Humanized antibodies against cd3

Abstract

The invention provides bispecific antibodies with selective cytotoxicity against malignant B-cells. The bispecific antibodies bind to an effector cell antigen and to a 28/32 kDa heterodimeric protein on the surface of malignant B-cells. The invention also includes the monospecific components of the bispecific antibodies, humanized versions thereof, and humanized bispecific antibodies. The invention further provides therapeutic and diagnostic methods employing these antibodies.

1 . A bispecific antibody that binds to: 
 (a) a first antigen on the surface of effector cells selected from the group consisting of T-cells and natural killer cells, and    (b) a second antigen on a 28/32 kDa heterodimeric protein on the surface of malignant B cells, which second antigen specifically binds to an antibody designated 1D10,    wherein the binding of the bispecific antibody to the first and second antigens results in killing of the malignant B cells.    
     
     
         2 - 15 . (canceled)  
     
     
         16 . A humanized antibody, the antibody comprising a humanized heavy chain and a humanized light chain: 
 (1) the humanized light chain comprising three complementarity determining regions (CDR1, CDR2 and CDR3) having amino acid sequences from the corresponding complementarity determining regions of the mouse M291 immunoglobulin light chain, and a variable region framework from a human kappa light chain variable region framework sequence, and 
 (2) the humanized heavy chain comprising three complementarity determining regions (CDR1, CDR2 and CDR3) having amino acid sequences from the corresponding complementarity determining regions of the mouse M291 immunoglobulin heavy chain, and a variable region framework from a human heavy chain variable region framework sequence except in at least one position selected from a second group consisting of H30, H67, H68, H70, H72 and H74 wherein the amino acid position is occupied by the same amino acid present in the equivalent position of the mouse M291 immunoglobulin heavy chain variable region framework;  
   wherein the immunoglobulin specifically binds to a CD3 antigen on the surface of T cells with a binding affinity having a lower limit of about 10 7  M −1  and an upper limit of about five-times the binding affinity of the M291 immunoglobulin.    
     
     
         17 . The humanized antibody of  claim 16 , wherein the humanized light chain variable region framework is from the light chain variable region framework of the HF2-1/17 antibody in subgroup I; 
 the humanized heavy chain region framework is from the heavy chain region variable framework of the 21/28 antibody except in at least one position selected from the second group, and except at position 44, wherein the amino acid position is occupied by the same amino acid present in the equivalent position of a human immunoglobulin subgroup I consensus sequence.    
     
     
         18 . The humanized antibody of  claim 17 , wherein the humanized light chain comprises the amino acid sequence of  FIG. 5A  (upper) and 
 the humanized heavy chain comprises the amino acid sequence of  FIG. 5B  (upper).    
     
     
         19 - 33 . (canceled)  
     
     
         34 . A method of treating or preventing a disorder of the immune system, comprising administering a therapeutically or prophylactically effective dose of a humanized anti-CD3 antibody to a patient affected by or at risk of the disorder.  
     
     
         35 . The method according to  claim 34 , wherein the disorder of the immune system is an autoimmune disease, inflammation, graft vs. host disease, or host vs. graft disease.  
     
     
         36 . The method of  claim 35 , wherein the disorder of the immune system is graft vs. host disease.  
     
     
         37 . The method according to  claim 34 , wherein the humanized antibody comprises a pair of humanized heavy chains and humanized light chains, wherein the humanized light chain variable region comprises the amino acid sequence of  FIG. 5A  (upper lines) (SEQ. ID. No. 8) and the humanized heavy chain variable region comprises the amino acid sequence of  FIG. 5B  (upper lines) (SEQ. ID. No. 10)  
     
     
         38 . The method according to  claim 34 , wherein the antibody comprises a constant domain of the IgG2 isotype.  
     
     
         39 . The method according to  claim 34 , wherein the antibody has an affinity for the CD3 antigen having a lower limit of about 10 7  M −1  and an upper limit of five-times the binding affinity of the M291 immunoglobulin, wherein the mouse M291 antibody has a heavy chain with a variable region of sequence SEQ. ID. No. 11 and a light chain with a variable region of sequence SEQ. ID. No. 9.  
     
     
         40 . The method according to  claim 34 , wherein the antibody comprises a humanized heavy chain and a humanized light chain: 
 (1) the humanized light chain comprising three complementarity determining regions, CDR1, CDR2 and CDR3, of the mouse M291 immunoglobulin light chain, and a variable region framework from a human kappa light chain variable region framework sequence, and    (2) the humanized heavy chain comprising three complementarity determining regions, CDR1, CDR2 and CDR3, of the mouse M291 immunoglobulin heavy chain, and a variable region framework from a human heavy chain variable region framework sequence provided that at least one position selected from a group consisting of H30, H67, H68, H70, H72 and H74 is occupied by the same amino acid present in the equivalent position of the mouse M291 immunoglobulin heavy chain variable region framework;    wherein the immunoglobulin specifically binds to a CD3 antigen on the surface of T cells with a binding affinity having a lower limit of about 10 7 M −1  and an upper limit of about five-times the binding affinity of the M291 immunoglobulin wherein the mouse antibody has an IgG1 heavy chain with a variable domain designated SEQ. ID. No. 11 and a kappa light chain with a variable domain designated SEQ. ID. No. 9.    
     
     
         41 . The method according to  claim 34 , wherein the antibody is administered subcutaneously, intramuscularly or intravenously.  
     
     
         42 . The method according to  claim 34 , wherein the patient is affected by the disorder and the dose is 0.01 to 100 mg  
     
     
         43 . The method according to  claim 34 , wherein the patient is affected by the disorder and the dose is 1-10 mg.  
     
     
         44 . The method according to  claim 34 , wherein the patient is at risk of the disorder and the dose is 0.1 to 100 mg.  
     
     
         45 . The method according to  claim 34 , wherein the patient is at risk of the disorder and the dose is 1 to 10 mg.  
     
     
         46 . The method of  claim 34 , wherein the humanized antibody is a bispecific antibody comprising: 
 a first Fab′fragment comprising the humanized heavy chain variable region comprises the amino acid sequence of  FIG. 5B  (upper lines) (SEQ. ID. No. 10) and the humanized light chain variable region comprises the amino acid sequence of  FIG. 5A  (upper lines) (SEQ. ID. No. 8);    a second Fab′ fragment comprising the heavy chain variable region shown in  FIG. 4B  (upper) (SEQ ID NO. 3) and the light chain variable region shown in  FIG. 4A  (upper) (SEQ ID NO. 1);    wherein the first Fab′fragment specifically binds to the CD3 antigen and the second Fab′ fragment specifically binds to the 28/32 kDa heterodimeric antigen on the surface of the malignant B cells.