Tri- or tetraspecific antibodies

Abstract

The present invention relates to tri- or tetraspecific antibodies, their manufacture and use.

1 - 26 . (canceled) 
     
     
         27 . One or more nucleic acid molecules encoding a trispecific or tetraspecific antibody, wherein the antibody comprises:
 a) a light chain and heavy chain of a full length antibody which specifically binds to a first antigen; and   b) a modified light chain and modified heavy chain of a full length antibody which specifically binds to a second antigen, wherein the variable domains VL and VH are replaced by each other, and/or wherein the constant domains CL and CH1 are replaced by each other; and   c) one to four antigen binding peptides fused via a peptide connector to the C- or N-terminus of the light chains or heavy chains of a) and/or b) wherein said antigen binding peptides specifically bind one or two further antigens.   
     
     
         28 . The one or more nucleic acid molecules of  claim 27 , wherein the antigen binding peptides comprise one or two antigen binding peptides which specifically bind to one or two further antigens. 
     
     
         29 . The one or more nucleic acid molecules of  claim 27 , wherein the antigen binding peptides comprise one or two antigen binding peptides which specifically bind to a third antigen. 
     
     
         30 . The one or more nucleic acid molecules of  claim 27 , wherein the antigen binding peptides comprise two identical antigen binding peptides which specifically bind to a third antigen. 
     
     
         31 . The one or more nucleic acid molecules of  claim 27 , wherein the antigen binding peptides comprise one antigen binding peptide which specifically binds to a third antigen and one antigen binding peptide which specifically binds to a fourth antigen. 
     
     
         32 . The one or more nucleic acid molecules of  claim 27 , wherein the antigen binding peptides are selected from the group consisting of a scFv fragment and a scFab fragment. 
     
     
         33 . The one or more nucleic acid molecules of  claim 27 , wherein the antigen binding peptides are scFv fragments. 
     
     
         34 . The one or more nucleic acid molecules of  claim 27 , wherein the antigen binding peptides are scFab fragments. 
     
     
         35 . The one or more nucleic acid molecules of  claim 27 , wherein the antigen binding peptides are fused to the C-terminus of the heavy chains of a) and/or b). 
     
     
         36 . The one or more nucleic acid molecules of  claim 27 , wherein the CH3 domain of the heavy chain of the full length antibody of a) and the CH3 domain of the modified heavy chain of the full length antibody of b) which meet at an interface which has been altered to promote the formation of the trispecific or tetraspecific antibody, wherein the interface of the first CH3 domain comprises a protuberance which is positionable in a cavity in the interface of the second CH3 domain, and wherein the amino acid sequence of an original interface has been altered so as to introduce the protuberance and/or cavity into the altered interface;
 wherein said interface is altered to promote the formation of the trispecific or tetraspecific antibody, wherein the alteration comprises:   i) a CH3 domain of one heavy chain is altered, so that within the original interface of the CH3 domain of one heavy chain that meets the original interface of the CH3 domain of the other heavy chain within the tri- or tetraspecific antibody, an amino acid residue is replaced with an amino acid residue having a larger side chain volume, thereby generating a protuberance within the interface of the CH3 domain of one heavy chain which is positionable in a cavity within the interface of the CH3 domain of the other heavy chain; and   ii) a CH3 domain of the other heavy chain is altered, so that within the original interface of the second CH3 domain that meets the original interface of the first CH3 domain within the tri- or tetraspecific antibody, an amino acid residue is replaced with an amino acid residue having a smaller side chain volume, thereby generating a cavity within the interface of the second CH3 domain within which a protuberance within the interface of the first CH3 domain is positionable.   
     
     
         37 . The one or more nucleic acid molecules of  claim 36 , wherein said amino acid residue having a larger side chain volume is selected from the group consisting of arginine (R), phenylalanine (F), tyrosine (Y), and tryptophan (W) and said amino acid residue having a smaller side chain volume is selected from the group consisting of alanine (A), serine (S), threonine (T), and valine (V). 
     
     
         38 . The one or more nucleic acid molecules of  claim 36 , characterized in that both CH3 domains are further altered by the introduction of cysteine (C) as amino acid in the corresponding positions of each CH3 domain such that a disulfide bridge between both CH3 domains can be formed. 
     
     
         39 . One or more nucleic acid molecules encoding a tetraspecific antibody, wherein the antibody comprises:
 a) a light chain comprising the amino acid sequence of SEQ ID NO: 1;   b) a modified heavy chain comprising the amino acid sequence of SEQ ID NO: 2;   c) a modified light chain comprising the amino acid sequence of SEQ ID NO: 3; and   d) a modified heavy chain comprising the amino acid sequence of SEQ ID NO: 4.   
     
     
         40 . One or more vectors comprising the one or more nucleic acid molecules of  claim 27 . 
     
     
         41 . One or more vectors comprising the one or more nucleic acid molecules of  claim 39 . 
     
     
         42 . A host cell comprising the one or more nucleic acid molecules of  claim 27 . 
     
     
         43 . The host cell of  claim 42 , wherein the host cell comprises one or more vectors comprising the one or more nucleic acid molecules. 
     
     
         44 . A method for the preparation of a trispecific or tetraspecific antibody using the host cell of  claim 42 , comprising:
 (a) culturing the host cell of  claim 42  under conditions that allow synthesis of the antibody; and   (b) recovering the antibody from the culture.   
     
     
         45 . A method for the treatment of a patient in need of therapy, comprising administering to the patient a therapeutically effective amount of an antibody prepared by the method according to  claim 44 . 
     
     
         46 . The method of  claim 45 , wherein the antibody is in a composition comprising the antibody and at least one pharmaceutically acceptable excipient.