Antibody constant domain regions and uses thereof

Abstract

The invention provides recombinant protein (e.g., a recombinant antibody or soluble receptor) having (i) a binding domain capable of specific binding to an epitope (for example an antibody variable domains, a receptor, a growth factor, a cytokine, or a fragment of any of the foregoing which is capable of specifically binding the desired epitope) and (ii) an effector domain comprising a constant domain region which is derived from immunoglobulin of a first species which is a companion mammal, e.g. dog, cat, or horse, having engineered substitutions at one or more positions and having an altered interaction with one or more FcRs or other ligands, and optionally enhanced effector function, relative to the parent constant domain region.

1 . A recombinant protein comprising
 a. a binding domain, or a fragment thereof, that specifically binds to an epitope; and   b. an effector domain comprising a constant domain region; wherein the constant domain region is derived from immunoglobulin of a companion mammal, selected from dogs, cats and horses, comprising a deletion, addition, or substitution for at least one amino acid position and has an altered enhanced biophysical property or effector function interaction with at least one ligand.   
     
     
         2 . The recombinant protein of  claim 1  which is a member of the immunoglobulin superfamily comprising an Ig fold. 
     
     
         3 . The recombinant protein of  claim 1 , wherein the binding domain comprises
 a. an antibody variable region or an epitope-binding fragment thereof, or   b. a soluble receptor binding domain or an epitope-binding fragment thereof.   
     
     
         4 . The recombinant protein of  claim 1 , wherein the epitope is selected from the group consisting of: a receptor, a growth factor, and a cytokine. 
     
     
         5 . The recombinant protein of  claim 1 , wherein the companion animal is a dog. 
     
     
         6 . The recombinant protein of  claim 1 , wherein the effector domain binds to at least one ligand is an FcR. 
     
     
         7 . The recombinant protein of  claim 1 , wherein the effector domain has altered effector function when compared to a parent constant domain region. 
     
     
         8 . The recombinant protein of  claim 1 , wherein the effector function is antibody-dependent cell-mediated cytotoxicity (ADCC) or complement dependent cytotoxicity (CDC). 
     
     
         9 . The recombinant protein of  claim 1 , wherein the recombinant protein has increased ADCC effector function relative to dog gamma globulin. 
     
     
         10 . The recombinant protein of  claim 1 , wherein the recombinant protein has increased CDC effector function relative to dog gamma globulin. 
     
     
         11 . The recombinant protein of  claim 1 , wherein the recombinant protein exhibits a TM of at least about the TM of dog gamma globulin. 
     
     
         12 . The recombinant protein of  claim 1 , wherein the constant domain region comprises an amino acid sequence selected from SEQ ID NOs:1-7. 
     
     
         13 . The recombinant protein of  claim 12 , wherein the effector domain further comprises a constant domain region of a kappa light chain comprising the sequence set forth in SEQ ID NO:8. 
     
     
         14 . The recombinant protein of  claim 12 , wherein the effector domain further comprises a constant domain region of a lambda light chain comprising the sequence set forth in SEQ ID NO:9. 
     
     
         15 . The recombinant protein of  claim 1  wherein the constant domain comprises any of SEQ ID NOS: 10-21. 
     
     
         16 . A recombinant protein according to  claim 1  wherein the binding domain comprises hypervariable regions from an antibody from a species other than the companion animal. 
     
     
         17 . A recombinant protein according to  claim 16  which is a heterochimeric antibody. 
     
     
         18 . A polynucleotide encoding the recombinant protein of  claim 1 . 
     
     
         19 . A method of treating a disease or condition in a companion animal selected from dogs, cats, and horses, comprising administering to a companion animal in need of such treatment a therapeutically effective amount of a recombinant protein of  claim 1 , thereby treating the disease or disorder. 
     
     
         20 . The method of  claim 19 , wherein the companion animal is a dog. 
     
     
         21 . The method of  claim 19 , wherein the disease or condition is selected from an inflammatory disorder and cancer. 
     
     
         22 . The method of  claim 19  further comprising administering a therapeutically effective amount of a second agent. 
     
     
         23 . The method of  claim 22  wherein the second agent is a chemotherapeutic agent. 
     
     
         24 . A pharmaceutical composition comprising the recombinant protein according to  claim 1 , in combination or association with a pharmaceutically acceptable diluent of carrier. 
     
     
         25 . A recombinant protein comprising any of the sequences of SEQ ID NOS 1-9. 
     
     
         26 . A nucleic acid encoding a protein according to  claim 25 . 
     
     
         27 . A recombinant protein comprising any of the sequences of SEQ ID NOS 10-21. 
     
     
         28 . A nucleic acid encoding a protein according to  claim 27 . 
     
     
         29 . A method of engineering an antibody for use in treating a dog, cat or horse, comprising:
 a. Isolating spleen cells or peripheral blood mononuclear cells from the species to be treated;   b. Making cDNA in said cells and amplifying the cDNA encoding the antibody constant domains from said cells;   c. Transforming a host cell with an expression vector comprising cDNA for the constant domain together with DNA encoding a variable region having affinity for an epitope;   d. Expressing and retrieving antibody from the host cell thus transformed, and   e. Assessing the effector function of the constant regions of antibodies thus obtained,
 i. by directly assessing the binding of the antibody to Fc; and/or 
 ii. by assessing the ability of the antibody to kill a target cell, wherein the variable region of the antibody recognizes an epitope on the target cell, in the absence of complement (ADCC) and/or presence of complement (CDC); 
   f. Selecting a constant region having the desired effector function;   g. Making an antibody comprising the selected constant region and a variable region having affinity for the desired target.   
     
     
         30 . An antibody engineered by the method of  claim 29 . 
     
     
         31 . A recombinant protein comprising
 a. a binding domain, or a fragment thereof, that specifically binds to an epitope; the binding domain comprising (i) a non-canine antibody hypervariable region or an epitope-binding fragment thereof, or (ii) a soluble receptor binding domain or an epitope-binding fragment thereof; and   b. an effector domain that provides ADCC and/or CDC effector function to the recombinant protein; the effector domain comprising a constant domain region; wherein the constant domain exhibits at least 80% amino acid sequence identity to any of SEQ ID NO: 1-7.   
     
     
         32 . The recombinant protein of  claim 31  wherein the constant domain region exhibits at least 90% amino acid sequence identity to any of SEQ ID NO: 1-7. 
     
     
         33 . The recombinant protein of  claim 31  wherein the constant domain region exhibits at least 95% amino acid sequence identity to any of SEQ ID NO: 1-7. 
     
     
         34 . The recombinant protein of  claim 31  wherein the constant domain region comprises SEQ ID NO: 1. 
     
     
         35 . The recombinant protein of  claim 31  wherein the constant domain region comprises SEQ ID NO: 2. 
     
     
         36 . The recombinant protein of  claim 31  wherein the constant domain region comprises SEQ ID NO: 4. 
     
     
         37 . The recombinant protein of  claim 31  wherein the constant domain region comprises SEQ ID NO: 6. 
     
     
         38 . The recombinant protein of  claim 31  wherein the constant domain region comprises SEQ ID NO: 7.