Patent Publication Number: US-2022228167-A1

Title: Peptide-Modified Hybrid Recombinant Adeno-Associated Virus Serotype Between AAV9 and AAVrh74 with Reduced Liver Tropism and Increased Muscle Transduction

Description:
FIELD OF THE INVENTION 
     The present invention relates to a recombinant adeno-associated virus (AAV) capsid, which is a peptide-modified hybrid between AAV serotype 9 (AAV9) and AAV serotype rh74 (AAVrh74) capsid proteins having a further reduced liver tropism and an increased muscle transduction compared to the hybrid AAV capsid protein not having the peptide. The invention relates also to the derived peptide-modified hybrid AAV serotype vector particles packaging a gene of interest, and their use in gene therapy, in particular for treating neuromuscular genetic diseases, in particular muscular genetic diseases. 
     BACKGROUND OF THE INVENTION 
     Recombinant Adeno-Associated Virus (rAAV) vectors are widely used for in vivo gene transfer. rAAV vectors are non-enveloped vectors composed of a capsid of 20 nm of diameter and a single strand DNA of 4.7 kb. The genome carries two genes, rep and cap, flanked by two palindromic regions named Inverted terminal Repeats (ITR). The cap gene codes for three structural proteins VP1, VP2 and VP3 that compose the AAV capsid. VP1, VP2 and VP3 share the same C-terminal end which is all of VP3. Using AAV2 has a reference, VP1 has a 735 amino acid sequence (GenBank YP_680426); VP2 (598 amino acids) starts at the Threonine 138 (T138) and VP3 (533 amino acids) starts at the methionine 203 (M203). AAV serotypes are defined by their capsid. Different serotypes exist, each of them displaying its own tissue targeting specificity. Therefore, the choice of using a serotype depends on the tissue to transduce. Skeletal muscle and liver tissues are infected and transduced efficiently by different serotypes of AAV vectors such as AAV8, AAV9 and 
     AAV-rh74. 
     Chimeric or hybrid AAV serotypes have been generated by exchanging fragments of capsid sequences between capsids of different naturally occurring AAV serotypes, in order to increase AAV transduction efficiency or increase AAV tropism to a cell or tissue type of interest. 
     Hybrid AAV capsids were generated by combining structural domains of capsids of AAV8 and AAV serotypes isolated from primate brain. The resulting AAV hybrid serotypes can transduce retinal tissue in human and mice with no increase in efficiency compared to AAV2 and AAV5 vectors (Charbel Issa et al., PLOS ONE, 2013, 8, e60361). However, one of the hybrid AAV serotype shows improved transduction efficiency for fat tissue compared to AAV1, AAV8 and AAV9 (Liu et al., Molecular Therapy, 2014, 1, 8, doi:10.1038/mtm). WO 2015/191508 discloses recombinant hybrid AAV capsids generated by exchanging variable regions of AAV capsids from various species (human, primate, avian, snake, bovine), in particular AAV capsids with central nervous system tropism to generate CNS specific chimeric capsids. 
     WO 2017/096164 discloses recombinant hybrid AAV capsids between AAV1, AAV2, AAV3b, AAV6 and AAV8 serotypes exhibiting enhanced human skeletal muscle tropism. However, all naturally occurring AAV serotypes and variants tested to date have a propensity to accumulate within the liver. This causes problems, in particular when the AAV vector is administered by the systemic route. Firstly, a transgene aimed to be expressed in muscle may have toxic effects on the liver. Secondly, AAV vector entry in liver reduces the amount of vector available for skeletal muscles. Consequently, higher doses of AAV vectors are required. This increases the possibility to induce liver toxicity and the cost of vector production. 
     Tissue-specific promoters and microRNA-based gene regulation strategies have been used to segregate gene expression patterns among different tissue types. However, such regulatory strategies do not preclude sequestration of AAV vector genomes in off-target organs such as the liver after systemic administration. 
     Attenuation of heparin binding by mutating the basic residues R585 or R588 of the capsid protein was shown to abolish heparin sulfate binding and reduce the liver tropism of AAV2-derived vectors (Asokan et al., Nat. Biotechnol., 2010, 28, 79-82). However, this strategy can only work for serotypes like AAV2 and AAV6 whose liver tropism is determined by basic residues binding to heparin. 
     Therefore, there is a need for new AAV vectors, having a reduced liver tropism and a concomitant increased muscle transduction. 
     An AAV2 vector displaying a RGDLGLS (SEQ ID NO: 8) peptide inserted into AAV capsid (position R588 relative to VP1 protein numbering) efficiently transduces primary murine breast cancer cells in vitro but fails to transduce breast cancer tumor cells in vivo (Michelfelder et al., PLoS ONE, 2009, 4, e5122). AAV9 displaying the same surface peptide (RGDLGLS or P1) targets efficiently human astrocytes in vitro (Kunze et al., Glia, 2018, 66, 413-427). 
     SUMMARY OF THE INVENTION 
     The inventors have generated a new peptide-modified hybrid AAV serotype using a combination of two serotypes that infect efficiently the muscle and liver tissues, AAV9 and AAV-rh74. The new peptide-modified hybrid AAV serotype was generated by the insertion of a peptide comprising the RGD motif into a variable region of the cap gene between the AAV9 and AAVrh74 serotypes ( FIG. 1 ). The inventors have found that surprisingly, the insertion of the peptide comprising the RGD motif on the hybrid capsid protein increased muscle transduction and ameliorated liver detargeting ( FIG. 2 ). 
     The new peptide-modified hybrid AAV serotypes are useful in gene therapy of diseases affecting muscle tissues, in particular skeletal muscle tissue and/or cardiac tissue, such as neuromuscular disorders, in particular muscular disorders, including genetic diseases, autoimmune diseases, neurodegenerative diseases and cancer. 
     Therefore, the invention encompasses a peptide-modified hybrid recombinant AAV capsid between AAV9 and AAVrh74 capsids with increased muscle transduction and reduced liver tropism, AAV vector particles comprising the peptide-modified hybrid recombinant AAV capsid, compositions comprising the peptide-modified hybrid AAV serotype vector particles, and methods of making and using said peptide-modified hybrid AAV serotype vector particles and compositions, in particular in gene therapy. 
     DETAILED DESCRIPTION OF THE INVENTION 
     Peptide-Modified Recombinant Hybrid AAV Capsid Protein 
     One aspect of the invention relates to a recombinant adeno-associated virus (AAV) capsid protein, which is a peptide-modified hybrid between AAV serotype 9 (AAV9) and AAV serotype 74 (AAVrh74) capsid proteins comprising at least one copy of a peptide comprising the RGD motif, wherein said recombinant peptide-modified hybrid AAV capsid protein has a further reduced liver tropism and an increased muscle transduction compared to the hybrid AAV capsid protein not having the peptide. 
     As used herein, the term “tropism” refers to the specificity of an AAV capsid protein present in an AAV viral particle, for infecting or transducing a particular type of cell or tissue. 
     The tropism of an AAV capsid for a particular type of cell or tissue may be determined by measuring the ability of AAV vector particles comprising the peptide-modified hybrid AAV capsid protein to infect or to transduce a particular type of cell or tissue, using standard assays that are well-known in the art such as those disclosed in the examples of the present application. 
     As used herein, the term “liver tropism” or “hepatic tropism” refers to the tropism for liver or hepatic tissue and cells, including hepatocytes. 
     In some embodiments, the liver tropism of the peptide-modified hybrid AAV capsid protein is further reduced by at least 5%, 10%, 20%, 30%, 40%, 50% or more compared to the liver tropism of the hybrid AAV capsid protein not having the peptide. 
     According to the invention, the peptide-modified hybrid AAV capsid protein has increased tropism or transduction for muscle cells and tissues compared to the hybrid AAV capsid not having the peptide. 
     Muscle tissues include in particular cardiac and skeletal muscle tissues. 
     As used herein, the term “muscle cells” refers to myocytes, myotubes, myoblasts, and/or satellite cells. 
     As used herein, “or” means “and/or”. 
     In some embodiments, the muscle tropism or muscle transduction of the peptide-modified hybrid AAV capsid protein is increased by at least 5%, 10%, 20%, 30%, 40%, 50% or more; preferably at least 50%, 60%, 70%, 80%, 90%, 99%, 100% or more compared to the hybrid AAV capsid protein not having the peptide. 
     In some embodiments, the peptide-modified hybrid AAV capsid protein is a peptide-modified hybrid VP1, VP2 or VP3 protein. 
     In some embodiments, the peptide-modified hybrid AAV capsid protein has tropism for at least skeletal muscle tissue. In some preferred embodiments, the peptide-modified hybrid AAV capsid protein has tropism for both skeletal and cardiac muscle tissues. An example of this type of peptide-modified hybrid is the peptide-modified hybrid AAV capsid of SEQ ID NO: 5 (named AAV-MT in the examples). This type of peptide-modified hybrid AAV capsid is useful for the treatment of cardiac and skeletal muscle disorders. 
     The peptide-modified hybrid AAV capsid protein according to the invention may be derived from any AAV9 and AAVrh74 capsid protein sequences; such sequences are well-known in the art and available in public sequence data base. For example, AAV9 capsid protein corresponds to GenBank accession numbers: AY530579.1; SEQ ID NO: 123 of WO 2005/033321; SEQ ID NO: 1 of WO 2012/112832; clade F AAV of WO 2016049230; AAV9 capsid variants in which one or more of the native residues at positions 271 (D), 446(Y), and 470 (N) are replaced with another amino acid, preferably alanine as disclosed in WO 2012/112832; AAV9 capsid variants at one or more of positions K143R, T251A, S499A, S669A and S490A as disclosed in US 2014/0162319. AAVrh74 capsid protein corresponds to SEQ ID NO: 1 of WO 2015/013313; SEQ ID NO: 6 of WO 2006/110689; SEQ ID NO: 1 of WO 2013/123503; SEQ ID NO: 4 of WO 2013/158879; and K137R, K333R, K550R, K552R, K569R, K691R, K695R, K709R variants and combination thereof. 
     In some embodiments, the peptide-modified hybrid AAV capsid protein according to the invention is derived from the AAV9 capsid protein of SEQ ID NO: 1 (GenBank AY530579.1) and the AAVrh74 protein of SEQ ID NO: 2. 
     In some embodiments, the peptide-modified hybrid AAV capsid protein according to the invention results from the replacement of a variable region in the AAV9 or AAVrh74 capsid sequence with the corresponding variable region of the other AAV serotype capsid sequence,
         wherein the variable region of AAV9 capsid corresponds to the sequence situated from any one of positions 331 to 493 to any one of positions 556 to 736 in AAV9 capsid of SEQ ID NO: 1 (reference sequence), or a fragment of at least 10, 15, 20, 25, 30, 35, 40, 45, 50, 55 or 60 consecutive amino acids of the sequence situated from positions 493 to 556 in AAV9 capsid of SEQ ID NO: 1, and   the variable region of AAVrh74 capsid corresponds to the sequence situated from any one of positions 332 to 495 to any one of positions 558 to 738 in AAVrh74 capsid of SEQ ID NO: 2 (reference sequence), or a fragment of at least 10, 15, 20, 25, 30, 35, 40, 45, 50, 55 or 60 consecutive amino acids of the sequence situated from positions 495 to 558 in AAVrh74 capsid of SEQ ID NO: 2.       

     The invention encompasses peptide-modified hybrid AAV capsid proteins derived from any AAV9 and AAVrh74 capsid protein sequences by replacement of a variable region in the AAV9 or AAVrh74 capsid sequence with the corresponding variable region of the other AAV serotype capsid sequence, as defined above. According to the invention, the variable region is defined using AAV9 capsid of SEQ ID NO: 1 and AAVrh74 capsid of SEQ ID NO: 2 as reference. After sequence alignment of any other AAV9 capsid sequence with SEQ ID NO: 1 or any of other AAVrh74 capsid sequence with SEQ ID NO: 2, using standard protein sequence alignment programs that are well-known in the art, such as for example BLAST, FASTA, CLUSTALW, and the like, a person skilled in the art can easily obtained the corresponding positions of the variable region in other AAV9 or AAVrh74 capsid sequences. 
     In some preferred embodiments, the peptide-modified hybrid AAV capsid protein according to the invention results from the replacement of the variable region corresponding to that situated from positions 449 to 609 in the AAV9 capsid sequence of SEQ ID NO: 1 or from positions 450 to 611 in the AAVrh74 capsid sequence of SEQ ID NO: 2 with the corresponding variable region of the other serotype. 
     In some preferred embodiments, the peptide-modified hybrid AAV capsid protein according to the invention is derived from a hybrid AAV capsid selected from the group consisting of: SEQ ID NO: 3, SEQ ID NO: 4, and the sequences having at least 85%, 90%, 95%, 97%, 98% or 99% identity with said sequences. SEQ ID NO: 3 is derived from AAV9 capsid protein of SEQ ID NO: 1 by replacement of AAV9 variable region (positions 449 to 609 of SEQ ID NO: 1) with the variable region of AAVrh74 capsid protein (positions 450 to 611 of SEQ ID NO: 2); the corresponding hybrid is named Hybrid Cap9-rh74 in the examples. VP2 corresponds to the amino acid sequence from T138 to the end of SEQ ID NO: 3. VP3 corresponds to the amino acid sequence from M203 to the end of SEQ ID NO: 3. SEQ ID NO: 4 is derived from AAVrh74 capsid protein of SEQ ID NO: 2 by replacement of rh74 variable region (positions 450 to 611 of SEQ ID NO: 2) with the variable region of AAV9 capsid protein (positions 449 to 609 of SEQ ID NO: 1); the corresponding hybrid is named Hybrid Caprh74-9 in the examples. VP2 corresponds to the amino acid sequence from T138 to the end of SEQ ID NO: 4. VP3 corresponds to the amino acid sequence from M204 to the end of SEQ ID NO: 4. 
     In some preferred embodiments, the peptide-modified hybrid AAV capsid protein according to the invention is derived from AAV9 capsid protein by replacement of a variable region of AAV9 capsid sequence with the corresponding variable region of AAVrh74 capsid sequence as defined above, preferably the peptide-modified hybrid AAV capsid protein comprises the replacement of the variable region corresponding to that situated from positions 449 to 609 in AAV9 capsid of SEQ ID NO: 1 with the variable region corresponding to that situated from positions 450 to 611 in AAVrh74 capsid of SEQ ID NO: 2. In some more preferred embodiments, the peptide-modified hybrid AAV capsid protein according to the invention is derived from a hybrid AAV capsid protein selected from the group consisting of: of SEQ ID NO: 3 and the sequences having at least 85%, 90%, 95%, 97%, 98% or 99% identity with said sequence; preferably SEQ ID NO: 3. 
     The term “identity” refers to the sequence similarity between two polypeptide molecules or between two nucleic acid molecules. When a position in both compared sequences is occupied by the same base or same amino acid residue, then the respective molecules are identical at that position. The percentage of identity between two sequences corresponds to the number of matching positions shared by the two sequences divided by the number of positions compared and multiplied by 100. Generally, a comparison is made when two sequences are aligned to give maximum identity. The identity may be calculated by alignment using, for example, the GCG (Genetics Computer Group, Program Manual for the GCG Package, Version 7, Madison, Wisconsin) pileup program, or any of sequence comparison algorithms such as BLAST, FASTA or CLUSTALW. 
     The peptide comprising the RGD motif is preferably of up to 30 amino acids. 
     In some embodiments the peptide of up to 30 amino acids comprises or consists of any one of: RGDLGLS (SEQ ID NO: 8), LRGDGLS (SEQ ID NO: 14), LGRGDLS (SEQ ID NO: 15), LGLRGDS (SEQ ID NO: 16), LGLSRGD (SEQ ID NO: 17) and RGDMSRE (SEQ ID NO: 18); preferably SEQ ID NO: 8. The sequences SEQ ID NO: 8 and 14 to 18 may be flanked by up to five or more amino acids at their N- and/or C-terminal ends, such as for example by GQSG (SEQ ID NO: 9) and AQAA (SEQ ID NO: 10), respectively at the N- and C-terminal end of the peptide. 
     The peptide-modified hybrid AAV capsid protein of the invention may comprise up to 5 copies of the peptide comprising the RGD motif, preferably 1 copy of said peptide. The peptide-modified hybrid AAV capsid protein may comprise multiple copies of the same peptide or one or more copies of different peptides. 
     The peptide-modified hybrid AAV capsid protein of the invention comprises the one or more peptide(s) comprising the RGD motif, inserted into a site exposed on the AAV capsid surface. Sites on the AAV capsid which are exposed on the capsid surface and tolerate peptide insertions, i.e. do not affect assembly and packaging of the virus capsid, are well-known in the art and include for example the AAV capsid surface loops or antigenic loops (Girod et al., Nat. Med., 1999, 5, 1052-1056; Grifman et al., Molecular Therapy, 2001, 3, 964-975); other sites are disclosed in Rabinowitz et al., Virology, 1999, 265, 274-285; Wu et al., J. Virol., 2000, 74, 8635-8647. 
     In particular, the peptide(s) comprising the RGD motif are inserted around any of positions 261, 383, 449, 575 or 590 according to the numbering in SEQ ID NO: 3, preferably around position 449 or 590, more preferably around position 590. The positions are indicated by reference to SEQ ID NO: 3; one skilled in the art will be able to find easily the corresponding positions in another sequence after alignment with SEQ ID NO: 3. 
     The insertion site is advantageously from positions 587 to 592 or 588 to 593 according to the numbering in SEQ ID NO: 3, preferably from positions 587 to 592. The insertion of the peptide may or may not cause the deletion of some or all of the residue(s) from the insertion site. The peptide advantageously replaces all the residues from positions 587 to 592 or 588 to 593 of the AAV capsid protein according to the numbering in SEQ ID NO: 3, preferably all of the residues from positions 587 to 592. 
     In some preferred embodiments, the peptide replaces all the residues from positions 587 to 592 of the AAV capsid protein according to the numbering in SEQ ID NO: 3. Preferably, said peptide comprises any one of RGDLGLS (SEQ ID NO: 8), LRGDGLS (SEQ ID NO: 14), LGRGDLS (SEQ ID NO: 15), LGLRGDS (SEQ ID NO: 16), LGLSRGD (SEQ ID NO: 17) and RGDMSRE (SEQ ID NO: 18); preferably SEQ ID NO: 8. A more preferred peptide is RGDLGLS (SEQ ID NO: 8) flanked by GQSG (SEQ ID NO: 9) and AQAA (SEQ ID NO: 10), respectively at its N- and C-terminal end, corresponding to GQSGRGDLGLSAQAA (SEQ ID NO: 13) . 
     In some preferred embodiment, the peptide-modified hybrid AAV capsid protein comprises or consists of a sequence selected from the group consisting of SEQ ID NO: 5, the sequences having at least 85%, 90%, 95%, 97%, 98% or 99% identity with said sequence and the fragment thereof corresponding to VP2 or VP3 capsid protein. VP2 corresponds to the amino acid sequence from T138 to the end of SEQ ID NO: 5. VP3 corresponds to the amino acid sequence from M203 to the end of SEQ ID NO: 5. In some preferred embodiments, said peptide-modified hybrid AAV capsid protein comprises the sequence SEQ ID NO: 5 or a fragment thereof corresponding to VP2 or VP3 capsid protein. SEQ ID NO: 5 is derived from the hybrid Cap9-rh74 of SEQ ID NO: 3 by the insertion of the peptide of SEQ ID NO: 8. 
     In some preferred embodiments, said peptide-modified hybrid AAV capsid protein comprises a sequence selected from the group consisting of the sequence SEQ ID NO: 5, the sequences having at least 85%, 90%, 95%, 97%, 98% or 99% identity with said sequence, and the fragment thereof corresponding to VP2 or VP3 capsid protein. VP2 corresponds to the amino acid sequence from T138 to the end of SEQ ID NO: 5. VP3 corresponds to the amino acid sequence from M203 to the end of SEQ ID NO: 5. In some preferred embodiments, said peptide-modified hybrid AAV capsid protein comprises the sequence SEQ ID NO: 5 or a fragment thereof corresponding to VP2 or VP3 capsid protein. 
     The invention encompasses also AAV VP1 and VP2 chimeric capsid proteins derived from the peptide-modified AAV9/rh74 hybrid VP3 capsid protein according to the invention, wherein the VP1-specific N-terminal region and/or VP2-specific N-terminal region are from a natural or artificial AAV serotype other than AAV9 and AAVrh74. 
     In some embodiments, the peptide-modified AAV VP1 chimeric capsid protein comprises:
         (i) a VP1-specific N-terminal region having a sequence from natural or artificial AAV serotype other than AAV9 and AAVrh74,   (ii) a VP2-specific N-terminal region having a sequence from AAV9, AAVrh74 or natural or artificial AAV serotype other than AAV9 and AAVrh74, and   (iii) a VP3 C-terminal region having the sequence of a peptide-modified hybrid VP3 protein according to the invention.       

     In some embodiments, the AAV VP2 chimeric capsid proteins comprises
         (i) a VP2-specific N-terminal region having a sequence from natural or artificial AAV serotype other than AAV9 and AAVrh74, and   (ii) a VP3 C-terminal region having the sequence of a peptide-modified hybrid VP3 protein according to the invention.       

     Polynucleotide, Vector, and Use for AAV Vector Production 
     Another aspect of the invention is a polynucleotide encoding the recombinant peptide-modified hybrid AAV capsid protein in expressible form. The polynucleotide may be DNA, RNA or a synthetic or semi-synthetic nucleic acid. 
     In some embodiments, the polynucleotide is a AAV9/rh74 hybrid cap gene encoding peptide-modified hybrid VP1, VP2 and VP3 capsid proteins according to the invention. In some preferred embodiments, the polynucleotide comprises the sequence SEQ ID NO: 6 (encoding the peptide-modified hybrid AAV capsid protein of SEQ ID NO: 5). 
     In some other embodiments, the polynucleotide is a chimeric cap gene which encodes for a peptide-modified AAV9/rh74 hybrid VP3 capsid protein according to the invention and a chimeric VP1 capsid protein, and maybe also a chimeric VP2 capsid protein wherein the VP1-specific N-terminal region, and maybe also the VP2-specific N-terminal region, are from a natural or artificial AAV serotype other than AAV9 and AAVrh74. Such chimeric cap gene may be generated by any suitable technique, using the coding sequence for a peptide-modified AAV9/rh74 hybrid VP3 capsid protein according to the invention in combination with heterologous sequences which may be obtained from different selected AAV serotypes, non-contiguous portions of the same AAV serotypes, from a non-viral AAV source or from a non-viral source. 
     In some embodiments, the polynucleotide further encodes AAV Replicase (Rep) protein in expressible form, preferably Rep from AAV2. 
     The polynucleotide is advantageously inserted into a recombinant vector, which includes, in a non-limiting manner, linear or circular DNA or RNA molecules consisting of chromosomal, non-chromosomal, synthetic or semi-synthetic nucleic acids, such as in particular viral vectors, plasmid or RNA vectors. 
     Numerous vectors into which a nucleic acid molecule of interest can be inserted in order to introduce it into and maintain it in a eukaryotic host cell are known per se; the choice of an appropriate vector depends on the use envisioned for this vector (for example, replication of the sequence of interest, expression of this sequence, maintaining of this sequence in extrachromosomal form, or else integration into the chromosomal material of the host), and also on the nature of the host cell. 
     In some embodiments, the vector is a plasmid. 
     The recombinant vector for use in the present invention is an expression vector comprising appropriate means for expression of the peptide-modified hybrid AAV capsid protein, and maybe also AAV Rep protein. Usually, each coding sequence (hybrid AAV Cap and AAV Rep) is inserted in a separate expression cassette either in the same vector or separately. Each expression cassette comprises the coding sequence (open reading frame or ORF) functionally linked to the regulatory sequences which allow the expression of the corresponding protein in AAV producer cells, such as in particular promoter, promoter/enhancer, initiation codon (ATG), stop codon, transcription termination signal. Alternatively, the hybrid AAV Cap and the AAV Rep proteins may be expressed from a unique expression cassette using an Internal Ribosome Entry Site (IRES) inserted between the two coding sequences or a viral 2A peptide. In addition, the codon sequences encoding the hybrid AAV Cap, and AAV Rep if present, are advantageously optimized for expression in AAV producer cells, in particular human producer cells. 
     The vector, preferably a recombinant plasmid, is useful for producing hybrid AAV vectors comprising the peptide-modified hybrid AAV capsid protein of the invention, using standard AAV production methods that are well-known in the art (Review in Aponte-Ubillus et al., Applied Microbiology and Biotechnology, 2018, 102: 1045-1054). 
     Following co-transfection, the cells are incubated for a time sufficient to allow the production of AAV vector particles, the cells are then harvested, lysed, and AAV vector particles are purified by standard purification methods such as affinity chromatography or Cesium Chloride density gradient ultracentrifugation. 
     AAV Particle, Pharmaceutical Composition and Therapeutic Uses 
     Another aspect of the invention is an AAV particle comprising the peptide-modified hybrid recombinant AAV capsid protein of the invention. The AAV particle may comprise peptide-modified hybrid VP1, VP2 and VP3 capsid proteins encoded by a hybrid cap gene according to the invention. Alternatively or additionally, the AAV particle may comprise chimeric VP1 and VP2 capsid proteins and a peptide-modified hybrid VP3 protein encoded by a chimeric cap gene according to the invention. 
     In some embodiments, the AAV particle is a mosaic AAV particle further comprising another AAV capsid protein from a natural or artificial AAV serotype other than AAV9 and AAVrh74 serotype, wherein the mosaic AAV particle has a reduced liver tropism compared to AAV9 and AAVrh74 serotypes. An artificial AAV serotype may be with no limitation, a chimeric AAV capsid, a recombinant AAV capsid, or a humanized AAV capsid. Such an artificial capsid may be generated by any suitable technique, using a selected AAV sequence (e.g. a fragment of a VP1 capsid protein) in combination with heterologous sequences which may be obtained from a different selected AAV serotype, non-contiguous portions of the same AAV serotype, from a non-viral AAV source or from a non-viral source. 
     Preferably, the AAV particle is an AAV vector particle. The genome of the AAV vector may either be a single-stranded or self-complementary double-stranded genome (McCarty et al, Gene Therapy, 2003, Dec., 10(26), 2112-2118). Self-complementary vectors are generated by deleting the terminal resolution site (trs) from one of the AAV terminal repeats. These modified vectors, whose replicating genome is half the length of the wild-type AAV genome have the tendency to package DNA dimers. The AAV genome is flanked by ITRs. In particular embodiments, the AAV vector is a pseudotyped vector, i.e. its genome and capsid are derived from AAVs of different serotypes. In some preferred embodiments, the genome of the pseudotyped vector is derived from AAV2. 
     In some preferred embodiments, the AAV vector particle is packaging a gene of interest. 
     The AAV particle may be obtained using the method of producing recombinant AAV vector particles of the invention. 
     By “gene of interest”, it is meant a gene useful for a particular application, such as with no limitation, diagnosis, reporting, modifying, therapy and genome editing. 
     For example, the gene of interest may be a therapeutic gene, a reporter gene or a genome-editing enzyme. 
     By “gene of interest for therapy”, “gene of therapeutic interest”, or “heterologous gene of interest”, it is meant a therapeutic gene or a gene encoding a therapeutic protein, peptide or RNA. 
     The gene of interest is any nucleic acid sequence capable of modifying a target gene or target cellular pathway, in particular in muscle cells. For example, the gene may modify the expression, sequence or regulation of the target gene or cellular pathway. In some embodiments, the gene of interest is a functional version of a gene or a fragment thereof. The functional version of said gene includes the wild-type gene, a variant gene such as variants belonging to the same family and others, or a truncated version, which preserves the functionality of the encoded protein at least partially. A functional version of a gene is useful for replacement or additive gene therapy to replace a gene, which is deficient or non-functional in a patient. In other embodiments, the gene of interest is a gene which inactivates a dominant allele causing an autosomal dominant genetic disease. A fragment of a gene is useful as recombination template for use in combination with a genome editing enzyme. 
     Alternatively, the gene of interest may encode a protein of interest for a particular application, (for example an antibody or antibody fragment, a genome-editing enzyme) or a RNA. In some embodiments, the protein is a therapeutic protein including a therapeutic antibody or antibody fragment, or a genome-editing enzyme. In some embodiments, the RNA is a therapeutic RNA. The gene of interest is a functional gene able to produce the encoded protein, peptide or RNA in the target cells of the disease, in particular muscle cells. 
     The AAV viral vector comprises the gene of interest in a form expressible in muscle cells, including cardiac and skeletal muscle cells. In particular, the gene of interest is operatively linked to a ubiquitous, tissue-specific or inducible promoter which is functional in muscle cells. The gene of interest may be inserted in an expression cassette further comprising polyA sequences. 
     The RNA is advantageously complementary to a target DNA or RNA sequence or binds to a target protein. For example, the RNA is an interfering RNA such as a shRNA, a microRNA, a guide RNA (gRNA) for use in combination with a Cas enzyme or similar enzyme for genome editing, an antisense RNA capable of exon skipping such as a modified small nuclear RNA (snRNA) or a long non-coding RNA. The interfering RNA or microRNA may be used to regulate the expression of a target gene involved in muscle disease. The guide RNA in complex with a Cas enzyme or similar enzyme for genome editing may be used to modify the sequence of a target gene, in particular to correct the sequence of a mutated/deficient gene or to modify the expression of a target gene involved in a disease, in particular a neuromuscular disease. The antisense RNA capable of exon skipping is used in particular to correct a reading frame and restore expression of a deficient gene having a disrupted reading frame. In some embodiments, the RNA is a therapeutic RNA. 
     The genome-editing enzyme according to the invention is any enzyme or enzyme complex capable of modifying a target gene or target cellular pathway, in particular in muscle cells. For example, the genome-editing enzyme may modify the expression, sequence or regulation of the target gene or cellular pathway. The genome-editing enzyme is advantageously an engineered nuclease, such as with no limitations, a meganuclease, zinc finger nuclease (ZFN), transcription activator-like effector-based nuclease (TALENs), Cas enzyme from clustered regularly interspaced palindromic repeats (CRISPR)-Cas system and similar enzymes. The genome-editing enzyme, in particular an engineered nuclease such as Cas enzyme and similar enzymes, may be a functional nuclease which generates a double-strand break (DSB) in the target genomic locus and is used for site-specific genome editing applications, including with no limitations: gene correction, gene replacement, gene knock-in, gene knock-out, mutagenesis, chromosome translocation, chromosome deletion, and the like. For site-specific genome editing applications, the genome-editing enzyme, in particular an engineered nuclease such as Cas enzyme and similar enzymes may be used in combination with a homologous recombination (HR) matrix or template (also named DNA donor template) which modifies the target genomic locus by double-strand break (DSB)-induced homologous recombination. In particular, the HR template may introduce a transgene of interest into the target genomic locus or repair a mutation in the target genomic locus, preferably in an abnormal or deficient gene causing a neuromuscular disease. Alternatively, the genome-editing enzyme, such as Cas enzyme and similar enzymes may be engineered to become nuclease-deficient and used as DNA-binding protein for various genome engineering applications such as with no limitation: transcriptional activation, transcriptional repression, epigenome modification, genome imaging, DNA or RNA pull-down and the like. 
     Another aspect of the invention is a pharmaceutical composition comprising a therapeutically effective amount of AAV particles comprising the peptide-modified hybrid recombinant AAV capsid protein of the invention, preferably AAV vector particles packaging a therapeutic gene of interest. 
     In some embodiments of the invention, the pharmaceutical composition of the invention is for use as a medicament, in particular in gene therapy. The invention encompasses the use of the pharmaceutical composition of the invention as a medicament, in particular for the treatment of a disease by gene therapy. 
     Gene therapy can be performed by gene transfer, gene editing, exon skipping, RNA-interference, trans-splicing or any other genetic modification of any coding or regulatory sequences in the cell, including those included in the nucleus, mitochondria or as commensal nucleic acid such as with no limitation viral sequences contained in cells. 
     The two main types of gene therapy are the following:
         a therapy aiming to provide a functional replacement gene for a deficient/abnormal gene: this is replacement or additive gene therapy;   a therapy aiming at gene or genome editing: in such a case, the purpose is to provide to a cell the necessary tools to correct the sequence or modify the expression or regulation of a deficient/abnormal gene so that a functional gene is expressed or an abnormal gene is suppressed (inactivated): this is gene editing therapy.       

     In additive gene therapy, the gene of interest may be a functional version of a gene, which is deficient or mutated in a patient, as is the case for example in a genetic disease. In such a case, the gene of interest will restore the expression of a functional gene. 
     Gene or genome editing uses one or more gene(s) of interest, such as:
         (i) a gene encoding a therapeutic RNA as defined above such as an interfering RNA like a shRNA or a microRNA, a guide RNA (gRNA) for use in combination with a Cas enzyme or similar enzyme, or an antisense RNA capable of exon skipping such as a modified small nuclear RNA (snRNA); and   (ii) a gene encoding a genome-editing enzyme as defined above such as an engineered nuclease like a meganuclease, zinc finger nuclease (ZFN), transcription activator-like effector-based nuclease (TALENs), Cas enzyme or similar enzymes; or a combination of such genes, and maybe also a fragment of a functional version of a gene for use as recombination template, as defined above.       

     Gene therapy is used for treating various diseases, including with no limitations, genetic diseases, in particular neuromuscular genetic disorders such as muscular genetic disorders; cancer; neurodegenerative diseases and auto-immune diseases. 
     In some embodiments, gene therapy is used for treating diseases affecting muscle tissues, in particular skeletal muscle tissue and/or cardiac tissue, such as with no-limitations: neuromuscular genetic disorders, in particular muscular genetic disorders such as. 
     Examples of mutated genes in neuromuscular genetic disorders, including muscular genetic disorders that can be targeted by gene therapy using the pharmaceutical composition of the invention are listed in the following tables: 
     Muscular Dystrophies 
       
     
       
         
           
               
               
             
               
                   
               
               
                 Gene 
                 Protein 
               
               
                   
               
             
            
               
                 DMD 
                 Dystrophin 
               
               
                 EMD 
                 Emerin 
               
               
                 FHL1 
                 Four and a half LIM domain 1 
               
               
                 LMNA 
                 Lamin A/C 
               
               
                 SYNE1 
                 Spectrin repeat containing, nuclear envelope 1 (nesprin 1) 
               
               
                 SYNE2 
                 Spectrin repeat containing, nuclear envelope 2 (nesprin 2) 
               
               
                 TMEM43 
                 Transmembrane protein 43 
               
               
                 TOR1AIP1 
                 Torsin A interacting protein 1 
               
               
                 DUX4 
                 Double homeobox 4 
               
               
                 SMCHD1 
                 Structural maintenance of chromosomes flexible hinge 
               
               
                   
                 domain containing 1 
               
               
                 PTRF 
                 Polymerase I and transcript release factor 
               
               
                 MYOT 
                 Myotilin 
               
               
                 CAV3 
                 Caveolin 3 
               
               
                 DNAJB6 
                 HSP-40 homologue, subfamily B, number 6 
               
               
                 DES 
                 Desmin 
               
               
                 TNPO3 
                 Transportin 3 
               
               
                 HNRNPDL 
                 Heterogeneous nuclear ribonucleoprotein D-like 
               
               
                 CAPN3 
                 Calpain 3 
               
               
                 DYSF 
                 Dysferlin 
               
               
                 SGCG 
                 Gamma sarcoglycan 
               
               
                 SGCA 
                 Alpha sarcoglycan 
               
               
                 SGCB 
                 Beta sarcoglycan 
               
               
                 SGCD 
                 Delta-sarcoglycan 
               
               
                 TCAP 
                 Telethonin 
               
               
                 TRIM32 
                 Tripartite motif-containing 32 
               
               
                 FKRP 
                 Fukutin-related protein 
               
               
                 TTN 
                 Titin 
               
               
                 POMT1 
                 Protein-O-mannosyltransferase 1 
               
               
                 ANO5 
                 Anoctamin 5 
               
               
                 FKTN 
                 Fukutin 
               
               
                 POMT2 
                 Protein-O-mannosyltransferase 2 
               
               
                 POMGNT1 
                 O-linked mannose beta1,2-N-acetylglucosaminyltransferase 
               
               
                 PLEC 
                 Plectin 
               
               
                 TRAPPC11 
                 trafficking protein particle complex 11 
               
               
                 GMPPB 
                 GDP-mannose pyrophosphorylase B 
               
               
                 DAG1 
                 Dystroglycan1 
               
               
                 DPM3 
                 Dolichyl-phosphate mannosyltransferase polypeptide 3 
               
               
                 ISPD 
                 Isoprenoid synthase domain containing 
               
               
                 VCP 
                 Valosin-containing protein 
               
               
                 LIMS2 
                 LIM and senescent cell antigen-like domains 2 
               
               
                 GAA 
                 Glucosidase alpha, acid 
               
               
                   
               
            
           
         
       
     
     Congenital Muscular Dystrophies 
       
     
       
         
           
               
               
             
               
                   
               
               
                 Gene 
                 Protein 
               
               
                   
               
             
            
               
                 LAMA2 
                 Laminin alpha 2 chain of merosin 
               
               
                 COL6A1 
                 Alpha 1 type VI collagen 
               
               
                 COL6A2 
                 Alpha 2 type VI collagen 
               
               
                 COL6A3 
                 Alpha 3 type VI collagen 
               
               
                 SEPN1 
                 Selenoprotein N1 
               
               
                 FHL1 
                 Four and a half LIM domain 1 
               
               
                 ITGA7 
                 Integrin alpha 7 precursor 
               
               
                 DNM2 
                 Dynamin 2 
               
               
                 TCAP 
                 Telethonin 
               
               
                 LMNA 
                 Lamin A/C 
               
               
                 FKTN 
                 Fukutin 
               
               
                 POMT1 
                 Protein-O-mannosyltransferase 1 
               
               
                 POMT2 
                 Protein-O-mannosyltransferase 2 
               
               
                 FKRP 
                 Fukutin-related protein 
               
               
                 POMGNT1 
                 O-linked mannose beta1,2-N-acetylglucosaminyltransferase 
               
               
                 ISPD 
                 Isoprenoid synthase domain containing 
               
               
                 POMGNT2 
                 protein O-linked mannose N-acetylglucosaminyltransferase 
               
               
                   
                 2 
               
               
                 B3GNT1 
                 UDP-GlcNAc:betaGal beta-1,3-N-acetylglucosaminyl- 
               
               
                   
                 transferase 1 
               
               
                 GMPPB 
                 GDP-mannose pyrophosphorylase B 
               
               
                 LARGE 
                 Like-glycosyltransferase 
               
               
                 DPM1 
                 Dolichyl-phosphate mannosyltransferase 1, catalytic subunit 
               
               
                 DPM2 
                 Dolichyl-phosphate mannosyltransferase polypeptide 2, 
               
               
                   
                 regulatory subunit 
               
               
                 ALG13 
                 UDP-N-acetylglucosami-nyltransferase 
               
               
                 B3GALNT2 
                 Beta-1,3-N-acetylgalacto-saminyltransferase 2 
               
               
                 TMEM5 
                 Transmembrane protein 5 
               
               
                 POMK 
                 Protein-O-mannose kinase 
               
               
                 CHKB 
                 Choline kinase beta 
               
               
                 ACTA1 
                 Alpha actin, skeletal muscle 
               
               
                 TRAPPC11 
                 trafficking protein particle complex 11 
               
               
                   
               
            
           
         
       
     
     Congenital Myopathies 
       
     
       
         
           
               
               
             
               
                   
               
               
                 Gene 
                 Protein 
               
               
                   
               
             
            
               
                 TPM3 
                 Tropomyosin 3 
               
               
                 NEB 
                 Nebulin 
               
               
                 ACTA1 
                 Alpha actin, skeletal muscle 
               
               
                 TPM2 
                 Tropomyosin 2 (beta) 
               
               
                 TNNT1 
                 Slow troponin T 
               
               
                 KBTBD13 
                 Kelch repeat and BTB (POZ) domain containing 13 
               
               
                 CFL2 
                 Cofilin 2 (muscle) 
               
               
                 KLHL40 
                 Kelch-like family member 40 
               
               
                 KLHL41 
                 Kelch-like family member 41 
               
               
                 LMOD3 
                 Leiomodin 3 (fetal) 
               
               
                 SEPN1 
                 Selenoprotein N1 
               
               
                 RYR1 
                 Ryanodine receptor 1 (skeletal) 
               
               
                 MYH7 
                 Myosin, heavy polypeptide 7, cardiac muscle, beta 
               
               
                 MTM1 
                 Myotubularin 
               
               
                 DNM2 
                 Dynamin 2 
               
               
                 BIN1 
                 Amphiphysin 
               
               
                 TTN 
                 Titin 
               
               
                 SPEG 
                 SPEG complex locus 
               
               
                 MEGF10 
                 Multiple EGF-like-domains 10 
               
               
                 MYH2 
                 Myosin, heavy polypeptide 2, skeletal muscle 
               
               
                 MYBPC3 
                 Cardiac myosin binding protein-C 
               
               
                 CNTN1 
                 Contactin-1 
               
               
                 TRIM32 
                 Tripartite motif-containing 32 
               
               
                 PTPLA 
                 Protein tyrosine phosphatase-like (3-Hydroxyacyl-CoA 
               
               
                   
                 dehydratase 
               
               
                 CACNA1S 
                 Calcium channel, voltage-dependent, L type, alpha 1S 
               
               
                   
                 subunit 
               
               
                   
               
            
           
         
       
     
     Distal Myopathies 
       
     
       
         
           
               
               
             
               
                   
               
               
                 Gene 
                   
               
               
                 symbol 
                 protein 
               
               
                   
               
             
            
               
                 DYSF 
                 Dysferlin 
               
               
                 TTN 
                 Titin 
               
               
                 GNE 
                 UDP-N-acetylglucosamine-2- epimerase/N-acetylmannosamine 
               
               
                   
                 kinase 
               
               
                 MYH7 
                 Myosin, heavy polypeptide 7, cardiac muscle, beta 
               
               
                 MATR3 
                 Matrin 3 
               
               
                 TIA1 
                 Cytotoxic granuleassociated RNA binding protein 
               
               
                 MYOT 
                 Myotilin 
               
               
                 NEB 
                 Nebulin 
               
               
                 CAV3 
                 Caveolin 3 
               
               
                 LDB3 
                 LIM domain binding 3 
               
               
                 ANO5 
                 Anoctamin 5 
               
               
                 DNM2 
                 Dynamin 2 
               
               
                 KLHL9 
                 Kelch-like homologue 9 
               
               
                 FLNC 
                 Filamin C, gamma (actin-binding protein - 280) 
               
               
                 VCP 
                 Valosin-containing protein 
               
               
                   
               
            
           
         
       
     
     Other Myopathies 
       
     
       
         
           
               
               
             
               
                   
               
               
                 Gene 
                   
               
               
                 symbol 
                 protein 
               
               
                   
               
             
            
               
                 ISCU 
                 Iron-sulfur cluster scaffold homolog ( E. coli ) 
               
               
                 MSTN 
                 Myostatin 
               
               
                 FHL1 
                 Four and a half LIM domain 1 
               
               
                 BAG3 
                 BCL2-associated athanogene 3 
               
               
                 ACVR1 
                 Activin A receptor, type II-like kinase 2 
               
               
                 MYOT 
                 Myotilin 
               
               
                 FLNC 
                 Filamin C, gamma (actin-binding protein - 280) 
               
               
                 LDB3 
                 LIM domain binding 3 
               
               
                 LAMP2 
                 Lysosomal-associated membrane protein 2 precursor 
               
               
                 VCP 
                 Valosin-containing protein 
               
               
                 CAV3 
                 Caveolin 3 
               
               
                 SEPN1 
                 Selenoprotein N1 
               
               
                 CRYAB 
                 Crystallin, alpha B 
               
               
                 DES 
                 Desmin 
               
               
                 VMA21 
                 VMA21 Vacuolar H+-ATPase Homolog ( S. Cerevisiae ) 
               
               
                 PLEC 
                 plectin 
               
               
                 PABPN1 
                 Poly(A) binding protein, nuclear 1 
               
               
                 TTN 
                 Titin 
               
               
                 RYR1 
                 Ryanodine receptor 1 (skeletal) 
               
               
                 CLN3 
                 Ceroid-lipofuscinosis, neuronal 3 (=battenin) 
               
               
                 TRIM54 
               
               
                 TRIM63 
                 Tripartite motif containing 63, E3 ubiquitin protein ligase 
               
               
                   
               
            
           
         
       
     
     Myotonic Syndromes 
       
     
       
         
           
               
               
             
               
                   
               
               
                 Gene 
                 protein 
               
               
                   
               
             
            
               
                 DMPK 
                 Myotonic dystrophy protein kinase 
               
               
                 CNPB 
                 Cellular nucleic acid-binding protein 
               
               
                 CLCN1 
                 Chloride channel 1, skeletal muscle (Thomsen disease, 
               
               
                   
                 autosomal dominant) 
               
               
                 CAV3 
                 Caveolin 3 
               
               
                 HSPG2 
                 Perlecan 
               
               
                 ATP2A1 
                 ATPase, Ca++ transporting, fast twitch 1 
               
               
                   
               
            
           
         
       
     
     Ion Channel Muscle Diseases 
       
     
       
         
           
               
               
             
               
                   
               
               
                 Gene 
                 protein 
               
               
                   
               
             
            
               
                 CLCN1 
                 Chloride channel 1, skeletal muscle (Thomsen disease, 
               
               
                   
                 autosomal dominant) 
               
               
                 SCN4A 
                 Sodium channel, voltage-gated, type IV, alpha 
               
               
                 SCN5A 
                 Voltage-gated sodium channel type V alpha 
               
               
                 CACNA1S 
                 Calcium channel, voltage-dependent, L type, alpha 1S 
               
               
                   
                 subunit 
               
               
                 CACNA1A 
                 Calcium channel, voltage-dependent, P/Q type, alpha 1A 
               
               
                   
                 subunit 
               
               
                 KCNE3 
                 Potassium voltage-gated channel, Isk-related family, 
               
               
                   
                 member 3 
               
               
                 KCNA1 
                 Potassium voltage-gated channel, shaker-related subfamily, 
               
               
                   
                 member 1 
               
               
                 KCNJ18 
                 Kir2.6 (inwardly rectifying potassium channel 2.6) 
               
               
                 KCNJ2 
                 Potassium inwardly-rectifying channel J2 
               
               
                 KCNH2 
                 Voltage-gated potassium channel, subfamily H, member 2 
               
               
                 KCNQ1 
                 Potassium voltage-gated channel, KQT-like subfamily, 
               
               
                   
                 member 1 
               
               
                 KCNE2 
                 Potassium voltage-gated channel, Isk-related family, 
               
               
                   
                 member 2 
               
               
                 KCNE1 
                 Potassium voltage-gated channel, Isk-related family, 
               
               
                   
                 member 1 
               
               
                   
               
            
           
         
       
     
     Malignant Hyperthermia 
       
     
       
         
           
               
               
             
               
                   
               
               
                 Gene 
                 protein 
               
               
                   
               
             
            
               
                 RYR1 
                 Ryanodine receptor 1 (skeletal) 
               
               
                 CACNA1S 
                 Calcium channel, voltage-dependent, L type, alpha 
               
               
                   
                 1S subunit 
               
               
                   
               
            
           
         
       
     
     Metabolic Myopathies 
       
     
       
         
           
               
               
             
               
                   
               
               
                 Gene 
                 protein 
               
               
                   
               
             
            
               
                 GAA 
                 Acid alpha-glucosidase preproprotein 
               
               
                 AGL 
                 Amylo-1,6-glucosidase, 4-alpha-glucanotransferase 
               
               
                 GBE1 
                 Glucan (1,4-alpha-), branching enzyme 1 (glycogen 
               
               
                   
                 branching enzyme, Andersen disease, glycogen storage 
               
               
                   
                 disease type IV) 
               
               
                 PYGM 
                 Glycogen phosphorylase 
               
               
                 PFKM 
                 Phosphofructokinase, muscle 
               
               
                 PHKA1 
                 Phosphorylase b kinase, alpha submit 
               
               
                 PGM1 
                 Phosphoglucomutase 1 
               
               
                 GYG1 
                 Glycogenin 1 
               
               
                 GYS1 
                 Glycogen synthase 3 glycogen synthase 1 (muscle) 
               
               
                   
                 glycogen synthase 1 (muscle) 
               
               
                 PRKAG2 
                 Protein kinase, AMP-activated, gamma 2 non-catalytic 
               
               
                   
                 subunit 
               
               
                 RBCK1 
                 RanBP-type and C3HC4-type zinc finger containing 1 
               
               
                   
                 (heme-oxidized IRP2 ubiquitin ligase 1) 
               
               
                 PGK1 
                 Phosphoglycerate kinase 1 
               
               
                 PGAM2 
                 Phosphoglycerate mutase 2 (muscle) 
               
               
                 LDHA 
                 Lactate dehydrogenase A 
               
               
                 ENO3 
                 Enolase 3, beta muscle specific 
               
               
                 CPT2 
                 Carnitine palmitoyltransferase II 
               
               
                 SLC22A5 
                 Solute carrier family 22 member 5 
               
               
                 SLC25A20 
                 Carnitine-acylcarnitine translocase 
               
               
                 ETFA 
                 Electron-transfer-flavoprotein, alpha polypeptide 
               
               
                 ETFB 
                 Electron-transfer-flavoprotein, beta polypeptide 
               
               
                 ETFDH 
                 Electron-transferring-flavoprotein dehydrogenase 
               
               
                 ACADVL 
                 Acyl-Coenzyme A dehydrogenase, very long chain 
               
               
                 ABHD5 
                 Abhydrolase domain containing 5 
               
               
                 PNPLA2 
                 Adipose triglyceride lipase (desnutrin) 
               
               
                 LPIN1 
                 Lipin 1 (phosphatidic acid phosphatase 1) 
               
               
                 PNPLA8 
                 Patatin-like phospholipase domain containing 8 
               
               
                   
               
            
           
         
       
     
     Hereditary Cardiomyopathies 
       
     
       
         
           
               
               
             
               
                   
               
               
                 Gene 
                 protein 
               
               
                   
               
             
            
               
                 MYH6 
                 Myosin heavy chain 6 
               
               
                 MYH7 
                 Myosin, heavy polypeptide 7, cardiac muscle, beta 
               
               
                 TNNT2 
                 Troponin T2, cardiac 
               
               
                 TPM1 
                 Tropomyosin 1 (alpha) 
               
               
                 MYBPC3 
                 Cardiac myosin binding protein-C 
               
               
                 PRKAG2 
                 Protein kinase, AMP-activated, gamma 2 non-catalytic 
               
               
                   
                 subunit 
               
               
                 TNNI3 
                 Troponin I, cardiac 
               
               
                 MYL3 
                 Myosin light chain 3 
               
               
                 TTN 
                 Titin 
               
               
                 MYL2 
                 Myosin light chain 2 
               
               
                 ACTC1 
                 Actin, alpha, cardiac muscle precursor 
               
               
                 CSRP3 
                 Cysteine and glycine-rich protein 3 (cardiac LIM protein) 
               
               
                 TNNC1 
                 Slow troponin C 
               
               
                 VCL 
                 Vinculin 
               
               
                 MYLK2 
                 Myosin light chain kinase 2 
               
               
                 CAV3 
                 Caveolin 3 
               
               
                 MYOZ2 
                 Myozenin 2, or calsarcin 1, a Z disk protein 
               
               
                 JPH2 
                 Junctophilin-2 
               
               
                 PLN 
                 Phospholamban 
               
               
                 NEXN 
                 Nexilin(F-actin binding protein) 
               
               
                 ANKRD1 
                 Ankyrin repeat domain 1 (cardiac muscle) 
               
               
                 ACTN2 
                 Actinin alpha2 
               
               
                 NDUFAF1 
                 NADH-ubiquinone oxidoreductase 1 alpha subcomplex 
               
               
                 TSFM 
                 Ts translation elongation factor, mitochondrial 
               
               
                 AARS2 
                 Alanyl-tRNA synthetase 2, mitochondrial 
               
               
                 MRPL3 
                 Mitochondrial ribosomal protein L3 
               
               
                 COX15 
                 COX15 homolog, cytochrome c oxidase assembly protein 
               
               
                   
                 (yeast) 
               
               
                 MTO1 
                 Mitochondrial tRNA translation optimization 1 
               
               
                 MRPL44 
                 Mitochondrial ribosomal protein L44 
               
               
                 LMNA 
                 Lamin A/C 
               
               
                 LDB3 
                 LIM domain binding 3 
               
               
                 SCN5A 
                 Voltage-gated sodium channel type V alpha 
               
               
                 DES 
                 Desmin 
               
               
                 EYA4 
                 Eyes absent 4 
               
               
                 SGCD 
                 Delta-sarcoglycan 
               
               
                 TCAP 
                 Telethonin 
               
               
                 ABCC9 
                 ATP-binding cassette, sub-family C (member 9) 
               
               
                 TMPO 
                 Lamina-associated polypeptide 2 
               
               
                 PSEN2 
                 Presenilin 2 
               
               
                 CRYAB 
                 Crystallin, alpha B 
               
               
                 FKTN 
                 Fukutin 
               
               
                 TAZ 
                 Tafazzin 
               
               
                 DMD 
                 Dystrophin 
               
               
                 LAMA4 
                 Laminin alpha 4 
               
               
                 ILK 
                 Integrin-linked kinase 
               
               
                 MYPN 
                 Myopalladin 
               
               
                 RBM20 
                 RNA binding motif protein 20 
               
               
                 SYNE1 
                 Spectrin repeat containing, nuclear envelope 1 (nesprin 1) 
               
               
                 MURC 
                 Muscle-related coiled-coil protein 
               
               
                 DOLK 
                 Dolichol kinase 
               
               
                 GATAD1 
                 GATA zinc finger domain containing 1 
               
               
                 SDHA 
                 succinate dehydrogenase complex, subunit A, 
               
               
                   
                 flavoprotein (Fp) 
               
               
                 GAA 
                 Acid alpha-glucosidase preproprotein 
               
               
                 DTNA 
                 Dystrobrevin, alpha 
               
               
                 FLNA 
                 Filamin A, alpha (actin binding protein 280) 
               
               
                 TGFB3 
                 Transforming growth factor, beta 3 
               
               
                 RYR2 
                 Ryanodine receptor 2 
               
               
                 TMEM43 
                 Transmembrane protein 43 
               
               
                 DSP 
                 Desmoplakin 
               
               
                 PKP2 
                 Plakophilin 2 
               
               
                 DSG2 
                 Desmoglein 2 
               
               
                 DSC2 
                 Desmocollin 2 
               
               
                 JUP 
                 Junction plakoglobin 
               
               
                 CASQ2 
                 Calsequestrin 2 (cardiac muscle) 
               
               
                 KCNQ1 
                 Potassium voltage-gated channel, KQT-like subfamily, 
               
               
                   
                 member 1 
               
               
                 KCNH2 
                 Voltage-gated potassium channel, subfamily H, member 2 
               
               
                 ANK2 
                 Ankyrin 2 
               
               
                 KCNE1 
                 Potassium voltage-gated channel, Isk-related family, 
               
               
                   
                 member 1 
               
               
                 KCNE2 
                 Potassium voltage-gated channel, Isk-related family, 
               
               
                   
                 member 2 
               
               
                 KCNJ2 
                 Potassium inwardly-rectifying channel J2 
               
               
                 CACNA1C 
                 Calcium channel, voltage-dependent, L type, alpha 1C 
               
               
                   
                 subunit 
               
               
                 SCN4B 
                 Sodium channel, voltage-gated, type IV, beta subunit 
               
               
                 AKAP9 
                 A kinase (PRKA) anchor protein (yotiao) 9 
               
               
                 SNTA1 
                 Syntrophin, alpha 1 
               
               
                 KCNJ5 
                 Potassium inwardly-rectifying channel, subfamily J, 
               
               
                   
                 member 5 
               
               
                 NPPA 
                 Natriuretic peptide precursor A 
               
               
                 KCNA5 
                 Potassium voltage-gated channel, shaker-related subfamily, 
               
               
                   
                 member 5 
               
               
                 GJA5 
                 Connexin 40 
               
               
                 SCN1B 
                 Sodium channel, voltage-gated, type I, beta subunit 
               
               
                 SCN2B 
                 Sodium channel, voltage-gated, type II, beta subunit 
               
               
                 NUP155 
                 Nucleoporin 155 kDa 
               
               
                 GPD1L 
                 Glycerol-3-phosphate dehydrogenase 1-like 
               
               
                 CACNB2 
                 Calcium channel, voltage-dependent, beta 2 subunit 
               
               
                 KCNE3 
                 Potassium voltage-gated channel, Isk-related family, 
               
               
                   
                 member 3 
               
               
                 SCN3B 
                 Sodium channel, voltage-gated, type III, beta subunit 
               
               
                 HCN4 
                 Hyperpolarization activated cyclic nucleotide-gated 
               
               
                   
                 potassium channel 4 
               
               
                   
               
            
           
         
       
     
     Congenital Myasthenic Syndromes 
       
     
       
         
           
               
               
             
               
                   
               
               
                 Gene 
                 protein 
               
               
                   
               
             
            
               
                 CHRNA1 
                 Cholinergic receptor, nicotinic, alpha polypeptide 1 
               
               
                 CHRNB1 
                 Cholinergic receptor, nicotinic, beta 1 muscle 
               
               
                 CHRND 
                 Cholinergic receptor, nicotinic, delta 
               
               
                 CHRNE 
                 Cholinergic receptor, nicotinic, epsilon 
               
               
                 RAPSN 
                 Rapsyn 
               
               
                 CHAT 
                 Choline acetyltransferase isoform 
               
               
                 COLQ 
                 Acetylcholinesterase collagen-like tail subunit 
               
               
                 MUSK 
                 muscle, skeletal, receptor tyrosine kinase 
               
               
                 DOK7 
                 Docking protein 7 
               
               
                 AGRN 
                 Agrin 
               
               
                 GFPT1 
                 Glutamine-fructose-6-phosphate transaminase 1 
               
               
                 DPAGT1 
                 Dolichyl-phosphate (UDP-N-acetylglucosamine) N- 
               
               
                   
                 acetylglucosaminephosphotransferase 1 (GlcNAc-1-P 
               
               
                   
                 transferase) 
               
               
                 LAMB2 
                 Laminin, beta 2 (laminin S) 
               
               
                 SCN4A 
                 Sodium channel, voltage-gated, type IV, alpha 
               
               
                 CHRNG 
                 Cholinergic receptor, nicotinic, gamma polypeptide 
               
               
                 PLEC 
                 plectin 
               
               
                 ALG2 
                 Alpha-1,3/1,6-mannosyltransferase 
               
               
                 ALG14 
                 UDP-N-acetylglucosaminyltransferase 
               
               
                 SYT2 
                 Synaptotagmin II 
               
               
                 PREPL 
                 Prolyl endopeptidase-like 
               
               
                   
               
            
           
         
       
     
     Motor Neuron Diseases 
       
     
       
         
           
               
               
             
               
                   
               
               
                 Gene 
                 protein 
               
               
                   
               
             
            
               
                 SMN1 
                 Survival of motor neuron 1, telomeric 
               
               
                 IGHMBP2 
                 Immunoglobulin mu binding protein 2 
               
               
                 PLEKHG5 
                 Pleckstrin homology domain containing, family G (with 
               
               
                   
                 RhoGef domain) member 5 
               
               
                 HSPB8 
                 Heat shock 27 kDa protein 8 
               
               
                 HSPB1 
                 Heat shock 27 kDa protein 1 
               
               
                 HSPB3 
                 Heat shock 27 kDa protein 3 
               
               
                 AARS 
                 Alanyl-tRNA synthetase 
               
               
                 GARS 
                 Glycyl-tRNA synthetase 
               
               
                 BSCL2 
                 Seipin 
               
               
                 REEP1 
                 Receptor accessory protein 1 
               
               
                 SLC5A7 
                 Solute carrier family 5 (sodium/choline cotransporter), 
               
               
                   
                 member 7 
               
               
                 DCTN1 
                 Dynactin 1 
               
               
                 UBA1 
                 Ubiquitin-activating enzyme 1 
               
               
                 ATP7A 
                 ATPase, Cu++ transporting, alpha polypeptide 
               
               
                 DNAJB2 
                 DnaJ (Hsp40) homolog, subfamily B, member 2 
               
               
                 TRPV4 
                 Transient receptor potential cation channel, subfamily V, 
               
               
                   
                 member 4 
               
               
                 DYNC1H1 
                 Dynein, cytoplasmic 1, heavy chain 1 
               
               
                 BICD2 
                 Bicaudal D homolog 2 ( Drosophila ) 
               
               
                 FBXO38 
                 F-box protein 38 
               
               
                 ASAH1 
                 N-acylsphingosine amidohydrolase (acid ceramidase) 1 
               
               
                 VAPB 
                 Vesicle-associated membrane protein-associated protein B 
               
               
                   
                 and C 
               
               
                 EXOSC8 
                 Exosome component 8 
               
               
                 SOD1 
                 Superoxide dismutase 1, soluble 
               
               
                 ALS2 
                 Alsin 
               
               
                 SETX 
                 Senataxin 
               
               
                 FUS 
                 Fusion (involved in t(12; 16) in malignant liposarcoma) 
               
               
                 ANG 
                 Angiogenin 
               
               
                 TARDBP 
                 TAR DNA binding protein 
               
               
                 FIG4 
                 Sac domain-containing inositol phosphatase 3 
               
               
                 OPTN 
                 Optineurin 
               
               
                 ATXN2 
                 Ataxin 2 
               
               
                 VCP 
                 Valosin-containing protein 
               
               
                 UBQLN2 
                 Ubiquilin 2 
               
               
                 SIGMAR1 
                 Sigma non-opioid intracellular receptor 1 
               
               
                 CHMP2B 
                 Charged multivesicular body protein 2B 
               
               
                 PFN1 
                 Profilin 1 
               
               
                 MATR3 
                 Matrin 3 
               
               
                 NEFH 
                 Neurofilament, heavy polypeptide 
               
               
                 PRPH 
                 Peripherin 
               
               
                 C9orf72 
                 Chromosome 9 open reading frame 72 
               
               
                 CHCHD10 
                 Coiled-coil-helix-coiled-coil-helix domain containing 10 
               
               
                 SQSTM1 
                 Sequestosome 1 
               
               
                 AR 
                 Androgen receptor 
               
               
                 GLE1 
                 GLE1 RNA export mediator homolog (yeast) 
               
               
                 ERBB3 
                 V-erb-b2 erythroblastic leukemia viral oncogene homolog 3 
               
               
                   
                 (avian) 
               
               
                 PIP5K1C 
                 Phosphatidylinositol-4-phosphate 5-kinase, type I, gamma 
               
               
                 EXOSC3 
                 Exosome component 3 
               
               
                 VRK1 
                 Vaccinia related kinase 1 
               
               
                 SLC52A3 
                 Solute carrier family 52, riboflavin transporter, member 3 
               
               
                 SLC52A2 
                 Solute carrier family 52, riboflavin transporter, member 2 
               
               
                 HEXB 
                 Hexosaminidase B 
               
               
                   
               
            
           
         
       
     
     Hereditary Motor and Sensory Neuropathies 
       
     
       
         
           
               
               
             
               
                   
               
               
                 Gene 
                 Protein 
               
               
                   
               
             
            
               
                 PMP22 
                 Peripheral myelin protein 22 
               
               
                 MPZ 
                 Myelin protein zero 
               
               
                 LITAF 
                 Lipopolysaccharide-induced TNF factor 
               
               
                 EGR2 
                 Early growth response 2 protein 
               
               
                 NEFL 
                 Neurofilament, light polypeptide 68 kDa 
               
               
                 HOXD10 
                 Homeobox D10 
               
               
                 ARHGEF10 
                 Rho guanine nucleotide exchange factor 10 
               
               
                 FBLN5 
                 Fibulin 5 (extra-cellular matrix) 
               
               
                 DNM2 
                 Dynamin 2 
               
               
                 YARS 
                 Tyrosyl-tRNA synthetase 
               
               
                 INF2 
                 Inverted formin 2 
               
               
                 GNB4 
                 Guanine nucleotidebinding protein, beta-4 
               
               
                 GDAP1 
                 Ganglioside-induced differentiation-associated protein 1 
               
               
                 MTMR2 
                 Myotubularin-related protein 2 
               
               
                 SBF2 
                 SET binding factor 2 
               
               
                 SBF1 
                 SET binding factor 1 
               
               
                 SH3TC2 
                 KIAA1985 protein 
               
               
                 NDRG1 
                 N-myc downstream regulated gene 1 
               
               
                 PRX 
                 Periaxin 
               
               
                 HK1 
                 Hexokinase 1 
               
               
                 FGD4 
                 Actin-filament binding protein Frabin 
               
               
                 FIG4 
                 Sac domain-containing inositol phosphatase 3 
               
               
                 SURF1 
                 surfeit 1 
               
               
                 GJB1 
                 Gap junction protein, beta 1, 32 kDa (connexin 32) 
               
               
                 AIFM1 
                 Apoptosis-inducing factor, mitochondrionassociated 1 
               
               
                 PRPS1 
                 Phosphoribosyl pyrophosphate synthetase 1 
               
               
                 PDK3 
                 Pyruvate dehydrogenase kinase, isoenzyme 3 
               
               
                 KIF1B 
                 Kinesin family member 1B 
               
               
                 MFN2 
                 Mitofusin 2 
               
               
                 RAB7A 
                 RAB7, member RAS oncogene family 
               
               
                 TRPV4 
                 Transient receptor potential cation channel, subfamily V, 
               
               
                   
                 member 4 
               
               
                 GARS 
                 Glycyl-tRNA synthetase 
               
               
                 HSPB1 
                 Heat shock 27 kDa protein 1 
               
               
                 HSPB8 
                 Heat shock 27 kDa protein 8 
               
               
                 AARS 
                 Alanyl-tRNA synthetase 
               
               
                 DYNC1H1 
                 Dynein, cytoplasmic 1, heavy chain 1 
               
               
                 LRSAM1 
                 leucine rich repeat and sterile alpha motif containing 1 
               
               
                 DHTKD1 
                 dehydrogenase E1 and transketolase domain containing 1 
               
               
                 TRIM2 
                 Tripartite motif containing 2 
               
               
                 TFG 
                 TRK-fused gene 
               
               
                 MARS 
                 methionyl-tRNA synthetase 
               
               
                 KIF5A 
                 Kinesin family member 5A 
               
               
                 LMNA 
                 Lamin A/C 
               
               
                 MED25 
                 Mediator complex subunit 25 
               
               
                 DNAJB2 
                 DnaJ (Hsp40) homolog, subfamily B, member 2 
               
               
                 HINT1 
                 Histidine triad nucleotide binding protein 1 
               
               
                 KARS 
                 Lysyl-tRNA synthetase 
               
               
                 PLEKHG5 
                 Pleckstrin homology domain containing, family G (with 
               
               
                   
                 RhoGef domain) member 5 
               
               
                 COX6A1 
                 Cytochrome c oxidase subunit VIa polypeptide 1 
               
               
                 IGHMBP2 
                 Immunoglobulin mu binding protein 2 
               
               
                 SPTLC1 
                 Serine palmitoyltransferase subunit 1 
               
               
                 SPTLC2 
                 Serine palmitoyltransferase long chain base subunit 2 
               
               
                 ATL1 
                 Atlastin GTPase 1 
               
               
                 KIF1A 
                 Kinesin family member 1A 
               
               
                 WNK1 
                 WNK lysine deficient protein kinase 1 
               
               
                 IKBKAP 
                 Inhibitor of kappa light polypeptide gene enhancer in 
               
               
                   
                 B-cells, kinase complex-associated protein 
               
               
                 NGF 
                 Nerve growth factor (beta polypeptide) 
               
               
                 DNMT1 
                 DNA (cytosine-5)-methyltransferase 1 
               
               
                 SLC12A6 
                 Potassium chloride cotransporter KCC3 
               
               
                 GJB3 
                 Gap junction protein, beta 3, 31 kDa (=connexin 31) 
               
               
                 sept-09 
                 Septin 9 
               
               
                 GAN 
                 Gigaxonin 
               
               
                 CTDP1 
                 CTD phosphatase subunit 1 
               
               
                 VRK1 
                 Vaccinia related kinase 1 
               
               
                   
               
            
           
         
       
     
     Hereditary Paraplegia 
       
     
       
         
           
               
               
             
               
                   
               
               
                 Gene 
                   
               
               
                 symbol 
                 protein 
               
               
                   
               
             
            
               
                 ATL1 
                 Atlastin 
               
               
                 SPAST 
                 Spastin 
               
               
                 NIPA1 
                 Non-imprinted in Prader-Willi/Angelman syndrome 1 
               
               
                 KIAA0196 
                 Strumpellin 
               
               
                 KIF5A 
                 Kinesin family member 5A 
               
               
                 RTN2 
                 Reticulon 2 
               
               
                 HSPD1 
                 Heat shock 60 kDa protein 1 (chaperonin) 
               
               
                 BSCL2 
                 Seipin 
               
               
                 REEP1 
                 Receptor accessory protein 1 
               
               
                 ZFYVE27 
                 Protrudin 
               
               
                 SLC33A1 
                 Solute carrier family 33 (acetyl- CoA transporter) 
               
               
                 CYP7B1 
                 Cytochrome P450, family 7, subfamily B, polypeptide 1 
               
               
                 SPG7 
                 Paraplegin 
               
               
                 SPG11 
                 Spatacsin 
               
               
                 ZFYVE26 
                 Spastizin 
               
               
                 ERLIN2 
                 ER lipid raft associated 2 
               
               
                 SPG20 
                 Spartin 
               
               
                 SPG21 
                 Maspardin 
               
               
                 B4GALNT1 
                 beta-1,4-N-acetyl-galactosaminyl transferase 1 
               
               
                 DDHD1 
                 DDHD domain containing 1 
               
               
                 KIF1A 
                 Kinesin family member 1A 
               
               
                 FA2H 
                 Fatty acid 2-hydroxylase 
               
               
                 PNPLA6 
                 Patatin-like phospholipase domain containing 6 
               
               
                 C19orf12 
                 chromosome 19 open reading frame 12 
               
               
                 GJC2 
                 gap junction protein, gamma 2, 47 kDa 
               
               
                 NT5C2 
                 5′-nucleotidase, cytosolic II 
               
               
                 GBA2 
                 glucosidase, beta (bile acid) 2 
               
               
                 AP4B1 
                 adaptor-related protein complex 4, beta 1 subunit 
               
               
                 AP5Z1 
                 Hypothetical protein LOC9907 
               
               
                 TECPR2 
                 tectonin beta-propeller repeat containing 2 
               
               
                 AP4M1 
                 Adaptor-related protein complex 4, mu 1 subunit 
               
               
                 AP4E1 
                 Adaptor-related protein complex 5, zeta 1 subunit 
               
               
                 AP4S1 
                 adaptor-related protein complex 4, sigma 1 subunit 
               
               
                 DDHD2 
                 DDHD domain containing 2 
               
               
                 C12orf65 
                 adaptor-related protein complex 4, sigma 1 subunit 
               
               
                 CYP2U1 
                 cytochrome P450, family 2, subfamily U, polypeptide 1 
               
               
                 ARL6IP1 
                 ADP-ribosylation factor-like 6 interacting protein 1 
               
               
                 AMPD2 
                 adenosine monophosphate deaminase 2 
               
               
                 ENTPD1 
                 ectonucleoside triphosphate diphosphohydrolase 1 
               
               
                 ALDH3A2 
                 Aldehyde dehydrogenase 3A2 
               
               
                 ALS2 
                 Alsin 
               
               
                 L1CAM 
                 L1 cell adhesion molecule 
               
               
                 PLP1 
                 Proteolipid protein 1 
               
               
                 MTPAP 
                 mitochondrial poly(A) polymerase 
               
               
                 AFG3L2 
                 AFG3 ATPase family gene 3-like 2 ( S. cerevisiae ) 1 
               
               
                 SACS 
                 Sacsin 
               
               
                   
               
            
           
         
       
     
     Other Neuromuscular Disorders 
       
     
       
         
           
               
               
             
               
                   
               
               
                 Gene 
                 protein 
               
               
                   
               
             
            
               
                 TOR1A 
                 Torsin A 
               
               
                 SGCE 
                 Sarcoglycan, epsilon 
               
               
                 IKBKAP 
                 Inhibitor of kappa light polypeptide gene enhancer in B- 
               
               
                   
                 cells, kinase complex-associated protein 
               
               
                 TTR 
                 Transthyretin (prealbumin, amyloidosis type I) 
               
               
                 KIF21A 
                 Kinesin family member 21A 
               
               
                 PHOX2A 
                 Paired-like aristaless homeobox protein 2A 
               
               
                 TUBB3 
                 Tubulin, beta 3 
               
               
                 TPM2 
                 Tropomyosin 2 (beta) 
               
               
                 MYH3 
                 Myosine, heavy chain 3, skeletal muscle, embryonic 
               
               
                 TNNI2 
                 Troponin I, type 2 
               
               
                 TNNT3 
                 Troponin T3, skeletal 
               
               
                 SYNE1 
                 Spectrin repeat containing, nuclear envelope 1 (nesprin 1) 
               
               
                 MYH8 
                 Myosin heavy chain, 8, skeletal muscle, perinatal 
               
               
                 POLG 
                 Polymerase (DNA directed), gamma 
               
               
                 SLC25A4 
                 Mitochondrial carrier; adenine nucleotide translocator 
               
               
                 C10orf2 
                 chromosome 10 open reading frame 2 
               
               
                 POLG2 
                 Mitochondrial DNA polymerase, accessory subunit 
               
               
                 RRM2B 
                 Ribonucleotide reductase M2 B (TP53 inducible) 
               
               
                 TK2 
                 Thymidine kinase 2, mitochondrial 
               
               
                 SUCLA2 
                 Succinate-CoA ligase, ADP-forming, beta subunit 
               
               
                 OPA1 
                 optic atrophy 1 
               
               
                 STIM1 
                 Stromal interaction molecule 1 
               
               
                 ORAI1 
                 ORAI calcium release-activated calcium modulator 1 
               
               
                 PUS1 
                 Pseudouridylate synthase 1 
               
               
                 CHCHD10 
                 Coiled-coil-helix-coiled-coil-helix domain containing 10 
               
               
                 CASQ1 
                 Calsequestrin 1 (fast-twitch, skeletal muscle) 
               
               
                 YARS2 
                 tyrosyl-tRNA synthetase 2, mitochondrial 
               
               
                   
               
            
           
         
       
     
     Any one of the above listed genes may be targeted in replacement gene therapy, wherein the gene of interest is a functional version of the deficient or mutated gene. 
     Alternatively, the above listed genes may be used as target for gene editing. Gene editing is used to correct the sequence of a mutated gene or modify the expression or regulation of a deficient/abnormal gene so that a functional gene is expressed in muscle cells. In such cases, the gene of interest is chosen from those encoding therapeutic RNAs such as interfering RNAs, guide RNAs for genome editing and antisense RNAs capable of exon skipping, wherein the therapeutic RNAs target the preceding list of genes. Tools such as CRISPR/Cas9 may be used for that purpose. 
     Thus, by gene editing or gene replacement a correct version of this gene is provided in muscle cells of affected patients, this may contribute to effective therapies against this disease. 
     In some embodiments, the target gene for gene therapy (additive gene therapy or gene editing) is a gene responsible for one of the neuromuscular diseases listed above, preferably selected from the group comprising Duchenne muscular dystrophy (DMD gene), Limb-girdle muscular dystrophies (LGMDs) (CAPN3, DYSF, FKRP, ANO5 genes and others), Spinal muscular atrophy (SMNI gene), myotubular myopathy (MTMI gene), Pompe disease (GAA gene) and Glycogen storage disease III (GSD3) (AGL gene). 
     Dystrophinopathies are a spectrum of X-linked muscle diseases caused by pathogenic variants in DMD gene, which encodes the protein dystrophin. Dystrophinopathies comprises Duchenne muscular dystrophy (DMD), Becker muscular dystrophy (BMD) and DMD-associated dilated cardiomyopathy. 
     The Limb-girdle muscular dystrophies (LGMDs) are a group of disorders that are clinically similar to DMD but occur in both sexes as a result of autosomal recessive and autosomal dominant inheritance. Limb-girdle dystrophies are caused by mutation of genes that encode sarcoglycans and other proteins associated with the muscle cell membrane, which interact with dystrophin. The term LGMD1 refers to genetic types showing dominant inheritance (autosomal dominant), whereas LGMD2 refers to types with autosomal recessive inheritance. Pathogenic variants at more than 50 loci have been reported (LGMD1A to LGMD1G; LGMD2A to LGMD2W).Calpainopathy (LGMD2A) is caused by mutation of the gene CAPN3 with more than 450 pathogenic variants described. Contributing genes to LGMD phenotype include: anoctamin 5 (ANO5), blood vessel epicardial substance (BVES), calpain 3 (CAPN3), caveolin 3 (CAV3), CDP-L-ribitol pyrophosphorylase A (CRPPA), dystroglycan 1 (DAG1), desmin (DES), DnaJ heat shock protein family (Hsp40) homolog, subfamily B, member 6 (DNAJB6), dysferlin (DYSF), fukutin related protein (FKRP), fukutin (FKT), GDP-mannose pyrophosphorylase B (GMPPB), heterogeneous nuclear ribonucleoprotein D like (HNRNPDL), LIM zinc finger domain containing 2 (LIMS2), lain A:C (LMNA), myotilin (MYOT), plectin (PLEC), protein O-glucosyltransferase 1 (PLOGLUT1), protein O-linked mannose N-acetylglucosaminyltransferase 1 (beta 1,2-) 
     (POMGNT1), protein O-mannose kinase (POMK), protein O-mannosyltransferase 1 (POMT1), protein O-mannosyltransferase 2 (POMT2), sarcoglycan alpha (SGCA), sarcoglycan beta (SGCB), sarcoglycan delta (SGCD), sarcoglycan gamma (SGCG), titin-cap (TCAP), transportin 3 (TNPO3), torsin 1A interacting protein (TOR1AIP1), trafficking protein particle complex 11 (TRAPPC11), tripartite motif containing 32 (TRIM 32) and titin (TTN). Major contributing genes to LGMD phenotype include CAPN3, DYSF, FKRP and ANO5 (Babi Ramesh Reddy Nallamilli et al., Annals of Clinical and Translational Neurology, 2018, 5, 1574-1587. 
     Spinal muscular atrophy is a genetic disorder caused by mutations in the Survival Motor Neuron 1 (SMN1) gene which is characterized by weakness and wasting (atrophy) in muscles used for movement. 
     X-linked myotubular myopathy is a genetic disorder caused by mutations in the myotubularin (MTM1) gene which affects muscles used for movement (skeletal muscles) and occurs almost exclusively in males. This condition is characterized by muscle weakness (myopathy) and decreased muscle tone (hypotonia). 
     Pompe disease is a genetic disorder caused by mutations in the acid alpha-glucosidase (GAA) gene. Mutations in the GAA gene prevent acid alpha-glucosidase from breaking down glycogen effectively, which allows this sugar to build up to toxic levels in lysosomes. This buildup damages organs and tissues throughout the body, particularly the muscles, leading to the progressive signs and symptoms of Pompe disease. 
     Glycogen storage disease III (GSD3) is an autosomal recessive metabolic disorder caused by homozygous or compound heterozygous mutation in the Amylo-Alpha-1, 6-Glucosidase, 4-Alpha-Glucanotransferase (AGL) gene which encodes the glycogen debrancher enzyme and associated with an accumulation of abnormal glycogen with short outer chains. Clinically, patients with GSD III present in infancy or early childhood with hepatomegaly, hypoglycemia, and growth retardation. Muscle weakness in those with Ma is minimal in childhood but can become more severe in adults; some patients develop cardiomyopathy. 
     In some embodiments, the pharmaceutical composition of the invention is for use for treating muscular diseases (i.e., myopathies) or muscular injuries, in particular neuromuscular genetic disorders, with no liver damage, such as for example: Muscular dystrophies, Congenital muscular dystrophies, Congenital myopathies, Distal myopathies, Other myopathies, Myotonic syndromes, Ion Channel muscle diseases, Malignant hyperthermia, Metabolic myopathies, Hereditary Cardiomyopathies, Congenital myasthenic syndromes, Motor Neuron diseases, Hereditary paraplegia, Hereditary motor and sensory neuropathies and other neuromuscular disorders. In some preferred embodiments, the pharmaceutical composition of the invention is for use for treating muscular diseases (i.e., myopathies) or muscular injuries, in particular muscular genetic disorders, with no liver damage, such as for example: Muscular dystrophies, Congenital muscular dystrophies, Congenital myopathies, Distal myopathies, Other myopathies, Myotonic syndromes, Ion Channel muscle diseases, Malignant hyperthermia, Metabolic myopathies, Hereditary Cardiomyopathies and Congenital myasthenic syndromes as defined above; more particularly Muscular dystrophies, Congenital muscular dystrophies, Congenital myopathies, Distal myopathies, Ion Channel muscle diseases, Malignant hyperthermia, Metabolic myopathies and Hereditary Cardiomyopathies as defined above. 
     Replacement or additive gene therapy may be used to treat cancer, in particular rhabdomyosarcomas. Genes of interest in cancer could regulate the cell cycle or the metabolism and migration of the tumor cells, or induce tumor cell death. For instance, inducible caspase-9 could be expressed in muscle cells to trigger cell death, preferably in combination therapy to elicit durable anti-tumor immune responses. 
     Gene editing may be used to modify gene expression in muscle cells, in the case of auto-immunity or cancer, or to perturb the cycle of viruses in such cells. In such cases, preferably, the gene of interest is chosen from those encoding guide RNA (gRNA), site-specific endonucleases (TALEN, meganucleases, zinc finger nucleases, Cas nuclease), DNA templates and RNAi components, such as shRNA and microRNA. Tools such as CRISPR/Cas9 may be used for this purpose. 
     In some embodiments, gene therapy is used for treating diseases affecting other tissues, by expression of a therapeutic gene in muscle tissue. This is useful to avoid expression of the therapeutic gene in the liver, in particular in patients having a concurrent hepatic disorder such as hepatitis. The therapeutic gene encodes preferably a therapeutic protein, peptide or antibody which is secreted from the muscle cells into the blood stream where it can be delivered to other target tissues such as for example the liver. Examples of therapeutic genes include with no limitation: Factor VIII, Factor IX and GAA genes. 
     The pharmaceutical composition of the invention which comprises AAV vector particles with reduced liver tropism may be administered to patients having concurrent liver degeneration such as fibrosis, non-alcoholic fatty liver disease, non-alcoholic steatohepatitis, viral or toxic hepatitis or underlying genetic disorders inducing liver degeneration. 
     In the context of the invention, a therapeutically effective amount refers to a dose sufficient for reversing, alleviating or inhibiting the progress of the disorder or condition to which such term applies, or reversing, alleviating or inhibiting the progress of one or more symptoms of the disorder or condition to which such term applies. 
     The effective dose is determined and adjusted depending on factors such as the composition used, the route of administration, the physical characteristics of the individual under consideration such as sex, age and weight, concurrent medication, and other factors, that those skilled in the medical arts will recognize. 
     In the various embodiments of the present invention, the pharmaceutical composition comprises a pharmaceutically acceptable carrier and/or vehicle. 
     A “pharmaceutically acceptable carrier” refers to a vehicle that does not produce an adverse, allergic or other untoward reaction when administered to a mammal, especially a human, as appropriate. A pharmaceutically acceptable carrier or excipient refers to a non-toxic solid, semi-solid or liquid filler, diluent, encapsulating material or formulation auxiliary of any type. 
     Preferably, the pharmaceutical composition contains vehicles, which are pharmaceutically acceptable for a formulation capable of being injected. These may be in particular isotonic, sterile, saline solutions (monosodium or disodium phosphate, sodium, potassium, calcium or magnesium chloride and the like or mixtures of such salts), or dry, especially freeze-dried compositions which upon addition, depending on the case, of sterilized water or physiological saline, permit the constitution of injectable solutions. 
     The pharmaceutical forms suitable for injectable use include sterile aqueous solutions or suspensions. The solution or suspension may comprise additives which are compatible with viral vectors and do not prevent viral vector particle entry into target cells. In all cases, the form must be sterile and must be fluid to the extent that easy syringe ability exists. It must be stable under the conditions of manufacture and storage and must be preserved against the contaminating action of microorganisms, such as bacteria and fungi. An example of an appropriate solution is a buffer, such as phosphate buffered saline (PBS) or Ringer lactate. 
     The invention provides also a method for treating a disease affecting muscle tissue in particular skeletal muscle tissue and/or cardiac tissue, comprising: administering to a patient a therapeutically effective amount of the pharmaceutical composition as described above. 
     The invention provides also a method for treating a disease by expression of a therapeutic gene in muscle tissue, comprising: administering to a patient a therapeutically effective amount of the pharmaceutical composition as described above. 
     As used herein, the term “patient” or “individual” denotes a mammal. Preferably, a patient or individual according to the invention is a human. 
     In the context of the invention, the term “treating” or “treatment”, as used herein, means reversing, alleviating or inhibiting the progress of the disorder or condition to which such term applies, or reversing, alleviating or inhibiting the progress of one or more symptoms of the disorder or condition to which such term applies. 
     The pharmaceutical composition of the present invention, is generally administered according to known procedures, at dosages and for periods of time effective to induce a therapeutic effect in the patient. 
     The administration may be parenteral, oral, local, or loco-regional. The parenteral administration is advantageously by injection or perfusion, such as e subcutaneous (SC), intramuscular (IM), intravascular such as intravenous (IV), intraperitoneal (IP), intradermal (ID) or else. Preferably, the administration produces a systemic effect in the whole body, i.e., all the muscles of the patient, including the diaphragm and the heart. Preferably, the administration is systemic, more preferably parenteral. 
     The practice of the present invention will employ, unless otherwise indicated, conventional techniques, which are within the skill of the art. Such techniques are explained fully in the literature. 
     The invention will now be exemplified with the following examples, which are not limitative, with reference to the attached drawings in which: 
    
    
     
       FIGURE LEGENDS 
         FIG. 1 : Design of peptide insertion in the hybrid AAV serotype between AAV9 and AAVrh74. 
       A. Schematic representation of the Cap genes (VP1) of AAV9, AAVrh74 and the AAV9-rh74 hybrid capsid highlighting the sequences of the variable region. B. The VP1 protein of the AAV-MT capsid derives from the insertion of 15 amino acid (15-mer) between amino acid 586 and 593 of the VP1 of the AAV9-rh74 capsid. 
         FIG. 2 : Comparative biodistribution of AAV9-RH74 and AAV-MT in GSDIII mice. GSDIII mice were tail vein injected with 2×10 12  vector genome/mouse of the indicated AAV vectors. Three months after injection, tissues were collected. PBS-injected mice were used as controls. (A-F) Vector genome copy number per cell (VGCN/cell) measured in liver (A); heart (B); diaphragm (C); quadriceps (D); extensor digitorum longus (EDL) (E) and gastrocnemius (F). Data were presented as mean ±standard deviation. Statistical analyses were performed by ANOVA (*=p&lt;0.05 vs. PBS; §=p&lt;0.05 vs. ALL; n=4-5 mice per group). 
     
    
    
     EXAMPLE 
     Peptide-Modified Hybrid AAV9-rh74 Serotype Vector 
     1. Material and Methods 
     Plasmid Construction for New Serotypes 
     To construct the plasmid containing AAV2 Rep sequence and Hybrid Cap 9-rh74, a fragment of 1029 nt, containing the highly variable part of AAV-rh74 Cap flanked with AAV9 Cap sequence fragments and restriction sites BsiWI in 5′ and Eco47III in 3′, was synthesized (GENEWIZ). This fragment was then inserted using the mentioned restriction sites in the plasmid pAAV2-9, which contains AAV2 Rep and AAV9 Cap, to replace the AAV9 Cap corresponding sequence. Peptide engraftment was performed by replacing the QQNAAP hexapeptide (SEQ ID NO: 11) in the AAV9-rh74 capsid with the GQSGRGDLGLSAQAA (SEQ ID NO: 13) amino acid sequence. 
     AAV Production 
     HEK293T cells were grown in suspension in 250 mL of serum-free medium. The cells were transfected with 3 plasmids: i) a transgene plasmid, containing AAV2 ITRs flanking an expression cassette ii) the helper plasmid pXX6, containing adenoviral sequences necessary for AAV production, and iii) a plasmid containing AAV Rep and Cap genes, defining the serotype of AAV. Two days after transfection, the cells were lysed to release the AAV particles. 
     The viral lysate was purified by affinity chromatography. Viral genomes were quantified by a TaqMan real-time PCR assay using primers and probes corresponding to the ITRs of the AAV vector genome (Rohr et al., J. Virol. Methods, 2002, 106, 81-88). 
     In Vivo Studies 
     All mouse studies were performed according to the French and European legislation on animal care and experimentation (2010/63/EU) and approved by the local institutional ethical committee (protocol no. 2016-002C). AAV vectors were administered intravenously via the tail vein to three month-old male GDE knockout mice. PBS-injected littermates were used as controls. Three months after vector injections, tissues were harvested and homogenized in DNAse/RNAse free water using Fastprep tubes (6.5 m/s; 60 seconds). 
     Vector Genome Copy Number (VGCN) Quantification 
     For vector genome copy number (VGCN) quantification in samples, DNA was extracted from samples using MagNA Pure 96 Instrument (Roche). Real-time PCR was performed on 1μL of DNA, using the protocol for AAV vectors titration described above. Exon Mex5 of titin gene was used as genomic DNA loading control. 
     2. Results 
     The AAV9-rh74 hybrid capsid was engineered by inserting a peptide in the common region between VP1, VP2 and VP3 between Q at positions 586 and I at position 593 of SEQ ID NO: 3 ( FIG. 1 ). AAV capsid modification was performed according to Kienle EC (Dissertation for the degree of Doctor of natural Sciences, Combined Faculties for the Natural Sciences and for Mathematics of the Ruperto-Carola University of Heidelberg, Germany, 2014) using a peptide as disclosed in Michelfelder et al. (PLoS ONE, 2009, 4, e5122). Briefly, the hexapeptide QQNAAP (SEQ ID NO: 11) present in the common region between VP1, VP2 and VP3 of the hybrid Cap9-rh74 (positions 587 to 592 of SEQ ID NO: 3) is mutated to the octapeptide GQSGAQAA (SEQ ID NO: 12) and peptide P1 (RGDLGLS; SEQ ID NO: 8) is inserted between glycine at position 4 and alanine at position 5. The peptide-modified hybrid Cap9-rh74 having the insertion of the peptide P1 has the amino acid sequence SEQ ID NO: 5 and the corresponding coding sequence is SEQ ID NO: 6. Vectors are produced by triple transfection in HEK293 cells grown in suspension and purified by affinity chromatography as described in example 1. The resulting AAV Vector called AAV-Muscle Transducer (AAV-MT) was injected in GDE knockout mice, an animal model of GSDIII, in parallel with AAV9-rh74 at the dose of 2×10 12  vg/mouse. Tissues were obtained three months post-injection and vector genome copy number (VGCN) was measured by quantitative PCR to assess tissue targeting ( FIG. 2 ). In the liver, injection of AAV9-rh74 resulted in significantly increased vector genome copies compared to PBS injected mice (p&lt;0.05;  FIG. 2A ) while liver transduced with AAV-MT did not showed a significant increase in VGCN compared to the PBS group. In general, AAV-MT showed better transduction efficacy in muscles when compared to the parental capsid AAV9-rh74 ( FIG. 2B-F ) Importantly, in diaphragm, quadriceps and gastrocnemius, the AAV-MT outperformed the parental capsid (p&lt;0.05 vs. AAV9-rh74 group). Finally, in heart and extensor digitorum longus (EDL) a significantly higher VGCN were measured after injection with the AAV-MT capsid but not with the AAV9-rh74 when compared with PBS injected mice. These data indicate that the AAV-MT capsid outperforms the AAV9-rh74 in muscle transduction and maintains or even ameliorates the pronounced liver detargeting observed for the parental capsid.