Humanized anti-egfl7 antibodies and methods using same

Abstract

The present invention concerns antibodies to EGFL 7 and the uses of same.

What is claimed is: 
     
         1 . An anti-EGFL7 antibody comprising the following six hypervariable region (HVR) sequences:
 (i) HVR-L1 comprising KX 1 SX 2 SX 3 DYX 4 GDSYX 5 S, wherein X 1  is A or R; X 2  is H or Q; X 3  is G or V; X 4  is selected from the group consisting of D, L, R, S, and W; and X 5  is M or V (SEQ ID NO: 210);   (ii) HVR-L2 comprising GASX 1 X 2 EX 3 , wherein X 1  is N or Y; X 2  is selected from the group consisting of L, R and Y; and X 3  is Q or S (SEQ ID NO: 211);   (iii) HVR-L3 comprising QQNNEX 1 PX 2 T, wherein X 1  is D or E; and X 2  is F or Y (SEQ ID NO: 212);   (iv) HVR-H1 comprising GX 1 X 2 X 3 X 4 TYGX 5 S, wherein X 1  is H or V; X 2  is R or T; X 3  is selected from the group consisting of F, G, R, and S; X 4  is selected from the group consisting of D, G, R, and T; and X 5  is M or Y (SEQ ID NO: 213);   (v) HVR-H2 comprising GWINX 1 X 2 SGVPTX 3 AX 4 X 5 X 6 X 7 X 8 , wherein X 1  is selected from the group consisting of I, M, T, and W; X 2  is H or R; X 3  is selected from group consisting of I, M, T, and Y; X 4  is D or H; X 5  is selected from group consisting of D, M and T; X 6  is F or Y; X 7  is K or S; and X 8  is G or R (SEQ ID NO: 214, and   (vi) HVR-H3 comprising AX 1 LGSX 2 AVDX 3 , wherein X 1  is N or R; X 2  is selected from the group consisting of C, R, H, and Y; and X 3  is A or Y (SEQ ID NO: 215).   
     
     
         2 . An anti-EGFL7 antibody comprising the following six HVR sequences:
 HVR-L1 comprising an amino acid sequence selected from SEQ ID NOs: 37-43 and 234; HVR-L2 comprising an amino acid sequence selected from the group consisting of SEQ ID NOs: 32, 44-47, and 235; HVR-L3 comprising an amino acid sequence selected from the group consisting of SEQ ID NO: 33, 48, and 236; HVR-H1 comprising an amino acid sequence selected from the group consisting of SEQ ID NOs: 34 and 49-57; HVR-H2 comprising an amino acid sequence selected from the group consisting of SEQ ID NOs: 35 and 58-73; and HVR-H3 comprising an amino acid sequence selected from the group consisting of SEQ ID NOs: 36, 74-77, and 237.   
     
     
         3 . The antibody of  claim 1 , wherein HVR-L1 comprises the amino acid sequence KASQSVDYSGDSYMS (SEQ ID NO: 234), HVR-L2 comprises the amino acid sequence GASYRES (SEQ ID NO: 235), HVR-L3 comprises the amino acid sequence QQNNEEPYT (SEQ ID NO: 236), HVR-H1 comprises the amino acid sequence GHTFTTYGMS (SEQ ID NO: 34), HVR-H2 comprises the amino acid sequence GWINTHSGVPTYADDFKG (SEQ ID NO: 35), and HVR-H3 comprises the amino acid sequence ARLGSYAVDY (SEQ ID NO: 237). 
     
     
         4 . The antibody of  claim 1 , wherein the heavy chain comprises the following framework sequences: FR-H1 comprises EX 1 QLVESGGGLVQPGGSLRLSCAAS, wherein X 1  is I or V (SEQ ID NO: 216); FR-H2 comprises WVRQAPGKGLEWX 1 , wherein X 1  is I, M, or V (SEQ ID NO: 217); FR-H3 comprises
 RFTX 1 SX 2 DX 3 SX 4 X 5 TX 6 YLQMNSLRAEDTAVYX 7 CAR, wherein X 1  is F or I; X 2  is L or R; X 3  is N or T, X 4  is selected from the group consisting of A, E, K and T; X 5  is N or S; X 6  is selected from the group consisting of A, L, M, T and V; and X 7  is F or Y (SEQ ID NO: 218); and FR-H4 comprises WGQGTLVTVSS (SEQ ID NO: 219).   
     
     
         5 . The antibody of  claim 4 , wherein the heavy chain comprises the following framework sequences: FR-H1 comprises EVQLVESGGGLVQPGGSLRLSCAAS (SEQ ID NO: 197); FR-H2 comprises WVRQAPGKGLEWV (SEQ ID NO: 198); FR-H3 comprises RFTISX 1 DNSKNTX 2 YLQMNSLRAEDTAVYYCAR, wherein X 1  L or R; X 2  is selected from the group consisting of A, L, M, T and V (SEQ ID NO: 220); and FR-H4 comprises WGQGTLVTVSS (SEQ ID NO: 200). 
     
     
         6 . The antibody of  claim 4 , wherein the heavy chain comprises the following framework sequences: FR-H1 comprises EIQLVESGGGLVQPGGSLRLSCAAS (SEQ ID NO: 238); FR-H2 comprises WVRQAPGKGLEWM (SEQ ID NO: 239); FR-H3 comprises RFTISX 1 DNSKSTX 2 YLQMNSLRAEDTAVYFCAR, wherein X 1  is L or R; X 2  is selected from the group consisting of A, L, M, T and V (SEQ ID NO: 240); and FR-H4 comprises WGQGTLVTVSS (SEQ ID NO: 219). 
     
     
         7 . The antibody of  claim 1 , wherein light chain comprises the following framework sequences: FR-L1 comprises DIQMTQSPSSLSASVGDRVTITC (SEQ ID NO: 201), FR-L2 comprises WYQQKPGKAPKLLIY (SEQ ID NO: 202), FR-L3 comprises GVPSRFSGSGSGTDFTLTISSLQPEDFATYYC (SEQ ID NO: 203), FR-L4 comprises FGQGTKVEIK (SEQ ID NO: 221) or FGQGTKVEIKR (SEQ ID NO: 204). 
     
     
         8 . The antibody of  claim 1 , wherein the light chain comprises the variable domain sequence of 4F11.v17 as shown in  FIG. 15  (SEQ ID NO: 82) or 4F11.v22 as shown in  FIG. 15  (SEQ ID NO: 83). 
     
     
         9 . The antibody of  claim 1 , wherein the heavy chain comprises the variable domain sequence of 4F11.v17 as shown in  FIG. 16  (SEQ ID NO: 84) or 4F11.v22 as shown in  FIG. 16  (SEQ ID NO: 85). 
     
     
         10 . The antibody of  claim 1 , wherein the light chain comprises the variable domain sequence of 4F11.v17 as shown in  FIG. 15  (SEQ ID NO: 82) and the heavy chain comprises the variable domain sequence of 4F11.v17 as shown in  FIG. 16  (SEQ ID NO: 84). 
     
     
         11 . The antibody of  claim 1 , wherein the light chain comprises the variable domain sequence of 4F11.v22 as shown in  FIG. 15  (SEQ ID NO: 83) and the heavy chain comprises the variable domain sequence of 4F11.v22 as shown in  FIG. 16  (SEQ ID NO: 85). 
     
     
         12 . The antibody of  claim 1 , wherein at least a portion of the framework sequence is a human consensus framework sequence. 
     
     
         13 . An anti-EGFL7 antibody comprising the following six HVR sequences:
 (i) HVR-L1 comprising X 1 X 2 X 3 X 4 X 5 X 6 VX 7 X 8 X 9 X 10 ITYLX 11 , wherein X 1  is selected from the group consisting of L, Q, R, S, and T; X 2  is selected from the group consisting of P, T, and W; X 3  is H or S; X 4  is D or Q; X 5  is G or S; X 6  is L or V; X 7  is H or P; X 8  is selected from the group consisting of I, L, P, T, and Y; X 9  is selected from the group consisting of N, Q or S; X 10  is selected from the group consisting of A, G, and S; and X 11  is G or H (SEQ ID NO: 222);   (ii) HVR-L2 comprising RVSNX I X 2 S, wherein X 1  is D or R; and X 2  is selected from the group consisting of A, G, F, I, and T (SEQ ID NO: 223);   (iii) HVR-L3 comprising X 1 QSX 2 X 3 VPLT, wherein X 1  is selected from the group consisting of A, G, I, K, L, N, T, and V; X 2  is C or T; and X 3  is F or H (SEQ ID NO: 224);   (iv) HVR-H1 comprising GYX 1 X 2 X 3 DX 4 YX 5 N, wherein X 1  is N or T; X 2  is F or V; X 3  is selected from the group consisting of I, M, R, and S; X 4  is selected from the group consisting of Y, Q, and K; and X 5  is I or M (SEQ ID NO: 225);   (v) HVR-H2 comprising GDINX 1 X 2 X 3 X 4 X 5 X 6 HX 7 X 8 X 9 X 10 X 11 X 12 X 13 , wherein X 1  is selected from the group consisting of A, L, N, and P; X 2  is selected from the group consisting of D, L, and R; X 3  is selected from the group consisting of G, K, N, R, S, and Y; X 4  is G or S; X 5  is selected from the group consisting of G, I, K, R, S, T, and V; X 6  is selected from the group consisting of G, R, and T; X 7  is selected from the group consisting of I, V, L, and Y; X 8  is N or S; X 9  is selected from the group consisting of A, N, and Q; X 10  is K or V; X 11  is F or Q; X 12  is K or T; and X 13  is selected from the group consisting of G, H, R, and S (SEQ ID NO: 226); and   (vi) HVR-H3 comprising X 1 REGVYHX 2 YDDYAX 3 DY, wherein X 1  is selected from the group consisting of A, N, and T; X 2  is D or P; and X 3  is M or W (SEQ ID NO: 227).   
     
     
         14 . An anti-EGFL7 antibody comprising the following HVR sequences:
 HVR-L1 comprising an amino acid sequence selected from SEQ ID NOs: 106-124 and 241; HVR-L2 comprising an amino acid sequence selected from the group consisting of SEQ ID NOs: 101 and 125-129; HVR-L3 comprising an amino acid sequence selected from the group consisting of SEQ ID NOs: 102 and 130-145; HVR-H1 comprising an amino acid sequence selected from the group consisting of SEQ ID NOs: 103 and 146-153; HVR-H2 comprising an amino acid sequence selected from the group consisting of SEQ ID NOs: 104, 154-187, and 247; and HVR-H3 comprising an amino acid sequence selected from the group consisting of SEQ ID NOs: 105 and 188-192.   
     
     
         15 . The antibody of  claim 13 , wherein the heavy chain comprises the following framework sequences: FR-H1 comprises EVQLVESGGGLVQPGGSLRLSCAAS (SEQ ID NO: 197); FR-H2 comprises WVRQAPGKGLEWX 1 , wherein X 1  is I or V (SEQ ID NO: 228); FR-H3 comprises RX 1 TX 2 SX 3 DX 4 SX 5 X 6 TX 7 YX 8 QMNSLRAEDTAVYYC, wherein X 1  is F or V; X 2  is I or L; X 3  is selected from the group consisting of L, R, and V; X 4  is K or N; X 5  is selected from the group consisting of K, N, R, and S; X 6  is N or S; X 7  is selected from the group consisting of A, L, and V; and X 8  is L or M (SEQ ID NO: 229); and FR-H4 comprises WGQGTLVTVSS (SEQ ID NO: 200). 
     
     
         16 . The antibody of  claim 15 , wherein the heavy chain comprises the following framework sequences: FR-H1 comprises EVQLVESGGGLVQPGGSLRLSCAAS (SEQ ID NO: 197); FR-H2 comprises WVRQAPGKGLEWV (SEQ ID NO: 198); FR-H3 comprises RFTISRDX 1 SKNTX 2 YLQMNSLRAEDTAVYYCAR, wherein X 1  is N or K; and X 2  is selected from the group consisting of A, L, and V (SEQ ID NO: 230); and FR-H4 comprises WGQGTLVTVSS (SEQ ID NO: 200). 
     
     
         17 . The antibody of  claim 13 , wherein light chain comprises the following framework sequences: FR-L1 comprises DIQMTQSPSSLSASVGDRVTITC (SEQ ID NO: 201), FR-L2 comprises WYQQKPGKAPKLLIY (SEQ ID NO: 202), FR-L3 comprises GVPSRFSGSGSGTDFTLTISSLQPEDFATYYC (SEQ ID NO: 203), FR-L4 comprises FGQGTKVEIK (SEQ ID NO: 221) or FGQGTKVEIKR (SEQ ID NO: 204). 
     
     
         18 . The antibody of  claim 13 , wherein at least a portion of the framework sequence is a human consensus framework sequence. 
     
     
         19 . The antibody of  claim 1  or  13 , wherein said antibody is a bispecific antibody. 
     
     
         20 . The bispecific antibody of  claim 19 , wherein said bispecific antibody binds to vascular endothelial growth factor (VEGF).