Therapeutic tissue-targeting extracellular vesicles loaded with biomolecules

Abstract

Described herein are compositions of therapeutic tissue-targeting extracellular vesicles, and methods and systems of producing the therapeutic tissue -targeting extracellular vesicles. Also described herein are methods of treating a disease or condition with the therapeutic tissue-targeting extracellular vesicles.

Description

WSGR Ref. No. 54841-719.601THERAPEUTIC TISSUE-TARGETING EXTRACELLULAR VESICLES LOADED WITH BIOMOLECULESCROSS-REFERENCE

[0001] This application claims the benefit of U.S. Provisional Application No 63 / 756,000, filed on February 7, 2025, which is incorporated herein by reference in its entirety.BACKGROUND

[0002] DMD can be caused by mutations in the gene encoding the dystrophin protein, which can render DMD patients unable to synthesize normal dystrophin mRNA and / or protein that is expressed in cells throughout the body, including in the heart, skeletal muscles, smooth muscles, retina, and brain.

[0003] Exon deletions and duplications can be DMD-associated mutations in the dystrophin gene, which can account for about 79% of mutations in large DNA segments spanning > 1 exons. Such deletions and duplications can be located in exons 45-55 and 3-9 of the dystrophin gene. Other types of small mutations (<1 exon) can account for approximately 21%, of which point mutations can account for about 11%, small deletions can account for about 5%, small insertions can account for about 2%, and splice site mutations can account for about 3% of total frequency of dystrophin mutations. The severity of the DMD phenotype can be affected by whether mutations in the dystrophin gene have disrupted the open reading frame or cause premature truncation of the dystrophin. Out- frame mutations can lead to transcription and translation disorders of the dystrophin gene, which can result in a more severe DMD phenotype. In -frame mutations, while retaining an open reading frame, can transcribe and translate into truncated, partially functional dystrophin proteins, which can result in milder phenotypes, such as Becker muscular dystrophy (BMD).

[0004] Other gene therapies for DMD have been developed, which can be designed to reduce the sensitivity of ribosomes to early termination codons, thereby allowing mRNA to continue translation without stopping at the termination codon during translation; by adeno-associated viral integration of partial dystrophin DNA into the genome; or by “exon skipping” to direct RNA splicing using antisense oligonucleotides (ASOs) for evading incorporation of mutated or misaligned exons within DMD mRNA, resulting in a truncated but partially functional dystrophin protein. Such therapeutics can be met withWSGR Ref. No. 54841-719.601challenges of reduced bioavailability because they can have a low affinity for proteins and can be quickly cleared in the body after systemic injection. It can be more difficult for these drugs to enter the cell, and thus administration of larger drug doses or high frequency of administration can be necessary for improving bioavailability. Moreover, AAV vectors can have packaging size limits, which can be too small to accommodate the full-length dystrophin DNA sequence. Exon skipping can be patient-precise and unique ASOs can be designed based on the identified location of the patient’s dystrophin mutation.

[0005] The hDMD-mRNA-tEXOs described herein can be advantageous in that they can be natural vesicles that contain full-length dystrophin mRNA and can be secreted by all cells in the human body with low immunogenicity and high biocompatibility. Here, hDMD- mRNA-tEXOs can circumvent the complications associated with viral vectors and do not require development of patient-precise molecules, like ASOs, because the hDMD- mRNA-tEXOs can supplement a patient with normal, full-length dystrophin mRNA. The hDMD-mRNA-tEXOs can have the added benefit of delivering mRNA, which can stay in the cytosol of targeted muscle cells without entering the nucleus, which can eliminate the risk of foreign gene integration that can plague AAV-mediated gene delivery. The hDMD-mRNA-tEXOs can further express molecular muscle tissue -targeting polypeptides that can target and generate full-length dystrophin in muscle cells following administration to a subject.SUMMARY

[0006] Described and provided herein is an extracellular vesicle comprising: (i) an exogenous nucleic acid molecule encoding a dystrophin protein that comprises an amino acid sequence having at least 70% sequence identity to the amino acid sequence set forth in any one of SEQ ID NOs: 1-6; and (ii) a targeting polypeptide configured to bind to a targeted cell. In some embodiments, the nucleic acid comprises a mRNA. In some embodiments, the dystrophin protein comprises an amino acid sequence having at least 70% sequence identity to the amino acid sequence set forth in SEQ ID NO: 1. In some embodiments, the dystrophin protein is a full-length dystrophin protein. In some embodiments, the dystrophin protein comprises the amino acid sequence set forth in SEQ ID NO: 1. In some embodiments, the dystrophin protein consists of the amino acid sequence set forth in SEQ ID NO: 1. In some embodiments, the nucleic acid comprises a full-length human dystrophin mRNA. In some embodiments, the targeting polypeptide comprises a targeting domain configured to bind to a surface protein of the targeted cell.WSGR Ref. No. 54841-719.601In some embodiments, the targeting domain comprises a muscle cell -targeting polypeptide. In some embodiments, the muscle cell-targeting polypeptide is a skeletal muscle-targeting polypeptide, a cardiac muscle -targeting polypeptide, or a smooth muscle-targeting polypeptide. In some embodiments, the muscle cell-targeting polypeptide is the skeletal muscle -targeting polypeptide. In some embodiments, the targeting domain is a M12 peptide, an RGD motif, a dystrophin -binding peptide, a T7 peptide, a T9 peptide, a ASSLNIA peptide, a cardiomyocyte -specific peptide (CMP), an ischemic myocardium-targeting polypeptide (IMTP), a cardiac homing peptide (CHP), or a CRPPR peptide. In some embodiments, the targeting domain comprises the M12 peptide, the RGD motif, the dystrophin -binding peptide, the T7 peptide, the T9 peptide, an GPGRGDQTTL peptide or the ASSLNIA peptide. In some embodiments, the targeting domain comprises the M12 peptide. In some embodiments, the targeting domain comprises the RGD motif. In some embodiments, the targeting domain comprises the T7 peptide. In some embodiments, the T7 peptide comprises an amino acid sequence having no more than 4 amino acid substitutions with reference to the amino acid sequence set forth in SEQ ID NO: 42. In some embodiments, the T7 peptide comprises the amino acid sequence set forthin SEQ ID NO: 42. In some embodiments, the T7 peptide consists of the amino acid sequence set forth in SEQ ID NO: 42. In some embodiments, the targeting domain comprises the T9 peptide. In some embodiments, the T9 peptide comprises an amino acid sequence having no more than 4 amino acid substitutions with reference to the amino acid sequence set forth in SEQ ID NO: 44. In some embodiments, the T9 peptide comprises the amino acid sequence set forth in SEQ ID NO: 44. In some embodiments, the T9 peptide consists of the amino acid sequence set forth in SEQ ID NO: 44. In some embodiments, the targeting domain is coupled to an adapter polypeptide. In some embodiments, the adapter polypeptide comprises a SIRPa domain. In some embodiments, the adapter polypeptide comprises an extracellular vesicle surface protein. In some embodiments, the extracellular vesicle surface protein comprises CD47, CD63, CD81, CD82, CD315, a hetero trimeric G protein, MHC class I, an integrin, a transferrin receptor (TFR2), a LAMP1 / 2, a heparan sulfate proteoglycan, EMMPRIN, ADAM10, a GPI-anchored 5 'nucleotidase, CD73, CD55, CD59, sonic hedgehog (SHH), TSPAN8, CD37, CD53, CD9, PECAM1, ERBB2, EPCAM, CD90, CD45, CD41, CD42a, Glycophorin A, CD 14, MHC class II, CD3, Acetylcholinesterase / AChE-S, AChE-E, amyloid beta A4 / APP, PTGFRN, or multidrug resistance-associated protein. In some embodiments, the adapter polypeptide comprises the CD47. In some embodiments, the CD47 comprises aWSGR Ref. No. 54841-719.601human CD47. In some embodiments, the human CD47 comprises an amino acid sequence comprising at least 70% sequence identity to the amino acid sequence set forth in SEQ ID NO: 41. In some embodiments, the human CD47 comprises an amino acid sequence that comprises the amino acid sequence set forth in SEQ ID NO: 41. In some embodiments, the human CD47 consists of the amino acid sequence set forth in SEQ ID NO: 41. In some embodiments, the adapter polypeptide is non-covalently coupled to the targeting domain. In some embodiments, the adapter polypeptide is covalently coupled to the targeting domain. In some embodiments, the targeting domain is covalently coupled to an N-terminus of the adapter polypeptide. In some embodiments, the targeting polypeptide comprises a T7 peptide covalently coupled to a human CD47 polypeptide. In some embodiments, the targeting polypeptide comprises a T9 peptide covalently coupled to a human CD47 polypeptide. In some embodiments, the targeting polypeptide comprises an amino acid sequence having at least 70% sequence identity to the amino acid sequence set forth in SEQ ID NO: 43 or SEQ ID NO: 45. In some embodiments, the targeting polypeptide comprises the amino acid sequence set forth in SEQ ID NO: 43 or SEQ ID NO: 45. In some embodiments, the targeting polypeptide consists of the amino acid sequence set forth in SEQ ID NO: 43 or SEQ ID NO: 45. In some embodiments, the extracellular vesicle further comprises a second targeting polypeptide, wherein the targeting polypeptide consists of the sequence set forth in SEQ ID NO: 43 and the second targeting polypeptide consists of the sequence set forth in SEQ ID NO: 45. In some embodiments, the mRNA comprises a sequence encoded by the sequence set forth in SEQ ID NO: 31. In some embodiments, the extracellular vesicle comprises a diameter of greater than 20 nm. In some embodiments, the extracellular vesicle comprises a size distribution with a peak at 50 nm to 200 nm in diameter. In some embodiments, the extracellular vesicle expresses a marker of naturally occurring extracellular vesicles, wherein the marker comprises CD9, CD63, TSG101, or ARF6. In some embodiments, the extracellular vesicle comprises an exosome, microvesicle, or an apop to tic body. In some embodiments, the extracellular vesicle comprises the exosome. In some embodiments, the extracellular vesicle comprises a plurality of extracellular vesicles. In some embodiments, the extracellular vesicle comprises at least 1x106copies of the nucleic acid molecule. In some embodiments, the extracellular vesicle comprises from about 2x106to about 5x1012copies of the nucleic acid molecule. In some embodiments, the targeted cell is an adherent cell, a multinucleated cell, a mononucleated cell, a contractile cell, a muscle cell,WSGR Ref. No. 54841-719.601a skeletal muscle cell, a cardiomyocyte, a smooth muscle cell, a myofibroblast. In some embodiments, the targeted cell is the skeletal muscle cell.

[0007] Also provided herein is a composition comprising: (i) a first extracellular vesicle comprising a first targeting domain; and (ii) a second extracellular vesicle comprising a second targeting domain, wherein the first extracellular vesicle and the second extracellular vesicle comprise exogenous messenger RNA (mRNA) encoding a dystrophin protein. In some embodiments, the first targeting domain comprises a first muscle-targeting peptide and the second targeting domain comprises a second muscle - targeting peptide. In some embodiments, the first muscle -targeting domain is a T7 targeting domain; and the second muscle -targeting domain is a T9 targeting domain. In some embodiments, the T7 targeting domain comprises an amino acid sequence having no more than 4 amino acid substitutions with reference to the amino acid sequence set forth in SEQ ID NO: 42, and wherein the T9 targeting domain comprises an amino acid sequence having no more than 4 amino acid substitutions with reference to the amino acid sequence set forth in SEQ ID NO: 44. In some embodiments, the T7 targeting domain comprises the amino acid sequence set forth in SEQ ID NO: 42; and wherein the T9 targeting domain comprises the amino acid sequence set forth in SEQ ID NO: 44. In some embodiments, the T7 targeting domain consists of the amino acid sequence set forth in SEQ ID NO: 42; and wherein the T9 targeting domain consists of the amino acid sequence set forth in SEQ ID NO: 44. In some embodiments, the first targeting domain is coupled to a first adapter polypeptide, and the second targeting domain is coupled to a second adapter polypeptide. In some embodiments, the first adapter polypeptide and the second adapter polypeptide comprise a CD47 polypeptide. In some embodiments, the CD47 polypeptide comprises a human CD47 polypeptide. In some embodiments, the human CD47 polypeptide comprises an amino acid sequence comprising at least 70% sequence identity to the amino acid sequence set forth in SEQ ID NO: 41. In some embodiments, the human CD47 polypeptide comprises the amino acid sequence set forth in SEQ ID NO: 41. In some embodiments, the human CD47 polypeptide consists of the amino acid sequence set forth in SEQ ID NO: 41. In some embodiments, the first extracellular vesicle comprises a first targeting polypeptide, wherein the first targeting polypeptide comprises the first targeting domain and a first adapter polypeptide, and wherein the first targeting polypeptide comprises a sequence comprising at least 70% sequence identity to the amino acid sequence set forth in SEQ ID NO: 43. In some embodiments, the first targeting polypeptide comprises an amino acid sequenceWSGR Ref. No. 54841-719.601consisting of the amino acid sequence set forth in SEQ ID NO: 43. In some embodiments, the second extracellular vesicle comprises a second targeting polypeptide, wherein the second targeting polypeptide comprises the second targeting domain and a second ad apter polypeptide, wherein the second targeting polypeptide comprises an amino acid sequence that has at least 70% sequence identity to the amino acid sequence set forth in SEQ ID NO: 45. In some embodiments, the second targeting polypeptide comprises an amino acid sequence consisting of the amino acid sequence set forth in SEQ ID NO: 45. In some embodiments, the first extracellular vesicle and the second extracellular vesicle comprise an exogenous nucleic acid molecule encoding a dystrophin protein. In some embodiments, the dystrophin protein comprises an amino acid sequence having at least 70% sequence identity to the amino acid sequence set forth in any one of SEQ ID NOs: 1 -6. In some embodiments, the dystrophin protein is a full-length dystrophin protein. In some embodiments, the dystrophin protein comprises the amino acid sequence set forth in any one of SEQ ID NOs: 1-6. In some embodiments, the dystrophin protein consists of the amino acid sequence set forth in any one of SEQ ID NOs: 1 -6. In some embodiments, the dystrophin protein consists of the amino acid sequence set forth in SEQ ID NO: 1. In some embodiments, the exogenous nucleic acid molecule comprises an mRNA, wherein the mRNA comprises a nucleic acid sequence encoded by the sequence set forth in SEQ ID NO: 31. In some embodiments, the first muscle -targeting domain the second muscletargeting domain are configured to deliver the first extracellular vesicle and the second extracellular vesicle to a first target cell and a second target cell, respectively. In some embodiments, the first target cell and the second target cell are the same cell. In some embodiments, the first target cell and the second target cell are different cells. In some embodiments, the first extracellular vesicle and the second extracellular vesicle comprise an exosome, microvesicle, or an apoptotic body. In some embodiments, the first extracellular vesicle and the second extracellular vesicle comprise the exosome. In some embodiments, the composition further comprises a third extracellular vesicle, wherein the third extracellular vesicle comprises a third targeting polypeptide consisting of the amino acid sequence of SEQ ID NO: 43; and a fourth targeting polypeptide consisting of the amino acid sequence of SEQ ID NO: 45, and a second exogenous nucleic acid molecule encoding a dystrophin protein consisting of the amino acid sequence set forth in SEQ ID NO: 1. In some embodiments, the second exogenous nucleic acid molecule comprises an mRNA, and wherein the mRNA comprises a sequence encoded by the sequence set forth in SEQ ID NO: 31. In some embodiments, the composition comprises at least 1x103WSGR Ref. No. 54841-719.601extracellular vesicles. In some embodiments, the composition comprises at least 1x106copies of the exogenous nucleic acid molecule. In some embodiments, the composition comprises from about 2x106to about 5x1012copies of the exogenous nucleic acid molecule. In some embodiments, the composition is formulated for intravenous injection, systemic injection, parenteral injection, intracardiac, or intramuscular injection.

[0008] In another aspect, disclosed herein is an extracellular vesicle comprising: a heterologous adapter polypeptide; and an exogenous nucleic acid molecule encoding a dystrophin protein that comprises an amino acid sequence having at least 70% sequence identity to the amino acid sequence set forth in any one of SEQ ID NOs: 1 -6. In some embodiments, the heterologous adapter polypeptide comprises an extracellular vesicle surface protein. In some embodiments, the extracellular vesicle surface protein comprises CD47 polypeptide. In some embodiments, the CD47 polypeptide comprises a human CD47 polypeptide. In some embodiments, the human CD47 polypeptide comprises an amino acid sequence comprising at least 70% sequence identity to the amino acid sequence set forth in SEQ ID NO: 41. In some embodiments, the human CD47 polypeptide comprises an amino sequence consisting of the amino acid sequence set forth in SEQ ID NO: 41. In some embodiments, the dystrophin protein comprises an amino acid sequence having at least 70% sequence identity to the amino acid sequence set forth in SEQ ID NO: 1. In some embodiments, the dystrophin protein comprises the amino acid sequence set forth SEQ ID NO: 1. In some embodiments, the dystrophin protein comprises an amino acid sequence consisting of the amino acid sequence set forth in SEQ ID NO: 1. In some embodiments, the nucleic acid molecule comprises an exogenous messenger RNA (mRNA). In some embodiments, the extracellular vesicle comprises an exosome, microvesicle, or an apoptotic body. In some embodiments, the extracellular vesicle comprises the exosome. In some embodiments, the extracellular vesicle comprises at least 1x103extracellular vesicles. In some embodiments, the extracellular vesicle does not comprise a heterologous targeting domain.

[0009] In yet another aspect, the disclosure also describes a pharmaceutical composition comprising an extracellular vesicle provided herein, and a pharmaceutically acceptable excipient. In some embodiments, the exogenous nucleic acid molecule is present in the pharmaceutical composition at a concentration of at least 1×107copies / mL. In some embodiments, the exogenous nucleic acid molecule is present in the pharmaceutical composition at a concentration of from about 1×107copies / mL to about 1×1014copies / mL. In some embodiments, the exogenous nucleic acid molecule is present in theWSGR Ref. No. 54841-719.601pharmaceutical composition at a concentration of from about 1×108copies / mL to about 1×1012copies / mL. In some embodiments, the exogenous nucleic acid molecule is present in the pharmaceutical composition at a concentration of from about 1×109copies / mL to about 1×1010copies / mL. In some embodiments, the exogenous nucleic acid molecule is present in the pharmaceutical composition at a concentration is about 1×109copies / mL.

[0010] Also described herein is a pharmaceutical composition comprising a composition of the disclosure and a pharmaceutically acceptable excipient. In some embodiments, the pharmaceutical composition is a liquid solution. In some embodiments, the pharmaceutically acceptable excipient comprises a saline solution or a buffered saline solution. In some embodiments, the pharmaceutical composition comprises a pH of at least 5.5 when measured at room temperature. In some embodiments, the pH of the pharmaceutical composition is from about 6.5 to about 8.5 when measured at room temperature. In some embodiments, the pH of the pharmaceutical composition is about 7.7 when measured at room temperature. In some embodiments, the pharmaceutical composition further comprises human serum albumin. In some embodiments, the human serum albumin is present at a concentration at least 0.1% w / v. In some embodiments, the human serum albumin is present at a concentration of from about 0.1% w / v to about 2% w / v. In some embodiments, the human serum albumin is present at a concentration of about 0.2% w / v. In some embodiments, the human serum albumin is present at a concentration of about 1% w / v. In some embodiments, the pharmaceutical composition further comprises a cryoprotection medium. In some embodiments, the cryoprotection medium is CryoStor® CS10, glycerol, ethylene glycol, propylene glycol, trehalose, sucrose, Dimethyl sulfoxide (DMSO), polyvinylpyrrolidone (PVP), and hydroxyethyl starch (HES), mFreSR™, NB-KUL 10, STEM-CELLBANKER, CryoSOfree™, PRIME - XVFreezIS, Optibumin® 25, orXT-Thrive®. In some embodiments, the cryoprotection medium is present in the pharmaceutical composition at a concentration of at least 2% v / v. In some embodiments, the cryoprotection medium is present in the pharmaceutical composition at a concentration of from about 2% v / v to about 30% v / v. In some embodiments, the cryoprotection medium is present in the pharmaceutical composition at a concentration of from about 3% v / v to about 10% v / v. In some embodiments, the cryoprotection medium is present in the pharmaceutical composition at a concentration of from about 3% v / v to about 7% v / v. In some embodiments, the cryoprotection medium is present in the pharmaceutical composition at a concentration of from about 4% v / v toWSGR Ref. No. 54841-719.601about 6% v / v. In some embodiments, the cryoprotection medium is present in the pharmaceutical composition at a concentration is about 5% v / v. In some embodiments, the cryoprotection medium is present in the pharmaceutical composition at a concentration of from about 10% v / v to about 30% v / v. In some embodiments, the cryoprotection medium is present in the pharmaceutical composition at a concentration of from about 20% v / v to about 30% v / v. In some embodiments, the cryoprotection medium is present in the pharmaceutical composition at a concentration is about 25% v / v. In some embodiments, the pharmaceutical composition is formulated for injection via a systemic route, an intravenous route, a parenteral route, an intramuscular route, a subcutaneous route, or an intraperitoneal route, intracardiac route. In some embodiments, the pharmaceutical composition is formulated for injection via a coronary artery catheter.

[0011] In one aspect, described herein is a catheter comprising the extracellular vesicle. In some embodiments, the catheter comprises a heart catheter.

[0012] In another aspect, described herein is a syringe comprising the extracellular vesicle.

[0013] In yet another aspect, this disclosure also describes a kit, comprising the (a) an extracellular vesicle, a composition, or a pharmaceutical composition described herein; (b) a catheter or a syringe; and (c) instructions.

[0014] Further provided herein is a method of increasing an expression level of human dystrophin in a subject, the method comprising: administering to the subject the extracellular vesicle, the composition, or the pharmaceutical composition described throughout the instant disclosure, wherein the administering results in an elevated expression level of the human dystrophin in the human subject. In some embodiments, the administering comprises administering to a cardiac tissue of the subject. In some embodiments, the cardiac tissue is a pericardium, an artery, a heart ventricle, a heart atrium, or a myocardium of the subject. In some embodiments, the administering comprises administering into the cardiac tissue of the subject via an injection. In some embodiments, the injection comprises injections into the cardiac tissue of the subject via an intracardiac injection, an intra-arterial injection, an intrapericardial injection, or an intramyocardial injection. In some embodiments, the administering comprises administering into the cardiac tissue of the subject via a catheter. In some embodiments, the catheter comprises a balloon catheter, a microvalve catheter, a guide catheter, or a stent. In some embodiments, the administering comprises administering to the subject via an intravenous route, a parenteral route, an intramuscular route. In some embodiments, the method does not comprise administering to the subject an immunosuppressive agent.WSGR Ref. No. 54841-719.601In some embodiments, the method comprises administering to the subject an immunosuppressive agent. In some embodiments, the immunosuppressive agent comprises a mTOR inhibitor. In some embodiments, the mTOR inhibitor comprises sirolimus. In some embodiments, the immunosuppressive agent comprises a calcineurin inhibitor. In some embodiments, the calcineurin inhibitor comprises tacrolimus or cyclosporine. In some embodiments, the administering treats a disorder of the subject. In some embodiments, the disorder comprises a muscle disorder. In some embodiments, the muscle disorder comprises a muscular dystrophy. In some embodiments, the muscular dystrophy comprises Duchenne muscular dystrophy or Becker muscular dystrophy. In some embodiments, the disorder comprises the Duchenne muscular dystrophy. In some embodiments, the subject is a human subject. In some embodiments, the method results in at least a 2-fold increase in the expression level of the human dystrophin protein in the subject compared to an expression level of human dystrophin protein in the subject prior to the administering. In some embodiments, the method results in at least a 2 -fold increase in a level of human dystrophin mRNA in the subject compared to an expression level of human dystrophin mRNA in the subject prior to the administering. In some embodiments, the method results in at least a 2 -fold increase in a level of full-length human dystrophin mRNA in the subject compared to an expression level of human dystrophin mRNA in the subject prior to the administering. In some embodiments, the method results in at least a 2-fold increase in a level of full-length human dystrophin protein in the subject compared to an expression level of full-length human dystrophin protein in the subject prior to administration of the extracellular vesicle, the extracellular vesicle, or the pharmaceutical composition. In some embodiments, the method results in an increase in the level of human dystrophin or the level of full-length human dystrophin in a muscle tissue of the subject. In some embodiments, the muscle tissue is a skeletal muscle of the subject. In some embodiments, the skeletal muscle tissue is a diaphragm, a quadricep, a bicep, a tricep, a gastrocnemius, a tibialis, a fibularis, a deltoid, a gluteus maximus, or an abdominal muscle of the subject. In some embodiments, the method results in a reduction of a level ofN-terminalpro-B-type natriuretic peptide (NT-proBNP) in the subject compared to the level of NT-proBNP in the subject prior to the administering. In some embodiments, the reduction of the level comprises at least a 20% reduction in the level of NT-proBNP in the subject compared to the level of NT-proBNP in the subject prior to the administering. In some embodiments, the level is of a blood sample. In some embodiments, the method results in a reduction of a level of CreatineWSGR Ref. No. 54841-719.601Kinase (CK) in the subject compared to the level of CK in the subject prior to the administering. In some embodiments, the method results in at least a 10% reduction in the level of CK in the subject compared to the level of CK in the subject prior to the administering. In some embodiments, the method results in at least a 20% reduction in the level of CK in a blood sample obtained from the subject compared to the level of CK in the subject prior to administration of the extracellular vesicle, the extracellular vesicle, or the pharmaceutical composition. In some embodiments, the method results in at least a 5% increase in muscle strength of the subject compared to a control subject. In some embodiments, the method results in at least a 50% increase in muscle strength of the subject within 7 days of the administering. In some embodiments, the method results in at least a 2-fold increase in grip strength of the subject within 14 days of the administering. In some embodiments, the method results in at least a 10% increase in grip strength of the subject within about 14 days of the administering. In some embodiments, the method results in at least a 70% increase in grip strength of the subject within 14 days of the administering. In some embodiments, the method results in at least a 10% increase in a number of muscle cells in a sample obtained from the subject compared to a similar sample obtained from a control subject. In some embodiments, the method results in an increased localization of dystrophin protein to a cell membrane of a muscle cell of the subject compared to a level of localization of dystrophin protein to a cell membrane of a muscle cell of the control subject. In some embodiments, the method results in a longer lifespan of the subject compared to a lifespan of a control subject who has the tissue dystrophy. In some embodiments, the method results in the subject have a lifespan that is at least 10% longer compared to a lifespan of a control subject. In some embodiments, the method results in the subject having a time to rise from a laying down position to a standing position of less than about 10 seconds. In some embodiments, the method results in the subject having a time to rise from a laying down position to a standing position of less than about 10 seconds since the administering. In some embodiments, the method results in the subject exhibiting at least a 10% decrease in a time to rise compared to a time to rise of the subject before the administering. In some embodiments, the method results in the subject being measured to have a North Star Ambulatory Assessment (NSAA) score of at least 20 compared to a NSAA score measured of the subject before the administering. In some embodiments, the method results in the subject exhibiting at least a 10% increase in a NSAA score compared to a NSAA score measured of the subject prior to the administering. In some embodiments, the method results in the subjectWSGR Ref. No. 54841-719.601being measured to walk a ten-meter distance at a speed greater than a speed of the subject prior to the administering. In some embodiments, the method results in the subject being measured to walk a distance over a time period of ten minutes that is at least 10% compared to a distance walked by the subject over the ten -minute time period prior to the administering. In some embodiments, subsequent to the administering, the subject can walk up at least four steps of a stairway in no more than about 6 seconds. In some embodiments, subsequent to the administering, the subject walks up at least four steps of a stairway over a time period that is reduced by at least 10% compared to a similar time period over which the subject was able to walk up the at least four steps of the stairway prior to the administering. In some embodiments, the method results in the subject having a four-stair climb time of less than about 10 seconds since the administering. In some embodiments, the method further comprises measuring a level of the human full-length dystrophin in a sample of the subject. In some embodiments, the measuring comprises quantifying a level of human full-length dystrophin mRNA of at least 2.6 x10⁵ copies / mL in the sample obtained from the subject, or processed therefrom, after the administering. In some embodiments, the measuring comprises quantifying a level of human full-length dystrophin mRNA of at least 2.6 x10⁵ copies / mL in the sample obtained from the subject, or processed therefrom, after the administering. In some embodiments, the measuring comprises quantifying a level of human full-length dystrophin protein in the sample obtained from the subject, or processed therefrom, of at least 0.4% of a level of full-length dystrophin protein measured in a healthy control subject, or a control subject who does not have or is not suspected of having a muscle disorder, a muscular dystrophy, or Duchenne Muscular Dystrophy. In some embodiments, the subject has a level of human full-length dystrophin mRNA of at least 2.6 x10⁵ copies / mL after the administering. In some embodiments, the elevated expression level of the human full-length dystrophin is detected by western blot, mass spectrometry, histology, immunohistochemistry, reverse transcription polymerase chain reaction (RT-PCR), RNA sequencing, or northern blot. In some embodiments, the administering further comprises administering at least one dose of the extracellular vesicle, or the pharmaceutical composition to the subject. In some embodiments, the administering further comprises administering at least one dose of the extracellular vesicle or the pharmaceutical composition to the subject. In some embodiments, the administering further comprises administering to the subject a second dose of the extracellular vesicle, or the pharmaceutical composition after at least a period of at least one hour after the administering. In some embodiments, the administeringWSGR Ref. No. 54841-719.601further comprises administering the extracellular vesicle or the pharmaceutical composition to the subject at least once per week. In some embodiments, the administering further comprises administering the extracellular vesicle or the pharmaceutical composition to the subject at least twice per week. In some embodiments, the administering further comprises administering the extracellular vesicle or the pharmaceutical composition to the subject at least once per day. In some embodiments, the administering further comprises administering the extracellular vesicle or the pharmaceutical composition to the subject at least once per month. In some embodiments, the administering further comprises administering at least 4 doses of the extracellular vesicle or the pharmaceutical composition to the subject over a period of at least 2 weeks. In some embodiments, the administering further comprises administering at least 8 doses of the extracellular vesicle or the pharmaceutical composition to the subject over a period of at least one month. In some embodiments, the administering comprises administering the extracellular vesicle or the pharmaceutical composition to a tissue of the subject. In some embodiments, the tissue is a skeletal muscle. In some embodiments, the tissue is a quadricep, a tibialis anterior, a gastrocnemius, a diaphragm, a heart, a cardiac ventricle, a cardiac atrium, or a muscle of a face, a shoulder, an arm, a chest, a back, calves, or a lower extremity of the subject. In some embodiments, the method further comprises obtaining a tissue biopsy from the subject. In some embodiments, the tissue biopsy comprises a muscle tissue biopsy. In some embodiments, the method further comprises obtaining a blood sample from the subject. In some embodiments, the method further comprises obtaining a sample from the subject after a period of at least one hour after the administering.

[0015] In yet another aspect, the instant disclosure further provides a method of producing the extracellular vesicle described herein, wherein the method comprises: introducing a vector encoding the exogenous nucleic acid molecule into an extracellular vesicle donor cell via transfection; culturing the extracellular vesicle donor cell after the introducing for a sufficient amount of time in a culture medium for the production of the extracellular vesicle; and collecting the extracellular vesicle from the culture medium. In some embodiments, the introducing comprises electroporating the vector into the extracellular vesicle donor cell. In some embodiments, the introducing comprises electroporating a second vector into the extracellular vesicle donor cell. In some embodiments, the second vector comprises a polynucleotide sequence encoding the targeting polypeptide. In some embodiments, the introducing comprises applying an electric field across a plurality ofWSGR Ref. No. 54841-719.601vessels configured for culturing the extracellular vesicle donor cell. In some embodiments, the culturing further comprises replenishing the cell culture medium in contact with the extracellular vesicle donor cell. In some embodiments, the culturing further comprises culturing the extracellular vesicle donor cell for one or more cell culture passages. In some embodiments, the method further comprises collecting the extracellular vesicle from the culture medium. In some embodiments, the method further comprises isolating the extracellular vesicle, thereby generating an isolated extracellular vesicle. In some embodiments, the method further comprises concentrating the isolated extracellular vesicle, thereby generating isolated and concentrated extracellular vesicle. In some embodiments, the method further comprises cryopreserving the isolated and concentrated extracellular vesicle. In some embodiments, the method further comprises resuspending the isolated and concentrated extracellular vesicle. In some embodiments, the extracellular vesicle donor cell is a primary cell or a cell obtained from a cell line. In some embodiments, the extracellular vesicle donor cell is a somatic cell. In some embodiments, the extracellular vesicle donor cell is a fibroblast. In some embodiments, the extracellular vesicle donor cell is a dermal fibroblast. In some embodiments, the extracellular vesicle donor cell is a stem cell. In some embodiments, the stem cell is a mesenchymal stem cell. In some embodiments, the extracellular vesicle donor cell is a human cell. Further described and provided herein is a protein comprising at least 97% sequence identity to the sequence set forth in SEQ ID NO: 43. In some embodiments, the protein comprises the sequence set forth in SEQ ID NO: 43. In some embodiments, the protein consists of the sequence set forth in SEQ ID NO: 43.

[0016] Also described herein is a protein comprising at least 97% sequence identity to the sequence set forth in SEQ ID NO: 45. In some embodiments, the protein comprises the sequence set forth in SEQ ID NO: 45. In some embodiments, the protein consists of the sequence set forth in SEQ ID NO: 45.

[0017] In one aspect, this disclosure further provides a polynucleotide encoding a protein of described herein.INCORPORATION BY REFERENCE

[0018] All publications, patents, and patent applications mentioned in this specification are herein incorporated by reference to the same extent as if each individual publication, patent, or patent application was specifically and individually indicated to be incorporated by reference.WSGR Ref. No. 54841-719.601BRIEF DESCRIPTION OF THE DRAWINGS

[0019] This patent application contains at least one drawing executed in color. Copies of this patent or patent application with color drawing(s) will be provided by the Office upon request and payment of the necessary fee.

[0020] The novel features of the disclosure are set forth with particularity in the appended claims. A better understanding of the features and advantages of the present disclosure will be obtained by reference to the following detailed description that sets forth illustrative embodiments.

[0021] FIG. 1 illustrates Cellular Nanoporation (“CNP,” also referred to herein as cellular nanoelectroporation, or “NEP”) generating extracellular vesicles (EVs) loaded with transcribed mRNAs for therapeutic administration.

[0022] FIG.2 depicts in vivo delivery and therapeutic effects induced by administration of extracellular vesicles (EVs) loaded with full-length dystrophin mRNA. FIG.2A shows an experimental schematic and timeline over which Dystrophin -knockout mice (mdx) are intramuscularly administered an injection of EVs containing full-length dystrophin mRNA (Dystrophin-mRNA-EV) on day 1. Mice are sacrificed on day 14, and their tissue is subsequently analyzed for dystrophin expression. FIG. 2B depicts quantification of paw grip strength measured in grams of force in mdx mice injected with Dystrophin-mRNA-EVs (Mdx + EV mRNA; DMD t-EVs) in comparison with wildtype (WT) mice and uninjected mdx mice.

[0023] FIG.3 shows quantification of grip strength measured in untreated DMD KO mice, DMD KO mice treated with DMD t-EVs, and untreated D / V / D-WT littermates at weekly intervals from 0 weeks to 16 weeks after intravenous injection of DMD t-EVs. DMD t-EV treatment result a ~2 -fold increase, or doubling, in the grams of force of grip strength of DMD KO mice compared to untreated DMD KO mice, and further increased grip strength to be much more comparable to the untreated D / V / D-WT littermates. DMD t-EVs are targeting extracellular vesicles that express the human CD47-T7 and / or human CD47 -T9 targeting polypeptides and are loaded with full-length human dystrophin mRNA cargo.

[0024] FIG.4 depicts representative fluorescent immunohistochemical data in situ showing localization and expression intensity of GFP-labelled human dystrophin (hHDMD-GFP) mRNA DMD t-EVs and full-length dystrophin in the tibialis anterior (FIG. 4A) and quadriceps (FIG. 4B) of WT, untreated mdx KO, and DMD t-EV-treated mdx KO mice after at least 1,2, 3, and 4 months after administration of the first of biweekly intravenousWSGR Ref. No. 54841-719.601injections of DMD t-EVs to mdx KO mice. DMD t-EVs were administered every 3-4 days throughout the course of treatment.

[0025] FIG. 5 depicts representative fluorescent immunohistochemical data in situ showing engraftment of DMD-GFP protein and dystrophin protein levels in the gastrocnemius (FIG. 5A), triceps (FIG. 5B), and diaphragm (FIG. 5C), and heart (FIG. 5D) in WT control, untreated mdx KO mice, and mdx KO mice that received biweekly (every 3-4 days) intravenous injections of DMD t-EVs after for at least 1 month or 4 months. The mdx KO mice that received injections of DMD t-EVs were also intravenously administered FK506 (tacrolimus, a calcineurin inhibitor) daily throughout the course of the experiment. All test animals were 3-4 weeks of age at the time the DMD t-EV-treated mdx KO mice received the first DMD t-EV injection.

[0026] FIG. 6A shows representative fluorescent immunohistochemical images in situ showing that 3.5 weeks after intravenous administration of DMD t-EVs loaded with GFP- labelled human dystrophin (hDMD-GFP) mRNA results in skeletal muscle protein engraftment for non-human primate (NHP) models. Three fields of view of quadricep tissue sections were assessed. Arrow in FOV3 indicates colocalization of dystrophin protein (red) with HDMD-GFP (green).

[0027] FIG. 6B depicts quantification of the percentage of HDMD-GFP fluorescence area relative to total dystrophin fluorescence area in FOV1, FOV2, and FOV3.

[0028] FIG. 6C depicts quantification of normalized total fluorescence intensity of DIR- labeled DMD t-EVs in various organs following intravenous administration of DMD t- EVs to marmosets. Marmosets were administered 7 doses of DMD t-EVs every 3-4 days before wide-field fluorescence imaging. D = Diaphragm, Q = Quadriceps (leg muscle), T = Triceps, G = Gastrocnemius (leg muscle), TA = Tibialis Anterior (leg muscle), H = Heart, Lu = Lung, K = Kidney, Li = Liver, B = Brain.

[0029] FIG. 7 shows a schematic of an ongoing Phase I clinical trial administering the extracellular vesicles provided and described herein for treating muscular dystrophy in pediatric male patients in accordance with some embodiments.

[0030] FIGs.8 shows results of a Complete Blood Count (CBC) hematological analysis of blood samples obtained from Patient 1 of cohort 1 over a time course that spans pre - and post-treatment timepoints evaluated in this study. At 16 days before treatment (D-16), Patient 1 was not yet taking oral tacrolimus. Hematology parameters measured are: white blood cell count (WBC; FIG. 8A), red blood cell count (RBC; FIG. 8B), Red Cell Distribution Width - Coefficient of Variation (RDW-CV; FIG. 8C), platelet count (PLT;WSGR Ref. No. 54841-719.601FIG. 8D), Platelet Distribution Width (PDW; FIG. 8E), N-terminal pro-B-type natriuretic peptide (NT-proBNP; FIG.8F), Hematocrit (HCT; FIG. 8G), basophil count (BASO count; FIG.8H), eosinophil count (EO; FIG. 8I), C-reactive protein (CRP; FIG.8J), basophil percentage (BASO%; FIG. 8K), eosinophil percentage (EO%; FIG. 8L), Mean Corpuscular Volume (MCV; FIG. 8M), lymphocyte count (LYMPH; FIG. 8N), Mean Corpuscular Hemoglobin (MCH; FIG. 8O), Mean Corpuscular Hemoglobin Concentration (MCHC; FIG. 8P), lymphocyte percentage (LYMPH%; FIG. 8Q), hemoglobin (HGB; FIG.8R), neutrophil count (NEUT; FIG. 8S), monocyte percentage (MONO%; FIG. 8T), Mean Platelet Volume (MPV; FIG. 8U), neutrophil percentage (NEUT%; FIG. 8V), and monocyte count (MONO; FIG. 8W). Shaded region of each graph indicates the normal range of each respective CBC parameter.

[0031] FIG. 9 shows results of a comprehensive metabolic panel (CMP) in blood samples obtained from Patient 1 over a time course that spanned before, during, and after treatment with the hDMD-mRNA-tEXOs. Patient 1 received doses of hDMD-mRNA- tEXOs for cohort 1 in the study. Blood parameters that were measured included in the CMP were Alanine Transaminase (ALT; FIG. 9A), Aspartate Aminotransferase (AST;FIG. 9B), Creatine Kinase (CK; FIG. 9C), Alkaline Phosphatase (ALP; FIG. 9D), Calculated Globulin (CG; FIG.9E), creatine kinase-MB isoenzyme (CK-MB; FIG. 9F), prealbumin (PA; FIG. 9G), Glutathione Reductase (GR; FIG. 9H), lactate dehydrogenase (LDH; FIG. 9I), phosphate (PH; FIG. 9J), glucose (GLU; FIG. 9K), lactate (FIG. 9L), albumin (ALB; FIG. 9M), Globulin (GLB; FIG. 9N), albumin-to-globulin ratio (ALB / GLB; FIG. 9O), Gamma-glutamyl transferase (GGT; FIG. 9P), Direct Bilirubin (DBIL; FIG. 9Q), Cystatin C (CysC; FIG. 9R), Total Bilirubin (TILB;FIG. 9S), total protein (TP; FIG. 9T), urea (FIG. 9U), troponin (FIG. 9V), uric acid (UA; FIG. 9W). Shaded region of each graph indicates the normal range of each respective CMP parameter.

[0032] FIG. 10 shows results of an electrolyte panel in blood samples obtained from Patient 1 over a time course that spanned before, during, and after treatment with the hDMD- mRNA-tEXOs. Parameters that were measured included in the electrolyte panel were sodium (FIG. 10A), potassium (FIG. 10B), chloride (FIG. 10C), calcium (FIG. 10D), enzymatic carbon dioxide (ECO2; FIG. 10E), and magnesium (FIG. 10F). Shaded region of each graph indicates the normal range of each respective electrolyte.

[0033] FIG. 11 shows results of a cytokine multiplex assays in blood samples obtained from Patient 1 over a time course that spanned before, during, and after treatment with theWSGR Ref. No. 54841-719.601hDMD-mRNA-tEXOs. At 12 days before treatment (D-12), a baseline blood sample was obtained from the Patient 1. Levels of the following cytokines were measured in the cytokine panel: IL-1β (FIG. 11A), IL-2 (FIG. 11B), IL-2R (FIG. 11C), IL-4 (FIG. 11D), IL-5 (FIG. 11E), IL-6 (FIG. 11F), IL-8 (FIG. 11G), IL-10 (FIG. 11H), IL-17 (FIG. 11I), IL-18 (FIG. 11J), IL-12p70 (FIG. 11K), TNFα (FIG. 11L), INF-α (FIG. 11M), IFN-γ (FIG. 11N), and INF-β (FIG. 11O). Shaded region of each graph indicates the normal range of each respective cytokine.

[0034] FIG. 12 shows results of vital signs measured in Patient 1 spanning the time from the day of the initial infusion of hDMD-mRNA-tEXOs until day 54 (D54). Measured vital signs are respiration frequency (FIG. 12A), sphygmus / heart rate (FIG. 12B), body temperature (FIG. 12C), peripheral capillary oxygen saturation (SPO2; FIG. 12D), Systolic Blood Pressure (SBP; FIG. 12E), and Diastolic Blood Pressure (DBP; FIG. 12F).Day and / or time stamps of each vital sign measured are indicated on the x-axis.

[0035] FIG. 13 A depicts a representative normalized reporter (Rn) intensity curve of human dystrophin mRNA amplification measured by quantitative RT-PCR (qRT-PCR) in gastrocnemius and bicep tissue biopsies obtained from Patient 1 before and after treatment with hDMD-mRNA-tEXOs. One biopsy was obtained from the gastrocnemius and bicep at baseline (before treatment with hDMD-mRNA-tEXOs), and then two biopsy samples were obtained from each of the gastrocnemius and bicep at D29 after treatment.

[0036] FIG. 13B depicts a standard curve of synthetic human dystrophin mRNA copies per milliliter (copies / mL) generated to achieve the indicated cycle threshold (Ct) value. Positive Template Control (PTC) recovery of a non-DMD (normal) tissue bank biopsy sample was 134.77%. No-template control (NTC) was measured at 30.29 (Ctvalue).

[0037] FIG. 13C depicts quantification of dystrophin (DMD) mRNA concentration detected in gastrocnemius and bicep biopsy tissue obtained from Patient 1 before and after treatment with 8 doses of 5×109hDMD-mRNA-tEXOs per kg. Before treatment: n = 6; average 4.00×105DMD mRNA copy number / mL ± 6.59×104SD. After treatment: n = 6; average 3.89×106DMD mRNA copy number / mL ± 9.76×105SD.

[0038] FIG. 13D depicts a representative western blot of dystrophin protein expression (top) and loading control (bottom) detected in muscle biopsy tissue from Patient 1 before and after treatment with hDMD-mRNA-tEXOs.

[0039] FIG. 13E depicts a full-field view of the western blot shown in FIG. 13D.

[0040] FIG. 13F depicts a curve of dystrophin protein band intensity in muscle biopsy samples obtained from Patient 1 before and after treatment with hDMD-mRNA-tEXOs.WSGR Ref. No. 54841-719.601Standard curve is based on dystrophin expression across 5 dilutions of normal (non-DMD) tissue biopsy sample. Arrows indicate the dystrophin levels measured in muscle biopsy samples obtained from Patient 1 before and after treatment.

[0041] FIG. 13G depicts quantification of dystrophin protein expression levels in muscle biopsies obtained from Patient 1 before and after treatment with hDMD-mRNA-tEXOs. Dystrophin protein expression level was normalized to the average dystrophin protein levels detected in pre-treatment biopsy samples.

[0042] FIG. 13H shows representative immunofluorescence images of laminin, DAPI, and dystrophin protein expression and localization in tissue sections prepared from human muscle biopsies obtained from Patient 1 before and after treatment with hDMD-mRNA- tEXOs.

[0043] FIG. 131 depicts representative immunofluorescence images of laminin, DAPI, and dystrophin protein expression and localization in tissue sections prepared from human muscle biopsies obtained from Patient 1 before and 3 days after the final dose of treatment with hDMD-mRNA-tEXOs across 3 fields of view (FOV) for each treatment condition.

[0044] FIG. 13J and FIG. 13K show quantification of normalized dystrophin immunofluorescence intensity and the fold percentage change of dystrophin immunofluorescence intensity in the FOVs shown in FIG. 131, respectively.Immunofluorescence intensity of dystrophin expression detected in each FOV for before and after treatment conditions was normalized to the average dystrophin levels measured in tissues sections of the before -treatment muscle biopsies. FIG. 13K Treatment with hDMD-mRNA-tEXOs resulted in a statistically significant (***p < 0.001) increase of dystrophin levels detected in after-treatment muscle biopsies compared to before- treatment samples.

[0045] FIG. 14A shows quantification of the time to rise task performed by Patient 1 and Patient 2 throughout the study period, including before, during, and after administration of the 8-dose treatment regimen with HDMD-mRNA-EXOs. The time to rise was defined as the time needed for the subject to rise from laying flat on the ground to standing in the upright position. Patient 1 ’s time to rise was measured at 0 (baseline), 7, 14, 24, 56, and 114 days relative to the day on which Patient 1 received the first dose of HDMD-mRNA- EXOs. Patient 2’s time to rise was measured at 0 (baseline), 7, and 14 days relative to the day on which Patient 2 received the first dose of HDMD-mRNA-EXOs. On each day of assessment, 3 trials were performed. Error bars represent standard deviation of the mean. Shaded region indicates the first assessment measured after patients were administeredWSGR Ref. No. 54841-719.601HDMD-mRNA-EXOs. Baseline time to rise measurements were obtained before administration of the first dose of HDMD-mRNA-EXOs.

[0046] FIG. 14B depicts quantification of overall NSAA scores of Patient 1 and Patient 2 throughout the study period, including before, during, and after administration of the 8 - dose treatment regimen with HDMD-mRNA-EXOs. Patient 1 ’s NSAA score was measured at 0 (baseline), 7, 14,24, 56, and 114 days relative to the day on which Patient 1 received the first dose of HDMD-mRNA-EXOs. Patient 2 ’s NSAA score was measured at 0 (baseline), 7, and 14 days relative to the day on which Patient 2 received the first dose of HDMD-mRNA-EXOs. NSAA scores were based on the combined additive scores of each patient’s ability to perform their respective assigned NSAA tasks. Shaded region indicates the first assessment measured after patients were administered HDMD-mRNA-EXOs. The baseline NSAA score for each patient was obtained before administration of the first dose of HDMD-mRNA-EXOs.

[0047] FIG. 14C shows quantification of the 4-stair climb test (4SCT) task performed by Patient 1 and Patient 2 throughout the study period, including before, during, and after administration of the 8 -dose treatment regimen with HDMD-mRNA-EXOs. The 4SCT was defined as the time needed for the subject to climb 4 stairs of at least 15 cm in height. Patient 1 ’s 4SCT was measured at 24, 56, and 114 days relative to the day on which Patient 1 received the first dose of HDMD-mRNA-EXOs. Patient 2 ’s 4SCT was measured at 0 (baseline), 7, and 14 days relative to the day on which Patient 2 received the first dose of HDMD-mRNA-EXOs. On each day of assessment, 3 trials were performed. Error bars represent standard deviation of the mean. Shaded region indicates the first assessment measured after patients were administered HDMD-mRNA-EXOs. Baseline time to rise measurements were obtained before administration of the first dose of HDMD-mRNA- EXOs.DETAILED DESCRIPTION OF THE INVENTION

[0048] While various embodiments of the invention have been shown and described herein, it will be obvious to those skilled in the art that such embodiments are provided by way of example only. Numerous variations, changes, and substitutions may occur to those skilled in the art without departing from the invention. It should be understood that various alternatives to the embodiments of the invention described herein may be employed.WSGR Ref. No. 54841-719.601

[0049] This disclosure provides extracellular vesicles that carry exogenous RNA cargo and that can be used to treat a variety of tissue disorders, often with long-lasting effects. Further provided herein are extracellular vesicles that contain mRNA, particularly mRNA encoding a dystrophin protein, and that can be used to treat dystrophies, such as muscular dystrophies or cardiac dystrophies. The extracellular vesicles of this disclosure may be therapeutic. Accordingly, also described herein is a method of treating a tissue dystrophy by administering to a subject the extracellular vesicles that carry exogenous RNA cargo. The present disclosure provides an extracellular vesicle that expresses, encapsulates, or carries an RNA cargo. In certain embodiments, the extracellular vesicle is an exosome. In some cases, the extracellular vesicle expresses CD9, CD63, TSG101, or ARF6 protein markersDefinitions

[0050] Unless defined otherwise, all terms of art, notations and other technical and scientific terms or terminology used herein are intended to have the same meaning as is commonly understood by one of ordinary skill in the art to which the claimed subject matter pertains. In some cases, terms with commonly understood meanings are defined herein for clarity and / or for ready reference, and the inclusion of such definitions herein should not necessarily be construed to represent a substantial difference over what is generally understood in the art.

[0051] In the following description, certain specific details are set forth in order to provide a thorough understanding of various embodiments. However, one skilled in the art will understand that the embodiments provided canbe practiced without these details. Unless the context requires otherwise, throughout the specification and claims which follow, the word “comprise” and variations thereof, such as, “can comprise” and “comprising” are to be construed in an open, inclusive sense, that is, as “including, but not limited to.” As used herein, open terms, for example, “comprise”, “contain”, “include”, “including”, “have”, “having” and the like refer to comprising unless otherwise indicated.

[0052] As used in this specification and the appended claims, the singular forms “a,” “an,” and “the” include plural referents unless the content clearly dictates otherwise. Further, headings provided herein are for convenience only and do not interpret the scope or meaning of the claimed embodiments.

[0053] As used herein, “or” may refer to “and”, “or,” or “and / or” and may be used both exclusively and inclusively. For example, the term “A or B” may refer to “A or B”, “A but not B”, “B but not A”, and “A and B”. In some cases, context may dictate a particularWSGR Ref. No. 54841-719.601meaning. As used herein, the phrases “at least one”, “one or more”, and “and / or” are open-ended expressions that are both conjunctive and disjunctive in operation. For example, each of the expressions “at least one of A, B and C”, “at least one of A, B, or C”, “one or more of A, B, and C”, “one or more of A, B, or C” and “A, B, and / or C” means A alone, B alone, C alone, A and B together, A and C together, B and C together, or A, B and C together.

[0054] The term “about” or “approximately” means within an acceptable error range for the particular value as determined by one of ordinary skill in the art, which will depend in part on how the value is measured or determined, e.g., the limitations of the measurement system. For example, “about” can mean within 1 or more than 1 standard deviation, per the practice in the given value. Where particular values are described in the application and claims, unless otherwise stated the term “about” should be assumed to mean an acceptable error range for the particular value.Numeric ranges

[0055] Whenever the term “at least,” “greater than,” or “greater than or equal to” precedes the first numerical value in a series of two or more numerical values, the term “at least,” “greater than” or “greater than or equal to” applies to each of the numerical values in that series of numerical values. For example, greater than or equal to 1, 2, or 3 is equivalent to greater than or equal to 1, greater than or equal to 2, or greater than or equal to 3.

[0056] Whenever the term “no more than,” “less than,” or “less than or equal to” precedes the first numerical value in a series of two or more numerical values, the term “no more than,” “less than,” or “less than or equal to” can apply to each of the numerical values in that series of numerical values. For example, less than or equal to 3, 2, or 1 is equivalent to less than or equal to 3, less than or equal to 2, or less than or equal to 1.

[0057] Throughout this application, various cases may be presented in a range format. It should be understood that the description in range format is merely for convenience and brevity and should not be construed as an inflexible limitation on the scope of the disclosure. Accordingly, the description of a range should be considered to have specifically disclosed all the possible subranges as well as individual numerical values within that range. For example, description of a range such as from 1 to 6 should be considered to have specifically disclosed subranges such as from 1 to 3, from 1 to 4, from 1 to 5, from 2 to 4, from 2 to 6, from 3 to 6 etc., as well as individual numbers within that range, for example, 1, 2, 3, 4, 5, and 6. This applies regardless of the breadth of the range.WSGR Ref. No. 54841-719.601

[0058] “Optional” or “optionally” means that the subsequently described event or circumstance may or may not occur, and that the description includes instances where said event or circumstance occurs and instances where it does not.

[0059] “Exemplary” means “an example of’ and is not intended to convey an indication of a preferred or ideal aspect. “Such as” is not used in a restrictive sense, but for explanatory purposes.

[0060] As used herein, the terms “essentially” or “substantially,” when describing a relative value, a relative amount or a relative degree between two subjects, generally refers to within 80%, 81%, 82%, 83%, 84%, 85%, 86%, 87%, 88%, 89% 90%, 91%, 92%, 93%, 94%, 95%, 96%, 97%, 98%, 99%, 100%, 101%, 102%, 103%, 104%, 105%, 106%, 107%, 108%, 109%, or 110% of each other in value, amount or degree.Increasing

[0061] The terms “increased”, “increasing”, or “increase” are used herein to generally mean an increase by a statically significant amount. In some cases, the terms “increased,” or “increase,” mean an increase of at least 10% as compared to a reference level, for example an increase of at least 10%, at least 20%, or at least 30%, or at least 40%, or at least 50%, or at least 60%, or at least 70%, or at least 80%, or at least 90% or up to and including a 100% increase or any increase between 10-100% as compared to a reference level, standard, or control. Other examples of “increase” include an increase of at least 2 - fold, at least 5 -fold, at least 10-fold, at least 20-fold, at least 50-fold, at least 100-fold, at least 1000-fold or more as compared to a reference level.Decreasing

[0062] The terms, “decreased”, “decreasing”, or “decrease” are used herein generally to mean a decrease by a statistically significant amount. In some cases, “decreased” or “decrease” means a reduction by at least 10% as compared to a reference level, for example a decrease by at least 20%, or at least 30%, or at least 40%, or at least 50%, or at least 60%, or atleast70%, oratleast80%, orat least 90% orup to and including a 100% decrease (e.g., absent level or non-detectable level as compared to a reference level), or any decrease between 10-100% as compared to a reference level. In the context of a marker or symptom, by these terms is meant a statistically significant decrease in such level. The decrease can be, for example, at least 10%, at least 20%, at least 30%, at leastWSGR Ref. No. 54841-719.60140% or more, and is preferably down to a level accepted as within the range of normal for an individual without a given disease.Polynucleotide

[0063] The terms “polynucleotide,” “oligonucleotide,” and “nucleic acid” are used interchangeably to refer to a polymeric form of nucleotides of any length, either deoxyribonucleotides or ribonucleotides, or analogs thereof, either in single-, double-, or multi-stranded form. In some cases, a polynucleotide is exogenous (e.g. a heterologous polynucleotide). In some cases, a polynucleotide is endogenous to a cell. In some cases, a polynucleotide can exist in a cell-free environment. In some cases, a polynucleotide is a gene or fragment thereof. In some cases, a polynucleotide is DNA. In some cases, a polynucleotide is RNA. A polynucleotide canhave any three-dimensional structure, and can perform any function, known or unknown. In some cases, a polynucleotide comprises one or more analogs (e.g. altered backbone, sugar, or nucleobase). If present, modifications to the nucleotide structure may be imparted before or after assembly of the polymer. Some non-limiting examples of analogs include: 5 -bromouracil, peptide nucleic acid, xeno nucleic acid, morpholinos, locked nucleic acids, glycol nucleic acids, threose nucleic acids, dideoxynucleotides, cordycepin, 7-deaza-GTP, fluorophores (e.g. rhodamine or fluorescein linked to the sugar), thiol containing nucleotides, biotin linked nucleotides, fluorescent base analogs, CpG islands, methyl-7-guanosine, methylated nucleotides, inosine, thiouridine, pseudouridine, dihydrouridine, queuosine, and wyosine. In some cases, a polynucleotide can comprise one or more analogs (e.g. altered backbone, sugar, or nucleobase). Non-limiting examples of polynucleotides include coding or noncoding regions of a gene or gene fragment, loci (locus) defined from linkage analysis, exons, introns, messenger RNA (mRNA), transfer RNA (tRNA), ribosomal RNA (rRNA), short interfering RNA (siRNA), short-hairpin RNA (shRNA), micro-RNA (miRNA), non-coding RNA, ribozymes, cDNA, recombinant polynucleotides, branched polynucleotides, plasmids, vectors, isolated DNA of any sequence, isolated RNA of any sequence, cell- free polynucleotides including cell-free DNA (cfDNA) and cell-free RNA (cfRNA), nucleic acid probes, and primers. In some cases, the sequence of nucleotides is interrupted by non-nucleotide components.WSGR Ref. No. 54841-719.601Nucleotide

[0064] The term “nucleotide,” as used herein, generally refers to a base-sugar-phosphate combination. A nucleotide comprises a synthetic nucleotide. A nucleotide comprises a synthetic nucleotide analog. Nucleotides is monomeric units of a nucleic acid sequence (e.g. deoxyribonucleic acid (DNA) and ribonucleic acid (RNA)). The term nucleotide can include ribonucleoside triphosphates adenosine triphosphate (ATP), uridine triphosphate (UTP), cytosine triphosphate (CTP), guanosine triphosphate (GTP) and deoxyribonucleoside triphosphates such as dATP, dCTP, diTP, dUTP, dGTP, dTTP, or derivatives thereof. Such derivatives can include, for example, [aS]dATP, 7 -deaza-dGTP and 7-deaza-dATP, and nucleotide derivatives that confer nuclease resistance on the nucleic acid molecule containing them. The term nucleotide as used herein can refer to dideoxyribonucleoside triphosphates (ddNTPs) and their derivatives. Illustrative examples of dideoxyribonucleoside triphosphates can include, but are not limited to, ddATP, ddCTP, ddGTP, ddITP, and ddTTP.Intact

[0065] “Fully intact” and “substantially intact” refer to a nucleic acid described herein having a nucleic acid sequence that can be transcribed and / or translated into a therapeutic polypeptide described herein. Fully intact nucleic acid refers to full-length nucleic acid sequence, which is not partially degraded or fragmented. For example, a fully intact nucleic acid can be a messenger RNA that can be translated into a full-length protein such as any one of the therapeutic polypeptides described herein. In general, a fully intact or substantially intact messenger RNA is capable of being translated into a polypeptide. Generally, messenger RNA can comprise a 5’ cap which may assist with binding to a ribosome and a poly (A) tail, which may be useful for translation. The term “substantially intact” refers to a nucleic acid sequence that can be partially degraded or fragmented but still can be transcribed and / or translated into any one of the therapeutic polypeptides described herein. For example, a substantially intact nucleic acid can be a partially degraded or fragmented messenger RNA that can be translated into any one of the therapeutic polypeptides described herein.Fragment

[0066] As used herein, the terms “fragment” or equivalent terms can refer to a locus of a protein that has less than the full length of the protein and optionally maintains theWSGR Ref. No. 54841-719.601function of the protein. “Percent identity” and “% identity” refers to the extent to which two sequences (nucleotide or amino acid) have the same residue at the same positions in an alignment. For example, “an amino acid sequence is X% identical to SEQ ID NO: Y” refers to % identity of the amino acid sequence to SEQ ID NO: Y and is elaborated as X% of residues in the amino acid sequence are identical to the residues of sequence disclosed in SEQ ID NO: Y. Generally, computer programs are employed for such calculations. Exemplary programs that compare and align pairs of sequences, include ALIGN, FASTA, gapped BLAST, BLASTP, BLASTN, or GCG.Polypeptide

[0067] As used herein, the terms “polypeptide”, “peptide”, and “protein” can be used interchangeably herein in reference to a polymer of amino acid residues. A polypeptide can refer to a full-length polypeptide as translated from a coding open reading frame, or as processed to its mature form. A polypeptide can refer to a degradation fragment or a processing fragment of a protein that nonetheless uniquely or identifiably maps to a particular protein. A polypeptide can be a single linear polymer chain of amino acids bonded together by peptide bonds between the carboxyl and amino groups of adjacent amino acid residues. A polypeptide can be modified, for example, by the addition of carbohydrate, phosphorylation, etc. A polypeptide can be a heterologous polypeptide.

[0068] The term “full-length dystrophin” as used herein may refer to any dystrophin isoform.In certain embodiments, the term “full-length dystrophin” refers to the isoform 4 (Dp427). Possible nucleic acid sequence encoding Dp427 or any other isoform or homolog of dystrophin is available publicly, See, e.g., NCBI Reference Sequences: NM 000109.3, NM_004006.2, NM_004009.3, NM_004010.3, NM_004011.3, NM_004012.3, NM_004013.2, NM_004014.2, NM_004015.2, NM_004016.2 NM_004017.2, NM_004018.2, NM_004019.2, NM_004020.3, NM 004021.2, NM_004022.2, NM_004023.2, NM_004007.2, XM_006724468.2, XM_006724469.3, XM_006724470.3, XM_006724473.2, XM_006724474.3, XM_006724475.2, XM 011545467.1, XM_011545468.2, XM_011545469.1, XM_017029328.1, XM_017029329.1, XM 017029330.1, and XM_017029331.1, each of which is incorporated herein. Additional sequence encoding Dp427 or any other isoform or homolog of dystrophin may be generated via tools for reverse -translation, e.g., www.ebi.ac.uk / Tools / st / , www.ebi.ac.uk / Tools / st / emboss_transeq / , www.ebi.ac.uk / Tools / st / emboss_sixpack / , www.ebi.ac.uk / Tools / st / emboss_backtranseq / ,WSGR Ref. No. 54841-719.601and www.ebi.ac.uk / Tools / st / emboss_backtranambig / . Furthermore, the coding sequences might be codon-optimized for expression in a subject.Percentage sequence identity

[0069] “Percentage of sequence identity”, as used herein, can be determined by comparing two optimally locally aligned sequences over a comparison window defined by the length of the local alignment between the two sequences. The amino acid sequence in the comparison window can comprise additions or deletions (e. g., gaps or overhangs) as compared to the reference sequence (which does not comprise additions or deletions) for optimal alignment of the two sequences.

[0070] Local alignment between two sequencescan include segments of each sequence that are deemed to be sufficiently similar according to a criterion that depends on the algorithm used to perform the alignment (e. g. BLAST). The percentage of sequence identity is calculated by determining the number of positions at which the identical nucleic acid base or amino acid residue occurs in both sequences to yield the number of matched positions, dividing the number of matched positions by the total number of positions in the window of comparison and multiplying the result by 100. Optimal alignment of sequences for comparison can be conducted by the local homology algorithm of Smith and Waterman (Add. APL. Math. 2:482, 1981), by the global homology alignment algorithm of Needleman and Wunsch (J Mol. Biol. 48:443, 1970), by the search for similarity method of Pearson and Lipman (Proc. Natl. Acad. Sci. USA 85:2444, 1988), by heuristic implementations of these algorithms (NCBI BLAST, WU- BLAST, BLAT, SIM, BLASTZ), or by inspection. Given that two sequences have been identified for comparison, GAP and BESTFIT can be employed to determine their optimal alignment. Typically, the default values of 5.00 for gap weight and 0.30 for gap weight length are used. The term “substantial sequence identity” between polynucleotide or polypeptide sequences refers to polynucleotide or polypeptide comprising a sequence that has at least 50% sequence identity, at least 60% sequence identity, at least 70% sequence identity, at least 80% sequence identity, at least 85% sequence identity, at least 90% sequence identity, at least 95% sequence identity, at least 96% sequence identity, at least 97% sequence identity, at least 98% sequence identity, or at least 99% sequence identity compared to a reference sequence using the programs. In addition, pairwise sequence homology or sequence similarity, as used, refers to the percentage of residues that are similar between two sequences aligned. Families of amino acid residues havingWSGR Ref. No. 54841-719.601similar side chains have been well defined in the art. These families include amino acids with basic side chains (e.g., lysine, arginine, histidine), acidic side chains (e.g., aspartic acid, glutamic acid), uncharged polar side chains (e.g., glycine, asparagine, glutamine, serine, threonine, tyrosine, cysteine), nonpolar side chains (e.g., alanine, valine, leucine, isoleucine, proline, phenylalanine, methionine, tryptophan), beta-branched side chains (e.g., threonine, valine, isoleucine) and aromatic side chains (e.g., tyrosine, phenylalanine, tryptophan, histidine). Query nucleic acid and amino acid sequences can be searched against subject nucleic acid or amino acid sequences residing in public or proprietary databases. Such searches can be done using the National Center for Biotechnology Information Basic Local Alignment Search Tool (NCBI BLAST v 2.18) program. The NCBI BLAST program is available on the internet from the National Center for Biotechnology Information (blast.ncbi.nlm.nih.gov / Blast.cgi). Typically the following parameters for NCBI BLAST can be used: Filter options set to “default”, the Comparison Matrix set to “BLOSUM62”, the Gap Costs set to “Existence: 11, Extension: 1”, the Word Size set to 3, the Expect (E threshold) set to le-3, and the minimum length of the local alignment set to 50% of the query sequence length. Sequence identity and similarity can also be determined using GenomeQuest™ software (Gene-IT, Worcester Mass. USA).Functional isoform

[0071] As used herein, the terms “isoform”, “functional isoform”, or “functional fragment” or equivalent terms are used herein interchangeably to refer to a locus of an RNA molecule that encodes a protein that has less than the full length of an RNA molecule that encodes the full-length protein, or a protein that has an amino acid sequence that is less than the full length of the protein, and maintains the function of the protein. Examples of functional isoforms can be a splicing isoform of dystrophin that retains dystrophin function in muscle cells to confer essentially normal muscle development in a subject or a cell. For example, Dystrophin-3, Dp427c (UniprotPl 1532-4) and Dystrophin-4, Dp427m (UniProt: Pl 1532-1) are functional splicing isoforms that can retain dystrophin function in muscle development.Culturing

[0072] The term “culturing”, as used herein, can refer to the propagation of cells and / or cellular byproducts or secreted extracellular vesicles thereof on or in media of variousWSGR Ref. No. 54841-719.601kinds. Non-limiting examples of suitable media include Dulbecco's Phosphate Buffered Saline (DPBS), Dulbecco's Modified Eagle's Medium (DMEM), Minimum Essential Medium (MEM), and other types of cell culture media known in the art.Extracellular vesicle donor cell

[0073] The terms “extracellular vesicle donor cell”, “donor cell”, or “source cell” can refer to a type of cell that can be cultured in vitro or sourced from a biological sample or commercial cell line that produces and secretes extracellular vesicles (e.g., exosomes, microvesicles, or apoptotic bodies) into their surrounding environment, such as in a culture medium or buffer.Transfection

[0074] The terms “transfection” or “transfected” generally refers to introduction of a nucleic acid construct into a cell by non-viral or viral-based methods. In some cases, the nucleic acid molecules are gene sequences encoding complete proteins or functional portions thereof. In some cases, the nucleic acid molecules are non-coding sequences. In some cases, the transfection methods are utilized for introducing nucleic acid molecules into a cell for generating a transgenic animal. Such techniques can include pronuclear micro injection, retrovirus mediated gene transfer into germ lines, gene targeting into embryonic stem cells, electroporation of embryos, sperm mediated gene transfer, and in vitro transformation of somatic cells, such as cumulus or mammary cells, or adult, fetal, or embryonic stem cells, followed by nuclear transplantation.Nanoelectroporation

[0075] The term “cell nanoporation”, “CNP”, “nanoelectroporation”, or “NEP” are used interchangeably herein to refer to a method of transfecting a cell, such as a donor cell, with at least one heterologous polynucleotide such as a vector by loading the at least o ne heterologous polynucleotide into a nanochannel and accelerating the at least one heterologous polynucleotide into the cell by generating an electric field. The cell to be transfected canbe situated at an opening of the nanochannel, where the electric field of the nanoelectroporation can create pores in the cell membrane to allow the at least one heterologous polynucleotide to be introduced into the cell. By applying an electric field across a porous membrane containing channels for transducing a plasmid, drug, viral vector, or nucleic acid into the cell to express an exogenous molecule or therapeutic molecule. The channels of the porous membrane can span the thickness of the porousWSGR Ref. No. 54841-719.601membrane and can have a diameter that can be less than 20 μm, 10 μm, 1 μm, 1000 nm, 900 nm, 800 nm, 700 nm, 600 nm, 500 nm, 400 nm, 300 nm, 200 nm, or less than 100 nm. The terms “pore”, “nanochannel”, and “micro channel” are used interchangeably herein to refer to the channels of the CNP membrane for delivering the plasmids, vectors, drugs, or nucleic acids described throughout this disclosure. It shall be understood that instances of “nanochannels” and “microchannels” recited herein are not intended to be limited to the metric range of nanometers or micrometers. For example, a “microchannel” can be used to describe a channel of a CNP membrane that are 500 nm is diameter. In another example, a “nanochannel” can describe a channel of a CNP membrane that is 1500 nm, or 1.5 μm.Plasmid

[0076] A “plasmid,” as used herein, generally refers to a non-viral expression vector, e.g., a nucleic acid molecule that encodes for genes and / or regulatory elements necessary for the expression of genes. The term “vector,” as used herein, generally refers to a nucleic acid molecule capable transferring or transporting a payload nucleic acid molecule. The payload nucleic acid molecule can be generally linked to, e.g., inserted into, the vector nucleic acid molecule. A vector can include sequences that direct autonomous replication in a cell, or can include sequences sufficient to allow integration into host cell gene (e.g., host cell DNA). Examples of a vector can include, but are not limited to, plasmids (e.g., DNA plasmids or RNA plasmids), transposons, cosmids, bacterial artificial chromosomes, and viral vectors. A “viral vector,” as used herein, generally refers to a viral-derived nucleic acid that is capable of transporting another nucleic acid into a cell. A viral vector is capable of directing expression of a protein or proteins encoded by one or more genes carried by the vector when it is present in the appropriate environment. Examples for viral vectors include, but are not limited to Gamma -retroviral, Alpha- retroviral, Foamy viral, lentiviral, adenoviral, or adeno-associated viral vectors. A vector of any of the aspects of the present disclosure can comprise exogenous, endogenous, or heterologous control sequences such as promoters and / or enhancers.Exogenous RNA

[0077] As used here, the term “extracellular vesicle”, abbreviated as EV, can encompass exosomes, microvesicles, apoptotic bodies, and the like. The terms “extracellular vesicles” and “EVs,” as used herein, may in some embodiments refer to a membranousWSGR Ref. No. 54841-719.601particle having a diameter (or largest dimension where the particles is not spheroid) of between about 10 nm to about 5000 nm, more typically between 30 nm and 1000 nm, and most typically between about 50 nm and 750 nm. Most commonly, EVs will have a size (average diameter) that is up to 5% of the size of the donor cell. Therefore, especially contemplated EVs include those that are shed, exocytosed, or secreted from a cell (e.g., an extracellular vesicle donor cell). In certain contexts, an “extracellular vesicle” can encompass one or a plurality of extracellular vesicles. For example, reference to an “extracellular vesicle” in the context of administering an effective amount or dose of the extracellular vesicle, or preparing a composition comprising the extracellular vesicle, shall be understood to encompass a plurality of extracellular vesicles. In some cases, an “extracellular vesicle” can refer to a population of extracellular vesicles, such as a population of extracellular vesicles having a distinct characteristic or set of characteristics (e.g., a targeting polypeptide, an adapter polypeptide, a heterologous targeting domain, a tissue-specific targeting capability, etc.).Poly(A) tail

[0078] As used herein, the term “poly(A) tail” refers to a continuous or discontinuous sequence of adenylate residues typically located at the 3' end of an RNA molecule. The poly(A) tail may follow the 3', for example, the 3' UTR of mRNA. Such a poly(A) tail may consist of or contain about 20 or more, 25 or more, 40 or more, 60 or more, 80 or more, or 100 or more adenyl (A) nucleotides. The poly(A) tail may contain 20 to 400 adenines (A). For example, the poly(A) tail may contain20 to 300, 40 to 300, 100 to 300, 150 to 300, 180 to 280, 200 to 250, or 200 to 250 adenines. For example, the poly(A) tail may contain about 200-250 adenines, but is not limited thereto. The number of adenine nucleotides present in the poly(A) tail are designated as A(n), where “n” indicates the number of adenines in an mRNA sequence of any integer value ranging from 1-400. For example, a SEQ ID NO: Y representing an RNA molecule with an indicated “A(n)” or “- A(n)” feature possesses a poly(A) tail having a number of 200, 201, 202,... or 250 adenines. The polyA tail may have, based on the number of nucleotides of the poly(A) tail, typically at least 75%, at least 80%, at least 85%, at least 90%, at least 95%, at least 96%, at least 97%, at least 98%, or at least 99% are A nucleotides, but the remaining nucleotides may be nucleotides other than A nucleotides, for example, U, T or C. As an example, the poly(A) tail may contain modifications that delay degradation of mRNA. The poly(A) tail can be added to an RNA molecule having a poly(A) signal, such as aWSGR Ref. No. 54841-719.601Bovine Growth Hormone (bGH) poly(A) signal or other any poly(A) signal commonly known in the art.Extracellular vesicles

[0079] As used here, the term “extracellular vesicle”, abbreviated as EV, can encompass exosomes, microvesicles, apoptotic bodies, and the like. The terms “extracellular vesicles” and “EVs,” as used herein, may in some embodiments refer to a membranous particle having a diameter (or largest dimension where the particles is not spheroid) of between about 10 nm to about 5000 nm, more typically between 30 nm and 1000 nm, and most typically between about 50 nm and 750 nm. Most commonly, EVs will have a size (average diameter) that is up to 5% of the size of the donor cell. Therefore, especially contemplated EVs include those that are shed, exocytosed, or secreted from a cell (e.g., an extracellular vesicle donor cell). In certain contexts, an “extracellular vesicle” may may encompass one or a plurality of extracellular vesicles. For example, reference to an “extracellular vesicle” in the context of administering an effective amount or dose of the extracellular vesicle, or preparing a composition comprising the extracellular vesicle, shall be understood to encompass a plurality of extracellular vesicles. In some cases, an “extracellular vesicle” may refer to a population of extracellular vesicles, such as a population of extracellular vesicles having a distinct characteristic or set of characteristics (e.g., a targeting polypeptide, an adapter polypeptide, a heterologous targeting domain, a tissue-specific targeting capability, etc.).Composition

[0080] In some cases, the term “composition” as used herein can refer to a combination of an active agent (e.g., an extracellular vesicle described herein) and at least one other compound, carrier, or composition, which can be inert (for example, a detectable agent or liquid carrier) or active, such as, but not limited to, a cryopreservation agent, an injectable buffer or saline diluent, or an adjuvant. An extracellular vesicle composition can refere to a composition comprising at least one extracellular vesicle. A composition can comprise extracellular vesicles (e.g., exosomes, microvesicles, apoptotic bodies, or combinations thereof) secreted from any cell capable of producing and secreting vesicular structures, including any of the extracellular vesicle donor cells described herein (e.g., mesenchymal stem cells, fibroblasts, dermal fibroblasts) in a culture medium or buffer described herein.WSGR Ref. No. 54841-719.601The terms “composition”, “product”, “formulation”, “solution”, and “mixture” can be used interchangeably herein.In vivo, ex vivo, and in vitro

[0081] As used herein, the term “zh vivo” is used to describe an event that takes place in a subject’s body.

[0082] As used herein, the term “ex vivo” is used to describe an event that takes place outside of a subject’s body. An “ex vivo” assay cannot be performed directly on a subject. Rather, it is performed upon a sample separate from a subject, such as a biological sample obtained from the subject. Ex vivo is used to describe an event occurring in an intact cell or other type of biological sample outside a subject’s body.As used herein, the term “zh vitro” is used to describe an event that takes place contained in a container for holding a laboratory reagent such that it is separated from the living biological source organism from which the material is obtained. In vitro assays can encompass cellbased assays in which live or dead cells or other biological materials are employed. In vitro assays can also encompass a cell-free assay in which no intact cells are employed.Treating

[0083] The terms “treating” or “treatment” or “to treat” or “alleviating” or “to alleviate” refer to therapeutic measures that cure, slow down, lessen symptoms of, and / or halt progression of a diagnosed, or suspected, pathological condition or disorder. Thus, a subject in need of treatment includes a subject already diagnosed with, or presenting symptoms of, the disorder, as well as those prone to have the disorder, or those in whom the disorder is to be prevented. In some cases, treating may refer to reducing, or ameliorating a disorder and / or symptoms associated therewith, including a condition or disorder associated with tissue degeneration (e.g., a muscular dystrophy, a cardiomyopathy, Duchenne Muscular Dystrophy, Becker Muscular Dystrophy, a dystrophin-related disorder). “Treating” can refer to administration of the therapy (e.g., administration of therapeutic extracellular vesicles) to a subject after the onset, or suspected onset, of a disease (e.g., a muscular dystrophy, a cardiomyopathy, Duchenne Muscular Dystrophy, Becker Muscular Dystrophy, a dystrophin -related disorder). “Treating” includes the concepts of “alleviating”, which refers to lessening the frequency of occurrence or recurrence, or the severity, of any symptoms or other ill effects related to the disease and / or the side effects associated with therapy, including any of theWSGR Ref. No. 54841-719.601therapeutic benefits described herein. The term “treating” may also encompass the concept of “managing” which refers to reducing the severity of a disease or disorder in a patient, e.g., extending the life or prolonging the survivability of a patient with the disease, or delaying its recurrence, e.g., lengthening the period of remission in a patient who had suffered from the disease. It is appreciated that, although not precluded, treating a disorder or condition does not require that the disorder, condition, or symptoms associated therewith be completely eliminated.Prevention

[0084] The terms “prevent” or “prevention” refer to prophylactic or preventative measures that slow down the development of a targeted pathologic condition or disorder. Thus, those in need of prevention include those prone to have the disorder or those in whom the disorder is to be prevented.Therapeutic extracellular vesicle

[0085] The terms “therapeutic extracellular vesicle,” “therapeutic EV,” “therapeutic exosome,” “therapeutic microvesicle,” “therapeutic apoptotic body,” and the like, as used herein, generally refer to an extracellular vesicle (or, where applicable, an exosome, microvesicle, or apoptotic body) that can be used to treat a condition or disorder. Depending on the context in which the terms are used, they may, at times, refer to an extracellular vesicle (or, exosome, microvesicle, or apoptotic body) that can be used to treat a condition or disorder associated with tissue degeneration, such as a tissue dystrophy (e.g., a muscular dystrophy). It shall be understood that reference to a therapeutic extracellular vesicle, or any extracellular vesicle described herein, in the context of administration to a subject.Therapeutic benefit

[0086] As used herein, the term “therapeutic benefit” or “therapeutically effective” refers to anything that promotes or enhances the well-being of the subject with respect to the medical treatment of the condition, including, but not limited to, a reduction in the frequency or severity of signs or symptoms of the underlying disorder (e.g., muscular dystrophy, a cardiomyopathy, Duchenne Muscular Dystrophy, Becker Muscular Dystrophy, a dystrophin-related disorder). In the context of gene therapy, a therapeutic benefit may be manifested as a measurable or observable improvement in the subject following administration of the therapeutic extracellular vesicles of this disclosure (e.g.,WSGR Ref. No. 54841-719.601exosomes, microvesicles). Such improvements may include, without limitation, alleviation of disease-associated symptoms (e.g., reduced fatigue, reduced swelling in extremities, reduced instances and / or severity of arrhythmia or arrhythmic events, increased exercise tolerance (e.g., increased walking distance, increased walking speed, increased running speed, increased duration of running or walking), reduced ventricular enlargement to resemble a more normal ventricular size, and reduced thickening of ventricular walls), or restoration of physiological function. In some cases, therapeutic improvements can include inducing changes in physiological parameters in the subject that more closely resemble the normal physiological state. For example, administration of the therapeutic extracellular vesicles provided and described herein can result in improved physiological parameters in a subject who has a muscular dystrophy, including but not limited to increased dystrophin mRNA levels, increased dystrophin expression levels, increased dystrophin protein levels, increased motor coordination, increased grip strength, increased ambulation, increased walking speed, increased agility, increased muscle mass, decreased QRS prolongation, decreased QRS duration, decreased QTc intervals, decreased QT dispersion, and decreased Tpeak-Tend intervals, reduced resting heart rate, increased cardiac conduction time, and / or slowed progression of cardiac or muscle fibrosis and / or scarring compared to the subject prior to treatment or to an earlier timepoint within a treatment regimen, or compared to a control subject who has a muscular dystrophy but was not administered the therapeutic extracellular vesicles. The therapeutic benefit may be determined by clinical assessment (e.g., NSAA and associated behavioral tasks), subject-reported outcomes, biomarker analysis, imaging, or other objective measures demonstrating an improvement in patient condition relative to pretreatment levels or untreated controls. Improvement may cover any range of change from a measured numerical value indicative of the severity of signs and symptoms for a disease. In some cases, the disease is a muscular dystrophy (e.g., muscular dystrophy, a cardiomyopathy, Duchenne Muscular Dystrophy, Becker Muscular Dystrophy, a dystrophin-related disorder). For example, the improvement may be measured as a percentile change or a fold-change from a stated numerical value prior to treatment or a surgical procedure or as compared to an untreated subject or subject treated with a placebo or vehicle. As a nonlimiting example, the improvement may be measured as a percentile change or a fold-change from a stated numerical value prior to infusion of therapeutic extracellular vesicles. As another example, the improvement be measured as a percentile change or a fold-change from a stated numerical value of a treated subject orWSGR Ref. No. 54841-719.601subject receiving gene therapy as compared to another subject that did not receive treatment or gene therapy. In some cases, an improvement is measured by the absence or reduction of a pathological phenotype following treatment as compared within the same subject prior to treatment or as compared to a diseased or afflicted control subject. In some cases, an improvement is measured by the presence or increase of a non-pathological phenotype following treatment as compared within the same subject prior to treatment or as compared to a diseased or afflicted control subject. In some cases, an improvement is measured by a lesser or statistically insignificant percentile change or a fold-change from a stated numerical value of a treated subject or subject receiving gene therapy as compared to the same subject prior to treatment or as compared to another healthy unafflicted control subject. A therapeutic benefit can refer to eradication or amelioration of symptoms or of an underlying disorder being treated. Also, a therapeutic benefit is achieved with the eradication or amelioration of one or more of the physiological symptoms associated with the underlying disorder such that an improvement is observed in the subject, notwithstanding that the subject can still be afflicted with the underlying disorder. A prophylactic effect can include delaying, preventing, or eliminating the appearance of a disease or condition, delaying or eliminating the onset of symptoms of a disease or condition, slowing, halting, or reversing the progression of a disease or condition, or any combination thereof. A prophylactic benefit, a subject at risk of developing a particular disease, or to a subject reporting one or more of the physiological symptoms of a disease can undergo treatment, even though a diagnosis of this disease cannot have been made. A subject is successfully "treated" if, after receiving a therapeutic amount of a therapeutic agent or drug (e.g., therapeutic extracellular vesicles), the subject shows observable and / or measurable reduction in or absence of one or more signs and symptoms of the particular condition. Reduction of the signs or symptoms of a condition may also be felt or experienced by the subject. A subject is also considered treated if the subject experiences stable condition. In some embodiments, treatment with a therapeutic agent or drug (e.g., therapeutic extracellular vesicles) is effective to result in the subjects having ameliorated symptoms after treatment. These parameters for assessing successful treatment and improvement in the condition are readily measurable by routine procedures familiar to a physician of appropriate skill in the art.WSGR Ref. No. 54841-719.601Effective amount

[0087] The terms “effective amount”, “effective dose”, “therapeutic dose”, and “therapeutically effective amount” are used interchangeably herein to refer to an amount sufficient to effect therapeutic, beneficial, and / or desired results. Such an amount or dose may vary depending on the therapeutic agent used within the subject, as well as subject factors including, but not limited to age, weight, height or general health of the subject in need of treatment. An effective amount can be administered in one or more administrations. An “effective extracellular vesicle” can refer to an extracellular vesicle that, when administered to a subject, results in a degree of improvement in cardiovascular and / or muscular function, ambulation, motor coordination, voluntary muscle movement, breathing, pulmonary function; promotion and / or generation of physiologically active muscle cells, cardiomyocytes, and / or tissues thereof; promotion of dystrophin expression in a subject (e.g., in blood, tissue, muscle, heart); delayed onset and / or severity of cardiac and / or muscular fibrosis or scarring; prolonged lifespan; delayed onset and / or severity of cardiovascular or muscular dysfunction; promotion of exercise tolerance; and other therapeutic benefits described herein. In some instances, the expression “an effective amount” can be used herein in reference to that quantity of extracellular vesicle treatment which can be used to obtain a beneficial or desired result relative to that occurring in an untreated control subject, or the subject prior to administration or treatment with the extracellular vesicle, under suitable conditions of treatment as described herein. A therapeutically effective dose further refers to that amount of the therapeutic agent or drug sufficient to result in amelioration of symptoms, e.g., treatment, healing, prevention, or amelioration of the relevant medical condition, or an increase in rate of treatment, healing, prevention, or amelioration of such conditions. When applied to an individual active ingredient administered alone, a therapeutically effective dose refers to that ingredient alone. When applied to a combination, a therapeutically effective dose refers to combined amounts of the active ingredients that result in the therapeutic effect, whether administered in combination, serially or simultaneously. In some embodiments, an effective dose is characterized as a number of extracellular vesicles or therapeutic mRNA copy numbers administered to a subject. For example, the effective dose can be a mass of mRNA or a mass of extracellular vesicles per unit mass of the subject (e.g., mg / kg). In some cases, an effective dose can be defined as a concentration of mRNA copy number per unit volume of a solution, buffer, medium, or biological sample. An effective dose can, in some embodiments, refer to an amount, such as a unit mass, number of extracellular vesicles, of a therapeutic mRNA, protein or drug delivered by the extracellular vesicles of this disclosure to a subject. In some embodiments, an effectiveWSGR Ref. No. 54841-719.601dose can be expressed as a concentration of a therapeutic protein or drug in a biological sample, medium, buffer, or solution. In some cases, when applied to delivering the therapeutic mRNA, protein, or drug to the subject, an effective dose refers to a mass of the therapeutic protein or drug, a mRNA copy number, or a number of extracellular vesicles per unit mass of the subject.Pharmaceutical acceptable carrier

[0088] The term “pharmaceutically acceptable carrier,” “pharmaceutically acceptable excipient,” “physiologically acceptable carrier,” or “physiologically acceptable excipienf’ refers to a pharmaceutically acceptable material, composition, or vehicle, such as a liquid or solid filler, diluent, excipient, solvent, or encapsulating material. A component is “pharmaceutically acceptable” in the sense of being compatible with the other ingredients of a pharmaceutical formulation. It can also be suitable for use in contact with the tissue or organ of humans and non-human mammals without excessive toxicity, irritation, allergic response, immunogenicity, or other problems or complications, commensurate with a reasonable benefit / risk ratio.Pharmaceutical composition

[0089] The term “pharmaceutical composition” refers to the systems or a mixture of the systems or compositions comprising each component of the systems disclosed herein with other chemical components, such as diluents or carriers. The pharmaceutical composition can facilitate administration of the systems or components of the systems to the subject. Multiple techniques of administering a compound exist in the art including, but not limited to, oral, injection, parenteral, and surgical administration.Formulated

[0090] The terms “formulated”, “prepared”, or “mixed” generally refer to a formulation containing a composition as described and provided within the disclosure which contains a technical element or limitation that renders said composition suitable for use in a subject in need thereof. Such formulations contain additional elements or limitations that further distinguish the composition over other compositions that lack such technical elements or limitations. In some non-limiting examples, the formulation may comprise a therapeutic extracellular vesicle of the present disclosure, or a therapeutic molecule thereof (e.g., full- length dystrophin mRNA).WSGR Ref. No. 54841-719.601Administer

[0091] The terms “administer”, “introduce”, “inject”, and “infuse” are generally used herein to encompass any technical methods of placing compositions of the disclosure (e.g., extracellular vesicles, exosomes) into the body of a subject in need thereof. For example, extracellular vesicles can be administered into the bloodstream of a subject by intravenous injection, parenteral injection, systemic injection, intramuscular injection, local injection, or intracardiac injection. In some cases, the method involves administering extracellular vesicles to a subject via a syringe, infusion device, or catheter. In some cases, the extracellular vesicles are directly injected into the bloodstream of the subject. As a further, non-limiting example, compositions, formulations, and methods of the disclosure may be introduced into the subject as a liquid solution or as a semi-solid biomaterial.Subject

[0092] The terms “patient” or “subject” are used interchangeably herein and encompass mammals. Non-limiting examples of mammals include any member of the mammalian class: humans, non-human primates such as chimpanzees, marmosets, and other apes and monkey species; farm animals such as cattle, horses, sheep, goats, or swine; domestic animals such as rabbits, dogs, and cats; laboratory animals including rodents, such as rats, mice and guinea pigs, and the like.Control Subject

[0093] A “control subject”, as used herein, can provide a reference point for measuring changes in phenotype of the subject, and can be any suitable subject, including any mammalian species such as, for example, a human, non-human primate, rodent, pig, dog, or cat. A control subject can also refer to a biological sample obtained from a subject that can be used to measure phenotypic changes in the subject, such as a sample that is obtained from the subject at an earlier timepoint. The control subject can be, in some cases, a subject that does not have a dystrophin mutation or dysregulated dystrophin expression, a muscular dystrophy, a cardiomyopathy. In some cases, the control subject can be a healthy or otherwise normal individual. A control subject can comprise, but is not limited to, a subject having or suspected of having a dystrophin -related condition, a deficit in muscular or cardiac function, a mutation in the dystrophin gene that results in mRNA and / or protein with reduced cellular function, a muscular dystrophy (e.g.,WSGR Ref. No. 54841-719.601Duchenne Muscular Dystrophy, Becker Muscular Dystrophy, Facioscapulohumeral Muscular Dystrophy, Congenital Muscular Dystrophy, Distal Muscular Dystrophy, Emery-Dreifuss Muscular Dystrophy), or a cardiomyopathy (e.g., X-linked Dilated Cardiomyopathy) who was not administered the therapeutic extracellular vesicles described herein.Engraftment

[0094] The term “engraftment” is used herein to refer to the ability of an extracellular vesicle to deliver therapeutic cargo to a tissue, whether such extracellular vesicle is provided by infusion via a syringe catheter, or other surgical delivery device. The term encompasses all events surrounding or leading up to engraftment, such as tissue homing of an extracellular vesicle within the tissue of interest (e.g., skeletal muscle). The engraftment efficiency or rate of engraftment can be evaluated or quantified using any clinically acceptable parameter as known to those of skill in the art and can include, for example, assessment of mRNA copy number or protein expression levels of a therapeutic extracellular vesicle cargo incorporated into tissue(s) into which the extracellular vesicle has homed, colonized, or become engrafted; or by evaluation of the progress of a subject through disease progression, biodistribution of the extracellular vesicle, or survival of a recipient. In one embodiment, engraftment is determined by measuring a level of a therapeutic extracellular vesicle cargo in peripheral blood, plasma, muscle tissue (e.g., skeletal muscle), or cardiac tissue during a period of time after administration.Determining

[0095] The terms “determining,” “measuring,” “evaluating,” “assessing,” “assaying,” and “analyzing” are often used interchangeably herein to refer to forms of measurement. The terms include determining if an element is present or not (for example, detection). These terms can include quantitative, qualitative or quantitative and qualitative determinations. Assessing can be relative or absolute. “Detecting the presence of’ can include determining the amount of something present in addition to determining whether it is present or absent depending on the context.Monitoring

[0096] As used herein, the term “monitoring”, when used with respect to therapeutic engraftment, refers to the surveying or measurement of a therapeutic extracellular vesicle cargo (e.g., human full-length dystrophin) in a subject relative to administering a dose ofWSGR Ref. No. 54841-719.601the therapeutic extracellular vesicle described herein. Monitoring involves collecting biological samples at multiple time points — such as prior to treatment (baseline) and at one or more post-treatment intervals — and determining, relative to baseline or control values, the level or change in a marker indicative of therapeutic engraftment. Such markers may include, without limitation, genetic chimerism analysis (e.g., percentage of therapeutic extracellular vesicles cargo mRNA or protein), expression of a uniquely identifiable extracellular vesicle marker (e.g., transgene, cell-surface receptor, targeting polypeptide, tissue -targeting domain, adapter polypeptide), or other quantifiable signal attributable to the administered extracellular vesicles. In some cases, the therapeutic extracellular vesicle cargo can be a mRNA molecule, or a protein encoded by the mRNA molecule, or an otherwise quantifiable signal that is associated with an absence of administering the therapeutic extracellular vesicle, including a marker that is indicative of a diseased state of a subject (e.g., dystrophin), whereby various increasing levels of therapeutic extracellular vesicle cargo engraftment would decrease a level of the marker in a subject. Changes in the level of the therapeutic extracellular vesicle cargo, or a nucleic acid molecule or a protein encoded by the cargo, are used to establish engraftment kinetics — including maximal engraftment — and to inform decisions regarding the need for further doses of therapeutic extracellular vesicles. In embodiments, monitoring can comprise serial chimerism or marker-quantification assays at defined intervals to determine whether further therapeutic dosing is required based on plateau or decline in engraftment levels.

[0097] Any methods described herein are modular and not limited to sequential steps.Accordingly, terms such as “first” and “second” do not necessarily imply priority, order of importance, or order of acts.

[0098] Use of absolute or sequential terms, for example, “will,” “will not,” “shall,” “shall not,” “must,” “must not,” “first,” “initially,” “next,” “subsequently,” “before,” “after,” “lastly,” and “finally,” are not meant to limit scope of the present embodiments disclosed herein but as exemplary.Overview

[0099] The present disclosure provides an extracellular vesicle that expresses, encapsulates, or carries a cargo molecule.

[0100] In some cases, the cargo molecule can be an RNA cargo encodes a full-length dystrophin protein. In some cases, the RNA cargo is a mRNA transcribed from aWSGR Ref. No. 54841-719.601dystrophin DNA. In some cases, the RNA cargo is a mRNA transcribed from a human dystrophin gene. In some cases, the RNA cargo is a dystrophin mRNA (e.g., a human dystrophin mRNA), particularly a full-length dystrophin mRNA.

[0101] The extracellular vesicles can be designed to carry a payload, such as a therapeutic, to be delivered to the targeted cell. In some cases, the therapeutic delivered by the extracellular vesicles can include a therapeutic compound (e.g., a therapeutic polynucleotide, therapeutic DNA, therapeutic RNA, or therapeutic mRNA). In some cases, the extracellular vesicles may carry a non -therapeutic compound (e.g., non- therapeutic polynucleotide). In some cases, the therapeutic compound is an RNA cargo described herein, particularly a human dystrophin mRNA (e.g., a full-length human dystrophin mRNA).

[0102] In some cases, the extracellular vesicle (e.g., exosome) further can comprise a targeting polypeptide that mediates delivery of the extracellular vesicle to a target tissue, particularly a muscle tissue (e.g., skeletal muscle, smooth muscle, or cardiac muscle), for the purpose of increasing accumulation of the extracellular vesicles in the target tissue or in cells of the target tissue. In some cases, the targeting polypeptide targets muscle cells. In some cases, the targeting polypeptide targets skeletal muscle cells. In some cases, the targeting polypeptide is a peptide that targets or binds a surface protein of a cell of a target tissue (i.e., targeted cell). The surface protein of the targeted cell can be a receptor, a dystrophin protein, or a myofiber of the targeted cell. For example, the targeting polypeptide can comprise a heterologous targeting domain that can bind to a surface protein, such as a transferrin receptor, an integrin receptor, a cell adhesion molecule, or a proteoglycan (e.g., a dystroglycan, a sacroglycan). Non-limiting examples of heterologous targeting domain of the targeting polypeptides provided herein include a M12 peptide, an RGD motif, a dystrophin-binding peptide, a T7 peptide, a T9 peptide, a ASSLNIA peptide, a cardiomyocyte-specific peptide (CMP), an ischemic myocardiumtargeting polypeptide (IMTP), a cardiac homing peptide (CHP), a CRPPR peptide, or any other targeting polypeptide known in the art to bind to a surface protein of a targeted cell. In some cases, the targeting polypeptide can comprise a T7 peptide. In some cases, the targeting polypeptide can comprise a T9 peptide.

[0103] In some cases, this disclosure provides a composition comprising one or more exosomes (or combination of exosomes and microvesicles) comprising: (1) an adapter polypeptide (e.g., CD47, or protein at least 70% identical to CD47), wherein the adapter polypeptide can comprise a transmembrane domain and an extracellular domain whereinWSGR Ref. No. 54841-719.601the extracellular domain is attached (e.g., covalently attached) to a muscle targeting peptide (e.g., a peptide targeting skeletal muscle, a peptide with an RGD domain as described herein, a peptide include T7 targeting protein, or a peptide including T9 targeting protein) and / or a peptide at least 70%, at least 80%, at least 85%, at least 90%, at least 95%, or at least 99% identical to a muscle -targeting domain described herein) and (2) an RNA molecule (e.g., a mRNA molecule encoding full-length dystrophin protein). In some cases, the muscle targeting polypeptide is directly attached to the exosome, in the absence of an adapter protein. In some embodiments, the muscle -targeting polypeptide can comprise a transmembrane domain that anchors the polypeptide to the membrane of the extracellular vesicle. In some cases, the muscle targeting domain is attached (e.g., covalently linked) to the N terminus of the extracellular domain of the adapter protein. In some cases, it is attached to the C terminus of the extracellular domain of the adapter protein, or to an internal region of the extracellular domain of the adapter protein. In some cases, the exosome is targeted to muscle tissue (e.g., skeletal muscle, cardiac muscle, tibalis anterior, quadriceps, gastrocenemius, or triceps) in a preferential manner, particularly preferential over its targeting to diaphragm, heart, liver, spleen, lung, kidney, brain, or any combination thereof (including at least two of such tissue, at least three of such tissue, at least four of such tissue, at least five of such tissue, at least six of such tissue, or seven of such tissue). In some cases, the extracellular vesicle is significantly less targeted to lung tissue when it includes the muscle -targeting vector compared to when it lacks the muscle- targeting polypeptide.

[0104] In another aspect, this disclosure provides an extracellular vesicle that can comprise an exogenous messenger RNA (mRNA) encoding a full-length dystrophin protein; and a targeting polypeptide configured to bind to a targeted cell.

[0105] In yet another aspect, an extracellular vesicle is provided, wherein the extracellular vesicle can comprise a targeting polypeptide comprising a T7 peptide configured to deliver an extracellular vesicle of the extracellular vesicle to a skeletal muscle cell.

[0106] An extracellular vesicle comprising the extracellular vesicle of any one of the preceding claims, wherein the extracellular vesicle can comprise a targeting polypeptide having a T9 peptide configured to deliver an extracellular vesicle of the extracellular vesicle to a skeletal muscle cell.

[0107] Further provided herein are polynucleotides comprising a nucleic acid sequence encoding a therapeutic cargo molecule. In some cases, the therapeutic cargo molecule can be a full-length dystrophin protein. In some instances, the therapeutic cargo molecule canWSGR Ref. No. 54841-719.601comprise a nucleic acid sequence having the nucleic acid of any one of SEQ ID NOs: 1- 6.

[0108] An additional aspect described herein is a cell comprising a polynucleotide that encodes a therapeutic cargo molecule of this disclosure. In some cases, the cell can be an extracellular vesicle donor cell described throughout this disclosure that has been electroporated or transfected with any polynucleotide comprising a therapeutic cargo, as described herein.

[0109] In yet another aspect, provided and described herein is a protein comprising a targeting polypeptide.

[0110] Also described herein are compositions comprising the extracellular vesicle(s), which can comprise homogenous or heterogenous pluralities of extracellular vesicles. In some embodiments, the extracellular vesicles can be targeting extracellular vesicles that comprise a targeting polypeptide. In other cases, the extracellular vesicles can comprise non-targeting extracellular vesicles which lack heterologous targeting domains that would otherwise direct the extracellular vesicles to a particular tissue or cell type.

[0111] Also provided and described herein are unit doses, kits, and devices that can be used for delivering or administering the extracellular vesicles and compositions disclosed herein.

[0112] This disclosure also discloses pharmaceutical compositions comprising extracellular vesicles that can be used to treat the muscle disorders described, such as muscular dystrophies, including Duchenne Muscular Dystrophy.

[0113] The extracellular vesicles provided herein can be produced by a number of methods and approaches. One approach provided herein involves introducing at least one heterologous polynucleotide such as a vector (e.g., plasmid, viral vector, or DNA) into an extracellular vesicle donor cell, where the at least one heterologous polynucleotide encodes a dystrophin-glycoprotein complex protein or mRNA for such complex (or a portion of such complex), particularly a dystrophin RNA. Introduction of the vector (e.g., DNA plasmid) can be via a variety of methods, including transfection, calcium phosphate transfection, electroporation, nanoelectroporation, lipofection, cellular nanoelectroporation, viral delivery, or other methods. In some cases, the extracellular vesicle donor cell is a primary cell (e.g., a primary adherent cell, a primary multinucleated cell, a primary mononucleated cell, a primary contractile cell, a primary muscle cell, a primary skeletal muscle cell, a primary cardiomyocyte, a primary smooth muscle cell, a myofibroblast, etc.). In some cases, the extracellular vesicle donor cell is from a cell lineWSGR Ref. No. 54841-719.601(e.g., human cell, human myocyte line, mammalian myocyte line, human cardiomyocyte line, a mammalian cardiomyocyte line, a smooth muscle cell line, etc.). In some cases, the extracellular vesicle donor cell is not genetically modified prior to the nanoelectroporation. In some cases, the extracellular vesicle donor cell is genetically modified prior to the nano electrop oration. In some cases, this disclosure provides methods of producing large number of exosomes containing a high quantity of mRNA transcripts, even from cells with otherwise low basal secretion of exosomes.

[0114] Described herein are methods of treating a tissue dystrophy (e.g., a muscular dystrophy) comprising administering one or more of the extracellular vesicles provided in this disclosure. The methods provided herein are especially useful for reversing, reducing, improving, or ameliorating the progression or development of a tissue dystrophy or a symptom thereof.

[0115] In some cases, the methods provided herein reduce or ameliorate muscular degeneration or weakness in a subject (e.g., reduce loss of muscle mass, reduce progression of muscular degeneration, improve muscle strength, reduce or ameliorate cardiac degeneration, reduce or halt development of cardiomyopathy, heart failure, or cardia arrhythmia) over a period of time, such as at least 1, 2, 3, or 4 weeks following administration of extracellular vesicles provided herein. In some cases, the methods provided herein can be used for improving or maintaining muscular function or muscle mass in a subject. In some cases, the extracellular vesicles comprise at least one therapeutic polynucleotide (e.g., therapeutic mRNA, etc.). In some cases, the extracellular vesicles comprising therapeutic polynucleotides can be obtained by nanoelectroporating at least one extracellular vesicle donor cell with at least a first vector (e.g. a plasmid), wherein the first vector encodes a therapeutic polypeptide (e.g., dystrophin protein). In some embodiments, the therapeutic polynucleotide can comprise an RNA encoding a protein of the dystrophin-glycoprotein complex (e.g., dystrophin RNA). In some cases, the therapeutic polynucleotide is a dystrophin mRNA, particularly a full-length dystrophin mRNA. In some cases, the therapeutic polynucleotide is a human dystrophin RNA (e.g., human dystrophin mRNA). The therapeutic polynucleotide can, in some cases, increase the amount of dystrophin mRNA and / or dystrophin protein in cells of the subject, wherein the subject has or is suspected of having a dystrophy, such as a muscular dystrophy (e.g., Duchenne muscular dystrophy (DMD), Becker muscular dystrophy (BMD), etc.).WSGR Ref. No. 54841-719.601

[0116] In some instances, the first vectors can be expressed in the extracellular vesicle donor cells to obtain the therapeutic polynucleotides. In some embodiments, the extracellular vesicles released from the extracellular vesicle donor cells comprise the therapeutic polynucleotides. In some cases, the extracellular vesicles are collected and systematically administered to the subject (e.g., via intraperitoneal injection, intravenous injection, etc.). In some cases, the extracellular vesicles are collected and locally administered to the subject (e.g., via subcutaneous injection, intramuscular injection, intraventricular injection, etc.). In some cases, the extracellular vesicles are administered into a muscle of the subject, such as a skeletal muscle, a cardiac muscle, or a smooth muscle of the subject. In some cases, the extracellular vesicles are administered into a quadricep, a tibialis anterior, a gastrocnemius, a diaphragm, a heart, a cardiac ventricle, a cardiac atrium, an intestine, or any muscle of a face, shoulder, arm, chest, back, calves, or lower extremity of the subject.Extracellular Vesicle Cargo

[0117] The present disclosure provides an extracellular vesicle that expresses, encapsulates, or carries a cargo molecule. The cargo molecule of the extracellular vesicle can comprise a therapeutic cargo molecule, including a nucleic acid molecule or a polypeptide. The nucleic acid molecule can, in some cases, comprise a therapeutic nucleic acid molecule. In some cases, the nucleic acid molecule can comprise a DNA molecule or an RNA molecule. In some cases, the DNA molecule can comprise a plasmid, a vector, or a viral vector that encodes an extracellular vesicle cargo molecule, such as a therapeutic extracellular vesicle cargo molecule in the form of an RNA molecule or polypeptide. In some cases, the cargo molecule can be an exogenous nucleic acid or polypeptide. In some embodiments, the cargo molecule can be a synthetic nucleic acid molecule or polypeptide. In some embodiments, the extracellular vesicle cargo molecule is encoded by a plasmid encoding a protein that interacts with a protein of a dystrophin -glycoprotein complex.

[0118] The extracellular vesicle can be an exosome, a microvesicle, or an apoptotic body. In certain embodiments, the extracellular vesicle can be an exosome. In some cases, the extracellular vesicle can express CD9, CD63, TSG101, or ARF6 protein markers. The extracellular vesicle can, in some instances, be from about 10 nanometers (nm) to about 10 micrometers (µm) in diameter, particularly from about 30 nm to about 500 nm in diameter.WSGR Ref. No. 54841-719.601

[0119] The RNA cargo that is expressed, encapsulated, or carried by the extracellular vesicle can encode a therapeutic molecule, such as a therapeutic polypeptide. In some cases, the RNA cargo can be a messenger RNA (mRNA) molecule. The RNA molecule can be an exogenous RNA molecule, such as an exogenous mRNA molecule. In some cases, the RNA cargo can be one or more molecules of messenger RNA (mRNA). In some cases, the RNA cargo encodes a protein that binds to a cytoskeletal protein of a cell. In some cases, the RNA cargo encodes a protein that binds to a protein that interacts with a cell adhesion molecule. In some cases, the RNA cargo encodes a protein that binds to a signaling protein of a dystrophin of a cell. In some cases, the RNA cargo encodes a protein that interacts with a protein of a dystrophin-glycoprotein complex (e.g., a dystroglycan). In some cases, the RNA cargo encodes a protein of a dystrophin- glycoprotein complex, particularly a dystrophin protein or a fragment thereof. In some cases, the RNA cargo can comprise a dystrophin mRNA that is at least 2 kilobases (kbs), at least 4 kbs, at least 5 kbs, at least 7 kbs, at least 10 kbs, at least 12 kbs, at least 13 kbs, or at least 14 kbs in length. In some cases, the RNA cargo can be a dystrophin mRNA molecule that is about 13.7 kbs in length. In some cases, the RNA cargo can be a full- length dystrophin mRNA. The RNA cargo can be, in some cases, a full-length human dystrophin mRNA molecule.

[0120] In some embodiments, the RNA cargo can be an mRNA molecule encoding a dystrophin protein. The mRNA molecule can encode a dystrophin protein, including a human dystrophin protein. In some cases, the mRNA molecule can encode a full-length dystrophin protein. In a particular embodiment, the mRNA molecule can encode a full- length human dystrophin protein.

[0121] The RNA molecule can encode a dystrophin protein having a dystrophin amino acid sequence (e.g., UniProt P11532-1, NP_003997.2). In some cases, the dystrophin protein can include an amino acid substitution. For example, the amino acid substitution in the dystrophin protein can be compared to a reference dystrophin amino acid sequence, such as that of a canonical dystrophin sequence or a functional fragment thereof, or a functional variant thereof. Non-limiting examples of dystrophin variants can include Dp427c (Uniprot P11532-4), Dp427m (UniProt P11532-1), Dp427p (UniProt P11532-11), Dp260-1 (UniProt P11532-2), and Dp260-2 (UniProt P11532-3). The dystrophin protein can comprise at least 1, 2, 3, 4, 5, 6, 7, 8, 9, 10, or more amino acid substitutions relative to a reference an amino acid sequence of a canonical dystrophin protein or functional fragment thereof.WSGR Ref. No. 54841-719.601

[0122] In some embodiments, the RNA molecule can encode a dystrophin protein having an amino acid substitution at an amino acid residue corresponding to an aspartate residue at position 882 of a reference dystrophin protein consisting of the amino acid sequence set forth in SEQ ID NO: 2. In some embodiments, the RNA molecule can encode a dystrophin protein having an aspartate -to-glycine amino acid substitution at an amino acid residue corresponding to the aspartate residue at position 882 (D882G) of a dystrophin protein consisting of the amino acid sequence set forth in SEQ ID NO: 2.

[0123] In some embodiments, the RNA molecule can encode a dystrophin protein having an amino acid substitution at an amino acid residue corresponding to the lysine residue at position 2366 of a reference dystrophin protein consisting of the amino acid sequence set forth in SEQ ID NO: 2. In some embodiments, the RNA molecule can encode a dystrophin protein having a lysine -to -glutamine amino acid substitution at an amino acid residue corresponding to the lysine residue at position 2366 (K2366Q) of a dystrophin protein consisting of the amino acid sequence set forth in SEQ ID NO: 2.

[0124] In some embodiments, the RNA molecule can encode a dystrophin protein having an arginine -to -glutamine amino acid substitution at an amino acid residue corresponding to the arginine residue at position 2937 (R2937Q) of a reference dystrophin protein consisting of the amino acid sequence set forth in SEQ ID NO: 2.

[0125] In some embodiments, the RNA molecule can encode a dystrophin protein having a plurality of amino acid substitutions of a dystrophin protein consisting of the amino acid sequence set forth in SEQ ID NO: 2. In some embodiments, the RNA molecule can encode a dystrophin protein having D882G, K2366Q, and R2937Q amino acid substitutions of a dystrophin protein consisting of the amino acid sequence set forth in SEQ ID NO: 2.

[0126] In some cases, the RNA cargo can comprise a polynucleotide encoding a polypeptide having an amino acid sequence that has at least 70%, at least 80%, at least 90%, at least 95%, at least 96%, or at least 99% sequence identity to an amino acid sequence set forth in SEQ ID NO: 1. In some embodiments, the RNA cargo can comprise a nucleic acid molecule encoding a dystrophin polypeptide having an amino acid sequence that has at least 50%, 60%, 70%, 75%, 80%, 85%, 90%, 95%, 96%, 97%, 98%, 99%, or 100% sequence identity to an amino acid sequence set forth in SEQ ID NO: 1. In some embodiments, the RNA cargo can comprise a nucleic acid molecule encoding a dystrophin polypeptide having an amino acid sequence that is at least 50%, 60%, 70%, 75%, 80%, 85%, 90%, 95%, 96%, 97%, 98%, 99%, or 100% identical to the amino acid sequence setWSGR Ref. No. 54841-719.601forth in SEQ ID NO: 1. In some embodiments, the RNA cargo can comprise a nucleic acid molecule encoding a dystrophin polypeptide having an amino acid sequence that comprises the amino acid sequence set forth in SEQ ID NO: 1. In some embodiments, the RNA cargo can comprise a nucleic acid molecule encoding a dystrophin polypeptide having an amino acid sequence that consists of the amino acid sequence set forth in SEQ ID NO: 1.

[0127] In some cases, the RNA cargo can comprise a polynucleotide encoding a polypeptide having an amino acid sequence that has at least 70%, at least 80%, at least 90%, at least 95%, at least 96%, or at least 99% sequence identity to an amino acid sequence set forth in SEQ ID NO: 2 (P11532-1, UniProt). In some embodiments, the RNA cargo can comprise a nucleic acid molecule encoding a dystrophin polypeptide having an amino acid sequence that has at least 50%, 60%, 70%, 75%, 80%, 85%, 90%, 95%, 96%, 97%, 98%, 99%, or 100% sequence identity to an amino acid sequence set forth in SEQ ID NO: 2. In some embodiments, the RNA cargo can comprise a nucleic acid molecule encoding a dystrophin polypeptide having an amino acid sequence that is at least 50%, 60%, 70%, 75%, 80%, 85%, 90%, 95%, 96%, 97%, 98%, 99%, or 100% identical to the amino acid sequence set forth in SEQ ID NO: 2. In some embodiments, the RNA cargo can comprise a nucleic acid molecule encoding a dystrophin polypeptide having an amino acid sequence that comprises the amino acid sequence set forth in SEQ ID NO: 2. In some embodiments, the RNA cargo can comprise a nucleic acid molecule encoding a dystrophin polypeptide having an amino acid sequence that consists of the amino acid sequence set forth in SEQ ID NO: 2.

[0128] In some cases, the RNA cargo can comprise a polynucleotide encoding a polypeptide having an amino acid sequence that has at least 70%, at least 80%, at least 90%, at least 95%, at least 96%, or at least 99% sequence identity to an amino acid sequence set forth in SEQ ID NO: 3 (UniProt P11532-4). In some embodiments, the RNA cargo can comprise a nucleic acid molecule encoding a dystrophin polypeptide having an amino acid sequence that has at least 50%, 60%, 70%, 75%, 80%, 85%, 90%, 95%, 96%, 97%, 98%, 99%, or 100% sequence identity to an amino acid sequence set forth in SEQ ID NO: 3. In some embodiments, the RNA cargo can comprise a nucleic acid molecule encoding a dystrophin polypeptide having an amino acid sequence that is at least 50%, 60%, 70%, 75%, 80%, 85%, 90%, 95%, 96%, 97%, 98%, 99%, or 100% identical to the amino acid sequence set forth in SEQ ID NO: 3. In some embodiments, the RNA cargo can comprise a nucleic acid molecule encoding a dystrophin polypeptide having an amino acid sequenceWSGR Ref. No. 54841-719.601that comprises the amino acid sequence set forth in SEQ ID NO: 3. In some embodiments, the RNA cargo can comprise a nucleic acid molecule encoding a dystrophin polypeptide having an amino acid sequence that consists of the amino acid sequence set forth in SEQ ID NO: 3.

[0129] In some cases, the RNA cargo can comprise a polynucleotide encoding a polypeptide having an amino acid sequence that has at least 70%, at least 80%, at least 90%, at least 95%, at least 96%, or at least 99% sequence identity to an amino acid sequence set forth in SEQ ID NO: 4 (UniProt P11532-11). In some embodiments, the RNA cargo can comprise a nucleic acid molecule encoding a dystrophin polypeptide having an amino acid sequence that has at least 50%, 60%, 70%, 75%, 80%, 85%, 90%, 95%, 96%, 97%, 98%, 99%, or 100% sequence identity to an amino acid sequence set forth in SEQ ID NO: 4. In some embodiments, the RNA cargo can comprise a nucleic acid molecule encoding a dystrophin polypeptide having an amino acid sequence that is at least 50%, 60%, 70%, 75%, 80%, 85%, 90%, 95%, 96%, 97%, 98%, 99%, or 100% identical to the amino acid sequence set forth in SEQ ID NO: 4. In some embodiments, the RNA cargo can comprise a nucleic acid molecule encoding a dystrophin polypeptide having an amino acid sequence that comprises the amino acid sequence set forth in SEQ ID NO: 4. In some embodiments, the RNA cargo can comprise a nucleic acid molecule encoding a dystrophin polypeptide having an amino acid sequence that consists of the amino acid sequence set forth in SEQ ID NO: 4.

[0130] In some cases, the RNA cargo can comprise a polynucleotide encoding a polypeptide having an amino acid sequence that has at least 70%, at least 80%, at least 90%, at least 95%, at least 96%, or at least 99% sequence identity to an amino acid sequence set forth in SEQ ID NO: 4 (UniProt P11532-2). In some embodiments, the RNA cargo can comprise a nucleic acid molecule encoding a dystrophin polypeptide having an amino acid sequence that has at least 50%, 60%, 70%, 75%, 80%, 85%, 90%, 95%, 96%, 97%, 98%, 99%, or 100% sequence identity to an amino acid sequence set forth in SEQ ID NO: 4. In some embodiments, the RNA cargo can comprise a nucleic acid molecule encoding a dystrophin polypeptide having an amino acid sequence that is at least 50%, 60%, 70%, 75%, 80%, 85%, 90%, 95%, 96%, 97%, 98%, 99%, or 100% identical to the amino acid sequence set forth in SEQ ID NO: 4. In some embodiments, the RNA cargo can comprise a nucleic acid molecule encoding a dystrophin polypeptide having an amino acid sequence that comprises the amino acid sequence set forth in SEQ ID NO: 4. In some embodiments, the RNA cargo can comprise a nucleic acid molecule encoding a dystrophin polypeptideWSGR Ref. No. 54841-719.601having an amino acid sequence that consists of the amino acid sequence set forth in SEQ ID NO: 4.

[0131] In some cases, the RNA cargo can comprise a polynucleotide encoding a polypeptide having an amino acid sequence that has at least 70%, at least 80%, at least 90%, at least 95%, at least 96%, or at least 99% sequence identity to an amino acid sequence set forth in SEQ ID NO: 5 (UniProt P11532-2). In some embodiments, the RNA cargo can comprise a nucleic acid molecule encoding a dystrophin polypeptide having an amino acid sequence that has at least 50%, 60%, 70%, 75%, 80%, 85%, 90%, 95%, 96%, 97%, 98%, 99%, or 100% sequence identity to an amino acid sequence set forth in SEQ ID NO: 5. In some embodiments, the RNA cargo can comprise a nucleic acid molecule encoding a dystrophin polypeptide having an amino acid sequence that is at least 50%, 60%, 70%, 75%, 80%, 85%, 90%, 95%, 96%, 97%, 98%, 99%, or 100% identical to the amino acid sequence set forth in SEQ ID NO: 5. In some embodiments, the RNA cargo can comprise a nucleic acid molecule encoding a dystrophin polypeptide having an amino acid sequence that comprises the amino acid sequence set forth in SEQ ID NO: 5. In some embodiments, the RNA cargo can comprise a nucleic acid molecule encoding a dystrophin polypeptide having an amino acid sequence that consists of the amino acid sequence set forth in SEQ ID NO: 5.

[0132] In some cases, the RNA cargo can comprise a polynucleotide encoding a polypeptide having an amino acid sequence that has at least 70%, at least 80%, at least 90%, at least 95%, at least 96%, or at least 99% sequence identity to an amino acid sequence set forth in SEQ ID NO: 6 (UniProt P11532-3). In some embodiments, the RNA cargo can comprise a nucleic acid molecule encoding a dystrophin polypeptide having an amino acid sequence that has at least 50%, 60%, 70%, 75%, 80%, 85%, 90%, 95%, 96%, 97%, 98%, 99%, or 100% sequence identity to an amino acid sequence set forth in SEQ ID NO: 6. In some embodiments, the RNA cargo can comprise a nucleic acid molecule encoding a dystrophin polypeptide having an amino acid sequence that is at least 50%, 60%, 70%, 75%, 80%, 85%, 90%, 95%, 96%, 97%, 98%, 99%, or 100% identical to the amino acid sequence set forth in SEQ ID NO: 6. In some embodiments, the RNA cargo can comprise a nucleic acid molecule encoding a dystrophin polypeptide having an amino acid sequence that comprises the amino acid sequence set forth in SEQ ID NO: 6. In some embodiments, the RNA cargo can comprise a nucleic acid molecule encoding a dystrophin polypeptide having an amino acid sequence that consists of the amino acid sequence set forth in SEQ ID NO: 6.WSGR Ref. No. 54841-719.601

[0133] In some embodiments, the RNA cargo can be an mRNA molecule having a nucleic acid sequence that encodes a dystrophin polypeptide. In some embodiments, the RNA cargo can be a can be an mRNA molecule having a nucleic acid sequence that encodes a full-length dystrophin polypeptide (e.g., a human full-length dystrophin polypeptide).

[0134] In some embodiments, the RNA cargo can be an mRNA molecule comprising a nucleic acid sequence having at least 50%, 60%, 70%, 75%, 80%, 85%, 90%, 95%, 96%, 97%, 98%, 99%, or 100% sequence identity to an mRNA molecule comprising the nucleic acid sequence set forth in SEQ ID NO: 21. In some embodiments, the RNA cargo can be an mRNA molecule having a nucleic acid sequence having at least 50%, 60%, 70%, 75%, 80%, 85%, 90%, 95%, 96%, 97%, 98%, 99%, or 100% sequence identity to an mRNA nucleic acid sequence set forth in SEQ ID NO: 21. In some embodiments, the RNA cargo can be an mRNA molecule having a nucleic acid sequence that is at least 50%, 60%, 70%, 75%, 80%, 85%, 90%, 95%, 96%, 97%, 98%, 99%, or 100% identical to the nucleic acid sequence set forth in SEQ ID NO: 21. In some embodiments, the RNA cargo can be an mRNA molecule consisting of the nucleic acid sequence set forth in SEQ ID NO: 21.

[0135] In some embodiments, the RNA cargo can be an mRNA molecule comprising a nucleic acid sequence having at least 50%, 60%, 70%, 75%, 80%, 85%, 90%, 95%, 96%, 97%, 98%, 99%, or 100% sequence identity to an mRNA molecule comprising the nucleic acid sequence set forth in SEQ ID NO: 22 (Ref. Seq. NM 004006.3). In some embodiments, the RNA cargo can be an mRNA molecule having a nucleic acid sequence having at least 50%, 60%, 70%, 75%, 80%, 85%, 90%, 95%, 96%, 97%, 98%, 99%, or 100% sequence identity to an mRNA molecule comprising the nucleic acid sequence set forth in SEQ ID NO: 22. In some embodiments, the RNA cargo can be an mRNA molecule having a nucleic acid sequence that is at least 50%, 60%, 70%, 75%, 80%, 85%, 90%, 95%, 96%, 97%, 98%, 99%, or 100% identical to an mRNA molecule comprising the nucleic acid sequence set forth in SEQ ID NO: 22. In some embodiments, the RNA cargo can be an mRNA molecule having a nucleic acid sequence consisting of an mRNA molecule comprising the nucleic acid sequence set forth in SEQ ID NO: 22.

[0136] In some embodiments, the RNA cargo can be an mRNA molecule comprising a nucleic acid sequence having at least 50%, 60%, 70%, 75%, 80%, 85%, 90%, 95%, 96%, 97%, 98%, 99%, or 100% sequence identity to an mRNA molecule comprising the nucleic acid sequence set forth in SEQ ID NO: 23 (Ref. Seq. NM 000109.3). In some embodiments, the RNA cargo can be an mRNA molecule having a nucleic acid sequence having at least 50%, 60%, 70%, 75%, 80%, 85%, 90%, 95%, 96%, 97%, 98%, 99%, orWSGR Ref. No. 54841-719.601100% sequence identity to an mRNA molecule comprising the nucleic acid sequence set forth in SEQ ID NO: 23. In some embodiments, the RNA cargo can be an mRNA molecule having a nucleic acid sequence that is at least 50%, 60%, 70%, 75%, 80%, 85%, 90%, 95%, 96%, 97%, 98%, 99%, or 100% identical to an mRNA molecule comprising the nucleic acid sequence set forth in SEQ ID NO: 23. In some embodiments, the RNA cargo can be an mRNA molecule having a nucleic acid sequence consisting of an mRNA molecule comprising the nucleic acid sequence set forth in SEQ ID NO: 23.

[0137] In some embodiments, the RNA cargo can be an mRNA molecule comprising a nucleic acid sequence having at least 50%, 60%, 70%, 75%, 80%, 85%, 90%, 95%, 96%, 97%, 98%, 99%, or 100% sequence identity to an mRNA molecule comprising the nucleic acid sequence set forth in SEQ ID NO: 24 (Ref. Seq. NM 004009.3). In some embodiments, the RNA cargo can be an mRNA molecule having a nucleic acid sequence having at least 50%, 60%, 70%, 75%, 80%, 85%, 90%, 95%, 96%, 97%, 98%, 99%, or 100% sequence identity to an mRNA molecule comprising the nucleic acid sequence set forth in SEQ ID NO: 24. In some embodiments, the RNA cargo can be an mRNA molecule having a nucleic acid sequence that is at least 50%, 60%, 70%, 75%, 80%, 85%, 90%, 95%, 96%, 97%, 98%, 99%, or 100% identical to an mRNA molecule comprising the nucleic acid sequence set forth in SEQ ID NO: 24. In some embodiments, the RNA cargo can be an mRNA molecule having a nucleic acid sequence consisting of an mRNA molecule comprising the nucleic acid sequence set forth in SEQ ID NO: 24.

[0138] In some embodiments, the RNA cargo can be an mRNA molecule comprising a nucleic acid sequence having at least 50%, 60%, 70%, 75%, 80%, 85%, 90%, 95%, 96%, 97%, 98%, 99%, or 100% sequence identity to an mRNA molecule comprising the nucleic acid sequence set forth in SEQ ID NO: 25 (Ref. Seq. NM 004011.4). In some embodiments, the RNA cargo can be an mRNA molecule having a nucleic acid sequence having at least 50%, 60%, 70%, 75%, 80%, 85%, 90%, 95%, 96%, 97%, 98%, 99%, or 100% sequence identity to an mRNA molecule comprising the nucleic acid sequence set forth in SEQ ID NO: 25. In some embodiments, the RNA cargo can be an mRNA molecule having a nucleic acid sequence that is at least 50%, 60%, 70%, 75%, 80%, 85%, 90%, 95%, 96%, 97%, 98%, 99%, or 100% identical to an mRNA molecule comprising the nucleic acid sequence set forth in SEQ ID NO: 21. In some embodiments, the RNA cargo can be an mRNA molecule having a nucleic acid sequence that is at least 50%, 60%, 70%, 75%, 80%, 85%, 90%, 95%, 96%, 97%, 98%, 99%, or 100% identical to an mRNA molecule comprising the nucleic acid sequence set forth in SEQ ID NO: 22. InWSGR Ref. No. 54841-719.601some embodiments, the RNA cargo can be an mRNA molecule having a nucleic acid sequence consisting of an mRNA molecule comprising the nucleic acid sequence set forth in SEQ ID NO: 25.

[0139] In some embodiments, the RNA cargo can be an mRNA molecule comprising a nucleic acid sequence having at least 50%, 60%, 70%, 75%, 80%, 85%, 90%, 95%, 96%, 97%, 98%, 99%, or 100% sequence identity to an mRNA molecule comprising the nucleic acid sequence set forth in SEQ ID NO: 26 (NCBI Reference Sequence: NM_004012.4). In some embodiments, the RNA cargo can be an mRNA molecule having a nucleic acid sequence having at least 50%, 60%, 70%, 75%, 80%, 85%, 90%, 95%, 96%, 97%, 98%, 99%, or 100% sequence identity to an mRNA molecule comprising the nucleic acid sequence set forth in SEQ ID NO: 26. In some embodiments, the RNA cargo can be an mRNA molecule having a nucleic acid sequence that is at least 50%, 60%, 70%, 75%, 80%, 85%, 90%, 95%, 96%, 97%, 98%, 99%, or 100% identical to an mRNA molecule comprising the nucleic acid sequence set forth in SEQ ID NO: 26. In some embodiments, the RNA cargo can be an mRNA molecule having a nucleic acid sequence consisting of an mRNA molecule comprising the nucleic acid sequence set forth in SEQ ID NO: 26.

[0140] In some cases, the RNA cargo can be encoded by a DNA molecule of a dystrophin gene. The RNA cargo can, in some cases, be encoded by a DNA molecule of a human dystrophin gene, such as a full-length human dystrophin gene or a functional variant thereof.

[0141] In some cases, the RNA cargo can be encoded by a DNA molecule comprising a nucleic acid sequence having at least 50%, 60%, 70%, 75%, 80%, 85%, 90%, 95%, 96%, 97%, 98%, 99%, or 100% sequence identity to a DNA molecule having the nucleic acid sequence set forth in SEQ ID NO: 31. In some cases, the RNA cargo can be encoded by a DNA molecule comprising a nucleic acid sequence that is at least 50%, 60%, 70%, 75%, 80%, 85%, 90%, 95%, 96%, 97%, 98%, 99%, or 100% identical to a DNA molecule having the nucleic acid sequence set forth in SEQ ID NO: 31. In some cases, the RNA cargo can be encoded by a DNA molecule comprising the nucleic acid sequence set forth in SEQ ID NO: 31. In some cases, the RNA cargo can be encoded by a DNA molecule consisting of the nucleic acid sequence set forth in SEQ ID NO: 31.

[0142] In some cases, the RNA cargo can be encoded by a DNA molecule comprising a nucleic acid sequence having at least 50%, 60%, 70%, 75%, 80%, 85%, 90%, 95%, 96%, 97%, 98%, 99%, or 100% sequence identity to a DNA molecule having the nucleic acid sequence set forth in SEQ ID NO: 32. In some cases, the RNA cargo can be encoded by aWSGR Ref. No. 54841-719.601DNA molecule comprising a nucleic acid sequence that is at least 50%, 60%, 70%, 75%, 80%, 85%, 90%, 95%, 96%, 97%, 98%, 99%, or 100% identical to a DNA molecule having the nucleic acid sequence set forth in SEQ ID NO: 32. In some cases, the RNA cargo can be encoded by a DNA molecule comprising the nucleic acid sequence set forth in SEQ ID NO: 32. In some cases, the RNA cargo can be encoded by a DNA molecule consisting of the nucleic acid sequence set forth in SEQ ID NO: 32.

[0143] In some cases, the RNA cargo can be encoded by a DNA molecule comprising a nucleic acid sequence having at least 50%, 60%, 70%, 75%, 80%, 85%, 90%, 95%, 96%, 97%, 98%, 99%, or 100% sequence identity to a DNA molecule having the nucleic acid sequence set forth in SEQ ID NO: 33. In some cases, the RNA cargo can be encoded by a DNA molecule comprising a nucleic acid sequence that is at least 50%, 60%, 70%, 75%, 80%, 85%, 90%, 95%, 96%, 97%, 98%, 99%, or 100% identical to a DNA molecule having the nucleic acid sequence set forth in SEQ ID NO: 33. In some cases, the RNA cargo can be encoded by a DNA molecule comprising the nucleic acid sequence set forth in SEQ ID NO: 33. In some cases, the RNA cargo can be encoded by a DNA molecule consisting of the nucleic acid sequence set forth in SEQ ID NO: 33.

[0144] In some cases, the RNA cargo can be encoded by a DNA molecule comprising a nucleic acid sequence having at least 50%, 60%, 70%, 75%, 80%, 85%, 90%, 95%, 96%, 97%, 98%, 99%, or 100% sequence identity to a DNA molecule having the nucleic acid sequence set forth in SEQ ID NO: 34. In some cases, the RNA cargo can be encoded by a DNA molecule comprising a nucleic acid sequence that is at least 50%, 60%, 70%, 75%, 80%, 85%, 90%, 95%, 96%, 97%, 98%, 99%, or 100% identical to a DNA molecule having the nucleic acid sequence set forth in SEQ ID NO: 34. In some cases, the RNA cargo can be encoded by a DNA molecule comprising the nucleic acid sequence set forth in SEQ ID NO: 34. In some cases, the RNA cargo can be encoded by a DNA molecule consisting of the nucleic acid sequence set forth in SEQ ID NO: 34.

[0145] In some cases, the RNA cargo can be encoded by a DNA molecule comprising a nucleic acid sequence having at least 50%, 60%, 70%, 75%, 80%, 85%, 90%, 95%, 96%, 97%, 98%, 99%, or 100% sequence identity to a DNA molecule having the nucleic acid sequence set forth in SEQ ID NO: 35. In some cases, the RNA cargo can be encoded by a DNA molecule comprising a nucleic acid sequence that is at least 50%, 60%, 70%, 75%, 80%, 85%, 90%, 95%, 96%, 97%, 98%, 99%, or 100% identical to a DNA molecule having the nucleic acid sequence set forth in SEQ ID NO: 35. In some cases, the RNA cargo can be encoded by a DNA molecule comprising the nucleic acid sequence set forthWSGR Ref. No. 54841-719.601in SEQ ID NO: 35. In some cases, the RNA cargo can be encoded by a DNA molecule consisting of the nucleic acid sequence set forth in SEQ ID NO: 35.

[0146] In some cases, the RNA cargo can be encoded by a DNA molecule comprising a nucleic acid sequence having at least 50%, 60%, 70%, 75%, 80%, 85%, 90%, 95%, 96%, 97%, 98%, 99%, or 100% sequence identity to a DNA molecule having the nucleic acid sequence set forth in SEQ ID NO: 36. In some cases, the RNA cargo can be encoded by a DNA molecule comprising a nucleic acid sequence that is at least 50%, 60%, 70%, 75%, 80%, 85%, 90%, 95%, 96%, 97%, 98%, 99%, or 100% identical to a DNA molecule having the nucleic acid sequence set forth in SEQ ID NO: 36. In some cases, the RNA cargo can be encoded by a DNA molecule comprising the nucleic acid sequence set forth in SEQ ID NO: 36. In some cases, the RNA cargo can be encoded by a DNA molecule consisting of the nucleic acid sequence set forth in SEQ ID NO: 36.

[0147] In some cases, the RNA cargo can be encoded by a DNA molecule comprising a nucleic acid sequence having at least 50%, 60%, 70%, 75%, 80%, 85%, 90%, 95%, 96%, 97%, 98%, 99%, or 100% sequence identity to a DNA molecule having the nucleic acid sequence set forth in SEQ ID NO: 37. In some cases, the RNA cargo can be encoded by a DNA molecule comprising a nucleic acid sequence that is at least 50%, 60%, 70%, 75%, 80%, 85%, 90%, 95%, 96%, 97%, 98%, 99%, or 100% identical to a DNA molecule having the nucleic acid sequence set forth in SEQ ID NO: 37. In some cases, the RNA cargo can be encoded by a DNA molecule comprising the nucleic acid sequence set forth in SEQ ID NO: 37. In some cases, the RNA cargo can be encoded by a DNA molecule consisting of the nucleic acid sequence set forth in SEQ ID NO: 37. In some cases, the RNA cargo can be encoded by a coding sequence of the DNA molecule having the nucleic acid sequence set forth in SEQ ID NO: 37.

[0148] In some cases, the RNA cargo encodes a dystrophin protein that is at least 40 kilodaltons (kDa), at least 70 kDa, at least 120 kDa, at least 140 kDa, at least 250 kDa, at least 400 kDa, or at least 420 kDa in size. In some cases, the RNA cargo encodes a dystrophin protein that is detectable by western blot. In some cases, the RNA cargo encodes a protein that has at least 70%, at least 80%, at least 90%, at least 95%, at least 96%, or at least 99% sequence identity to a polypeptide sequence of SEQ ID NO: 1, SEQ ID NO: 2 (P11532-1, UniProt), SEQ ID NO: 3 (Pl 1532-4, UniProt), SEQ ID NO: 4 (Pl 1532-11, UniProt), SEQ ID NO: 5 (Pl 1532-2, UniProt), or SEQ ID NO: 6 (Pl 1532-3, UniProt). In some cases, the RNA cargo encodes a dystrophin protein that is a full-length dystrophin protein. In some cases, the RNA cargo encodes a protein that has atWSGR Ref. No. 54841-719.601least 70%, at least 80%, at least 90%, at least 95%, at least 96%, or at least 99% identical to a polypeptide sequence of SEQ ID NO: 1, SEQ ID NO: 2 (P11532-1, UniProt), SEQ ID NO: 3 (Pl 1532-4, UniProt), SEQ ID NO: 4 (Pl 1532-11, UniProt), SEQ ID NO: 5 (Pl 1532-2, UniProt), or SEQ ID NO: 6 (Pl 1532-3, UniProt). In some cases, the RNA cargo encodes a protein that has the polypeptide sequence of SEQ ID NO: 1, SEQ ID NO: 2 (P11532-1, UniProt), SEQ ID NO: 3 (Pl 1532-4, UniProt), SEQ ID NO: 4 (Pl 1532-11, UniProt), SEQ ID NO: 5 (Pl 1532-2, UniProt), or SEQ ID NO: 6 (Pl 1532- 3, UniProt).

[0149] In some cases, the extracellular vesicle can comprise a mass of a cargo molecule. For example, the extracellular vesicle can comprise a mass of an RNA molecule (e.g., an exogenous mRNA molecule) of at least 1 fg, 10 fg, 100 fg, 1 pg, 10 pg, 100 pg, 1 ng, 10 ng, 20 ng, 40 ng, 50 ng, 100 ng, 500 ng, 1 μg, 10 μg, 20 μg, 30 μg, 40 μg, or 50 μg. In some cases, the extracellular vesicle can comprise a mass of an RNA molecule of no more than 1 fg, 10 fg, 100 fg, 1 pg, 10 pg, 100 pg, 1 ng, 10 ng, 20 ng, 40 ng, 50 ng, 100 ng, 500 ng, 1 μg, 10 μg, 20 μg, 30 μg, 40 μg, or 50 μg.Therapeutics

[0150] In some cases, the extracellular vesicle described herein can comprise a therapeutic cargo. In some cases, the at least one therapeutic can be within (e.g. encapsulated) the extracellular vesicle. In some cases, the therapeutic can be a therapeutic polynucleotide, such as an RNA molecule (e.g., an mRNA molecule encoding a dystrophin protein). In some cases, the therapeutic cargo can be a therapeutic polypeptide. In some instances, the therapeutic is a therapeutic compound. In some cases, the therapeutic cargo can comprise a therapeutic polynucleotide, therapeutic polypeptide, therapeutic compound, or a combination thereof. In some instances, the extracellular vesicle can comprise a plurality of therapeutics, where the plurality of therapeutic cargos can comprise a therapeutic polynucleotide, therapeutic polypeptide, therapeutic compound, or a combination thereof.

[0151] In some cases, the extracellular vesicles described herein comprise at least one targeting polypeptide. In some cases, the targeting polypeptide is a muscle cell -targeting polypeptide comprising a muscle cell -targeting domain. In some cases, the accumulation of the extracellular vesicles comprising the muscle cell-targeting polypeptides comprising the muscle cell-targeting domain at the muscle tissue is higher compared to accumulation of extracellular vesicles without the muscle cell-targeting polypeptides. In some instances, the accumulation of the extracellular vesicles comprising the muscle celltargeting polypeptides at the muscle is at least 2-fold, 5-fold, 10-fold, 50-fold, 100-fold,WSGR Ref. No. 54841-719.601200-fold, 500-fold, 1,000-fold, 5,000-fold, or 10,000-fold higher compared to accumulation of extracellular vesicles lacking the muscle cell -targeting polypeptide. In some instances, the accumulation of the extracellular vesicles comprising the muscle celltargeting polypeptides at the muscle is at least 100 -fold higher compared to accumulation of extracellular vesicles lacking the muscle cell-targeting polypeptide.

[0152] In some cases, the muscle cell-targeting polypeptides comprise at least one muscle cell-targeting domain. In some cases, the muscle cell-targeting domains can be on an N- terminus of the muscle cell-targeting polypeptides. In some cases, the muscle celltargeting domains can be on a C-terminus of the muscle cell-targeting polypeptides. In some cases, the muscle cell-targeting domains can at any peptide location of the muscle cell-targeting polypeptides. In some cases, at least two targeting domains can be on the same muscle cell-targeting polypeptides. In some cases, the at least two targeting domains on the same muscle cell-targeting polypeptides can be the same. In some cases, the at least two targeting domains on the same muscle cell-targeting polypeptides can be different. In some instances, the targeting domains comprise at least 3, 4, 5, 6, 7, 8, 9, 10, 15, 20, 25, 30, 40, 50, or 100 amino acids.

[0153] In some cases, the extracellular vesicles comprising the extracellular vesicle surface proteins comprise increased half-life in circulation compared to half-life of extracellular vesicles without the extracellular vesicle surface proteins. In some cases, the half-life of the extracellular vesicles increased by the extracellular vesicle surface proteins is at least 90 minutes, 2 hours, 3 hours, 4 hours, 5 hours, 6 hours, 7 hours, 8 hours, 9 hours, 10 hours, 11 hours, 12 hours, 18 hours, 24 hours, 36 hours, 48 hours, 3 days, 4 days, 5 days, 6 days, 7 days, 10 days, 12 days, 14 days, 21 days, 28 days, 30 days, or longer than half - life of extracellular vesicles lacking the extracellular vesicle surface proteins.

[0154] In some cases, the extracellular vesicles comprising the extracellular vesicle surface proteins have decreased toxicity compared to the extracellular vesicles lacking the extracellular vesicle surface proteins. In such cases, often the extracellular vesicle surface proteins specifically bind to a target and do not have significant off-target binding. In some cases, the toxicity can comprise toxicity to cells that are not targeted by the tumor targeting polypeptides. In some cases, the extracellular vesicles comprising the extracellular vesicle surface proteins have decreased toxicity that is at least is 1 -fold, 2- fold, 3-fold, 4-fold, 5-fold, 6-fold, 7-fold, 8-fold, 9-fold, 10-fold, 20-fold, 30-fold, 50- fold, 100-fold, or more decreased compared to the extracellular vesicles lacking the extracellular vesicle surface proteins. In some cases, the decreased toxicity of theWSGR Ref. No. 54841-719.601extracellular vesicles comprising the extracellular vesicle surface proteins is at least 10%, 20%, 30%, 40%, 50%, 60%, 70%, 80%, 90%, or more decreased compared to the extracellular vesicles lacking the extracellular vesicle surface proteins.

[0155] In some cases, the extracellular vesicles (e.g., exosomes) are tolerated by the subject following administration of the extracellular vesicles. For example, in some cases, the extracellular vesicles do not induce an immune response, or are not immunogenic.Therapeutic Polynucleotides

[0156] Described herein, in some cases, are extracellular vesicles comprising a therapeutic polynucleotide. In some instances, the therapeutic polynucleotide can be encoded by the at least one heterologous polynucleotide, plasmid, or vector transfected into the extracellular vesicle donor cell. In some cases, the therapeutic polynucleotide can comprise a nucleic acid sequence that can be translated into a therapeutic polypeptide by the cell targeted and bound by the targeting polypeptide described herein. In some embodiments, the therapeutic polynucleotide can comprise a nucleic acid sequence that can be translated into a dystrophin protein by a targeted cell and bound by the targeting polypeptide described herein.

[0157] In some cases, the extracellular vesicle can comprise at least one therapeutic polynucleotide. In some cases, the extracellular vesicle can comprise at least 1, 2, 3, 4, 5, 6, 7, 8, 9, 10, 50, 100, 500, 1,000, 5,000, 10,000, 50,000, 100,000, 500,000, 1,000,000 or more copies of the therapeutic polynucleotides (e.g., an exogenous mRNA molecule encoding a full-length dystrophin protein). In some cases, each extracellular vesicle can comprise at least 1, 2, 3, 4, 5, 6, 7, 8, 9, 10, 50, 100, 500, 1,000, 5,000, 10,000, 50,000, 100,000, 500,000, 1,000,000 or more copies of the therapeutic mRNA described herein. In some instances, the extracellular vesicle can comprise at least two therapeutic polynucleotides. In some instances, the extracellular vesicle can comprise at least two therapeutic polynucleotides, where the at least two therapeutic polynucleotides are different or the same. In some cases, the at least two different therapeutic polynucleotides encapsulated by the extracellular vesicles comprise different ratio. For example, the ratio between the first and the second of the two different therapeutic polynucleotides can be 1:1,000,000, 1:500,000, 1:100,000, 1:50,000, 1:10,000, 1:5,000, 1:1,000, 1:500, 1:100, 1:50, 1:10, 1:5, 1:4, 1:3, 1:2, or 1:1. In some instances, the extracellular vesicle can comprise at least two, three, four, five, six, seven, right, nine, ten or more therapeutic polynucleotides encapsulated in the same extracellular vesicle.WSGR Ref. No. 54841-719.601

[0158] In some cases, the therapeutic polynucleotide can comprise RNA, particularly mRNA.In some embodiments, the therapeutic polynucleotide can be any of the RNA cargos described herein. In some cases, the therapeutic polynucleotide can comprise mRNA. In some cases, the mRNA is fully intact or substantially intact. In some cases, the mRNA encodes a portion of the protein. In some cases, the mRNA encodes a full-length protein. In some cases, the therapeutic polynucleotide encodes a therapeutic polypeptide that is detectable by western blot. In some cases, the mRNA can comprise at least 50, 100, 200, 500, 1,000, 5,000, 10,000, 50,000, 100,000, 500,000, or 1,000,000 of RNA nucleotides. In some cases, the therapeutic polynucleotide can comprise at least 2 kilobases (kbs), at least 4 kbs, at least 5 kbs, at least 7 kbs, at least 10 kbs, at least 12 kbs, at least 13 kbs, or at least 14 kbs of RNA nucleotides. In some instances, therapeutic polynucleotides comprise DNA. In some instances, therapeutic polynucleotides comprise DNA such as vectors that encode therapeutic polypeptide or RNA therapeutic. The therapeutic polynucleotide can encode therapeutic polypeptide including but not limited to: a dystrophin protein or peptide, a scaffolding protein or peptide, a cytoskeleton-associated protein, a protein that binds to a cell adhesion molecule, a protein that interacts with a protein of a dystrophin-glycoprotein complex, a protein that interacts with a proteoglycan (e.g., a dystroglycan), or a protein of dystrophin-glycoprotein complex. In some cases, the therapeutic polynucleotide described herein can encode a dystrophin protein, particularly a full-length dystrophin protein. In some cases, the therapeutic polynucleotide described herein can encode a human dystrophin protein. In some cases, the therapeutic polynucleotide is a dystrophin mRNA that is about 13.7 kbs in length. In some cases, the therapeutic polynucleotide is a full-length human dystrophin mRNA. In some cases, the therapeutic polynucleotide encode a fragment of human dystrophin protein.

[0159] In some cases, the therapeutic polynucleotide encodes a therapeutic polypeptide. The therapeutic polypeptide can be any protein encoded by RNA cargo or therapeutic polynucleotide of this disclosure. In some cases, the therapeutic polynucleotide encodes a protein that is detectable by western blot. In some cases, the therapeutic polynucleotide encodes a protein that is at least 40 kilodaltons (kDa), at least 70 kDa, at least 120 kDa, at least 140 kDa, at least 250 kDa, at least 400 kDa, or at least 420 kDa in size. Particularly, the therapeutic polynucleotide can encode a protein that is about 420 kDa in size.

[0160] In some cases, the therapeutic polynucleotide encodes a protein that has at least 70%, at least 80%, at least 90%, at least 95%, at least 96%, or at least 99% sequence identity to a polypeptide sequence of SEQ ID NO: 1, SEQ ID NO: 2 (P11532-1, UniProt), SEQ IDWSGR Ref. No. 54841-719.601NO: 3 (Pl 1532-4, UniProt), SEQ ID NO: 4 (Pl 1532-11, UniProt), SEQ ID NO: 5 (Pl 1532-2, UniProt), or SEQ ID NO: 6 (Pl 1532-3, UniProt). In some cases, the therapeutic polynucleotide encodes a protein that has at least 70%, at least 80%, at least 90%, at least 95%, at least 96%, or at least 99% identical to a polypeptide sequence of SEQ ID NO: 1, SEQ ID NO: 2 (P11532-1, UniProt), SEQ ID NO: 3 (Pl 1532-4, UniProt), SEQ ID NO: 4 (Pl 1532-11, UniProt), SEQ ID NO: 5 (Pl 1532-2, UniProt), or SEQ ID NO: 6 (Pl 1532-3, UniProt). In some cases, the therapeutic polynucleotide encodes a protein that has the polypeptide sequence of SEQ ID NO: 1, SEQ ID NO: 2 (P11532-1, UniProt), SEQ ID NO: 3 (Pl 1532-4, UniProt), SEQ ID NO: 4 (Pl 1532-11, UniProt), SEQ ID NO: 5 (Pl 1532-2, UniProt), or SEQ ID NO: 6 (Pl 1532-3, UniProt). In some cases, the RNA cargo encodes a full-length dystrophin protein. In some cases, the RNA cargo is a mRNA transcribed from a dystrophin DNA. In some cases, the RNA cargo is a mRNA transcribed from a human dystrophin gene. In some cases, the RNA cargo is a dystrophin mRNA (e.g., a human dystrophin mRNA), particularly a full-length dystrophin mRNA.

[0161] In some cases, the RNA cargo can comprise a dystrophin mRNA that is at least about 2 kilobases (kbs), at least about 4 kbs, at least about 5 kbs, at least about 7 kbs, at least about 10 kbs, at least about 12 kbs, at least about 13 kbs, or at least about 14 kbs in length. In some cases, the RNA cargo is a dystrophin mRNA that is about 13.7 kbs in length. In some cases, the RNA cargo is a full-length human dystrophin mRNA. In some cases, the RNA cargo is a polynucleotide having at least about 70%, at least about 80%, at least about 90%, at least about 95%, at least about 96%, or at least about 99% sequence identity to a polynucleotide sequence of SEQ ID NO: 27.

[0162] In some cases, the RNA cargo encodes a dystrophin protein that is at least about 40 kilodaltons (kDa), at least about 70 kDa, at least about 120 kDa, at least about 140 kDa, at least about 250 kDa, at least about 400 kDa, or at least about 420 kDa in size. In some cases, the RNA cargo encodes a dystrophin protein that is detectable by western blot. In some cases, the RNA cargo encodes a protein that has at least about 70%, at least about 80%, at least about 90%, at least about 95%, at least about 96%, or at least about 99% sequence identity to a polypeptide sequence of SEQ ID NO: 1 (Ref. Seq. NP 003997.2), SEQ ID NO: 2 (P11532-1, UniProt), SEQ ID NO: 3 (Pl 1532-4, UniProt), SEQ ID NO: 4 (Pl 1532-11, UniProt), SEQ ID NO: 5 (Pl 1532-2, UniProt), or SEQ ID NO: 6 (Pl 1532-3, UniProt). In some cases, the RNA cargo encodes a dystrophin protein that is a full-length dystrophin protein. In some cases, the RNA cargo encodes a protein that has at least aboutWSGR Ref. No. 54841-719.60170%, at least about 80%, at least about 90%, at least about 95%, at least about 96%, or at least about 99% identical to a polypeptide sequence of SEQ ID NO: 1 (Ref. Seq.NP_003997.2), SEQ ID NO: 2 (P11532-1, UniProt), SEQ ID NO: 3 (Pl 1532-4, UniProt), SEQ ID NO: 4 (Pl 1532-11, UniProt), SEQ ID NO: 5 (Pl 1532-2, UniProt), or SEQ ID NO: 6 (Pl 1532-3, UniProt). In some cases, the RNA cargo encodes a protein that has the polypeptide sequence of SEQ ID NO: 1 (Ref. Seq. NP 003997.2), SEQ ID NO: 2 (P11532-1, UniProt), SEQ ID NO: 3 (Pl 1532-4, UniProt), SEQ ID NO: 4 (Pl 1532-11, UniProt), SEQ ID NO: 5 (Pl 1532-2, UniProt), or SEQ ID NO: 6 (Pl 1532-3, UniProt).

[0163] In some cases, the therapeutic polynucleotides comprise at least one modified nucleic acid or nucleic acid analog. Exemplary modified nucleic acids include, but are not limited to, uracil-5-yl, hypoxanthin-9-yl (I), 2-aminoadenin-9-yl, 5 -methylcytosine (5-me-C), 5- hydroxymethyl cytosine, xanthine, hypoxanthine, 2 -aminoadenine, 6-methyl and other alkyl derivatives of adenine and guanine, 2 -propyl and other alkyl derivatives of adenine and guanine, 2-thiouracil, 2-thiothymine and2-thiocytosine, 5-halouracil and cytosine, 5- propynyl uracil and cytosine, 6 -azo uracil, cytosine and thymine, 5 -uracil (pseudouracil), 4-thiouracil, 8-halo, 8-amino, 8-thiol, 8-thioalkyl, 8-hydroxyl and other 8-substituted adenines and guanines, 5 -halo particularly 5 -bromo, 5-trifiuoromethyl and other 5- substituted uracils and cytosines, 7 -methylguanine and 7-methyladenine, 8-azaguanine and 8 -azaadenine, 7-deazaguanine and 7 -deazaadenine and 3 -deazaguanine and 3- deazaadenine. Certain modified nucleic acids, such as 5 -substituted pyrimidines, 6- azapyrimidines and N-2 substituted purines, N-6 substituted purines, 0-6 substituted purines, 2 -aminopropyladenine, 5-propynyluracil, 5-propynylcytosine, 5 -methylcytosine, those that increase the stability of duplex formation, universal nucleic acids, hydrophobic nucleic acids, promiscuous nucleic acids, size -expanded nucleic acids, fluorinated nucleic acids, 5-substituted pyrimidines, 6 -azapyrimidines and N-2, N-6 and 0-6 substituted purines, including 2 -aminopropyladenine, 5-propynyluracil and 5-propynylcytosine. 5- methylcytosine (5-me-C), 5- hydroxymethyl cytosine, xanthine, hypoxanthine, 2- aminoadenine, 6-methyl, other alkyl derivatives of adenine and guanine, 2 -propyl and other alkyl derivatives of adenine and guanine, 2-thiouracil, 2-thiothymine and 2- thiocytosine, 5-halouracil, 5-halocytosine, 5-propynyl (-C=C-CH3) uracil, 5-propynyl cytosine, other alkynyl derivatives of pyrimidine nucleic acids, 6 -azo uracil, 6-azo cytosine, 6-azo thymine, 5 -uracil (pseudouracil), 4-thiouracil, 8-halo, 8-amino, 8-thiol, 8- thioalkyl, 8-hydroxyl and other 8-substituted adenines and guanines, 5 -halo particularly 5-bromo, 5 -trifluoro methyl, other 5-substituted uracils and cytosines, 7-methylguanine, 7-WSGR Ref. No. 54841-719.601methyladenine, 2-F-adenine, 2-amino-adenine, 8-azaguanine, 8-azaadenine, 7- deazaguanine, 7- deazaadenine, 3 -deazaguanine, 3 -deazaadenine, tricyclic pyrimidines, phenoxazine cytidine( [5,4-b][l,4]benzoxazin-2(3H)-one), phenothiazine cytidine (1H- pyrimido[5,4-b][l,4]benzothiazin-2(3H)-one), G-clamps, phenoxazine cytidine (e.g. 9- (2- aminoethoxy)-H-pyrimido[5,4-b][l,4]benzoxazin-2(3H)-one), carbazole cytidine (2H- pyrimido[4,5- b]indol-2-one), pyridoindole cytidine (H-pyrido[3’,2’:4,5]pyrrolo[2,3- d]pyrimidin-2-one), those in which the purine or pyrimidine base is replaced with other heterocycles, 7-deaza-adenine, 7-deazaguanosine, 2 -aminopyridine, 2-pyridone, azacytosine, 5 -bromocytosine, bromouracil, 5 -chlorocytosine, chlorinated cytosine, cyclocytosine, cytosine arabinoside, 5 -fluoro cytosine, fluoropyrimidine, fluorouracil, 5,6- dihydrocytosine, 5 -iodocytosine, hydroxyurea, iodouracil, 5 -nitrocytosine, 5- bromouracil, 5 -chloro uracil, 5 -fluorouracil, and 5-iodouracil, 2-amino-adenine, 6-thio- guanine, 2-thio-thymine, 4-thio-thymine, 5-propynyl-uracil, 4 -thio -uracil, N4- ethylcytosine, 7-deazaguanine, 7-deaza-8- azaguanine, 5 -hydroxycytosine, 2’- deoxyuridine, 2-amino-2’-deoxyadenosine Modified nucleic acids comprising various heterocyclic bases and various sugar moieties (and sugar analogs) are available in the art, and the nucleic acids in some cases include one or several heterocyclic bases other than the principal five base components of naturally-occurring nucleic acids. For example, the heterocyclic base includes, in some cases, uracil-5-yl, cytosin-5-yl, adenin-7-yl, adenin-8- yl, guanin-7-yl, guanin-8-yl, 4- aminopyrrolo [2.3-d] pyrimidin-5-yl, 2-amino-4- oxopyrolo [2, 3-d] pyrimidin-5-yl, 2- amino -4-oxopyrrolo [2.3-d] pyrimidin-3-yl groups, where the purines are attached to the sugar moiety of the nucleic acid via the 9 -position, the pyrimidines via the 1 -position, the pyrrolopyrimidines via the 7-position and the pyrazolopyrimidines via the 1 -position.

[0164] In some cases, nucleotide analogs are also modified at the phosphate moiety.Modified phosphate moieties include, but are not limited to, those with modification at the linkage between two nucleotides and contains, for example, a phosphorothioate, chiral phosphorothioate, phosphorodithioate, phosphotriester, aminoalkylphosphotriester, methyl and other alkyl phosphonates including 3 ’-alkylene phosphonate and chiral phosphonates, phosphinates, phosphoramidates including 3 ’ -amino phosphoramidate and aminoalkylphosphoramidates, thionophosphoramidates, thionoalkylphosphonates, thionoalkylphosphotriesters, and boranophosphates. It is understood that these phosphate or modified phosphate linkage between two nucleotides are through a 3 ’-5’ linkage or aWSGR Ref. No. 54841-719.6012 ’-5’ linkage, and the linkage contains inverted polarity such as 3 ’-5’ to 5 ’-3’ or 2 ’-5’ to 5 ’-2’. Various salts, mixed salts and free acid forms are also included.

[0165] In some cases, modified nucleic acids include 2’,3’-dideoxy-2’,3’-didehydro- nucleo sides 5 ’-substituted DN A and RNA derivatives or 5 ’-substituted monomers made as the monophosphate with modified bases.

[0166] In some cases, modified nucleic acids include modifications at the 5 ’ -position and the 2 ’-position of the sugar ring (, such as 5’-CH2-substituted2’-O-protected nucleosides. In some cases, modified nucleic acids include amide linked nucleoside dimers have been prepared for incorporation into oligonucleotides wherein the 3’ linked nucleoside in the dimer (5’ to 3’) comprises a 2’-OCH3and a 5’-(S)-CH3. Modified nucleic acids can include 2 ’-substituted 5’-CH2(or O) modified nucleosides. Modified nucleic acids can include 5’-methylenephosphonate DNA and RNA monomers, and dimers. Modified nucleic acids can include 5 ’-phosphonate monomers having a 2 ’-substitution and other modified 5 ’-phosphonate monomers. Modified nucleic acids can include 5 ’-modified methylenephosphonate monomers. Modified nucleic acids can include analogs of 5’ or 6 ’-phosphonate ribonucleosides comprising a hydroxyl group at the 5 ’ and / or 6 ’ -position. Modified nucleic acidscan include 5 ’-phosphonate deoxyribonucleoside monomers and dimers having a 5 ’-phosphate group. Modified nucleic acids can include nucleosides having a 6 ’-phosphonate group wherein the 5’ or / and 6’-position is unsubstituted or substituted with a thio -tert-butyl group (SC(CH3)3) (and analogs thereof); a methyleneamino group (CH2NH2) (and analogs thereof) or a cyano group (CN) (and analogs thereof).

[0167] In some cases, modified nucleic acids also include modifications of the sugar moiety.In some cases, nucleic acids contain one or more nucleosides wherein the sugar group has been modified. Such sugar modified nucleosides may impart enhanced nuclease stability, increased binding affinity, or some other beneficial biological property. In certain cases, nucleic acids comprise a chemically modified ribofuranose ring moiety. Examples of chemically modified ribofuranose rings include, without limitation, addition of substituent groups (including 5’ and / or 2’ substituent groups; bridging of two ring atoms to form bicyclic nucleic acids (BNA); replacement of the ribosyl ring oxygen atom with S, N(R), or C(RI)(R2) (R = H, C1-C12 alkyl or a protecting group); and combinations thereof.

[0168] In some instances, a modified nucleic acid comprises modified sugars or sugar analogs. Thus, in addition to ribose and deoxyribose, the sugar moiety can be pentose,WSGR Ref. No. 54841-719.601deoxypentose, hexose, deoxyhexose, glucose, arabinose, xylose, lyxose, or a sugar “analog” cyclopentyl group. The sugar can be in a pyranosyl or furan osyl form. The sugar moiety may be the furanoside of ribose, deoxyribose, arabinose or 2’ -O-alkylribose, and the sugar can be attached to the respective heterocyclic bases either in [alpha] or [beta] anomeric configuration. Sugar modifications include, but are not limited to, 2’-alkoxy- RNA analogs, 2’-amino-RNA analogs, 2’-fluoro-DNA, and 2’-alkoxy- or amino- RNAZDNA chimeras. For example, a sugar modification may include 2’-O-methyl- uridine or 2’-O-methyl-cytidine. Sugar modifications include 2’-O-alkyl-substituted deoxyribonucleosides and2’-O-ethyleneglycol like ribonucleosides. The preparation of these sugars or sugar analogs and the respective “nucleosides” wherein such sugars or analogs are attached to a heterocyclic base (nucleic acid base) is known. Sugar modifications may also be made and combined with other modifications.

[0169] Modifications to the sugar moiety include natural modifications of the ribose and deoxy ribose as well as modified modifications. Sugar modifications include, but are not limited to, the following modifications at the 2’ position: OH; F; O-, S-, orN-alkyl; O-, S-, or N-alkenyl; O-, S- orN-alkynyl; or O-alkyl-O-alkyl, wherein the alkyl, alkenyl and alkynyl may be substituted or unsubstituted Ci to Cio, alkyl or C2 to Cio alkenyl and alkynyl. 2’ sugar modifications also include but are not limited to -O[(CH2)nO]mCH3, - O(CH2)nOCH3, -O(CH2)nNH2, -O(CH2)nCH3, -O(CH2)nONH2, and -O(CH2)nON[(CH2)n CH3)]2, where n and m are from 1 to about 10.

[0170] Other modifications at the 2’ position include but are not limited to: Ci to Cio lower alkyl, substituted lower alkyl, alkaryl, aralkyl, O-alkaryl, O-aralkyl, SH, SCH3, OCN, Cl, Br, CN, CF3, OCF3, SOCH3, SO2CH3, ONO2, NO2, N3, NH2, heterocycloalkyl, hetero cyclo alkaryl, amino alkylamino, polyalkylamino, substituted silyl, an RNA cleaving group, a reporter group, an intercalator, a group for improving the pharmacokinetic properties of an oligonucleotide, or a group for improving the pharmacodynamic properties of an oligonucleotide, and other substituents having similar properties. Similar modifications may also be made at other positions on the sugar, particularly the 3’ position of the sugar on the 3 ’ terminal nucleotide or in 2 ’ -5 ’ linked oligonucleotides and the 5’ position of the 5’ terminal nucleotide. Modified sugars also include those that contain modifications at the bridging ring oxygen, such as CH2and S. Nucleotide sugar analogs may also have sugar mimetics such as cyclobutyl moieties in place of the pentofuranosyl sugar.WSGR Ref. No. 54841 -719.601

[0171] Examples of nucleic acids having modified sugar moieties include, without limitation, nucleic acids comprising 5 ’-vinyl, 5 ’-methyl (R or S), 4’-S, 2’-F, 2’-OCH3, and 2’- O(CH2)2OCH3substituent groups. The substituent at the 2’ position can also be selected from allyl, amino, azido, thio, O-allyl, O-(Ci-Cio alkyl), OCF3, O(CH2)2SCH3, O(CH2)2- O-N(Rm)(Rn), and O-CH2-C(=O)-N(Rm)(Rn), where each Rmand Rnis, independently, H or substituted or unsubstituted Ci-Cio alkyl.

[0172] In certain cases, nucleic acids described herein include one or more bicyclic nucleic acids. In certain such cases, the bicyclic nucleic acid comprises a bridge between the 4’ and the 2’ ribosyl ring atoms. In certain cases, nucleic acids provided herein include one or more bicyclic nucleic acids wherein the bridge comprises a 4’ to 2’ bicyclic nucleic acid. Examples of such 4’ to 2’ bicyclic nucleic acids include, but are not limited to, one of the formulae: 4’-(CH2)-O-2’ (LNA); 4’-(CH2)-S-2’; 4’-(CH2)2-O-2’ (ENA); 4’- CH(CH3)-O-2’ and 4’-CH(CH2OCH3)-O-2’, and analogs thereof; 4’-C(CH3)(CH3)-O- 2 ’and analogs thereof.

[0173] In certain cases, nucleic acids comprise linked nucleic acids. Nucleic acids can be linked together using any inter nucleic acid linkage. The two main classes of inter nucleic acid linking groups are defined by the presence or absence of a phosphorus atom. Representative phosphorus containing inter nucleic acid linkages include, but are not limited to, phosphodiesters, phosphotriesters, methylphosphonates, phosphoramidate, and phosphorothioates (P=S). Representative non-phosphorus containing inter nucleic acid linking groups include, but are not limited to, methylenemethylimino (-CH2-N(CH3)-O- CH2-), thiodiester (-O-C(O)-S-), thionocarbamate (-O-C(O)(NH)-S-); siloxane (-0- Si(H)2-O-); and N, N* -dimethylhydrazine (-CH2-N(CH3)-N(CH3)). In certain aspects, inter nucleic acids linkages having a chiral atom can be prepared as a racemic mixture, as separate enantiomers, e.g., alkylphosphonates and phosphorothioates. Modified nucleic acids can contain a single modification. Modified nucleic acids can contain multiple modifications within one of the moieties or between different moieties.

[0174] Backbone phosphate modifications to nucleic acid include, but are not limited to, methyl phosphonate, phosphoro thio ate, phosphoramidate (bridging or non -bridging), phosphotriester, phosphorodithioate, phosphodithioate, and boranophosphate, and may be used in any combination. Other non- phosphate linkages may also be used.

[0175] In some cases, backbone modifications e.g., methylphosphonate, phosphorothioate, phosphoroamidate and phosphorodithioate intemucleotide linkages) can conferWSGR Ref. No. 54841-719.601immunomodulatory activity on the modified nucleic acid and / or enhance their stability in vivo.

[0176] In some instances, a phosphorous derivative (or modified phosphate group) is attached to the sugar or sugar analog moiety in and can be a monophosphate, diphosphate, triphosphate, alkylphosphonate, phosphorothioate, phosphorodithioate, phosphoramidate or the like.

[0177] In some cases, backbone modification comprises replacing the phosphodiester linkage with an alternative moiety such as an anionic, neutral or cationic group. Examples of such modifications include: anionic internucleoside linkage; N3’ to P5’ phosphoramidate modification; boranophosphate DNA; prooligonucleotides; neutral internucleoside linkages such as methylphosphonates; amide linked DNA; methylene(methylimino) linkages; formacetal and thio formacetal linkages; backbones containing sulfonyl groups; morpholino oligos; peptide nucleic acids (PNA); and positively charged deoxyribonucleic guanidine (DNG) oligos (Micklefield, 2001, Current Medicinal Chemistry 8: 1157-1179). A modified nucleic acid may comprise a chimeric or mixed backbone comprising one or more modifications, e.g. a combination of phosphate linkages such as a combination of phosphodiester and phosphorothioate linkages.

[0178] Substitutes for the phosphate include, for example, short chain alkyl or cycloalkyl intemucleoside linkages, mixed heteroatom and alkyl or cycloalkyl intemucleoside linkages, or one or more short chain heteroatomic or heterocyclic intemucleoside linkages. These include those having morpholino linkages (formed in part from the sugar portion of a nucleoside); siloxane backbones; sulfide, sulfoxide and sulfone backbones; formacetyl and thioformacetyl backbones; methylene formacetyl and thioformacetyl backbones; alkene containing backbones; sulfamate backbones; methyleneimino and methylenehydrazino backbones; sulfonate and sulfonamide backbones; amide backbones; and others having mixed N, O, S and CH2 component parts. It is also understood in a nucleotide substitute that both the sugar and the phosphate moieties of the nucleotide can be replaced, by for example an amide type linkage (aminoethylglycine) (PNA). United States Patent Nos. 5,539,082; 5,714,331; and 5,719,262 teach how to make and use PNA molecules, each of which is herein incorporated by reference. See also Nielsen et al., Science, 1991, 254, 1497-1500. It is also possible to link other types of molecules (conjugates) to nucleotides or nucleotide analogs to enhance for example, cellular uptake. Conjugates can be chemically linked to the nucleotide or nucleotide analogs. Such conjugates include but are not limited to lipid moieties such as a cholesterol moiety,WSGR Ref. No. 54841-719.601cholic acid, a thioether, e.g., hexyl-S-tritylthiol, a thiocholesterol, an aliphatic chain, e.g., dodecandiol or undecyl residues (, a phospholipid, e.g., di-hexadecyl-rac-glycerol or triethylammonium 1-di-O-hexadecyl-rac-glycero-S-H-phosphonate, a polyamine or a polyethylene glycol chain, or adamantane acetic acid, a palmityl moiety, or an octadecylamine or hexylamino-carbonyl-oxycholesterol moiety.

[0179] In some cases, the at least one modified nucleotide or nucleotide analogue described herein can be resistant toward nucleases such as for example ribonuclease such as RNase H, deoxyribonuclease such as DNase, or exonuclease such as 5 ’ -3 ’ exonuclease and 3 ’-5 ’ exonuclease when compared to natural nucleic acid molecules. In some instances, the at least one modified nucleotide or nucleotide analogue comprises 2 ’ -O-methy 1, 2’-O- methoxyethyl (2’-O-MOE), 2’-O-aminopropyl, 2'-deoxy, T-deoxy-2'-fluoro, 2'-O- aminopropyl (2'-O-AP), 2'-O-dimethylaminoethyl (2'-O-DMAOE), 2'-O- dimethylaminopropyl (2'-O-DMAP), T-O- dimethylaminoethyloxyethyl (2'-O- DMAEOE), or 2'-O-N-methylacetamido (2'-O-NMA) modified, LNA, ENA, PNA, HNA, morpholino, methylphosphonate nucleotides, thiolphosphonate nucleotides, 2’-fluoro N3- P5 ’-phosphoramidites, or combinations thereof are resistant toward nucleases such as for example ribonuclease such as RNase H, deoxyribonuclease such as DNase, or exonuclease such as 5 ’-3’ exonuclease and 3’-5’ exonuclease. In some instances, 2’-O- methyl modified nucleic acid molecule is nuclease resistance (e.g., RNase H, DNase, 5’- 3’ exonuclease or 3’-5’ exonuclease resistance). In some instances, 2’0-methoxyethyl (2’-O-MOE) modified nucleic acid molecule is nuclease resistance (e.g., RNase H, DNase, 5’-3’ exonuclease or 3’-5’ exonuclease resistance). In some instances, 2’-O- aminopropyl modified nucleic acid molecule is nuclease resistance (e.g., RNase H, DNase, 5 ’-3’ exonuclease or 3 ’-5’ exonuclease resistance). In some instances, 2'-deoxy modified nucleic acid molecule is nuclease resistance (e.g., RNase H, DNase, 5 ’-3’ exonuclease or 3 ’-5’ exonuclease resistance). In some instances, T-deoxy-2'-fluoro modified nucleic acid molecule is nuclease resistance (e.g., RNase H, DNase, 5 ’-3’ exonuclease or 3 ’-5 ’ exonuclease resistance). In some instances, 2'-O-aminopropyl (2'-O- AP) modified nucleic acid molecule is nuclease resistance (e.g., RNase H, DNase, 5 ’-3’ exonuclease or 3 ’-5’ exonuclease resistance). In some instances, 2'-O- dimethylamino ethyl (2'-O-DMAOE) modified nucleic acid molecule is nuclease resistance (e.g., RNase H, DNase, 5 ’-3’ exonuclease or 3 ’-5’ exonuclease resistance). In some instances, 2'-O-dimethylaminopropyl (2'-O-DMAP) modified nucleic acid molecule is nuclease resistance (e.g., RNase H, DNase, 5’-3’ exonuclease or 3’-5’ exonucleaseWSGR Ref. No. 54841-719.601resistance). In some instances, T-O- dimethylaminoethyloxyethyl (2'-O-DMAEOE) modified nucleic acid molecule is nuclease resistance (e.g., RNase H, DNase, 5 ’-3’ exonuclease or 3 ’-5 ’ exonuclease resistance). In some instances, 2'-O-N-methylacetamido (2'-0-NMA) modified nucleic acid molecule is nuclease resistance (e.g., RNase H, DNase, 5 ’-3’ exonuclease or 3 ’-5’ exonuclease resistance). In some instances, LNA modified nucleic acid molecule is nuclease resistance (e.g., RNase H, DNase, 5 ’-3’ exonuclease or 3 ’-5’ exonuclease resistance). In some instances, ENA modified nucleic acid molecule is nuclease resistance (e.g., RNase H, DNase, 5 ’-3’ exonuclease or 3 ’-5’ exonuclease resistance). In some instances, HNA modified nucleic acid molecule is nuclease resistance (e.g., RNase H, DNase, 5’-3’ exonuclease or 3’-5’ exonuclease resistance). In some instances, morph olinos is nuclease resistance (e.g., RNase H, DNase, 5 ’-3’ exonuclease or 3 ’-5’ exonuclease resistance). In some instances, PNA modified nucleic acid molecule is resistant to nucleases (e.g., RNase H, DNase, 5 ’-3’ exonuclease or 3 ’-5’ exonuclease resistance). In some instances, methylphosphonate nucleotides modified nucleic acid molecule is nuclease resistance (e.g., RNase H, DNase, 5 ’-3’ exonuclease or 3 ’-5’ exonuclease resistance). In some instances, thiolphosphonate nucleotides modified nucleic acid molecule is nuclease resistance (e.g., RNase H, DNase, 5 ’-3’ exonuclease or 3 ’-5’ exonuclease resistance). In some instances, nucleic acid molecule comprising 2’-fluoro N3-P5’-phosphoramidites is nuclease resistance (e.g., RNase H, DNase, 5 ’-3’ exonuclease or 3 ’-5’ exonuclease resistance). In some instances, the 5’ conjugates described herein inhibit 5 ’-3’ exonucleo lytic cleavage. In some instances, the 3’ conjugates described herein inhibit 3 ’-5’ exonucleolytic cleavage.

[0180] In additional cases, the modified nucleotide or nucleotide analogue described herein is modified to increase its stability. In some embodiment, the nucleic acid molecule is RNA (e.g., mRNA). In some instances, the mRNA can be modified by one or more of the modifications to increase its stability. In some cases, the mRNA can be modified at the 2 ’ hydroxyl position, such as by 2’-O-methyl, 2’-O-methoxyethyl (2’-O-MOE), 2’-O- aminopropyl, 2'-deoxy, T-deoxy-2'-fluoro, 2'-O-aminopropyl (2'-0-AP), 2'-O- dimethylaminoethyl (2'-O-DMAOE), 2'-O-dimethylaminopropyl (2'-O-DMAP), T-O- dimethylamino ethyloxyethyl (2'-O-DMAEOE), or 2'-O-N-methylacetamido (2'-O-NMA) modification or by a locked or bridged ribose conformation (e.g., LNA or ENA). In some cases, the at least one modified nucleotide or nucleotide analogue is modified by 2’ -O- methyl and / or 2’-O-methoxyethyl ribose. In some cases, the at least one modified nucleotide or nucleotide analogue also includes morpholinos, PNAs, HNA,WSGR Ref. No. 54841-719.601methylphosph onate nucleotides, thiolphosphonate nucleotides, and / or 2 ’-fluoro N3-P5’- phosphoramidites to increase its stability. In some instances, the at least one modified nucleotide or nucleotide analogue is a chirally pure (or stereo pure) nucleic acid molecule. In some instances, the chirally pure (or stereo pure) nucleic acid molecule is modified to increase its stabilityTABLE 1: Amino Acid Sequences of Dystrophin Encoded by mRNA Cargo in Extracellular VesiclesSEQ Description SequenceID NO:1 Human MLWWEEVEDCYEREDVQKKTFTKWVNAQFSKFGKQHIENL dystrophin FSDLQDGRRLLDLLEGLTGQKLPKEKGSTRVHALNNVNKAL isoform RVLQNNNVDLVNIGSTDIVDGNHKLTLGLIWNIILHWQVKN Dp427m VMKNIMAGLQQTNSEKILLSWVRQSTRNYPQVNVINFTTSW SDGLALNALIHSHRPDLFDWNSWCQQSATQRLEHAFNIARRef. Seq. YQLGIEKLLDPEDVDTTYPDKKSILMYITSLFQVLPQQVSIEAI NP 00399 QEVEMLPRPPKVTKEEHFQLHHQMHYSQQITVSLAQGYERT 7.1 SSPKPRFKSYAYTQAAYVTTSDPTRSPFPSQHLEAPEDKSFGS SLMESEVNLDRYQTALEEVLSWLLSAEDTLQAQGEISNDVE VVKDQFHTHEGYMMDLTAHQGRVGNILQLGSKLIGTGKLSE DEETEVQEQMNLLNSRWECLRVASMEKQSNLHRVLMDLQN QKLKELNDWLTKTEERTRKMEEEPLGPDLEDLKRQVQQHK VLQEDLEQEQVRVNSLTHMWVVDESSGDHATAALEEQLK VLGDRWANICRWTEDRWVLLQDILLKWQRLTEEQCLFSAW LSEKEDAVNKIHTTGFKDQNEMLSSLQKLAVLKADLEKKKQ SMGKLYSLKQDLLSTLKNKSVTQKTEAWLDNFARCWDNLV QKLEKSTAQISQAVTTTQPSLTQTTVMETVTTVTTREQILVK HAQEELPPPPPQKKRQITVDSEIRKRLDVDITELHSWITRSEA VLQSPEFAIFRKEGNFSDLKEKVNAIEREKAEKFRKLQDASRS AQALVEQMVNEGVNADSIKQASEQLNSRWIEFCQLLSERLN WLEYQNNIIAFYNQLQQLEQMTTTAENWLKIQPTTPSEPTAI KSQLKICKDEVNRLSGLQPQIERLKIQSIALKEKGQGPMFLDA DFVAFTNHFKQVFSDVQAREKELQTIFDTLPPMRYQETMSAI RTWVQQSETKLSIPQLSVTDYEIMEQRLGELQALQSSLQEQQ SGLYYLSTTVKEMSKKAPSEISRKYQSEFEEIEGRWKKLSSQL VEHCQKLEEQMNKLRKIQNHIQTLKKWMAEVDVFLKEEWP ALGDSEILKKQLKQCRLLVSDIQTIQPSLNSVNEGGQKIKNEA EPEFASRLETELKELNTQWDHMCQQVYARKEALKGGLEKT VSLQKDLSEMHEWMTQAEEEYLERDFEYKTPDELQKAVEE MKRAKEEAQQKEAKVKLLTESVNSVIAQAPPVAQEALKKEL ETLTTNYQWLCTRLNGKCKTLEEVWACWHELLSYLEKANK WLNEVEFKLKTTENIPGGAEEISEVLDSLENLMRHSEDNPNQI RILAQTLTDGGVMDELINEELETFNSRWRELHEEAVRRQKLL EQSIQSAQETEKSLHLIQESLTFIDKQLAAYIADKVDAAQMPQ EAQKIQSDLTSHEISLEEMKKHNQGKEAAQRVLSQIDVAQK KLQDVSMKFRLFQKPANFEQRLQESKMILDEVKMHLPALETKSVEQEVVQSQLNHCVNLYKSLSEVKSEVEMVIKTGRQIVQWSGR Ref. No. 54841-719.601KKQTENPKELDERVTALKLHYNELGAKVTERKQQLEKCLKL SRKMRKEMNVLTEWLAATDMELTKRSAVEGMPSNLDSEVA WGKATQKEIEKQKVHLKSITEVGEALKTVLGKKETLVEDKL SLLNSNWIAVTSRAEEWLNLLLEYQKHMETFDQNVDHITKW IIQADTLLDESEKKKPQQKEDVLKRLKAELNDIRPKVDSTRD QAANLMANRGDHCRKLVEPQISELNHRFAAISHRIKTGKASI PLKELEQFNSDIQKLLEPLEAEIQQGVNLKEEDFNKDMNEDN EGTVKELLQRGDNLQQRITDERKREEIKIKQQLLQTKHNALK DLRSQRRKKALEISHQWYQYKRQADDLLKCLDDIEKKLASL PEPRDERKIKEIDRELQKKKEELNAVRRQAEGLSEDGAAMA VEPTQIQLSKRWREIESKFAQFRRLNFAQIHTVREETMMVMT EDMPLEISYVPSTYLTEITHVSQALLEVEQLLNAPDLCAKDFE DLFKQEESLKNIKDSLQQSSGRIDIIHSKKTAALQSATPVERV KLQEALSQLDFQWEKVNKMYKDRQGRFDRSVEKWRRFHY DIKIFNQWLTEAEQFLRKTQIPENWEHAKYKWYLKELQDGI GQRQTWRTLNATGEEIIQQSSKTDASILQEKLGSLNLRWQE VCKQLSDRKKRLEEQKNILSEFQRDLNEFVLWLEEADNIASIP LEPGKEQQLKEKLEQVKLLVEELPLRQGILKQLNETGGPVLV SAPISPEEQDKLENKLKQTNLQWIKVSRALPEKQGEIEAQIKD LGQLEKKLEDLEEQLNHLLLWLSPIRNQLEIYNQPNQEGPFD VQETEIAVQAKQPDVEEILSKGQHLYKEKPATQPVKRKLEDL SSEWKAVNRLLQELRAKQPDLAPGLTTIGASPTQTVTLVTQP VVTKETAISKLEMPSSLMLEVPALADFNRAWTELTDWLSLL DQVIKSQRVMVGDLEDINEMIIKQKATMQDLEQRRPQLEELI TAAQNLKNKTSNQEARTIITDRIERIQNQWDEVQEHLQNRRQ QLNEMLKDSTQWLEAKEEAEQVLGQARAKLESWKEGPYTV DAIQKKITETKQLAKDLRQWQTNVDVANDLALKLLRDYSA DDTRKVHMITENINASWRSIHKRVSEREAALEETHRLLQQFP LDLEKFLAWLTEAETTANVLQDATRKERLLEDSKGVKELMK QWQDLQGEIEAHTDVYHNLDENSQKILRSLEGSDDAVLLQR RLDNMNFKWSELRKKSLNIRSHLEASSDQWKRLHLSLQELL VWLQLKDDELSRQAPIGGDFPAVQKQNDVHRAFKRELKTKE PVIMSTLETVRIFLTEQPLEGLEKLYQEPRELPPEERAQNVTR LLRKQAEEVNTEWEKLNLHSADWQRKIDETLERLQELQEAT DELDLKLRQAEVIKGSWQPVGDLLIDSLQDHLEKVKALRGEI APLKENVSHVNDLARQLTTLGIQLSPYNLSTLEDLNTRWKLL QVAVEDRVRQLHEAHRDFGPASQHFLSTSVQGPWERAISPN KVPYYINHETQTTCWDHPKMTELYQSLADLNNVRFSAYRTA MKLRRLQKALCLDLLSLSAACDALDQHNLKQNDQPMDILQI INCLTTIYDRLEQEHNNLVNVPLCVDMCLNWLLNVYDTGRT GRIRVLSFKTGIISLCKAHLEDKYRYLFKQVASSTGFCDQRRL GLLLHDSIQIPRQLGEVASFGGSNIEPSVRSCFQFANNKPEIEA ALFLDWMRLEPQSMVWLPVLHRVAAAETAKHQAKCNICKE CPIIGFRYRSLKHFNYDICQSCFFSGRVAKGHKMHYPMVEYC TPTTSGEDVRDFAKVLKNKFRTKRYFAKHPRMGYLPVQTVL EGDNMETPVTLINFWPVDSAPASSPQLSHDDTHSRIEHYASR LAEMENSNGSYLNDSISPNESIDDEHLLIQHYCQSLNQDSPLS QPRSPAQILISLESEERGELERILADLEEENRNLQAEYDRLKQ QHEHKGLSPLPSPPEMMPTSPQSPRDAELIAEAKLLRQHKGRLEARMQILEDHNKQLESQLHRLRQLLEQPQAEAKVNGTTVSWSGR Ref. No. 54841-719.601SPSTSLQRSDSSQPMLLRWGSQTSDSMGEEDLLSPPQDTSTG LEEVMEQLNNSFPSSRGRNTPGKPMREDTM2 Pl 1532-1 MLWWEEVEDCYEREDVQKKTFTKWVNAQFSKFGKQHIENL (UniProt); FSDLQDGRRLLDLLEGLTGQKLPKEKGSTRVHALNNVNKAL human RVLQNNNVDLVNIGSTDIVDGNHKLTLGLIWNIILHWQVKN Dystrophin VMKNIMAGLQQTNSEKILLSWVRQSTRNYPQVNVINFTTSW SDGLALNALIHSHRPDLFDWNSWCQQSATQRLEHAFNIAR YQLGIEKLLDPEDVDTTYPDKKSILMYITSLFQVLPQQVSIEAI QEVEMLPRPPKVTKEEHFQLHHQMHYSQQITVSLAQGYERT SSPKPRFKSYAYTQAAYVTTSDPTRSPFPSQHLEAPEDKSFGS SLMESEVNLDRYQTALEEVLSWLLSAEDTLQAQGEISNDVE VVKDQFHTHEGYMMDLTAHQGRVGNILQLGSKLIGTGKLSE DEETEVQEQMNLLNSRWECLRVASMEKQSNLHRVLMDLQN QKLKELNDWLTKTEERTRKMEEEPLGPDLEDLKRQVQQHK VLQEDLEQEQVRVNSLTHMWVVDESSGDHATAALEEQLK VLGDRWANICRWTEDRWVLLQDILLKWQRLTEEQCLFSAW LSEKEDAVNKIHTTGFKDQNEMLSSLQKLAVLKADLEKKKQ SMGKLYSLKQDLLSTLKNKSVTQKTEAWLDNFARCWDNLV QKLEKSTAQISQAVTTTQPSLTQTTVMETVTTVTTREQILVK HAQEELPPPPPQKKRQITVDSEIRKRLDVDITELHSWITRSEA VLQSPEFAIFRKEGNFSDLKEKVNAIEREKAEKFRKLQDASRS AQALVEQMVNEGVNADSIKQASEQLNSRWIEFCQLLSERLN WLEYQNNIIAFYNQLQQLEQMTTTAENWLKIQPTTPSEPTAI KSQLKICKDEVNRLSDLQPQIERLKIQSIALKEKGQGPMFLDA DFVAFTNHFKQVFSDVQAREKELQTIFDTLPPMRYQETMSAI RTWVQQSETKLSIPQLSVTDYEIMEQRLGELQALQSSLQEQQ SGLYYLSTTVKEMSKKAPSEISRKYQSEFEEIEGRWKKLSSQL VEHCQKLEEQMNKLRKIQNHIQTLKKWMAEVDVFLKEEWP ALGDSEILKKQLKQCRLLVSDIQTIQPSLNSVNEGGQKIKNEA EPEFASRLETELKELNTQWDHMCQQVYARKEALKGGLEKT VSLQKDLSEMHEWMTQAEEEYLERDFEYKTPDELQKAVEE MKRAKEEAQQKEAKVKLLTESVNSVIAQAPPVAQEALKKEL ETLTTNYQWLCTRLNGKCKTLEEVWACWHELLSYLEKANK WLNEVEFKLKTTENIPGGAEEISEVLDSLENLMRHSEDNPNQI RILAQTLTDGGVMDELINEELETFNSRWRELHEEAVRRQKLL EQSIQSAQETEKSLHLIQESLTFIDKQLAAYIADKVDAAQMPQ EAQKIQSDLTSHEISLEEMKKHNQGKEAAQRVLSQIDVAQK KLQDVSMKFRLFQKPANFEQRLQESKMILDEVKMHLPALET KSVEQEVVQSQLNHCVNLYKSLSEVKSEVEMVIKTGRQIVQ KKQTENPKELDERVTALKLHYNELGAKVTERKQQLEKCLKL SRKMRKEMNVLTEWLAATDMELTKRSAVEGMPSNLDSEVA WGKATQKEIEKQKVHLKSITEVGEALKTVLGKKETLVEDKL SLLNSNWIAVTSRAEEWLNLLLEYQKHMETFDQNVDHITKW IIQADTLLDESEKKKPQQKEDVLKRLKAELNDIRPKVDSTRD QAANLMANRGDHCRKLVEPQISELNHRFAAISHRIKTGKASI PLKELEQFNSDIQKLLEPLEAEIQQGVNLKEEDFNKDMNEDN EGTVKELLQRGDNLQQRITDERKREEIKIKQQLLQTKHNALK DLRSQRRKKALEISHQWYQYKRQADDLLKCLDDIEKKLASL PEPRDERKIKEIDRELQKKKEELNAVRRQAEGLSEDGAAMAVEPTQIQLSKRWREIESKFAQFRRLNFAQIHTVREETMMVMTWSGR Ref. No. 54841-719.601EDMPLEISYVPSTYLTEITHVSQALLEVEQLLNAPDLCAKDFE DLFKQEESLKNIKDSLQQSSGRIDIIHSKKTAALQSATPVERV KLQEALSQLDFQWEKVNKMYKDRQGRFDRSVEKWRRFHY DIKIFNQWLTEAEQFLRKTQIPENWEHAKYKWYLKELQDGI GQRQTWRTLNATGEEIIQQSSKTDASILQEKLGSLNLRWQE VCKQLSDRKKRLEEQKNILSEFQRDLNEFVLWLEEADNIASIP LEPGKEQQLKEKLEQVKLLVEELPLRQGILKQLNETGGPVLV SAPISPEEQDKLENKLKQTNLQWIKVSRALPEKQGEIEAQIKD LGQLEKKLEDLEEQLNHLLLWLSPIRNQLEIYNQPNQEGPFD VKETEIAVQAKQPDVEEILSKGQHLYKEKPATQPVKRKLEDL SSEWKAVNRLLQELRAKQPDLAPGLTTIGASPTQTVTLVTQP VVTKETAISKLEMPSSLMLEVPALADFNRAWTELTDWLSLL DQVIKSQRVMVGDLEDINEMIIKQKATMQDLEQRRPQLEELI TAAQNLKNKTSNQEARTIITDRIERIQNQWDEVQEHLQNRRQ QLNEMLKDSTQWLEAKEEAEQVLGQARAKLESWKEGPYTV DAIQKKITETKQLAKDLRQWQTNVDVANDLALKLLRDYSA DDTRKVHMITENINASWRSIHKRVSEREAALEETHRLLQQFP LDLEKFLAWLTEAETTANVLQDATRKERLLEDSKGVKELMK QWQDLQGEIEAHTDVYHNLDENSQKILRSLEGSDDAVLLQR RLDNMNFKWSELRKKSLNIRSHLEASSDQWKRLHLSLQELL VWLQLKDDELSRQAPIGGDFPAVQKQNDVHRAFKRELKTKE PVIMSTLETVRIFLTEQPLEGLEKLYQEPRELPPEERAQNVTR LLRKQAEEVNTEWEKLNLHSADWQRKIDETLERLRELQEAT DELDLKLRQAEVIKGSWQPVGDLLIDSLQDHLEKVKALRGEI APLKENVSHVNDLARQLTTLGIQLSPYNLSTLEDLNTRWKLL QVAVEDRVRQLHEAHRDFGPASQHFLSTSVQGPWERAISPN KVPYYINHETQTTCWDHPKMTELYQSLADLNNVRFSAYRTA MKLRRLQKALCLDLLSLSAACDALDQHNLKQNDQPMDILQI INCLTTIYDRLEQEHNNLVNVPLCVDMCLNWLLNVYDTGRT GRIRVLSFKTGIISLCKAHLEDKYRYLFKQVASSTGFCDQRRL GLLLHDSIQIPRQLGEVASFGGSNIEPSVRSCFQFANNKPEIEA ALFLDWMRLEPQSMVWLPVLHRVAAAETAKHQAKCNICKE CPIIGFRYRSLKHFNYDICQSCFFSGRVAKGHKMHYPMVEYC TPTTSGEDVRDFAKVLKNKFRTKRYFAKHPRMGYLPVQTVL EGDNMETPVTLINFWPVDSAPASSPQLSHDDTHSRIEHYASR LAEMENSNGSYLNDSISPNESIDDEHLLIQHYCQSLNQDSPLS QPRSPAQILISLESEERGELERILADLEEENRNLQAEYDRLKQ QHEHKGLSPLPSPPEMMPTSPQSPRDAELIAEAKLLRQHKGR LEARMQILEDHNKQLESQLHRLRQLLEQPQAEAKVNGTTVS SPSTSLQRSDSSQPMLLRWGSQTSDSMGEEDLLSPPQDTSTG LEEVMEQLNNSFPSSRGRNTPGKPMREDTM3 Pl 1532-4 MEDEREDVQKKTFTKWVNAQFSKFGKQHIENLFSDLQDGRR (UniProt); LLDLLEGLTGQKLPKEKGSTRVHALNNVNKALRVLQNNNV human DLVNIGSTDIVDGNHKLTLGLIWNIILHWQVKNVMKNIMAG Dystrophin LQQTNSEKILLSWVRQSTRNYPQVNVINFTTSWSDGLALNAL -3, Dp427c IHSHRPDLFDWNSWCQQSATQRLEHAFNIARYQLGIEKLLD amino acid PEDVDTTYPDKKSILMYITSLFQVLPQQVSIEAIQEVEMLPRP sequence PKVTKEEHFQLHHQMHYSQQITVSLAQGYERTSSPKPRFKSYAYTQAAYVTTSDPTRSPFPSQHLEAPEDKSFGSSLMESEVNLDRYQTALEEVLSWLLSAEDTLQAQGEISNDVEWKDQFHTHWSGR Ref. No. 54841-719.601EGYMMDLTAHQGRVGNILQLGSKLIGTGKLSEDEETEVQEQ MNLLNSRWECLRVASMEKQSNLHRVLMDLQNQKLKELND WLTKTEERTRKMEEEPLGPDLEDLKRQVQQHKVLQEDLEQE QVRVNSLTHMVWVDESSGDHATAALEEQLKVLGDRWANI CRWTEDRWVLLQDILLKWQRLTEEQCLFSAWLSEKEDAVN KIHTTGFKDQNEMLSSLQKLAVLKADLEKKKQSMGKLYSLK QDLLSTLKNKSVTQKTEAWLDNFARCWDNLVQKLEKSTAQI SQAVTTTQPSLTQTTVMETVTTVTTREQILVKHAQEELPPPPP QKKRQITVDSEIRKRLDVDITELHSWITRSEAVLQSPEFAIFRK EGNFSDLKEKVNAIEREKAEKFRKLQDASRSAQALVEQMVN EGVNADSIKQASEQLNSRWIEFCQLLSERLNWLEYQNNIIAF YNQLQQLEQMTTTAENWLKIQPTTPSEPTAIKSQLKICKDEV NRLSDLQPQIERLKIQSIALKEKGQGPMFLDADFVAFTNHFK QVFSDVQAREKELQTIFDTLPPMRYQETMSAIRTWVQQSETK LSIPQLSVTDYEIMEQRLGELQALQSSLQEQQSGLYYLSTTVK EMSKKAPSEISRKYQSEFEEIEGRWKKLSSQLVEHCQKLEEQ MNKLRKIQNHIQTLKKWMAEVDVFLKEEWPALGDSEILKKQ LKQCRLLVSDIQTIQPSLNSVNEGGQKIKNEAEPEFASRLETE LKELNTQWDHMCQQVYARKEALKGGLEKTVSLQKDLSEM HEWMTQAEEEYLERDFEYKTPDELQKAVEEMKRAKEEAQQ KEAKVKLLTESVNSVIAQAPPVAQEALKKELETLTTNYQWL CTRLNGKCKTLEEVWACWHELLSYLEKANKWLNEVEFKLK TTENIPGGAEEISEVLDSLENLMRHSEDNPNQIRILAQTLTDG GVMDELINEELETFNSRWRELHEEAVRRQKLLEQSIQSAQET EKSLHLIQESLTFIDKQLAAYIADKVDAAQMPQEAQKIQSDL TSHEISLEEMKKHNQGKEAAQRVLSQIDVAQKKLQDVSMKF RLFQKPANFEQRLQESKMILDEVKMHLPALETKSVEQEWQ SQLNHCVNLYKSLSEVKSEVEMVIKTGRQIVQKKQTENPKEL DERVTALKLHYNELGAKVTERKQQLEKCLKLSRKMRKEMN VLTEWLAATDMELTKRSAVEGMPSNLDSEVAWGKATQKEI EKQKVHLKSITEVGEALKTVLGKKETLVEDKLSLLNSNWIA VTSRAEEWLNLLLEYQKHMETFDQNVDHITKWIIQADTLLD ESEKKKPQQKEDVLKRLKAELNDIRPKVDSTRDQAANLMAN RGDHCRKLVEPQISELNHRFAAISHRIKTGKASIPLKELEQFN SDIQKLLEPLEAEIQQGVNLKEEDFNKDMNEDNEGTVKELLQ RGDNLQQRITDERKREEIKIKQQLLQTKHNALKDLRSQRRKK ALEISHQWYQYKRQADDLLKCLDDIEKKLASLPEPRDERKIK EIDRELQKKKEELNAVRRQAEGLSEDGAAMAVEPTQIQLSK RWREIESKFAQFRRLNFAQIHTVREETMMVMTEDMPLEISYV PSTYLTEITHVSQALLEVEQLLNAPDLCAKDFEDLFKQEESLK NIKDSLQQSSGRIDIIHSKKTAALQSATPVERVKLQEALSQLD FQWEKVNKMYKDRQGRFDRSVEKWRRFHYDIKIFNQWLTE AEQFLRKTQIPENWEHAKYKWYLKELQDGIGQRQTWRTLN ATGEEIIQQSSKTDASILQEKLGSLNLRWQEVCKQLSDRKKR LEEQKNILSEFQRDLNEFVLWLEEADNIASIPLEPGKEQQLKE KLEQVKLLVEELPLRQGILKQLNETGGPVLVSAPISPEEQDKL ENKLKQTNLQWIKVSRALPEKQGEIEAQIKDLGQLEKKLEDL EEQLNHLLLWLSPIRNQLEIYNQPNQEGPFDVKETEIAVQAK QPDVEEILSKGQHLYKEKPATQPVKRKLEDLSSEWKAVNRLLQELRAKQPDLAPGLTTIGASPTQTVTLVTQPVVTKETAISKLWSGR Ref. No. 54841-719.601EMPSSLMLEVPALADFNRAWTELTDWLSLLDQVIKSQRVMV GDLEDINEMIIKQKATMQDLEQRRPQLEELITAAQNLKNKTS NQEARTIITDRIERIQNQWDEVQEHLQNRRQQLNEMLKDSTQ WLEAKEEAEQVLGQARAKLESWKEGPYTVDAIQKKITETKQ LAKDLRQWQTNVDVANDLALKLLRDYSADDTRKVHMITEN INASWRSIHKRVSEREAALEETHRLLQQFPLDLEKFLAWLTE AETTANVLQDATRKERLLEDSKGVKELMKQWQDLQGEIEA HTDVYHNLDENSQKILRSLEGSDDAVLLQRRLDNMNFKWSE LRKKSLNIRSHLEASSDQWKRLHLSLQELLVWLQLKDDELS RQAPIGGDFPAVQKQNDVHRAFKRELKTKEPVIMSTLETVRI FLTEQPLEGLEKLYQEPRELPPEERAQNVTRLLRKQAEEVNT EWEKLNLHSADWQRKIDETLERLRELQEATDELDLKLRQAE VIKGSWQPVGDLLIDSLQDHLEKVKALRGEIAPLKENVSHVN DLARQLTTLGIQLSPYNLSTLEDLNTRWKLLQVAVEDRVRQ LHEAHRDFGPASQHFLSTSVQGPWERAISPNKVPYYINHETQ TTCWDHPKMTELYQSLADLNNVRFSAYRTAMKLRRLQKAL CLDLLSLSAACDALDQHNLKQNDQPMDILQIINCLTTIYDRL EQEHNNLVNVPLCVDMCLNWLLNVYDTGRTGRIRVLSFKT GIISLCKAHLEDKYRYLFKQVASSTGFCDQRRLGLLLHDSIQI PRQLGEVASFGGSNIEPSVRSCFQFANNKPEIEAALFLDWMR LEPQSMVWLPVLHRVAAAETAKHQAKCNICKECPIIGFRYRS LKHFNYDICQSCFFSGRVAKGHKMHYPMVEYCTPTTSGEDV RDFAKVLKNKFRTKRYFAKHPRMGYLPVQTVLEGDNMETP VTLINFWPVDSAPASSPQLSHDDTHSRIEHYASRLAEMENSN GSYLNDSISPNESIDDEHLLIQHYCQSLNQDSPLSQPRSPAQILI SLESEERGELERILADLEEENRNLQAEYDRLKQQHEHKGLSP LPSPPEMMPTSPQSPRDAELIAEAKLLRQHKGRLEARMQILE DHNKQLESQLHRLRQLLEQPQAEAKVNGTTVSSPSTSLQRSD SSQPMLLRWGSQTSDSMGEEDLLSPPQDTSTGLEEVMEQLN NSFPSSRGRNTPGKPMREDTM4 Pl 1532-11 MSEVSSDEREDVQKKTFTKWVNAQFSKFGKQHIENLFSDLQ (UniProt); DGRRLLDLLEGLTGQKLPKEKGSTRVHALNNVNKALRVLQ human NNNVDLVNIGSTDIVDGNHKLTLGLIWNIILHWQVKNVMKN Dystrophin IMAGLQQTNSEKILLSWVRQSTRNYPQVNVINFTTSWSDGLA Dp427p LNALIHSHRPDLFDWNSVVCQQSATQRLEHAFNIARYQLGIE amino acid KLLDPEDVDTTYPDKKSILMYITSLFQVLPQQVSIEAIQEVEM sequence LPRPPKVTKEEHFQLHHQMHYSQQITVSLAQGYERTSSPKPR FKSYAYTQAAYVTTSDPTRSPFPSQHLEAPEDKSFGSSLMESE VNLDRYQTALEEVLSWLLSAEDTLQAQGEISNDVEVVKDQF HTHEGYMMDLTAHQGRVGNILQLGSKLIGTGKLSEDEETEV QEQMNLLNSRWECLRVASMEKQSNLHRVLMDLQNQKLKEL NDWLTKTEERTRKMEEEPLGPDLEDLKRQVQQHKVLQEDL EQEQVRVNSLTHMWVVDESSGDHATAALEEQLKVLGDRW ANICRWTEDRWVLLQDILLKWQRLTEEQCLFSAWLSEKEDA VNKIHTTGFKDQNEMLSSLQKLAVLKADLEKKKQSMGKLY SLKQDLLSTLKNKSVTQKTEAWLDNFARCWDNLVQKLEKS TAQISQAVTTTQPSLTQTTVMETVTTVTTREQILVKHAQEEL PPPPPQKKRQITVDSEIRKRLDVDITELHSWITRSEAVLQSPEF AIFRKEGNFSDLKEKVNAIEREKAEKFRKLQDASRSAQALVEQMVNEGVNADSIKQASEQLNSRWIEFCQLLSERLNWLEYQNWSGR Ref. No. 54841-719.601NIIAFYNQLQQLEQMTTTAENWLKIQPTTPSEPTAIKSQLKIC KDEVNRLSDLQPQIERLKIQSIALKEKGQGPMFLDADFVAFT NHFKQVFSDVQAREKELQTIFDTLPPMRYQETMSAIRTWVQ QSETKLSIPQLSVTDYEIMEQRLGELQALQSSLQEQQSGLYYL STTVKEMSKKAPSEISRKYQSEFEEIEGRWKKLSSQLVEHCQ KLEEQMNKLRKIQNHIQTLKKWMAEVDVFLKEEWPALGDS EILKKQLKQCRLLVSDIQTIQPSLNSVNEGGQKIKNEAEPEFA SRLETELKELNTQWDHMCQQVYARKEALKGGLEKTVSLQK DLSEMHEWMTQAEEEYLERDFEYKTPDELQKAVEEMKRAK EEAQQKEAKVKLLTESVNSVIAQAPPVAQEALKKELETLTTN YQWLCTRLNGKCKTLEEVWACWHELLSYLEKANKWLNEV EFKLKTTENIPGGAEEISEVLDSLENLMRHSEDNPNQIRILAQ TLTDGGVMDELINEELETFNSRWRELHEEAVRRQKLLEQSIQ SAQETEKSLHLIQESLTFIDKQLAAYIADKVDAAQMPQEAQK IQSDLTSHEISLEEMKKHNQGKEAAQRVLSQIDVAQKKLQD VSMKFRLFQKPANFEQRLQESKMILDEVKMHLPALETKSVE QEVVQSQLNHCVNLYKSLSEVKSEVEMVIKTGRQIVQKKQT ENPKELDERVTALKLHYNELGAKVTERKQQLEKCLKLSRKM RKEMNVLTEWLAATDMELTKRSAVEGMPSNLDSEVAWGK ATQKEIEKQKVHLKSITEVGEALKTVLGKKETLVEDKLSLLN SNWIAVTSRAEEWLNLLLEYQKHMETFDQNVDHITKWIIQA DTLLDESEKKKPQQKEDVLKRLKAELNDIRPKVDSTRDQAA NLMANRGDHCRKLVEPQISELNHRFAAISHRIKTGKASIPLKE LEQFNSDIQKLLEPLEAEIQQGVNLKEEDFNKDMNEDNEGTV KELLQRGDNLQQRITDERKREEIKIKQQLLQTKHNALKDLRS QRRKKALEISHQWYQYKRQADDLLKCLDDIEKKLASLPEPR DERKIKEIDRELQKKKEELNAVRRQAEGLSEDGAAMAVEPT QIQLSKRWREIESKFAQFRRLNFAQIHTVREETMMVMTEDM PLEISYVPSTYLTEITHVSQALLEVEQLLNAPDLCAKDFEDLF KQEESLKNIKDSLQQSSGRIDIIHSKKTAALQSATPVERVKLQ EALSQLDFQWEKVNKMYKDRQGRFDRSVEKWRRFHYDIKI FNQWLTEAEQFLRKTQIPENWEHAKYKWYLKELQDGIGQR QTVVRTLNATGEEIIQQSSKTDASILQEKLGSLNLRWQEVCK QLSDRKKRLEEQKNILSEFQRDLNEFVLWLEEADNIASIPLEP GKEQQLKEKLEQVKLLVEELPLRQGILKQLNETGGPVLVSAP ISPEEQDKLENKLKQTNLQWIKVSRALPEKQGEIEAQIKDLG QLEKKLEDLEEQLNHLLLWLSPIRNQLEIYNQPNQEGPFDVK ETEIAVQAKQPDVEEILSKGQHLYKEKPATQPVKRKLEDLSS EWKAVNRLLQELRAKQPDLAPGLTTIGASPTQTVTLVTQPV VTKETAISKLEMPSSLMLEVPALADFNRAWTELTDWLSLLD QVIKSQRVMVGDLEDINEMIIKQKATMQDLEQRRPQLEELIT AAQNLKNKTSNQEARTIITDRIERIQNQWDEVQEHLQNRRQQ LNEMLKDSTQWLEAKEEAEQVLGQARAKLESWKEGPYTVD AIQKKITETKQLAKDLRQWQTNVDVANDLALKLLRDYSAD DTRKVHMITENINASWRSIHKRVSEREAALEETHRLLQQFPL DLEKFLAWLTEAETTANVLQDATRKERLLEDSKGVKELMK QWQDLQGEIEAHTDVYHNLDENSQKILRSLEGSDDAVLLQR RLDNMNFKWSELRKKSLNIRSHLEASSDQWKRLHLSLQELL VWLQLKDDELSRQAPIGGDFPAVQKQNDVHRAFKRELKTKEPVIMSTLETVRIFLTEQPLEGLEKLYQEPRELPPEERAQNVTRWSGR Ref. No. 54841-719.601LLRKQAEEVNTEWEKLNLHSADWQRKIDETLERLRELQEAT DELDLKLRQAEVIKGSWQPVGDLLIDSLQDHLEKVKALRGEI APLKENVSHVNDLARQLTTLGIQLSPYNLSTLEDLNTRWKLL QVAVEDRVRQLHEAHRDFGPASQHFLSTSVQGPWERAISPN KVPYYINHETQTTCWDHPKMTELYQSLADLNNVRFSAYRTA MKLRRLQKALCLDLLSLSAACDALDQHNLKQNDQPMDILQI INCLTTIYDRLEQEHNNLVNVPLCVDMCLNWLLNVYDTGRT GRIRVLSFKTGIISLCKAHLEDKYRYLFKQVASSTGFCDQRRL GLLLHDSIQIPRQLGEVASFGGSNIEPSVRSCFQFANNKPEIEA ALFLDWMRLEPQSMVWLPVLHRVAAAETAKHQAKCNICKE CPIIGFRYRSLKHFNYDICQSCFFSGRVAKGHKMHYPMVEYC TPTTSGEDVRDFAKVLKNKFRTKRYFAKHPRMGYLPVQTVL EGDNMETPVTLINFWPVDSAPASSPQLSHDDTHSRIEHYASR LAEMENSNGSYLNDSISPNESIDDEHLLIQHYCQSLNQDSPLS QPRSPAQILISLESEERGELERILADLEEENRNLQAEYDRLKQ QHEHKGLSPLPSPPEMMPTSPQSPRDAELIAEAKLLRQHKGR LEARMQILEDHNKQLESQLHRLRQLLEQPQAEAKVNGTTVS SPSTSLQRSDSSQPMLLRWGSQTSDSMGEEDLLSPPQDTSTG LEEVMEQLNNSFPSSRGRNTPGKPMREDTM5 Pl 1532-2 MTEIILLIFFPAYFLNAVRRQKLLEQSIQSAQETEKSLHLIQES (UniProt); LTFIDKQLAAYIADKVDAAQMPQEAQKIQSDLTSHEISLEEM human KKHNQGKEAAQRVLSQIDVAQKKLQDVSMKFRLFQKPANF Dystrophin EQRLQESKMILDEVKMHLPALETKSVEQEWQSQLNHCVNL -1, Dp260- YKSLSEVKSEVEMVIKTGRQIVQKKQTENPKELDERVTALKL 1 amino HYNELGAKVTERKQQLEKCLKLSRKMRKEMNVLTEWLAAT acid DMELTKRSAVEGMPSNLDSEVAWGKATQKEIEKQKVHLKSI sequence TEVGEALKTVLGKKETLVEDKLSLLNSNWIAVTSRAEEWLN LLLEYQKHMETFDQNVDHITKWIIQADTLLDESEKKKPQQK EDVLKRLKAELNDIRPKVDSTRDQAANLMANRGDHCRKLV EPQISELNHRFAAISHRIKTGKASIPLKELEQFNSDIQKLLEPLE AEIQQGVNLKEEDFNKDMNEDNEGTVKELLQRGDNLQQRIT DERKREEIKIKQQLLQTKHNALKDLRSQRRKKALEISHQWY QYKRQADDLLKCLDDIEKKLASLPEPRDERKIKEIDRELQKK KEELNAVRRQAEGLSEDGAAMAVEPTQIQLSKRWREIESKF AQFRRLNFAQIHTVREETMMVMTEDMPLEISYVPSTYLTEIT HVSQALLEVEQLLNAPDLCAKDFEDLFKQEESLKNIKDSLQQ SSGRIDIIHSKKTAALQSATPVERVKLQEALSQLDFQWEKVN KMYKDRQGRFDRSVEKWRRFHYDIKIFNQWLTEAEQFLRKT QIPENWEHAKYKWYLKELQDGIGQRQTWRTLNATGEEIIQ QSSKTDASILQEKLGSLNLRWQEVCKQLSDRKKRLEEQKNIL SEFQRDLNEFVLWLEEADNIASIPLEPGKEQQLKEKLEQVKL LVEELPLRQGILKQLNETGGPVLVSAPISPEEQDKLENKLKQT NLQWIKVSRALPEKQGEIEAQIKDLGQLEKKLEDLEEQLNHL LLWLSPIRNQLEIYNQPNQEGPFDVKETEIAVQAKQPDVEEIL SKGQHLYKEKPATQPVKRKLEDLSSEWKAVNRLLQELRAK QPDLAPGLTTIGASPTQTVTLVTQPVVTKETAISKLEMPSSLM LEVPALADFNRAWTELTDWLSLLDQVIKSQRVMVGDLEDIN EMIIKQKATMQDLEQRRPQLEELITAAQNLKNKTSNQEARTII TDRIERIQNQWDEVQEHLQNRRQQLNEMLKDSTQWLEAKEEAEQVLGQARAKLESWKEGPYTVDAIQKKITETKQLAKDLRWSGR Ref. No. 54841-719.601QWQTNVDVANDLALKLLRDYSADDTRKVHMITENINASWR SIHKRVSEREAALEETHRLLQQFPLDLEKFLAWLTEAETTAN VLQDATRKERLLEDSKGVKELMKQWQDLQGEIEAHTDVYH NLDENSQKILRSLEGSDDAVLLQRRLDNMNFKWSELRKKSL NIRSHLEASSDQWKRLHLSLQELLVWLQLKDDELSRQAPIGG DFPAVQKQNDVHRAFKRELKTKEPVIMSTLETVRIFLTEQPL EGLEKLYQEPRELPPEERAQNVTRLLRKQAEEVNTEWEKLN LHSADWQRKIDETLERLRELQEATDELDLKLRQAEVIKGSW QPVGDLLIDSLQDHLEKVKALRGEIAPLKENVSHVNDLARQL TTLGIQLSPYNLSTLEDLNTRWKLLQVAVEDRVRQLHEAHR DFGPASQHFLSTSVQGPWERAISPNKVPYYINHETQTTCWDH PKMTELYQSLADLNNVRFSAYRTAMKLRRLQKALCLDLLSL SAACDALDQHNLKQNDQPMDILQIINCLTTIYDRLEQEHNNL VNVPLCVDMCLNWLLNVYDTGRTGRIRVLSFKTGIISLCKAH LEDKYRYLFKQVASSTGFCDQRRLGLLLHDSIQIPRQLGEVA SFGGSNIEPSVRSCFQFANNKPEIEAALFLDWMRLEPQSMVW LPVLHRVAAAETAKHQAKCNICKECPIIGFRYRSLKHFNYDI CQSCFFSGRVAKGHKMHYPMVEYCTPTTSGEDVRDFAKVL KNKFRTKRYFAKHPRMGYLPVQTVLEGDNMETPVTLINFWP VDSAPASSPQLSHDDTHSRIEHYASRLAEMENSNGSYLNDSIS PNESIDDEHLLIQHYCQSLNQDSPLSQPRSPAQILISLESEERG ELERILADLEEENRNLQAEYDRLKQQHEHKGLSPLPSPPEMM PTSPQSPRDAELIAEAKLLRQHKGRLEARMQILEDHNKQLES QLHRLRQLLEQPQAEAKVNGTTVSSPSTSLQRSDSSQPMLLR VVGSQTSDSMGEEDLLSPPQDTSTGLEEVMEQLNNSFPSSRG RNTPGKPMREDTM6 Pl 1532-3 MSARKLRNLSYKKAVRRQKLLEQSIQSAQETEKSLHLIQESL (UniProt); TFIDKQLAAYIADKVDAAQMPQEAQKIQSDLTSHEISLEEMK human KHNQGKEAAQRVLSQIDVAQKKLQDVSMKFRLFQKPANFE Dystrophin QRLQESKMILDEVKMHLPALETKSVEQEWQSQLNHCVNLY -2, Dp260- KSLSEVKSEVEMVIKTGRQIVQKKQTENPKELDERVTALKLH 2 amino YNELGAKVTERKQQLEKCLKLSRKMRKEMNVLTEWLAATD acid MELTKRSAVEGMPSNLDSEVAWGKATQKEIEKQKVHLKSIT sequence EVGEALKTVLGKKETLVEDKLSLLNSNWIAVTSRAEEWLNL LLEYQKHMETFDQNVDHITKWIIQADTLLDESEKKKPQQKE DVLKRLKAELNDIRPKVDSTRDQAANLMANRGDHCRKLVE PQISELNHRFAAISHRIKTGKASIPLKELEQFNSDIQKLLEPLE AEIQQGVNLKEEDFNKDMNEDNEGTVKELLQRGDNLQQRIT DERKREEIKIKQQLLQTKHNALKDLRSQRRKKALEISHQWY QYKRQADDLLKCLDDIEKKLASLPEPRDERKIKEIDRELQKK KEELNAVRRQAEGLSEDGAAMAVEPTQIQLSKRWREIESKF AQFRRLNFAQIHTVREETMMVMTEDMPLEISYVPSTYLTEIT HVSQALLEVEQLLNAPDLCAKDFEDLFKQEESLKNIKDSLQQ SSGRIDIIHSKKTAALQSATPVERVKLQEALSQLDFQWEKVN KMYKDRQGRFDRSVEKWRRFHYDIKIFNQWLTEAEQFLRKT QIPENWEHAKYKWYLKELQDGIGQRQTWRTLNATGEEIIQ QSSKTDASILQEKLGSLNLRWQEVCKQLSDRKKRLEEQKNIL SEFQRDLNEFVLWLEEADNIASIPLEPGKEQQLKEKLEQVKL LVEELPLRQGILKQLNETGGPVLVSAPISPEEQDKLENKLKQTNLQWIKVSRALPEKQGEIEAQIKDLGQLEKKLEDLEEQLNHLWSGR Ref. No. 54841-719.601LLWLSPIRNQLEIYNQPNQEGPFDVKETEIAVQAKQPDVEEIL SKGQHLYKEKPATQPVKRKLEDLSSEWKAVNRLLQELRAK QPDLAPGLTTIGASPTQTVTLVTQPVVTKETAISKLEMPSSLM LEVPALADFNRAWTELTDWLSLLDQVIKSQRVMVGDLEDIN EMIIKQKATMQDLEQRRPQLEELITAAQNLKNKTSNQEARTII TDRIERIQNQWDEVQEHLQNRRQQLNEMLKDSTQWLEAKE EAEQVLGQARAKLESWKEGPYTVDAIQKKITETKQLAKDLR QWQTNVDVANDLALKLLRDYSADDTRKVHMITENINASWR SIHKRVSEREAALEETHRLLQQFPLDLEKFLAWLTEAETTAN VLQDATRKERLLEDSKGVKELMKQWQDLQGEIEAHTDVYH NLDENSQKILRSLEGSDDAVLLQRRLDNMNFKWSELRKKSL NIRSHLEASSDQWKRLHLSLQELLVWLQLKDDELSRQAPIGG DFPAVQKQNDVHRAFKRELKTKEPVIMSTLETVRIFLTEQPL EGLEKLYQEPRELPPEERAQNVTRLLRKQAEEVNTEWEKLN LHSADWQRKIDETLERLRELQEATDELDLKLRQAEVIKGSW QPVGDLLIDSLQDHLEKVKALRGEIAPLKENVSHVNDLARQL TTLGIQLSPYNLSTLEDLNTRWKLLQVAVEDRVRQLHEAHR DFGPASQHFLSTSVQGPWERAISPNKVPYYINHETQTTCWDH PKMTELYQSLADLNNVRFSAYRTAMKLRRLQKALCLDLLSL SAACDALDQHNLKQNDQPMDILQIINCLTTIYDRLEQEHNNL VNVPLCVDMCLNWLLNVYDTGRTGRIRVLSFKTGIISLCKAH LEDKYRYLFKQVASSTGFCDQRRLGLLLHDSIQIPRQLGEVA SFGGSNIEPSVRSCFQFANNKPEIEAALFLDWMRLEPQSMVW LPVLHRVAAAETAKHQAKCNICKECPIIGFRYRSLKHFNYDI CQSCFFSGRVAKGHKMHYPMVEYCTPTTSGEDVRDFAKVL KNKFRTKRYFAKHPRMGYLPVQTVLEGDNMETPVTLINFWP VDSAPASSPQLSHDDTHSRIEHYASRLAEMENSNGSYLNDSIS PNESIDDEHLLIQHYCQSLNQDSPLSQPRSPAQILISLESEERG ELERILADLEEENRNLQAEYDRLKQQHEHKGLSPLPSPPEMM PTSPQSPRDAELIAEAKLLRQHKGRLEARMQILEDHNKQLES QLHRLRQLLEQPQAEAKVNGTTVSSPSTSLQRSDSSQPMLLR VVGSQTSDSMGEEDLLSPPQDTSTGLEEVMEQLNNSFPSSRGRNTPGKPMREDTMTABLE 2: Nucleic Acid Sequences of Dystrophin mRNA Cargo in Extracellular Vesicles SEQ Description Nucleic Acid SequenceID NO:21 Human AUGCUUUGGUGGGAAGAAGUAGAGGACUGUUAUGAAAG Dystrophin AGAAGAUGUUCAAAAGAAAACAUUCACAAAAUGGGUAA (DMD) AUGCACAAUUUUCUAAGUUUGGGAAGCAGCAUAUUGAG mRNA AACCUCUUCAGUGACCUACAGGAUGGGAGGCGCCUCCUA coding GACCUCCUCGAAGGCCUGACAGGGCAAAAACUGCCAAAA sequence GAAAAAGGAUCCACAAGAGUUCAUGCCCUGAACAAUGU CAACAAGGCACUGCGGGUUUUGCAGAACAAUAAUGUUGRef. Seq. AUUUAGUGAAUAUUGGAAGUACUGACAUCGUAGAUGGA NM 00400 AAUCAUAAACUGACUCUUGGUUUGAUUUGGAAUAUAAU 6.2 CCUCCACUGGCAGGUCAAAAAUGUAAUGAAAAAUAUCAUGGCUGGAUUGCAACAAACCAACAGUGAAAAGAUUCUCCUGAGCUGGGUCCGACAAUCAACUCGUAAUUAUCCACAGWSGR Ref. No. 54841-719.601GUUAAUGUAAUCAACUUCACCACCAGCUGGUCUGAUGG CCUGGCUUUGAAUGCUCUCAUCCAUAGUCAUAGGCCAGA CCUAUUUGACUGGAAUAGUGUGGUUUGCCAGCAGUCAG CCACACAACGACUGGAACAUGCAUUCAACAUCGCCAGAU AUCAAUUAGGCAUAGAGAAACUACUCGAUCCUGAAGAU GUUGAUACCACCUAUCCAGAUAAGAAGUCCAUCUUAAU GUACAUCACAUCACUCUUCCAAGUUUUGCCUCAACAAGU GAGCAUUGAAGCCAUCCAGGAAGUGGAAAUGUUGCCAA GGCCACCUAAAGUGACUAAAGAAGAACAUUUUCAGUUA CAUCAUCAAAUGCACUAUUCUCAACAGAUCACGGUCAGU CUAGCACAGGGAUAUGAGAGAACUUCUUCCCCUAAGCCU CGAUUCAAGAGCUAUGCCUACACACAGGCUGCUUAUGUC ACCACCUCUGACCCUACACGGAGCCCAUUUCCUUCACAG CAUUUGGAAGCUCCUGAAGACAAGUCAUUUGGCAGUUC AUUGAUGGAGAGUGAAGUAAACCUGGACCGUUAUCAAA CAGCUUUAGAAGAAGUAUUAUCGUGGCUUCUUUCUGCU GAGGACACAUUGCAAGCACAAGGAGAGAUUUCUAAUGA UGUGGAAGUGGUGAAAGACCAGUUUCAUACUCAUGAGG GGUACAUGAUGGAUUUGACAGCCCAUCAGGGCCGGGUU GGUAAUAUUCUACAAUUGGGAAGUAAGCUGAUUGGAAC AGGAAAAUUAUCAGAAGAUGAAGAAACUGAAGUACAAG AGCAGAUGAAUCUCCUAAAUUCAAGAUGGGAAUGCCUC AGGGUAGCUAGCAUGGAAAAACAAAGCAAUUUACAUAG AGUUUUAAUGGAUCUCCAGAAUCAGAAACUGAAAGAGU UGAAUGACUGGCUAACAAAAACAGAAGAAAGAACAAGG AAAAUGGAGGAAGAGCCUCUUGGACCUGAUCUUGAAGA CCUAAAACGCCAAGUACAACAACAUAAGGUGCUUCAAG AAGAUCUAGAACAAGAACAAGUCAGGGUCAAUUCUCUC ACUCACAUGGUGGUGGUAGUUGAUGAAUCUAGUGGAGA UCACGCAACUGCUGCUUUGGAAGAACAACUUAAGGUAU UGGGAGAUCGAUGGGCAAACAUCUGUAGAUGGACAGAA GACCGCUGGGUUCUUUUACAAGACAUCCUUCUCAAAUG GCAACGUCUUACUGAAGAACAGUGCCUUUUUAGUGCAU GGCUUUCAGAAAAAGAAGAUGCAGUGAACAAGAUUCAC ACAACUGGCUUUAAAGAUCAAAAUGAAAUGUUAUCAAG UCUUCAAAAACUGGCCGUUUUAAAAGCGGAUCUAGAAA AGAAAAAGCAAUCCAUGGGCAAACUGUAUUCACUCAAA CAAGAUCUUCUUUCAACACUGAAGAAUAAGUCAGUGAC CCAGAAGACGGAAGCAUGGCUGGAUAACUUUGCCCGGU GUUGGGAUAAUUUAGUCCAAAAACUUGAAAAGAGUACA GCACAGAUUUCACAGGCUGUCACCACCACUCAGCCAUCA CUAACACAGACAACUGUAAUGGAAACAGUAACUACGGU GACCACAAGGGAACAGAUCCUGGUAAAGCAUGCUCAAG AGGAACUUCCACCACCACCUCCCCAAAAGAAGAGGCAGA UUACUGUGGAUUCUGAAAUUAGGAAAAGGUUGGAUGUU GAUAUAACUGAACUUCACAGCUGGAUUACUCGCUCAGA AGCUGUGUUGCAGAGUCCUGAAUUUGCAAUCUUUCGGA AGGAAGGCAACUUCUCAGACUUAAAAGAAAAAGUCAAU GCCAUAGAGCGAGAAAAAGCUGAGAAGUUCAGAAAACUGCAAGAUGCCAGCAGAUCAGCUCAGGCCCUGGUGGAACAWSGR Ref. No. 54841-719.601GAUGGUGAAUGAGGGUGUUAAUGCAGAUAGCAUCAAAC AAGCCUCAGAACAACUGAACAGCCGGUGGAUCGAAUUC UGCCAGUUGCUAAGUGAGAGACUUAACUGGCUGGAGUA UCAGAACAACAUCAUCGCUUUCUAUAAUCAGCUACAACA AUUGGAGCAGAUGACAACUACUGCUGAAAACUGGUUGA AAAUCCAACCCACCACCCCAUCAGAGCCAACAGCAAUUA AAAGUCAGUUAAAAAUUUGUAAGGAUGAAGUCAACCGG CUAUCAGGUCUUCAACCUCAAAUUGAACGAUUAAAAAU UCAAAGCAUAGCCCUGAAAGAGAAAGGACAAGGACCCA UGUUCCUGGAUGCAGACUUUGUGGCCUUUACAAAUCAU UUUAAGCAAGUCUUUUCUGAUGUGCAGGCCAGAGAGAA AGAGCUACAGACAAUUUUUGACACUUUGCCACCAAUGC GCUAUCAGGAGACCAUGAGUGCCAUCAGGACAUGGGUC CAGCAGUCAGAAACCAAACUCUCCAUACCUCAACUUAGU GUCACCGACUAUGAAAUCAUGGAGCAGAGACUCGGGGA AUUGCAGGCUUUACAAAGUUCUCUGCAAGAGCAACAAA GUGGCCUAUACUAUCUCAGCACCACUGUGAAAGAGAUG UCGAAGAAAGCGCCCUCUGAAAUUAGCCGGAAAUAUCA AUCAGAAUUUGAAGAAAUUGAGGGACGCUGGAAGAAGC UCUCCUCCCAGCUGGUUGAGCAUUGUCAAAAGCUAGAG GAGCAAAUGAAUAAACUCCGAAAAAUUCAGAAUCACAU ACAAACCCUGAAGAAAUGGAUGGCUGAAGUUGAUGUUU UUCUGAAGGAGGAAUGGCCUGCCCUUGGGGAUUCAGAA AUUCUAAAAAAGCAGCUGAAACAGUGCAGACUUUUAGU CAGUGAUAUUCAGACAAUUCAGCCCAGUCUAAACAGUG UCAAUGAAGGUGGGCAGAAGAUAAAGAAUGAAGCAGAG CCAGAGUUUGCUUCGAGACUUGAGACAGAACUCAAAGA ACUUAACACUCAGUGGGAUCACAUGUGCCAACAGGUCU AUGCCAGAAAGGAGGCCUUGAAGGGAGGUUUGGAGAAA ACUGUAAGCCUCCAGAAAGAUCUAUCAGAGAUGCACGA AUGGAUGACACAAGCUGAAGAAGAGUAUCUUGAGAGAG AUUUUGAAUAUAAAACUCCAGAUGAAUUACAGAAAGCA GUUGAAGAGAUGAAGAGAGCUAAAGAAGAGGCCCAACA AAAAGAAGCGAAAGUGAAACUCCUUACUGAGUCUGUAA AUAGUGUCAUAGCUCAAGCUCCACCUGUAGCACAAGAG GCCUUAAAAAAGGAACUUGAAACUCUAACCACCAACUAC CAGUGGCUCUGCACUAGGCUGAAUGGGAAAUGCAAGAC UUUGGAAGAAGUUUGGGCAUGUUGGCAUGAGUUAUUGU CAUACUUGGAGAAAGCAAACAAGUGGCUAAAUGAAGUA GAAUUUAAACUUAAAACCACUGAAAACAUUCCUGGCGG AGCUGAGGAAAUCUCUGAGGUGCUAGAUUCACUUGAAA AUUUGAUGCGACAUUCAGAGGAUAACCCAAAUCAGAUU CGCAUAUUGGCACAGACCCUAACAGAUGGCGGAGUCAU GGAUGAGCUAAUCAAUGAGGAACUUGAGACAUUUAAUU CUCGUUGGAGGGAACUACAUGAAGAGGCUGUAAGGAGG CAAAAGUUGCUUGAACAGAGCAUCCAGUCUGCCCAGGA GACUGAAAAAUCCUUACACUUAAUCCAGGAGUCCCUCAC AUUCAUUGACAAGCAGUUGGCAGCUUAUAUUGCAGACA AGGUGGACGCAGCUCAAAUGCCUCAGGAAGCCCAGAAAAUCCAAUCUGAUUUGACAAGUCAUGAGAUCAGUUUAGAWSGR Ref. No. 54841-719.601AGAAAUGAAGAAACAUAAUCAGGGGAAGGAGGCUGCCC AAAGAGUCCUGUCUCAGAUUGAUGUUGCACAGAAAAAA UUACAAGAUGUCUCCAUGAAGUUUCGAUUAUUCCAGAA ACCAGCCAAUUUUGAGCAGCGUCUACAAGAAAGUAAGA UGAUUUUAGAUGAAGUGAAGAUGCACUUGCCUGCAUUG GAAACAAAGAGUGUGGAACAGGAAGUAGUACAGUCACA GCUAAAUCAUUGUGUGAACUUGUAUAAAAGUCUGAGUG AAGUGAAGUCUGAAGUGGAAAUGGUGAUAAAGACUGGA CGUCAGAUUGUACAGAAAAAGCAGACGGAAAAUCCCAA AGAACUUGAUGAAAGAGUAACAGCUUUGAAAUUGCAUU AUAAUGAGCUGGGAGCAAAGGUAACAGAAAGAAAGCAA CAGUUGGAGAAAUGCUUGAAAUUGUCCCGUAAGAUGCG AAAGGAAAUGAAUGUCUUGACAGAAUGGCUGGCAGCUA CAGAUAUGGAAUUGACAAAGAGAUCAGCAGUUGAAGGA AUGCCUAGUAAUUUGGAUUCUGAAGUUGCCUGGGGAAA GGCUACUCAAAAAGAGAUUGAGAAACAGAAGGUGCACC UGAAGAGUAUCACAGAGGUAGGAGAGGCCUUGAAAACA GUUUUGGGCAAGAAGGAGACGUUGGUGGAAGAUAAACU CAGUCUUCUGAAUAGUAACUGGAUAGCUGUCACCUCCCG AGCAGAAGAGUGGUUAAAUCUUUUGUUGGAAUACCAGA AACACAUGGAAACUUUUGACCAGAAUGUGGACCACAUC ACAAAGUGGAUCAUUCAGGCUGACACACUUUUGGAUGA AUCAGAGAAAAAGAAACCCCAGCAAAAAGAAGACGUGC UUAAGCGUUUAAAGGCAGAACUGAAUGACAUACGCCCA AAGGUGGACUCUACACGUGACCAAGCAGCAAACUUGAU GGCAAACCGCGGUGACCACUGCAGGAAAUUAGUAGAGC CCCAAAUCUCAGAGCUCAACCAUCGAUUUGCAGCCAUUU CACACAGAAUUAAGACUGGAAAGGCCUCCAUUCCUUUG AAGGAAUUGGAGCAGUUUAACUCAGAUAUACAAAAAUU GCUUGAACCACUGGAGGCUGAAAUUCAGCAGGGGGUGA AUCUGAAAGAGGAAGACUUCAAUAAAGAUAUGAAUGAA GACAAUGAGGGUACUGUAAAAGAAUUGUUGCAAAGAGG AGACAACUUACAACAAAGAAUCACAGAUGAGAGAAAGC GAGAGGAAAUAAAGAUAAAACAGCAGCUGUUACAGACA AAACAUAAUGCUCUCAAGGAUUUGAGGUCUCAAAGAAG AAAAAAGGCUCUAGAAAUUUCUCAUCAGUGGUAUCAGU ACAAGAGGCAGGCUGAUGAUCUCCUGAAAUGCUUGGAU GACAUUGAAAAAAAAUUAGCCAGCCUACCUGAGCCCAG AGAUGAAAGGAAAAUAAAGGAAAUUGAUCGGGAAUUGC AGAAGAAGAAAGAGGAGCUGAAUGCAGUGCGUAGGCAA GCUGAGGGCUUGUCUGAGGAUGGGGCCGCAAUGGCAGU GGAGCCAACUCAGAUCCAGCUCAGCAAGCGCUGGCGGGA AAUUGAGAGCAAAUUUGCUCAGUUUCGAAGACUCAACU UUGCACAAAUUCACACUGUCCGUGAAGAAACGAUGAUG GUGAUGACUGAAGACAUGCCUUUGGAAAUUUCUUAUGU GCCUUCUACUUAUUUGACUGAAAUCACUCAUGUCUCACA AGCCCUAUUAGAAGUGGAACAACUUCUCAAUGCUCCUG ACCUCUGUGCUAAGGACUUUGAAGAUCUCUUUAAGCAA GAGGAGUCUCUGAAGAAUAUAAAAGAUAGUCUACAACAAAGCUCAGGUCGGAUUGACAUUAUUCAUAGCAAGAAGAWSGR Ref. No. 54841-719.601CAGCAGCAUUGCAAAGUGCAACGCCUGUGGAAAGGGUG AAGCUACAGGAAGCUCUCUCCCAGCUUGAUUUCCAAUGG GAAAAAGUUAACAAAAUGUACAAGGACCGACAAGGGCG AUUUGACAGAUCUGUUGAGAAAUGGCGGCGUUUUCAUU AUGAUAUAAAGAUAUUUAAUCAGUGGCUAACAGAAGCU GAACAGUUUCUCAGAAAGACACAAAUUCCUGAGAAUUG GGAACAUGCUAAAUACAAAUGGUAUCUUAAGGAACUCC AGGAUGGCAUUGGGCAGCGGCAAACUGUUGUCAGAACA UUGAAUGCAACUGGGGAAGAAAUAAUUCAGCAAUCCUC AAAAACAGAUGCCAGUAUUCUACAGGAAAAAUUGGGAA GCCUGAAUCUGCGGUGGCAGGAGGUCUGCAAACAGCUG UCAGACAGAAAAAAGAGGCUAGAAGAACAAAAGAAUAU CUUGUCAGAAUUUCAAAGAGAUUUAAAUGAAUUUGUUU UAUGGUUGGAGGAAGCAGAUAACAUUGCUAGUAUCCCA CUUGAACCUGGAAAAGAGCAGCAACUAAAAGAAAAGCU UGAGCAAGUCAAGUUACUGGUGGAAGAGUUGCCCCUGC GCCAGGGAAUUCUCAAACAAUUAAAUGAAACUGGAGGA CCCGUGCUUGUAAGUGCUCCCAUAAGCCCAGAAGAGCAA GAUAAACUUGAAAAUAAGCUCAAGCAGACAAAUCUCCA GUGGAUAAAGGUUUCCAGAGCUUUACCUGAGAAACAAG GAGAAAUUGAAGCUCAAAUAAAAGACCUUGGGCAGCUU GAAAAAAAGCUUGAAGACCUUGAAGAGCAGUUAAAUCA UCUGCUGCUGUGGUUAUCUCCUAUUAGGAAUCAGUUGG AAAUUUAUAACCAACCAAACCAAGAAGGACCAUUUGAC GUUCAGGAAACUGAAAUAGCAGUUCAAGCUAAACAACC GGAUGUGGAAGAGAUUUUGUCUAAAGGGCAGCAUUUGU ACAAGGAAAAACCAGCCACUCAGCCAGUGAAGAGGAAG UUAGAAGAUCUGAGCUCUGAGUGGAAGGCGGUAAACCG UUUACUUCAAGAGCUGAGGGCAAAGCAGCCUGACCUAG CUCCUGGACUGACCACUAUUGGAGCCUCUCCUACUCAGA CUGUUACUCUGGUGACACAACCUGUGGUUACUAAGGAA ACUGCCAUCUCCAAACUAGAAAUGCCAUCUUCCUUGAUG UUGGAGGUACCUGCUCUGGCAGAUUUCAACCGGGCUUG GACAGAACUUACCGACUGGCUUUCUCUGCUUGAUCAAG UUAUAAAAUCACAGAGGGUGAUGGUGGGUGACCUUGAG GAUAUCAACGAGAUGAUCAUCAAGCAGAAGGCAACAAU GCAGGAUUUGGAACAGAGGCGUCCCCAGUUGGAAGAAC UCAUUACCGCUGCCCAAAAUUUGAAAAACAAGACCAGCA AUCAAGAGGCUAGAACAAUCAUUACGGAUCGAAUUGAA AGAAUUCAGAAUCAGUGGGAUGAAGUACAAGAACACCU UCAGAACCGGAGGCAACAGUUGAAUGAAAUGUUAAAGG AUUCAACACAAUGGCUGGAAGCUAAGGAAGAAGCUGAG CAGGUCUUAGGACAGGCCAGAGCCAAGCUUGAGUCAUG GAAGGAGGGUCCCUAUACAGUAGAUGCAAUCCAAAAGA AAAUCACAGAAACCAAGCAGUUGGCCAAAGACCUCCGCC AGUGGCAGACAAAUGUAGAUGUGGCAAAUGACUUGGCC CUGAAACUUCUCCGGGAUUAUUCUGCAGAUGAUACCAG AAAAGUCCACAUGAUAACAGAGAAUAUCAAUGCCUCUU GGAGAAGCAUUCAUAAAAGGGUGAGUGAGCGAGAGGCUGCUUUGGAAGAAACUCAUAGAUUACUGCAACAGUUCCCWSGR Ref. No. 54841-719.601CCUGGACCUGGAAAAGUUUCUUGCCUGGCUUACAGAAG CUGAAACAACUGCCAAUGUCCUACAGGAUGCUACCCGUA AGGAAAGGCUCCUAGAAGACUCCAAGGGAGUAAAAGAG CUGAUGAAACAAUGGCAAGACCUCCAAGGUGAAAUUGA AGCUCACACAGAUGUUUAUCACAACCUGGAUGAAAACA GCCAAAAAAUCCUGAGAUCCCUGGAAGGUUCCGAUGAU GCAGUCCUGUUACAAAGACGUUUGGAUAACAUGAACUU CAAGUGGAGUGAACUUCGGAAAAAGUCUCUCAACAUUA GGUCCCAUUUGGAAGCCAGUUCUGACCAGUGGAAGCGU CUGCACCUUUCUCUGCAGGAACUUCUGGUGUGGCUACAG CUGAAAGAUGAUGAAUUAAGCCGGCAGGCACCUAUUGG AGGCGACUUUCCAGCAGUUCAGAAGCAGAACGAUGUAC AUAGGGCCUUCAAGAGGGAAUUGAAAACUAAAGAACCU GUAAUCAUGAGUACUCUUGAGACUGUACGAAUAUUUCU GACAGAGCAGCCUUUGGAAGGACUAGAGAAACUCUACC AGGAGCCCAGAGAGCUGCCUCCUGAGGAGAGAGCCCAGA AUGUCACUCGGCUUCUACGAAAGCAGGCUGAGGAGGUC AAUACUGAGUGGGAAAAAUUGAACCUGCACUCCGCUGA CUGGCAGAGAAAAAUAGAUGAGACCCUUGAAAGACUCC AGGAACUUCAAGAGGCCACGGAUGAGCUGGACCUCAAG CUGCGCCAAGCUGAGGUGAUCAAGGGAUCCUGGCAGCCC GUGGGCGAUCUCCUCAUUGACUCUCUCCAAGAUCACCUC GAGAAAGUCAAGGCACUUCGAGGAGAAAUUGCGCCUCU GAAAGAGAACGUGAGCCACGUCAAUGACCUUGCUCGCCA GCUUACCACUUUGGGCAUUCAGCUCUCACCGUAUAACCU CAGCACUCUGGAAGACCUGAACACCAGAUGGAAGCUUCU GCAGGUGGCCGUCGAGGACCGAGUCAGGCAGCUGCAUG AAGCCCACAGGGACUUUGGUCCAGCAUCUCAGCACUUUC UUUCCACGUCUGUCCAGGGUCCCUGGGAGAGAGCCAUCU CGCCAAACAAAGUGCCCUACUAUAUCAACCACGAGACUC AAACAACUUGCUGGGACCAUCCCAAAAUGACAGAGCUCU ACCAGUCUUUAGCUGACCUGAAUAAUGUCAGAUUCUCA GCUUAUAGGACUGCCAUGAAACUCCGAAGACUGCAGAA GGCCCUUUGCUUGGAUCUCUUGAGCCUGUCAGCUGCAUG UGAUGCCUUGGACCAGCACAACCUCAAGCAAAAUGACCA GCCCAUGGAUAUCCUGCAGAUUAUUAAUUGUUUGACCA CUAUUUAUGACCGCCUGGAGCAAGAGCACAACAAUUUG GUCAACGUCCCUCUCUGCGUGGAUAUGUGUCUGAACUG GCUGCUGAAUGUUUAUGAUACGGGACGAACAGGGAGGA UCCGUGUCCUGUCUUUUAAAACUGGCAUCAUUUCCCUGU GUAAAGCACAUUUGGAAGACAAGUACAGAUACCUUUUC AAGCAAGUGGCAAGUUCAACAGGAUUUUGUGACCAGCG CAGGCUGGGCCUCCUUCUGCAUGAUUCUAUCCAAAUUCC AAGACAGUUGGGUGAAGUUGCAUCCUUUGGGGGCAGUA ACAUUGAGCCAAGUGUCCGGAGCUGCUUCCAAUUUGCU AAUAAUAAGCCAGAGAUCGAAGCGGCCCUCUUCCUAGAC UGGAUGAGACUGGAACCCCAGUCCAUGGUGUGGCUGCCC GUCCUGCACAGAGUGGCUGCUGCAGAAACUGCCAAGCAU CAGGCCAAAUGUAACAUCUGCAAAGAGUGUCCAAUCAUUGGAUUCAGGUACAGGAGUCUAAAGCACUUUAAUUAUGWSGR Ref. No. 54841-719.601ACAUCUGCCAAAGCUGCUUUUUUUCUGGUCGAGUUGCA AAAGGCCAUAAAAUGCACUAUCCCAUGGUGGAAUAUUG CACUCCGACUACAUCAGGAGAAGAUGUUCGAGACUUUG CCAAGGUACUAAAAAACAAAUUUCGAACCAAAAGGUAU UUUGCGAAGCAUCCCCGAAUGGGCUACCUGCCAGUGCAG ACUGUCUUAGAGGGGGACAACAUGGAAACUCCCGUUAC UCUGAUCAACUUCUGGCCAGUAGAUUCUGCGCCUGCCUC GUCCCCUCAGCUUUCACACGAUGAUACUCAUUCACGCAU UGAACAUUAUGCUAGCAGGCUAGCAGAAAUGGAAAACA GCAAUGGAUCUUAUCUAAAUGAUAGCAUCUCUCCUAAU GAGAGCAUAGAUGAUGAACAUUUGUUAAUCCAGCAUUA CUGCCAAAGUUUGAACCAGGACUCCCCCCUGAGCCAGCC UCGUAGUCCUGCCCAGAUCUUGAUUUCCUUAGAGAGUG AGGAAAGAGGGGAGCUAGAGAGAAUCCUAGCAGAUCUU GAGGAAGAAAACAGGAAUCUGCAAGCAGAAUAUGACCG UCUAAAGCAGCAGCACGAACAUAAAGGCCUGUCCCCACU GCCGUCCCCUCCUGAAAUGAUGCCCACCUCUCCCCAGAG UCCCCGGGAUGCUGAGCUCAUUGCUGAGGCCAAGCUACU GCGUCAACACAAAGGCCGCCUGGAAGCCAGGAUGCAAAU CCUGGAAGACCACAAUAAACAGCUGGAGUCACAGUUAC ACAGGCUAAGGCAGCUGCUGGAGCAACCCCAGGCAGAGG CCAAAGUGAAUGGCACAACGGUGUCCUCUCCUUCUACCU CUCUACAGAGGUCCGACAGCAGUCAGCCUAUGCUGCUCC GAGUGGUUGGCAGUCAAACUUCGGACUCCAUGGGUGAG GAAGAUCUUCUCAGUCCUCCCCAGGACACAAGCACAGGG UUAGAGGAGGUGAUGGAGCAACUCAACAACUCCUUCCC UAGUUCAAGAGGAAGAAAUACCCCUGGAAAGCCAAUGA GAGAGGACACAAUG22 Homo aucaguuacuguguugacucacucaguguugggaucacucacuuucccccuacaggac sapiens ucagaucugggaggcaauuaccuucggagaaaaacgaauaggaaaaacugaaguguua dystrophin cuuuuuuuaaagcugcugaaguuuguugguuucucauuguuuuuaagccuacuggag (DMD), caauaaaguuugaagaacuuuuaccagguuuuuuuuaucgcugccuugauauacacuu transcript uucaaaAUGCUUUGGUGGGAAGAAGUAGAGGACUGUUAUG variant AAAGAGAAGAUGUUCAAAAGAAAACAUUCACAAAAUGG Dp427m, GUAAAUGCACAAUUUUCUAAGUUUGGGAAGCAGCAUAU mRNA UGAGAACCUCUUCAGUGACCUACAGGAUGGGAGGCGCC UCCUAGACCUCCUCGAAGGCCUGACAGGGCAAAAACUGC(Ref. Seq. CAAAAGAAAAAGGAUCCACAAGAGUUCAUGCCCUGAAC NM 00400 AAUGUCAACAAGGCACUGCGGGUUUUGCAGAACAAUAA 6.3); UGUUGAUUUAGUGAAUAUUGGAAGUACUGACAUCGUAG AUGGAAAUCAUAAACUGACUCUUGGUUUGAUUUGGAAU(CODING AUAAUCCUCCACUGGCAGGUCAAAAAUGUAAUGAAAAA SEQUENC UAUCAUGGCUGGAUUGCAACAAACCAACAGUGAAAAGAE) UUCUCCUGAGCUGGGUCCGACAAUCAACUCGUAAUUAUC CACAGGUUAAUGUAAUCAACUUCACCACCAGCUGGUCUG AUGGCCUGGCUUUGAAUGCUCUCAUCCAUAGUCAUAGG CCAGACCUAUUUGACUGGAAUAGUGUGGUUUGCCAGCA GUCAGCCACACAACGACUGGAACAUGCAUUCAACAUCGC CAGAUAUCAAUUAGGCAUAGAGAAACUACUCGAUCCUGAAGAUGUUGAUACCACCUAUCCAGAUAAGAAGUCCAUCWSGR Ref. No. 54841-719.601UUAAUGUACAUCACAUCACUCUUCCAAGUUUUGCCUCAA CAAGUGAGCAUUGAAGCCAUCCAGGAAGUGGAAAUGUU GCCAAGGCCACCUAAAGUGACUAAAGAAGAACAUUUUC AGUUACAUCAUCAAAUGCACUAUUCUCAACAGAUCACG GUCAGUCUAGCACAGGGAUAUGAGAGAACUUCUUCCCC UAAGCCUCGAUUCAAGAGCUAUGCCUACACACAGGCUGC UUAUGUCACCACCUCUGACCCUACACGGAGCCCAUUUCC UUCACAGCAUUUGGAAGCUCCUGAAGACAAGUCAUUUG GCAGUUCAUUGAUGGAGAGUGAAGUAAACCUGGACCGU UAUCAAACAGCUUUAGAAGAAGUAUUAUCGUGGCUUCU UUCUGCUGAGGACACAUUGCAAGCACAAGGAGAGAUUU CUAAUGAUGUGGAAGUGGUGAAAGACCAGUUUCAUACU CAUGAGGGGUACAUGAUGGAUUUGACAGCCCAUCAGGG CCGGGUUGGUAAUAUUCUACAAUUGGGAAGUAAGCUGA UUGGAACAGGAAAAUUAUCAGAAGAUGAAGAAACUGAA GUACAAGAGCAGAUGAAUCUCCUAAAUUCAAGAUGGGA AUGCCUCAGGGUAGCUAGCAUGGAAAAACAAAGCAAUU UACAUAGAGUUUUAAUGGAUCUCCAGAAUCAGAAACUG AAAGAGUUGAAUGACUGGCUAACAAAAACAGAAGAAAG AACAAGGAAAAUGGAGGAAGAGCCUCUUGGACCUGAUC UUGAAGACCUAAAACGCCAAGUACAACAACAUAAGGUG CUUCAAGAAGAUCUAGAACAAGAACAAGUCAGGGUCAA UUCUCUCACUCACAUGGUGGUGGUAGUUGAUGAAUCUA GUGGAGAUCACGCAACUGCUGCUUUGGAAGAACAACUU AAGGUAUUGGGAGAUCGAUGGGCAAACAUCUGUAGAUG GACAGAAGACCGCUGGGUUCUUUUACAAGACAUCCUUC UCAAAUGGCAACGUCUUACUGAAGAACAGUGCCUUUUU AGUGCAUGGCUUUCAGAAAAAGAAGAUGCAGUGAACAA GAUUCACACAACUGGCUUUAAAGAUCAAAAUGAAAUGU UAUCAAGUCUUCAAAAACUGGCCGUUUUAAAAGCGGAU CUAGAAAAGAAAAAGCAAUCCAUGGGCAAACUGUAUUC ACUCAAACAAGAUCUUCUUUCAACACUGAAGAAUAAGU CAGUGACCCAGAAGACGGAAGCAUGGCUGGAUAACUUU GCCCGGUGUUGGGAUAAUUUAGUCCAAAAACUUGAAAA GAGUACAGCACAGAUUUCACAGGCUGUCACCACCACUCA GCCAUCACUAACACAGACAACUGUAAUGGAAACAGUAA CUACGGUGACCACAAGGGAACAGAUCCUGGUAAAGCAU GCUCAAGAGGAACUUCCACCACCACCUCCCCAAAAGAAG AGGCAGAUUACUGUGGAUUCUGAAAUUAGGAAAAGGUU GGAUGUUGAUAUAACUGAACUUCACAGCUGGAUUACUC GCUCAGAAGCUGUGUUGCAGAGUCCUGAAUUUGCAAUC UUUCGGAAGGAAGGCAACUUCUCAGACUUAAAAGAAAA AGUCAAUGCCAUAGAGCGAGAAAAAGCUGAGAAGUUCA GAAAACUGCAAGAUGCCAGCAGAUCAGCUCAGGCCCUGG UGGAACAGAUGGUGAAUGAGGGUGUUAAUGCAGAUAGC AUCAAACAAGCCUCAGAACAACUGAACAGCCGGUGGAUC GAAUUCUGCCAGUUGCUAAGUGAGAGACUUAACUGGCU GGAGUAUCAGAACAACAUCAUCGCUUUCUAUAAUCAGC UACAACAAUUGGAGCAGAUGACAACUACUGCUGAAAACUGGUUGAAAAUCCAACCCACCACCCCAUCAGAGCCAACAWSGR Ref. No. 54841-719.601GCAAUUAAAAGUCAGUUAAAAAUUUGUAAGGAUGAAGU CAACCGGCUAUCAGAUCUUCAACCUCAAAUUGAACGAUU AAAAAUUCAAAGCAUAGCCCUGAAAGAGAAAGGACAAG GACCCAUGUUCCUGGAUGCAGACUUUGUGGCCUUUACA AAUCAUUUUAAGCAAGUCUUUUCUGAUGUGCAGGCCAG AGAGAAAGAGCUACAGACAAUUUUUGACACUUUGCCAC CAAUGCGCUAUCAGGAGACCAUGAGUGCCAUCAGGACA UGGGUCCAGCAGUCAGAAACCAAACUCUCCAUACCUCAA CUUAGUGUCACCGACUAUGAAAUCAUGGAGCAGAGACU CGGGGAAUUGCAGGCUUUACAAAGUUCUCUGCAAGAGC AACAAAGUGGCCUAUACUAUCUCAGCACCACUGUGAAA GAGAUGUCGAAGAAAGCGCCCUCUGAAAUUAGCCGGAA AUAUCAAUCAGAAUUUGAAGAAAUUGAGGGACGCUGGA AGAAGCUCUCCUCCCAGCUGGUUGAGCAUUGUCAAAAGC UAGAGGAGCAAAUGAAUAAACUCCGAAAAAUUCAGAAU CACAUACAAACCCUGAAGAAAUGGAUGGCUGAAGUUGA UGUUUUUCUGAAGGAGGAAUGGCCUGCCCUUGGGGAUU CAGAAAUUCUAAAAAAGCAGCUGAAACAGUGCAGACUU UUAGUCAGUGAUAUUCAGACAAUUCAGCCCAGUCUAAA CAGUGUCAAUGAAGGUGGGCAGAAGAUAAAGAAUGAAG CAGAGCCAGAGUUUGCUUCGAGACUUGAGACAGAACUC AAAGAACUUAACACUCAGUGGGAUCACAUGUGCCAACA GGUCUAUGCCAGAAAGGAGGCCUUGAAGGGAGGUUUGG AGAAAACUGUAAGCCUCCAGAAAGAUCUAUCAGAGAUG CACGAAUGGAUGACACAAGCUGAAGAAGAGUAUCUUGA GAGAGAUUUUGAAUAUAAAACUCCAGAUGAAUUACAGA AAGCAGUUGAAGAGAUGAAGAGAGCUAAAGAAGAGGCC CAACAAAAAGAAGCGAAAGUGAAACUCCUUACUGAGUC UGUAAAUAGUGUCAUAGCUCAAGCUCCACCUGUAGCAC AAGAGGCCUUAAAAAAGGAACUUGAAACUCUAACCACC AACUACCAGUGGCUCUGCACUAGGCUGAAUGGGAAAUG CAAGACUUUGGAAGAAGUUUGGGCAUGUUGGCAUGAGU UAUUGUCAUACUUGGAGAAAGCAAACAAGUGGCUAAAU GAAGUAGAAUUUAAACUUAAAACCACUGAAAACAUUCC UGGCGGAGCUGAGGAAAUCUCUGAGGUGCUAGAUUCAC UUGAAAAUUUGAUGCGACAUUCAGAGGAUAACCCAAAU CAGAUUCGCAUAUUGGCACAGACCCUAACAGAUGGCGG AGUCAUGGAUGAGCUAAUCAAUGAGGAACUUGAGACAU UUAAUUCUCGUUGGAGGGAACUACAUGAAGAGGCUGUA AGGAGGCAAAAGUUGCUUGAACAGAGCAUCCAGUCUGC CCAGGAGACUGAAAAAUCCUUACACUUAAUCCAGGAGU CCCUCACAUUCAUUGACAAGCAGUUGGCAGCUUAUAUU GCAGACAAGGUGGACGCAGCUCAAAUGCCUCAGGAAGCC CAGAAAAUCCAAUCUGAUUUGACAAGUCAUGAGAUCAG UUUAGAAGAAAUGAAGAAACAUAAUCAGGGGAAGGAGG CUGCCCAAAGAGUCCUGUCUCAGAUUGAUGUUGCACAG AAAAAAUUACAAGAUGUCUCCAUGAAGUUUCGAUUAUU CCAGAAACCAGCCAAUUUUGAGCAGCGUCUACAAGAAA GUAAGAUGAUUUUAGAUGAAGUGAAGAUGCACUUGCCUGCAUUGGAAACAAAGAGUGUGGAACAGGAAGUAGUACAWSGR Ref. No. 54841-719.601GUCACAGCUAAAUCAUUGUGUGAACUUGUAUAAAAGUC UGAGUGAAGUGAAGUCUGAAGUGGAAAUGGUGAUAAAG ACUGGACGUCAGAUUGUACAGAAAAAGCAGACGGAAAA UCCCAAAGAACUUGAUGAAAGAGUAACAGCUUUGAAAU UGCAUUAUAAUGAGCUGGGAGCAAAGGUAACAGAAAGA AAGCAACAGUUGGAGAAAUGCUUGAAAUUGUCCCGUAA GAUGCGAAAGGAAAUGAAUGUCUUGACAGAAUGGCUGG CAGCUACAGAUAUGGAAUUGACAAAGAGAUCAGCAGUU GAAGGAAUGCCUAGUAAUUUGGAUUCUGAAGUUGCCUG GGGAAAGGCUACUCAAAAAGAGAUUGAGAAACAGAAGG UGCACCUGAAGAGUAUCACAGAGGUAGGAGAGGCCUUG AAAACAGUUUUGGGCAAGAAGGAGACGUUGGUGGAAGA UAAACUCAGUCUUCUGAAUAGUAACUGGAUAGCUGUCA CCUCCCGAGCAGAAGAGUGGUUAAAUCUUUUGUUGGAA UACCAGAAACACAUGGAAACUUUUGACCAGAAUGUGGA CCACAUCACAAAGUGGAUCAUUCAGGCUGACACACUUUU GGAUGAAUCAGAGAAAAAGAAACCCCAGCAAAAAGAAG ACGUGCUUAAGCGUUUAAAGGCAGAACUGAAUGACAUA CGCCCAAAGGUGGACUCUACACGUGACCAAGCAGCAAAC UUGAUGGCAAACCGCGGUGACCACUGCAGGAAAUUAGU AGAGCCCCAAAUCUCAGAGCUCAACCAUCGAUUUGCAGC CAUUUCACACAGAAUUAAGACUGGAAAGGCCUCCAUUCC UUUGAAGGAAUUGGAGCAGUUUAACUCAGAUAUACAAA AAUUGCUUGAACCACUGGAGGCUGAAAUUCAGCAGGGG GUGAAUCUGAAAGAGGAAGACUUCAAUAAAGAUAUGAA UGAAGACAAUGAGGGUACUGUAAAAGAAUUGUUGCAAA GAGGAGACAACUUACAACAAAGAAUCACAGAUGAGAGA AAGCGAGAGGAAAUAAAGAUAAAACAGCAGCUGUUACA GACAAAACAUAAUGCUCUCAAGGAUUUGAGGUCUCAAA GAAGAAAAAAGGCUCUAGAAAUUUCUCAUCAGUGGUAU CAGUACAAGAGGCAGGCUGAUGAUCUCCUGAAAUGCUU GGAUGACAUUGAAAAAAAAUUAGCCAGCCUACCUGAGC CCAGAGAUGAAAGGAAAAUAAAGGAAAUUGAUCGGGAA UUGCAGAAGAAGAAAGAGGAGCUGAAUGCAGUGCGUAG GCAAGCUGAGGGCUUGUCUGAGGAUGGGGCCGCAAUGG CAGUGGAGCCAACUCAGAUCCAGCUCAGCAAGCGCUGGC GGGAAAUUGAGAGCAAAUUUGCUCAGUUUCGAAGACUC AACUUUGCACAAAUUCACACUGUCCGUGAAGAAACGAU GAUGGUGAUGACUGAAGACAUGCCUUUGGAAAUUUCUU AUGUGCCUUCUACUUAUUUGACUGAAAUCACUCAUGUC UCACAAGCCCUAUUAGAAGUGGAACAACUUCUCAAUGC UCCUGACCUCUGUGCUAAGGACUUUGAAGAUCUCUUUA AGCAAGAGGAGUCUCUGAAGAAUAUAAAAGAUAGUCUA CAACAAAGCUCAGGUCGGAUUGACAUUAUUCAUAGCAA GAAGACAGCAGCAUUGCAAAGUGCAACGCCUGUGGAAA GGGUGAAGCUACAGGAAGCUCUCUCCCAGCUUGAUUUCC AAUGGGAAAAAGUUAACAAAAUGUACAAGGACCGACAA GGGCGAUUUGACAGAUCUGUUGAGAAAUGGCGGCGUUU UCAUUAUGAUAUAAAGAUAUUUAAUCAGUGGCUAACAGAAGCUGAACAGUUUCUCAGAAAGACACAAAUUCCUGAGWSGR Ref. No. 54841-719.601AAUUGGGAACAUGCUAAAUACAAAUGGUAUCUUAAGGA ACUCCAGGAUGGCAUUGGGCAGCGGCAAACUGUUGUCA GAACAUUGAAUGCAACUGGGGAAGAAAUAAUUCAGCAA UCCUCAAAAACAGAUGCCAGUAUUCUACAGGAAAAAUU GGGAAGCCUGAAUCUGCGGUGGCAGGAGGUCUGCAAAC AGCUGUCAGACAGAAAAAAGAGGCUAGAAGAACAAAAG AAUAUCUUGUCAGAAUUUCAAAGAGAUUUAAAUGAAUU UGUUUUAUGGUUGGAGGAAGCAGAUAACAUUGCUAGUA UCCCACUUGAACCUGGAAAAGAGCAGCAACUAAAAGAA AAGCUUGAGCAAGUCAAGUUACUGGUGGAAGAGUUGCC CCUGCGCCAGGGAAUUCUCAAACAAUUAAAUGAAACUG GAGGACCCGUGCUUGUAAGUGCUCCCAUAAGCCCAGAAG AGCAAGAUAAACUUGAAAAUAAGCUCAAGCAGACAAAU CUCCAGUGGAUAAAGGUUUCCAGAGCUUUACCUGAGAA ACAAGGAGAAAUUGAAGCUCAAAUAAAAGACCUUGGGC AGCUUGAAAAAAAGCUUGAAGACCUUGAAGAGCAGUUA AAUCAUCUGCUGCUGUGGUUAUCUCCUAUUAGGAAUCA GUUGGAAAUUUAUAACCAACCAAACCAAGAAGGACCAU UUGACGUUAAGGAAACUGAAAUAGCAGUUCAAGCUAAA CAACCGGAUGUGGAAGAGAUUUUGUCUAAAGGGCAGCA UUUGUACAAGGAAAAACCAGCCACUCAGCCAGUGAAGA GGAAGUUAGAAGAUCUGAGCUCUGAGUGGAAGGCGGUA AACCGUUUACUUCAAGAGCUGAGGGCAAAGCAGCCUGA CCUAGCUCCUGGACUGACCACUAUUGGAGCCUCUCCUAC UCAGACUGUUACUCUGGUGACACAACCUGUGGUUACUA AGGAAACUGCCAUCUCCAAACUAGAAAUGCCAUCUUCCU UGAUGUUGGAGGUACCUGCUCUGGCAGAUUUCAACCGG GCUUGGACAGAACUUACCGACUGGCUUUCUCUGCUUGA UCAAGUUAUAAAAUCACAGAGGGUGAUGGUGGGUGACC UUGAGGAUAUCAACGAGAUGAUCAUCAAGCAGAAGGCA ACAAUGCAGGAUUUGGAACAGAGGCGUCCCCAGUUGGA AGAACUCAUUACCGCUGCCCAAAAUUUGAAAAACAAGA CCAGCAAUCAAGAGGCUAGAACAAUCAUUACGGAUCGA AUUGAAAGAAUUCAGAAUCAGUGGGAUGAAGUACAAGA ACACCUUCAGAACCGGAGGCAACAGUUGAAUGAAAUGU UAAAGGAUUCAACACAAUGGCUGGAAGCUAAGGAAGAA GCUGAGCAGGUCUUAGGACAGGCCAGAGCCAAGCUUGA GUCAUGGAAGGAGGGUCCCUAUACAGUAGAUGCAAUCC AAAAGAAAAUCACAGAAACCAAGCAGUUGGCCAAAGAC CUCCGCCAGUGGCAGACAAAUGUAGAUGUGGCAAAUGA CUUGGCCCUGAAACUUCUCCGGGAUUAUUCUGCAGAUG AUACCAGAAAAGUCCACAUGAUAACAGAGAAUAUCAAU GCCUCUUGGAGAAGCAUUCAUAAAAGGGUGAGUGAGCG AGAGGCUGCUUUGGAAGAAACUCAUAGAUUACUGCAAC AGUUCCCCCUGGACCUGGAAAAGUUUCUUGCCUGGCUUA CAGAAGCUGAAACAACUGCCAAUGUCCUACAGGAUGCU ACCCGUAAGGAAAGGCUCCUAGAAGACUCCAAGGGAGU AAAAGAGCUGAUGAAACAAUGGCAAGACCUCCAAGGUG AAAUUGAAGCUCACACAGAUGUUUAUCACAACCUGGAUGAAAACAGCCAAAAAAUCCUGAGAUCCCUGGAAGGUUCWSGR Ref. No. 54841-719.601CGAUGAUGCAGUCCUGUUACAAAGACGUUUGGAUAACA UGAACUUCAAGUGGAGUGAACUUCGGAAAAAGUCUCUC AACAUUAGGUCCCAUUUGGAAGCCAGUUCUGACCAGUG GAAGCGUCUGCACCUUUCUCUGCAGGAACUUCUGGUGU GGCUACAGCUGAAAGAUGAUGAAUUAAGCCGGCAGGCA CCUAUUGGAGGCGACUUUCCAGCAGUUCAGAAGCAGAA CGAUGUACAUAGGGCCUUCAAGAGGGAAUUGAAAACUA AAGAACCUGUAAUCAUGAGUACUCUUGAGACUGUACGA AUAUUUCUGACAGAGCAGCCUUUGGAAGGACUAGAGAA ACUCUACCAGGAGCCCAGAGAGCUGCCUCCUGAGGAGAG AGCCCAGAAUGUCACUCGGCUUCUACGAAAGCAGGCUGA GGAGGUCAAUACUGAGUGGGAAAAAUUGAACCUGCACU CCGCUGACUGGCAGAGAAAAAUAGAUGAGACCCUUGAA AGACUCCGGGAACUUCAAGAGGCCACGGAUGAGCUGGA CCUCAAGCUGCGCCAAGCUGAGGUGAUCAAGGGAUCCUG GCAGCCCGUGGGCGAUCUCCUCAUUGACUCUCUCCAAGA UCACCUCGAGAAAGUCAAGGCACUUCGAGGAGAAAUUG CGCCUCUGAAAGAGAACGUGAGCCACGUCAAUGACCUUG CUCGCCAGCUUACCACUUUGGGCAUUCAGCUCUCACCGU AUAACCUCAGCACUCUGGAAGACCUGAACACCAGAUGGA AGCUUCUGCAGGUGGCCGUCGAGGACCGAGUCAGGCAGC UGCAUGAAGCCCACAGGGACUUUGGUCCAGCAUCUCAGC ACUUUCUUUCCACGUCUGUCCAGGGUCCCUGGGAGAGAG CCAUCUCGCCAAACAAAGUGCCCUACUAUAUCAACCACG AGACUCAAACAACUUGCUGGGACCAUCCCAAAAUGACAG AGCUCUACCAGUCUUUAGCUGACCUGAAUAAUGUCAGA UUCUCAGCUUAUAGGACUGCCAUGAAACUCCGAAGACU GCAGAAGGCCCUUUGCUUGGAUCUCUUGAGCCUGUCAGC UGCAUGUGAUGCCUUGGACCAGCACAACCUCAAGCAAAA UGACCAGCCCAUGGAUAUCCUGCAGAUUAUUAAUUGUU UGACCACUAUUUAUGACCGCCUGGAGCAAGAGCACAACA AUUUGGUCAACGUCCCUCUCUGCGUGGAUAUGUGUCUG AACUGGCUGCUGAAUGUUUAUGAUACGGGACGAACAGG GAGGAUCCGUGUCCUGUCUUUUAAAACUGGCAUCAUUU CCCUGUGUAAAGCACAUUUGGAAGACAAGUACAGAUAC CUUUUCAAGCAAGUGGCAAGUUCAACAGGAUUUUGUGA CCAGCGCAGGCUGGGCCUCCUUCUGCAUGAUUCUAUCCA AAUUCCAAGACAGUUGGGUGAAGUUGCAUCCUUUGGGG GCAGUAACAUUGAGCCAAGUGUCCGGAGCUGCUUCCAA UUUGCUAAUAAUAAGCCAGAGAUCGAAGCGGCCCUCUU CCUAGACUGGAUGAGACUGGAACCCCAGUCCAUGGUGU GGCUGCCCGUCCUGCACAGAGUGGCUGCUGCAGAAACUG CCAAGCAUCAGGCCAAAUGUAACAUCUGCAAAGAGUGU CCAAUCAUUGGAUUCAGGUACAGGAGUCUAAAGCACUU UAAUUAUGACAUCUGCCAAAGCUGCUUUUUUUCUGGUC GAGUUGCAAAAGGCCAUAAAAUGCACUAUCCCAUGGUG GAAUAUUGCACUCCGACUACAUCAGGAGAAGAUGUUCG AGACUUUGCCAAGGUACUAAAAAACAAAUUUCGAACCA AAAGGUAUUUUGCGAAGCAUCCCCGAAUGGGCUACCUGCCAGUGCAGACUGUCUUAGAGGGGGACAACAUGGAAACWSGR Ref. No. 54841-719.601UCCCGUUACUCUGAUCAACUUCUGGCCAGUAGAUUCUGC GCCUGCCUCGUCCCCUCAGCUUUCACACGAUGAUACUCA UUCACGCAUUGAACAUUAUGCUAGCAGGCUAGCAGAAA UGGAAAACAGCAAUGGAUCUUAUCUAAAUGAUAGCAUC UCUCCUAAUGAGAGCAUAGAUGAUGAACAUUUGUUAAU CCAGCAUUACUGCCAAAGUUUGAACCAGGACUCCCCCCU GAGCCAGCCUCGUAGUCCUGCCCAGAUCUUGAUUUCCUU AGAGAGUGAGGAAAGAGGGGAGCUAGAGAGAAUCCUAG CAGAUCUUGAGGAAGAAAACAGGAAUCUGCAAGCAGAA UAUGACCGUCUAAAGCAGCAGCACGAACAUAAAGGCCU GUCCCCACUGCCGUCCCCUCCUGAAAUGAUGCCCACCUC UCCCCAGAGUCCCCGGGAUGCUGAGCUCAUUGCUGAGGC CAAGCUACUGCGUCAACACAAAGGCCGCCUGGAAGCCAG GAUGCAAAUCCUGGAAGACCACAAUAAACAGCUGGAGU CACAGUUACACAGGCUAAGGCAGCUGCUGGAGCAACCCC AGGCAGAGGCCAAAGUGAAUGGCACAACGGUGUCCUCU CCUUCUACCUCUCUACAGAGGUCCGACAGCAGUCAGCCU AUGCUGCUCCGAGUGGUUGGCAGUCAAACUUCGGACUCC AUGGGUGAGGAAGAUCUUCUCAGUCCUCCCCAGGACACA AGCACAGGGUUAGAGGAGGUGAUGGAGCAACUCAACAA CUCCUUCCCUAGUUCAAGAGGAAGAAAUACCCCUGGAAA GCCAAUGAGAGAGGACACAAUGuaggaagucuuuuccacauggcaga ugauuugggcagagcgauggaguccuuaguaucagucaugacagaugaagaaggagca gaauaaauguuuuacaacuccugauucccgcaugguuuuuauaauauucauacaacaa agaggauuagacaguaagaguuuacaagaaauaaaucuauauuuuugugaaggguagu gguauuauacuguagauuucaguaguuucuaagucuguuauuguuuuguuaacaaug gcagguuuuacacgucuaugcaauuguacaaaaaaguuauaagaaaacuacauguaaaa ucuugauagcuaaauaacuugccauuucuuuauauggaacgcauuuuggguuguuua aaaauuuauaacaguuauaaagaaagauuguaaacuaaagugugcuuuauaaaaaaaag uuguuuauaaaaaccccuaaaaacaaaacaaacacacacacacacacauacacacacacac acaaaacuuugaggcagcgcauuguuuugcauccuuuuggcgugauauccauaugaaa uucauggcuuuuucuuuuuuugcauauuaaagauaagacuuccucuaccaccacacca aaugacuacuacacacugcucauuugagaacugucagcugaguggggcaggcuugagu uuucauuucauauaucuauaugucuauaaguauauaaauacuauaguuauauagauaa agagauacgaauuucuauagacugacuuuuuccauuuuuuaaauguucaugucacauc cuaauagaaagaaauuacuucuagucagucauccaggcuuaccugcuuggucuagaau ggauuuuucccggagccggaagccaggaggaaacuacaccacacuaaaacauugucuac agcuccagauguuucucauuuuaaacaacuuuccacugacaacgaaaguaaaguaaagu auuggauuuuuuuaaagggaacaugugaaugaauacacaggacuuauuauaucagagu gaguaaucgguugguugguugauugauugauugauugauacauucagcuuccugcug cuagcaaugccacgauuuagauuuaaugaugcuucaguggaaaucaaucagaagguau ucugaccuugugaacaucagaagguauuuuuuaacucccaagcaguagcaggacgaug auagggcuggagggcuauggauucccagcccaucccugugaaggaguaggccacucuu uaagugaaggauuggaugauuguucauaauacauaaaguucucuguaauuacaacuaa auuauuaugcccucuucucacagucaaaaggaacugggugguuugguuuuuguugcu uuuuuagauuuauugucccaugugggaugaguuuuuaaaugccacaagacauaauuu aaaauaaauaaacuuugggaaaagguguaaaacaguagccccaucacauuugugauacu gacagguaucaacccagaagcccaugaacuguguuuccauccuuugcauuucucugcg aguaguuccacacagguuuguaaguaaguaagaaagaaggcaaauugauucaaauguuacaaaaaaacccuucuugguggauuagacagguuaaauauauaaacaaacaaacaaaaaWSGR Ref. No. 54841-719.601uugcucaaaaaagaggagaaaagcucaagaggaaaagcuaaggacugguaggaaaaagc uuuacucuuucaugccauuuuauuucuuuuugauuuuuaaaucauucauucaauaga uaccaccgugugaccuauaauuuugcaaaucuguuaccucugacaucaaguguaauua gcuuuuggagagugggcugacaucaaguguaauuagcuuuuggagaguggguuuug uccauuauuaauaauuaauuaauuaacaucaaacacggcuucucaugcuauuucuacc ucacuuugguuuugggguguuccugauaauugugcacaccugaguucacagcuucac cacuuguccauugcguuauuuucuuuuuccuuuauaauucuuucuuuuuccuucaua auuuucaaaagaaaacccaaagcucuaagguaacaaauuaccaaauuacaugaagauuu gguuuuugucuugcauuuuuuuccuuuaugugacgcuggaccuuuucuuuacccaag gauuuuuaaaacucagauuuaaaacaagggguuacuuuacauccuacuaagaaguuua aguaaguaaguuucauucuaaaaucagagguaaauagagugcauaaauaauuuuguuu uaaucuuuuuguuuuucuuuuagacacauuagcucuggagugagucugucauaauau uugaacaaaaauugagagcuuuauugcugcauuuuaagcauaauuaauuuggacauua uuucguguuguguucuuuauaaccaccaaguauuaaacuguaaaucauaauguaacug aagcauaaacaucacauggcauguuuugucauuguuuucagguacugaguucuuacuu gaguaucauaauauauuguguuuuaacaccaacacuguaacauuuacgaauuauuuuu uuaaacuucaguuuuacugcauuuucacaacauaucagacuucaccaaauauaugccu uacuauuguauuauaguacugcuuuacuguguaucucaauaaagcacgcaguuauguu acaaaaaa23 Human Cguuaaaugcaaacgcugcucuggcucauguguuugcuccgagguauagguuuuguu dystrophin cgacugacguaucagauagucagagugguuaccacaccgacguuguagcagcugcaua (DMD), auaaaugacugaaagaaucauguuaggcaugcccaccuaaccuaacuugaaucaugcgq transcript aaggggagcuguuggaauucaaauagacuuucugguucccagcagucggcaguaauag variant aaugcuuucaggaagaugacagaaucaggagaaagaugcuguuuugcacuaucuugau Dp427c, uuguuacagcagccaacuuauuggcaugauggagugacaggaaaaacagcuggcAU mRNA GGAAGAUGAAAGAGAAGAUGUUCAAAAGAAAACAUUCA CAAAAUGGGUAAAUGCACAAUUUUCUAAGUUUGGGAAG(Ref. Seq. CAGCAUAUUGAGAACCUCUUCAGUGACCUACAGGAUGG NM 00010 GAGGCGCCUCCUAGACCUCCUCGAAGGCCUGACAGGGCA 9.3) AAAACUGCCAAAAGAAAAAGGAUCCACAAGAGUUCAUG CCCUGAACAAUGUCAACAAGGCACUGCGGGUUUUGCAG(CODING AACAAUAAUGUUGAUUUAGUGAAUAUUGGAAGUACUGA SEQUENC CAUCGUAGAUGGAAAUCAUAAACUGACUCUUGGUUUGAE) UUUGGAAUAUAAUCCUCCACUGGCAGGUCAAAAAUGUA AUGAAAAAUAUCAUGGCUGGAUUGCAACAAACCAACAG UGAAAAGAUUCUCCUGAGCUGGGUCCGACAAUCAACUC GUAAUUAUCCACAGGUUAAUGUAAUCAACUUCACCACC AGCUGGUCUGAUGGCCUGGCUUUGAAUGCUCUCAUCCA UAGUCAUAGGCCAGACCUAUUUGACUGGAAUAGUGUGG UUUGCCAGCAGUCAGCCACACAACGACUGGAACAUGCAU UCAACAUCGCCAGAUAUCAAUUAGGCAUAGAGAAACUA CUCGAUCCUGAAGAUGUUGAUACCACCUAUCCAGAUAA GAAGUCCAUCUUAAUGUACAUCACAUCACUCUUCCAAGU UUUGCCUCAACAAGUGAGCAUUGAAGCCAUCCAGGAAG UGGAAAUGUUGCCAAGGCCACCUAAAGUGACUAAAGAA GAACAUUUUCAGUUACAUCAUCAAAUGCACUAUUCUCA ACAGAUCACGGUCAGUCUAGCACAGGGAUAUGAGAGAA CUUCUUCCCCUAAGCCUCGAUUCAAGAGCUAUGCCUACA CACAGGCUGCUUAUGUCACCACCUCUGACCCUACACGGAGCCCAUUUCCUUCACAGCAUUUGGAAGCUCCUGAAGACAWSGR Ref. No. 54841-719.601AGUCAUUUGGCAGUUCAUUGAUGGAGAGUGAAGUAAAC CUGGACCGUUAUCAAACAGCUUUAGAAGAAGUAUUAUC GUGGCUUCUUUCUGCUGAGGACACAUUGCAAGCACAAG GAGAGAUUUCUAAUGAUGUGGAAGUGGUGAAAGACCAG UUUCAUACUCAUGAGGGGUACAUGAUGGAUUUGACAGC CCAUCAGGGCCGGGUUGGUAAUAUUCUACAAUUGGGAA GUAAGCUGAUUGGAACAGGAAAAUUAUCAGAAGAUGAA GAAACUGAAGUACAAGAGCAGAUGAAUCUCCUAAAUUC AAGAUGGGAAUGCCUCAGGGUAGCUAGCAUGGAAAAAC AAAGCAAUUUACAUAGAGUUUUAAUGGAUCUCCAGAAU CAGAAACUGAAAGAGUUGAAUGACUGGCUAACAAAAAC AGAAGAAAGAACAAGGAAAAUGGAGGAAGAGCCUCUUG GACCUGAUCUUGAAGACCUAAAACGCCAAGUACAACAAC AUAAGGUGCUUCAAGAAGAUCUAGAACAAGAACAAGUC AGGGUCAAUUCUCUCACUCACAUGGUGGUGGUAGUUGA UGAAUCUAGUGGAGAUCACGCAACUGCUGCUUUGGAAG AACAACUUAAGGUAUUGGGAGAUCGAUGGGCAAACAUC UGUAGAUGGACAGAAGACCGCUGGGUUCUUUUACAAGA CAUCCUUCUCAAAUGGCAACGUCUUACUGAAGAACAGU GCCUUUUUAGUGCAUGGCUUUCAGAAAAAGAAGAUGCA GUGAACAAGAUUCACACAACUGGCUUUAAAGAUCAAAA UGAAAUGUUAUCAAGUCUUCAAAAACUGGCCGUUUUAA AAGCGGAUCUAGAAAAGAAAAAGCAAUCCAUGGGCAAA CUGUAUUCACUCAAACAAGAUCUUCUUUCAACACUGAA GAAUAAGUCAGUGACCCAGAAGACGGAAGCAUGGCUGG AUAACUUUGCCCGGUGUUGGGAUAAUUUAGUCCAAAAA CUUGAAAAGAGUACAGCACAGAUUUCACAGGCUGUCAC CACCACUCAGCCAUCACUAACACAGACAACUGUAAUGGA AACAGUAACUACGGUGACCACAAGGGAACAGAUCCUGG UAAAGCAUGCUCAAGAGGAACUUCCACCACCACCUCCCC AAAAGAAGAGGCAGAUUACUGUGGAUUCUGAAAUUAGG AAAAGGUUGGAUGUUGAUAUAACUGAACUUCACAGCUG GAUUACUCGCUCAGAAGCUGUGUUGCAGAGUCCUGAAU UUGCAAUCUUUCGGAAGGAAGGCAACUUCUCAGACUUA AAAGAAAAAGUCAAUGCCAUAGAGCGAGAAAAAGCUGA GAAGUUCAGAAAACUGCAAGAUGCCAGCAGAUCAGCUC AGGCCCUGGUGGAACAGAUGGUGAAUGAGGGUGUUAAU GCAGAUAGCAUCAAACAAGCCUCAGAACAACUGAACAGC CGGUGGAUCGAAUUCUGCCAGUUGCUAAGUGAGAGACU UAACUGGCUGGAGUAUCAGAACAACAUCAUCGCUUUCU AUAAUCAGCUACAACAAUUGGAGCAGAUGACAACUACU GCUGAAAACUGGUUGAAAAUCCAACCCACCACCCCAUCA GAGCCAACAGCAAUUAAAAGUCAGUUAAAAAUUUGUAA GGAUGAAGUCAACCGGCUAUCAGGUCUUCAACCUCAAA UUGAACGAUUAAAAAUUCAAAGCAUAGCCCUGAAAGAG AAAGGACAAGGACCCAUGUUCCUGGAUGCAGACUUUGU GGCCUUUACAAAUCAUUUUAAGCAAGUCUUUUCUGAUG UGCAGGCCAGAGAGAAAGAGCUACAGACAAUUUUUGAC ACUUUGCCACCAAUGCGCUAUCAGGAGACCAUGAGUGCCAUCAGGACAUGGGUCCAGCAGUCAGAAACCAAACUCUCCWSGR Ref. No. 54841-719.601AUACCUCAACUUAGUGUCACCGACUAUGAAAUCAUGGA GCAGAGACUCGGGGAAUUGCAGGCUUUACAAAGUUCUC UGCAAGAGCAACAAAGUGGCCUAUACUAUCUCAGCACCA CUGUGAAAGAGAUGUCGAAGAAAGCGCCCUCUGAAAUU AGCCGGAAAUAUCAAUCAGAAUUUGAAGAAAUUGAGGG ACGCUGGAAGAAGCUCUCCUCCCAGCUGGUUGAGCAUUG UCAAAAGCUAGAGGAGCAAAUGAAUAAACUCCGAAAAA UUCAGAAUCACAUACAAACCCUGAAGAAAUGGAUGGCU GAAGUUGAUGUUUUUCUGAAGGAGGAAUGGCCUGCCCU UGGGGAUUCAGAAAUUCUAAAAAAGCAGCUGAAACAGU GCAGACUUUUAGUCAGUGAUAUUCAGACAAUUCAGCCC AGUCUAAACAGUGUCAAUGAAGGUGGGCAGAAGAUAAA GAAUGAAGCAGAGCCAGAGUUUGCUUCGAGACUUGAGA CAGAACUCAAAGAACUUAACACUCAGUGGGAUCACAUG UGCCAACAGGUCUAUGCCAGAAAGGAGGCCUUGAAGGG AGGUUUGGAGAAAACUGUAAGCCUCCAGAAAGAUCUAU CAGAGAUGCACGAAUGGAUGACACAAGCUGAAGAAGAG UAUCUUGAGAGAGAUUUUGAAUAUAAAACUCCAGAUGA AUUACAGAAAGCAGUUGAAGAGAUGAAGAGAGCUAAAG AAGAGGCCCAACAAAAAGAAGCGAAAGUGAAACUCCUU ACUGAGUCUGUAAAUAGUGUCAUAGCUCAAGCUCCACC UGUAGCACAAGAGGCCUUAAAAAAGGAACUUGAAACUC UAACCACCAACUACCAGUGGCUCUGCACUAGGCUGAAUG GGAAAUGCAAGACUUUGGAAGAAGUUUGGGCAUGUUGG CAUGAGUUAUUGUCAUACUUGGAGAAAGCAAACAAGUG GCUAAAUGAAGUAGAAUUUAAACUUAAAACCACUGAAA ACAUUCCUGGCGGAGCUGAGGAAAUCUCUGAGGUGCUA GAUUCACUUGAAAAUUUGAUGCGACAUUCAGAGGAUAA CCCAAAUCAGAUUCGCAUAUUGGCACAGACCCUAACAGA UGGCGGAGUCAUGGAUGAGCUAAUCAAUGAGGAACUUG AGACAUUUAAUUCUCGUUGGAGGGAACUACAUGAAGAG GCUGUAAGGAGGCAAAAGUUGCUUGAACAGAGCAUCCA GUCUGCCCAGGAGACUGAAAAAUCCUUACACUUAAUCCA GGAGUCCCUCACAUUCAUUGACAAGCAGUUGGCAGCUU AUAUUGCAGACAAGGUGGACGCAGCUCAAAUGCCUCAG GAAGCCCAGAAAAUCCAAUCUGAUUUGACAAGUCAUGA GAUCAGUUUAGAAGAAAUGAAGAAACAUAAUCAGGGGA AGGAGGCUGCCCAAAGAGUCCUGUCUCAGAUUGAUGUU GCACAGAAAAAAUUACAAGAUGUCUCCAUGAAGUUUCG AUUAUUCCAGAAACCAGCCAAUUUUGAGCAGCGUCUAC AAGAAAGUAAGAUGAUUUUAGAUGAAGUGAAGAUGCAC UUGCCUGCAUUGGAAACAAAGAGUGUGGAACAGGAAGU AGUACAGUCACAGCUAAAUCAUUGUGUGAACUUGUAUA AAAGUCUGAGUGAAGUGAAGUCUGAAGUGGAAAUGGUG AUAAAGACUGGACGUCAGAUUGUACAGAAAAAGCAGAC GGAAAAUCCCAAAGAACUUGAUGAAAGAGUAACAGCUU UGAAAUUGCAUUAUAAUGAGCUGGGAGCAAAGGUAACA GAAAGAAAGCAACAGUUGGAGAAAUGCUUGAAAUUGUC CCGUAAGAUGCGAAAGGAAAUGAAUGUCUUGACAGAAUGGCUGGCAGCUACAGAUAUGGAAUUGACAAAGAGAUCAWSGR Ref. No. 54841-719.601GCAGUUGAAGGAAUGCCUAGUAAUUUGGAUUCUGAAGU UGCCUGGGGAAAGGCUACUCAAAAAGAGAUUGAGAAAC AGAAGGUGCACCUGAAGAGUAUCACAGAGGUAGGAGAG GCCUUGAAAACAGUUUUGGGCAAGAAGGAGACGUUGGU GGAAGAUAAACUCAGUCUUCUGAAUAGUAACUGGAUAG CUGUCACCUCCCGAGCAGAAGAGUGGUUAAAUCUUUUG UUGGAAUACCAGAAACACAUGGAAACUUUUGACCAGAA UGUGGACCACAUCACAAAGUGGAUCAUUCAGGCUGACA CACUUUUGGAUGAAUCAGAGAAAAAGAAACCCCAGCAA AAAGAAGACGUGCUUAAGCGUUUAAAGGCAGAACUGAA UGACAUACGCCCAAAGGUGGACUCUACACGUGACCAAGC AGCAAACUUGAUGGCAAACCGCGGUGACCACUGCAGGA AAUUAGUAGAGCCCCAAAUCUCAGAGCUCAACCAUCGAU UUGCAGCCAUUUCACACAGAAUUAAGACUGGAAAGGCC UCCAUUCCUUUGAAGGAAUUGGAGCAGUUUAACUCAGA UAUACAAAAAUUGCUUGAACCACUGGAGGCUGAAAUUC AGCAGGGGGUGAAUCUGAAAGAGGAAGACUUCAAUAAA GAUAUGAAUGAAGACAAUGAGGGUACUGUAAAAGAAUU GUUGCAAAGAGGAGACAACUUACAACAAAGAAUCACAG AUGAGAGAAAGCGAGAGGAAAUAAAGAUAAAACAGCAG CUGUUACAGACAAAACAUAAUGCUCUCAAGGAUUUGAG GUCUCAAAGAAGAAAAAAGGCUCUAGAAAUUUCUCAUC AGUGGUAUCAGUACAAGAGGCAGGCUGAUGAUCUCCUG AAAUGCUUGGAUGACAUUGAAAAAAAAUUAGCCAGCCU ACCUGAGCCCAGAGAUGAAAGGAAAAUAAAGGAAAUUG AUCGGGAAUUGCAGAAGAAGAAAGAGGAGCUGAAUGCA GUGCGUAGGCAAGCUGAGGGCUUGUCUGAGGAUGGGGC CGCAAUGGCAGUGGAGCCAACUCAGAUCCAGCUCAGCAA GCGCUGGCGGGAAAUUGAGAGCAAAUUUGCUCAGUUUC GAAGACUCAACUUUGCACAAAUUCACACUGUCCGUGAA GAAACGAUGAUGGUGAUGACUGAAGACAUGCCUUUGGA AAUUUCUUAUGUGCCUUCUACUUAUUUGACUGAAAUCA CUCAUGUCUCACAAGCCCUAUUAGAAGUGGAACAACUUC UCAAUGCUCCUGACCUCUGUGCUAAGGACUUUGAAGAU CUCUUUAAGCAAGAGGAGUCUCUGAAGAAUAUAAAAGA UAGUCUACAACAAAGCUCAGGUCGGAUUGACAUUAUUC AUAGCAAGAAGACAGCAGCAUUGCAAAGUGCAACGCCU GUGGAAAGGGUGAAGCUACAGGAAGCUCUCUCCCAGCU UGAUUUCCAAUGGGAAAAAGUUAACAAAAUGUACAAGG ACCGACAAGGGCGAUUUGACAGAUCUGUUGAGAAAUGG CGGCGUUUUCAUUAUGAUAUAAAGAUAUUUAAUCAGUG GCUAACAGAAGCUGAACAGUUUCUCAGAAAGACACAAA UUCCUGAGAAUUGGGAACAUGCUAAAUACAAAUGGUAU CUUAAGGAACUCCAGGAUGGCAUUGGGCAGCGGCAAAC UGUUGUCAGAACAUUGAAUGCAACUGGGGAAGAAAUAA UUCAGCAAUCCUCAAAAACAGAUGCCAGUAUUCUACAG GAAAAAUUGGGAAGCCUGAAUCUGCGGUGGCAGGAGGU CUGCAAACAGCUGUCAGACAGAAAAAAGAGGCUAGAAG AACAAAAGAAUAUCUUGUCAGAAUUUCAAAGAGAUUUAAAUGAAUUUGUUUUAUGGUUGGAGGAAGCAGAUAACAUWSGR Ref. No. 54841-719.601UGCUAGUAUCCCACUUGAACCUGGAAAAGAGCAGCAAC UAAAAGAAAAGCUUGAGCAAGUCAAGUUACUGGUGGAA GAGUUGCCCCUGCGCCAGGGAAUUCUCAAACAAUUAAA UGAAACUGGAGGACCCGUGCUUGUAAGUGCUCCCAUAA GCCCAGAAGAGCAAGAUAAACUUGAAAAUAAGCUCAAG CAGACAAAUCUCCAGUGGAUAAAGGUUUCCAGAGCUUU ACCUGAGAAACAAGGAGAAAUUGAAGCUCAAAUAAAAG ACCUUGGGCAGCUUGAAAAAAAGCUUGAAGACCUUGAA GAGCAGUUAAAUCAUCUGCUGCUGUGGUUAUCUCCUAU UAGGAAUCAGUUGGAAAUUUAUAACCAACCAAACCAAG AAGGACCAUUUGACGUUCAGGAAACUGAAAUAGCAGUU CAAGCUAAACAACCGGAUGUGGAAGAGAUUUUGUCUAA AGGGCAGCAUUUGUACAAGGAAAAACCAGCCACUCAGCC AGUGAAGAGGAAGUUAGAAGAUCUGAGCUCUGAGUGGA AGGCGGUAAACCGUUUACUUCAAGAGCUGAGGGCAAAG CAGCCUGACCUAGCUCCUGGACUGACCACUAUUGGAGCC UCUCCUACUCAGACUGUUACUCUGGUGACACAACCUGUG GUUACUAAGGAAACUGCCAUCUCCAAACUAGAAAUGCC AUCUUCCUUGAUGUUGGAGGUACCUGCUCUGGCAGAUU UCAACCGGGCUUGGACAGAACUUACCGACUGGCUUUCUC UGCUUGAUCAAGUUAUAAAAUCACAGAGGGUGAUGGUG GGUGACCUUGAGGAUAUCAACGAGAUGAUCAUCAAGCA GAAGGCAACAAUGCAGGAUUUGGAACAGAGGCGUCCCC AGUUGGAAGAACUCAUUACCGCUGCCCAAAAUUUGAAA AACAAGACCAGCAAUCAAGAGGCUAGAACAAUCAUUAC GGAUCGAAUUGAAAGAAUUCAGAAUCAGUGGGAUGAAG UACAAGAACACCUUCAGAACCGGAGGCAACAGUUGAAU GAAAUGUUAAAGGAUUCAACACAAUGGCUGGAAGCUAA GGAAGAAGCUGAGCAGGUCUUAGGACAGGCCAGAGCCA AGCUUGAGUCAUGGAAGGAGGGUCCCUAUACAGUAGAU GCAAUCCAAAAGAAAAUCACAGAAACCAAGCAGUUGGC CAAAGACCUCCGCCAGUGGCAGACAAAUGUAGAUGUGG CAAAUGACUUGGCCCUGAAACUUCUCCGGGAUUAUUCU GCAGAUGAUACCAGAAAAGUCCACAUGAUAACAGAGAA UAUCAAUGCCUCUUGGAGAAGCAUUCAUAAAAGGGUGA GUGAGCGAGAGGCUGCUUUGGAAGAAACUCAUAGAUUA CUGCAACAGUUCCCCCUGGACCUGGAAAAGUUUCUUGCC UGGCUUACAGAAGCUGAAACAACUGCCAAUGUCCUACA GGAUGCUACCCGUAAGGAAAGGCUCCUAGAAGACUCCA AGGGAGUAAAAGAGCUGAUGAAACAAUGGCAAGACCUC CAAGGUGAAAUUGAAGCUCACACAGAUGUUUAUCACAA CCUGGAUGAAAACAGCCAAAAAAUCCUGAGAUCCCUGG AAGGUUCCGAUGAUGCAGUCCUGUUACAAAGACGUUUG GAUAACAUGAACUUCAAGUGGAGUGAACUUCGGAAAAA GUCUCUCAACAUUAGGUCCCAUUUGGAAGCCAGUUCUG ACCAGUGGAAGCGUCUGCACCUUUCUCUGCAGGAACUUC UGGUGUGGCUACAGCUGAAAGAUGAUGAAUUAAGCCGG CAGGCACCUAUUGGAGGCGACUUUCCAGCAGUUCAGAA GCAGAACGAUGUACAUAGGGCCUUCAAGAGGGAAUUGAAAACUAAAGAACCUGUAAUCAUGAGUACUCUUGAGACUWSGR Ref. No. 54841-719.601GUACGAAUAUUUCUGACAGAGCAGCCUUUGGAAGGACU AGAGAAACUCUACCAGGAGCCCAGAGAGCUGCCUCCUGA GGAGAGAGCCCAGAAUGUCACUCGGCUUCUACGAAAGC AGGCUGAGGAGGUCAAUACUGAGUGGGAAAAAUUGAAC CUGCACUCCGCUGACUGGCAGAGAAAAAUAGAUGAGAC CCUUGAAAGACUCCAGGAACUUCAAGAGGCCACGGAUG AGCUGGACCUCAAGCUGCGCCAAGCUGAGGUGAUCAAG GGAUCCUGGCAGCCCGUGGGCGAUCUCCUCAUUGACUCU CUCCAAGAUCACCUCGAGAAAGUCAAGGCACUUCGAGGA GAAAUUGCGCCUCUGAAAGAGAACGUGAGCCACGUCAA UGACCUUGCUCGCCAGCUUACCACUUUGGGCAUUCAGCU CUCACCGUAUAACCUCAGCACUCUGGAAGACCUGAACAC CAGAUGGAAGCUUCUGCAGGUGGCCGUCGAGGACCGAG UCAGGCAGCUGCAUGAAGCCCACAGGGACUUUGGUCCAG CAUCUCAGCACUUUCUUUCCACGUCUGUCCAGGGUCCCU GGGAGAGAGCCAUCUCGCCAAACAAAGUGCCCUACUAUA UCAACCACGAGACUCAAACAACUUGCUGGGACCAUCCCA AAAUGACAGAGCUCUACCAGUCUUUAGCUGACCUGAAU AAUGUCAGAUUCUCAGCUUAUAGGACUGCCAUGAAACU CCGAAGACUGCAGAAGGCCCUUUGCUUGGAUCUCUUGA GCCUGUCAGCUGCAUGUGAUGCCUUGGACCAGCACAACC UCAAGCAAAAUGACCAGCCCAUGGAUAUCCUGCAGAUU AUUAAUUGUUUGACCACUAUUUAUGACCGCCUGGAGCA AGAGCACAACAAUUUGGUCAACGUCCCUCUCUGCGUGGA UAUGUGUCUGAACUGGCUGCUGAAUGUUUAUGAUACGG GACGAACAGGGAGGAUCCGUGUCCUGUCUUUUAAAACU GGCAUCAUUUCCCUGUGUAAAGCACAUUUGGAAGACAA GUACAGAUACCUUUUCAAGCAAGUGGCAAGUUCAACAG GAUUUUGUGACCAGCGCAGGCUGGGCCUCCUUCUGCAUG AUUCUAUCCAAAUUCCAAGACAGUUGGGUGAAGUUGCA UCCUUUGGGGGCAGUAACAUUGAGCCAAGUGUCCGGAG CUGCUUCCAAUUUGCUAAUAAUAAGCCAGAGAUCGAAG CGGCCCUCUUCCUAGACUGGAUGAGACUGGAACCCCAGU CCAUGGUGUGGCUGCCCGUCCUGCACAGAGUGGCUGCUG CAGAAACUGCCAAGCAUCAGGCCAAAUGUAACAUCUGCA AAGAGUGUCCAAUCAUUGGAUUCAGGUACAGGAGUCUA AAGCACUUUAAUUAUGACAUCUGCCAAAGCUGCUUUUU UUCUGGUCGAGUUGCAAAAGGCCAUAAAAUGCACUAUC CCAUGGUGGAAUAUUGCACUCCGACUACAUCAGGAGAA GAUGUUCGAGACUUUGCCAAGGUACUAAAAAACAAAUU UCGAACCAAAAGGUAUUUUGCGAAGCAUCCCCGAAUGG GCUACCUGCCAGUGCAGACUGUCUUAGAGGGGGACAAC AUGGAAACUCCCGUUACUCUGAUCAACUUCUGGCCAGUA GAUUCUGCGCCUGCCUCGUCCCCUCAGCUUUCACACGAU GAUACUCAUUCACGCAUUGAACAUUAUGCUAGCAGGCU AGCAGAAAUGGAAAACAGCAAUGGAUCUUAUCUAAAUG AUAGCAUCUCUCCUAAUGAGAGCAUAGAUGAUGAACAU UUGUUAAUCCAGCAUUACUGCCAAAGUUUGAACCAGGA CUCCCCCCUGAGCCAGCCUCGUAGUCCUGCCCAGAUCUUGAUUUCCUUAGAGAGUGAGGAAAGAGGGGAGCUAGAGAWSGR Ref. No. 54841-719.601GAAUCCUAGCAGAUCUUGAGGAAGAAAACAGGAAUCUG CAAGCAGAAUAUGACCGUCUAAAGCAGCAGCACGAACA UAAAGGCCUGUCCCCACUGCCGUCCCCUCCUGAAAUGAU GCCCACCUCUCCCCAGAGUCCCCGGGAUGCUGAGCUCAU UGCUGAGGCCAAGCUACUGCGUCAACACAAAGGCCGCCU GGAAGCCAGGAUGCAAAUCCUGGAAGACCACAAUAAAC AGCUGGAGUCACAGUUACACAGGCUAAGGCAGCUGCUG GAGCAACCCCAGGCAGAGGCCAAAGUGAAUGGCACAACG GUGUCCUCUCCUUCUACCUCUCUACAGAGGUCCGACAGC AGUCAGCCUAUGCUGCUCCGAGUGGUUGGCAGUCAAAC UUCGGACUCCAUGGGUGAGGAAGAUCUUCUCAGUCCUCC CCAGGACACAAGCACAGGGUUAGAGGAGGUGAUGGAGC AACUCAACAACUCCUUCCCUAGUUCAAGAGGAAGAAAU ACCCCUGGAAAGCCAAUGAGAGAGGACACAAUGuaggaagu cuuuuccacauggcagaugauuugggcagagcgauggaguccuuaguaucagucaugg cagaugaagaaggagcagaauaaauguuuuacaacuccugauucccgcaugguuuuua uaauauucauacaacaaagaggauuagacaguaagaguuuacaagaaauaaaucuauau uuuugugaaggguagugguauuauacuguagauuucaguaguuucuaagucuguuau uguuuuguuaacaauggcagguuuuacacgucuaugcaauuguacaaaaaaguuauaa gaaaacuacauguaaaaucuugauagcuaaauaacuugccauuucuuuauauggaacg cauuuuggguuguuuaaaaauuuauaacaguuauaaagaaagauuguaaacuaaagug ugcuuuauaaaaaaaaguuguuuauaaaaaccccuaaaaacaaaacaaacacacacacac acacauacacacacacacacaaaacuuugaggcagcgcauuguuuugcauccuuuuggc gugauauccauaugaaauucauggcuuuuucuuuuuuugcauauuaaagauaagacu uccucuaccaccacaccaaaugacuacuacacacugcucauuugagaacugucagcuga guggggcaggcuugaguuuucauuucauauaucuauaugucuauaaguauauaaaua cuauaguuauauagauaaagagauacgaauuucuauagacugacuuuuuccauuuuuu aaauguucaugucacauccuaauagaaagaaauuacuucuagucagucauccaggcuu accugcuuggucuagaauggauuuuucccggagccggaagccaggaggaaacuacacc acacuaaaacauugucuacagcuccagauguuucucauuuuaaacaacuuuccacugac aacgaaaguaaaguaaaguauuggauuuuuuuaaagggaacaugugaaugaauacaca ggacuuauuauaucagagugaguaaucgguugguugguugauugauugauugauugg uacauucagcuuccugcugcuagcaaugccacgauuuagauuuaaugaugcuucagug gaaaucaaucagaagguauucugaccuugugaacaucagaagguauuuuuuaacuccc aagcaguagcaggacgaugauagggcuggagggcuauggauucccagcccaucccugu gaaggaguaggccacucuuuaagugaaggauuggaugauuguucauaauacauaaagu ucucuguaauuacaacuaaauuauuaugcccucuucucacagucaaaaggaacugggu gguuugguuuuuguugcuuuuuuagauuuauugucccaugugggaugaguuuuuaa augccacaagacauaauuuaaaauaaauaaacuuugggaaaagguguaaaacaguagcc ccaucacauuugugauacugacagguaucaacccagaagcccaugaacuguguuucca uccuuugcauuucucugcgaguaguuccacacagguuuguaaguaaguaagaaagaag gcaaauugauucaaauguuacaaaaaaacccuucuugguggauuagacagguuaaaua uauaaacaaacaaacaaaaauugcucaaaaaagaggagaaaagcucaagaggaaaagcua aggacugguaggaaaaagcuuuacucuuucaugccauuuuauuucuuuuugauuuuu aaaucauucauucaauagauaccaccgugugaccuauaauuuugcaaaucuguuaccu cugacaucaaguguaauuagcuuuuggagagugggcugacaucaaguguaauuagcu uuuggagaguggguuuuguccauuauuaauaauuaauuaauuaacaucaaacacggcu ucucaugcuauuucuaccucacuuugguuuugggguguuccugauaauugugcacac cugaguucacagcuucaccacuuguccauugcguuauuuucuuuuuccuuuauaauucuuucuuuuuccuucauaauuuucaaaagaaaacccaaagcucuaagguaacaaauuaccWSGR Ref. No. 54841-719.601aaauuacaugaagauuugguuuuugucuugcauuuuuuuccuuuaugugacgcugga ccuuuucuuuacccaaggauuuuuaaaacucagauuuaaaacaagggguuacuuuaca uccuacuaagaaguuuaaguaaguaaguuucauucuaaaaucagagguaaauagagug cauaaauaauuuuguuuuaaucuuuuuguuuuucuuuuagacacauuagcucuggag ugagucugucauaauauuugaacaaaaauugagagcuuuauugcugcauuuuaagcau aauuaauuuggacauuauuucguguuguguucuuuauaaccaccaaguauuaaacugu aaaucauaauguaacugaagcauaaacaucacauggcauguuuugucauuguuuucag guacugaguucuuacuugaguaucauaauauauuguguuuuaacaccaacacuguaac auuuacgaauuauuuuuuuaaacuucaguuuuacugcauuuucacaacauaucagacu ucaccaaauauaugccuuacuauuguauuauaguacugcuuuacuguguaucucaaua aagcacgcaguuauguuac24 Human augcugucugugaagcugaaucugugagaacaccucacuauucacggcaaccggagug dystrophin gaagaaacaggugcaaaaagauuguguguuugucugcuuuugugaggcuggucagag (DMD), auucugugccugcuuuaucugugcuuggcuaugacucuaccuccagguuuaccauacc transcript ccauagaauguguaagagaaaaguaccaacagggaaaucagcaaaaagcuuuccuauga variant agguguguagccagccuccgcagaauuugaaaugucugaggucucaucugaugaaaga Dp427pl, gaagauguucaaaagaaaacauucacaaaaugggUAAAUGCACAAUUUUC mRNA UAAGUUUGGGAAGCAGCAUAUUGAGAACCUCUUCAGUG ACCUACAGGAUGGGAGGCGCCUCCUAGACCUCCUCGAAG(Ref. Seq. GCCUGACAGGGCAAAAACUGCCAAAAGAAAAAGGAUCC NM 00400 ACAAGAGUUCAUGCCCUGAACAAUGUCAACAAGGCACU 9.3) GCGGGUUUUGCAGAACAAUAAUGUUGAUUUAGUGAAUA UUGGAAGUACUGACAUCGUAGAUGGAAAUCAUAAACUG(CODING ACUCUUGGUUUGAUUUGGAAUAUAAUCCUCCACUGGCA SEQUENC GGUCAAAAAUGUAAUGAAAAAUAUCAUGGCUGGAUUGCE) AACAAACCAACAGUGAAAAGAUUCUCCUGAGCUGGGUC CGACAAUCAACUCGUAAUUAUCCACAGGUUAAUGUAAU CAACUUCACCACCAGCUGGUCUGAUGGCCUGGCUUUGAA UGCUCUCAUCCAUAGUCAUAGGCCAGACCUAUUUGACUG GAAUAGUGUGGUUUGCCAGCAGUCAGCCACACAACGAC UGGAACAUGCAUUCAACAUCGCCAGAUAUCAAUUAGGC AUAGAGAAACUACUCGAUCCUGAAGAUGUUGAUACCAC CUAUCCAGAUAAGAAGUCCAUCUUAAUGUACAUCACAU CACUCUUCCAAGUUUUGCCUCAACAAGUGAGCAUUGAA GCCAUCCAGGAAGUGGAAAUGUUGCCAAGGCCACCUAA AGUGACUAAAGAAGAACAUUUUCAGUUACAUCAUCAAA UGCACUAUUCUCAACAGAUCACGGUCAGUCUAGCACAGG GAUAUGAGAGAACUUCUUCCCCUAAGCCUCGAUUCAAG AGCUAUGCCUACACACAGGCUGCUUAUGUCACCACCUCU GACCCUACACGGAGCCCAUUUCCUUCACAGCAUUUGGAA GCUCCUGAAGACAAGUCAUUUGGCAGUUCAUUGAUGGA GAGUGAAGUAAACCUGGACCGUUAUCAAACAGCUUUAG AAGAAGUAUUAUCGUGGCUUCUUUCUGCUGAGGACACA UUGCAAGCACAAGGAGAGAUUUCUAAUGAUGUGGAAGU GGUGAAAGACCAGUUUCAUACUCAUGAGGGGUACAUGA UGGAUUUGACAGCCCAUCAGGGCCGGGUUGGUAAUAUU CUACAAUUGGGAAGUAAGCUGAUUGGAACAGGAAAAUU AUCAGAAGAUGAAGAAACUGAAGUACAAGAGCAGAUGA AUCUCCUAAAUUCAAGAUGGGAAUGCCUCAGGGUAGCUAGCAUGGAAAAACAAAGCAAUUUACAUAGAGUUUUAAUWSGR Ref. No. 54841-719.601GGAUCUCCAGAAUCAGAAACUGAAAGAGUUGAAUGACU GGCUAACAAAAACAGAAGAAAGAACAAGGAAAAUGGAG GAAGAGCCUCUUGGACCUGAUCUUGAAGACCUAAAACG CCAAGUACAACAACAUAAGGUGCUUCAAGAAGAUCUAG AACAAGAACAAGUCAGGGUCAAUUCUCUCACUCACAUG GUGGUGGUAGUUGAUGAAUCUAGUGGAGAUCACGCAAC UGCUGCUUUGGAAGAACAACUUAAGGUAUUGGGAGAUC GAUGGGCAAACAUCUGUAGAUGGACAGAAGACCGCUGG GUUCUUUUACAAGACAUCCUUCUCAAAUGGCAACGUCU UACUGAAGAACAGUGCCUUUUUAGUGCAUGGCUUUCAG AAAAAGAAGAUGCAGUGAACAAGAUUCACACAACUGGC UUUAAAGAUCAAAAUGAAAUGUUAUCAAGUCUUCAAAA ACUGGCCGUUUUAAAAGCGGAUCUAGAAAAGAAAAAGC AAUCCAUGGGCAAACUGUAUUCACUCAAACAAGAUCUU CUUUCAACACUGAAGAAUAAGUCAGUGACCCAGAAGAC GGAAGCAUGGCUGGAUAACUUUGCCCGGUGUUGGGAUA AUUUAGUCCAAAAACUUGAAAAGAGUACAGCACAGAUU UCACAGGCUGUCACCACCACUCAGCCAUCACUAACACAG ACAACUGUAAUGGAAACAGUAACUACGGUGACCACAAG GGAACAGAUCCUGGUAAAGCAUGCUCAAGAGGAACUUC CACCACCACCUCCCCAAAAGAAGAGGCAGAUUACUGUGG AUUCUGAAAUUAGGAAAAGGUUGGAUGUUGAUAUAACU GAACUUCACAGCUGGAUUACUCGCUCAGAAGCUGUGUU GCAGAGUCCUGAAUUUGCAAUCUUUCGGAAGGAAGGCA ACUUCUCAGACUUAAAAGAAAAAGUCAAUGCCAUAGAG CGAGAAAAAGCUGAGAAGUUCAGAAAACUGCAAGAUGC CAGCAGAUCAGCUCAGGCCCUGGUGGAACAGAUGGUGA AUGAGGGUGUUAAUGCAGAUAGCAUCAAACAAGCCUCA GAACAACUGAACAGCCGGUGGAUCGAAUUCUGCCAGUU GCUAAGUGAGAGACUUAACUGGCUGGAGUAUCAGAACA ACAUCAUCGCUUUCUAUAAUCAGCUACAACAAUUGGAG CAGAUGACAACUACUGCUGAAAACUGGUUGAAAAUCCA ACCCACCACCCCAUCAGAGCCAACAGCAAUUAAAAGUCA GUUAAAAAUUUGUAAGGAUGAAGUCAACCGGCUAUCAG GUCUUCAACCUCAAAUUGAACGAUUAAAAAUUCAAAGC AUAGCCCUGAAAGAGAAAGGACAAGGACCCAUGUUCCU GGAUGCAGACUUUGUGGCCUUUACAAAUCAUUUUAAGC AAGUCUUUUCUGAUGUGCAGGCCAGAGAGAAAGAGCUA CAGACAAUUUUUGACACUUUGCCACCAAUGCGCUAUCAG GAGACCAUGAGUGCCAUCAGGACAUGGGUCCAGCAGUC AGAAACCAAACUCUCCAUACCUCAACUUAGUGUCACCGA CUAUGAAAUCAUGGAGCAGAGACUCGGGGAAUUGCAGG CUUUACAAAGUUCUCUGCAAGAGCAACAAAGUGGCCUA UACUAUCUCAGCACCACUGUGAAAGAGAUGUCGAAGAA AGCGCCCUCUGAAAUUAGCCGGAAAUAUCAAUCAGAAU UUGAAGAAAUUGAGGGACGCUGGAAGAAGCUCUCCUCC CAGCUGGUUGAGCAUUGUCAAAAGCUAGAGGAGCAAAU GAAUAAACUCCGAAAAAUUCAGAAUCACAUACAAACCC UGAAGAAAUGGAUGGCUGAAGUUGAUGUUUUUCUGAAGGAGGAAUGGCCUGCCCUUGGGGAUUCAGAAAUUCUAAAWSGR Ref. No. 54841-719.601AAAGCAGCUGAAACAGUGCAGACUUUUAGUCAGUGAUA UUCAGACAAUUCAGCCCAGUCUAAACAGUGUCAAUGAA GGUGGGCAGAAGAUAAAGAAUGAAGCAGAGCCAGAGUU UGCUUCGAGACUUGAGACAGAACUCAAAGAACUUAACA CUCAGUGGGAUCACAUGUGCCAACAGGUCUAUGCCAGA AAGGAGGCCUUGAAGGGAGGUUUGGAGAAAACUGUAAG CCUCCAGAAAGAUCUAUCAGAGAUGCACGAAUGGAUGA CACAAGCUGAAGAAGAGUAUCUUGAGAGAGAUUUUGAA UAUAAAACUCCAGAUGAAUUACAGAAAGCAGUUGAAGA GAUGAAGAGAGCUAAAGAAGAGGCCCAACAAAAAGAAG CGAAAGUGAAACUCCUUACUGAGUCUGUAAAUAGUGUC AUAGCUCAAGCUCCACCUGUAGCACAAGAGGCCUUAAAA AAGGAACUUGAAACUCUAACCACCAACUACCAGUGGCUC UGCACUAGGCUGAAUGGGAAAUGCAAGACUUUGGAAGA AGUUUGGGCAUGUUGGCAUGAGUUAUUGUCAUACUUGG AGAAAGCAAACAAGUGGCUAAAUGAAGUAGAAUUUAAA CUUAAAACCACUGAAAACAUUCCUGGCGGAGCUGAGGA AAUCUCUGAGGUGCUAGAUUCACUUGAAAAUUUGAUGC GACAUUCAGAGGAUAACCCAAAUCAGAUUCGCAUAUUG GCACAGACCCUAACAGAUGGCGGAGUCAUGGAUGAGCU AAUCAAUGAGGAACUUGAGACAUUUAAUUCUCGUUGGA GGGAACUACAUGAAGAGGCUGUAAGGAGGCAAAAGUUG CUUGAACAGAGCAUCCAGUCUGCCCAGGAGACUGAAAA AUCCUUACACUUAAUCCAGGAGUCCCUCACAUUCAUUGA CAAGCAGUUGGCAGCUUAUAUUGCAGACAAGGUGGACG CAGCUCAAAUGCCUCAGGAAGCCCAGAAAAUCCAAUCUG AUUUGACAAGUCAUGAGAUCAGUUUAGAAGAAAUGAAG AAACAUAAUCAGGGGAAGGAGGCUGCCCAAAGAGUCCU GUCUCAGAUUGAUGUUGCACAGAAAAAAUUACAAGAUG UCUCCAUGAAGUUUCGAUUAUUCCAGAAACCAGCCAAU UUUGAGCAGCGUCUACAAGAAAGUAAGAUGAUUUUAGA UGAAGUGAAGAUGCACUUGCCUGCAUUGGAAACAAAGA GUGUGGAACAGGAAGUAGUACAGUCACAGCUAAAUCAU UGUGUGAACUUGUAUAAAAGUCUGAGUGAAGUGAAGUC UGAAGUGGAAAUGGUGAUAAAGACUGGACGUCAGAUUG UACAGAAAAAGCAGACGGAAAAUCCCAAAGAACUUGAU GAAAGAGUAACAGCUUUGAAAUUGCAUUAUAAUGAGCU GGGAGCAAAGGUAACAGAAAGAAAGCAACAGUUGGAGA AAUGCUUGAAAUUGUCCCGUAAGAUGCGAAAGGAAAUG AAUGUCUUGACAGAAUGGCUGGCAGCUACAGAUAUGGA AUUGACAAAGAGAUCAGCAGUUGAAGGAAUGCCUAGUA AUUUGGAUUCUGAAGUUGCCUGGGGAAAGGCUACUCAA AAAGAGAUUGAGAAACAGAAGGUGCACCUGAAGAGUAU CACAGAGGUAGGAGAGGCCUUGAAAACAGUUUUGGGCA AGAAGGAGACGUUGGUGGAAGAUAAACUCAGUCUUCUG AAUAGUAACUGGAUAGCUGUCACCUCCCGAGCAGAAGA GUGGUUAAAUCUUUUGUUGGAAUACCAGAAACACAUGG AAACUUUUGACCAGAAUGUGGACCACAUCACAAAGUGG AUCAUUCAGGCUGACACACUUUUGGAUGAAUCAGAGAAAAAGAAACCCCAGCAAAAAGAAGACGUGCUUAAGCGUUWSGR Ref. No. 54841-719.601UAAAGGCAGAACUGAAUGACAUACGCCCAAAGGUGGAC UCUACACGUGACCAAGCAGCAAACUUGAUGGCAAACCGC GGUGACCACUGCAGGAAAUUAGUAGAGCCCCAAAUCUC AGAGCUCAACCAUCGAUUUGCAGCCAUUUCACACAGAAU UAAGACUGGAAAGGCCUCCAUUCCUUUGAAGGAAUUGG AGCAGUUUAACUCAGAUAUACAAAAAUUGCUUGAACCA CUGGAGGCUGAAAUUCAGCAGGGGGUGAAUCUGAAAGA GGAAGACUUCAAUAAAGAUAUGAAUGAAGACAAUGAGG GUACUGUAAAAGAAUUGUUGCAAAGAGGAGACAACUUA CAACAAAGAAUCACAGAUGAGAGAAAGCGAGAGGAAAU AAAGAUAAAACAGCAGCUGUUACAGACAAAACAUAAUG CUCUCAAGGAUUUGAGGUCUCAAAGAAGAAAAAAGGCU CUAGAAAUUUCUCAUCAGUGGUAUCAGUACAAGAGGCA GGCUGAUGAUCUCCUGAAAUGCUUGGAUGACAUUGAAA AAAAAUUAGCCAGCCUACCUGAGCCCAGAGAUGAAAGG AAAAUAAAGGAAAUUGAUCGGGAAUUGCAGAAGAAGAA AGAGGAGCUGAAUGCAGUGCGUAGGCAAGCUGAGGGCU UGUCUGAGGAUGGGGCCGCAAUGGCAGUGGAGCCAACU CAGAUCCAGCUCAGCAAGCGCUGGCGGGAAAUUGAGAG CAAAUUUGCUCAGUUUCGAAGACUCAACUUUGCACAAA UUCACACUGUCCGUGAAGAAACGAUGAUGGUGAUGACU GAAGACAUGCCUUUGGAAAUUUCUUAUGUGCCUUCUAC UUAUUUGACUGAAAUCACUCAUGUCUCACAAGCCCUAU UAGAAGUGGAACAACUUCUCAAUGCUCCUGACCUCUGU GCUAAGGACUUUGAAGAUCUCUUUAAGCAAGAGGAGUC UCUGAAGAAUAUAAAAGAUAGUCUACAACAAAGCUCAG GUCGGAUUGACAUUAUUCAUAGCAAGAAGACAGCAGCA UUGCAAAGUGCAACGCCUGUGGAAAGGGUGAAGCUACA GGAAGCUCUCUCCCAGCUUGAUUUCCAAUGGGAAAAAG UUAACAAAAUGUACAAGGACCGACAAGGGCGAUUUGAC AGAUCUGUUGAGAAAUGGCGGCGUUUUCAUUAUGAUAU AAAGAUAUUUAAUCAGUGGCUAACAGAAGCUGAACAGU UUCUCAGAAAGACACAAAUUCCUGAGAAUUGGGAACAU GCUAAAUACAAAUGGUAUCUUAAGGAACUCCAGGAUGG CAUUGGGCAGCGGCAAACUGUUGUCAGAACAUUGAAUG CAACUGGGGAAGAAAUAAUUCAGCAAUCCUCAAAAACA GAUGCCAGUAUUCUACAGGAAAAAUUGGGAAGCCUGAA UCUGCGGUGGCAGGAGGUCUGCAAACAGCUGUCAGACA GAAAAAAGAGGCUAGAAGAACAAAAGAAUAUCUUGUCA GAAUUUCAAAGAGAUUUAAAUGAAUUUGUUUUAUGGUU GGAGGAAGCAGAUAACAUUGCUAGUAUCCCACUUGAAC CUGGAAAAGAGCAGCAACUAAAAGAAAAGCUUGAGCAA GUCAAGUUACUGGUGGAAGAGUUGCCCCUGCGCCAGGG AAUUCUCAAACAAUUAAAUGAAACUGGAGGACCCGUGC UUGUAAGUGCUCCCAUAAGCCCAGAAGAGCAAGAUAAA CUUGAAAAUAAGCUCAAGCAGACAAAUCUCCAGUGGAU AAAGGUUUCCAGAGCUUUACCUGAGAAACAAGGAGAAA UUGAAGCUCAAAUAAAAGACCUUGGGCAGCUUGAAAAA AAGCUUGAAGACCUUGAAGAGCAGUUAAAUCAUCUGCUGCUGUGGUUAUCUCCUAUUAGGAAUCAGUUGGAAAUUUWSGR Ref. No. 54841-719.601AUAACCAACCAAACCAAGAAGGACCAUUUGACGUUCAG GAAACUGAAAUAGCAGUUCAAGCUAAACAACCGGAUGU GGAAGAGAUUUUGUCUAAAGGGCAGCAUUUGUACAAGG AAAAACCAGCCACUCAGCCAGUGAAGAGGAAGUUAGAA GAUCUGAGCUCUGAGUGGAAGGCGGUAAACCGUUUACU UCAAGAGCUGAGGGCAAAGCAGCCUGACCUAGCUCCUGG ACUGACCACUAUUGGAGCCUCUCCUACUCAGACUGUUAC UCUGGUGACACAACCUGUGGUUACUAAGGAAACUGCCA UCUCCAAACUAGAAAUGCCAUCUUCCUUGAUGUUGGAG GUACCUGCUCUGGCAGAUUUCAACCGGGCUUGGACAGA ACUUACCGACUGGCUUUCUCUGCUUGAUCAAGUUAUAA AAUCACAGAGGGUGAUGGUGGGUGACCUUGAGGAUAUC AACGAGAUGAUCAUCAAGCAGAAGGCAACAAUGCAGGA UUUGGAACAGAGGCGUCCCCAGUUGGAAGAACUCAUUA CCGCUGCCCAAAAUUUGAAAAACAAGACCAGCAAUCAAG AGGCUAGAACAAUCAUUACGGAUCGAAUUGAAAGAAUU CAGAAUCAGUGGGAUGAAGUACAAGAACACCUUCAGAA CCGGAGGCAACAGUUGAAUGAAAUGUUAAAGGAUUCAA CACAAUGGCUGGAAGCUAAGGAAGAAGCUGAGCAGGUC UUAGGACAGGCCAGAGCCAAGCUUGAGUCAUGGAAGGA GGGUCCCUAUACAGUAGAUGCAAUCCAAAAGAAAAUCA CAGAAACCAAGCAGUUGGCCAAAGACCUCCGCCAGUGGC AGACAAAUGUAGAUGUGGCAAAUGACUUGGCCCUGAAA CUUCUCCGGGAUUAUUCUGCAGAUGAUACCAGAAAAGU CCACAUGAUAACAGAGAAUAUCAAUGCCUCUUGGAGAA GCAUUCAUAAAAGGGUGAGUGAGCGAGAGGCUGCUUUG GAAGAAACUCAUAGAUUACUGCAACAGUUCCCCCUGGAC CUGGAAAAGUUUCUUGCCUGGCUUACAGAAGCUGAAAC AACUGCCAAUGUCCUACAGGAUGCUACCCGUAAGGAAA GGCUCCUAGAAGACUCCAAGGGAGUAAAAGAGCUGAUG AAACAAUGGCAAGACCUCCAAGGUGAAAUUGAAGCUCA CACAGAUGUUUAUCACAACCUGGAUGAAAACAGCCAAA AAAUCCUGAGAUCCCUGGAAGGUUCCGAUGAUGCAGUC CUGUUACAAAGACGUUUGGAUAACAUGAACUUCAAGUG GAGUGAACUUCGGAAAAAGUCUCUCAACAUUAGGUCCC AUUUGGAAGCCAGUUCUGACCAGUGGAAGCGUCUGCAC CUUUCUCUGCAGGAACUUCUGGUGUGGCUACAGCUGAA AGAUGAUGAAUUAAGCCGGCAGGCACCUAUUGGAGGCG ACUUUCCAGCAGUUCAGAAGCAGAACGAUGUACAUAGG GCCUUCAAGAGGGAAUUGAAAACUAAAGAACCUGUAAU CAUGAGUACUCUUGAGACUGUACGAAUAUUUCUGACAG AGCAGCCUUUGGAAGGACUAGAGAAACUCUACCAGGAG CCCAGAGAGCUGCCUCCUGAGGAGAGAGCCCAGAAUGUC ACUCGGCUUCUACGAAAGCAGGCUGAGGAGGUCAAUAC UGAGUGGGAAAAAUUGAACCUGCACUCCGCUGACUGGC AGAGAAAAAUAGAUGAGACCCUUGAAAGACUCCAGGAA CUUCAAGAGGCCACGGAUGAGCUGGACCUCAAGCUGCGC CAAGCUGAGGUGAUCAAGGGAUCCUGGCAGCCCGUGGG CGAUCUCCUCAUUGACUCUCUCCAAGAUCACCUCGAGAAAGUCAAGGCACUUCGAGGAGAAAUUGCGCCUCUGAAAGWSGR Ref. No. 54841-719.601AGAACGUGAGCCACGUCAAUGACCUUGCUCGCCAGCUUA CCACUUUGGGCAUUCAGCUCUCACCGUAUAACCUCAGCA CUCUGGAAGACCUGAACACCAGAUGGAAGCUUCUGCAG GUGGCCGUCGAGGACCGAGUCAGGCAGCUGCAUGAAGCC CACAGGGACUUUGGUCCAGCAUCUCAGCACUUUCUUUCC ACGUCUGUCCAGGGUCCCUGGGAGAGAGCCAUCUCGCCA AACAAAGUGCCCUACUAUAUCAACCACGAGACUCAAACA ACUUGCUGGGACCAUCCCAAAAUGACAGAGCUCUACCAG UCUUUAGCUGACCUGAAUAAUGUCAGAUUCUCAGCUUA UAGGACUGCCAUGAAACUCCGAAGACUGCAGAAGGCCCU UUGCUUGGAUCUCUUGAGCCUGUCAGCUGCAUGUGAUG CCUUGGACCAGCACAACCUCAAGCAAAAUGACCAGCCCA UGGAUAUCCUGCAGAUUAUUAAUUGUUUGACCACUAUU UAUGACCGCCUGGAGCAAGAGCACAACAAUUUGGUCAA CGUCCCUCUCUGCGUGGAUAUGUGUCUGAACUGGCUGCU GAAUGUUUAUGAUACGGGACGAACAGGGAGGAUCCGUG UCCUGUCUUUUAAAACUGGCAUCAUUUCCCUGUGUAAA GCACAUUUGGAAGACAAGUACAGAUACCUUUUCAAGCA AGUGGCAAGUUCAACAGGAUUUUGUGACCAGCGCAGGC UGGGCCUCCUUCUGCAUGAUUCUAUCCAAAUUCCAAGAC AGUUGGGUGAAGUUGCAUCCUUUGGGGGCAGUAACAUU GAGCCAAGUGUCCGGAGCUGCUUCCAAUUUGCUAAUAA UAAGCCAGAGAUCGAAGCGGCCCUCUUCCUAGACUGGAU GAGACUGGAACCCCAGUCCAUGGUGUGGCUGCCCGUCCU GCACAGAGUGGCUGCUGCAGAAACUGCCAAGCAUCAGGC CAAAUGUAACAUCUGCAAAGAGUGUCCAAUCAUUGGAU UCAGGUACAGGAGUCUAAAGCACUUUAAUUAUGACAUC UGCCAAAGCUGCUUUUUUUCUGGUCGAGUUGCAAAAGG CCAUAAAAUGCACUAUCCCAUGGUGGAAUAUUGCACUCC GACUACAUCAGGAGAAGAUGUUCGAGACUUUGCCAAGG UACUAAAAAACAAAUUUCGAACCAAAAGGUAUUUUGCG AAGCAUCCCCGAAUGGGCUACCUGCCAGUGCAGACUGUC UUAGAGGGGGACAACAUGGAAACUCCCGUUACUCUGAU CAACUUCUGGCCAGUAGAUUCUGCGCCUGCCUCGUCCCC UCAGCUUUCACACGAUGAUACUCAUUCACGCAUUGAACA UUAUGCUAGCAGGCUAGCAGAAAUGGAAAACAGCAAUG GAUCUUAUCUAAAUGAUAGCAUCUCUCCUAAUGAGAGC AUAGAUGAUGAACAUUUGUUAAUCCAGCAUUACUGCCA AAGUUUGAACCAGGACUCCCCCCUGAGCCAGCCUCGUAG UCCUGCCCAGAUCUUGAUUUCCUUAGAGAGUGAGGAAA GAGGGGAGCUAGAGAGAAUCCUAGCAGAUCUUGAGGAA GAAAACAGGAAUCUGCAAGCAGAAUAUGACCGUCUAAA GCAGCAGCACGAACAUAAAGGCCUGUCCCCACUGCCGUC CCCUCCUGAAAUGAUGCCCACCUCUCCCCAGAGUCCCCG GGAUGCUGAGCUCAUUGCUGAGGCCAAGCUACUGCGUC AACACAAAGGCCGCCUGGAAGCCAGGAUGCAAAUCCUGG AAGACCACAAUAAACAGCUGGAGUCACAGUUACACAGG CUAAGGCAGCUGCUGGAGCAACCCCAGGCAGAGGCCAAA GUGAAUGGCACAACGGUGUCCUCUCCUUCUACCUCUCUACAGAGGUCCGACAGCAGUCAGCCUAUGCUGCUCCGAGUGWSGR Ref. No. 54841-719.601GUUGGCAGUCAAACUUCGGACUCCAUGGGUGAGGAAGA UCUUCUCAGUCCUCCCCAGGACACAAGCACAGGGUUAGA GGAGGUGAUGGAGCAACUCAACAACUCCUUCCCUAGUUC AAGAGGAAGAAAUACCCCUGGAAAGCCAAUGAGAGAGG ACACAAUGuaggaagucuuuuccacauggcagaugauuugggcagagcgaugg aguccuuaguaucagucaugacagaugaagaaggagcagaauaaauguuuuacaacuc cugauucccgcaugguuuuuauaauauucauacaacaaagaggauuagacaguaagag uuuacaagaaauaaaucuauauuuuugugaaggguagugguauuauacuguagauuu caguaguuucuaagucuguuauuguuuuguuaacaauggcagguuuuacacgucuau gcaauuguacaaaaaaguuauaagaaaacuacauguaaaaucuugauagcuaaauaacu ugccauuucuuuauauggaacgcauuuuggguuguuuaaaaauuuauaacaguuaua aagaaagauuguaaacuaaagugugcuuuauaaaaaaaaguuguuuauaaaaaccccua aaaacaaaacaaacacacacacacacacauacacacacacacacaaaacuuugaggcagcg cauuguuuugcauccuuuuggcgugauauccauaugaaauucauggcuuuuucuuuu uuugcauauuaaagauaagacuuccucuaccaccacaccaaaugacuacuacacacugc ucauuugagaacugucagcugaguggggcaggcuugaguuuucauuucauauaucua uaugucuauaaguauauaaauacuauaguuauauagauaaagagauacgaauuucuau agacugacuuuuuccauuuuuuaaauguucaugucacauccuaauagaaagaaauuac uucuagucagucauccaggcuuaccugcuuggucuagaauggauuuuucccggagccg gaagccaggaggaaacuacaccacacuaaaacauugucuacagcuccagauguuucuca uuuuaaacaacuuuccacugacaacgaaaguaaaguaaaguauuggauuuuuuuaaag ggaacaugugaaugaauacacaggacuuauuauaucagagugaguaaucgguugguug guugauugauugauugauugauacauucagcuuccugcugcuagcaaugccacgauu uagauuuaaugaugcuucaguggaaaucaaucagaagguauucugaccuugugaacau cagaagguauuuuuuaacucccaagcaguagcaggacgaugauagggcuggagggcua uggauucccagcccaucccugugaaggaguaggccacucuuuaagugaaggauuggau gauuguucauaauacauaaaguucucuguaauuacaacuaaauuauuaugcccucuuc ucacagucaaaaggaacugggugguuugguuuuuguugcuuuuuuagauuuauuguc ccaugugggaugaguuuuuaaaugccacaagacauaauuuaaaauaaauaaacuuugg gaaaagguguaaaacaguagccccaucacauuugugauacugacagguaucaacccagagcccaugaacuguguuuccauccuuugcauuucucugcgaguaguuccacacagguu uguaaguaaguaagaaagaaggcaaauugauucaaauguuacaaaaaaacccuucuug guggauuagacagguuaaauauauaaacaaacaaacaaaaauugcucaaaaaagaggag aaaagcucaagaggaaaagcuaaggacugguaggaaaaagcuuuacucuuucaugcca uuuuauuucuuuuugauuuuuaaaucauucauucaauagauaccaccgugugaccuau aauuuugcaaaucuguuaccucugacaucaaguguaauuagcuuuuggagagugggc ugacaucaaguguaauuagcuuuuggagaguggguuuuguccauuauuaauaauuaa uuaauuaacaucaaacacggcuucucaugcuauuucuaccucacuuugguuuuggggu guuccugauaauugugcacaccugaguucacagcuucaccacuuguccauugcguuau uuucuuuuuccuuuauaauucuuucuuuuuccuucauaauuuucaaaagaaaacccaa agcucuaagguaacaaauuaccaaauuacaugaagauuugguuuuugucuugcauuuu uuuccuuuaugugacgcuggaccuuuucuuuacccaaggauuuuuaaaacucagauuu aaaacaagggguuacuuuacauccuacuaagaaguuuaaguaaguaaguuucauucua aaaucagagguaaauagagugcauaaauaauuuuguuuuaaucuuuuuguuuuucuu uuagacacauuagcucuggagugagucugucauaauauuugaacaaaaauugagagcu uuauugcugcauuuuaagcauaauuaauuuggacauuauuucguguuguguucuuua uaaccaccaaguauuaaacuguaaaucauaauguaacugaagcauaaacaucacauggc auguuuugucauuguuuucagguacugaguucuuacuugaguaucauaauauauuguguuuuaacaccaacacuguaacauuuacgaauuauuuuuuuaaacuucaguuuuacugWSGR Ref. No. 54841-719.601cauuuucacaacauaucagacuucaccaaauauaugccuuacuauuguauuauaguac ugcuuuacuguguaucucaauaaagcacgcaguuauguuac25 Human cugagaaagacagauugcaAUGACUGAGAUGAUUUUGCUAAUUUU dystrophin UUUUCCAGCCUAUUUCCUUAAUGCUGUAAGGAGGCAAA (DMD) AGUUGCUUGAACAGAGCAUCCAGUCUGCCCAGGAGACU transcript GAAAAAUCCUUACACUUAAUCCAGGAGUCCCUCACAUUC variant AUUGACAAGCAGUUGGCAGCUUAUAUUGCAGACAAGGU Dp260-l, GGACGCAGCUCAAAUGCCUCAGGAAGCCCAGAAAAUCCA mRNA AUCUGAUUUGACAAGUCAUGAGAUCAGUUUAGAAGAAA (Ref. Seq. UGAAGAAACAUAAUCAGGGGAAGGAGGCUGCCCAAAGA NM 00401 GUCCUGUCUCAGAUUGAUGUUGCACAGAAAAAAUUACA 1-4) AGAUGUCUCCAUGAAGUUUCGAUUAUUCCAGAAACCAG CCAAUUUUGAGCAGCGUCUACAAGAAAGUAAGAUGAUU(CODING UUAGAUGAAGUGAAGAUGCACUUGCCUGCAUUGGAAAC SEQUENC AAAGAGUGUGGAACAGGAAGUAGUACAGUCACAGCUAAE) AUCAUUGUGUGAACUUGUAUAAAAGUCUGAGUGAAGUG AAGUCUGAAGUGGAAAUGGUGAUAAAGACUGGACGUCA GAUUGUACAGAAAAAGCAGACGGAAAAUCCCAAAGAAC UUGAUGAAAGAGUAACAGCUUUGAAAUUGCAUUAUAAU GAGCUGGGAGCAAAGGUAACAGAAAGAAAGCAACAGUU GGAGAAAUGCUUGAAAUUGUCCCGUAAGAUGCGAAAGG AAAUGAAUGUCUUGACAGAAUGGCUGGCAGCUACAGAU AUGGAAUUGACAAAGAGAUCAGCAGUUGAAGGAAUGCC UAGUAAUUUGGAUUCUGAAGUUGCCUGGGGAAAGGCUA CUCAAAAAGAGAUUGAGAAACAGAAGGUGCACCUGAAG AGUAUCACAGAGGUAGGAGAGGCCUUGAAAACAGUUUU GGGCAAGAAGGAGACGUUGGUGGAAGAUAAACUCAGUC UUCUGAAUAGUAACUGGAUAGCUGUCACCUCCCGAGCA GAAGAGUGGUUAAAUCUUUUGUUGGAAUACCAGAAACA CAUGGAAACUUUUGACCAGAAUGUGGACCACAUCACAA AGUGGAUCAUUCAGGCUGACACACUUUUGGAUGAAUCA GAGAAAAAGAAACCCCAGCAAAAAGAAGACGUGCUUAA GCGUUUAAAGGCAGAACUGAAUGACAUACGCCCAAAGG UGGACUCUACACGUGACCAAGCAGCAAACUUGAUGGCA AACCGCGGUGACCACUGCAGGAAAUUAGUAGAGCCCCAA AUCUCAGAGCUCAACCAUCGAUUUGCAGCCAUUUCACAC AGAAUUAAGACUGGAAAGGCCUCCAUUCCUUUGAAGGA AUUGGAGCAGUUUAACUCAGAUAUACAAAAAUUGCUUG AACCACUGGAGGCUGAAAUUCAGCAGGGGGUGAAUCUG AAAGAGGAAGACUUCAAUAAAGAUAUGAAUGAAGACAA UGAGGGUACUGUAAAAGAAUUGUUGCAAAGAGGAGACA ACUUACAACAAAGAAUCACAGAUGAGAGAAAGCGAGAG GAAAUAAAGAUAAAACAGCAGCUGUUACAGACAAAACA UAAUGCUCUCAAGGAUUUGAGGUCUCAAAGAAGAAAAA AGGCUCUAGAAAUUUCUCAUCAGUGGUAUCAGUACAAG AGGCAGGCUGAUGAUCUCCUGAAAUGCUUGGAUGACAU UGAAAAAAAAUUAGCCAGCCUACCUGAGCCCAGAGAUG AAAGGAAAAUAAAGGAAAUUGAUCGGGAAUUGCAGAAG AAGAAAGAGGAGCUGAAUGCAGUGCGUAGGCAAGCUGAGGGCUUGUCUGAGGAUGGGGCCGCAAUGGCAGUGGAGCWSGR Ref. No. 54841-719.601CAACUCAGAUCCAGCUCAGCAAGCGCUGGCGGGAAAUUG AGAGCAAAUUUGCUCAGUUUCGAAGACUCAACUUUGCA CAAAUUCACACUGUCCGUGAAGAAACGAUGAUGGUGAU GACUGAAGACAUGCCUUUGGAAAUUUCUUAUGUGCCUU CUACUUAUUUGACUGAAAUCACUCAUGUCUCACAAGCCC UAUUAGAAGUGGAACAACUUCUCAAUGCUCCUGACCUC UGUGCUAAGGACUUUGAAGAUCUCUUUAAGCAAGAGGA GUCUCUGAAGAAUAUAAAAGAUAGUCUACAACAAAGCU CAGGUCGGAUUGACAUUAUUCAUAGCAAGAAGACAGCA GCAUUGCAAAGUGCAACGCCUGUGGAAAGGGUGAAGCU ACAGGAAGCUCUCUCCCAGCUUGAUUUCCAAUGGGAAA AAGUUAACAAAAUGUACAAGGACCGACAAGGGCGAUUU GACAGAUCUGUUGAGAAAUGGCGGCGUUUUCAUUAUGA UAUAAAGAUAUUUAAUCAGUGGCUAACAGAAGCUGAAC AGUUUCUCAGAAAGACACAAAUUCCUGAGAAUUGGGAA CAUGCUAAAUACAAAUGGUAUCUUAAGGAACUCCAGGA UGGCAUUGGGCAGCGGCAAACUGUUGUCAGAACAUUGA AUGCAACUGGGGAAGAAAUAAUUCAGCAAUCCUCAAAA ACAGAUGCCAGUAUUCUACAGGAAAAAUUGGGAAGCCU GAAUCUGCGGUGGCAGGAGGUCUGCAAACAGCUGUCAG ACAGAAAAAAGAGGCUAGAAGAACAAAAGAAUAUCUUG UCAGAAUUUCAAAGAGAUUUAAAUGAAUUUGUUUUAUG GUUGGAGGAAGCAGAUAACAUUGCUAGUAUCCCACUUG AACCUGGAAAAGAGCAGCAACUAAAAGAAAAGCUUGAG CAAGUCAAGUUACUGGUGGAAGAGUUGCCCCUGCGCCA GGGAAUUCUCAAACAAUUAAAUGAAACUGGAGGACCCG UGCUUGUAAGUGCUCCCAUAAGCCCAGAAGAGCAAGAU AAACUUGAAAAUAAGCUCAAGCAGACAAAUCUCCAGUG GAUAAAGGUUUCCAGAGCUUUACCUGAGAAACAAGGAG AAAUUGAAGCUCAAAUAAAAGACCUUGGGCAGCUUGAA AAAAAGCUUGAAGACCUUGAAGAGCAGUUAAAUCAUCU GCUGCUGUGGUUAUCUCCUAUUAGGAAUCAGUUGGAAA UUUAUAACCAACCAAACCAAGAAGGACCAUUUGACGUU AAGGAAACUGAAAUAGCAGUUCAAGCUAAACAACCGGA UGUGGAAGAGAUUUUGUCUAAAGGGCAGCAUUUGUACA AGGAAAAACCAGCCACUCAGCCAGUGAAGAGGAAGUUA GAAGAUCUGAGCUCUGAGUGGAAGGCGGUAAACCGUUU ACUUCAAGAGCUGAGGGCAAAGCAGCCUGACCUAGCUCC UGGACUGACCACUAUUGGAGCCUCUCCUACUCAGACUGU UACUCUGGUGACACAACCUGUGGUUACUAAGGAAACUG CCAUCUCCAAACUAGAAAUGCCAUCUUCCUUGAUGUUGG AGGUACCUGCUCUGGCAGAUUUCAACCGGGCUUGGACA GAACUUACCGACUGGCUUUCUCUGCUUGAUCAAGUUAU AAAAUCACAGAGGGUGAUGGUGGGUGACCUUGAGGAUA UCAACGAGAUGAUCAUCAAGCAGAAGGCAACAAUGCAG GAUUUGGAACAGAGGCGUCCCCAGUUGGAAGAACUCAU UACCGCUGCCCAAAAUUUGAAAAACAAGACCAGCAAUCA AGAGGCUAGAACAAUCAUUACGGAUCGAAUUGAAAGAA UUCAGAAUCAGUGGGAUGAAGUACAAGAACACCUUCAGAACCGGAGGCAACAGUUGAAUGAAAUGUUAAAGGAUUCWSGR Ref. No. 54841-719.601AACACAAUGGCUGGAAGCUAAGGAAGAAGCUGAGCAGG UCUUAGGACAGGCCAGAGCCAAGCUUGAGUCAUGGAAG GAGGGUCCCUAUACAGUAGAUGCAAUCCAAAAGAAAAU CACAGAAACCAAGCAGUUGGCCAAAGACCUCCGCCAGUG GCAGACAAAUGUAGAUGUGGCAAAUGACUUGGCCCUGA AACUUCUCCGGGAUUAUUCUGCAGAUGAUACCAGAAAA GUCCACAUGAUAACAGAGAAUAUCAAUGCCUCUUGGAG AAGCAUUCAUAAAAGGGUGAGUGAGCGAGAGGCUGCUU UGGAAGAAACUCAUAGAUUACUGCAACAGUUCCCCCUG GACCUGGAAAAGUUUCUUGCCUGGCUUACAGAAGCUGA AACAACUGCCAAUGUCCUACAGGAUGCUACCCGUAAGGA AAGGCUCCUAGAAGACUCCAAGGGAGUAAAAGAGCUGA UGAAACAAUGGCAAGACCUCCAAGGUGAAAUUGAAGCU CACACAGAUGUUUAUCACAACCUGGAUGAAAACAGCCA AAAAAUCCUGAGAUCCCUGGAAGGUUCCGAUGAUGCAG UCCUGUUACAAAGACGUUUGGAUAACAUGAACUUCAAG UGGAGUGAACUUCGGAAAAAGUCUCUCAACAUUAGGUC CCAUUUGGAAGCCAGUUCUGACCAGUGGAAGCGUCUGC ACCUUUCUCUGCAGGAACUUCUGGUGUGGCUACAGCUG AAAGAUGAUGAAUUAAGCCGGCAGGCACCUAUUGGAGG CGACUUUCCAGCAGUUCAGAAGCAGAACGAUGUACAUA GGGCCUUCAAGAGGGAAUUGAAAACUAAAGAACCUGUA AUCAUGAGUACUCUUGAGACUGUACGAAUAUUUCUGAC AGAGCAGCCUUUGGAAGGACUAGAGAAACUCUACCAGG AGCCCAGAGAGCUGCCUCCUGAGGAGAGAGCCCAGAAUG UCACUCGGCUUCUACGAAAGCAGGCUGAGGAGGUCAAU ACUGAGUGGGAAAAAUUGAACCUGCACUCCGCUGACUG GCAGAGAAAAAUAGAUGAGACCCUUGAAAGACUCCGGG AACUUCAAGAGGCCACGGAUGAGCUGGACCUCAAGCUGC GCCAAGCUGAGGUGAUCAAGGGAUCCUGGCAGCCCGUG GGCGAUCUCCUCAUUGACUCUCUCCAAGAUCACCUCGAG AAAGUCAAGGCACUUCGAGGAGAAAUUGCGCCUCUGAA AGAGAACGUGAGCCACGUCAAUGACCUUGCUCGCCAGCU UACCACUUUGGGCAUUCAGCUCUCACCGUAUAACCUCAG CACUCUGGAAGACCUGAACACCAGAUGGAAGCUUCUGCA GGUGGCCGUCGAGGACCGAGUCAGGCAGCUGCAUGAAG CCCACAGGGACUUUGGUCCAGCAUCUCAGCACUUUCUUU CCACGUCUGUCCAGGGUCCCUGGGAGAGAGCCAUCUCGC CAAACAAAGUGCCCUACUAUAUCAACCACGAGACUCAAA CAACUUGCUGGGACCAUCCCAAAAUGACAGAGCUCUACC AGUCUUUAGCUGACCUGAAUAAUGUCAGAUUCUCAGCU UAUAGGACUGCCAUGAAACUCCGAAGACUGCAGAAGGC CCUUUGCUUGGAUCUCUUGAGCCUGUCAGCUGCAUGUG AUGCCUUGGACCAGCACAACCUCAAGCAAAAUGACCAGC CCAUGGAUAUCCUGCAGAUUAUUAAUUGUUUGACCACU AUUUAUGACCGCCUGGAGCAAGAGCACAACAAUUUGGU CAACGUCCCUCUCUGCGUGGAUAUGUGUCUGAACUGGCU GCUGAAUGUUUAUGAUACGGGACGAACAGGGAGGAUCC GUGUCCUGUCUUUUAAAACUGGCAUCAUUUCCCUGUGUAAAGCACAUUUGGAAGACAAGUACAGAUACCUUUUCAAWSGR Ref. No. 54841-719.601GCAAGUGGCAAGUUCAACAGGAUUUUGUGACCAGCGCA GGCUGGGCCUCCUUCUGCAUGAUUCUAUCCAAAUUCCAA GACAGUUGGGUGAAGUUGCAUCCUUUGGGGGCAGUAAC AUUGAGCCAAGUGUCCGGAGCUGCUUCCAAUUUGCUAA UAAUAAGCCAGAGAUCGAAGCGGCCCUCUUCCUAGACUG GAUGAGACUGGAACCCCAGUCCAUGGUGUGGCUGCCCGU CCUGCACAGAGUGGCUGCUGCAGAAACUGCCAAGCAUCA GGCCAAAUGUAACAUCUGCAAAGAGUGUCCAAUCAUUG GAUUCAGGUACAGGAGUCUAAAGCACUUUAAUUAUGAC AUCUGCCAAAGCUGCUUUUUUUCUGGUCGAGUUGCAAA AGGCCAUAAAAUGCACUAUCCCAUGGUGGAAUAUUGCA CUCCGACUACAUCAGGAGAAGAUGUUCGAGACUUUGCC AAGGUACUAAAAAACAAAUUUCGAACCAAAAGGUAUUU UGCGAAGCAUCCCCGAAUGGGCUACCUGCCAGUGCAGAC UGUCUUAGAGGGGGACAACAUGGAAACUCCCGUUACUC UGAUCAACUUCUGGCCAGUAGAUUCUGCGCCUGCCUCGU CCCCUCAGCUUUCACACGAUGAUACUCAUUCACGCAUUG AACAUUAUGCUAGCAGGCUAGCAGAAAUGGAAAACAGC AAUGGAUCUUAUCUAAAUGAUAGCAUCUCUCCUAAUGA GAGCAUAGAUGAUGAACAUUUGUUAAUCCAGCAUUACU GCCAAAGUUUGAACCAGGACUCCCCCCUGAGCCAGCCUC GUAGUCCUGCCCAGAUCUUGAUUUCCUUAGAGAGUGAG GAAAGAGGGGAGCUAGAGAGAAUCCUAGCAGAUCUUGA GGAAGAAAACAGGAAUCUGCAAGCAGAAUAUGACCGUC UAAAGCAGCAGCACGAACAUAAAGGCCUGUCCCCACUGC CGUCCCCUCCUGAAAUGAUGCCCACCUCUCCCCAGAGUC CCCGGGAUGCUGAGCUCAUUGCUGAGGCCAAGCUACUGC GUCAACACAAAGGCCGCCUGGAAGCCAGGAUGCAAAUCC UGGAAGACCACAAUAAACAGCUGGAGUCACAGUUACAC AGGCUAAGGCAGCUGCUGGAGCAACCCCAGGCAGAGGCC AAAGUGAAUGGCACAACGGUGUCCUCUCCUUCUACCUCU CUACAGAGGUCCGACAGCAGUCAGCCUAUGCUGCUCCGA GUGGUUGGCAGUCAAACUUCGGACUCCAUGGGUGAGGA AGAUCUUCUCAGUCCUCCCCAGGACACAAGCACAGGGUU AGAGGAGGUGAUGGAGCAACUCAACAACUCCUUCCCUA GUUCAAGAGGAAGAAAUACCCCUGGAAAGCCAAUGAGA GAGGACACAAUGuaggaagucuuuuccacauggcagaugauuugggcagag cgauggaguccuuaguaucagucaugacagaugaagaaggagcagaauaaauguuuua caacuccugauucccgcaugguuuuuauaauauucauacaacaaagaggauuagacag uaagaguuuacaagaaauaaaucuauauuuuugugaaggguagugguauuauacugu agauuucaguaguuucuaagucuguuauuguuuuguuaacaauggcagguuuuacac gucuaugcaauuguacaaaaaaguuauaagaaaacuacauguaaaaucuugauagcuaa auaacuugccauuucuuuauauggaacgcauuuuggguuguuuaaaaauuuauaacag uuauaaagaaagauuguaaacuaaagugugcuuuauaaaaaaaaguuguuuauaaaaa ccccuaaaaacaaaacaaacacacacacacacacauacacacacacacacaaaacuuugag gcagcgcauuguuuugcauccuuuuggcgugauauccauaugaaauucauggcuuuu ucuuuuuuugcauauuaaagauaagacuuccucuaccaccacaccaaaugacuacuaca cacugcucauuugagaacugucagcugaguggggcaggcuugaguuuucauuucaua uaucuauaugucuauaaguauauaaauacuauaguuauauagauaaagagauacgaauuucuauagacugacuuuuuccauuuuuuaaauguucaugucacauccuaauagaaagaWSGR Ref. No. 54841-719.601aauuacuucuagucagucauccaggcuuaccugcuuggucuagaauggauuuuucccg gagccggaagccaggaggaaacuacaccacacuaaaacauugucuacagcuccagaugu uucucauuuuaaacaacuuuccacugacaacgaaaguaaaguaaaguauuggauuuuu uuaaagggaacaugugaaugaauacacaggacuuauuauaucagagugaguaaucggu ugguugguugauugauugauugauugauacauucagcuuccugcugcuagcaaugcc acgauuuagauuuaaugaugcuucaguggaaaucaaucagaagguauucugaccuugu gaacaucagaagguauuuuuuaacucccaagcaguagcaggacgaugauagggcugga gggcuauggauucccagcccaucccugugaaggaguaggccacucuuuaagugaagga uuggaugauuguucauaauacauaaaguucucuguaauuacaacuaaauuauuaugcc cucuucucacagucaaaaggaacugggugguuugguuuuuguugcuuuuuuagauuu auugucccaugugggaugaguuuuuaaaugccacaagacauaauuuaaaauaaauaaa cuuugggaaaagguguaaaacaguagccccaucacauuugugauacugacagguauca acccagaagcccaugaacuguguuuccauccuuugcauuucucugcgaguaguuccac acagguuuguaaguaaguaagaaagaaggcaaauugauucaaauguuacaaaaaaaccc uucuugguggauuagacagguuaaauauauaaacaaacaaacaaaaauugcucaaaaaa gaggagaaaagcucaagaggaaaagcuaaggacugguaggaaaaagcuuuacucuuuc augccauuuuauuucuuuuugauuuuuaaaucauucauucaauagauaccaccgugug accuauaauuuugcaaaucuguuaccucugacaucaaguguaauuagcuuuuggagag ugggcugacaucaaguguaauuagcuuuuggagaguggguuuuguccauuauuaaua auuaauuaauuaacaucaaacacggcuucucaugcuauuucuaccucacuuugguuuu gggguguuccugauaauugugcacaccugaguucacagcuucaccacuuguccauugc guuauuuucuuuuuccuuuauaauucuuucuuuuuccuucauaauuuucaaaagaaa acccaaagcucuaagguaacaaauuaccaaauuacaugaagauuugguuuuugucuug cauuuuuuuccuuuaugugacgcuggaccuuuucuuuacccaaggauuuuuaaaacuc agauuuaaaacaagggguuacuuuacauccuacuaagaaguuuaaguaaguaaguuuc auucuaaaaucagagguaaauagagugcauaaauaauuuuguuuuaaucuuuuuguu uuucuuuuagacacauuagcucuggagugagucugucauaauauuugaacaaaaauug agagcuuuauugcugcauuuuaagcauaauuaauuuggacauuauuucguguugugu ucuuuauaaccaccaaguauuaaacuguaaaucauaauguaacugaagcauaaacauca cauggcauguuuugucauuguuuucagguacugaguucuuacuugaguaucauaaua uauuguguuuuaacaccaacacuguaacauuuacgaauuauuuuuuuaaacuucaguu uuacugcauuuucacaacauaucagacuucaccaaauauaugccuuacuauuguauua uaguacugcuuuacuguguaucucaauaaagcacgcaguuauguuacaaaaaa 26 Human cugagaaagacagauugcaaugacugagaugauuuugcuaauuuuuuuuccagccuau DMD, uuccuuaauguacgugauauugugauugauuuucaugcagagaucccugauccuaua transcript guuuuguuugcuauuuauuuuucuccuucacauuuuuuuucuaucaacagagcu^A variant UGAGUGCCAGGAAGCUGCGAAAUCUGUCUUACAAAAAG Dp260-2, GCUGUAAGGAGGCAAAAGUUGCUUGAACAGAGCAUCCA mRNA GUCUGCCCAGGAGACUGAAAAAUCCUUACACUUAAUCCA GGAGUCCCUCACAUUCAUUGACAAGCAGUUGGCAGCUU(Ref. Seq. AUAUUGCAGACAAGGUGGACGCAGCUCAAAUGCCUCAG NM_00401 GAAGCCCAGAAAAUCCAAUCUGAUUUGACAAGUCAUGA 2.4) GAUCAGUUUAGAAGAAAUGAAGAAACAUAAUCAGGGGA AGGAGGCUGCCCAAAGAGUCCUGUCUCAGAUUGAUGUU(CODING GCACAGAAAAAAUUACAAGAUGUCUCCAUGAAGUUUCG SEQUENC AUUAUUCCAGAAACCAGCCAAUUUUGAGCAGCGUCUACE) AAGAAAGUAAGAUGAUUUUAGAUGAAGUGAAGAUGCACUUGCCUGCAUUGGAAACAAAGAGUGUGGAACAGGAAGUAGUACAGUCACAGCUAAAUCAUUGUGUGAACUUGUAUAWSGR Ref. No. 54841-719.601AAAGUCUGAGUGAAGUGAAGUCUGAAGUGGAAAUGGUG AUAAAGACUGGACGUCAGAUUGUACAGAAAAAGCAGAC GGAAAAUCCCAAAGAACUUGAUGAAAGAGUAACAGCUU UGAAAUUGCAUUAUAAUGAGCUGGGAGCAAAGGUAACA GAAAGAAAGCAACAGUUGGAGAAAUGCUUGAAAUUGUC CCGUAAGAUGCGAAAGGAAAUGAAUGUCUUGACAGAAU GGCUGGCAGCUACAGAUAUGGAAUUGACAAAGAGAUCA GCAGUUGAAGGAAUGCCUAGUAAUUUGGAUUCUGAAGU UGCCUGGGGAAAGGCUACUCAAAAAGAGAUUGAGAAAC AGAAGGUGCACCUGAAGAGUAUCACAGAGGUAGGAGAG GCCUUGAAAACAGUUUUGGGCAAGAAGGAGACGUUGGU GGAAGAUAAACUCAGUCUUCUGAAUAGUAACUGGAUAG CUGUCACCUCCCGAGCAGAAGAGUGGUUAAAUCUUUUG UUGGAAUACCAGAAACACAUGGAAACUUUUGACCAGAA UGUGGACCACAUCACAAAGUGGAUCAUUCAGGCUGACA CACUUUUGGAUGAAUCAGAGAAAAAGAAACCCCAGCAA AAAGAAGACGUGCUUAAGCGUUUAAAGGCAGAACUGAA UGACAUACGCCCAAAGGUGGACUCUACACGUGACCAAGC AGCAAACUUGAUGGCAAACCGCGGUGACCACUGCAGGA AAUUAGUAGAGCCCCAAAUCUCAGAGCUCAACCAUCGAU UUGCAGCCAUUUCACACAGAAUUAAGACUGGAAAGGCC UCCAUUCCUUUGAAGGAAUUGGAGCAGUUUAACUCAGA UAUACAAAAAUUGCUUGAACCACUGGAGGCUGAAAUUC AGCAGGGGGUGAAUCUGAAAGAGGAAGACUUCAAUAAA GAUAUGAAUGAAGACAAUGAGGGUACUGUAAAAGAAUU GUUGCAAAGAGGAGACAACUUACAACAAAGAAUCACAG AUGAGAGAAAGCGAGAGGAAAUAAAGAUAAAACAGCAG CUGUUACAGACAAAACAUAAUGCUCUCAAGGAUUUGAG GUCUCAAAGAAGAAAAAAGGCUCUAGAAAUUUCUCAUC AGUGGUAUCAGUACAAGAGGCAGGCUGAUGAUCUCCUG AAAUGCUUGGAUGACAUUGAAAAAAAAUUAGCCAGCCU ACCUGAGCCCAGAGAUGAAAGGAAAAUAAAGGAAAUUG AUCGGGAAUUGCAGAAGAAGAAAGAGGAGCUGAAUGCA GUGCGUAGGCAAGCUGAGGGCUUGUCUGAGGAUGGGGC CGCAAUGGCAGUGGAGCCAACUCAGAUCCAGCUCAGCAA GCGCUGGCGGGAAAUUGAGAGCAAAUUUGCUCAGUUUC GAAGACUCAACUUUGCACAAAUUCACACUGUCCGUGAA GAAACGAUGAUGGUGAUGACUGAAGACAUGCCUUUGGA AAUUUCUUAUGUGCCUUCUACUUAUUUGACUGAAAUCA CUCAUGUCUCACAAGCCCUAUUAGAAGUGGAACAACUUC UCAAUGCUCCUGACCUCUGUGCUAAGGACUUUGAAGAU CUCUUUAAGCAAGAGGAGUCUCUGAAGAAUAUAAAAGA UAGUCUACAACAAAGCUCAGGUCGGAUUGACAUUAUUC AUAGCAAGAAGACAGCAGCAUUGCAAAGUGCAACGCCU GUGGAAAGGGUGAAGCUACAGGAAGCUCUCUCCCAGCU UGAUUUCCAAUGGGAAAAAGUUAACAAAAUGUACAAGG ACCGACAAGGGCGAUUUGACAGAUCUGUUGAGAAAUGG CGGCGUUUUCAUUAUGAUAUAAAGAUAUUUAAUCAGUG GCUAACAGAAGCUGAACAGUUUCUCAGAAAGACACAAAUUCCUGAGAAUUGGGAACAUGCUAAAUACAAAUGGUAUWSGR Ref. No. 54841-719.601CUUAAGGAACUCCAGGAUGGCAUUGGGCAGCGGCAAAC UGUUGUCAGAACAUUGAAUGCAACUGGGGAAGAAAUAA UUCAGCAAUCCUCAAAAACAGAUGCCAGUAUUCUACAG GAAAAAUUGGGAAGCCUGAAUCUGCGGUGGCAGGAGGU CUGCAAACAGCUGUCAGACAGAAAAAAGAGGCUAGAAG AACAAAAGAAUAUCUUGUCAGAAUUUCAAAGAGAUUUA AAUGAAUUUGUUUUAUGGUUGGAGGAAGCAGAUAACAU UGCUAGUAUCCCACUUGAACCUGGAAAAGAGCAGCAAC UAAAAGAAAAGCUUGAGCAAGUCAAGUUACUGGUGGAA GAGUUGCCCCUGCGCCAGGGAAUUCUCAAACAAUUAAA UGAAACUGGAGGACCCGUGCUUGUAAGUGCUCCCAUAA GCCCAGAAGAGCAAGAUAAACUUGAAAAUAAGCUCAAG CAGACAAAUCUCCAGUGGAUAAAGGUUUCCAGAGCUUU ACCUGAGAAACAAGGAGAAAUUGAAGCUCAAAUAAAAG ACCUUGGGCAGCUUGAAAAAAAGCUUGAAGACCUUGAA GAGCAGUUAAAUCAUCUGCUGCUGUGGUUAUCUCCUAU UAGGAAUCAGUUGGAAAUUUAUAACCAACCAAACCAAG AAGGACCAUUUGACGUUAAGGAAACUGAAAUAGCAGUU CAAGCUAAACAACCGGAUGUGGAAGAGAUUUUGUCUAA AGGGCAGCAUUUGUACAAGGAAAAACCAGCCACUCAGCC AGUGAAGAGGAAGUUAGAAGAUCUGAGCUCUGAGUGGA AGGCGGUAAACCGUUUACUUCAAGAGCUGAGGGCAAAG CAGCCUGACCUAGCUCCUGGACUGACCACUAUUGGAGCC UCUCCUACUCAGACUGUUACUCUGGUGACACAACCUGUG GUUACUAAGGAAACUGCCAUCUCCAAACUAGAAAUGCC AUCUUCCUUGAUGUUGGAGGUACCUGCUCUGGCAGAUU UCAACCGGGCUUGGACAGAACUUACCGACUGGCUUUCUC UGCUUGAUCAAGUUAUAAAAUCACAGAGGGUGAUGGUG GGUGACCUUGAGGAUAUCAACGAGAUGAUCAUCAAGCA GAAGGCAACAAUGCAGGAUUUGGAACAGAGGCGUCCCC AGUUGGAAGAACUCAUUACCGCUGCCCAAAAUUUGAAA AACAAGACCAGCAAUCAAGAGGCUAGAACAAUCAUUAC GGAUCGAAUUGAAAGAAUUCAGAAUCAGUGGGAUGAAG UACAAGAACACCUUCAGAACCGGAGGCAACAGUUGAAU GAAAUGUUAAAGGAUUCAACACAAUGGCUGGAAGCUAA GGAAGAAGCUGAGCAGGUCUUAGGACAGGCCAGAGCCA AGCUUGAGUCAUGGAAGGAGGGUCCCUAUACAGUAGAU GCAAUCCAAAAGAAAAUCACAGAAACCAAGCAGUUGGC CAAAGACCUCCGCCAGUGGCAGACAAAUGUAGAUGUGG CAAAUGACUUGGCCCUGAAACUUCUCCGGGAUUAUUCU GCAGAUGAUACCAGAAAAGUCCACAUGAUAACAGAGAA UAUCAAUGCCUCUUGGAGAAGCAUUCAUAAAAGGGUGA GUGAGCGAGAGGCUGCUUUGGAAGAAACUCAUAGAUUA CUGCAACAGUUCCCCCUGGACCUGGAAAAGUUUCUUGCC UGGCUUACAGAAGCUGAAACAACUGCCAAUGUCCUACA GGAUGCUACCCGUAAGGAAAGGCUCCUAGAAGACUCCA AGGGAGUAAAAGAGCUGAUGAAACAAUGGCAAGACCUC CAAGGUGAAAUUGAAGCUCACACAGAUGUUUAUCACAA CCUGGAUGAAAACAGCCAAAAAAUCCUGAGAUCCCUGGAAGGUUCCGAUGAUGCAGUCCUGUUACAAAGACGUUUGWSGR Ref. No. 54841-719.601GAUAACAUGAACUUCAAGUGGAGUGAACUUCGGAAAAA GUCUCUCAACAUUAGGUCCCAUUUGGAAGCCAGUUCUG ACCAGUGGAAGCGUCUGCACCUUUCUCUGCAGGAACUUC UGGUGUGGCUACAGCUGAAAGAUGAUGAAUUAAGCCGG CAGGCACCUAUUGGAGGCGACUUUCCAGCAGUUCAGAA GCAGAACGAUGUACAUAGGGCCUUCAAGAGGGAAUUGA AAACUAAAGAACCUGUAAUCAUGAGUACUCUUGAGACU GUACGAAUAUUUCUGACAGAGCAGCCUUUGGAAGGACU AGAGAAACUCUACCAGGAGCCCAGAGAGCUGCCUCCUGA GGAGAGAGCCCAGAAUGUCACUCGGCUUCUACGAAAGC AGGCUGAGGAGGUCAAUACUGAGUGGGAAAAAUUGAAC CUGCACUCCGCUGACUGGCAGAGAAAAAUAGAUGAGAC CCUUGAAAGACUCCGGGAACUUCAAGAGGCCACGGAUG AGCUGGACCUCAAGCUGCGCCAAGCUGAGGUGAUCAAG GGAUCCUGGCAGCCCGUGGGCGAUCUCCUCAUUGACUCU CUCCAAGAUCACCUCGAGAAAGUCAAGGCACUUCGAGGA GAAAUUGCGCCUCUGAAAGAGAACGUGAGCCACGUCAA UGACCUUGCUCGCCAGCUUACCACUUUGGGCAUUCAGCU CUCACCGUAUAACCUCAGCACUCUGGAAGACCUGAACAC CAGAUGGAAGCUUCUGCAGGUGGCCGUCGAGGACCGAG UCAGGCAGCUGCAUGAAGCCCACAGGGACUUUGGUCCAG CAUCUCAGCACUUUCUUUCCACGUCUGUCCAGGGUCCCU GGGAGAGAGCCAUCUCGCCAAACAAAGUGCCCUACUAUA UCAACCACGAGACUCAAACAACUUGCUGGGACCAUCCCA AAAUGACAGAGCUCUACCAGUCUUUAGCUGACCUGAAU AAUGUCAGAUUCUCAGCUUAUAGGACUGCCAUGAAACU CCGAAGACUGCAGAAGGCCCUUUGCUUGGAUCUCUUGA GCCUGUCAGCUGCAUGUGAUGCCUUGGACCAGCACAACC UCAAGCAAAAUGACCAGCCCAUGGAUAUCCUGCAGAUU AUUAAUUGUUUGACCACUAUUUAUGACCGCCUGGAGCA AGAGCACAACAAUUUGGUCAACGUCCCUCUCUGCGUGGA UAUGUGUCUGAACUGGCUGCUGAAUGUUUAUGAUACGG GACGAACAGGGAGGAUCCGUGUCCUGUCUUUUAAAACU GGCAUCAUUUCCCUGUGUAAAGCACAUUUGGAAGACAA GUACAGAUACCUUUUCAAGCAAGUGGCAAGUUCAACAG GAUUUUGUGACCAGCGCAGGCUGGGCCUCCUUCUGCAUG AUUCUAUCCAAAUUCCAAGACAGUUGGGUGAAGUUGCA UCCUUUGGGGGCAGUAACAUUGAGCCAAGUGUCCGGAG CUGCUUCCAAUUUGCUAAUAAUAAGCCAGAGAUCGAAG CGGCCCUCUUCCUAGACUGGAUGAGACUGGAACCCCAGU CCAUGGUGUGGCUGCCCGUCCUGCACAGAGUGGCUGCUG CAGAAACUGCCAAGCAUCAGGCCAAAUGUAACAUCUGCA AAGAGUGUCCAAUCAUUGGAUUCAGGUACAGGAGUCUA AAGCACUUUAAUUAUGACAUCUGCCAAAGCUGCUUUUU UUCUGGUCGAGUUGCAAAAGGCCAUAAAAUGCACUAUC CCAUGGUGGAAUAUUGCACUCCGACUACAUCAGGAGAA GAUGUUCGAGACUUUGCCAAGGUACUAAAAAACAAAUU UCGAACCAAAAGGUAUUUUGCGAAGCAUCCCCGAAUGG GCUACCUGCCAGUGCAGACUGUCUUAGAGGGGGACAACAUGGAAACUCCCGUUACUCUGAUCAACUUCUGGCCAGUAWSGR Ref. No. 54841-719.601GAUUCUGCGCCUGCCUCGUCCCCUCAGCUUUCACACGAU GAUACUCAUUCACGCAUUGAACAUUAUGCUAGCAGGCU AGCAGAAAUGGAAAACAGCAAUGGAUCUUAUCUAAAUG AUAGCAUCUCUCCUAAUGAGAGCAUAGAUGAUGAACAU UUGUUAAUCCAGCAUUACUGCCAAAGUUUGAACCAGGA CUCCCCCCUGAGCCAGCCUCGUAGUCCUGCCCAGAUCUU GAUUUCCUUAGAGAGUGAGGAAAGAGGGGAGCUAGAGA GAAUCCUAGCAGAUCUUGAGGAAGAAAACAGGAAUCUG CAAGCAGAAUAUGACCGUCUAAAGCAGCAGCACGAACA UAAAGGCCUGUCCCCACUGCCGUCCCCUCCUGAAAUGAU GCCCACCUCUCCCCAGAGUCCCCGGGAUGCUGAGCUCAU UGCUGAGGCCAAGCUACUGCGUCAACACAAAGGCCGCCU GGAAGCCAGGAUGCAAAUCCUGGAAGACCACAAUAAAC AGCUGGAGUCACAGUUACACAGGCUAAGGCAGCUGCUG GAGCAACCCCAGGCAGAGGCCAAAGUGAAUGGCACAACG GUGUCCUCUCCUUCUACCUCUCUACAGAGGUCCGACAGC AGUCAGCCUAUGCUGCUCCGAGUGGUUGGCAGUCAAAC UUCGGACUCCAUGGGUGAGGAAGAUCUUCUCAGUCCUCC CCAGGACACAAGCACAGGGUUAGAGGAGGUGAUGGAGC AACUCAACAACUCCUUCCCUAGUUCAAGAGGAAGAAAU ACCCCUGGAAAGCCAAUGAGAGAGGACACAAUGuaggaagu cuuuuccacauggcagaugauuugggcagagcgauggaguccuuaguaucagucaugg cagaugaagaaggagcagaauaaauguuuuacaacuccugauucccgcaugguuuuua uaauauucauacaacaaagaggauuagacaguaagaguuuacaagaaauaaaucuauau uuuugugaaggguagugguauuauacuguagauuucaguaguuucuaagucuguuau uguuuuguuaacaauggcagguuuuacacgucuaugcaauuguacaaaaaaguuauaa gaaaacuacauguaaaaucuugauagcuaaauaacuugccauuucuuuauauggaacg cauuuuggguuguuuaaaaauuuauaacaguuauaaagaaagauuguaaacuaaagug ugcuuuauaaaaaaaaguuguuuauaaaaaccccuaaaaacaaaacaaacacacacacac acacauacacacacacacacaaaacuuugaggcagcgcauuguuuugcauccuuuuggc gugauauccauaugaaauucauggcuuuuucuuuuuuugcauauuaaagauaagacu uccucuaccaccacaccaaaugacuacuacacacugcucauuugagaacugucagcuga guggggcaggcuugaguuuucauuucauauaucuauaugucuauaaguauauaaaua cuauaguuauauagauaaagagauacgaauuucuauagacugacuuuuuccauuuuuu aaauguucaugucacauccuaauagaaagaaauuacuucuagucagucauccaggcuu accugcuuggucuagaauggauuuuucccggagccggaagccaggaggaaacuacacc acacuaaaacauugucuacagcuccagauguuucucauuuuaaacaacuuuccacugac aacgaaaguaaaguaaaguauuggauuuuuuuaaagggaacaugugaaugaauacaca ggacuuauuauaucagagugaguaaucgguugguugguugauugauugauugauugg uacauucagcuuccugcugcuagcaaugccacgauuuagauuuaaugaugcuucagug gaaaucaaucagaagguauucugaccuugugaacaucagaagguauuuuuuaacuccc aagcaguagcaggacgaugauagggcuggagggcuauggauucccagcccaucccugu gaaggaguaggccacucuuuaagugaaggauuggaugauuguucauaauacauaaagu ucucuguaauuacaacuaaauuauuaugcccucuucucacagucaaaaggaacugggu gguuugguuuuuguugcuuuuuuagauuuauugucccaugugggaugaguuuuuaa augccacaagacauaauuuaaaauaaauaaacuuugggaaaagguguaaaacaguagcc ccaucacauuugugauacugacagguaucaacccagaagcccaugaacuguguuucca uccuuugcauuucucugcgaguaguuccacacagguuuguaaguaaguaagaaagaag gcaaauugauucaaauguuacaaaaaaacccuucuugguggauuagacagguuaaauauauaaacaaacaaacaaaaauugcucaaaaaagaggagaaaagcucaagaggaaaagcuaWSGR Ref. No. 54841 -719.601aggacugguaggaaaaagcuuuacucuuucaugccauuuuauuucuuuuugauuuuu aaaucauucauucaauagauaccaccgugugaccuauaauuuugcaaaucuguuaccu cugacaucaaguguaauuagcuuuuggagagugggcugacaucaaguguaauuagcu uuuggagaguggguuuuguccauuauuaauaauuaauuaauuaacaucaaacacggcu ucucaugcuauuucuaccucacuuugguuuugggguguuccugauaauugugcacac cugaguucacagcuucaccacuuguccauugcguuauuuucuuuuuccuuuauaauuc uuucuuuuuccuucauaauuuucaaaagaaaacccaaagcucuaagguaacaaauuacc aaauuacaugaagauuugguuuuugucuugcauuuuuuuccuuuaugugacgcugga ccuuuucuuuacccaaggauuuuuaaaacucagauuuaaaacaagggguuacuuuaca uccuacuaagaaguuuaaguaaguaaguuucauucuaaaaucagagguaaauagagug cauaaauaauuuuguuuuaaucuuuuuguuuuucuuuuagacacauuagcucuggag ugagucugucauaauauuugaacaaaaauugagagcuuuauugcugcauuuuaagcau aauuaauuuggacauuauuucguguuguguucuuuauaaccaccaaguauuaaacugu aaaucauaauguaacugaagcauaaacaucacauggcauguuuugucauuguuuucag guacugaguucuuacuugaguaucauaauauauuguguuuuaacaccaacacuguaac auuuacgaauuauuuuuuuaaacuucaguuuuacugcauuuucacaacauaucagacu ucaccaaauauaugccuuacuauuguauuauaguacugcuuuacuguguaucucaaua aagcacgcaguuauguuacaaaaaa27 Full mature gucagauccucacucucuuccgcaucgcugucugcgagggccagcuguugggcucgcgguugaggacaa mRNA acucuucgcggucuuuccaguacucuuggaucggaaacccgucggccuccgaacgguacuccgccaccga gggaccugagcgaguccgcaucgaccggaucggaaaaccucucgagaaaggcgucuaaccagucacaguc encoding gca a gguguccacucccagguccaaguuuaaacuuggccgcca ccAUGCUUUGGUGGGAAG human AAGUAGAGGACUGUUAUGAAAGAGAAGAUGUUCAAAAGAAAACAUU Dystrophin CACAAAAUGGGUAAAUGCACAAUUUUCUAAGUUUGGGAAGCAGCAU AUUGAGAACCUCUUCAGUGACCUACAGGAUGGGAGGCGCCUCCUAGRef. Seq. ACCUCCUCGAAGGCCUGACAGGGCAAAAACUGCCAAAAGAAAAAGG AUCCACAAGAGUUCAUGCCCUGAACAAUGUCAACAAGGCACUGCGG NM 00400 GUUUUGCAGAACAAUAAUGUUGAUUUAGUGAAUAUUGGAAGUACUG 6.2 ACAUCGUAGAUGGAAAUCAUAAACUGACUCUUGGUUUGAUUUGGAA UAUAAUCCUCCACUGGCAGGUCAAAAAUGUAAUGAAAAAUAUCAUG(CODING GCUGGAUUGCAACAAACCAACAGUGAAAAGAUUCUCCUGAGCUGGG SEQUENC UCCGACAAUCAACUCGUAAUUAUCCACAGGUUAAUGUAAUCAACUU CACCACCAGCUGGUCUGAUGGCCUGGCUUUGAAUGCUCUCAUCCAUAE) GUCAUAGGCCAGACCUAUUUGACUGGAAUAGUGUGGUUUGCCAGCA GUCAGCCACACAACGACUGGAACAUGCAUUCAACAUCGCCAGAUAUC AAUUAGGCAUAGAGAAACUACUCGAUCCUGAAGAUGUUGAUACCAC CUAUCCAGAUAAGAAGUCCAUCUUAAUGUACAUCACAUCACUCUUC CAAGUUUUGCCUCAACAAGUGAGCAUUGAAGCCAUCCAGGAAGUGG AAAUGUUGCCAAGGCCACCUAAAGUGACUAAAGAAGAACAUUUUCA GUUACAUCAUCAAAUGCACUAUUCUCAACAGAUCACGGUCAGUCUA GCACAGGGAUAUGAGAGAACUUCUUCCCCUAAGCCUCGAUUCAAGA GCUAUGCCUACACACAGGCUGCUUAUGUCACCACCUCUGACCCUACA CGGAGCCCAUUUCCUUCACAGCAUUUGGAAGCUCCUGAAGACAAGU CAUUUGGCAGUUCAUUGAUGGAGAGUGAAGUAAACCUGGACCGUUA UCAAACAGCUUUAGAAGAAGUAUUAUCGUGGCUUCUUUCUGCUGAG GACACAUUGCAAGCACAAGGAGAGAUUUCUAAUGAUGUGGAAGUGG UGAAAGACCAGUUUCAUACUCAUGAGGGGUACAUGAUGGAUUUGAC AGCCCAUCAGGGCCGGGUUGGUAAUAUUCUACAAUUGGGAAGUAAG CUGAUUGGAACAGGAAAAUUAUCAGAAGAUGAAGAAACUGAAGUAC AAGAGCAGAUGAAUCUCCUAAAUUCAAGAUGGGAAUGCCUCAGGGU AGCUAGCAUGGAAAAACAAAGCAAUUUACAUAGAGUUUUAAUGGAU CUCCAGAAUCAGAAACUGAAAGAGUUGAAUGACUGGCUAACAAAAA CAGAAGAAAGAACAAGGAAAAUGGAGGAAGAGCCUCUUGGACCUGA UCUUGAAGACCUAAAACGCCAAGUACAACAACAUAAGGUGCUUCAA GAAGAUCUAGAACAAGAACAAGUCAGGGUCAAUUCUCUCACUCACAUGGUGGUGGUAGUUGAUGAAUCUAGUGGAGAUCACGCAACUGCUGCWSGR Ref. No. 54841-719.601UUUGGAAGAACAACUUAAGGUAUUGGGAGAUCGAUGGGCAAACAUC UGUAGAUGGACAGAAGACCGCUGGGUUCUUUUACAAGACAUCCUUC UCAAAUGGCAACGUCUUACUGAAGAACAGUGCCUUUUUAGUGCAUG GCUUUCAGAAAAAGAAGAUGCAGUGAACAAGAUUCACACAACUGGC UUUAAAGAUCAAAAUGAAAUGUUAUCAAGUCUUCAAAAACUGGCCG UUUUAAAAGCGGAUCUAGAAAAGAAAAAGCAAUCCAUGGGCAAACU GUAUUCACUCAAACAAGAUCUUCUUUCAACACUGAAGAAUAAGUCA GUGACCCAGAAGACGGAAGCAUGGCUGGAUAACUUUGCCCGGUGUU GGGAUAAUUUAGUCCAAAAACUUGAAAAGAGUACAGCACAGAUUUC ACAGGCUGUCACCACCACUCAGCCAUCACUAACACAGACAACUGUAA UGGAAACAGUAACUACGGUGACCACAAGGGAACAGAUCCUGGUAAA GCAUGCUCAAGAGGAACUUCCACCACCACCUCCCCAAAAGAAGAGGC AGAUUACUGUGGAUUCUGAAAUUAGGAAAAGGUUGGAUGUUGAUAU AACUGAACUUCACAGCUGGAUUACUCGCUCAGAAGCUGUGUUGCAG AGUCCUGAAUUUGCAAUCUUUCGGAAGGAAGGCAACUUCUCAGACU UAAAAGAAAAAGUCAAUGCCAUAGAGCGAGAAAAAGCUGAGAAGUU CAGAAAACUGCAAGAUGCCAGCAGAUCAGCUCAGGCCCUGGUGGAA CAGAUGGUGAAUGAGGGUGUUAAUGCAGAUAGCAUCAAACAAGCCU CAGAACAACUGAACAGCCGGUGGAUCGAAUUCUGCCAGUUGCUAAG UGAGAGACUUAACUGGCUGGAGUAUCAGAACAACAUCAUCGCUUUC UAUAAUCAGCUACAACAAUUGGAGCAGAUGACAACUACUGCUGAAA ACUGGUUGAAAAUCCAACCCACCACCCCAUCAGAGCCAACAGCAAUU AAAAGUCAGUUAAAAAUUUGUAAGGAUGAAGUCAACCGGCUAUCAG GUCUUCAACCUCAAAUUGAACGAUUAAAAAUUCAAAGCAUAGCCCU GAAAGAGAAAGGACAAGGACCCAUGUUCCUGGAUGCAGACUUUGUG GCCUUUACAAAUCAUUUUAAGCAAGUCUUUUCUGAUGUGCAGGCCA GAGAGAAAGAGCUACAGACAAUUUUUGACACUUUGCCACCAAUGCG CUAUCAGGAGACCAUGAGUGCCAUCAGGACAUGGGUCCAGCAGUCA GAAACCAAACUCUCCAUACCUCAACUUAGUGUCACCGACUAUGAAA UCAUGGAGCAGAGACUCGGGGAAUUGCAGGCUUUACAAAGUUCUCU GCAAGAGCAACAAAGUGGCCUAUACUAUCUCAGCACCACUGUGAAA GAGAUGUCGAAGAAAGCGCCCUCUGAAAUUAGCCGGAAAUAUCAAU CAGAAUUUGAAGAAAUUGAGGGACGCUGGAAGAAGCUCUCCUCCCA GCUGGUUGAGCAUUGUCAAAAGCUAGAGGAGCAAAUGAAUAAACUC CGAAAAAUUCAGAAUCACAUACAAACCCUGAAGAAAUGGAUGGCUG AAGUUGAUGUUUUUCUGAAGGAGGAAUGGCCUGCCCUUGGGGAUUC AGAAAUUCUAAAAAAGCAGCUGAAACAGUGCAGACUUUUAGUCAGU GAUAUUCAGACAAUUCAGCCCAGUCUAAACAGUGUCAAUGAAGGUG GGCAGAAGAUAAAGAAUGAAGCAGAGCCAGAGUUUGCUUCGAGACU UGAGACAGAACUCAAAGAACUUAACACUCAGUGGGAUCACAUGUGC CAACAGGUCUAUGCCAGAAAGGAGGCCUUGAAGGGAGGUUUGGAGA AAACUGUAAGCCUCCAGAAAGAUCUAUCAGAGAUGCACGAAUGGAU GACACAAGCUGAAGAAGAGUAUCUUGAGAGAGAUUUUGAAUAUAAA ACUCCAGAUGAAUUACAGAAAGCAGUUGAAGAGAUGAAGAGAGCUA AAGAAGAGGCCCAACAAAAAGAAGCGAAAGUGAAACUCCUUACUGA GUCUGUAAAUAGUGUCAUAGCUCAAGCUCCACCUGUAGCACAAGAG GCCUUAAAAAAGGAACUUGAAACUCUAACCACCAACUACCAGUGGC UCUGCACUAGGCUGAAUGGGAAAUGCAAGACUUUGGAAGAAGUUUG GGCAUGUUGGCAUGAGUUAUUGUCAUACUUGGAGAAAGCAAACAAG UGGCUAAAUGAAGUAGAAUUUAAACUUAAAACCACUGAAAACAUUC CUGGCGGAGCUGAGGAAAUCUCUGAGGUGCUAGAUUCACUUGAAAA UUUGAUGCGACAUUCAGAGGAUAACCCAAAUCAGAUUCGCAUAUUG GCACAGACCCUAACAGAUGGCGGAGUCAUGGAUGAGCUAAUCAAUG AGGAACUUGAGACAUUUAAUUCUCGUUGGAGGGAACUACAUGAAGA GGCUGUAAGGAGGCAAAAGUUGCUUGAACAGAGCAUCCAGUCUGCC CAGGAGACUGAAAAAUCCUUACACUUAAUCCAGGAGUCCCUCACAU UCAUUGACAAGCAGUUGGCAGCUUAUAUUGCAGACAAGGUGGACGC AGCUCAAAUGCCUCAGGAAGCCCAGAAAAUCCAAUCUGAUUUGACA AGUCAUGAGAUCAGUUUAGAAGAAAUGAAGAAACAUAAUCAGGGGAAGGAGGCUGCCCAAAGAGUCCUGUCUCAGAUUGAUGUUGCACAGAAWSGR Ref. No. 54841-719.601AAAAUUACAAGAUGUCUCCAUGAAGUUUCGAUUAUUCCAGAAACCA GCCAAUUUUGAGCAGCGUCUACAAGAAAGUAAGAUGAUUUUAGAUG AAGUGAAGAUGCACUUGCCUGCAUUGGAAACAAAGAGUGUGGAACA GGAAGUAGUACAGUCACAGCUAAAUCAUUGUGUGAACUUGUAUAAA AGUCUGAGUGAAGUGAAGUCUGAAGUGGAAAUGGUGAUAAAGACUG GACGUCAGAUUGUACAGAAAAAGCAGACGGAAAAUCCCAAAGAACU UGAUGAAAGAGUAACAGCUUUGAAAUUGCAUUAUAAUGAGCUGGGA GCAAAGGUAACAGAAAGAAAGCAACAGUUGGAGAAAUGCUUGAAAU UGUCCCGUAAGAUGCGAAAGGAAAUGAAUGUCUUGACAGAAUGGCU GGCAGCUACAGAUAUGGAAUUGACAAAGAGAUCAGCAGUUGAAGGA AUGCCUAGUAAUUUGGAUUCUGAAGUUGCCUGGGGAAAGGCUACUC AAAAAGAGAUUGAGAAACAGAAGGUGCACCUGAAGAGUAUCACAGA GGUAGGAGAGGCCUUGAAAACAGUUUUGGGCAAGAAGGAGACGUUG GUGGAAGAUAAACUCAGUCUUCUGAAUAGUAACUGGAUAGCUGUCA CCUCCCGAGCAGAAGAGUGGUUAAAUCUUUUGUUGGAAUACCAGAA ACACAUGGAAACUUUUGACCAGAAUGUGGACCACAUCACAAAGUGG AUCAUUCAGGCUGACACACUUUUGGAUGAAUCAGAGAAAAAGAAAC CCCAGCAAAAAGAAGACGUGCUUAAGCGUUUAAAGGCAGAACUGAA UGACAUACGCCCAAAGGUGGACUCUACACGUGACCAAGCAGCAAAC UUGAUGGCAAACCGCGGUGACCACUGCAGGAAAUUAGUAGAGCCCC AAAUCUCAGAGCUCAACCAUCGAUUUGCAGCCAUUUCACACAGAAU UAAGACUGGAAAGGCCUCCAUUCCUUUGAAGGAAUUGGAGCAGUUU AACUCAGAUAUACAAAAAUUGCUUGAACCACUGGAGGCUGAAAUUC AGCAGGGGGUGAAUCUGAAAGAGGAAGACUUCAAUAAAGAUAUGAA UGAAGACAAUGAGGGUACUGUAAAAGAAUUGUUGCAAAGAGGAGAC AACUUACAACAAAGAAUCACAGAUGAGAGAAAGCGAGAGGAAAUAA AGAUAAAACAGCAGCUGUUACAGACAAAACAUAAUGCUCUCAAGGA UUUGAGGUCUCAAAGAAGAAAAAAGGCUCUAGAAAUUUCUCAUCAG UGGUAUCAGUACAAGAGGCAGGCUGAUGAUCUCCUGAAAUGCUUGG AUGACAUUGAAAAAAAAUUAGCCAGCCUACCUGAGCCCAGAGAUGA AAGGAAAAUAAAGGAAAUUGAUCGGGAAUUGCAGAAGAAGAAAGAG GAGCUGAAUGCAGUGCGUAGGCAAGCUGAGGGCUUGUCUGAGGAUG GGGCCGCAAUGGCAGUGGAGCCAACUCAGAUCCAGCUCAGCAAGCGC UGGCGGGAAAUUGAGAGCAAAUUUGCUCAGUUUCGAAGACUCAACU UUGCACAAAUUCACACUGUCCGUGAAGAAACGAUGAUGGUGAUGAC UGAAGACAUGCCUUUGGAAAUUUCUUAUGUGCCUUCUACUUAUUUG ACUGAAAUCACUCAUGUCUCACAAGCCCUAUUAGAAGUGGAACAAC UUCUCAAUGCUCCUGACCUCUGUGCUAAGGACUUUGAAGAUCUCUU UAAGCAAGAGGAGUCUCUGAAGAAUAUAAAAGAUAGUCUACAACAA AGCUCAGGUCGGAUUGACAUUAUUCAUAGCAAGAAGACAGCAGCAU UGCAAAGUGCAACGCCUGUGGAAAGGGUGAAGCUACAGGAAGCUCU CUCCCAGCUUGAUUUCCAAUGGGAAAAAGUUAACAAAAUGUACAAG GACCGACAAGGGCGAUUUGACAGAUCUGUUGAGAAAUGGCGGCGUU UUCAUUAUGAUAUAAAGAUAUUUAAUCAGUGGCUAACAGAAGCUGA ACAGUUUCUCAGAAAGACACAAAUUCCUGAGAAUUGGGAACAUGCU AAAUACAAAUGGUAUCUUAAGGAACUCCAGGAUGGCAUUGGGCAGC GGCAAACUGUUGUCAGAACAUUGAAUGCAACUGGGGAAGAAAUAAU UCAGCAAUCCUCAAAAACAGAUGCCAGUAUUCUACAGGAAAAAUUG GGAAGCCUGAAUCUGCGGUGGCAGGAGGUCUGCAAACAGCUGUCAG ACAGAAAAAAGAGGCUAGAAGAACAAAAGAAUAUCUUGUCAGAAUU UCAAAGAGAUUUAAAUGAAUUUGUUUUAUGGUUGGAGGAAGCAGAU AACAUUGCUAGUAUCCCACUUGAACCUGGAAAAGAGCAGCAACUAA AAGAAAAGCUUGAGCAAGUCAAGUUACUGGUGGAAGAGUUGCCCCU GCGCCAGGGAAUUCUCAAACAAUUAAAUGAAACUGGAGGACCCGUG CUUGUAAGUGCUCCCAUAAGCCCAGAAGAGCAAGAUAAACUUGAAA AUAAGCUCAAGCAGACAAAUCUCCAGUGGAUAAAGGUUUCCAGAGC UUUACCUGAGAAACAAGGAGAAAUUGAAGCUCAAAUAAAAGACCUU GGGCAGCUUGAAAAAAAGCUUGAAGACCUUGAAGAGCAGUUAAAUC AUCUGCUGCUGUGGUUAUCUCCUAUUAGGAAUCAGUUGGAAAUUUAUAACCAACCAAACCAAGAAGGACCAUUUGACGUUCAGGAAACUGAAWSGR Ref. No. 54841-719.601AUAGCAGUUCAAGCUAAACAACCGGAUGUGGAAGAGAUUUUGUCUA AAGGGCAGCAUUUGUACAAGGAAAAACCAGCCACUCAGCCAGUGAA GAGGAAGUUAGAAGAUCUGAGCUCUGAGUGGAAGGCGGUAAACCGU UUACUUCAAGAGCUGAGGGCAAAGCAGCCUGACCUAGCUCCUGGAC UGACCACUAUUGGAGCCUCUCCUACUCAGACUGUUACUCUGGUGAC ACAACCUGUGGUUACUAAGGAAACUGCCAUCUCCAAACUAGAAAUG CCAUCUUCCUUGAUGUUGGAGGUACCUGCUCUGGCAGAUUUCAACC GGGCUUGGACAGAACUUACCGACUGGCUUUCUCUGCUUGAUCAAGU UAUAAAAUCACAGAGGGUGAUGGUGGGUGACCUUGAGGAUAUCAAC GAGAUGAUCAUCAAGCAGAAGGCAACAAUGCAGGAUUUGGAACAGA GGCGUCCCCAGUUGGAAGAACUCAUUACCGCUGCCCAAAAUUUGAA AAACAAGACCAGCAAUCAAGAGGCUAGAACAAUCAUUACGGAUCGA AUUGAAAGAAUUCAGAAUCAGUGGGAUGAAGUACAAGAACACCUUC AGAACCGGAGGCAACAGUUGAAUGAAAUGUUAAAGGAUUCAACACA AUGGCUGGAAGCUAAGGAAGAAGCUGAGCAGGUCUUAGGACAGGCC AGAGCCAAGCUUGAGUCAUGGAAGGAGGGUCCCUAUACAGUAGAUG CAAUCCAAAAGAAAAUCACAGAAACCAAGCAGUUGGCCAAAGACCU CCGCCAGUGGCAGACAAAUGUAGAUGUGGCAAAUGACUUGGCCCUG AAACUUCUCCGGGAUUAUUCUGCAGAUGAUACCAGAAAAGUCCACA UGAUAACAGAGAAUAUCAAUGCCUCUUGGAGAAGCAUUCAUAAAAG GGUGAGUGAGCGAGAGGCUGCUUUGGAAGAAACUCAUAGAUUACUG CAACAGUUCCCCCUGGACCUGGAAAAGUUUCUUGCCUGGCUUACAG AAGCUGAAACAACUGCCAAUGUCCUACAGGAUGCUACCCGUAAGGA AAGGCUCCUAGAAGACUCCAAGGGAGUAAAAGAGCUGAUGAAACAA UGGCAAGACCUCCAAGGUGAAAUUGAAGCUCACACAGAUGUUUAUC ACAACCUGGAUGAAAACAGCCAAAAAAUCCUGAGAUCCCUGGAAGG UUCCGAUGAUGCAGUCCUGUUACAAAGACGUUUGGAUAACAUGAAC UUCAAGUGGAGUGAACUUCGGAAAAAGUCUCUCAACAUUAGGUCCC AUUUGGAAGCCAGUUCUGACCAGUGGAAGCGUCUGCACCUUUCUCU GCAGGAACUUCUGGUGUGGCUACAGCUGAAAGAUGAUGAAUUAAGC CGGCAGGCACCUAUUGGAGGCGACUUUCCAGCAGUUCAGAAGCAGA ACGAUGUACAUAGGGCCUUCAAGAGGGAAUUGAAAACUAAAGAACC UGUAAUCAUGAGUACUCUUGAGACUGUACGAAUAUUUCUGACAGAG CAGCCUUUGGAAGGACUAGAGAAACUCUACCAGGAGCCCAGAGAGC UGCCUCCUGAGGAGAGAGCCCAGAAUGUCACUCGGCUUCUACGAAA GCAGGCUGAGGAGGUCAAUACUGAGUGGGAAAAAUUGAACCUGCAC UCCGCUGACUGGCAGAGAAAAAUAGAUGAGACCCUUGAAAGACUCC AGGAACUUCAAGAGGCCACGGAUGAGCUGGACCUCAAGCUGCGCCA AGCUGAGGUGAUCAAGGGAUCCUGGCAGCCCGUGGGCGAUCUCCUC AUUGACUCUCUCCAAGAUCACCUCGAGAAAGUCAAGGCACUUCGAG GAGAAAUUGCGCCUCUGAAAGAGAACGUGAGCCACGUCAAUGACCU UGCUCGCCAGCUUACCACUUUGGGCAUUCAGCUCUCACCGUAUAACC UCAGCACUCUGGAAGACCUGAACACCAGAUGGAAGCUUCUGCAGGU GGCCGUCGAGGACCGAGUCAGGCAGCUGCAUGAAGCCCACAGGGAC UUUGGUCCAGCAUCUCAGCACUUUCUUUCCACGUCUGUCCAGGGUCC CUGGGAGAGAGCCAUCUCGCCAAACAAAGUGCCCUACUAUAUCAACC ACGAGACUCAAACAACUUGCUGGGACCAUCCCAAAAUGACAGAGCU CUACCAGUCUUUAGCUGACCUGAAUAAUGUCAGAUUCUCAGCUUAU AGGACUGCCAUGAAACUCCGAAGACUGCAGAAGGCCCUUUGCUUGG AUCUCUUGAGCCUGUCAGCUGCAUGUGAUGCCUUGGACCAGCACAA CCUCAAGCAAAAUGACCAGCCCAUGGAUAUCCUGCAGAUUAUUAAU UGUUUGACCACUAUUUAUGACCGCCUGGAGCAAGAGCACAACAAUU UGGUCAACGUCCCUCUCUGCGUGGAUAUGUGUCUGAACUGGCUGCU GAAUGUUUAUGAUACGGGACGAACAGGGAGGAUCCGUGUCCUGUCU UUUAAAACUGGCAUCAUUUCCCUGUGUAAAGCACAUUUGGAAGACA AGUACAGAUACCUUUUCAAGCAAGUGGCAAGUUCAACAGGAUUUUG UGACCAGCGCAGGCUGGGCCUCCUUCUGCAUGAUUCUAUCCAAAUUC CAAGACAGUUGGGUGAAGUUGCAUCCUUUGGGGGCAGUAACAUUGA GCCAAGUGUCCGGAGCUGCUUCCAAUUUGCUAAUAAUAAGCCAGAGAUCGAAGCGGCCCUCUUCCUAGACUGGAUGAGACUGGAACCCCAGUCWSGR Ref. No. 54841 -719.601CAUGGUGUGGCUGCCCGUCCUGCACAGAGUGGCUGCUGCAGAAACU GCCAAGCAUCAGGCCAAAUGUAACAUCUGCAAAGAGUGUCCAAUCA UUGGAUUCAGGUACAGGAGUCUAAAGCACUUUAAUUAUGACAUCUG CCAAAGCUGCUUUUUUUCUGGUCGAGUUGCAAAAGGCCAUAAAAUG CACUAUCCCAUGGUGGAAUAUUGCACUCCGACUACAUCAGGAGAAG AUGUUCGAGACUUUGCCAAGGUACUAAAAAACAAAUUUCGAACCAA AAGGUAUUUUGCGAAGCAUCCCCGAAUGGGCUACCUGCCAGUGCAG ACUGUCUUAGAGGGGGACAACAUGGAAACUCCCGUUACUCUGAUCA ACUUCUGGCCAGUAGAUUCUGCGCCUGCCUCGUCCCCUCAGCUUUCA CACGAUGAUACUCAUUCACGCAUUGAACAUUAUGCUAGCAGGCUAG CAGAAAUGGAAAACAGCAAUGGAUCUUAUCUAAAUGAUAGCAUCUC UCCUAAUGAGAGCAUAGAUGAUGAACAUUUGUUAAUCCAGCAUUAC UGCCAAAGUUUGAACCAGGACUCCCCCCUGAGCCAGCCUCGUAGUCC UGCCCAGAUCUUGAUUUCCUUAGAGAGUGAGGAAAGAGGGGAGCUA GAGAGAAUCCUAGCAGAUCUUGAGGAAGAAAACAGGAAUCUGCAAG CAGAAUAUGACCGUCUAAAGCAGCAGCACGAACAUAAAGGCCUGUC CCCACUGCCGUCCCCUCCUGAAAUGAUGCCCACCUCUCCCCAGAGUC CCCGGGAUGCUGAGCUCAUUGCUGAGGCCAAGCUACUGCGUCAACAC AAAGGCCGCCUGGAAGCCAGGAUGCAAAUCCUGGAAGACCACAAUA AACAGCUGGAGUCACAGUUACACAGGCUAAGGCAGCUGCUGGAGCA ACCCCAGGCAGAGGCCAAAGUGAAUGGCACAACGGUGUCCUCUCCUU CUACCUCUCUACAGAGGUCCGACAGCAGUCAGCCUAUGCUGCUCCGA GUGGUUGGCAGUCAAACUUCGGACUCCAUGGGUGAGGAAGAUCUUC UCAGUCCUCCCCAGGACACAAGCACAGGGUUAGAGGAGGUGAUGGA GCAACUCAACAACUCCUUCCCUAGUUCAAGAGGAAGAAAUACCCCUG GAAAGCCAAUGAGAGAGGACACAAUGUAGaaucaaccucuggauuacaaaauuug ugaaagauugacugguauucuuaacuauguugcuccuuuuacgcuauguggauacgcugcuuuaaugcc uuuguaucaugcuauugcuucccguauggcuuucauuuucuccuccuuguauaaauccugguugcuggc ucuuuaugaggaguuguggcccguugucaggcaacguggcguggugugcacuguguuugcugacgcaa cccccacugguuggggcauugccaccaccugucagcuccuuuccgggacuuucgcuuucccccucccuau ugccacggcggaacucaucgccgccugccuugcccgcugcuggacaggggcucggcuguugggcacuga caauuccgugguguugucggggaaaucaucguccuuuccuuggcugcucgccuguguugccaccuggau ucugcgcgggacguccuucugcuacgucccuucggcccucaauccagcggaccuuccuucccgcggccug cugccggcucugcggccucuuccgcgucuucgccuucgcccucagacgagucggaucucccuuugggcc gccuccccgccugaucuagagcggccgccgaauucgggcccguuuaaacccgcugaucagccucgacugu gccuucuaguugccagccaucuguuguuugccccucccccgugccuuccuugacccuggaaggugccacu cccacuguccuuuccuaauaaaaugaggaaauugcaucgc28 Full gucagauccucacucucuuccgcaucgcugucugcgagggccagcuguugggcucgcgguugaggacaa Mature acucuucgcggucuuuccaguacucuuggaucggaaacccgucggccuccgaacgguacuccgccaccga gggaccugagcgaguccgcaucgaccggaucggaaaaccucucgagaaaggcgucuaaccagucacaguc mRNA gca a gguguccacucccagguccaaguuuaaacuuggccgcca cc AUGCUUUGGUGGGAAG encoding AAGUAGAGGACUGUUAUGAAAGAGAAGAUGUUCAAAAGAAAACAUU human CACAAAAUGGGUAAAUGCACAAUUUUCUAAGUUUGGGAAGCAGCAU DMD AUUGAGAACCUCUUCAGUGACCUACAGGAUGGGAGGCGCCUCCUAG ACCUCCUCGAAGGCCUGACAGGGCAAAAACUGCCAAAAGAAAAAGG AUCCACAAGAGUUCAUGCCCUGAACAAUGUCAACAAGGCACUGCGGpoly A tail GUUUUGCAGAACAAUAAUGUUGAUUUAGUGAAUAUUGGAAGUACUG A(n) ACAUCGUAGAUGGAAAUCAUAAACUGACUCUUGGUUUGAUUUGGAA UAUAAUCCUCCACUGGCAGGUCAAAAAUGUAAUGAAAAAUAUCAUGRef. Seq. GCUGGAUUGCAACAAACCAACAGUGAAAAGAUUCUCCUGAGCUGGG NM 00400 UCCGACAAUCAACUCGUAAUUAUCCACAGGUUAAUGUAAUCAACUU CACCACCAGCUGGUCUGAUGGCCUGGCUUUGAAUGCUCUCAUCCAUA6.2 GUCAUAGGCCAGACCUAUUUGACUGGAAUAGUGUGGUUUGCCAGCA GUCAGCCACACAACGACUGGAACAUGCAUUCAACAUCGCCAGAUAUC AAUUAGGCAUAGAGAAACUACUCGAUCCUGAAGAUGUUGAUACCAC CUAUCCAGAUAAGAAGUCCAUCUUAAUGUACAUCACAUCACUCUUC CAAGUUUUGCCUCAACAAGUGAGCAUUGAAGCCAUCCAGGAAGUGG AAAUGUUGCCAAGGCCACCUAAAGUGACUAAAGAAGAACAUUUUCA GUUACAUCAUCAAAUGCACUAUUCUCAACAGAUCACGGUCAGUCUAGCACAGGGAUAUGAGAGAACUUCUUCCCCUAAGCCUCGAUUCAAGAWSGR Ref. No. 54841-719.601GCUAUGCCUACACACAGGCUGCUUAUGUCACCACCUCUGACCCUACA CGGAGCCCAUUUCCUUCACAGCAUUUGGAAGCUCCUGAAGACAAGU CAUUUGGCAGUUCAUUGAUGGAGAGUGAAGUAAACCUGGACCGUUA UCAAACAGCUUUAGAAGAAGUAUUAUCGUGGCUUCUUUCUGCUGAG GACACAUUGCAAGCACAAGGAGAGAUUUCUAAUGAUGUGGAAGUGG UGAAAGACCAGUUUCAUACUCAUGAGGGGUACAUGAUGGAUUUGAC AGCCCAUCAGGGCCGGGUUGGUAAUAUUCUACAAUUGGGAAGUAAG CUGAUUGGAACAGGAAAAUUAUCAGAAGAUGAAGAAACUGAAGUAC AAGAGCAGAUGAAUCUCCUAAAUUCAAGAUGGGAAUGCCUCAGGGU AGCUAGCAUGGAAAAACAAAGCAAUUUACAUAGAGUUUUAAUGGAU CUCCAGAAUCAGAAACUGAAAGAGUUGAAUGACUGGCUAACAAAAA CAGAAGAAAGAACAAGGAAAAUGGAGGAAGAGCCUCUUGGACCUGA UCUUGAAGACCUAAAACGCCAAGUACAACAACAUAAGGUGCUUCAA GAAGAUCUAGAACAAGAACAAGUCAGGGUCAAUUCUCUCACUCACA UGGUGGUGGUAGUUGAUGAAUCUAGUGGAGAUCACGCAACUGCUGC UUUGGAAGAACAACUUAAGGUAUUGGGAGAUCGAUGGGCAAACAUC UGUAGAUGGACAGAAGACCGCUGGGUUCUUUUACAAGACAUCCUUC UCAAAUGGCAACGUCUUACUGAAGAACAGUGCCUUUUUAGUGCAUG GCUUUCAGAAAAAGAAGAUGCAGUGAACAAGAUUCACACAACUGGC UUUAAAGAUCAAAAUGAAAUGUUAUCAAGUCUUCAAAAACUGGCCG UUUUAAAAGCGGAUCUAGAAA...

Claims

1. WSGR Ref. No. 54841-719.601CLAIMS WHAT IS CLAIMED IS:

1. An extracellular vesicle comprising:(i) an exogenous nucleic acid molecule encoding a dystrophin protein that comprises an amino acid sequence having at least 70% sequence identity to the amino acid sequence set forth in any one of SEQ ID NOs: 1-6; and(ii) a targeting polypeptide configured to bind to a targeted cell.

2. The extracellular vesicle of claim 1, wherein the nucleic acid comprises a mRNA.

3. The extracellular vesicle of claim 1 or 2, wherein the dystrophin protein comprises an amino acid sequence having at least 70% sequence identity to the amino acid sequence set forth in SEQ ID NO: 1.

4. The extracellular vesicle of any one of claims 1-3, wherein the dystrophin protein is a full-length dystrophin protein.

5. The extracellular vesicle of any one of claims 1-4, wherein the dystrophin protein comprises the amino acid sequence set forth in SEQ ID NO: 1.

6. The extracellular vesicle of any one of claims 1-5, wherein the dystrophin protein consists of the amino acid sequence set forth in SEQ ID NO: 1.

7. The extracellular vesicle of any one of claims 1-6, wherein the nucleic acid comprises a full-length human dystrophin mRNA.

8. The extracellular vesicle of any one of claims 1-7, wherein the targeting polypeptide comprises a targeting domain configured to bind to a surface protein of the targeted cell.

9. The extracellular vesicle of claim 5, wherein the targeting domain comprises a muscle cell-targeting polypeptide.

10. The extracellular vesicle of claim 9, wherein the muscle cell-targeting polypeptide is a skeletal muscle-targeting polypeptide, a cardiac muscle-targeting polypeptide, or a smooth muscle-targeting polypeptide.

11. The extracellular vesicle of claim 10, wherein the muscle cell-targeting polypeptide is the skeletal muscle-targeting polypeptide.

12. The extracellular vesicle of claim 5, wherein the targeting domain is a M12 peptide, an RGD motif, a dystrophin-binding peptide, a T7 peptide, a T9 peptide, a ASSLNIA peptide, a cardiomyocyte-specific peptide (CMP), an ischemic myocardium -targeting polypeptide (IMTP), a cardiac homing peptide (CHP), or a CRPPR peptide.WSGR Ref. No. 54841-719.60113. The extracellular vesicle of claim 12, wherein the targeting domain comprises the M12 peptide, the RGD motif, the dystrophin-binding peptide, the T7 peptide, the T9 peptide, an GPGRGDQTTL peptide or the ASSLNIA peptide.

14. The extracellular vesicle of claim 13, wherein the targeting domain comprises the M12 peptide.

15. The extracellular vesicle of claim 13, wherein the targeting domain comprises the RGD motif.

16. The extracellular vesicle of claim 13, wherein the targeting domain comprises the T7 peptide.

17. The extracellular vesicle of claim 16, wherein the T7 peptide comprises an amino acid sequence having no more than 4 amino acid substitutions with reference to the amino acid sequence set forth in SEQ ID NO: 42.

18. The extracellular vesicle of claim 16, wherein the T7 peptide comprises the amino acid sequence set forth in SEQ ID NO: 42.

19. The extracellular vesicle of claim 16, wherein the T7 peptide consists of the amino acid sequence set forth in SEQ ID NO: 42.

20. The extracellular vesicle of claim 13, wherein the targeting domain comprises the T9 peptide.

21. The extracellular vesicle of claim 20, wherein the T9 peptide comprises an amino acid sequence having no more than 4 amino acid substitutions with reference to the amino acid sequence set forth in SEQ ID NO: 44.

22. The extracellular vesicle of claim 20, wherein the T9 peptide comprises the amino acid sequence set forth in SEQ ID NO: 44.

23. The extracellular vesicle of claim 20, wherein the T9 peptide consists of the amino acid sequence set forth in SEQ ID NO: 44.

24. The extracellular vesicle of any one of claims 8-24, wherein the targeting domain is coupled to an adapter polypeptide.

25. The extracellular vesicle of any one of claims 24, wherein the adapter polypeptide comprises a SIRPa domain.

26. The extracellular vesicle of claim 24, wherein the adapter polypeptide comprises an extracellular vesicle surface protein.

27. The extracellular vesicle of claim 25, wherein the extracellular vesicle surface protein comprises CD47, CD63, CD81, CD82, CD315, a heterotrimeric G protein, MHC class I, an integrin, a transferrin receptor (TFR2), a LAMP1 / 2, a heparan sulfate proteoglycan,WSGR Ref. No. 54841-719.601EMMPRIN, ADAM10, a GPI-anchored 5 'nucleotidase, CD73, CD55, CD59, sonic hedgehog (SHH), TSPAN8, CD37, CD53, CD9, PECAM1, ERBB2, EPCAM, CD90, CD45, CD41, CD42a, Glycophorin A, CD14, MHC class II, CD3, Acetylcholinesterase / AChE-S, AChE-E, amyloid beta A4 / APP, PTGFRN, or multidrug resistance-associated protein.

28. The extracellular vesicle of claim 27, wherein the adapter polypeptide comprises the CD47.

29. The extracellular vesicle of claim 28, wherein the CD47 comprises a human CD47.

30. The extracellular vesicle of claim 28, wherein the human CD47 comprises an amino acid sequence comprising at least 70% sequence identity to the amino acid sequence set forth in SEQ ID NO: 41.

31. The extracellular vesicle of claim 28, wherein the human CD47 comprises an amino acid sequence that comprises the amino acid sequence set forth in SEQ ID NO: 41.

32. The extracellular vesicle of claim 31, wherein the human CD47 consists of the amino acid sequence set forth in SEQ ID NO: 41.

33. The extracellular vesicle of any oneof claims 24-32, wherein the adapter polypeptide is non-covalently coupled to the targeting domain.

34. The extracellular vesicle of any oneof claims 24-32, wherein the adapter polypeptide is covalently coupled to the targeting domain.

35. The extracellular vesicle of claim 34, wherein the targeting domain is covalently coupled to an N-terminus of the adapter polypeptide.

36. The extracellular vesicle of any one of claims 1-9, wherein the targeting polypeptide comprises a T7 peptide covalently coupled to a human CD47 polypeptide.

37. The extracellular vesicle of any one of claims 1-9, wherein the targeting polypeptide comprises a T9 peptide covalently coupled to a human CD47 polypeptide.

38. The extracellular vesicle of any one of claims 1-32, wherein the targeting polypeptide comprises an amino acid sequence having at least 70% sequence identity to the amino acid sequence set forth in SEQ ID NO: 43 or SEQ ID NO: 45.

39. The extracellular vesicle of any one of claims 1-32, wherein the targeting polypeptide comprises the amino acid sequence set forth in SEQ ID NO: 43 or SEQ ID NO:

45.

40. The extracellular vesicle of claim 39, wherein the targeting polypeptide consists of the amino acid sequence set forth in SEQ ID NO: 43 or SEQ ID NO: 45.WSGR Ref. No. 54841-719.60141. The extracellular vesicle of claim 40, further comprising a second targeting polypeptide, wherein the targeting polypeptide consists of the sequence set forth in SEQ ID NO: 43 and the second targeting polypeptide consists of the sequence set forth in SEQ ID NO: 45.

42. The extracellular vesicle of any one of claims 1 to 41, wherein the mRNA comprises a sequence encoded by the sequence set forth in SEQ ID NO: 31.

43. The extracellular vesicle of any one of claims 1 -42, wherein the extracellular vesicle comprises a diameter of greater than 20 nm.

44. The extracellular vesicle of any one of claims 1 -42, wherein the extracellular vesicle comprises a size distribution with a peak at 50 nm to 200 nm in diameter.

45. The extracellular vesicle of any one of claims 1-44, wherein the extracellular vesicle expresses a marker of naturally occurring extracellular vesicles, wherein the marker comprises CD9, CD63, TSG101, or ARF6.

46. The extracellular vesicle of any one of claims 1-45, wherein the extracellular vesicle comprises an exosome, microvesicle, or an apoptotic body.

47. The extracellular vesicle of claim 46, wherein the extracellular vesicle comprises the exosome.

48. The extracellular vesicle of any one of claims 1 -47, wherein the extracellular vesicle comprises a plurality of extracellular vesicles.

49. The extracellular vesicle of claim 48, wherein the extracellular vesicle comprises at least 1x106copies of the nucleic acid molecule.

50. The extracellular vesicle of claim 48, wherein the extracellular vesicle comprises from about 2x106to about 5x1012copies of the nucleic acid molecule.

51. The extracellular vesicle of any one of claims 1 -50, wherein the targeted cell is an adherent cell, a multinucleated cell, a mononucleated cell, a contractile cell, a muscle cell, a skeletal muscle cell, a cardiomyocyte, a smooth muscle cell, a myofibroblast.

52. The extracellular vesicle of claim 51, wherein the targeted cell is the skeletal muscle cell.

53. A composition comprising:(i) a first extracellular vesicle comprising a first targeting domain; and (ii) a second extracellular vesicle comprising a second targeting domain, wherein the first extracellular vesicle and the second extracellular vesicle comprise exogenous messenger RNA (mRNA) encoding a dystrophin protein.

54. The composition of claim 53, wherein the first targeting domain comprisesWSGR Ref. No. 54841-719.601a first muscle-targeting peptide and the second targeting domain comprises a second muscletargeting peptide.

55. The composition of claim 53 or claim 54, wherein:(i) the first muscle-targeting domain is a T7 targeting domain; and (ii) the second muscle-targeting domain is a T9 targeting domain.

56. The composition of claim 55, wherein the T7 targeting domain comprises an amino acid sequence having no more than 4 amino acid substitutions with reference to the amino acid sequence set forth in SEQ ID NO: 42, and wherein the T9 targeting domain comprises an amino acid sequence having no more than 4 amino acid substitutions with reference to the amino acid sequence set forth in SEQ ID NO: 44.

57. The composition of claim 56, wherein the T7 targeting domain comprises the amino acid sequence set forth in SEQ ID NO: 42; and wherein the T9 targeting domain comprises the amino acid sequence set forth in SEQ ID NO: 44.

58. The composition of claim 57, wherein the T7 targeting domain consists of the amino acid sequence set forth in SEQ ID NO: 42; and wherein the T9 targeting domain consists of the amino acid sequence set forth in SEQ ID NO: 44.

59. The composition of any one of claims 53-58, wherein the first targeting domain is coupled to a first adapter polypeptide, and the second targeting domain is coupled to a second adapter polypeptide.

60. The composition of claim 59, wherein the first adapter polypeptide and the second adapter polypeptide comprise a CD47 polypeptide.

61. The composition of claim 60, wherein the CD47 polypeptide comprises a human CD47 polypeptide.

62. The composition of claim 61, wherein the human CD47 polypeptide comprises an amino acid sequence comprising at least 70% sequence identity to the amino acid sequence set forth in SEQ ID NO: 41.

63. The composition of claim 62, wherein the human CD47 polypeptide comprises the amino acid sequence set forth in SEQ ID NO: 41.

64. The composition of claim 63, wherein the human CD47 polypeptide consists of the amino acid sequence set forth in SEQ ID NO: 41.

65. The composition of claim 53, wherein the first extracellular vesicle comprises a first targeting polypeptide, wherein the first targeting polypeptide comprises the first targeting domain and a first adapter polypeptide, and wherein the first targeting polypeptideWSGR Ref. No. 54841-719.601comprises a sequence comprising at least 70% sequence identity to the amino acid sequence set forth in SEQ ID NO: 43.

66. The composition of claim 65, wherein the first targeting polypeptide comprises an amino acid sequence consisting of the amino acid sequence set forth in SEQ ID NO:

43.

67. The composition of any one of claims 53-66, wherein the second extracellular vesicle comprises a second targeting polypeptide, wherein the second targeting polypeptide comprises the second targeting domain and a second adapter polypeptide, wherein the second targeting polypeptide comprises an amino acid sequence that has at least 70% sequence identity to the amino acid sequence set forth in SEQ ID NO: 45.

68. The composition of claim 67, wherein the second targeting polypeptide comprises an amino acid sequence consisting of the amino acid sequence set forth in SEQ ID NO: 45.

69. The composition of any one of claims 53-68, wherein the first extracellular vesicle and the second extracellular vesicle comprise an exogenous nucleic acid molecule encoding a dystrophin protein.

70. The composition of claim 69, wherein the dystrophin protein comprises an amino acid sequence having at least 70% sequence identity to the amino acid sequence set forth in any one of SEQ ID NOs: 1-6.

71. The composition of claim 70, wherein the dystrophin protein is a full-length dystrophin protein.

72. The composition of claim 71, wherein the dystrophin protein comprises the amino acid sequence set forth in any one of SEQ ID NOs: 1-6.

73. The composition of claim 72, wherein the dystrophin protein consists of the amino acid sequence set forth in any one of SEQ ID NOs: 1-6.

74. The composition of claim 73, wherein the dystrophin protein consists of the amino acid sequence set forth in SEQ ID NO: 1.

75. The composition of claim 74, wherein the exogenous nucleic acid molecule comprises an mRNA, wherein the mRNA comprises a nucleic acid sequence encoded by the sequence set forth in SEQ ID NO: 31.

76. The composition of any one of claims 53-75, wherein the first muscle-targeting domain the second muscle-targeting domain are configured to deliver the first extracellular vesicle and the second extracellular vesicle to a first target cell and a second target cell, respectively.

77. The composition of claim 76, wherein the first target cell and the second target cell are the same cell.WSGR Ref. No. 54841-719.60178. The composition of claim 76, wherein the first target cell and the second target cell are different cells.

79. The composition of any one of claims 53-78, wherein the first extracellular vesicle and the second extracellular vesicle comprise an exosome, microvesicle, or an apoptotic body.

80. The composition of claim 79, wherein the first extracellular vesicle and the second extracellular vesicle comprise the exosome.

81. The composition of any one of claims 53-80, further comprising a third extracellular vesicle, wherein the third extracellular vesicle comprisesa third targeting polypeptide consisting of the amino acid sequence of SEQ ID NO: 43; anda fourth targeting polypeptide consisting of the amino acid sequence of SEQ ID NO: 45,and a second exogenous nucleic acid molecule encoding a dystrophin protein consisting of the amino acid sequence set forth in SEQ ID NO: 1.

82. The composition of claim 81, wherein the second exogenous nucleic acid molecule comprises an mRNA, and wherein the mRNA comprises a sequence encoded by the sequence set forth in SEQ ID NO: 31.

83. The composition of any one of claims 53-82, comprising at least IxlO3extracellular vesicles.

84. The composition of claim 83, wherein the composition comprises at least 1x106copies of the exogenous nucleic acid molecule.

85. The composition of claim 83, wherein the composition comprises from about 2x106to about 5x1012copies of the exogenous nucleic acid molecule.

86. The composition of any one of claims Error! Reference source not found.-84, wherein t he composition is formulated for intravenous injection, systemic injection, parenteral injection, intracardiac, or intramuscular injection.

87. An extracellular vesicle comprising:a heterologous adapter polypeptide; andan exogenous nucleic acid molecule encoding a dystrophin protein that comprises an amino acid sequence having at least 70% sequence identity to the amino acid sequence set forth in any one of SEQ ID NOs: 1-6.

88. The extracellular vesicle of claim 87, wherein the heterologous adapter polypeptide comprises an extracellular vesicle surface protein.WSGR Ref. No. 54841-719.60189. The extracellular vesicle of claim 88 wherein the extracellular vesicle surface protein comprises CD47 polypeptide.

90. The extracellular vesicle of claim 89, wherein the CD47 polypeptide comprises a human CD47 polypeptide.

91. The extracellular vesicle of claim 90, wherein the human CD47 polypeptide comprises an amino acid sequence comprising at least 70% sequence identity to the amino acid sequence set forth in SEQ ID NO: 41.

92. The extracellular vesicle of claim 91, wherein the human CD47 polypeptide comprises an amino sequence consisting of the amino acid sequence set forth in SEQ ID NO:

41.

93. The extracellular vesicle of anyone of claims 87-92, wherein the dystrophin protein comprises an amino acid sequence having at least 70% sequence identity to the amino acid sequence set forth in SEQ ID NO: 1.

94. The extracellular vesicle of claim 93, wherein the dystrophin protein comprises the amino acid sequence set forth SEQ ID NO: 1.

95. The extracellular vesicle of claim 94, wherein the dystrophin protein comprises an amino acid sequence consisting of the amino acid sequence set forth in SEQ ID NO: 1.

96. The extracellular vesicle of anyone of claims 87-95, wherein the nucleic acid molecule comprises an exogenous messenger RNA (mRNA).

97. The extracellular vesicle of any one of claims 87-96, wherein the extracellular vesicle comprises an exosome, microvesicle, or an apoptotic body.

98. The extracellular vesicle of claim 97, wherein the extracellular vesicle comprises the exosome.

99. The extracellular vesicle of any one of claims 87-98, wherein the extracellular vesicle comprises at least 1x103extracellular vesicles.

100. The extracellular vesicle of claim anyone of claims 87-99, wherein the extracellular vesicle does not comprise a heterologous targeting domain.

101. A pharmaceutical composition comprising the extracellular vesicle of any one of claims 1-52 or 87-100, and a pharmaceutically acceptable excipient.

102. The pharmaceutical composition of claim 101, wherein the exogenous nucleic acid molecule is present in the pharmaceutical composition at a concentration of at least 1×107copies / mL.

103. The pharmaceutical composition of claim 102, wherein the exogenous nucleic acid molecule is present in the pharmaceutical composition at a concentration of from about 1×107copies / mL to about IxlO14copies / mL.WSGR Ref. No. 54841-719.601104. The pharmaceutical composition of claim 102, wherein the exogenous nucleic acid molecule is present in the pharmaceutical composition at a concentration of from about 1x108 copies / mL to about 1×1012copies / mL.

105. The pharmaceutical composition of claim 102, wherein the exogenous nucleic acid molecule is present in the pharmaceutical composition at a concentration of from about 1x109copies / mL to about 1x1010copies / mL.

106. The pharmaceutical composition of claim 102, wherein the exogenous nucleic acid molecule is present in the pharmaceutical composition at a concentration is about 1x109copies / mL.

107. A pharmaceutical composition comprising the composition of any one of claims 53- 86 and a pharmaceutically acceptable excipient.

108. The pharmaceutical composition of any one of claims 101-107, wherein the pharmaceutical composition is a liquid solution.

109. The pharmaceutical composition of any one of claims 101-107, wherein the pharmaceutically acceptable excipient comprises a saline solution or a buffered saline solution.

110. The pharmaceutical composition of any one of claims 101-109, comprising a pH of at least 5.5 when measured at room temperature.

111. The pharmaceutical composition of claim 110, wherein the pH of the pharmaceutical composition is from about 6.5 to about 8.5 when measured at room temperature.

112. The pharmaceutical composition of claim 110, wherein the pH of the pharmaceutical composition is about 7.7 when measured at room temperature.

113. The pharmaceutical composition of any one of claims 101-112, further comprising human serum albumin.

114. The pharmaceutical composition of claim 113, wherein the human serum albumin is present at a concentration at least 0.1% w / v.

115. The pharmaceutical composition of claim 113, wherein the human serum albumin is present at a concentration of from about 0.1% w / v to about 2% w / v.

116. The pharmaceutical composition of claim 113, wherein the human serum albumin is present at a concentration of about 0.2% w / v.

117. The pharmaceutical composition of claim 113, wherein the human serum albumin is present at a concentration of about 1 % w / v.

118. The pharmaceutical composition of any one of claims 101-117, further comprising a cryoprotection medium.WSGR Ref. No. 54841-719.601119. The pharmaceutical composition of claim 118, wherein the cryoprotection medium is CryoStor® CS10, glycerol, ethylene glycol, propylene glycol, trehalose, sucrose, Dimethyl sulfoxide (DMSO), polyvinylpyrrolidone (PVP), and hydroxyethyl starch (HES), mFreSR™, NB-KUL 10, STEM-CELLBANKER, CryoSOfree™, PRIME-XV FreezIS, Optibumin® 25, or XT-Thrive®.

120. The pharmaceutical composition of claim 118 or claim 119, wherein the cryoprotection medium is present in the pharmaceutical composition at a concentration of at least 2% v / v.

121. The pharmaceutical composition of claim 118 or claim 119, wherein the cryoprotection medium is present in the pharmaceutical composition at a concentration of from about 2% v / v to about 30% v / v.

122. The pharmaceutical composition of claim 118 or claim 119, wherein the cryoprotection medium is present in the pharmaceutical composition at a concentration of from about 3% v / v to about 10% v / v.

123. The pharmaceutical composition of claim 118 or claim 119, wherein the cryoprotection medium is present in the pharmaceutical composition at a concentration of from about 3% v / v to about 7% v / v.

124. The pharmaceutical composition of claim 118 or claim 119, wherein the cryoprotection medium is present in the pharmaceutical composition at a concentration of from about 4% v / v to about 6% v / v.

125. The pharmaceutical composition of claim 118 or claim 119, wherein the cryoprotection medium is present in the pharmaceutical composition at a concentration is about 5% v / v.

126. The pharmaceutical composition of claim 118 or claim 119, wherein the cryoprotection medium is present in the pharmaceutical composition at a concentration of from about 10% v / v to about 30% v / v.

127. The pharmaceutical composition of claim 118 or claim 119, wherein the cryoprotection medium is present in the pharmaceutical composition at a concentration of from about 20% v / v to about 30% v / v.

128. The pharmaceutical composition of claim 118 or claim 119, wherein the cryoprotection medium is present in the pharmaceutical composition at a concentration is about 25% v / v.WSGR Ref. No. 54841-719.601129. The pharmaceutical composition of any one of claims 101-128, formulated for injection via a systemic route, an intravenous route, a parenteral route, an intramuscular route, a subcutaneous route, or an intraperitoneal route, intracardiac route.

130. The pharmaceutical composition of any one of claims 101-128, formulated for injection via a coronary artery catheter.

131. A catheter comprising the extracellular vesicle of any one of claims 1-52 or 87-100, the composition of any one of claims 53-86, or the pharmaceutical composition of any one of claims 101-129.

132. The catheter of claim 131, wherein the catheter comprises a heart catheter.

133. A syringe comprising the extracellular vesicle of any one of claims 1-52 or 87-100, the composition of any one of claims 53-86, or the pharmaceutical composition of any one of claims 101-129.

134. A kit, comprising:(a) the extracellular vesicle of anyone of claims 1-52 or 87-100, the composition of any one of claims 53-86, or the pharmaceutical composition of any one of claims 101-129;(b) a catheter or a syringe; and(c) instructions.

135. A method of increasing an expression level of human dystrophin in a subject, the method comprising:administering to the subject the extracellular vesicle any one of claims 1-52 or 87- 100, the composition of any one of claims 53-86, or the pharmaceutical composition of any one of claims 101-129, wherein the administering results in an elevated expression level of the human dystrophin in the human subject.

136. The method of claim 135, wherein the administering comprises administering to a cardiac tissue of the subject.

137. The method of claim 136, wherein the cardiac tissue is a pericardium, an artery, a heart ventricle, a heart atrium, or a myocardium of the subject.

138. The method of any one of claims 136 or claim 137, wherein the administering comprises administering into the cardiac tissue of the subject via an injection139. The method of any one of claims 138, wherein the injection comprises injections into the cardiac tissue of the subject via an intracardiac injection, an intra-arterial injection, an intrapericardial injection, or an intramyo cardial injection.WSGR Ref. No. 54841-719.601140. The method of any one of claims 136-138, wherein the administering comprises administering into the cardiac tissue of the subject via a catheter.

141. The method of claim 140, wherein the catheter comprises a balloon catheter, a microvalve catheter, a guide catheter, or a stent.

142. The method of claim 135, wherein the administering comprises administering to the subject via an intravenous route, a parenteral route, an intramuscular route.

143. The method of any one of claims 135-142, wherein the method does not comprise administering to the subject an immunosuppressive agent.

144. The method of any one of claims 135-142, wherein the method comprises administering to the subject an immunosuppressive agent.

145. The method of claim 144, wherein the immunosuppressive agent comprises a mTOR inhibitor.

146. The method of claim 145, wherein the mTOR inhibitor comprises sirolimus.

147. The method of claim 145, wherein the immunosuppressive agent comprises a calcineurin inhibitor.

148. The method of claim 146, wherein the calcineurin inhibitor comprises tacrolimus or cyclosporine.

149. The method of any one of claims 135-148, wherein the administering treats a disorder of the subject.

150. The method of claim 149, wherein the disorder comprises a muscle disorder.

151. The method of claim 150, wherein the muscle disorder comprises a muscular dystrophy.

152. The method of claim 151, wherein the muscular dystrophy comprises Duchenne muscular dystrophy or Becker muscular dystrophy.

153. The method of claim 152, wherein the disorder comprises the Duchenne muscular dystrophy.

154. The method of any one of claims 135-153, wherein the subject is a human subject.

155. The method of any one of claims 135-154, wherein the method results in at least a 2- fold increase in the expression level of the human dystrophin protein in the subject compared to an expression level of human dystrophin protein in the subject prior to the administering.

156. The method of any one of claims 135-155, wherein the method results in at least a 2- fold increase in a level of human dystrophin mRNA in the subject compared to an expression level of human dystrophin mRNA in the subject prior to the administering.WSGR Ref. No. 54841-719.601157. The method of any one of claims 135-155, wherein the method results in at least a 2- fold increase in a level of full-length human dystrophin mRNA in the subject compared to an expression level of human dystrophin mRNA in the subject prior to the administering.

158. The method of any one of claims 135-155, wherein the method results in at least a 2- fold increase in a level of full-length human dystrophin protein in the subject compared to an expression level of full-length human dystrophin protein in the subject prior to administration of the extracellular vesicle, the extracellular vesicle, or the pharmaceutical composition.

159. The method of any one of claims 155-158, wherein the method results in an increase in the level of human dystrophin or the level of full-length human dystrophin in a muscle tissue of the subject.

160. The method of claim 159, wherein the muscle tissue is a skeletal muscle of the subject.

161. The method of claim 160, wherein the skeletal muscle tissue is a diaphragm, a quadricep, a bicep, a tricep, a gastrocnemius, a tibialis, a fibularis, a deltoid, a gluteus maximus, or an abdominal muscle of the subject.

162. The method of any one of claims 135-161, wherein the method results in a reduction of a level of N-terminal pro-B-type natriuretic peptide (NT-proBNP) in the subject compared to the level of NT-proBNP in the subject prior to the administering.

163. The method of claim 162, wherein the reduction of the level comprises at least a 20% reduction in the level of NT-proBNP in the subject compared to the level of NT-proBNP in the subject prior to the administering.

164. The method of claim 162 or claim 163, wherein the level is of a blood sample.

165. The method of any one of claims 135 to 164, wherein the method results in a reduction of a level of Creatine Kinase (CK) in the subject compared to the level of CK in the subject prior to the administering.

166. The method of claim 165, wherein the method results in at least a 10% reduction in the level of CK in the subject compared to the level of CK in the subject prior to the administering.

167. The method of claim 165 or claim 166, wherein the method results in at least a 20% reduction in the level of CK in a blood sample obtained from the subject compared to the level of CK in the subject prior to administration of the extracellular vesicle, the extracellular vesicle, or the pharmaceutical composition.WSGR Ref. No. 54841-719.601168. The method of any one of claims 135-167, wherein the method results in at least a 5% increase in muscle strength of the subject compared to a control subject.

169. The method of any one of claims 135-167, wherein the method results in at least a 50% increase in muscle strength of the subject within 7 days of the administering.

170. The method of any one of claims 135-167, wherein the method results in at least a 2- fold increase in grip strength of the subject within 14 days of the administering.

171. The method of any one of claims 135-167, wherein the method results in at least a 10% increase in grip strength of the subject within about 14 days of the administering.

172. The method of any one of claims 135-167, wherein the method results in at least a 70% increase in grip strength of the subject within 14 days of the administering.

173. The method of any one of claims 135-172, wherein the method results in at least a 10% increase in a number of muscle cells in a sample obtained from the subject compared to a similar sample obtained from a control subject.

174. The method of any one of claims 135-173, wherein the method results in an increased localization of dystrophin protein to a cell membrane of a muscle cell of the subject compared to a level of localization of dystrophin protein to a cell membrane of a muscle cell of the control subject.

175. The method of any one of claims 135-174, wherein the method results in a longer lifespan of the subject compared to a lifespan of a control subject who has the tissue dystrophy.

176. The method of any one of claims 135-174, wherein the method results in the subject have a lifespan that is at least 10% longer compared to a lifespan of a control subject.

177. The method of any one of claims 135-176, wherein the method results in the subject having a time to rise from a laying down position to a standing position of less than about 10 seconds.

178. The method of any one of claims 135-176, wherein the method results in the subject having a time to rise from a laying down position to a standing position of less than about 10 seconds since the administering.

179. The method of any one of claims 135-176, wherein the method results in the subject exhibiting at least a 10% decrease in a time to rise compared to a time to rise of the subject before the administering.

180. The method of any one of claims 135-179, wherein the method results in the subject being measured to have a North Star Ambulatory Assessment (NSAA) score of at least 20 compared to a NSAA score measured of the subject before the administering.WSGR Ref. No. 54841-719.601181. The method of anyone of claims 135-179, wherein the method results in the subject exhibiting at least a 10% increase in a NSAA score compared to a NSAA score measured of the subject prior to the administering.

182. The method of any one of claims 135-181, wherein the method results in the subject being measured to walk a ten-meter distance at a speed greater than a speed of the subject prior to the administering.

183. The method of any one of claims 135-181, wherein the method results in the subject being measured to walk a distance over a time period of ten minutes that is at least 10% compared to a distance walked by the subject over the ten -minute time period prior to the administering.

184. The method of any one of claims 135-183, wherein, subsequent to the administering, the subject can walk up at least four steps of a stairway in no more than about 6 seconds.

185. The method of any one of claims 135-183, wherein, subsequent to the administering, the subject walks up at least four steps of a stairway over a time period that is reduced by at least 10% compared to a similar time period over which the subject was able to walk up the at least four steps of the stairway prior to the administering.

186. The method of any one of claims 135-183, wherein the method results in the subject having a four-stair climb time of less than about 10 seconds since the administering.

187. The method of any one of claims 135-186, wherein the method further comprises measuring a level of the human full-length dystrophin in a sample of the subject.

188. The method of claim 187, wherein the measuring comprises quantifying a level of human full-length dystrophin mRNA of at least 2.6 x105copies / mL in the sample obtained from the subject, or processed therefrom, after the administering.

189. The method of claim 187 or claim 188, wherein the measuring comprises quantifying a level of human full-length dystrophin mRNA of at least 2.6 x105copies / mL in the sample obtained from the subject, or processed therefrom, after the administering.

190. The method of any one of claims 187-189, wherein the measuring comprises quantifying a level of human full-length dystrophin protein in the sample obtained from the subject, or processed therefrom, of at least 0.4% of a level of full-length dystrophin protein measured in a healthy control subject, or a control subject who does not have or is not suspected of having a muscle disorder, a muscular dystrophy, or Duchenne Muscular Dystrophy.

191. The method of any one of claims 135-190, wherein the subject has a level of human full-length dystrophin mRNA of at least 2.6 x105copies / mL after the administering.WSGR Ref. No. 54841-719.601192. The method of any one of claims 135-191, wherein the elevated expression level of the human full-length dystrophin is detected by western blot, mass spectrometry, histology, immunohistochemistry, reverse transcription polymerase chain reaction (RT-PCR), RNA sequencing, or northern blot.

193. The method of any one of claims 135-192, wherein the administering further comprises administering at least one dose of the extracellular vesicle, or the pharmaceutical composition to the subject.

194. The method of any one of claims 135-192, wherein the administering further comprises administering at least one dose of the extracellular vesicle or the pharmaceutical composition to the subject.

195. The method of any one of claims 135-194, wherein the administering further comprises administering to the subject a second dose of the extracellular vesicle, or the pharmaceutical composition after at least a period of at least one hour after the administering.

196. The method of any one of claims 135-195, wherein the administering further comprises administering the extracellular vesicle or the pharmaceutical composition to the subject at least once per week.

197. The method of any one of claims 135-195, wherein the administering further comprises administering the extracellular vesicle or the pharmaceutical composition to the subject at least twice per week.

198. The method of any one of claims 135-195, wherein the administering further comprises administering the extracellular vesicle or the pharmaceutical composition to the subject at least once per day.

199. The method of any one of claims 135-195, wherein the administering further comprises administering the extracellular vesicle or the pharmaceutical composition to the subject at least once per month.

200. The method of any one of claims 135-195, wherein the administering further comprises administering at least 4 doses of the extracellular vesicle or the pharmaceutical composition to the subject over a period of at least 2 weeks.

201. The method of any one of claims 135-195, wherein the administering further comprises administering at least 8 doses of the extracellular vesicle or the pharmaceutical composition to the subject over a period of at least one month.WSGR Ref. No. 54841-719.601202. The method of any one of claims 135-201, wherein the administering comprises administering the extracellular vesicle or the pharmaceutical composition to a tissue of the subject.

203. The method of claim 202, wherein the tissue is a skeletal muscle.

204. The method of claim 202, wherein the tissue is a quadricep, a tibialis anterior, a gastrocnemius, a diaphragm, a heart, a cardiac ventricle, a cardiac atrium, or a muscle of a face, a shoulder, an arm, a chest, a back, calves, or a lower extremity of the subject.

205. The method of any one of claims 173-204, further comprising obtaining a tissue biopsy from the subject.

206. The method of claim 205, wherein the tissue biopsy comprises a muscle tissue biopsy.

207. The method of any one of claims 173-204, further comprising obtaining a blood sample from the subject.

208. The method of any one of claims 173-207, further comprising obtaining a sample from the subject after a period of at least one hour after the administering.

209. A method of producing the extracellular vesicle of any one of claims 1-52 or 87-100 comprising:introducing a vector encoding the exogenous nucleic acid molecule into an extracellular vesicle donor cell via transfection;culturing the extracellular vesicle donor cell after the introducing for a sufficient amount of time in a culture medium for the production of the extracellular vesicle; andcollecting the extracellular vesicle from the culture medium.

210. The method of claim 209, wherein the introducing comprises electroporating the vector into the extracellular vesicle donor cell.

211. The method of claim 209, wherein the introducing comprises electroporating a second vector into the extracellular vesicle donor cell.

212. The method of claim 211, wherein the second vector comprises a polynucleotide sequence encoding the targeting polypeptide.

213. The method of any one of claims 209-212, wherein the introducing comprises applying an electric field across a plurality of vessels configured for culturing the extracellular vesicle donor cell.

214. The method of any one of claims 209-213, wherein the culturing further comprises replenishing the cell culture medium in contact with the extracellular vesicle donor cell.WSGR Ref. No. 54841-719.601215. The method of any one of claims 209-214, wherein the culturing further comprises culturing the extracellular vesicle donor cell for one or more cell culture passages.

216. The method of any one of claims 209-215, wherein the method further comprises collecting the extracellular vesicle from the culture medium.

217. The method of any one of claims 209-217, wherein the method further comprises isolating the extracellular vesicle, thereby generating an isolated extracellular vesicle.

218. The method of claim 217, wherein the method further comprises concentrating the isolated extracellular vesicle, thereby generating isolated and concentrated extracellular vesicle.

219. The method of claim 218, wherein the method further comprises cryopreserving the isolated and concentrated extracellular vesicle.

220. The method of claim 219, wherein the method further comprises resuspending the isolated and concentrated extracellular vesicle.

221. The method of any one of claims 209-220, wherein the extracellular vesicle donor cell is a primary cell or a cell obtained from a cell line.

222. The method of any one of claims 209-221, wherein the extracellular vesicle donor cell is a somatic cell.

223. The method of any one of claims 209-222, wherein the extracellular vesicle donor cell is a fibroblast.

224. The method of claim 223, wherein the extracellular vesicle donor cell is a dermal fibroblast.

225. The method of any one of claims 209-221, wherein the extracellular vesicle donor cell is a stem cell.

226. The method of claim 225, wherein the stem cell is a mesenchymal stem cell.

227. The method of any one of claims 209-226, wherein the extracellular vesicle donor cell is a human cell.

228. A protein comprising at least 97% sequence identity to the sequence set forth in SEQ ID NO: 43.

229. The protein of claim 228, comprising the sequence set forth in SEQ ID NO:

43.

230. The protein of claim 229, consisting of the sequence set forth in SEQ ID NO: 43.

231. A protein comprising at least 97% sequence identity to the sequence set forth in SEQ ID NO: 45.

232. The protein of claim 231, comprising the sequence set forth in SEQ ID NO:

45.

233. The protein of claim 232, consisting of the sequence set forth in SEQ ID NO: 45.WSGR Ref. No. 54841-719.601234. A polynucleotide encoding the protein of any one of claims 228-233.

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