Recombinant aav1, aav5, and aav6 capsid mutants and uses thereof

Abstract

Provided herein are modified recombinant adeno-associated virus (rAAV) capsid proteins, such as modified rAAV1, rAAV5, and rAAV6 capsid proteins, rAAV particles comprising such capsid proteins, nucleic acid molecules encoding such capsid proteins, as well as compositions, kits and methods of use thereof.

1 . A modified adeno-associated virus (AAV) capsid protein, wherein a VP3 region of the modified AAV capsid protein comprises a replacement of tyrosine residues with non-tyrosine residues and/or a replacement of a threonine residue with a non-threonine residue at positions corresponding to:
 Y705, Y731, and T492 of a wild-type AAV1 capsid protein having the sequence of SEQ ID NO: 1,   Y436, Y693, and Y719 of a wild-type AAV5 capsid protein having the sequence of SEQ ID NO: 2, or   Y705, Y731, and T492 of a wild-type AAV6 capsid protein having the sequence of SEQ ID NO: 3.   
     
     
         2 . The modified AAV capsid protein of  claim 1 , wherein the modified AAV capsid protein is a modified AAV1 capsid protein and the modified AAV1 capsid protein comprises replacement of tyrosine residues with non-tyrosine residues and a replacement of a threonine residue with a non-threonine residue at each of the positions corresponding to Y705, Y731, and T492 of the wild-type AAV1 capsid protein having the sequence of SEQ ID NO: 1. 
     
     
         3 . (canceled) 
     
     
         4 . The modified AAV capsid protein of  claim 1 , wherein the modified AAV capsid protein is a modified AAV6 capsid protein and the modified AAV6 capsid protein comprises replacement of tyrosine residues with non-tyrosine residues and a replacement of a threonine residue with a non-threonine residue at each of the positions corresponding to Y705, Y731, and T492 of a wild-type AAV6 capsid protein having the sequence of SEQ ID NO: 3. 
     
     
         5 - 7 . (canceled) 
     
     
         8 . A recombinant AAV (rAAV) particle comprising a modified capsid protein of  claim 1 , optionally further comprising a nucleic acid segment encoding a gene of interest, optionally wherein the gene of interest is a therapeutic agent. 
     
     
         9 . (canceled) 
     
     
         10 . A method, comprising:
 contacting a host cell with an rAAV particle comprising the modified AAV capsid protein of  claim 1  or a composition comprising the rAAV particle and a pharmaceutically-acceptable carrier,   wherein the modified capsid protein comprises replacement of tyrosine residues with non-tyrosine residues and a replacement of a threonine residue with a non-threonine residue at each of the positions corresponding to Y705, Y731, and T492 of the wild-type AAV1 capsid protein or of the wild-type AAV6 capsid protein,   optionally wherein the modified AAV capsid protein is a modified AAV1 capsid protein or a modified AAV6 capsid protein.   
     
     
         11 - 17 . (canceled) 
     
     
         18 . A method of treating, preventing, or ameliorating one or more symptoms of a disease, a dysfunction, or a deficiency in a subject, the method comprising administering to a subject in need thereof the rAAV particle of  claim 8 , in an amount and for a time sufficient to prevent, treat, or ameliorate the one or more symptoms of the disease, dysfunction, or deficiency in the subject. 
     
     
         19 . The method of  claim 18 , wherein the disease, dysfunction, or deficiency is cancer, diabetes, autoimmune disease, articular disease, arthritis, renal disease or dysfunction, pancreatic disease, intestinal disease, hepatic disease or dysfunction, biliary disease or dysfunction, neurological disease, neuromuscular disorder, neuromotor deficit, neuroskeletal impairment, neurological disability, neurosensory dysfunction, muscular dystrophy, stroke, Batten disease, ischemia, an eating disorder, Huntington's disease, Parkinson's disease, Tay-Sach's disease, amyotrophic lateral sclerosis, multiple sclerosis, memory loss, neuropathy, palsy, skeletal disease, pulmonary disease, cystic fibrosis, muscular dystrophy, alpha-1-antitrypsin deficiency, breast cancer, prostate cancer, pancreatic cancer, Alzheimer's disease, sickle cell disease, beta-thalassemia, metabolic disease, hypotension, hypertension, atherosclerosis, hypercholesterolemia, vascular damage, vascular disease, or cardiovascular disease. 
     
     
         20 . A method comprising contacting a host cell with the rAAV particle of  claim 8 , optionally wherein the host cell is a blood cell, a blood progenitor cell, a hematopoietic stem cell, a dendritic cell, a monocyte, a monocyte-derived dendritic cell, a blood differentiated cell, a vascular cell, an endothelial cell, an epithelial cell, an eye cell, a retinal cell, a lung cell, an airway epithelial cell, a bone cell, a muscle cell, a heart cell, a liver cell, a pancreas cell, an intestinal cell, a brain cell, a microglial cell, a neuron or neural cell, an astrocyte, a cardiac or tumor cell, or a fibroblast, optionally wherein the host cell is a mammalian host cell or a human host cell. 
     
     
         21 . A method of treating or preventing a disease or disorder in a subject in need thereof, comprising administering to the subject an effective amount of a recombinant AAV (rAAV) particle or a composition thereof,
 wherein the rAAV particle comprises a modified AAV capsid protein comprising an amino acid substitution at one or more positions corresponding to Y705, Y731, and T492 of a wild-type AAV1 capsid protein having the sequence of SEQ ID NO: 1 or a wild-type AAV6 capsid protein having the sequence of SEQ ID NO: 3,   and wherein the disease or disorder is a neural disease, a musculoskeletal disease, or a neuromuscular disease.   
     
     
         22 . The method of  claim 21 , wherein the neural disease, musculoskeletal disease, or neuromuscular disease is selected from the group consisting of neurological deficit dysfunction, Alzheimer's disease, Huntington's disease, Tay-Sach's disease, Parkinson's disease, memory loss, trauma, motor impairment, neuropathy, arthritis, muscular dystrophy, amyotrophic lateral sclerosis (ALS), multiple sclerosis (MS), and palsy. 
     
     
         23 . The method of  claim 21 , wherein the modified AAV capsid protein comprises amino acid substitutions at each of the positions corresponding to Y705, Y731, and T492 of the wild-type AAV1 capsid protein or the wild-type AAV6 capsid protein, optionally wherein the amino acid substitutions correspond to Y705F, Y731F, and T492V. 
     
     
         24 . The method of  claim 21 , wherein the AAV capsid is of serotype AAV1, AAV2, AAV3, AAV4, AAV5, AAV6, AAV7, AAV8, AAV9, AAV10, AAV11, AAV12, or AAV13. 
     
     
         25 . The method of  claim 21 , wherein the rAAV particle comprises a nucleic acid segment encoding a therapeutic agent. 
     
     
         26 . A method of transducing a cell of interest, comprising contacting the cell with a recombinant AAV (rAAV) particle or a composition thereof,
 wherein the rAAV particle comprises a modified AAV capsid protein comprising an amino acid substitution at one or more positions corresponding to Y705, Y731, and T492 of a wild-type AAV1 capsid protein having the sequence of SEQ ID NO: 1 or a wild-type AAV6 capsid protein having the sequence of SEQ ID NO: 3,   and wherein the cell is selected from the group consisting of a hematopoietic stem cell, a dendritic cell, a monocyte, an airway epithelial cell, a retinal cell, a muscle cell, a liver cell, a pancreas cell, a neuron, a microglial cell, and an astrocyte.   
     
     
         27 . The method of  claim 26 , wherein the cell is selected from the group consisting of a neuron, a microglial cell, and an astrocyte. 
     
     
         28 . The method of  claim 26 , wherein the cell is a neuron. 
     
     
         29 . The method of  claim 26 , wherein the cell is a muscle cell. 
     
     
         30 . The method of  claim 26 , wherein the modified AAV capsid protein comprises amino acid substitutions at each of the positions corresponding to Y705, Y731, and T492 of the wild-type AAV1 capsid protein or the wild-type AAV6 capsid protein, optionally wherein the amino acid substitutions correspond to Y705F, Y731F, and T492V. 
     
     
         31 . The method of  claim 26 , wherein the AAV capsid is of serotype AAV1, AAV2, AAV3, AAV4, AAV5, AAV6, AAV7, AAV8, AAV9, AAV10, AAV11, AAV12, or AAV13. 
     
     
         32 . The method of  claim 26 , wherein the rAAV particle comprises a nucleic acid segment encoding a therapeutic agent.