Search results for: muscle-gene-therapy

Muscle Gene Therapy

Author : Dongsheng Duan
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Muscle disease represents an important health threat to the general population. There is essentially no cure. Gene therapy holds great promise to correct the genetic defects and eventually achieve full recovery in these diseases. Significant progresses have been made in the field of muscle gene therapy over the last few years. The development of novel gene delivery vectors has substantially enhanced specificity and efficiency of muscle gene delivery. The new knowledge on the immune response to viral vectors has added new insight in overcoming the immune obstacles. Most importantly, the field has finally moved from small experimental animal models to human patients. This book will bring together the leaders in the field of muscle gene transfer to provide an updated overview on the progress of muscle gene therapy. It will also highlight important clinical applications of muscle gene therapy.

Muscle Gene Therapy

Author : Dongsheng Duan
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Gene therapy offers many conceptual advantages to treat muscle diseases, especially various forms of muscular dystrophies; however, it faces a number of unique challenges, including the need to deliver a therapeutic vector to all muscles throughout the body. In Muscle Gene Therapy: Methods and Protocols, expert researchers in the field present a collection of techniques aimed at bridging the translational gap in muscle gene therapy between the prevalent rodent models and vitally important larger animal models. Divided into three sections, this volume examines basic protocols for optimizing the muscle gene expression cassette and for evaluating the therapeutic outcomes, new developments in muscle gene therapy technology such as adeno-associated viral vector (AAV), oligonucleotide-mediated exon-skipping, and novel RNA-based strategies, and step-by-step guidance on muscle gene delivery in swine, ovine, canine, and non-human primates. Written in the highly successful Methods in Molecular BiologyTM series format, chapters include introductions to their respective topics, lists of the necessary materials and reagents, detailed, readily reproducible laboratory protocols, and tips on troubleshooting and avoiding known pitfalls. Authoritative and cutting-edge, Muscle Gene Therapy: Methods and Protocols serves as an invaluable resource for graduate students, post-doctoral fellows, and principle investigators pursuing the crucial advancement of muscle disease gene therapy in the hope of someday curing these debilitating disorders.

Studies on Retroviral mediated Gene Transfer Into Skeletal Muscle for Gene Therapy of Duchenne Muscular Dystrophy

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Improved Plasmid based Muscle Gene Therapy

Author : Yasamine Modarresi
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Muscular dystrophies are a group of inherited disorders characterized by progressive muscle weakness, and degeneration. One approach to treatment would be the replacement of the deficient protein via gene therapy. For effective gene therapy, both efficiency of gene delivery and stable expression of the transferred gene are important factors. Our goal was to determine which of the following mammalian expression vectors would be more useful (more stable) for muscle gene therapy; pAcGFP1-C1 and pEPito. We were also interested in switching/substituting both vectors' CMV promoter/enhancer region with the muscle specific promoter, Desmin (DES), to increase their stability for muscle gene therapy. This was accomplished by transfecting C2C12 myotubes with the aforementioned vectors. Both vectors showed relatively continuous GFP expression. Myotubes transfected with pEPito continued to express GFP till day 8. Cells transfected with pACGFP1-C1 also showed continuous GFP expression till day 6. Our results show that both vectors are promising candidates for gene therapy in muscle cells as they maintained stable gene expression of the GFP reporter gene for at least 6 days. Further studies should be done in order to determine the maximum duration that the myotubes would be able to maintain the plasmids and show continuous expression. Future studies could be done to assess stability of these vectors in mice which may lead to future gene therapy trials for muscle disorders and improving gene therapy strategies for other disorders.

Duchenne Muscular Dystrophy

Author : Jeffrey S. Chamberlain
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Duchenne Muscular Dystrophy (DMD) is one of the most prevalent genetic disorders of childhood and currently stands as an incurable condition. This authoritative guide provides a clear overview of the latest current and experimental approaches to the treatment of DMD and examines the clinical, genetic, and pathophysiological aspects of the disease in the context of emerging therapeutic modalities. The only available source on the subject, this reference emphasizes the importance of accurate diagnosis, carrier detection, and genetic counseling, and supplies state-of-the-art contributions on pharmacological interventions, regenerative medicine, and gene therapy.

Studies on Retroviral mediated Gene Transfer in Skeletal Muscle for Gene Therapy of Duchenne Muscular Dystrophy

Author : Ariberto Fassati
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Gene Therapy for Muscular Dystrophy Evaluation of a Muscle specific Promoter for Adenovirus mediated Gene Transfer

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Development of Gene Therapy for Duchenne Muscular Dystrophy Heart Disease in the MDX Mouse Model

Author : Brian P. Bostick
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Duchenne muscular dystrophy (DMD) is a fatal genetic muscle disease with no cure. DMD results from mutations in a critical muscle protein called dystrophin. Children born with DMD suffer severe muscle wasting leading to progressive weakness and paralysis. Patients usually die of respiratory or heart failure before the age of thirty. Gene therapy raises the hope of a cure for DMD heart disease. While significant strides have been made towards therapy for skeletal muscle disease, development of heart gene therapy lags behind. The seminal questions for realization of heart gene therapy of DMD include; developing an animal model, determining dosage, finding the correct gene, developing the vehicle for gene therapy and optimizing gene delivery. This dissertation details critical advancements towards gene therapy for DMD heart disease. First, we developed an animal model of DMD heart disease in the mdx mouse. We then determined that 50% mosaic dystrophin expression was sufficient to prevent DMD heart disease in this model. Next, we established that the truncated mini-dystrophin gene was capable of ameliorating DMD heart disease in the mdx mouse through cardiac specific transgenic expression. Then, we established the adeno-associated virus (AAV) as a vehicle for DMD heart gene therapy regardless of mouse age or the route of administration. Finally, we discovered that AAV-mediated truncated dystrophin gene therapy prevented DMD heart disease in neonatal mdx mice and ameliorated heart disease in symptomatic mdx mice. This work represents significant progress towards realization of an effective therapy for DMD heart disease.

Improving Stem Cell Based Therapy and Developing a Novel Gene Therapy Approach for Treating Duchenne Muscular Dystrophy DMD

Author : Mohammadsharif Tabebordbar
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In gene therapy, engineered gene products are delivered directly to muscle fibers as transgenes carried by viral vectors, such as Adeno Associated Viruses (AAVs). Viral- mediated delivery of a normal copy of the mutated genes into dystrophic muscle fibers holds big promise as a therapeutic avenue for Muscular Dystrophies. However, considering the indispensible role of satellite cells in muscle regeneration, an effective and long-term therapy for genetic muscle diseases requires restoration of gene expression in both dystrophic muscle fibers and satellite cells. Conventional gene therapy approaches lack the potential for long-term restoration of the mutated gene expression in satellite cells. In order to address this limitation, this study provides the proof of concept evidence for the use of a novel gene editing approach, which allows irreversible correction of the mutations in both dystrophic skeletal muscle fibers and satellite cells.

Structural Flexibility of the Dystrophin Rod Domain

Author : Scott Q. Harper
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Analysis of S MAR Vectors for Gene Therapy in Muscle

Author : Samah Fakhro
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Naked Plasmid DNA as a Vector for Gene Therapy in Skeletal Muscle

Author : Jillian Mary Mcmahon
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Gene Transfer and Gene Expression in Skeletal Muscle in Vitro and in Vivo and the Implications for Human Gene Therapy

Author : Mark Roman
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Evaluating Properties of Dystrophin and Delivery Methods of RAAV Gene Therapy for Duchenne Muscular Dystrophy

Author : Julian N. Ramos
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Duchenne muscular dystrophy (DMD) is a recessive muscle wasting disease caused by a deleterious mutation in the gene encoding the dystrophin protein. Dystrophin is an integral component the dystrophin-glycoprotein complex (DGC) that stabilizes the sarcolemma and allows transmission of mechanical force in striated muscle. Recombinant adeno-associated viral (rAAV) vectors have shown promise as a method for delivering therapeutic genes to dystrophic muscles. Vectors expressing miniaturized, or micro-, dystrophin proteins have repeatedly demonstrated rescue of rodent dystrophic animal models as well as improvement in larger dystrophic animal models. However, current micro-dystrophin constructs do not restore full, wild type function to transduced skeletal muscles. To improve the functionality of micro-dystrophin, we designed novel constructs and evaluated rAAV vector-treated dystrophic mice expressing these micro-dystrophins. We observed an improvement in functionality in two novel micro-dystrophins when compared to a previously established construct serving as our standard. We also examined the consequences of ablating micro-dystrophin expression in a mouse model. After determining that adult skeletal muscle falls into a dystrophic condition by three months after ablation, we concluded that rAAV vector-mediated gene therapy for DMD may require persistent expression of micro-dystrophin for life. We expanded on a previously reported immunosuppressive regimen in order to allow readministration of rAAV vectors in both dystrophic and wild type mice. Additionally, rAAV vectors effectively transduced striated muscle tissues after repeated, systemic delivery into wild type mice at doses that would be therapeutic for neuromuscular diseases. In order to further understand the tropism and properties of AAV serotypes that exhibit a high degree of tropism for skeletal muscle, we compared their transduction properties in mice and canine animal models. We found that AAV serotypes 6, 8, and 9 all poorly transduce myogenic satellite cells. We also determined that AAV8 transduces mouse and canine skeletal muscle at a lower efficiency than AAV serotypes 6 and 9. Yet, serotypes 6 and 9 exhibited similar transduction when administered into the jugular vein of canines at sub-saturating doses. These results expand on several aspects of rAAV-mediated gene therapy for DMD involving the a) design and functionality of the therapeutic construct, b) consequences of lost expression of micro-dystrophin, c) immune responses in relation to repeat transduction of rAAV vectors, and d) properties of AAV serotypes and methods of delivery.

Muscle Targeted Gene Therapy of Charcot Marie Tooth Disease is Dependent on Muscle Activity

Author : Stephan Klossner
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Muscle-Targeted Gene Therapy of Charcot Marie-Tooth Disease is Dependent on Muscle Activity.

Muscular Dystrophy

Author : Paula Johanson
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Muscular dystrophy (MD) is a genetic disorder that gradually weakens a person’s muscles. It is caused by missing or incorrect genetic information in the body’s cells. The person’s body is unable to make the proteins needed to build and maintain healthy muscles. Although there is no cure for muscular dystrophy, researchers are learning about how to prevent and treat the condition. Doctors have learned a great deal about how to improve muscle and joint function and how to slow the deterioration of muscles. Their goal is to help children, teenagers, and adults with muscular dystrophy live active and independent lives for as long as possible. This book delves into the history of the disorder, the genetic basis of MD, the main types of MD, and living with MD and suggests some coping strategies. It also explores diagnostic tests for MD, genetic research, and the promising effects of gene therapy.

Gene Replacement Restores the Contractile and Passive Properties of Skeletal Muscle in Murine Models of Duchenne Muscular Dystrophy

Author : Chady H. Hakim
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Duchenne muscular dystrophy (DMD) is a lethal disease caused by the loss of the dystrophin protein. Loss of mobility is a key clinical presentation in DMD. It is believed that deterioration in the mechanical properties (contractile and passive properties) of skeletal muscle contribute to reduction in mobility. These two sets of properties are inseparable aspects of muscle function. For example, elbow flexion is accomplished by the contraction of muscles in the anterior compartment of the upper arm and the passive stretch of muscles in the posterior compartment of the upper arm. To improve mobility of patients, both contractile and passive properties must be restored. Gene therapy holds great promise for treating DMD. Restoration of dystrophin expression using gene replacement strategies has improved the contractile force in mouse models of DMD. However, it is not yet known if gene replacement can also improve the passive properties. To address this concern, I performed comprehensive studies in my dissertation that provided new information on the passive properties changes in skeletal muscles of murine models of DMD, and have also offered new insights on how different strategies of gene replacement therapy may help improve the passive muscle properties in DMD. Together, these studies provided support to further develop dystrophin gene therapy to improve the loss of mobility in DMD patients.

Incretin based Muscle targeted Gene Therapy for Type 2 Diabetes

Author : Gillian Patterson
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Follistatin Gene Therapy for the Treatment of Muscular Dystrophy

Author : Chalonda Renee Handy
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Abstract: The muscular dystrophies (MDs) represent a group of inherited myogenic disorders characterized by persistent muscle wasting and progressive functional decline. To date, there are no optimal therapies for MD. Most potential treatments have been targeted towards improving conditions in Duchene Muscular Dystrophy where loss of dystrophin is responsible for the muscle defect. However, a number of emerging therapies are currently under review with particular attention being paid to factors that enhance muscle growth. The myostatin inhibitor follistatin, which results in enhanced muscle growth and function appears to offer substantial promise. The efficiency of follistatin lies in its ability to bind myostatin thus preventing its receptor binding that would normally allow for the downregulation of genes responsible for myoblast proliferation and differentiation. Follistatin is effective in that it not only circumvents the muscle inhibitory effects of myostatin, but is also able to operate outside of the myostatin pathway to offer muscle protection and improve pathology during instances of muscle wasting. Here, we highlighted the unique muscle enhancing effects of adeno-associated virus (AAV) delivery of follistatin (FS344) by using rodent models and non-human primates. We saw that in young as well as aged dystrophin deficient mdx mice, follistatin was able to improve muscle conditions and decrease dystrophic symptoms. The relatedness of follistatin to muscle disease was also seen in both local and systemic administration of follistatin gene therapy to severely dystrophic laminin-alpha2 deficient mice that closely recapitulate Merosin Deficient Congenital Muscular dystrophy. To investigate if follistatin was equally effective and safe in larger animals, in that this is an important criterion for clinical evaluations in humans, we administered a one-time injection of an AAV bearing follistatin into the quadriceps muscle of Cynomolgus macaques. Like our observations in mice, we saw significant improvements in muscle size and function as assessed by a physiological measure of muscle strength. To understand the possible mechanism for these follistatin induced changes, we performed a series of binding experiments as well as microarray analysis to study at the protein and transcriptional level the effects of muscle follistatin overexpression. Although not conclusive, these characterization studies set the stage for continued investigations into the follistatin mediated muscle response. Taken together this study not only provided evidence for the ability of follistatin to enhance muscle size and strength in both normal and dystrophic conditions, but also provides new starting points for understanding the molecular and functional relevance of myostatin inhibition by follistatin overexpression.

Towards Plasmid mediated Muscle targeted Incretin based Gene Therapy for Diabetes

Author : Aiman Mahmoud
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