Anti-Glypican-3 Antibody Having Improved Kinetics in Plasma

Abstract

A method of modulating the plasma half-life of anti-glypican 3 antibody, a pharmaceutical composition comprising as an active ingredient the anti-glypican 3 antibody that has a plasma half-life that has been modulated, a method of preparing the anti-glypican 3 antibody and a pharmaceutical composition comprising the anti-glypican 3 antibody as an active ingredient are provided. Disclosed is a method of modulating the plasma half-life of anti-glypican 3 antibody by modifying an amino acid residue that is exposed on the surface of the anti-glypican 3 antibody; and anti-glypican 3 antibody that has a plasma half-life that has been modulated by amino acid residue modification, a pharmaceutical composition comprising as an active ingredient the anti-glypican 3 antibody, and a method of preparing the anti-glypican 3 antibody and producing a pharmaceutical composition comprising the anti-glypican 3 antibody as an active ingredient.

What is claimed is: 
     
         1 . A method for preparing an antibody with modulated cytotoxicity, comprising the steps of:
 (a) culturing a host cell bearing a nucleic acid that encodes the antibody under conditions allowing for expression of the nucleic acid, wherein the antibody has an amino acid sequence altered to causes a modification in the charge of at least one amino acid residue that can be exposed on the surface of a cytotoxic antibody; and   (b) recovering the antibody from the host cell culture.   
     
     
         2 . The method according to  claim 1 , wherein the modification in the charge of the amino acid residue is achieved by amino acid substitution. 
     
     
         3 . The method according to  claim 1 , wherein the amino acid residue that can be exposed on the surface of the antibody is located in a region in the antibody other than the FcRn binding region. 
     
     
         4 . The method according to  claim 3 , wherein the FcRn binding region comprises the Fe region. 
     
     
         5 . The method according to  claim 3 , wherein the FcRn binding region comprises the amino acid residues of the EU numbers 250, 253, 310, 311, 314, 428, 435, 436 according to the Kabat numbering. 
     
     
         6 . The method according to  claim 1 , wherein the antibody is an IgG antibody. 
     
     
         7 . The method according to  claim 2 , wherein the amino acid residue whose charge is modified is an amino acid residue present in the constant region of the antibody. 
     
     
         8 . The method according to  claim 2 , wherein the amino acid residue whose charge is modified is an amino acid residue present in the heavy chain variable region or the light chain variable region of the antibody. 
     
     
         9 . The method according to  claim 8 , wherein the antibody is an antibody that comprises a complementarity-determining region (CDR) derived from a non-human animal, a framework region (FR) derived from human, and a constant region derived from human, and wherein the modification in the charge of the amino acid residue is achieved by substitution of at least one amino acid residue that can be exposed on the antibody surface in the CDR or FR of the antibody with an amino acid residue that has a charge different from that of the amino acid residue. 
     
     
         10 . The method according to  claim 9 , wherein the modification in the charge of the amino acid residue is achieved by:
 (1) at least one substitution in the heavy chain variable region shown in SEQ ID NO: 1 selected from:
 (a) substitution of K that is the 19th amino acid residue with T, 
 (b) substitution of Q that is the 43rd amino acid residue with E, 
 (c) substitution of Q that is the 62nd amino acid residue with E, 
 (d) substitution of K that is the 63rd amino acid residue with s, 
 (e) substitution of K that is the 65th amino acid residue with Q, and 
 (f) substitution of G that is the 66th amino acid residue with D; 
 and/or, 
   (2) at least one substitution in the light chain variable region shown in SEQ ID NO: 7 selected from:
 (g) substitution of R that is the 24th amino acid residue with Q, 
 (h) substitution of Q that is the 27th amino acid residue with E, 
 (i) substitution of K that is the 79th amino acid residue with T, 
 (j) substitution of R that is the 82nd amino acid residue with S, and 
 (k) substitution of K that is the 112nd amino acid residue with E. 
   
     
     
         11 . The method according to  claim 10 , further comprising at least one substitution in the heavy chain constant region shown in SEQ ID NO: 31 selected from:
 (a) substitution of H that is the 151st amino acid residue with Q,   (b) substitution of K that is the 157th amino acid residue with Q,   (c) substitution of R that is the 238th amino acid residue with Q,   (d) substitution of D that is the 239th amino acid residue with E,   (e) substitution of L that is the 241st amino acid residue with M, and   (f) substitution of Q that is the 302nd amino acid residue with E.   
     
     
         12 . The method according to  claim 8 , wherein the antibody comprises a complementarity-determining region (CDR) derived from a non-human animal, a framework region (FR) derived from human, and a constant region derived from human, and wherein the modification in the charge of the amino acid residue is achieved by substitution of at least one amino acid residue that can be exposed on the antibody surface in the CDR or FR of the antibody with an amino acid residue that has a charge different from that of the amino acid residue and substitution of at least one amino acid residue that can be exposed on the antibody surface in the constant region of the antibody with an amino acid residue that has a charge different from that of the amino acid residue. 
     
     
         13 . The method according to  claim 12 , wherein the substitution is at least one substitution in the heavy chain constant region shown in SEQ ID NO: 31 selected from:
 (a) substitution of H that is the 151st amino acid residue with Q,   (b) substitution of K that is the 157th amino acid residue with Q,   (c) substitution of R that is the 238th amino acid residue with Q,   (d) substitution of D that is the 239th amino acid residue with E,   (e) substitution of L that is the 241st amino acid residue with M, and   (f) substitution of Q that is the 302nd amino acid residue with E.   
     
     
         14 . The method according to  claim 1 , wherein the antibody has a reduced content of fucose attached to the Fe region of the antibody.