Aspects of the disclosure relate to complexes comprising a muscle-targeting agent covalently linked to a molecular payload. In some embodiments, the muscle-targeting agent specifically binds to an internalizing cell surface receptor on muscle cells. In some embodiments, the molecular payload inhibits expression or activity of DUX4. In some embodiments, the molecular payload is an oligonucleotide, such as an antisense oligonucleotide or RNAi oligonucleotide.

The present invention relates to a method of isolating exosomes. Specifically, the invention relates to a method comprising the steps of providing a sample including exosomes; identifying a cell-surface polypeptide on the exosomes; and isolating the exosomes using the cell-surface polypeptide on the exosomes. The exosomes isolated from by the methods of the invention can be studied for the purposes of biomarker identification, for the understanding of biological function and disease, and to find ways to target them with therapeutics.

The disclosure relates to methods and compositions for regulating expression of DUX4. Specifically, the disclosure provides a recombinant gene editing complex comprising: a recombinant gene editing protein; and, a nucleic acid encoding a guide RNA (gRNA) that specifically hybridizes to a target nucleic acid sequence encoding a D4Z4 macrosatellite repeat region, wherein binding of the complex to the target nucleic acid sequence results in inhibition of DUX4 gene expression. In some aspects, methods described by the disclosure are useful for treating a disease associated with aberrant DUX4 expression (e.g., facioscapulohumeral muscular dystrophy, FSHD).

Nucleic acid regulatory elements that are able to enhance muscle-specific expression of genes, in particular expression in muscle cells and/or tissues such as in diaphragm, smooth muscle, heart and/or skeletal muscle, including at least two diaphragm-specific regulatory elements and a heart- and skeletal muscle-specific regulatory element. Expression cassettes and vectors containing these nucleic acid regulatory elements, as well as uses thereof such as applications using gene therapy of muscle-related disorders, more particularly diaphragm, heart and/or skeletal muscle-directed gene therapy, and for vaccination purposes.

The disclosure provides gene therapy vectors, such as adeno-associated virus (AAV), optimized for delivering a transgene to muscles. The optimized vectors contain constitutive or a muscle-specific promoter to deliver whole body or skeletal/heart muscle-specific transgene expression, respectively, in combination with a transgene cDNA to replace the gene mutation found in a muscle disease with a normal copy of the gene, an internal ribosomal entry site (IRES) to allow for production of a second protein from the same transcript, and a muscle growth factor, to build new muscle growth and strength. For example, the invention provides The disclosure provides gene therapy vectors, such as recombinant adeno-associated vims (rAAV), designed for treatment of GNE myopathy in which the rAAV expresses UDP-GlcNAc-epimerase/ManNAc-6 alone or in combination with a muscle growth factor or muscle transdifferentation factor. The provided AAV replace the mutated GNE gene expression while expressing proteins that stimulate muscle growth.

The invention provides gene therapy vectors, such as adeno-associated vims (AAV) vectors, expressing a miniaturized human micro-dystrophin gene and method of using these vectors to express micro-dystrophin in skeletal muscle including diaphragm and cardiac muscle and to protect muscle fibers from injury, increase muscle strength and reduce and/or prevent fibrosis in subjects suffering from muscular dystrophy.

Some embodiments of the invention include modified cells. Certain embodiments of the invention include methods of using modified cells. Other embodiments of the invention include methods of administering modified cells. Further embodiments of the invention include methods of administering modified cells to treat diseases. Additional embodiments of the invention are also discussed herein.

The present invention concerns an expression system for systemic administration comprising a sequence encoding a FKRP protein, and: —a promoter sequence allowing the expression at a therapeutically acceptable level of FKRP in the skeletal muscles and a target sequence of an miRNA expressed in the heart; or—a promoter sequence allowing the expression at a therapeutically acceptable level of FKRP in the skeletal muscles and presenting a promoter activity at a toxically acceptable level in the heart; and its use for the treatment of various diseases linked to FKRP deficiencies.

The invention provides gene therapy vectors, such as adeno-associated virus (AAV) vectors, expressing a functional fragment of the miniaturized human micro-dystrophin gene and method of using these vectors to express the fragment of micro-dystrophin in skeletal muscles including diaphragm and cardiac muscle and to protect muscle fibers from injury, increase muscle strength and reduce and/or prevent fibrosis in subjects suffering from muscular dystrophy.

The invention relates to methods and compositions for increasing introduction (e.g., transduction) efficiency. In particular, the invention relates to methods of increasing introduction (e.g., transduction) efficiency by contacting a host cell with a heterologous agent in the presence of a cell membrane fusion protein or a functional fragment or derivative thereof. The invention further relates to compositions that include a heterologous agent and a cell membrane fusion protein or a functional fragment or derivative thereof.