Dissecting Pathogenetic Mechanisms and Therapeutic Strategies in Drosophila Models of Myotonic Dystrophy Type 1.
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ABSTRACT:
SUBMITTER: Souidi A
PROVIDER: S-EPMC6321436 | biostudies-literature | 2018 Dec
REPOSITORIES: biostudies-literature
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Project description:Myotonic dystrophy (DM) is a dominantly inherited neurodegenerative disorder for which there is no cure or effective treatment. Investigation of DM pathogenesis has identified a novel disease mechanism that requires development of innovative therapeutic strategies. It is now clear that DM is not caused by expression of a mutant protein. Instead, DM is the first recognized example of an RNA-mediated disease. Expression of the mutated gene gives rise to an expanded repeat RNA that is directly toxic to cells. The mutant RNA is retained in the nucleus, forming ribonuclear inclusions in affected tissue. A primary consequence of RNA toxicity in DM is dysfunction of two classes of RNA binding proteins, which leads to abnormal regulation of alternative splicing, or spliceopathy, of select genes. Spliceopathy now is known to cause myotonia and insulin resistance in DM. As our understanding of pathogenesis continues to improve, therapy targeted directly at the RNA disease mechanism will begin to replace the supportive care currently available. New pharmacologic approaches to treat myotonia and muscle wasting in DM type 1 are already in early clinical trials, and therapies designed to reverse the RNA toxicity have shown promise in preclinical models by correcting spliceopathy and eliminating myotonia. The well-defined ribonuclear inclusions may serve as convenient therapeutic targets to identify new agents that modify RNA toxicity. Continued development of appropriate model systems will allow testing of additional therapeutic strategies as they become available. Although DM is a decidedly complex disorder, its RNA-mediated disease mechanism may prove to be highly susceptible to therapy.
| S-EPMC4514697 | biostudies-other
Project description:Myotonic dystrophy type 1 (DM1) is the most common muscular dystrophy that affects multiple systems including the muscle and heart. The mutant CTG expansion at the 3'-UTR of the DMPK gene causes the expression of toxic RNA that aggregate as nuclear foci. The foci then interfere with RNA-binding proteins, affecting hundreds of mis-spliced effector genes, leading to aberrant alternative splicing and loss of effector gene product functions, ultimately resulting in systemic disorders. In recent years, increasing clinical, imaging, and pathological evidence have indicated that DM1, though to a lesser extent, could also be recognized as true brain diseases, with more and more researchers dedicating to develop novel therapeutic tools dealing with it. In this review, we summarize the current advances in the pathogenesis and pathology of central nervous system (CNS) deficits in DM1, intervention measures currently being investigated are also highlighted, aiming to promote novel and cutting-edge therapeutic investigations.
| S-EPMC8634727 | biostudies-literature
Project description:Myotonic dystrophy 2 (DM2) is a genetic multi-systemic disease primarily affecting skeletal muscle. It is caused by CCTGn expansion in intron 1 of the CNBP gene, which encodes a zinc finger protein. DM2 disease has been successfully modeled in Drosophila melanogaster, allowing the identification and validation of new pathogenic mechanisms and potential therapeutic strategies. Here, we describe the principal tools used in Drosophila to study and dissect molecular pathways related to muscular dystrophies and summarize the main findings in DM2 pathogenesis based on DM2 Drosophila models. We also illustrate how Drosophila may be successfully used to generate a tractable animal model to identify novel genes able to affect and/or modify the pathogenic pathway and to discover new potential drugs.
| S-EPMC10532015 | biostudies-literature
Project description:Myotonic dystrophy type 1 (DM1) is caused by a CTG nucleotide repeat expansion within the 3' UTR of the Dystrophia Myotonica protein kinase gene. In this study, we explored therapeutic genome editing using CRISPR/Cas9 via targeted deletion of expanded CTG repeats and targeted insertion of polyadenylation signals in the 3' UTR upstream of the CTG repeats to eliminate toxic RNA CUG repeats. We found paired SpCas9 or SaCas9 guide RNA induced deletion of expanded CTG repeats. However, this approach incurred frequent inversion in both the mutant and normal alleles. In contrast, the insertion of polyadenylation signals in the 3' UTR upstream of the CTG repeats eliminated toxic RNA CUG repeats, which led to phenotype reversal in differentiated neural stem cells, forebrain neurons, cardiomyocytes, and skeletal muscle myofibers. We concluded that targeted insertion of polyadenylation signals in the 3' UTR is a viable approach to develop therapeutic genome editing for DM1.
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Project description:Myotonic dystrophy type 1 (DM1) is an autosomal dominant, CTG•CAG microsatellite expansion disease. Expanded CUG repeat RNA sequester the muscleblind-like (MBNL) family of RNA-binding proteins, thereby disrupting their normal cellular function which leads to global mis-regulation of RNA processing. Previously, the small molecule furamidine was shown to reduce CUG foci and rescue mis-splicing in a DM1 HeLa cell model and to rescue mis-splicing in the HSALR DM1 mouse model, but furamidine's mechanism of action was not explored. Here we use a combination of biochemical, cell toxicity, and genomic studies in DM1 patient-derived myotubes and the HSALR DM1 mouse model to investigate furamidine's mechanism of action. Mis-splicing rescue was observed in DM1 myotubes and the HSALR DM1 mouse with furamidine treatment. Interestingly, while furamidine was found to bind CTG•CAG repeat DNA with nanomolar affinity, a reduction in expanded CUG repeat transcript levels was observed in the HSALR DM1 mouse but not DM1 patient-derived myotubes. Further investigation in these cells revealed that furamidine treatment at nanomolar concentrations led to up-regulation of MBNL1 and MBNL2 protein levels and a reduction of ribonuclear foci. Additionally, furamidine was shown to bind CUG RNA with nanomolar affinity and disrupted the MBNL1 -CUG RNA complex in vitro at micromolar concentrations. Furamidine's likely promiscuous interactions in vitro and in vivo appear to affect multiple pathways in the DM1 mechanism to rescue mis-splicing, yet surprisingly furamidine was shown globally to rescue many mis-splicing events with only modest off-target effects on gene expression in the HSALR DM1 mouse model. Importantly, over 20% of the differentially expressed genes were shown to be returned, to varying degrees, to wild-type expression levels.
| S-EPMC6343479 | biostudies-literature
Project description:Myotonic dystrophy type 1 (DM1) is a disease characterized by errors in alternative splicing, or "mis-splicing". The causative agent of mis-splicing in DM1 is an inherited CTG repeat expansion located in the 3' untranslated region of the DM protein kinase gene. When transcribed, CUG repeat expansion RNA sequesters muscleblind-like (MBNL) proteins, which constitute an important family of alternative splicing regulators. Sequestration of MBNL proteins results in the mis-splicing of its regulated transcripts. Previous work has demonstrated that pentamidine, a diamidine which is currently FDA-approved as an antiparasitic agent, was able to partially reverse mis-splicing in multiple DM1 models, albeit at toxic concentrations. In this study, we characterized a series of pentamidine analogues to determine their ability to reverse mis-splicing and their toxicity in vivo. Experiments in cell and mouse models demonstrated that compound 13, also known as furamidine, effectively reversed mis-splicing with equal efficacy and reduced toxicity compared to pentamidine.
| S-EPMC4972181 | biostudies-literature
Project description:Myotonic dystrophy type I (DM1) is a disabling neuromuscular disease with no causal treatment available. This disease is caused by expanded CTG trinucleotide repeats in the 3' UTR of the dystrophia myotonica protein kinase gene. On the RNA level, expanded (CUG)n repeats form hairpin structures that sequester splicing factors such as muscleblind-like 1 (MBNL1). Lack of available MBNL1 leads to misregulated alternative splicing of many target pre-mRNAs, leading to the multisystemic symptoms in DM1. Many studies aiming to identify small molecules that target the (CUG)n-MBNL1 complex focused on synthetic molecules. In an effort to identify new small molecules that liberate sequestered MBNL1 from (CUG)n RNA, we focused specifically on small molecules of natural origin. Natural products remain an important source for drugs and play a significant role in providing novel leads and pharmacophores for medicinal chemistry. In a new DM1 mechanism-based biochemical assay, we screened a collection of isolated natural compounds and a library of over 2100 extracts from plants and fungal strains. HPLC-based activity profiling in combination with spectroscopic methods were used to identify the active principles in the extracts. The bioactivity of the identified compounds was investigated in a human cell model and in a mouse model of DM1. We identified several alkaloids, including the β-carboline harmine and the isoquinoline berberine, that ameliorated certain aspects of the DM1 pathology in these models. Alkaloids as a compound class may have potential for drug discovery in other RNA-mediated diseases.
| S-EPMC5016119 | biostudies-other
Project description:Myotonic dystrophy involves two types of chronically debilitating rare neuromuscular diseases: type 1 (DM1) and type 2 (DM2). Both share similarities in molecular cause, clinical signs, and symptoms with DM2 patients usually displaying milder phenotypes. It is well documented that key clinical symptoms in DM are associated with a strong mis-regulation of RNA metabolism observed in patient's cells. This mis-regulation is triggered by two leading DM-linked events: the sequestration of Muscleblind-like proteins (MBNL) and the mis-regulation of the CUGBP RNA-Binding Protein Elav-Like Family Member 1 (CELF1) that cause significant alterations to their important functions in RNA processing. It has been suggested that DM1 may be treatable through endogenous modulation of the expression of MBNL and CELF1 proteins. In this study, we analyzed the recent identification of the involvement of microRNA (miRNA) molecules in DM and focus on the modulation of these miRNAs to therapeutically restore normal MBNL or CELF1 function. We also discuss additional prospective miRNA targets, the use of miRNAs as disease biomarkers, and additional promising miRNA-based and miRNA-targeting drug development strategies. This review provides a unifying overview of the dispersed data on miRNA available in the context of DM.
| S-EPMC6888406 | biostudies-literature
Project description:After respiratory distress, cardiac dysfunction is the second most common cause of fatality associated with the myotonic dystrophy (DM) disease. Despite the prevalance of heart failure in DM, physiopathological studies on heart symptoms have been relatively scarce because few murine models faithfully reproduce the cardiac disease. Consequently, only a small number of candidate compounds have been evaluated in this specific phenotype. To help cover this gap Drosophila combines the amenability of its invertebrate genetics with the possibility of quickly acquiring physiological parameters suitable for meaningful comparisons with vertebrate animal models and humans. Here we review available descriptions of cardiac disease in DM type 1 and type 2, and three recent papers reporting the cardiac toxicity of non-coding CUG (DM1) and CCUG (DM2) repeat RNA in flies. Notably, flies expressing CUG or CCUG RNA in their hearts developed strong arrhythmias and had reduced fractional shortening, which correlates with similar phenotypes in DM patients. Overexpression of Muscleblind, which is abnormally sequestered by CUG and CCUG repeat RNA, managed to strongly suppress arrhythmias and fractional shortening, thus demonstrating that Muscleblind depletion causes cardiac phenotypes in flies. Importantly, small molecules pentamidine and daunorubicin were able to rescue cardiac phenotypes by releasing Muscleblind from sequestration. Taken together, fly heart models have the potential to make important contributions to the understanding of the molecular causes of cardiac dysfunction in DM and in the quick assessment of candidate therapeutics.
| S-EPMC6054993 | biostudies-literature
Project description:BackgroundMyotonic dystrophy is a multisystem disorder characterized by widespread organic involvement including central nervous system symptoms. Although myotonic dystrophy disease types 1 (DM1) and 2 (DM2) cover a similar spectrum of symptoms, more pronounced clinical and brain alterations have been described in DM1. Here, we investigated brain volumetric and white matter alterations in both disease types and compared to healthy controls (HC).MethodsMRI scans were obtained from 29 DM1, 27 DM2, and 56 HC. We assessed macro- and microstructural brain changes by surface-based analysis of cortical thickness of anatomical images and tract-based spatial statistics of fractional anisotropy (FA) obtained by diffusion-weighted imaging, respectively. Global MRI measures were related to clinical and neuropsychological scores to evaluate their clinical relevance.ResultsCortical thickness was reduced in both patient groups compared to HC, showing similar patterns of regional distribution in DM1 and DM2 (occipital, temporal, frontal) but more pronounced cortical thinning for DM1. Similarly, FA values showed a widespread decrease in DM1 and DM2 compared to HC. Interestingly, FA was significantly lower in DM1 compared to DM2 within most parts of the brain.ConclusionComparisons between DM1 and DM2 indicate a more pronounced cortical thinning of grey matter and a widespread reduction in microstructural integrity of white matter in DM1. Future studies are required to unravel the underlying and separating mechanisms for the disease courses of the two types and their neuropsychological symptoms.
| S-EPMC11319366 | biostudies-literature