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Objectives Clinical diagnostics in adults with hereditary neurological diseases is complicated by clinical and genetic heterogeneity, as well as lifestyle effects. Here, we evaluate the effectiveness of exome sequencing and clinical costs in our difficult-to-diagnose adult patient cohort. Additionally, we expand the phenotypic and genetic spectrum of hereditary neurological disorders in Finland. Methods We performed clinical exome sequencing (CES) to 100 adult patients from Finland with neurological symptoms of suspected genetic cause. The patients were classified as myopathy (n = 57), peripheral neuropathy (n = 16), ataxia (n = 15), spastic paraplegia (n = 4), Parkinsonism (n = 3), and mixed (n = 5). In addition, we gathered the costs of prior diagnostic work-up to retrospectively assess the cost-effectiveness of CES as a first-line diagnostic tool. Results The overall diagnostic yield of CES was 27%. Pathogenic variants were found for 14 patients (in genes ANO5, CHCHD10, CLCN1, DES, DOK7, FKBP14, POLG, PYROXD1, SCN4A, TUBB3, and TTN) and likely pathogenic previously undescribed variants for 13 patients (in genes ABCD1, AFG3L2, ATL1, CACNA1A, COL6A1, DYSF, IRF2BPL, KCNA1, MT-ATP6, SAMD9L, SGCB, and TPM2). Age of onset below 40 years increased the probability of finding a genetic cause. Our cost evaluation of prior diagnostic work-up suggested that early CES would be cost-effective in this patient group, in which diagnostic costs increase linearly with prolonged investigations. Conclusions Based on our results, CES is a cost-effective, powerful first-line diagnostic tool in establishing the molecular diagnosis in adult neurological patients with variable symptoms. Importantly, CES can markedly shorten the diagnostic odysseys of about one third of patients.

The aetiology of dystonia disorders is complex, and next-generation sequencing has become a useful tool in elucidating the variable genetic background of these diseases. Here we report a deleterious heterozygous truncating variant in the inosine monophosphate dehydrogenase gene (IMPDH2) by whole-exome sequencing, co-segregating with a dominantly inherited dystonia-tremor disease in a large Finnish family. We show that the defect results in degradation of the gene product, causing IMPDH2 deficiency in patient cells. IMPDH2 is the first and rate-limiting enzyme in the de novo biosynthesis of guanine nucleotides, a dopamine synthetic pathway previously linked to childhood or adolescence-onset dystonia disorders. We report IMPDH2 as a new gene to the dystonia disease entity. The evidence underlines the important link between guanine metabolism, dopamine biosynthesis and dystonia.

Mutations in mitochondrial DNA encoded subunit of ATP synthase, MT-ATP6, are frequent causes of neurological mitochondrial diseases with a range of phenotypes from Leigh syndrome and NARP to ataxias and neuropathies. Here we investigated the functional consequences of an unusual heteroplasmic truncating mutation m.9154C>T in MT-ATP6, which caused peripheral neuropathy, ataxia and IgA nephropathy. ATP synthase not only generates cellular ATP, but its dimerization is required for mitochondrial cristae formation. Accordingly, the MT-ATP6 truncating mutation impaired the assembly of ATP synthase and disrupted cristae morphology, supporting our molecular dynamics simulations that predicted destabilized a/c subunit subcomplex. Next, we modeled the effects of the truncating mutation using patient-specific induced pluripotent stem cells. Unexpectedly, depending on mutation heteroplasmy level, the truncation showed multiple threshold effects in cellular reprogramming, neurogenesis and in metabolism of mature motor neurons (MN). Interestingly, MN differentiation beyond progenitor stage was impaired by Notch hyperactivation in the MT-ATP6 mutant, but not by rotenone-induced inhibition of mitochondrial respiration, suggesting that altered mitochondrial morphology contributed to Notch hyperactivation. Finally, we also identified a lower mutation threshold for a metabolic shift in mature MN, affecting lactate utilization, which may be relevant for understanding the mechanisms of mitochondrial involvement in peripheral motor neuropathies. These results establish a critical and disease-relevant role for ATP synthase in human cell fate decisions and neuronal metabolism.

OBJECTIVE: ITPR3, encoding inositol 1,4,5-trisphosphate receptor type 3, was previously reported as a potential candidate disease gene for Charcot-Marie-Tooth neuropathy. Here, we present genetic and functional evidence that ITPR3 is a Charcot-Marie-Tooth disease gene. METHODS: Whole-exome sequencing of four affected individuals in an autosomal dominant family and one individual who was the only affected individual in his family was used to identify disease-causing variants. Skin fibroblasts from two individuals of the autosomal dominant family were analyzed functionally by western blotting, quantitative reverse transcription PCR, and Ca2+ imaging. RESULTS: Affected individuals in the autosomal dominant family had onset of symmetrical neuropathy with demyelinating and secondary axonal features at around age 30, showing signs of gradual progression with severe distal leg weakness and hand involvement in the proband at age 64. Exome sequencing identified a heterozygous ITPR3 p.Val615Met variant segregating with the disease. The individual who was the only affected in his family had disease onset at age 4 with demyelinating neuropathy. His condition was progressive, leading to severe muscle atrophy below knees and atrophy of proximal leg and hand muscles by age 16. Trio exome sequencing identified a de novo ITPR3 variant p.Arg2524Cys. Altered Ca2+ -transients in p.Val615Met patient fibroblasts suggested that the variant has a dominant-negative effect on inositol 1,4,5-trisphosphate receptor type 3 function. INTERPRETATION: Together with two previously identified variants, our report adds further evidence that ITPR3 is a disease-causing gene for CMT and indicates altered Ca2+ homeostasis in disease pathogenesis. ispartof: ANNALS OF CLINICAL AND TRANSLATIONAL NEUROLOGY vol:7 issue:10 pages:1962-1972 ispartof: location:United States status: published

Background and ObjectivesTo determine the genetic cause of the disease in the previously reported family with adult-onset autosomal dominant distal myopathy (myopathy, distal, 3; MPD3).MethodsContinued clinical evaluation including muscle MRI and muscle pathology. A linkage analysis with single nucleotide polymorphism arrays and genome sequencing were used to identify the genetic defect, which was verified by Sanger sequencing. RNA sequencing was used to investigate the transcriptional effects of the identified genetic defect.ResultsSmall hand muscles (intrinsic, thenar, and hypothenar) were first involved with spread to the lower legs and later proximal muscles. Dystrophic changes with rimmed vacuoles and cytoplasmic inclusions were observed in muscle biopsies at advanced stage. A single nucleotide polymorphism array confirmed the previous microsatellite-based linkage to 8p22-q11 and 12q13-q22. Genome sequencing of three affected family members combined with structural variant calling revealed a small heterozygous deletion of 160 base pairs spanning the second last exon 10 of the heterogeneous nuclear ribonucleoprotein A1 (HNRNPA1) gene, which is in the linked region on chromosome 12. Segregation of the mutation with the disease was confirmed by Sanger sequencing. RNA sequencing showed that the mutant allele produces a shorter mutant mRNA transcript compared with the wild-type allele. Immunofluorescence studies on muscle biopsies revealed small p62 and larger TDP-43 inclusions.DiscussionA small exon 10 deletion in the gene HNRNPA1 was identified as the cause of MPD3 in this family. The new HNRNPA1-related phenotype, upper limb presenting distal myopathy, was thus confirmed, and the family displays the complexities of gene identification.

Rhabdomyolysis is the acute breakdown of skeletal myofibres in response to an initiating factor, most commonly toxins and over exertion. A variety of genetic disorders predispose to rhabdomyolysis through different pathogenic mechanisms, particularly in patients with recurrent episodes. However, the majority of cases remain without a genetic diagnosis. Here we present six patients who presented with severe and recurrent rhabdomyolysis, usually with onset in the teenage years; other features included a history of myalgia and muscle cramps. We identified ten bi-allelic loss-of-function variants in the gene encoding obscurin (OBSCN) co-segregating with disease. We show reduced expression of OBSCN and loss of obscurin protein in patient muscle. Obscurin is proposed to be involved in SR function and Ca2+ handling. Patient cultured myoblasts appear more susceptible to starvation as evidenced by a greater decreased in SR Ca2+ content compared to control myoblasts. This likely reflects a lower efficiency when pumping Ca2+ back into the SR and/or a decrease in Ca2+ SR storage ability when metabolism is diminished. OSBCN variants have previously been associated with cardiomyopathies. None of the patients presented with a cardiomyopathy and cardiac examinations were normal in all cases in which cardiac function was assessed. There was also no history of cardiomyopathy in first degree relatives, in particular in any of the carrier parents. This cohort is relatively young, thus follow-up studies and the identification of additional cases with bi-allelic null OBSCN variants will further delineate OBSCN-related disease and the clinical course of disease.

ObjectiveTo characterize serum biomarkers in mitochondrial CHCHD10-linked spinal muscular atrophy Jokela type (SMAJ) for disease monitoring and for understanding of pathogenic mechanisms.MethodsWe collected serum samples from a cohort of 49 SMAJ patients, all carriers of the heterozygous c.197G>T p.G66V variant inCHCHD10. As controls, we used age- and sex-matched serum samples obtained from Helsinki Biobank. Neurofilament light (NfL) and glial fibrillary acidic protein (GFAP) were measured with Single molecule array (Simoa), and fibroblast growth factor 21 (FGF-21) and growth differentiation factor 15 (GDF-15) with enzyme-linked immunosorbent assay. For nontargeted serum metabolite profiling, samples were analyzed with liquid chromatography–high resolution mass spectrometry. Disease severity was evaluated retrospectively by calculating a symptom-based score.ResultsAxon degeneration marker NfL was unexpectedly not altered in the serum of SMAJ patients, whereas astrocytic activation marker GFAP was slightly decreased. Creatine kinase was elevated in most patients, in particular males. We identified six metabolites that were significantly altered in SMAJ patients’ serum compared to controls: increased creatine and pyruvate, and decreased creatinine, taurine, N-acetyl-carnosine and succinate. Creatine correlated with disease severity. Altered pyruvate and succinate indicated a metabolic response to mitochondrial dysfunction, however, lactate or mitochondrial myopathy markers FGF-21 or GDF-15 were not changed.ConclusionsBiomarkers of muscle mass and damage are altered in SMAJ serum, indicating a role for skeletal muscle in disease pathogenesis in addition to neurogenic damage. Despite the minimal mitochondrial pathology in skeletal muscle, signs of a metabolic shift can be detected in the serum of SMAJ patients.

PURPOSE: To compare the value of serum biomarkers, FGF21 and GDF15 with histological analysis of muscle in the diagnosis of mitochondrial disease. METHODS: We collected 194 serum samples from patients with a suspected or known mitochondrial disease. Biomarkers were analyzed blinded using enzyme labelled immunosorbent assay (ELISA). Clinical data was collected using a structured questionnaire. RESULTS: Only 39% of patients with genetically verified mitochondrial disease had mitochondrial pathology in their muscle histology. In contrast, biomarkers were elevated in 62% of patients with genetically verified mitochondrial disease. Those with both biomarkers elevated had a muscle manifesting disorder and a defect affecting mitochondrial DNA expression. If at least one of the biomarkers was induced and the patient had a myopathic disease, a mitochondrial DNA expression disease was the cause with 94% probability. Among patients with biomarker analysis and muscle biopsy taken

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The Polyneuropathy And Treatment with Hizentra (PATH) study required subjects with chronic inflammatory demyelinating polyneuropathy (CIDP) to show dependency on immunoglobulin G (IgG) and then be restabilized on IgG before being randomized to placebo or one of two doses of subcutaneous immunoglobulin (SCIG). Nineteen of the 51 subjects (37%) randomized to placebo did not relapse over the next 24 weeks. This article explores the reasons for this effect. A post‐hoc analysis of the PATH placebo group was undertaken. A literature search identified other placebo‐controlled CIDP trials for review and comparison. In PATH, subjects randomized to placebo who did not relapse were significantly older, had more severe disease, and took longer to deteriorate in the IgG dependency period compared with those who relapsed. Published trials in CIDP, whose primary endpoint was stability or deterioration, had a mean non‐deterioration (placebo effect) of 43%, while trials with a primary endpoint of improvement had a placebo response of only 11%. Placebo is an important variable in the design of CIDP trials. Trials designed to show clinical improvement will have a significantly lower effect of this phenomenon than those designed to show stability or deterioration.

NAD(+) is a redox-active metabolite, the depletion of which has been proposed to promote aging and degenerative diseases in rodents. However, whether NAD(+) depletion occurs in patients with degenerative disorders and whether NAD(+) repletion improves their symptoms has remained open. Here, we report systemic NAD(+) deficiency in adult-onset mitochondrial myopathy patients. We administered an increasing dose of NAD(+) booster niacin, a vitamin B3 form (to 750-1,000 mg/day; clinicaltrials.gov NCT03973203) for patients and their matched controls for 10 or 4 months, respectively. Blood NAD(+) increased in all subjects, up to 8-fold, and muscle-NAD(+) of patients reached the level of their controls. Some patients showed anemia tendency, while muscle strength and mitochondrial biogenesis increased in all subjects. In patients, muscle metabolome shifted toward controls and liver fat decreased even 50%. Our evidence indicates that blood analysis is useful in identifying NAD(+) deficiency and points niacin to be an efficient NAD(+) booster for treating mitochondrial myopathy.

Background: A deficiency of muscle phosphofructokinase (PFKM) causes a rare metabolic muscle disease, the Tarui disease (Glycogen storage disease type VII, GSD VII) characterized by exercise intolerance with myalgia due to an inability to use glucose as an energy resource. No medical treatment for GSD VII currently exists. The aim of this study was to determine whether a dietary intervention with excessive fat intake would benefit GSD VII.Patient and Methods: A ketogenic diet (KD) intervention implemented as a modified Atkins diet was established for one patient with PFKM deficiency, with a low late lactate response and very high ammonia levels associated with exercise. We recorded the KD intervention for a total of 5 years with clinical and physiotherapeutic evaluations and regular laboratory parameters. Cardiopulmonary exercise testing, including breath gas analysis and venous lactate and ammonia measurements, was performed before KD and at 3, 8 months and 5 years after initiation of KD.Results: During the 5 years on KD, the patient's muscle symptoms had alleviated and exercise tolerance had improved. In exercise testing, venous ammonia had normalized, the lactate profile remained similar, but oxygen uptake and mechanical efficiency had increased and parameters showing ventilation had improved.Conclusions: This study is the first to show a long-term effect of KD in GSD VII with an alleviation of muscle symptoms, beneficial effects on breathing, and improvement in exercise performance and oxygen uptake. Based on these findings, KD can be recommended under medical and nutritional supervision for selected patients with GSD VII, although further research of this rare disease is warranted.

Mitochondrial dysfunction elicits stress responses that safeguard cellular homeostasis against metabolic insults. Mitochondrial integrated stress response (ISRmt) is a major response to mitochondrial (mt)DNA expression stress (mtDNA maintenance, translation defects), but the knowledge of dynamics or interdependence of components is lacking. We report that in mitochondrial myopathy, ISRmt progresses in temporal stages and development from early to chronic and is regulated by autocrine and endocrine effects of FGF21, a metabolic hormone with pleiotropic effects. Initial disease signs induce transcriptional ISRmt (ATF5, mitochondrial one-carbon cycle, FGF21, and GDF15). The local progression to 2nd metabolic ISRmt stage (ATF3, ATF4, glucose uptake, serine biosynthesis, and transsulfuration) is FGF21 dependent. Mitochondrial unfolded protein response marks the 3rd ISRmt stage of failing tissue. Systemically, FGF21 drives weight loss and glucose preference, and modifies metabolism and respiratory chain deficiency in a specific hippocampal brain region. Our evidence indicates that FGF21 is a local and systemic messenger of mtDNA stress in mice and humans with mitochondrial disease.

INTRODUCTION: We aimed to explore the value of genetic screening for Fabry disease (FD) and hereditary ATTR amyloidosis in patients with idiopathic small fiber neuropathy (SFN) or mixed neuropathy in a clinical setting.METHODS: This was a Nordic multicenter study with 9 participating centers. Patients with idiopathic SFN or mixed neuropathy were included. Genetic sequencing of the TTR and GLA genes was performed.RESULTS: 172 patients were enrolled. Genetic screening was performed in 155 patients. No pathogenic mutations in the TTR gene were found. A single patient had a possible pathogenic variant, R118C, in the GLA gene, but the clinical investigation showed no firm signs of FD.DISCUSSION: Screening for hereditary ATTR amyloidosis and FD in patients with idiopathic SFN or mixed neuropathy without any additional disease-specific symptoms or clinical characteristics in a Nordic population appears to be of little value in a clinical setting. This article is protected by copyright. All rights reserved.

PATH study group., In patients with chronic inflammatory demyelinating polyneuropathy (CIDP), intravenous immunoglobulin (IVIG) is recommended to be periodically reduced to assess the need for ongoing therapy. However, little is known about the effectiveness of restabilization with IVIG in patients who worsen after IVIG withdrawal. In the Polyneuropathy And Treatment with Hizentra (PATH) study, the pre‐randomization period included sudden stopping of IVIG followed by 12 weeks of observation. Those deteriorating were then restabilized with IVIG. Of 245 subjects who stopped IVIG, 28 did not show signs of clinical deterioration within 12 weeks. Two hundred and seven received IVIG restabilization with an induction dose of 2 g/kg bodyweight (bw) IgPro10 (Privigen, CSL Behring, King of Prussia, Pennsylvania) and maintenance doses of 1 g/kg bw every 3 weeks for up to 13 weeks. Signs of clinical improvement were seen in almost all (n = 188; 91%) subjects. During IVIG restabilization, 35 subjects either did not show CIDP stability (n = 21, analyzed as n = 22 as an additional subject was randomized in error) or withdrew for other reasons (n = 14). Of the 22 subjects who did not achieve clinical stability, follow‐up information in 16 subjects after an additional 4 weeks was obtained. Nine subjects were reported to have improved, leaving a maximum of 27 subjects (13%) who either showed no signs of clinical improvement during the restabilization phase and 4 weeks post‐study or withdrew for other reasons. In conclusion, sudden IVIG withdrawal was effective in detecting ongoing immunoglobulin G dependency with a small risk for subjects not returning to their baseline 17 weeks after withdrawal., Editorial support was provided by Meridian HealthComms Ltd, funded by CSL Behring.

Dermal fibroblasts from patients harboring mutations in the gap junction connexin43 gene have distinctly different mechanistic causes for the clinical presentations of oculodentodigital dysplasia., Oculodentodigital dysplasia (ODDD) is a rare genetic disease that affects the development of multiple organs in the human body. More than 70 mutations in the gap junction connexin43 (Cx43) gene, GJA1, are associated with ODDD, most of which are inherited in an autosomal dominant manner. Many patients exhibit similar clinical presentations. However, there is high intrafamilial and interfamilial phenotypic variability. To better understand this variability, we established primary human dermal fibroblast cultures from several ODDD patients and unaffected controls. In the present study, we characterized three fibroblast lines expressing heterozygous p.L7V, p.G138R, and p.G143S Cx43 variants. All ODDD fibroblasts exhibited slower growth, reduced migration, and defective cell polarization, traits common to all ODDD fibroblasts studied so far. However, we found striking differences in overall expression levels, with p.L7V down-regulated at the mRNA and protein level. Although all of the Cx43 variants could traffic to the cell surface, there were stark differences in gap junction plaque formation, gap junctional intercellular communication, Cx43 phosphorylation, and hemichannel activity among Cx43 variants, as well as subtle differences in myofibroblast differentiation. Together these findings enabled us to discover mutation-specific pathologies that may help to predict future clinical outcomes.

Following the involvement of CHCHD10 in FrontoTemporal-Dementia-Amyotrophic Lateral Sclerosis (FTD-ALS) clinical spectrum, a founder mutation (p.Gly66Val) in the same gene was identified in Finnish families with late onset spinal motor neuronopathy (SMAJ). SMAJ is a slowly progressive form of spinal muscular atrophy with a life expectancy within normal range. In order to understand why the p.Ser59Leu mutation, responsible for severe FTD-ALS, and the p.Gly66Val mutation could lead to different levels of severity, we compared their effects in patient cells. Unlike affected individuals bearing the p.Ser59Leu mutation, patients presenting with SMAJ phenotype have neither mitochondrial myopathy nor mtDNA instability. The expression of CHCHD10(S59L) mutant allele leads to disassembly of mitochondrial contact site and cristae organizing system (MICOS) with mitochondria] dysfunction and loss of cristae in patient fibroblasts. We also show that G66V fibroblasts do not display the loss of MICOS complex integrity and mitochondrial damage found in S59L cells. However, S59L and G66V fibroblasts show comparable accumulation of phosphorylated mitochondrial TDP-43 suggesting that the severity of phenotype and mitochondrial damage do not depend on mitochondrial TDP-43 localization. The expression of the CHCHD10(G66V) allele is responsible for mitochondrial network fragmentation and decreased sensitivity towards apoptotic stimuli, but with a less severe effect than that found in cells expressing the CHCHD10(S59L) allele. Taken together, our data show that cellular phenotypes associated with p.Ser59Leu and p.Gly66Val mutations in CHCHD10 are different; loss of MICOS complex integrity and mitochondrial dysfunction, but not TDP-43 mitochondrial localization, being likely essential to develop a severe motor neuron disease.

Key Clinical Message Patients with unknown clinical or radiological asymmetry in the face structures combined with atrophy and weakness of the masticatory muscles should be comprehensively examined clinically and with MRI, neurophysiological measurements, and serologically. Malignant lesions or benign idiopathic unilateral trigeminal motor neuropathy should be considered as an etiological explanation for the asymmetry.

Mika Saarela työryhmän jäsenenä. Background Approximately two-thirds of patients with chronic inflammatory demyelinating polyneuropathy (CIDP) need long-term intravenous immunoglobulin. Subcutaneous immunoglobulin (SCIg) is an alternative option for immunoglobulin delivery, but has not previously been investigated in a large trial of CIDP. The PATH study compared relapse rates in patients given SCIg versus placebo. Methods Between March 12, 2012, and Sept 20, 2016, we studied patients from 69 neuromuscular centres in North America, Europe, Israel, Australia, and Japan. Adults with definite or probable CIDP who responded to intravenous immunoglobulin treatment were eligible. We randomly allocated participants to 0.2 g/kg or 0.4 g/kg of a 20% SCIg solution (IgPro20) weekly versus placebo (2% human albumin solution) for maintenance treatment for 24 weeks. We did randomisation in a 1: 1:1 ratio with an interactive voice and web response system with a block size of six, stratified by region (Japan or non-Japan). The primary outcome was the proportion of patients with a CIDP relapse or who were withdrawn for any other reason during 24 weeks of treatment. Patients, caregivers, and study personnel, including those assessing outcomes, were masked to treatment assignment. Analyses were done in the intention-to-treat and per-protocol sets. This trial is registered with ClinicalTrials. gov, number NCT01545076. Findings In this randomised, double-blind, placebo-controlled trial, we randomly allocated 172 patients: 57 (33%) to the placebo group, 57 (33%) to the low-dose group, and 58 (34%) to the high-dose group. In the intention-to-treat set, 36 (63% [95% CI 50-74]) patients on placebo, 22 (39% [27-52]) on low-dose SCIg, and 19 (33% [22-46]) on high-dose SCIg had a relapse or were withdrawn from the study for other reasons (p=0.0007). Absolute risk reductions were 25% (95% CI 6-41) for low-dose versus placebo (p=0.007), 30% (12-46) for high-dose versus placebo (p=0.001), and 6% (-11 to 23) for high-dose versus low-dose (p=0.32). Causally related adverse events occurred in 47 (27%) patients (ten [18%] in the placebo group, 17 [30%] in the low-dose group, and 20 [34%] in the high-dose group). Six (3%) patients had 11 serious adverse events: one (2%) patient in the placebo group, three (5%) in the low-dose group, and two (3%) in the high-dose group; only one (an acute allergic skin reaction in the low-dose group) was assessed to be causally related. Interpretation This study, which is to our knowledge, the largest trial of CIDP to date and the first to study two administrations of immunoglobulins and two doses, showed that both doses of SCIg IgPro20 were efficacious and well tolerated, suggesting that SCIg can be used as a maintenance treatment for CIDP.

DNAJB6 is the causative gene for limb-girdle muscular dystrophy 1D (LGMD1D). Four different coding missense mutations, p.F89I, p.F93I, p.F93L, and p.P96R, have been reported in families from Europe, North America and Asia. The previously known mutations cause mainly adult-onset proximal muscle weakness with moderate progression and without respiratory involvement. A Finnish family and a British patient have been studied extensively due to a severe muscular dystrophy. The patients had childhood-onset LGMD, loss of ambulation in early adulthood and respiratory involvement; one patient died of respiratory failure aged 32. Two novel mutations, c.271T > A (p.F91I) and c.271T > C (p.F91L), in DNAJB6 were identified by whole exome sequencing as a cause of this severe form of LGMD1D. The results were confirmed by Sanger sequencing. The anti-aggregation effect of the mutant DNAJB6 was investigated in a filter-trap based system using transient transfection of mammalian cell lines and polyQ-huntingtin as a model for an aggregation-prone protein. Both novel mutant proteins show a significant loss of ability to prevent aggregation. (C) 2015 Elsevier B.V. All rights reserved.

Variants in family 1 kinesin (KIF1A), which encodes a kinesin axonal motor protein, have been described to cause variable neurological manifestations. Recessive missense variants have led to spastic paraplegia, and recessive truncations to sensory and autonomic neuropathy. De novo missense variants cause developmental delay or intellectual disability, cerebellar atrophy and variable spasticity. We describe a family with father-to-son transmission of de novo variant in the KIF1A motor domain, in a phenotype of pure spastic paraplegia. Structural modeling of the predicted p.(Ser69Leu) amino acid change suggested that it impairs the stable binding of ATP to the KIF1A protein. Our study reports the first dominantly inherited KIF1A variant and expands the spectrum of phenotypes caused by heterozygous KIF1A motor domain variants to include pure spastic paraplegia. We conclude that KIF1A should be considered a candidate gene for hereditary paraplegias regardless of inheritance pattern.

Hereditary sensory neuropathy type 1 (HSAN1) may be the first genetic neuropathy amenable to a specific mechanism-based treatment, as L-serine supplementation can be used to lower the neurotoxic levels of 1-deoxysphingolipids (1-deoxySL) that cause the neurodegeneration. The treatment is so far untested in HSAN1C caused by variants in the serine palmitoyl transferase subunit 2 (SPTLC2) gene. The aim of this study was to establish whether oral L-serine lowers 1-deoxySL in a patient with HSAN1C, to perform a dose escalation to find the minimal effective dose, and to assess the safety profile and global metabolic effects of the treatment. Our patient underwent a 52-wk treatment in which the L-serine dose was titrated up to 400 mg/kg/day. She was followed up by repeated clinical examination, nerve conduction testing, and skin biopsies to document effects on small nerve fibers. Serum was assayed for 1-deoxySL and metabolomics analysis of 111 metabolites. We found a robust lowering of 1-deoxySL, which correlated in a near-linear fashion with increased serum L-serine levels. Metabolomics analysis showed a modest elevation in glycine and a marked reduction in the level of cytosine, whereas most of the other assayed metabolites did not change. There were no direct side effects from the treatment, but the patient developed a transitory toe ulceration during the course of the study. The Charcot–Marie–Tooth neuropathy score increased by 1 point. We conclude that oral supplementation of L-serine decreases 1-deoxySL in HSAN1C without major global effects on metabolism. L-serine is therefore a potential treatment for HSAN1C.

Mutations in the mitochondrially located protein CHCHD10 cause motoneuron disease by an unknown mechanism. In this study, we investigate the mutations p. R15L and p. G66V in comparison to wild-type CHCHD10 and the non-pathogenic variant p. P34S in vitro, in patient cells as well as in the vertebrate in vivo model zebrafish. We demonstrate a reduction of CHCHD10 protein levels in p. R15L and p. G66V mutant patient cells to approximately 50%. Quantitative real-time PCR revealed that expression of CHCHD10 p. R15L, but not of CHCHD10 p. G66V, is already abrogated at the mRNA level. Altered secondary structure and rapid protein degradation are observed with regard to the CHCHD10 p. G66V mutant. In contrast, no significant differences in expression, degradation rate or secondary structure of non-pathogenic CHCHD10 p. P34S are detected when compared with wild-type protein. Knockdown of CHCHD10 expression in zebrafish to about 50% causes motoneuron pathology, abnormal myofibrillar structure and motility deficits in vivo. Thus, our data show that the CHCHD10 mutations p. R15L and p. G66V cause motoneuron disease primarily based on haploinsufficiency of CHCHD10.

Motor neuron disorders (MNDs) are a heterogeneous group of diseases that result from degeneration of motor neurons. If both upper and lower motor neurons (UMNs and LMNs) are affected, the disease is classified as amyotrophic lateral sclerosis (ALS). Primary lateral sclerosis (PLS) and progressive muscular atrophy (PMA) selectively affect the UMNs or LMNs, respectively, but are sometimes considered to be incomplete ALS variants because their phenotype may evolve into typical ALS over time. Bulbar affection in a UMN disease would favour a diagnosis of PLS over hereditary spastic paraplegia (HSP), whereas rapid progression may separate PMA from adult-onset spinal muscular atrophy (SMA). Some SMA-like or even ALS-like phenotypes have been incorporated into the large category of sensorimotor axonal neuropathies (Charcot-Marie-Tooth disease type 2, CMT2),1 although sensory abnormalities may be subtle in some forms.2 On the other hand, spinal and bulbar muscular atrophy (Kennedy disease) is classified as a form of adult-onset SMA despite prominent sensory abnormalities. It has recently been argued that the current classification system of MNDs is unsatisfactory and should be revised to include genetically and prognostically important categories.3 We have recently identified a new form of motor neuron disease in 55 patients from 17 Finnish families, where distinctive phenotype did not match any pre-existing neuromuscular disease category.4–6 Spinal muscular atrophy Jokela type (SMAJ, OMIM #615048) is characterised by painful cramps, fasciculations, decreased or absent tendon reflexes, elevated creatine kinase and hand tremor. The first symptoms appear commonly after age 30–40. Electromyography (EMG) and muscle biopsy display widespread neurogenic findings and a proportion of patients show sensory abnormalities. Muscle weakness and atrophy appear much later in the disease course, and patients have remained ambulant for several decades and their life expectancy is within normal range.4 ,5 SMAJ is caused by a dominant …

Charcot–Marie–Tooth disease (CMT) is a group of hereditary peripheral neuropathies. The dominantly inherited axonal CMT2 displays striking genetic heterogeneity, with 17 presently known disease genes. The large number of candidate genes, combined with lack of genotype–phenotype correlations, has made genetic diagnosis in CMT2 time-consuming and costly. In Finland, 25% of dominant CMT2 is explained by either a GDAP1 founder mutation or private MFN2 mutations but the rest of the families have remained without molecular diagnosis. Whole-exome and genome sequencing are powerful techniques to find disease mutations for CMT patients but they require large amounts of sequencing to confidently exclude heterozygous variants in all candidate genes, and they generate a vast amount of irrelevant data for diagnostic needs. Here we tested a targeted next-generation sequencing approach to screen the CMT2 genes. In total, 15 unrelated patients from dominant CMT2 families from Finland, in whom MFN2 and GDAP1 mutations had been excluded, participated in the study. The targeted approach produced sufficient sequence coverage for 95% of the 309 targeted exons, the rest we excluded by Sanger sequencing. Unexpectedly, the screen revealed a disease mutation only in one family, in the HSPB1 gene. Thus, new disease genes underlie CMT2 in the remaining families, indicating further genetic heterogeneity. We conclude that targeted next-generation sequencing is an efficient tool for genetic screening in CMT2 that also aids in the selection of patients for genome-wide approaches.

Background Anoctaminopathies are muscle diseases caused by recessive mutations in the ANO5 gene. The effects of anoctaminopathy on oxidative capacity have not previously been studied in a controlled setting. Objective To characterize oxidative capacity in a clinically and genetically well-defined series of patients with anoctaminopathy. Methods We sequenced the ANO5 gene in 111 Finnish patients with suspected LGMD2. Patients with positive findings underwent close clinical examination, including electromyography, muscle MRI, and, in selected cases, muscle biopsy. Oxidative capacity was analyzed using spiroergometry and compared to age-matched healthy controls. Results We characterized 12 newly identified and 2 previously identified patients with ANO5 mutations from 11 families. Our material was genetically homogeneous with most patients homozygous for the Finnish founder variant c.2272C>T (p.Arg758Cys). In one family, we found a novel p.Met470Arg variant compound heterozygous with p.Arg758Cys. Lower limb muscle MRI revealed progressive fatty degeneration of specific posterior compartment muscles. Patients' spiroergometric profiles showed that anoctaminopathy significantly impaired oxidative capacity with increasing ventilation. Conclusions Our findings support earlier reports that anoctaminopathy progresses slowly and demonstrate that the disease impairs the capacity for aerobic exercise.

Mitochondrial myopathy (MM) with progressive external ophthalmoplegia (PEO) is a common manifestation of mitochondrial disease in adulthood, for which there is no curative therapy. In mice with MM, ketogenic diet significantly delayed progression of the disease. We asked in this pilot study what effects high‐fat, low‐carbohydrate “modified Atkins” diet (mAD) had for PEO/MM patients and control subjects and followed up the effects by clinical, morphological, transcriptomic, and metabolomic analyses. All of our five patients, irrespective of genotype, showed a subacute response after 1.5–2 weeks of diet, with progressive muscle pain and leakage of muscle enzymes, leading to premature discontinuation of the diet. Analysis of muscle ultrastructure revealed selective fiber damage, especially in the ragged‐red‐fibers (RRFs), a MM hallmark. Two years of follow‐up showed improvement of muscle strength, suggesting activation of muscle regeneration. Our results indicate that (i) nutrition can modify mitochondrial disease progression, (ii) dietary counseling should be part of MM care, (iii) short mAD is a tool to induce targeted RRF lysis, and (iv) mAD, a common weight‐loss method, may induce muscle damage in a population subgroup. ![][1] High‐fat, low‐carbohydrate modified Atkins diet (mAD) is a common weight‐loss method, found to ameliorate mitochondrial myopathy in mice. In human patients, mAD induces muscle damage, especially of ragged‐red fibers, the most affected by the disease. [1]: /embed/graphic-1.gif

Item does not contain fulltext Facioscapulohumeral dystrophy (FSHD) is associated with somatic chromatin relaxation of the D4Z4 repeat array and derepression of the D4Z4-encoded DUX4 retrogene coding for a germline transcription factor. Somatic DUX4 derepression is caused either by a 1-10 unit repeat-array contraction (FSHD1) or by mutations in SMCHD1, which encodes a chromatin repressor that binds to D4Z4 (FSHD2). Here, we show that heterozygous mutations in DNA methyltransferase 3B (DNMT3B) are a likely cause of D4Z4 derepression associated with low levels of DUX4 expression from the D4Z4 repeat and increased penetrance of FSHD. Recessive mutations in DNMT3B were previously shown to cause immunodeficiency, centromeric instability, and facial anomalies (ICF) syndrome. This study suggests that transcription of DUX4 in somatic cells is modified by variations in its epigenetic state and provides a basis for understanding the reduced penetrance of FSHD within families.

ObjectiveWe used patient-specific neuronal cultures to characterize the molecular genetic mechanism of recessive nonsense mutations in neurofilament light (NEFL) underlying early-onset Charcot-Marie-Tooth (CMT) disease.MethodsMotor neurons were differentiated from induced pluripotent stem cells of a patient with early-onset CMT carrying a novel homozygous nonsense mutation in NEFL. Quantitative PCR, protein analytics, immunocytochemistry, electron microscopy, and single-cell transcriptomics were used to investigate patient and control neurons.ResultsWe show that the recessive nonsense mutation causes a nearly total loss of NEFL messenger RNA (mRNA), leading to the complete absence of NEFL protein in patient's cultured neurons. Yet the cultured neurons were able to differentiate and form neuronal networks and neurofilaments. Single-neuron gene expression fingerprinting pinpointed NEFL as the most downregulated gene in the patient neurons and provided data of intermediate filament transcript abundancy and dynamics in cultured neurons. Blocking of nonsense-mediated decay partially rescued the loss of NEFL mRNA.ConclusionsThe strict neuronal specificity of neurofilament has hindered the mechanistic studies of recessive NEFL nonsense mutations. Here, we show that such mutation leads to the absence of NEFL, causing childhood-onset neuropathy through a loss-of-function mechanism. We propose that the neurofilament accumulation, a common feature of many neurodegenerative diseases, mimics the absence of NEFL seen in recessive CMT if aggregation prevents the proper localization of wild-type NEFL in neurons. Our results suggest that the removal of NEFL as a proposed treatment option is harmful in humans.

Hereditary sensory and autonomic neuropathy 1 (HSAN1) is an autosomal dominant disorder that can be caused by variants in SPTLC1 or SPTLC2, encoding subunits of serine palmitoyl-CoA transferase. Disease variants alter the enzyme's substrate specificity and lead to accumulation of neurotoxic 1-deoxysphingolipids. We describe two families with autosomal dominant HSAN1C caused by a new variant in SPTLC2, c.547C > T, p.(Arg183Trp). The variant changed a conserved amino acid and was not found in public variant databases. All patients had a relatively mild progressive distal sensory impairment, with onset after age 50. Small fibers were affected early, leading to abnormalities on quantitative sensory testing. Sural biopsy revealed a severe chronic axonal neuropathy with subtotal loss of myelinated axons, relatively preserved number of non-myelinated fibers and no signs for regeneration. Skin biopsy with PGP9.5 labeling showed lack of intraepidermal nerve endings early in the disease. Motor manifestations developed later in the disease course, but there was no evidence of autonomic involvement. Patients had elevated serum 1-deoxysphingolipids, and the variant protein produced elevated amounts of 1-deoxysphingolipids in vitro, which proved the pathogenicity of the variant. Our results expand the genetic spectrum of HSAN1C and provide further detail about the clinical characteristics. Sequencing of SPTLC2 should be considered in all patients presenting with mild late-onset sensory-predominant small or large fiber neuropathy.

Objectives Mutations in the CHCHD10 gene, which encodes a mitochondrially targeted protein, have emerged as an important cause of motor neuron disease and frontotemporal lobar degeneration. The aim of this study was to assess the clinical variability in a large family carrying the p.Gly66Val mutation of the CHCHD10 gene. This mutation has recently been reported to cause late-onset spinal muscular atrophy (SMAJ) or sensorimotor axonal Charcot–Marie–Tooth neuropathy (CMT2) in the Finnish population. Materials and methods Nine affected members of an extended Finnish pedigree were included in the study. Detailed clinical and neurophysiological examinations were performed. The CHCHD10 p.Gly66Val mutation was examined by Sanger sequencing. Results The heterozygous p.Gly66Val mutation was present in all affected individuals from whom a DNA sample was available. The clinical phenotype varied from proximal sensorimotor neuropathy to spinal muscular atrophy and in one case resembled motor neuron disease ALS at its early stages. The age of onset varied from 30 to 73 years. Conclusions Our data demonstrate that even within the same family, the p.Gly66Val variant can cause variable phenotypes ranging from CMT2-type axonal neuropathy to spinal muscular atrophy, which may also present as an ALS-like disease. The spectrum of CHCHD10-related neuromuscular disease has widened rapidly, and we recommend keeping the threshold for genetic testing low particularly when dominant inheritance or mitochondrial pathology is present.

Neuroligins are cell-adhesion molecules located at the postsynaptic side of the synapse. Neuroligins interact with beta-neurexins and this interaction is involved in the formation of functional synapses. Mutations in two X-linked neuroligin genes, NLGN3 and NLGN4, have recently been implicated in pathogenesis of autism. The neuroligin gene family consists of five members (NLGN1 at 3q26, NLGN2 at 17p13, NLGN3 at Xq13, NLGN4 at Xp22, and NLGN4Y at Yq11), of which NLGN1 and NLGN3 are located within the best loci observed in our previous genome-wide scan for autism in the Finnish sample. Here, we report a detailed molecular genetic analysis of NLGN1, NLGN3, NLGN4, and NLNG4Y in the Finnish autism sample. Mutation analysis of 30 probands selected from families producing linkage evidence for Xq13 and/or 3q26 loci revealed several polymorphisms, but none of these seemed to be functional. Family-based association analysis in 100 families with autism spectrum disorders yielded only modest associations at NLGN1 (rs1488545, P=0.002), NLGN3 (DXS7132, P=0.014), and NLGN4 (DXS996, P=0.031). We conclude that neuroligin mutations most probably represent rare causes of autism and that it is unlikely that the allelic variants in these genes would be major risk factors for autism.

Limb-girdle muscular dystrophy 2D (LGMD2D) is caused by mutations in the alpha-sarcoglycan gene (SGCA). The most frequently reported mutation, 229CGC>TGC (R77C) in exon 3 of SGCA, results in the substitution of arginine by cysteine. We present here the clinical, immunohistochemical, and genetic data of 11 Finnish patients with LGMD2D caused by mutations in SGCA. Mutational analysis showed 10 patients homozygous and 1 compound heterozygous for R77C. A wide spectrum of SGCA mutations has been reported previously. Our results show an enrichment of R77C in Finland, further underlined by the observed carrier frequency of 1 per 150. According to the annual birth rate of approximately 60,000 in Finland, one LGMD2D patient with a homozygous mutation is expected to be born every 1 or 2 years on average. The presence of an ancient founder mutation is indicated by the fact that all patients shared a short common haplotype extending > or = 790 kilobases. Our results emphasize the need to include the SGCA gene R77C mutation test in routine DNA analyses of severe dystrophinopathy-like muscular dystrophies in Finland, and suggest that the applicability of this test in other populations should be studied as well.

Nutrient availability is the major regulator of life and reproduction, and a complex cellular signaling network has evolved to adapt organisms to fasting. These sensor pathways monitor cellular energy metabolism, especially mitochondrial ATP production and NAD(+)/NADH ratio, as major signals for nutritional state. We hypothesized that these signals would be modified by mitochondrial respiratory chain disease, because of inefficient NADH utilization and ATP production. Oral administration of nicotinamide riboside (NR), a vitamin B3 and NAD(+) precursor, was previously shown to boost NAD(+) levels in mice and to induce mitochondrial biogenesis. Here, we treated mitochondrial myopathy mice with NR. This vitamin effectively delayed early- and late-stage disease progression, by robustly inducing mitochondrial biogenesis in skeletal muscle and brown adipose tissue, preventing mitochondrial ultrastructure abnormalities and mtDNA deletion formation. NR further stimulated mitochondrial unfolded protein response, suggesting its protective role in mitochondrial disease. These results indicate that NR and strategies boosting NAD(+) levels are a promising treatment strategy for mitochondrial myopathy.

Background: Tibial muscular dystrophy (TMD), a late-onset dominant distal myopathy, is caused by yet unknown mutations on chromosome 2q, whereas MD with myositis (MDM) is a muscular dystrophy of the mouse, also progressing with age and linked to mouse chromosome 2. For both disorders, linkage studies have implicated titin as a potential candidate gene.Methods: The authors analyzed major candidate regions in the titin gene by sequencing and Southern blot hybridization, and performed titin immunohistochemistry on TMD patient material to identify the underlying mutation. Western blot studies were performed on the known titin ligands in muscle samples of both disorders and controls, and analysis of apoptosis was also performed.Results: The authors identified almost complete loss of calpain3, a ligand of titin, in the patient with limb-girdle MD (LGMD) with a homozygous state of TMD haplotype when primary calpain3 gene defect was excluded. Apoptotic myonuclei with altered distribution of transcription factor NF-kB and its inhibitor IkBα were encountered in muscle samples of patients with either heterozygous or homozygous TMD haplotype. Similar findings were confirmed in the MDM mouse.Conclusions: These results imply that titin mutations may be responsible for TMD, and that the pathophysiologic pathway following calpain3 deficiency may overlap with LGMD2A. The loss of calpain3 could be a downstream effect of the deficient TMD gene product. The significance of the secondary calpain3 defect for the pathogenesis of TMD was emphasized by similar calpain3 deficiency in the MDM mouse, which is suggested to be a mouse model for TMD. Homozygous mutation at the 2q locus may thus be capable of producing yet another LGMD.

Muscle-eye-brain disease belongs to congenital muscular dystrophies with central nervous system abnormalities. The etiology of MEB is still unknown, but abnormal immunoreactivity for laminin-2 has been reported. To evaluate disease progression in muscle tissue, 32 biopsy specimens from 17 muscle-eye-brain patients were analysed. The samples of four patients were studied by immunohistochemical techniques and by quantitative Western blotting. The samples showed a great variation in the muscle pathology. Regenerative fibers and mild fiber size variation were present in over 60%. At infancy, necrotic and regenerative fibers were common, while fat infiltration was the most prominent finding in the age group over five years. In quantitative studies, the amount of laminin alpha 2 chain was clearly reduced to 10-20% of normal. In contrast, laminin beta 2 chain was overexpressed in the Western blotting studies. These findings may reflect a yet unidentified primary disturbance in the basement membrane composition and function.

Objective Several genome-wide screens have been performed in autism spectrum disorders resulting in the identification of numerous putative susceptibility loci. Analyses of pooled primary data should result in an increased sample size and the different study samples have a potential to strengthen the evidence for some earlier identified loci, reveal novel loci, and even to provide information of the general significance of the locus. The objective of this study was to search for potential susceptibility loci for autism, which are supported by two independent samples. Methods We performed a combined analysis of the primary genome scan data of the Autism Genetic Resource Exchange (AGRE) and Finnish autism samples to reveal susceptibility loci potentially shared by these study samples. Results In the initial combined data analysis, the best loci (p < 0.05) were observed at 1p12-q25, 3p24-26, 4q21-31, 5p15-q12, 6q14-21, 7q33-36, 8q22-24, 17p12-q21, and 19p13-q13. The combined analysis of Finnish and AGRE families showed the most promising shared locus on 3p24-26 with nonparametric logarithm of odds (NPL) score of 2.20 (p = 0.011). The combined data analysis did not provide increased linkage evidence for the earlier identified loci on 3q25-27 or 17p12-q21. However, the 17p12-q21 locus remained promising also in the combined sample (NPLall =2.38, p = 0.0076). Interpretation Our study of 314 autism families highlights the importance of further analyses on 3p24-26 locus involving comprehensive molecular genetic analyses of oxytocin receptor gene (OXTR), a positional and functional candidate gene for autism. Ann Neurol 2005

Recent molecular studies on autism and related disorders have supported a multilocus etiology for the disease spectrum. To maximize genetic and cultural homogeneity, we have focused our molecular studies to families originating from a subisolate of Central Finland. Genealogical studies enabled the identification of a megapedigree comprising of 12 core families with autism and Asperger syndrome (AS). We analyzed two chromosomal regions on Iq and 3q showing highest lod scores in our genome-wide scan, as well as the AUTS1 locus on chromosome 7q. For markers on 3q25-27, more significant association was observed in families from subisolate compared to families from the rest of Finland. In contrast, no clear evidence for association on AUTS1 locus was obtained. The wide interval showing association, in particular, on chromosome 3q suggests a locus for autism spectrum of disorders on this chromosomal region.

DNAJB6 was recently identified as a causative gene for limb-girdle muscular dystrophy type 1D (LGMD1D). DNAJB6 belongs to a class of co-chaperones characterized by a J-domain in the N-terminus. To date, six different mutations have been identified in several families from European, North American and Asian countries. The known mutations cause mainly adult onset, slowly progressive proximal muscle weakness, although occasional patients with earlier onset have been reported. The evolution of the disease has been very mild to moderate and respiratory involvement has not been reported. A Finnish family with four affected members, three siblings and their mother, has been studied extensively in the past decade. All patients presented with marked proximal lower limb weakness at the age of 10 to13 years and many reported some weakness from early childhood. Weakness progressed to proximal upper limbs and to some extent also to distal lower limb muscles causing walking difficulties in early adulthood. The mother became wheelchair bound at age 25. Respiratory involvement was a major problem in the family; two sisters had dyspnea in their late teens and their mother died of respiratory failure at age 33. A novel mutation in DNAJB6 was identified in the family as the cause of this very severe, early onset form of LGMD1D. Functional studies of the mutation indicate a more pronounced reduction of the antiaggregation capacity compared to previously known mutations.

The role of genetic factors in the etiology of the autistic spectrum of disorders has clearly been demonstrated. Ten chromosomal regions, on chromosomes 1p, 4p, 6q, 7q, 13q, 15q, 16p, 17q, 19q and 22q have potentially been linked to autism.1–8 We have analyzed these chromosomal regions in a total of 17 multiplex families with autism originating from the isolated Finnish population by pairwise linkage analysis and sib-pair analysis. Mild evidence for putative contribution was found only with the 1p chromosomal region in the susceptibility to autism. Our data suggest that additional gene loci exist for autism which will be detectable in and even restricted to the isolated Finnish population.

To identify genetic loci for autism-spectrum disorders, we have performed a two-stage genomewide scan in 38 Finnish families. The detailed clinical examination of all family members revealed infantile autism, but also Asperger syndrome (AS) and developmental dysphasia, in the same set of families. The most significant evidence for linkage was found on chromosome 3q25-27, with a maximum two-point LOD score of 4.31 ( Z max dom ) for D3S3037, using infantile autism and AS as an affection status. Six markers flanking over a 5-cM region on 3q gave Z max dom >3, and a maximum parametric multipoint LOD score (MLS) of 4.81 was obtained in the vicinity of D3S3715 and D3S3037. Association, linkage disequilibrium, and haplotype analyses provided some evidence for shared ancestor alleles on this chromosomal region among affected individuals, especially in the regional subisolate. Additional potential susceptibility loci with two-point LOD scores >2 were observed on chromosomes 1q21-22 and 7q. The region on 1q21-22 overlaps with the previously reported candidate region for infantile autism and schizophrenia, whereas the region on chromosome 7q provided evidence for linkage 58 cM distally from the previously described autism susceptibility locus (AUTS1).