Aav-mediated delivery of therapeutic antibodies to the inner ear

Abstract

Provided herein are methods that include introducing into an inner ear of a mammal a therapeutically effective amount of an adeno-associated virus (AAV) vector that includes a nucleotide sequence encoding (a) a polypeptide including an antibody heavy chain variable domain operably linked to a signal peptide and a polypeptide including an antibody light chain variable domain operably linked to a signal peptide; (b) a polypeptide including an antigen-binding antibody fragment operably linked to a signal peptide; or (c) a soluble vascular endothelial growth factor receptor operably linked to a signal peptide.

What is claimed is: 
     
         1 . A method comprising introducing into an inner ear of a mammal a therapeutically effective amount of an adeno-associated virus (AAV) vector that comprises a nucleotide sequence encoding
 (a) a polypeptide comprising an antibody heavy chain variable domain operably linked to a signal peptide and a polypeptide comprising an antibody light chain variable domain operably linked to a signal peptide; or   (b) a polypeptide comprising an antigen-binding antibody fragment operably linked to a signal peptide.   
     
     
         2 . A method for increasing the level of an antibody or an antigen-binding antibody fragment in an inner ear of a mammal in need thereof, the method comprising:
 introducing into the inner ear of the mammal a therapeutically effective amount of an AAV vector that comprises a nucleotide sequence encoding   (a) a polypeptide comprising an antibody heavy chain variable domain operably linked to a signal peptide and a polypeptide comprising an antibody light chain variable domain operably linked to a signal peptide; or   (b) a polypeptide comprising an antigen-binding antibody fragment linked to a signal peptide;   wherein the introducing results in an increase in the level of the antibody or the antigen-binding antibody fragment in the inner ear of the mammal.   
     
     
         3 . The method of  claim 1  or  2 , wherein the antibody or the antigen-binding antibody fragment binds specifically to vascular endothelial growth factor (VEGF). 
     
     
         4 . The method of  claim 3 , wherein the antibody or antigen-binding antibody fragment decreases VEGF activity. 
     
     
         5 . The method of any one of  claims 1 - 4 , wherein the AAV vector further comprises one or both of a promoter and a Kozak sequence that are operably linked to the sequence encoding the antibody or the antigen-binding antibody fragment. 
     
     
         6 . The method of  claim 5 , wherein the AAV vector comprises a promoter selected from the group consisting of: an inducible promoter, a constitutive promoter, and a tissue-specific promoter. 
     
     
         7 . The method of any one of  claims 1 - 6 , wherein the AAV vector further comprises a polyadenylation signal sequence. 
     
     
         8 . The method of any one of  claims 1 - 7 , wherein the mammal is a human. 
     
     
         9 . The method of any one of  claims 1 - 8 , wherein the mammal has been identified as having an inner ear disorder. 
     
     
         10 . The method of any one of  claims 1 - 8 , wherein the mammal has been diagnosed as having an inner ear disorder. 
     
     
         11 . The method of any one of  claims 1 - 10 , wherein the AAV vector comprises a nucleic acid sequence encoding a polypeptide comprising an antibody heavy chain variable domain operably linked to a signal peptide and a polypeptide comprising an antibody light chain variable domain operably linked to a signal peptide. 
     
     
         12 . The method of any one of  claims 1 - 10 , wherein the AAV vector comprises a nucleic acid sequence encoding a polypeptide comprising an antigen-binding antibody fragment operably linked to a signal. 
     
     
         13 . A method for treating an inner ear disorder in a mammal in need thereof, the method comprising:
 introducing into the inner ear of the mammal a therapeutically effective amount of an AAV vector that comprises a nucleotide sequence encoding:   (a) a polypeptide comprising an antibody heavy chain variable domain operably linked to a signal peptide and a polypeptide comprising an antibody light chain variable domain operably linked to a signal peptide; or   (b) a polypeptide comprising an antigen-binding antibody fragment linked to a signal peptide;   wherein the introducing results in the treatment of the inner ear disorder in the mammal.   
     
     
         14 . The method of  claim 13 , wherein the AAV vector further comprises one or both of a promoter and a Kozak sequence that are operably linked to the sequence encoding the antibody or the antigen-binding antibody fragment. 
     
     
         15 . The method of  claim 14 , wherein the AAV vector comprises a promoter selected from the group consisting of: an inducible promoter, a constitutive promoter, and a tissue-specific promoter. 
     
     
         16 . The method of any one of  claims 13 - 15 , wherein the AAV vector further comprises a polyadenylation signal sequence. 
     
     
         17 . The method of any one of  claims 13 - 16 , wherein the mammal is a human. 
     
     
         18 . The method of any one of  claims 13 - 17 , wherein the mammal has been identified as having an inner ear disorder. 
     
     
         19 . The method of any one of  claims 13 - 17 , wherein the mammal has been diagnosed as having an inner ear disorder. 
     
     
         20 . The method of any one of  claims 13 - 19 , wherein the AAV vector comprises a nucleic acid sequence encoding a polypeptide comprising an antibody heavy chain variable domain operably linked to a signal peptide and a polypeptide comprising an antibody light chain variable domain operably linked to a signal peptide. 
     
     
         21 . The method of any one of  claims 13 - 19 , wherein the AAV vector comprises a nucleic acid sequence encoding a polypeptide comprising an antigen-binding antibody fragment operably linked to a signal. 
     
     
         22 . A method of reducing VEGF activity in an inner ear of a mammal in need thereof, the method comprising:
 introducing into the inner ear of the mammal a therapeutically effective amount of an AAV vector that comprises a nucleotide sequence encoding   (a) a polypeptide comprising an antibody heavy chain variable domain operably linked to a signal peptide and a polypeptide comprising an antibody light chain variable domain operably linked to a signal peptide; or   (b) a polypeptide comprising an antigen-binding antibody fragment linked to a signal peptide;   wherein the polypeptide of (a) encodes an antibody that binds specifically to VEGF and reduces VEGF activity, the polypeptide of (b) encodes an antigen-binding antibody fragment that binds specifically to VEGF and reduces VEGF activity;   wherein the introducing results in a reduction in VEGF activity in the inner ear of the mammal.   
     
     
         23 . The method of  claim 22 , wherein the AAV vector further comprises one or both of a promoter and a Kozak sequence that are operably linked to the sequence encoding the antibody or the antigen-binding antibody fragment. 
     
     
         24 . The method of  claim 23 , wherein the AAV vector comprises a promoter selected from the group consisting of: an inducible promoter, a constitutive promoter, and a tissue-specific promoter. 
     
     
         25 . The method of any one of  claims 22 - 24 , wherein the AAV vector further comprises a polyadenylation signal sequence. 
     
     
         26 . The method of any one of  claims 22 - 25 , wherein the mammal is a human. 
     
     
         27 . The method of any one of  claims 22 - 26 , wherein the mammal has been identified or diagnosed as having an acoustic neuroma. 
     
     
         28 . The method of any one of  claims 22 - 26 , wherein the mammal has been identified or diagnosed as having a vestibular schwannoma. 
     
     
         29 . The method of any one of  claims 22 - 26 , wherein the mammal has been identified or diagnosed as having a neurofibromatosis type 2. 
     
     
         30 . The method of any one of  claims 22 - 29 , wherein the AAV vector comprises a nucleic acid sequence encoding a polypeptide comprising an antibody heavy chain variable domain operably linked to a signal peptide and a polypeptide comprising an antibody light chain variable domain operably linked to a signal peptide. 
     
     
         31 . The method of any one of  claims 22 - 29 , wherein the AAV vector comprises a nucleic acid sequence encoding a polypeptide comprising an antigen-binding antibody fragment operably linked to a signal peptide. 
     
     
         32 . A method of treating acoustic neuroma, vestibular schwannoma, or neurofibromatosis type 2 in an inner ear of a mammal, the method comprising:
 introducing into the inner ear of the mammal a therapeutically effective amount of an AAV vector that comprises a nucleotide sequence encoding   (a) a polypeptide comprising an antibody heavy chain variable domain operably linked to a signal peptide and a polypeptide comprising an antibody light chain variable domain operably linked to a signal peptide; or   (b) a polypeptide comprising an antigen-binding antibody fragment linked to a signal peptide;   wherein the polypeptide of (a) encodes an antibody that binds specifically to VEGF and reduces VEGF activity, the polypeptide of (b) encodes an antigen-binding antibody fragment that binds specifically to VEGF and reduces VEGF activity;   wherein the introducing results in treatment of acoustic neuroma, vestibular schwannoma, or neurofibromatosis type II, respectively, in the inner ear of the mammal.   
     
     
         33 . The method of  claim 32 , wherein the AAV vector further comprises one or both of a promoter and a Kozak sequence that are operably linked to the sequence encoding the antibody or the antigen-binding antibody fragment. 
     
     
         34 . The method of  claim 33 , wherein the AAV vector comprises a promoter selected from the group consisting of: an inducible promoter, a constitutive promoter, and a tissue-specific promoter. 
     
     
         35 . The method of any one of  claims 32 - 34 , wherein the AAV vector further comprises a polyadenylation signal sequence. 
     
     
         36 . The method of any one of  claims 32 - 35 , wherein the mammal is a human. 
     
     
         37 . The method of any one of  claims 32 - 36 , wherein the mammal has been identified or diagnosed as having an acoustic neuroma. 
     
     
         38 . The method of any one of  claims 32 - 36 , wherein the mammal has been identified or diagnosed as having a vestibular schwannoma. 
     
     
         39 . The method of any one of  claims 32 - 36 , wherein the mammal has been identified or diagnosed as having neurofibromatosis type 2. 
     
     
         40 . The method of any one of  claims 32 - 39 , wherein the AAV vector comprises a nucleic acid sequence encoding a polypeptide comprising an antibody heavy chain variable domain operably linked to a signal peptide and a polypeptide comprising an antibody light chain variable domain operably linked to a signal peptide. 
     
     
         41 . The method of any one of  claims 32 - 40 , wherein the AAV vector comprises a nucleic acid sequence encoding a polypeptide comprising an antigen-binding antibody fragment operably linked to a signal peptide. 
     
     
         42 . The method of any one of  claims 1 - 41 , wherein the antibody comprises a Fc region that includes one or more amino acid substitutions that decreases the half-life of the antibody in a mammal as compared to a control antibody; or
 the antigen-binding antibody fragment thereof has a decreased in vivo half-life as compared to a control antigen-binding antibody fragment.   
     
     
         43 . A method comprising introducing into an inner ear of a mammal a therapeutically effective amount of an adeno-associated virus (AAV) vector that comprises a nucleotide sequence encoding a soluble vascular endothelial growth factor (VEGF) receptor operably linked to a signal peptide. 
     
     
         44 . A method for increasing the level of a soluble vascular endothelial growth factor (VEGF) receptor in an inner ear of a mammal in need thereof, the method comprising:
 introducing into the inner ear of the mammal a therapeutically effective amount of an AAV vector that comprises a nucleotide sequence encoding a soluble VEGF receptor operably linked to a signal peptide;   wherein the introducing results in an increase in the level of the soluble VEGF receptor in the inner ear of the mammal.   
     
     
         45 . The method of  claim 43  or  44 , wherein the soluble VEGF receptor comprises a portion of an extracellular region of VEGF receptor-1 (VEGFR-1). 
     
     
         46 . The method of  claim 45 , wherein the portion of the extracellular region of VEGFR-1 comprises a contiguous sequence from wildtype human VEGFR-1. 
     
     
         47 . The method of  claim 46 , wherein the portion of the extracellular region of VEGFR-1 comprises one or more immunoglobulin-like domains in the extracellular region from wildtype human VEGFR-1. 
     
     
         48 . The method of  claim 45 , wherein the portion of the extracellular region of VEGFR-1 comprises a sequence that is at least 90% identical to a contiguous sequence from wildtype human VEGFR-1. 
     
     
         49 . The method of  claim 43  or  44 , wherein the soluble VEGF receptor comprises a portion of an extracellular region of VEGF receptor-2 (VEGFR-2). 
     
     
         50 . The method of  claim 49 , wherein the portion of the extracellular region of VEGFR-2 comprises a contiguous sequence from wildtype human VEGFR-2. 
     
     
         51 . The method of  claim 50 , wherein the portion of the extracellular region of VEGFR-2 comprises one or more immunoglobulin-like domains in the extracellular region from wildtype human VEGFR-2. 
     
     
         52 . The method of  claim 49 , wherein the portion of the extracellular region of VEGFR-2 comprises a sequence that is at least 90% identical to a contiguous sequence from wildtype human VEGFR-2. 
     
     
         53 . The method of  claim 43  or  44 , wherein the soluble VEGF receptor comprises a portion of an extracellular region of VEGFR-1 and a portion of an extracellular region of VEGFR-2. 
     
     
         54 . The method of  claim 53 , wherein:
 the portion of the extracellular region of VEGFR-1 comprises one or more immunoglobulin-like domains in the extracellular region from wildtype human VEGFR-1; and   the portion of the extracellular region of VEGFR-2 comprises one or more immunoglobulin-like domains in the extracellular region from wildtype human VEGFR-2.   
     
     
         55 . The method of  claim 54 , wherein the soluble VEGF receptor is aflibercept. 
     
     
         56 . The method of  claim 43  or  44 , wherein the soluble VEGF receptor comprises a portion of an extracellular region of VEGF receptor-3 (VEGFR-3). 
     
     
         57 . The method of  claim 56 , wherein the portion of the extracellular region of VEGFR-3 comprises a contiguous sequence from wildtype human VEGFR-3. 
     
     
         58 . The method of  claim 57 , wherein the portion of the extracellular region of VEGFR-3 comprises one or more immunoglobulin-like domains in the extracellular region from wildtype human VEGFR-3. 
     
     
         59 . The method of  claim 56 , wherein the portion of the extracellular region of VEGFR-3 comprises a sequence that is at least 90% identical to a contiguous sequence from wildtype human VEGFR-3. 
     
     
         60 . The method of any one of  claims 43 - 59 , wherein the soluble VEGF receptor comprises a Fc domain. 
     
     
         61 . The method of  claim 60 , wherein the Fc domain is an IgG1 Fc domain. 
     
     
         62 . The method of  claim 61 , wherein the IgG1 Fc domain is a human wildtype IgG1 Fc domain. 
     
     
         63 . The method of any one of  claims 43 - 62 , wherein the soluble VEGF receptor decreases the ability of a VEGF to bind to one or more of VEGFR-1, VEGFR-2, and VEGFR-3. 
     
     
         64 . The method of any one of  claims 43 - 63 , wherein the AAV vector further comprises one or both of a promoter and a Kozak sequence that are operably linked to the sequence encoding the soluble VEGF receptor. 
     
     
         65 . The method of  claim 64 , wherein the AAV vector comprises a promoter selected from the group consisting of: an inducible promoter, a constitutive promoter, and a tissue-specific promoter. 
     
     
         66 . The method of any one of  claims 43 - 65 , wherein the AAV vector further comprises a polyadenylation signal sequence. 
     
     
         67 . The method of any one of  claims 43 - 66 , wherein the mammal is a human. 
     
     
         68 . The method of any one of  claims 43 - 67 , wherein the mammal has been identified as having an inner ear disorder. 
     
     
         69 . The method of any one of  claims 43 - 67 , wherein the mammal has been diagnosed as having an inner ear disorder. 
     
     
         70 . A method for treating an inner ear disorder in a mammal in need thereof, the method comprising:
 introducing into the inner ear of the mammal a therapeutically effective amount of an AAV vector that comprises a nucleotide sequence encoding a soluble vascular endothelial growth factor (VEGF) receptor operably linked to a signal peptide;   wherein the introducing results in the treatment of the inner ear disorder in the mammal.   
     
     
         71 . The method of  claim 70 , wherein the AAV vector further comprises one or both of a promoter and a Kozak sequence that are operably linked to the sequence encoding the soluble VEGF receptor. 
     
     
         72 . The method of  claim 71 , wherein the AAV vector comprises a promoter selected from the group consisting of: an inducible promoter, a constitutive promoter, and a tissue-specific promoter. 
     
     
         73 . The method of any one of  claims 70 - 72 , wherein the AAV vector further comprises a polyadenylation signal sequence. 
     
     
         74 . The method of any one of  claims 70 - 73 , wherein the mammal is a human. 
     
     
         75 . The method of any one of  claims 70 - 74 , wherein the mammal has been identified as having an inner ear disorder. 
     
     
         76 . The method of any one of  claims 70 - 74 , wherein the mammal has been diagnosed as having an inner ear disorder. 
     
     
         77 . A method of reducing a VEGF activity in an inner ear of a mammal in need thereof, the method comprising:
 introducing into the inner ear of the mammal a therapeutically effective amount of an AAV vector that comprises a nucleotide sequence encoding a soluble vascular endothelial growth factor (VEGF) receptor operably linked to a signal peptide;   wherein the introducing results in a reduction in the VEGF activity in the inner ear of the mammal.   
     
     
         78 . The method of  claim 77 , wherein the AAV vector further comprises one or both of a promoter and a Kozak sequence that are operably linked to the sequence encoding the soluble VEGF receptor. 
     
     
         79 . The method of  claim 78 , wherein the AAV vector comprises a promoter selected from the group consisting of: an inducible promoter, a constitutive promoter, and a tissue-specific promoter. 
     
     
         80 . The method of any one of  claims 77 - 79 , wherein the AAV vector further comprises a polyadenylation signal sequence. 
     
     
         81 . The method of any one of  claims 77 - 80 , wherein the mammal is a human. 
     
     
         82 . The method of any one of  claims 77 - 81 , wherein the mammal has been identified or diagnosed as having an acoustic neuroma. 
     
     
         83 . The method of any one of  claims 77 - 81 , wherein the mammal has been identified or diagnosed as having a vestibular schwannoma. 
     
     
         84 . The method of any one of  claims 77 - 81 , wherein the mammal has been identified or diagnosed as having a neurofibromatosis type 2. 
     
     
         85 . A method of treating acoustic neuroma, vestibular schwannoma, or neurofibromatosis type 2 in an inner ear of a mammal, the method comprising:
 introducing into the inner ear of the mammal a therapeutically effective amount of an AAV vector that comprises a nucleotide sequence encoding a nucleotide sequence encoding a soluble vascular endothelial growth factor (VEGF) receptor operably linked to a signal peptide;   wherein the introducing results in treatment of acoustic neuroma, vestibular schwannoma, or neurofibromatosis type II, respectively, in the inner ear of the mammal.   
     
     
         86 . The method of  claim 85 , wherein the AAV vector further comprises one or both of a promoter and a Kozak sequence that are operably linked to the sequence encoding the soluble VEGF receptor. 
     
     
         87 . The method of  claim 86 , wherein the AAV vector comprises a promoter selected from the group consisting of: an inducible promoter, a constitutive promoter, and a tissue-specific promoter. 
     
     
         88 . The method of any one of  claims 85 - 87 , wherein the AAV vector further comprises a polyadenylation signal sequence. 
     
     
         89 . The method of any one of  claims 85 - 88 , wherein the mammal is a human. 
     
     
         90 . The method of any one of  claims 85 - 89 , wherein the mammal has been identified or diagnosed as having an acoustic neuroma. 
     
     
         91 . The method of any one of  claims 85 - 89 , wherein the mammal has been identified or diagnosed as having a vestibular schwannoma. 
     
     
         92 . The method of any one of  claims 85 - 89 , wherein the mammal has been identified or diagnosed as having neurofibromatosis type 2. 
     
     
         93 . The method of any one of  claims 70 - 92 , wherein the soluble VEGF receptor comprises a portion of an extracellular region of VEGF receptor-1 (VEGFR-1). 
     
     
         94 . The method of  claim 93 , wherein the portion of the extracellular region of VEGFR-1 comprises a contiguous sequence from wildtype human VEGFR-1. 
     
     
         95 . The method of  claim 94 , wherein the portion of the extracellular region of VEGFR-1 comprises one or more immunoglobulin-like domains in the extracellular region from wildtype human VEGFR-1. 
     
     
         96 . The method of  claim 93 , wherein the portion of the extracellular region of VEGFR-1 comprises a sequence that is at least 90% identical to a contiguous sequence from wildtype human VEGFR-1. 
     
     
         97 . The method of any one of  claims 70 - 92 , wherein the soluble VEGF receptor comprises a portion of an extracellular region of VEGF receptor-2 (VEGFR-2). 
     
     
         98 . The method of  claim 97 , wherein the portion of the extracellular region of VEGFR-2 comprises a contiguous sequence from wildtype human VEGFR-2. 
     
     
         99 . The method of  claim 98 , wherein the portion of the extracellular region of VEGFR-2 comprises one or more immunoglobulin-like domains in the extracellular region from wildtype human VEGFR-2. 
     
     
         100 . The method of  claim 97 , wherein the portion of the extracellular region of VEGFR-2 comprises a sequence that is at least 90% identical to a contiguous sequence from wildtype human VEGFR-2. 
     
     
         101 . The method of any one of  claims 70 - 92 , wherein the soluble VEGF receptor comprises a portion of an extracellular region of VEGFR-1 and a portion of an extracellular region of VEGFR-2. 
     
     
         102 . The method of  claim 101 , wherein:
 the portion of the extracellular region of VEGFR-1 comprises one or more immunoglobulin-like domains in the extracellular region from wildtype human VEGFR-1; and   the portion of the extracellular region of VEGFR-2 comprises one or more immunoglobulin-like domains in the extracellular region from wildtype human VEGFR-2.   
     
     
         103 . The method of  claim 102 , wherein the soluble VEGF receptor is aflibercept. 
     
     
         104 . The method of any one of  claims 70 - 92 , wherein the soluble VEGF receptor comprises a portion of an extracellular region of VEGF receptor-3 (VEGFR-3). 
     
     
         105 . The method of  claim 104 , wherein the portion of the extracellular region of VEGFR-3 comprises a contiguous sequence from wildtype human VEGFR-3. 
     
     
         106 . The method of  claim 105 , wherein the portion of the extracellular region of VEGFR-3 comprises one or more immunoglobulin-like domains in the extracellular region from wildtype human VEGFR-3. 
     
     
         107 . The method of  claim 104 , wherein the portion of the extracellular region of VEGFR-3 comprises a sequence that is at least 90% identical to a contiguous sequence from wildtype human VEGFR-3. 
     
     
         108 . The method of any one of  claims 70 - 107 , wherein the soluble VEGF receptor comprises a Fc domain. 
     
     
         109 . The method of  claim 108 , wherein the Fc domain is an IgG1 Fc domain. 
     
     
         110 . The method of  claim 109 , wherein the IgG1 Fc domain is a human wildtype IgG1 Fc domain. 
     
     
         111 . The method of any one of  claims 70 - 110 , wherein the soluble VEGF receptor decreases the ability of a VEGF to bind to one or more of VEGFR-1, VEGFR-2, and VEGFR-3. 
     
     
         112 . The method of any one of  claims 43 - 111 , wherein the AAV vector further comprises a secretion sequence.