SBIR-STTR Award

Gene therapeutics offer hope to many patients with rare muscle and neuromuscular diseases. Despite some early successes, several serious off-target safety concerns have compromised their development due to hepatic toxicities and related immune responses to the adeno-associated viral (AAV) vectors. The generation of novel capsids with superior muscle specificity could, therefore, revolutionize the muscle gene therapy space by avoiding the off-target effects that compromise drug efficacy and safety. Our objective is to engineer novel AAV capsids with muscle tropism that exceeds the current "muscle tropic" serotypes (e.g. AAV6, AAV8, AAV9, AAVrh74 & MyoAAVs) as none is actually "muscle-specific". Indeed, all of these serotypes can transduce other tissues, especially the liver, which is functionally linked to the noted clinical toxicities. In fact, the liver functions as a sink for these vectors, limiting muscle transduction and elevating the minimally effective dose. We hypothesize that de-targeting the liver while simultaneously enhancing muscle tropism is key to improving muscle gene therapy safety and efficacy. Other groups have sought to enhance muscle tropism using directed evolution. This high throughput method artificially selects capsids with improved muscle tropism, but cannot also de-target the liver. By contrast, we will use a rational design approach to simultaneously target known epitopes for liver de-targeting, enhanced AAV-receptor binding and improved muscle targeting. These include those for improved sialic acid/AAV-receptor binding, impaired liver targeting/heparin sulfate binding, improved integrin binding and capsids with combined properties. We will also use AAV6 as liver- and muscle-targeting epitopes are known for this serotype, but not for the other serotypes. Milestone 1 will develop liver de-targeted/muscle targeted AAV6 capsids using well-established in vivo and in vitro imaging approaches. Milestone 2 will demonstrate functional efficacy by comparing a Smad7 muscle gene therapeutic featuring a wild-type AAV6 (AVGN7) to one with a novel optimized AAV6 capsid. These studies are understandably high risk yet their significance is disproportionately much higher as they will create a liver de- targeted capsid with improved muscle tropism and as a result, vastly superior safety and efficacy profiles. This would substantially innovate the muscle gene therapy space primarily by reducing a manufacturing burden that limits drug use to younger or fewer patients and thus, reducing the overall treatment costs while expanding the number of potential patient therapies.

Public Health Relevance Statement:
RELEVANCE TO PUBLIC HEALTH. The loss of striated muscle mass and function in patients with a rare neuromuscular disease significantly compromises quality of life and is often fatal. The proposed studies will develop novel adeno-associated viral capsids for improving the efficacy and safety of muscle-focused gene therapeutics. Ultimately, these capsids will be incorporated into a gene therapy program for inclusion body myositis and Duchenne muscular dystrophy and will be licensed to other commercial programs targeting these and other neuromuscular disease indications.

Project Terms:
21+ years old; Adult Human; adulthood; Adult; ages; Age; Antigenic Determinants; Binding Determinants; Epitopes; Award; Complementary DNA; cDNA; Capsid; Clinical Trials; Codon Nucleotides; Codon; Diagnosis; Disease; Disorder; Pharmaceutical Preparations; Drugs; Medication; Pharmaceutic Preparations; drug/agent; Duchenne muscular dystrophy; Duchene; Duchenne; Duchenne-Griesinger syndrome; Ellis-van Creveld (EvC) syndrome; Pseudohypertrophic Muscular Dystrophy; X-linked dilated cardiomyopathy; X-linked muscular dystrophy; X-linked recessive muscular dystrophy; benign X-linked recessive muscular dystrophy; childhood pseudohypertrophic muscular dystrophy; classic X-linked recessive muscular dystrophy; mild X-linked recessive muscular dystrophy; progressive muscular dystrophy of childhood; pseudohypertrophic adult muscular dystrophy; pseudohypertrophic muscular paralysis; Engineering; gene therapy; DNA Therapy; Gene Transfer Clinical; Genetic Intervention; gene repair therapy; gene-based therapy; genetic therapy; genomic therapy; Goals; Heparin; Heparinic Acid; In Vitro; Liver; hepatic body system; hepatic organ system; Methods; Muscle; Muscle Tissue; muscular; Muscular Atrophy; Muscle Atrophy; muscle breakdown; muscle degradation; muscle deterioration; muscle loss; muscle wasting; Spinal Muscular Atrophy; Aran-Duchenne disease; Cruveilhier disease; Myopathy; Muscle Disease; Muscle Disorders; Muscular Diseases; Myopathic Conditions; Myopathic Diseases and Syndromes; Myopathic disease or syndrome; muscular disorder; Neuromuscular Diseases; myoneural disorder; neuromuscular degenerative disorder; neuromuscular disorder; Legal patent; Patents; Patients; Program Development; Public Health; Quality of life; QOL; Virus Receptors; Viral Receptor; Risk; Safety; Serotyping; Sialic Acids; N-Acetylneuraminic Acids; Specificity; Testing; Drug or chemical Tissue Distribution; Tissue Distribution; Tissues; Body Tissues; Wasting Syndrome; Wasting Disease; wasting condition; wasting disorder; Generations; Businesses; promotor; promoter; Treatment Cost; animal breeding; improved; Chronic; Clinical; Link; Muscle function; liver function; Hepatotoxic effect; Liver Toxicity; Toxic effect on liver cells; hepatic toxicity; hepatoxicity; Hepatotoxicity; inclusion-body myosytis; Inclusion Body Myositis; muscle bulk; muscle mass; muscle form; Licensing; drug use; Drug usage; Voluntary Muscle; Skeletal Muscle; Tropism; Collaborations; Intellectual Property; directed evolution; Directed Molecular Evolution; Immunological response; host response; immune system response; immunoresponse; Immune response; programs; Sulfate; Viral; Penetrance; receptor binding; receptor bound; success; drug efficacy; trait; Toxic effect; Toxicities; Smad7 protein; Mothers Against DPP Homolog 7 Protein; Mothers Against Decapentaplegic Homolog 7 Protein; Mothers Against Decapentaplegic, Drosophila Homolog 7 Protein; Mothers Against Decapentaplegic, Drosophila, Homolog 7 Protein; SMA- and MAD-Related Protein 7; hSMAD7; novel; Agreement; Limb-Girdle Muscular Dystrophies; limb-girdle muscular weakness and atrophy; limb-girdle syndrome; myopathic limb-girdle syndrome; Property; drug development; integrin bound; Integrin Binding; Molecular Interaction; Binding; Effectiveness; preventing; prevent; Striated Muscles; Dose; Dose Limiting; Recombinants; in vivo; Development; developmental; Cytokeratin-8 Staining Method; CK8; pre-clinical; preclinical; neuromuscular; vector; elderly patient; older patient; innovate; innovative; innovation; Impairment; gene therapeutics; gene-based therapeutic; gene-based therapeutics; genes therapeutic; genes therapeutics; therapeutic gene; AAV vector; adeno-associated virus vector; adeno-associated viral vector; therapeutic agent development; therapeutic development; high risk; comparable efficacy; compare efficacy; comparative efficacy; imaging based approach; imaging approach; drug safety; pharmaceutical safety; medication safety; rational design; manufacture; manufacturing cost; fabrication cost

Gene therapeutics offer hope to many patients with rare muscle and neuromuscular diseases. Despite some early successes, several serious off-target safety concerns have compromised their development due to hepatic toxicities and related immune responses to the adeno-associated viral (AAV) vectors. The generation of novel capsids with superior muscle specificity could, therefore, revolutionize the muscle gene therapy space by avoiding the off-target effects that compromise drug efficacy and safety. Our objective is to engineer novel AAV capsids with muscle tropism that exceeds the current "muscle tropic" serotypes (e.g. AAV6, AAV8, AAV9, AAVrh74 & MyoAAVs) as none is actually "muscle-specific". Indeed, all of these serotypes can transduce other tissues, especially the liver, which is functionally linked to the noted clinical toxicities. In fact, the liver functions as a sink for these vectors, limiting muscle transduction and elevating the minimally effective dose. We hypothesize that de-targeting the liver while simultaneously enhancing muscle tropism is key to improving muscle gene therapy safety and efficacy. Other groups have sought to enhance muscle tropism using directed evolution. This high throughput method artificially selects capsids with improved muscle tropism, but cannot also de-target the liver. By contrast, we will use a rational design approach to simultaneously target known epitopes for liver de-targeting, enhanced AAV-receptor binding and improved muscle targeting. These include those for improved sialic acid/AAV-receptor binding, impaired liver targeting/heparin sulfate binding, improved integrin binding and capsids with combined properties. We will also use AAV6 as liver- and muscle-targeting epitopes are known for this serotype, but not for the other serotypes. Milestone 1 will develop liver de-targeted/muscle targeted AAV6 capsids using well-established in vivo and in vitro imaging approaches. Milestone 2 will demonstrate functional efficacy by comparing a Smad7 muscle gene therapeutic featuring a wild-type AAV6 (AVGN7) to one with a novel optimized AAV6 capsid. These studies are understandably high risk yet their significance is disproportionately much higher as they will create a liver de- targeted capsid with improved muscle tropism and as a result, vastly superior safety and efficacy profiles. This would substantially innovate the muscle gene therapy space primarily by reducing a manufacturing burden that limits drug use to younger or fewer patients and thus, reducing the overall treatment costs while expanding the number of potential patient therapies.

Public Health Relevance Statement:
RELEVANCE TO PUBLIC HEALTH. The loss of striated muscle mass and function in patients with a rare neuromuscular disease significantly compromises quality of life and is often fatal. The proposed studies will develop novel adeno-associated viral capsids for improving the efficacy and safety of muscle-focused gene therapeutics. Ultimately, these capsids will be incorporated into a gene therapy program for inclusion body myositis and Duchenne muscular dystrophy and will be licensed to other commercial programs targeting these and other neuromuscular disease indications. Terms: <21+ years old; AAV vector; AAV-based vector; Adult; Adult Human; Age; Agreement; Antigenic Determinants; Aran-Duchenne disease; Award; Binding; Binding Determinants; Body Tissues; Businesses; CK8; Capsid; Chronic; Clinical; Clinical Trials; Codon; Codon Nucleotides; Collaborations; Complementary DNA; Cruveilhier disease; Cytokeratin-8 Staining Method; DNA Therapy; Development; Diagnosis; Directed Molecular Evolution; Disease; Disorder; Dose; Dose Limiting; Drug or chemical Tissue Distribution; Drug usage; Drugs; Duchene; Duchenne; Duchenne muscular dystrophy; Duchenne-Griesinger syndrome; Effectiveness; Ellis-van Creveld (EvC) syndrome; Engineering; Epitopes; Gene Transfer Clinical; Generations; Genetic Intervention; Goals; Heparin; Heparinic Acid; Hepatotoxic effect; Hepatotoxicity; Immune response; Immunological response; Impairment; In Vitro; Inclusion Body Myositis; Integrin Binding; Intellectual Property; Legal patent; Licensing; Limb-Girdle Muscular Dystrophies; Link; Liver; Liver Toxicity; Medication; Methods; Molecular Interaction; Mothers Against DPP Homolog 7 Protein; Mothers Against Decapentaplegic Homolog 7 Protein; Mothers Against Decapentaplegic, Drosophila Homolog 7 Protein; Mothers Against Decapentaplegic, Drosophila, Homolog 7 Protein; Muscle; Muscle Atrophy; Muscle Disease; Muscle Disorders; Muscle Tissue; Muscle function; Muscular Atrophy; Muscular Diseases; Myopathic Conditions; Myopathic Diseases and Syndromes; Myopathic disease or syndrome; Myopathy; N-Acetylneuraminic Acids; Neuromuscular Diseases; Patents; Patients; Penetrance; Pharmaceutical Preparations; Program Development; Property; Pseudohypertrophic Muscular Dystrophy; Public Health; QOL; Quality of life; Recombinants; Risk; SMA- and MAD-Related Protein 7; Safety; Serotyping; Sialic Acids; Skeletal Muscle; Smad7 protein; Specificity; Spinal Muscular Atrophy; Striated Muscles; Sulfate; Testing; Tissue Distribution; Tissues; Toxic effect; Toxic effect on liver cells; Toxicities; Treatment Cost; Tropism; Viral; Viral Receptor; Virus Receptors; Voluntary Muscle; Wasting Disease; Wasting Syndrome; X-linked dilated cardiomyopathy; X-linked muscular dystrophy; X-linked recessive muscular dystrophy; adeno-associated viral vector; adeno-associated virus vector; adulthood; ages; animal breeding; benign X-linked recessive muscular dystrophy; cDNA; childhood pseudohypertrophic muscular dystrophy; classic X-linked recessive muscular dystrophy; comparable efficacy; comparative efficacy; compare efficacy; developmental; directed evolution; drug development; drug efficacy; drug safety; drug use; drug/agent; elderly patient; fabrication cost; gene repair therapy; gene therapeutics; gene therapy; gene-based therapeutic; gene-based therapeutics; gene-based therapy; genes therapeutic; genes therapeutics; genetic therapy; genomic therapy; hSMAD7; hepatic body system; hepatic organ system; hepatic toxicity; hepatoxicity; high risk; host response; imaging approach; imaging based approach; immune system response; immunoresponse; improved; in vivo; inclusion-body myosytis; innovate; innovation; innovative; integrin bound; limb-girdle muscular weakness and atrophy; limb-girdle syndrome; liver function; manufacture; manufacturing cost; medication safety; mild X-linked recessive muscular dystrophy; muscle breakdown; muscle bulk; muscle degradation; muscle deterioration; muscle form; muscle loss; muscle mass; muscle wasting; muscular; muscular disorder; myoneural disorder; myopathic limb-girdle syndrome; neuromuscular; neuromuscular degenerative disorder; neuromuscular disorder; novel; older patient; pharmaceutical safety; pre-clinical; preclinical; prevent; preventing; programs; progressive muscular dystrophy of childhood; promoter; promotor; pseudohypertrophic adult muscular dystrophy; pseudohypertrophic muscular paralysis; rational design; receptor binding; receptor bound; success; therapeutic agent development; therapeutic development; therapeutic gene; trait; vector; wasting condition; wasting disorder