STABLE HETERODIMERIC ANTIBODY DESIGN WITH MUTATIONS IN THE Fc DOMAIN

Abstract

The provided scaffolds have heavy chains that are asymmetric in the various domains (e.g. CH2 and CH3) to accomplish selectivity between the various Fc receptors involved in modulating effector function, beyond those achievable with a natural homodimeric (symmetric) Fc molecule, and increased stability and purity of the resulting variant Fc heterodimers. These novel molecules comprise complexes of heterogeneous components designed to alter the natural way antibodies behave and that find use in therapeutics.

1 .- 36 . (canceled) 
     
     
         37 . An isolated heteromultimer comprising a modified heterodimeric CH3 domain comprising:
 a first CH3 domain polypeptide and a second CH3 domain polypeptide, each independently comprising amino acid modifications as compared to a wild-type CH3 domain polypeptide, wherein the amino acid modifications promote formation of a heterodimeric CH3 domain as compared to a homodimeric CH3 domain;   wherein at least one of the first and second CH3 domain polypeptides comprises a T350X modification, wherein X is selected from valine, isoleucine, leucine and methionine;   
       wherein
 the first CH3 domain polypeptide comprises amino acid modifications at positions F405 and Y407, and the second CH3 domain polypeptide comprises an amino acid modification at position T394, wherein the amino acid modification at position F405 is F405A, F405S, F405T or F405V; the amino acid modification at position Y407 is Y407I or Y407V, and the amino acid modification at position T394 is T394W; 
 wherein the modified heterodimeric CH3 domain is comprised by an Fc construct based on a type G immunoglobulin (IgG), and 
 wherein the numbering of amino acid residues is according to the EU index as set forth in Kabat. 
 
     
     
         38 . Nucleic acid encoding the isolated heteromultimer according to  claim 37 . 
     
     
         39 . A vector comprising the nucleic acid according to  claim 38 . 
     
     
         40 . A mammalian host cell comprising nucleic acid encoding the isolated heteromultimer according to  claim 37 . 
     
     
         41 . A composition comprising the isolated heteromultimer according to  claim 37  and a pharmaceutically acceptable carrier. 
     
     
         42 . (canceled) 
     
     
         43 . The isolated heteromultimer according to  claim 37 , wherein the T350X modification is a T350V modification. 
     
     
         44 . The isolated heteromultimer according to  claim 43 , wherein both the first and second CH3 domain polypeptides comprise a T350V modification. 
     
     
         45 . The isolated heteromultimer according to  claim 37 , wherein the second CH3 domain polypeptide further comprises an amino acid modification at position T366 selected from T366I, T366L, T366M and T366V. 
     
     
         46 . The isolated heteromultimer according to  claim 45 , wherein the amino acid modification at position T366 is T366I or T366L. 
     
     
         47 . The isolated heteromultimer according to  claim 37 , wherein the second CH3 domain polypeptide further comprises an amino acid modification at position K392 selected from K392F, K392L and K392M. 
     
     
         48 . The isolated heteromultimer according to  claim 47 , wherein the amino acid modification at position K392 is K392L or K392M. 
     
     
         49 . The isolated heteromultimer according to  claim 37 , wherein the first CH3 domain polypeptide or the second CH3 domain polypeptide or both the first and second CH3 domain polypeptides further comprises the amino acid modification L351Y. 
     
     
         50 . The isolated heteromultimer according to  claim 37 , wherein the amino acid modification at position F405 is F405A. 
     
     
         51 . The isolated heteromultimer according to  claim 37 , wherein the amino acid modification at position Y407 is Y407V. 
     
     
         52 . The isolated heteromultimer according to  claim 37 , wherein the amino acid modification at position F405 is F405A and the amino acid modification at position Y407 is Y407V. 
     
     
         53 . The isolated heteromultimer according to  claim 52 , wherein the second CH3 domain polypeptide further comprises:
 (a) an amino acid modification at position T366 selected from T366I, T366L, T366M and T366V, or   (b) an amino acid modification at position K392 selected from K392F, K392L and K392M, or   (c) an amino acid modification at position T366 selected from T366I, T366L, T366M and T366V, and an amino acid modification at position K392 selected from K392F, K392L and K392M.   
     
     
         54 . The isolated heteromultimer according to  claim 53 , wherein the amino acid modification at position T366 is T366I or T366L. 
     
     
         55 . The isolated heteromultimer according to  claim 53 , wherein the amino acid modification at position K392 is K392L or K392M. 
     
     
         56 . The isolated heteromultimer according to  claim 37 , wherein:
 (a) the first CH3 domain polypeptide further comprises the amino acid modification S400E; or   (b) the second CH3 domain polypeptide further comprises the amino acid modification N390R; or   (c) the first CH3 domain polypeptide further comprises the amino acid modification S400E and the second CH3 domain polypeptide further comprises the amino acid modification N390R.   
     
     
         57 . The isolated heteromultimer according to  claim 37 , wherein:
 (a) the first CH3 domain polypeptide further comprises the amino acid modification S400R; or   (b) the second CH3 domain polypeptide further comprises the amino acid modification N390D or N390E; or   (c) the first CH3 domain polypeptide further comprises the amino acid modification S400R and the second CH3 domain polypeptide further comprises the amino acid modification N390D or N390E.   
     
     
         58 . The isolated heteromultimer according to  claim 37 , wherein the IgG is IgG1. 
     
     
         59 . The isolated heteromultimer according to  claim 58 , wherein the IgG1 is a human IgG1. 
     
     
         60 . The isolated heteromultimer according to  claim 37 , wherein the heteromultimer is a bispecific antibody or a multispecific antibody. 
     
     
         61 . The isolated heteromultimer according to  claim 37 , wherein the heteromultimer comprises at least one antigen-binding domain. 
     
     
         62 . The isolated heteromultimer according to  claim 61 , wherein the antigen-binding domain binds a cancer antigen.