSK-3beta) is suspected to play roles in apoptosis and tau phosphorylation. Here we report the immunological expression of IGF-I, GSK-3beta, phosphorylated-GSK-3alpha/beta (p-GSK-3alpha/beta) and phosphorylated-tau in the spinal cord and hippocampus of Kii and Guam amyotrophic lateral sclerosis (ALS) patients. Sixteen ALS patients (10 Japanese sporadic, 3 Kii and 3 Guam ALS) and 14 neurological controls (10 Japanese and 4 Guamanian) were examined. The immunoreactivity for each antibody was rated by the percentages of positive neurons to total anterior horn neurons in each patient and was analyzed statistically. Many normal-looking neurons from Japanese sporadic ALS, Kii ALS and Guam ALS patients, as well as from Japanese and Guam controls, were positive for anti-IGF-I antibody. A positive correlation between IR scores for anti-IGF-I antibody and clinical durations of Japanese sporadic ALS patients was found in this study (P < 0.0001). This suggested that IGF-I might have a protective effect against ALS degeneration. In Japanese sporadic ALS patients, abnormal as well as normal-looking neurons showed significant high IR scores for anti-GSK-3beta antibody than those of controls. Anterior horn neurons from Guam and Kii ALS patients characteristically showed weak staining for anti-GSK-3beta antibody but were markedly positive for anti-pGSK-3alpha/beta antibody compared to those from both Japanese controls and Japanese sporadic ALS patients, and showed the co-localization of IGF-I and p-GSK-3alpha/beta. This suggested that the IGF-I signaling pathway in Guam and Kii ALS patients might function to phosphorylate GSK-3beta to protect neurons from ALS degeneration. Neurofibrillary tangles (NFTs) in the hippocampus and spinal cord from Kii and Guam ALS patients showed the co-localization of PHF-tau and p-GSK-3alpha/beta by a confocal laser scanning technique. The predominant expression of p-GSK-3alpha/beta compared to GSK-3beta in spinal motor neurons and the co-localization of p-GSK-3alpha/beta and PHF-tau in NFT-laden neurons in the hippocampus and spinal cord were characteristic findings of Kii and Guam ALS patients.