Three novel MTM1 pathogenic variants identified in Japanese patients with X-linked myotubular myopathy.
ABSTRACT: BACKGROUND:X-linked myotubular myopathy (XLMTM) is a form of the severest congenital muscle diseases characterized by marked muscle weakness, hypotonia, and feeding and breathing difficulties in male infants. It is caused by mutations in the myotubularin gene (MTM1). METHODS:Evaluation of clinical history and examination of muscle pathology of three patients and comprehensive genome analysis on our original targeted gene panel system for muscular diseases. RESULTS:We report three patients, each of whom presents distinct muscle pathological features. The three patients have novel hemizygous MTM1 variants, including c.527A>G (p.Gln176Arg), c.595C>G (p.Pro199Ala), or c.688T>C (p.Trp230Arg). CONCLUSIONS:All variants were assessed as "Class 4 (likely pathogenic)" on the basis of the guideline of American College of Medical Genetics and Genomics. These distinct pathological features among the patients with variants in the second cluster of PTP domain in MTM1 provides an insight into microheterogeneities in disease phenotypes in XLMTM.
Project description:X-linked recessive myotubular myopathy (XLMTM) is a severe congenital muscle disorder caused by mutations in the MTM1 gene and characterized by severe hypotonia and generalized muscle weakness in affected males. It is generally a fatal disorder during the neonatal period and early infancy. The diagnosis is based on typical histopathological findings on muscle biopsy, combined with suggestive clinical features. We experienced a case of a newborn who required intubation and ventilator care because of profound hypotonia and respiratory difficulty. The preliminary diagnosis at the time of request for retrieval was hypoxic ischemic encephalopathy, but the infant was clinically reevaluated for generalized weakness and muscle atrophy. Muscle biopsies showed variability in fiber size and centrally located nuclei in nearly all the fibers. We detected an MTM1 gene mutation of c.1261-1C>A in the intron 10 region, and diagnosed the neonate with myotubular myopathy. The same mutation was detected in his mother.
Project description:Mutations in the MTM1 gene encoding myotubularin cause X-linked myotubular myopathy (XLMTM), a well-defined subtype of human centronuclear myopathy. Seven male Labrador Retrievers, age 14-26 wk, were clinically evaluated for generalized weakness and muscle atrophy. Muscle biopsies showed variability in fiber size, centrally placed nuclei resembling fetal myotubes, and subsarcolemmal ringed and central dense areas highlighted with mitochondrial specific reactions. Ultrastructural studies confirmed the centrally located nuclei, abnormal perinuclear structure, and mitochondrial accumulations. Wild-type triads were infrequent, with most exhibiting an abnormal orientation of T tubules. MTM1 gene sequencing revealed a unique exon 7 variant in all seven affected males, causing a nonconservative missense change, p.N155K, which haplotype data suggest derives from a recent founder in the local population. Analysis of a worldwide panel of 237 unaffected Labrador Retrievers and 59 additional control dogs from 25 other breeds failed to identify this variant, supporting it as the pathogenic mutation. Myotubularin protein levels and localization were abnormal in muscles from affected dogs, and expression of GFP-MTM1 p.N155K in COS-1 cells showed that the mutant protein was sequestered in proteasomes, where it was presumably misfolded and prematurely degraded. These data demonstrate that XLMTM in Labrador Retrievers is a faithful genetic model of the human condition.
Project description:X-linked myotubular myopathy (XLMTM) is a severe congenital disorder in male infants that leads to generalized skeletal muscle weakness and is frequently associated with fatal respiratory failure. XLMTM is caused by loss-of-function mutations in the MTM1 gene, which encodes myotubularin, the founder member of a family of 15 homologous proteins in mammals. We recently demonstrated the therapeutic efficacy of intravenous delivery of rAAV vectors expressing MTM1 in animal models of myotubular myopathy. Here, we tested whether the closest homologues of MTM1, MTMR1, and MTMR2 (the latter being implicated in Charcot-Marie-Tooth neuropathy type 4B1) are functionally redundant and could represent a therapeutic target for XLMTM. Serotype 9 recombinant AAV vectors encoding either MTM1, MTMR1, or MTMR2 were injected into the tibialis anterior muscle of Mtm1-deficient knockout mice. Two weeks after vector delivery, a therapeutic effect was observed with Mtm1 and Mtmr2, but not Mtmr1; with Mtm1 being the most efficacious transgene. Furthermore, intravenous administration of a single dose of the rAAV9-Mtmr2 vector in XLMTM mice improved the motor activity and muscle strength and prolonged survival throughout a 3-month study. These results indicate that strategies aiming at increasing MTMR2 expression levels in skeletal muscle may be beneficial in the treatment of myotubular myopathy.
Project description:INTRODUCTION:X-linked myotubular myopathy (XLMTM), a devastating pediatric disease caused by the absence of the protein myotubularin, results from mutations in the MTM1 gene. While there is no cure for XLMTM, we previously reported effects of MTM1 gene therapy using adeno-associated virus (AAV) vector on muscle weakness and pathology in MTM1-mutant dogs. Here, we followed 2 AAV-infused dogs over 4 years. METHODS:We evaluated gait, strength, respiration, neurological function, muscle pathology, AAV vector copy number (VCN), and transgene expression. RESULTS:Four years following AAV-mediated gene therapy, gait, respiratory performance, neurological function and pathology in AAV-infused XLMTM dogs remained comparable to their healthy littermate controls despite a decline in VCN and muscle strength. CONCLUSIONS:AAV-mediated gene transfer of MTM1 in young XLMTM dogs results in long-term expression of myotubularin transgene with normal muscular performance and neurological function in the absence of muscle pathology. These findings support a clinical trial in patients. Muscle Nerve 56: 943-953, 2017.
Project description:Loss-of-function mutations in the myotubularin gene (MTM1) cause X-linked myotubular myopathy (XLMTM), a fatal, congenital pediatric disease that affects the entire skeletal musculature. Systemic administration of a single dose of a recombinant serotype 8 adeno-associated virus (AAV8) vector expressing murine myotubularin to Mtm1-deficient knockout mice at the onset or at late stages of the disease resulted in robust improvement in motor activity and contractile force, corrected muscle pathology, and prolonged survival throughout a 6-month study. Similarly, single-dose intravascular delivery of a canine AAV8-MTM1 vector in XLMTM dogs markedly improved severe muscle weakness and respiratory impairment, and prolonged life span to more than 1 year in the absence of toxicity or a humoral or cell-mediated immune response. These results demonstrate the therapeutic efficacy of AAV-mediated gene therapy for myotubular myopathy in small- and large-animal models, and provide proof of concept for future clinical trials in XLMTM patients.
Project description:Myotubular myopathy is a subtype of centronuclear myopathy with X-linked inheritance and distinctive clinical and pathologic features. Most boys with myotubular myopathy have MTM1 mutations. In remaining individuals, it is not clear if disease is due to an undetected alteration in MTM1 or mutation of another gene. We describe a boy with myotubular myopathy but without mutation in MTM1 by conventional sequencing. Array-CGH analysis of MTM1 uncovered a large MTM1 duplication. This finding suggests that at least some unresolved cases of myotubular myopathy are due to duplications in MTM1, and that array-CGH should be considered when MTM1 sequencing is unrevealing.
Project description:No effective treatment exists for patients with X-linked myotubular myopathy (XLMTM), a fatal congenital muscle disease caused by deficiency of the lipid phosphatase, myotubularin. The Mtm1?4 and Mtm1 p.R69C mice model severely and moderately symptomatic XLMTM, respectively, due to differences in the degree of myotubularin deficiency. Contractile function of intact extensor digitorum longus (EDL) and soleus muscles from Mtm1?4 mice, which produce no myotubularin, is markedly impaired. Contractile forces generated by chemically skinned single fiber preparations from Mtm1?4 muscle were largely preserved, indicating that weakness was largely due to impaired excitation contraction coupling. Mtm1 p.R69C mice, which produce small amounts of myotubularin, showed impaired contractile function only in EDL muscles. Short-term replacement of myotubularin with a prototypical targeted protein replacement agent (3E10Fv-MTM1) in Mtm1?4 mice improved contractile function and muscle pathology. These promising findings suggest that even low levels of myotubularin protein replacement can improve the muscle weakness and reverse the pathology that characterizes XLMTM.
Project description:Mutations in the MTM1 gene cause X-linked myotubular myopathy (XLMTM), characterized by neonatal hypotonia and respiratory failure, and are responsible for a premature mortality in affected males. Female carriers are usually asymptomatic but they may present with muscular weakness because of a hypothesized skewed pattern of X-chromosome inactivation. By combining next generation sequencing (NGS) and CGH array approaches, we have investigated the role of MTM1 variants in a large cohort of undiagnosed patients with a wide spectrum of myopathies. Seven novel XLMTM patients have been identified, including two girls with an unremarkable family history for myotubular myopathy. Moreover, we have detected and finely mapped a large deletion causing a myotubular myopathy with abnormal genital development. Our data confirm that the severe neonatal onset of the disease in male infants is sufficient to address the direct molecular testing toward the MTM1 gene and, above all, suggest that the number of undiagnosed symptomatic female carriers is probably underestimated.
Project description:We aimed to correlate pathologic findings with MTM1 mutation type in a series of molecularly defined XLMTM cases. Clinical data from 15 XLMTM patients and their corresponding 16 muscle biopsies were studied. All patients were infants (range: 6-217 days old) when initially biopsied. The proportion of myofibers with central nuclei did not correlate with clinical outcome, however, morphometric studies showed that survivors had larger myofiber diameters in infancy than those who died (10.4+/-3.9microm versus 8.9+/-3microm; p<0.001). As a corollary, patients with MTM1 missense mutations had larger myofiber diameters (11.1+/-4microm), than those with truncation/deletion mutations (8.6+/-2.7microm) (controls 11.7+/-2.5microm) (p<0.0001). These data indicate that differences in myofiber size correlate with MTM1 mutation type and patient outcome. Failure to attain and/or maintain myofiber size, along with fiber type perturbations and the misplacement of myofiber nuclei and other organelles, are important components of XLMTM muscle pathology.
Project description:Congenital and inherited myopathies in dogs are faithful models of human muscle diseases and are being recognized with increasing frequency. In fact, canine models of dystrophin deficient muscular dystrophy and X-linked myotubular myopathy are of tremendous value in the translation of new and promising therapies for the treatment of these diseases. We have recently identified a family of Australian Rottweilers in which male puppies were clinically affected with severe muscle weakness and atrophy that resulted in early euthanasia or death. X-linked myotubular myopathy was suspected based on the early and severe clinical presentation and histopathological changes within muscle biopsies. The aim of this study was to determine the genetic basis for myopathy in these dogs and compare and contrast the clinical presentation, histopathology, ultrastructure, and mutation in this family of Rottweiler dogs with the previously described myotubular myopathy in Labrador retrievers.Histopathology, histochemistry, and ultrastructural examination of muscle biopsies from affected Rottweiler puppies were consistent with an X-linked myotubular myopathy. An unusual finding that differed from the previously reported Labradors and similar human cases was the presence of excessive autophagy and prominent autophagic vacuoles. Molecular investigations confirmed a missense mutation in exon 11 of MTM1 that was predicted to result in a non-functional phosphatase activity. Although the clinical presentations and histopathology were similar, the MTM1 p.(Q384P) mutation is different from the p.(N155K) mutation in exon 7 affecting Labrador retrievers with X-linked myotubular myopathy.Here we describe a second pathogenic mutation in MTM1 causing X-linked myotubular myopathy in dogs. Our findings suggest a variety of MTM1 mutations in dogs as seen in human patients. The number of MTM1 mutations resulting in similar severe and progressive clinical myopathy and histopathological changes are likely to increase as canine myopathies are further characterized.