Anti-vegf antibodies

Abstract

Anti-VEGF antibodies and variants thereof, including those having high affinity for binding to VEGF, are disclosed. Also provided are methods of using phage display technology with naïve libraries to generate and select the anti-VEGF antibodies with desired binding and other biological activities. Further contemplated are uses of the antibodies in research, diagnostic and therapeutic applications.

1 - 25 . (canceled) 
     
     
         26 . A method of inhibiting angiogenesis in a human or a mouse, the method comprising administering to the human or mouse an antibody capable of binding to human VEGF and mouse VEGF with Kd values within 10-fold of the other, wherein the antibody binds to an epitope comprising residue G88 of human VEGF. 
     
     
         27 . The method of  claim 26 , wherein the epitope further comprises any one or more additional residues selected from the group consisting of residues F17, K48, L66, M81, 183, H86, Q89, and 191 of human VEGF. 
     
     
         28 . The method of  claim 26 , wherein the antibody contacts between about 50% to about 80% of the surface area of residue G88 of human VEGF. 
     
     
         29 . A method of inhibiting angiogenesis in a human or a mouse, the method comprising administering to the human or mouse an antibody capable of binding to human VEGF and mouse VEGF with Kd values within 10-fold of the other, wherein the antibody comprises one or more of the following complementarity determining regions (CDRs):
 (a) a CDR-H1 comprising the amino acid sequence GFAISDYDIH (SEQ ID NO: 505);   (b) a CDR-H2 comprising the amino acid sequence DIAPYAGATAYADSVKG (SEQ ID NO: 506);   (c) a CDR-H3 comprising the amino acid sequence SSYAYYAAMDY (SEQ ID NO: 507);   (d) a CDR-L1 comprising the amino acid sequence RASQSYAYAVA (SEQ ID NO: 493);   (e) a CDR-L2 comprising the amino acid sequence DASYLYS (SEQ ID NO: 494); and   (f) a CDR-L3 comprising the amino acid sequence QQAYSSPDT (SEQ ID NO: 495).   
     
     
         30 . The method of  claim 29 , wherein the heavy chain of the antibody comprises at least one, at least two, or all three of the CDR sequences selected from GFAISDYIH (SEQ ID NO: 505), DIAPYAGATAYADSVKG (SEQ ID NO: 506), and SSYAYYAAMDY (SEQ ID NO: 507); or
 wherein the light chain of the antibody comprises at least one, at least two, or all three of the CDR sequences selected from RASQSYAYAVA (SEQ ID NO: 493), DASYLYS (SEQ ID NO: 494), and QQAYSSPDT (SEQ ID NO: 495).   
     
     
         31 . The method of  claim 29 , wherein the heavy chain of the antibody comprises the sequence MKKNIAFLLASMFVFSIATNAYAEVQLVESGGGLVQPGGSLRLSCAASGFAISDYDIHWVRQAPGKGLEWVA DIAPYAGATAYADSVKGRFTISADTSKNTAYLQMNSLRAEDTAVYYCSRSSYAYYAAMDYWGQGTLVTVSS (SEQ ID NO: 949). 
     
     
         32 . The method of  claim 29 , wherein the light chain of the antibody comprises the sequence DIQMTQSPSSLSASVGDRVTITCRASQSYAYAVAWYQQKPGKAPKLLIYDASYLYSGVP SRFSGSGSGTDFTLTISSLQPEDFATYYCQQAYSSPDTFGQGTKVEIK (SEQ ID NO: 948). 
     
     
         33 . A method of inhibiting angiogenesis in a human or a mouse, the method comprising administering to the human or mouse an antibody capable of binding to human VEGF and mouse VEGF with Kd values within 10-fold of the other, wherein the antibody comprises one or more of the following complementarity determining regions (CDRs):
 (a) a CDR-H1 comprising the amino acid sequence DYDIH (SEQ ID NO: 947);   (b) a CDR-H2 comprising the amino acid sequence AIAPYSGSTYYADSVK (SEQ ID NO: 510);   (c) a CDR-H3 comprising the amino acid sequence SYAYYSAMDY (SEQ ID NO: 511);   (d) a CDR-L1 comprising the amino acid sequence CRASQASYYDVA (SEQ ID NO: 945);   (e) a CDR-L2 comprising the amino acid sequence AASYLYS (SEQ ID NO: 498); and   (f) a CDR-L3 comprising the amino acid sequence CQQYYYAPAT (SEQ ID NO: 946).   
     
     
         34 . The method of  claim 33 , wherein the heavy chain of the antibody comprises at least one, at least two, or all three of the CDR sequences selected from: DYDIH (SEQ ID NO: 947), AIAPYSGSTYYADSVK (SEQ ID NO: 510), and SYAYYSAMDY (SEQ ID NO: 511); or
 wherein the light chain of the antibody comprises at least one, at least two, or all three of the CDR sequences selected from CRASQASYYDVA (SEQ ID NO: 945), AASYLYS (SEQ ID NO: 498), and CQQYYYAPAT (SEQ ID NO: 946).   
     
     
         35 . The method of  claim 33 , wherein the heavy chain of the antibody comprises the sequence MKKN IAFLLASMFVFSIATNAYAEVQLVESGGGLVQPGGSLRLSCAASGFSISDYDIHWVRQAPGKGLEWVA AIAPYSGSTYYADSVKGRFTISADTSKNTAYLQMNSLRAEDTAVYYCSRSSYAYYSAMDYWGQGTLVTVSS (SEQ ID NO: 951). 
     
     
         36 . The method of  claim 33 , wherein the light chain of the antibody comprises the sequence DIQMTQSPSSLSASVGDRVTITCRASQASYYDVAWYQQKPGKAPKLLIYAASYLYSGVPSRFSGSGSGTDFT LTISSLQPEDFATYYCQQYYYAPATFGQGTKVEIK (SEQ ID NO: 950). 
     
     
         37 . The method of any one of  claims 26 ,  29 , and  33 , wherein the antibody is selected from the group consisting of a synthetic antibody, a chimeric antibody, a humanized antibody, an affinity matured antibody, and a bispecific antibody. 
     
     
         38 . The method of any one of  claims 26 ,  29 , and  33 , wherein the antibody is an antibody fragment. 
     
     
         39 . The method of any one of  claims 26 ,  29 , and  33 , wherein the antibody binds to mouse and human VEGF with Kd values of no more than about 10 nM. 
     
     
         40 . The method of any one of  claims 26 ,  29 , and  33 , wherein the human or mouse is suffering from cancer or a disease caused by ocular neovascularization. 
     
     
         41 . The method of  claim 40 , wherein the cancer is selected from the group consisting of breast cancer, colorectal cancer, non-small cell lung cancer, non-Hodgkin's lymphoma (NHL), renal cancer, prostate cancer, liver cancer, head and neck cancer, melanoma, ovarian cancer, mesothelioma, and glioblastoma. 
     
     
         42 . The method of  claim 40 , wherein the disease caused by ocular neovascularization is diabetic blindness, retinopathy, primary diabetic retinopathy, or age-induced macular degeneration. 
     
     
         43 . A method of alleviating cancer or a disease caused by ocular neovascularization in a human or a mouse, the method comprising administering to the human or mouse an antibody capable of binding to human VEGF and mouse VEGF with Kd values within 10-fold of the other, wherein the antibody binds to an epitope comprising residue G88 of human VEGF. 
     
     
         44 . The method of  claim 43 , wherein the epitope further comprises any one or more additional residues selected from the group consisting of residues F17, K48, L66, M81, 183, H86, Q89, and 191 of human VEGF. 
     
     
         45 . The method of  claim 43 , wherein the antibody contacts between about 50% to about 80% of the surface area of residue G88 of human VEGF. 
     
     
         46 . A method of alleviating cancer or a disease caused by ocular neovascularization in a human or a mouse, the method comprising administering to the human or mouse an antibody capable of binding to human VEGF and mouse VEGF with Kd values within 10-fold of the other, wherein the antibody comprises one or more of the following complementarity determining regions (CDRs):
 (a) a CDR-H1 comprising the amino acid sequence GFAISDYDIH (SEQ ID NO: 505);   (b) a CDR-H2 comprising the amino acid sequence DIAPYAGATAYADSVKG (SEQ ID NO: 506),   (c) a CDR-H3 comprising the amino acid sequence SSYAYYAAMDY (SEQ ID NO: 507);   (d) a CDR-L1 comprising the amino acid sequence RASQSYAYAVA (SEQ ID NO: 493);   (e) a CDR-L2 comprising the amino acid sequence DASYLYS (SEQ ID NO: 494); and   (f) a CDR-L3 comprising the amino acid sequence QQAYSSPDT (SEQ ID NO: 495).   
     
     
         47 . The method of  claim 46 , wherein the heavy chain of the antibody comprises at least one, at least two, or all three of the CDR sequences selected from GFAISDYIH (SEQ ID NO: 505), DIAPYAGATAYADSVKG (SEQ ID NO: 506), and SSYAYYAAMDY (SEQ ID NO: 507); or
 wherein the light chain of the antibody comprises at least one, at least two, or all three of the CDR sequences selected from RASQSYAYAVA (SEQ ID NO: 493), DASYLYS (SEQ ID NO: 494), and QQAYSSPDT (SEQ ID NO: 495).   
     
     
         48 . The method of  claim 46 , wherein the heavy chain of the antibody comprises the sequence MKKN IAFLLASMFVFSIATNAYAEVQLVESGGGLVQPGGSLRLSCAASGFAISDYDIHWVRQAPGKGLEWVA DIAPYAGATAYADSVKGRFTISADTSKNTAYLQMNSLRAEDTAVYYCSRSSYAYYAAMDYWGQGTLVTVSS (SEQ ID NO: 949). 
     
     
         49 . The method of  claim 46 , wherein the light chain of the antibody comprises the sequence DIQMTQSPSSLSASVGDRVTITCRASQSYAYAVAWYQQKPGKAPKLLIYDASYLYSGVP SRFSGSGSGTDFTLTISSLQPEDFATYYCQQAYSSPDTFGQGTKVEIK (SEQ ID NO: 948). 
     
     
         50 . A method of alleviating cancer or a disease caused by ocular neovascularization in a human or a mouse, the method comprising administering to the human or mouse an antibody capable of binding to human VEGF and mouse VEGF with Kd values within 10-fold of the other, wherein the antibody comprises one or more of the following complementarity determining regions (CDRs):
 (a) a CDR-H1 comprising the amino acid sequence DYDIH (SEQ ID NO: 947);   (b) a CDR-H2 comprising the amino acid sequence AIAPYSGSTYYADSVK (SEQ ID NO: 510);   (c) a CDR-H3 comprising the amino acid sequence SYAYYSAMDY (SEQ ID NO: 511);   (d) a CDR-L1 comprising the amino acid sequence CRASQASYYDVA (SEQ ID NO: 945);   (e) a CDR-L2 comprising the amino acid sequence AASYLYS (SEQ ID NO: 498); and   (f) a CDR-L3 comprising the amino acid sequence CQQYYYAPAT (SEQ ID NO: 946).   
     
     
         51 . The method of  claim 50 , wherein the heavy chain of the antibody comprises at least one, at least two, or all three of the CDR sequences selected from: DYDIH (SEQ ID NO: 947), AIAPYSGSTYYADSVK (SEQ ID NO: 510), and SYAYYSAMDY (SEQ ID NO: 511); or
 wherein the light chain of the antibody comprises at least one, at least two, or all three of the CDR sequences selected from CRASQASYYDVA (SEQ ID NO: 945), AASYLYS (SEQ ID NO: 498), and CQQYYYAPAT (SEQ ID NO: 946).   
     
     
         52 . The method of  claim 50 , wherein the heavy chain of the antibody comprises the sequence MKKN IAFLLASMFVFSIATNAYAEVQLVESGGGLVQPGGSLRLSCAASGFSISDYDIHWVRQAPGKGLEWVA AIAPYSGSTYYADSVKGRFTISADTSKNTAYLQMNSLRAEDTAVYYCSRSSYAYYSAMDYWGQGTLVTVSS (SEQ ID NO: 951). 
     
     
         53 . The method of  claim 50 , wherein the light chain of the antibody comprises the sequence DIQMTQSPSSLSASVGDRVTITCRASQASYYDVAWYQQKPGKAPKLLIYAASYLYSGVPSRFSGSGSGTDFT LTISSLQPEDFATYYCQQYYYAPATFGQGTKVEIK (SEQ ID NO: 950). 
     
     
         54 . The method of any one of  claims 43 ,  46 , and  50 , wherein the antibody is selected from the group consisting of a synthetic antibody, a chimeric antibody, a humanized antibody, an affinity matured antibody, and a bispecific antibody. 
     
     
         55 . The method of any one of  claims 43 ,  46 , and  50 , wherein the antibody is an antibody fragment. 
     
     
         56 . The method of any one of  claims 43 ,  46 , and  50 , wherein the antibody binds to mouse and human VEGF with Kd values of no more than about 10 nM. 
     
     
         57 . The method of any one of  claims 43 ,  46 , and  50 , wherein the cancer is selected from the group consisting of breast cancer, colorectal cancer, non-small cell lung cancer, non-Hodgkin's lymphoma (NHL), renal cancer, prostate cancer, liver cancer, head and neck cancer, melanoma, ovarian cancer, mesothelioma, and glioblastoma. 
     
     
         58 . The method of any one of  claims 43 ,  46 , and  50 , wherein the disease caused by ocular neovascularization is diabetic blindness, retinopathy, primary diabetic retinopathy, or age-induced macular degeneration.