METHODS AND COMPOSITIONS FOR NEEDLELESS DELIVERY OF ANTIBODIES
The present invention relates, in part, to methods and compositions for needleless delivery of antibodies to a subject. The present invention also relates, in part, to methods for needleless delivery of fusion proteins comprising a bioactive molecule and an antibody fragment to subject. In one aspect, the methods and compositions involve administering to the subject a delivery construct comprising a carrier construct non-covalently bound to the antibody or fusion protein to be delivered, wherein the carrier construct comprises a receptor-binding domain, a transcytosis domain, and an antibody-binding domain to which the antibody or the antibody fragment of the fusion protein non-covalently binds.
1 . A delivery construct, comprising a carrier construct non-covalently bound to an antibody, wherein said carrier construct comprises:
a)— a receptor-binding domain,
b)— a transcytosis domain, and
c)— an antibody-binding domain to which the antibody non-covalently binds.
2 . The delivery construct of claim 1 , wherein said antibody-binding domain is an Fc binding domain.
3 . The delivery construct of claim 2 , wherein said Fc binding domain is a Protein G, Protein A, an Fc receptor, or an antibody-binding fragment thereof.
4 . The delivery construct of claim 3 , wherein said Fc receptor is Fc receptor γ or neonatal Fc receptor.
5 . The delivery construct of claim 1 , wherein the antibody-binding domain is an antigen.
6 . The delivery construct of claim 1 , wherein the antibody is selected from the group consisting of a monoclonal antibody, a human antibody, a humanized antibody, a chimeric antibody, a single chain antibody, a single domain antibody, an antigen-binding antibody fragment, a Fab fragment and a F(ab′)2 fragment.
7 . The delivery construct of claim 1 , wherein the antibody is selected from RITUXAN, AVASTIN, HERCEPTIN, ERBITUX, TYSABRI, REMICADE, ENBREL, HUMIRA, XOLAIR, and RAPTUVA.
8 . The delivery construct of claim 1 , wherein the carrier construct further comprises a cleavable linker, wherein cleavage at said cleavable linker separates said antibody-binding domain from the remainder of said carrier construct, and wherein said cleavable linker is cleavable by an enzyme that i) exhibits greater activity at a basal-lateral membrane of a polarized epithelial cell than at an apical membrane of the polarized epithelial cell, or ii) exhibits greater activity in the plasma of a subject than at an apical membrane of the polarized epithelial cell of the subject.
9 . The delivery construct of claim 8 , wherein said cleavable linker comprises an amino acid sequence that is selected from the group consisting of Ala-Ala-Pro-Phe (SEQ ID NO.:4), Gly-Gly-Phe (SEQ ID NO.:5), Ala-Ala-Pro-Val (SEQ ID NO.:6), Gly-Gly-Leu (SEQ ID NO.:7), Ala-Ala-Leu (SEQ ID NO.:8), Phe-Val-Arg (SEQ ID NO:9), Val-Gly-Arg (SEQ ID NO.:10).
10 . The delivery construct of claim 8 , wherein the epithelial cell is selected from the group consisting of nasal epithelial cells, oral epithelial cells, intestinal epithelial cells, rectal epithelial cells, vaginal epithelial cells, and pulmonary epithelial cells.
11 . The delivery construct of claim 8 , wherein said enzyme that is present at a basal-lateral membrane of a polarized epithelial cell is selected from the group consisting of Cathepsin GI, Chymotrypsin I, Elastase I, Subtilisin AI, Subtilisin AII, Thrombin I, and Urokinase I.
12 . The delivery construct of claim 1 , wherein said receptor-binding domain is selected from the group consisting of a receptor-binding domain from Pseudomonas exotoxin A; cholera toxin; botulinum toxin; diptheria toxin; shiga toxin; shiga-like toxin; monoclonal antibodies; polyclonal antibodies; single-chain antibodies; TGF α; EGF; IGF-I; IGF-II; IGF-III; IL-1; IL-2; IL-3; IL-6; MIP-1a; MIP-1b; MCAF; and IL-8.
13 . The delivery construct of claim 1 , wherein said receptor-binding domain binds to a cell-surface receptor that is selected from the group consisting of α2-macroglobulin receptor, epidermal growth factor receptor, transferrin receptor, chemokine receptor, CD25, CD11B, CD11C, CD80, CD86, TNFα receptor, TOLL receptor, M-CSF receptor, GM-CSF receptor, scavenger receptor, and VEGF receptor.
14 . The delivery construct of claim 12 , wherein said receptor-binding domain of Pseudomonas exotoxin A is Domain Ia of Pseudomonas exotoxin A.
15 . The delivery construct of claim 12 , wherein said receptor-binding domain of Pseudomonas exotoxin A has an amino acid sequence that is SEQ ID NO.:1.
16 . The delivery construct of claim 1 , wherein said transcytosis domain is selected from the group consisting of a transcytosis domain from Pseudomonas exotoxin A, botulinum toxin, diptheria toxin, pertussis toxin, cholera toxin, heat-labile E. coli enterotoxin, shiga toxin, and shiga-like toxin.
17 . The delivery construct of claim 16 , wherein said transcytosis domain is a Pseudomonas exotoxin A transcytosis domain.
18 . The delivery construct of claim 17 , wherein said Pseudomonas exotoxin A transcytosis domain has an amino acid sequence that is SEQ ID NO.:2.
19 . A cell comprising a first polynucleotide and a second polynucleotide, wherein the first polynucleotide encodes an antibody and the second polynucleotide encodes a carrier construct comprising:
a)— a receptor-binding domain,
b)— a transcytosis domain, and
c)— an antibody-binding domain to which the antibody non-covalently binds.
20 . The cell of claim 19 , wherein said antibody-binding domain is an Fc binding domain.
21 . The cell of claim 20 , wherein said Fc binding domain is a Protein G, Protein A or an Fc receptor.
22 . The cell of claim 21 , wherein said Fc receptor is Fc receptor γ or neonatal Fc receptor.
23 . The cell of claim 19 , wherein the antibody-binding domain is an antigen.
24 . The cell of claim 19 , wherein the antibody is selected from the group consisting of a monoclonal antibody, a human antibody, a humanized antibody, a chimeric antibody, a single chain antibody, a single domain antibody, a Fab fragment and a F(ab′)2 fragment.
25 . The cell of claim 19 , wherein the antibody is selected from RITUXAN, AVASTIN, HERCEPTIN, ERBITUX, TYSABRI, REMICADE, ENBREL, HUMIRA, XOLAIR, and RAPTIVA.
26 . The cell of claim 19 , wherein the carrier construct further comprises a cleavable linker, wherein cleavage at said cleavable linker separates said antibody-binding domain from the remainder of said carrier construct, and wherein said cleavable linker is cleavable by an enzyme that i) exhibits greater activity at a basal-lateral membrane of a polarized epithelial cell than at an apical membrane of the polarized epithelial cell, or ii) exhibits greater activity in the plasma of a subject than at an apical membrane of the polarized epithelial cell of the subject.
27 . The cell of claim 26 , wherein said cleavable linker comprises an amino acid sequence that is selected from the group consisting of Ala-Ala-Pro-Phe (SEQ ID NO.:4), Gly-Gly-Phe (SEQ ID NO.:5), Ala-Ala-Pro-Val (SEQ ID NO.:6), Gly-Gly-Leu (SEQ ID NO.:7), Ala-Ala-Leu (SEQ ID NO.:8), Phe-Val-Arg (SEQ ID NO.:9), Val-Gly-Arg (SEQ ID NO.:10).
28 . The cell of claim 26 , wherein the epithelial cell is selected from the group consisting of nasal epithelial cells, oral epithelial cells, intestinal epithelial cells, rectal epithelial cells, vaginal epithelial cells, and pulmonary epithelial cells.
29 . The cell of claim 26 , wherein said enzyme that is present at a basal-lateral membrane of a polarized epithelial cell is selected from the group consisting of Cathepsin GI, Chymotrypsin I, Elastase I, Subtilisin AI, Subtilisin AII, Thrombin I, and Urokinase I.
30 . The cell of claim 19 , wherein said receptor-binding domain is selected from the group consisting of a receptor-binding domain from Pseudomonas exotoxin A; cholera toxin; botulinum toxin; diptheria toxin; shiga toxin; shiga-like toxin; monoclonal antibodies; polyclonal antibodies; single-chain antibodies; TGF α; EGF; IGF-I; IGF-II; IGF-III; IL-1; IL-2; IL-3; IL-6; MIP-1a; MIP-Ib; MCAF; and IL-8.
31 . The cell of claim 19 , wherein said receptor-binding domain binds to a cell-surface receptor that is selected from the group consisting of α2-macroglobulin receptor, epidermal growth factor receptor, transferrin receptor, chemokine receptor, CD25, CD11B, CD11C, CD80, CD96, TNFα receptor, TOLL receptor, M-CSF receptor, GM-CSF receptor, scavenger receptor, and VEGF receptor.
32 . The cell of claim 30 , wherein said receptor-binding domain of Pseudomonas exotoxin A is Domain Ia of Pseudomonas exotoxin A.
33 . The cell of claim 30 , wherein said receptor-binding domain of Pseudomonas exotoxin A has an amino acid sequence that is SEQ ID NO.:1.
34 . The cell of claim 19 , wherein said transcytosis domain is selected from the group consisting of a transcytosis domain from Pseudomonas exotoxin A, botulinum toxin, diptheria toxin, pertussis toxin, cholera toxin, heat-labile E. coli enterotoxin, shiga toxin, and shiga-like toxin.
35 . The cell of claim 34 , wherein said transcytosis domain is Pseudomonas exotoxin A transcytosis domain.
36 . The cell of claim 35 , wherein said Pseudomonas exotoxin A transcytosis domain has an amino acid sequence that is SEQ ID NO.:2.
37 . A composition comprising a delivery construct of claim 1 .
38 . The composition of claim 37 , wherein said composition further comprises a pharmaceutically acceptable diluent, excipient, vehicle, or carrier.
39 . The composition of claim 37 , wherein said composition is formulated for nasal or oral administration.
40 . A method for delivering an antibody to a subject, the method comprising contacting an apical surface of a polarized epithelial cell of the subject with a delivery construct comprising a carrier construct non-covalently bound to the antibody, wherein said carrier construct comprises a receptor-binding domain, a transcytosis domain, and an antibody-binding domain to which the antibody non-covalently binds, and wherein the transcytosis domain transcytoses the binding partner to and through the basal-lateral membrane of said epithelial cell.
41 . The method of claim 40 , wherein said antibody-binding domain is an Fc binding domain.
42 . The method of claim 41 , wherein said Fc binding domain is a Protein G, Protein A or an Fc receptor.
43 . The method of claim 42 , wherein said Fc receptor is Fc receptor γ or neonatal Fc receptor.
44 . The method of claim 40 , wherein the antibody-binding domain is an antigen.
45 . The method of claim 40 , wherein the antibody is selected from the group consisting of a monoclonal antibody, a human antibody, a humanized antibody, a chimeric antibody, a single chain antibody, a single domain antibody, a Fab fragment and a F(ab′)2 fragment.
46 . The method of claim 40 , wherein the antibody is selected from RITUXAN, AVASTIN, HERCEPTIN, ERBITUX, TYSABRI, REMICADE, ENBREL, HUMIRA, XOLAIR, and RAPTIVA.
47 . The method of claim 40 , wherein the carrier construct further comprises a cleavable linker, wherein cleavage at said cleavable linker separates said antibody-binding domain from the remainder of said carrier construct, and wherein said cleavable linker is cleavable by an enzyme that i) exhibits greater activity at a basal-lateral membrane of a polarized epithelial cell than at an apical membrane of the polarized epithelial cell, or ii) exhibits greater activity in the plasma of the subject than at an apical membrane of the polarized epithelial cell of the subject.
48 . The method of claim 47 , wherein said cleavable linker comprises an amino acid sequence that is selected from the group consisting of Ala-Ala-Pro-Phe (SEQ ID NO.:4), Gly-Gly-Phe (SEQ ID NO.:5), Ala-Ala-Pro-Val (SEQ ID NO.:6), Gly-Gly-Leu (SEQ ID NO.:7), Ala-Ala-Leu (SEQ ID NO.:8), Phe-Val-Arg (SEQ ID NO.:9), Val-Gly-Arg (SEQ ID NO.:10).
49 . The method of claim 47 , wherein said enzyme that is present at a basal-lateral membrane of a polarized epithelial cell is selected from the group consisting of Cathepsin GI, Chymotrypsin I, Elastase I, Subtilisin AI, Subtilisin AII, Thrombin I, and Urokinase I.
50 . The method of claim 47 , wherein the epithelial cell is selected from the group consisting of nasal epithelial cells, oral epithelial cells, intestinal epithelial cells, rectal epithelial cells, vaginal epithelial cells, and pulmonary epithelial cells.
51 . The method of claim 40 , wherein said receptor-binding domain is selected from the group consisting of a receptor-binding domain from Pseudomonas exotoxin A; cholera toxin; botulinum toxin; diptheria toxin; shiga toxin; shiga-like toxin; monoclonal antibodies; polyclonal antibodies; single-chain antibodies; TGF α; EGF; IGF-I; IGF-II; IGF-II; IL-1; IL-2; IL-3; IL-6; MIP-1a; MIP-1b; MCAF; and IL-8.
52 . The method of claim 40 , wherein said receptor-binding domain binds to a cell-surface receptor that is selected from the group consisting of α2-macroglobulin receptor, epidermal growth factor receptor, transferrin receptor, chemokine receptor, CD25, CD11B, CD11C, CD80, CD86, TNFα receptor, TOLL receptor, M-CSF receptor, GM-CSF receptor, scavenger receptor, and VEGF receptor.
53 . The method of claim 51 , wherein said receptor-binding domain of Pseudomonas exotoxin A is Domain Ia of Pseudomonas exotoxin A.
54 . The method of claim 51 , wherein said receptor-binding domain of Pseudomonas exotoxin A has an amino acid sequence that is SEQ ID NO.:1.
55 . The method of claim 40 , wherein said transcytosis domain is selected from the group consisting of a transcytosis domain from Pseudomonas exotoxin A, botulinum toxin, diptheria toxin, pertussis toxin, cholera toxin, heat-labile E. coli enterotoxin, shiga toxin, and shiga-like toxin.
56 . The method of claim 55 , wherein said transcytosis domain is Pseudomonas exotoxin A transcytosis domain.
57 . The method of claim 56 , wherein said Pseudomonas exotoxin A transcytosis domain has an amino acid sequence that is SEQ ID NO.:2.
58 . The method of claim 40 , wherein said subject is a mammal.
59 . The method of claim 58 , wherein said mammal is a human.
60 . A method for delivering an antibody to the bloodstream of a subject, the method comprising contacting the delivery construct of claim 1 to an apical surface of a polarized epithelial cell of the subject, such that the antibody is delivered to the bloodstream of the subject.
61 . The method of claim 60 , wherein said antibody-binding domain is an Fc binding domain.
62 . The method of claim 61 , wherein said Fc binding domain is a Protein G, Protein A or an Fc receptor.
63 . The method of claim 62 , wherein said Fc receptor is Fc receptor γ or neonatal Fc receptor.
64 . The method of claim 60 , wherein the antibody-binding domain is an antigen.
65 . The method of claim 60 , wherein the antibody is selected from the group consisting of a monoclonal antibody, a human antibody, a humanized antibody, a chimeric antibody, a single chain antibody, a single domain antibody, a Fab fragment and a F(ab′)2 fragment.
66 . The method of claim 60 , wherein the antibody is selected from RITUXAN, AVASTIN, HERCEPTIN, ERBITUX, TYSABRI, REMICADE, ENBREL, HUMIRA, XOLAIR, and RAPTIVA.
67 . The method of claim 60 , wherein the carrier construct further comprises a cleavable linker, wherein cleavage at said cleavable linker separates said antibody-binding domain from the remainder of said carrier construct, and wherein said cleavable linker is cleavable by an enzyme that i) exhibits greater activity at a basal-lateral membrane of a polarized epithelial cell than at an apical membrane of the polarized epithelial cell, or ii) exhibits greater activity in the plasma of the subject than at an apical membrane of the polarized epithelial cell of the subject.
68 . The method of claim 67 , wherein said cleavable linker comprises an amino acid sequence that is selected from the group consisting of Ala-Ala-Pro-Phe (SEQ ID NO.:4), Gly-Gly-Phe (SEQ ID NO.:5), Ala-Ala-Pro-Val (SEQ ID NO.:6), Gly-Gly-Leu (SEQ ID NO.:7), Ala-Ala-Leu (SEQ ID NO.:8), Phe-Val-Arg (SEQ ID NO.:9), Val-Gly-Arg (SEQ ID NO.:10).
69 . The method of claim 67 , wherein said enzyme that is present at a basal-lateral membrane of a polarized epithelial cell is selected from the group consisting of Cathepsin GI, Chymotrypsin I, Elastase I, Subtilisin AI, Subtilisin AII, Thrombin I, and Urokinase I.
70 . The method of claim 67 , wherein the epithelial cell is selected from the group consisting of nasal epithelial cells, oral epithelial cells, intestinal epithelial cells, rectal epithelial cells, vaginal epithelial cells, and pulmonary epithelial cells.
71 . The method of claim 60 , wherein said receptor-binding domain is selected from the group consisting of a receptor-binding domain from Pseudomonas exotoxin A; cholera toxin; botulinum toxin; diptheria toxin; shiga toxin; shiga-like toxin; monoclonal antibodies; polyclonal antibodies; single-chain antibodies; TGF α; EGF; IGF-I; IGF-II; IGF-III; IL-1; IL-2; IL-3; IL-6; MIP-1a; MIP-1b; MCAF; and IL-8.
72 . The method of claim 60 , wherein said receptor-binding domain binds to a cell-surface receptor that is selected from the group consisting of α2-macroglobulin receptor, epidermal growth factor receptor, transferrin receptor, chemokine receptor, CD25, CD11B, CD11C, CD80, CD86, TNFα receptor, TOLL receptor, M-CSF receptor, GM-CSF receptor, scavenger receptor, and VEGF receptor.
73 . The method of claim 71 , wherein said receptor-binding domain of Pseudomonas exotoxin A is Domain Ia of Pseudomonas exotoxin A.
74 . The method of claim 71 , wherein said receptor-binding domain of Pseudomonas exotoxin A has an amino acid sequence that is SEQ ID NO.:1.
75 . The method of claim 60 , wherein said transcytosis domain is selected from the group consisting of a transcytosis domain from Pseudomonas exotoxin A, botulinum toxin, diptheria toxin, pertussis toxin, cholera toxin, heat-labile E. coli enterotoxin, shiga toxin, and shiga-like toxin.
76 . The method of claim 75 , wherein said transcytosis domain is Pseudomonas exotoxin A transcytosis domain.
77 . The method of claim 76 , wherein said Pseudomonas exotoxin A transcytosis domain has an amino acid sequence that is SEQ ID NO.:2.
78 . The method of claim 60 , wherein said subject is a mammal.
79 . The method of claim 78 , wherein said mammal is a human.