Therapeutic canine immunoglobulins and methods of using same

Abstract

A method of preparing a canine antibody suitable for use in the therapeutic treatment of a canine is provided. In particular, there is provided immunoglobulins which can be selected for the characteristic of whether they mediate downstream complement mediated immune activation when bound to a target antigen. Canine derived antibodies comprising specific heavy chain isotypes are provided. The invention extends to the use of the immunoglobulins of the invention in methods of treating conditions such as pain, inflammatory conditions and cancerous conditions in a canine.

1 .- 48 . (canceled) 
     
     
         49 . An antibody or fusion protein, or a binding fragment thereof, wherein the antibody, fusion protein or binding fragment specifically binds to a target antigen and has a heavy chain constant domain comprising the amino acid sequence of SEQ ID NO:8, SEQ ID NO:11 or SEQ ID NO:13, wherein the heavy chain constant domain minimizes activation of downstream immune system effector functions when the antibody, fusion protein or binding fragment is bound to the target antigen 
     
     
         50 . The antibody, fusion protein or binding fragment of  claim 49 , wherein the downstream immune system effector functions are selected from complement dependent cytotoxicity, antibody dependent cell mediated cytotoxicity, and antibody dependent cellular pathogenesis. 
     
     
         51 . A method for treating, inhibiting or ameliorating pain or inflammation in a canine subject in need thereof, comprising administering a therapeutically effective amount of an antibody, fusion protein, or binding fragment thereof according to  claim 49 , wherein the heavy chain constant domain minimizes the activation of downstream immune system effector functions when the antibody, fusion protein or binding fragment is bound to the target antigen. 
     
     
         52 . The method of  claim 51 , wherein the pain is selected from the group consisting of neuropathic pain, oncologic pain, pain associated with, or resulting from, rheumatoid arthritis, pain associated with, or resulting from, osteoarthritis, pain associated with, or resulting from, inflammation and pain associated with, or resulting from, pruritis. 
     
     
         53 . The method of  claim 51 , wherein the downstream immune system effector functions are selected from complement dependent cytotoxicity, antibody dependent cell mediated cytotoxicity, and antibody dependent cellular pathogenesis. 
     
     
         54 . An antibody or fusion protein, or a binding fragment thereof, wherein the antibody, fusion protein or binding fragment specifically binds to a target antigen and has a heavy chain constant domain comprising the amino acid sequence of SEQ ID NO:9, SEQ ID NO:10, or SEQ ID NO:14 wherein the heavy chain constant domain mediates activation of downstream immune system effector functions when the antibody, fusion protein or binding fragment is bound to the target antigen. 
     
     
         55 . The antibody, fusion protein, or binding fragment of  claim 54 , wherein the downstream immune system effector functions are selected from complement dependent cytotoxicity, antibody dependent cell mediated cytotoxicity, and antibody dependent cellular pathogenesis. 
     
     
         56 . A method for the treatment or prevention of a cancerous or malignant condition in a canine subject in need thereof, comprising administering a therapeutically effective amount of an antibody, fusion protein, or binding fragment according to  claim 54 , wherein the heavy chain constant domain mediates activation of downstream immune system effector functions when the antibody, fusion protein or binding fragment is bound to the target antigen. 
     
     
         57 . The method of  claim 56 , wherein the downstream immune system effector functions are selected from complement dependent cytotoxicity, antibody dependent cell mediated cytotoxicity, and antibody dependent cellular pathogenesis. 
     
     
         58 . An antibody or fusion protein, or a binding fragment thereof, wherein the antibody, fusion protein, or binding fragment specifically binds to a target antigen and has a heavy chain constant domain which does not bind to C1q when the antibody, fusion protein or binding fragment is bound to its target antigen, and wherein the antibody, fusion protein or binding fragment can be purified using Protein A chromatography. 
     
     
         59 . The antibody, fusion protein or binding fragment of  claim 58 , wherein the antibody has a heavy chain constant domain comprising the amino acid sequence of SEQ ID NO:6 or SEQ ID NO:13. 
     
     
         60 . A method for treating, inhibiting or ameliorating pain or inflammation in a canine subject in need thereof, comprising administering a therapeutically effective amount of the antibody, fusion protein, or binding fragment of  claim 58 . 
     
     
         61 . The method of  claim 60 , wherein the antibody, fusion protein, or binding fragment has a heavy chain constant domain comprising the amino acid sequence of SEQ ID NO:6 or SEQ ID NO:13. 
     
     
         62 . A pharmaceutical composition comprising the antibody, fusion protein, or binding fragment of  claim 49 , and at least one pharmaceutically acceptable carrier, excipient or diluent. 
     
     
         63 . A pharmaceutical composition comprising the antibody, fusion protein, or binding fragment of  claim 54 , and at least one pharmaceutically acceptable carrier, excipient or diluent. 
     
     
         64 . A pharmaceutical composition comprising the antibody, fusion protein, or binding fragment of  claim 58 , and at least one pharmaceutically acceptable carrier, excipient or diluent. 
     
     
         65 . A method for the purification of the antibody, fusion protein, or binding fragment of  claim 49  from a source mixture, wherein the antibody, fusion protein or binding fragment thereof comprises a canine heavy chain constant domain of isotype A having an amino acid sequence of SEQ ID NO:8 or a canine immunoglobulin heavy chain constant domain of isotype D having an amino acid sequence of SEQ ID NO:11, comprising:
 (i) subjecting a source mixture comprising target immunoglobulins or fusion proteins, or antigen-binding fragments thereof, to anion exchange chromatography; 
 (ii) subjecting the source mixture to hydrophobic interaction chromatography; and 
 (iii) subjecting the source mixture to size exclusion chromatography. 
 
     
     
         66 . A method for the purification of the antibody, fusion protein, or binding fragment of  claim 49  from a source mixture, wherein the antibody, fusion protein or binding fragment thereof comprises a canine heavy chain constant domain of isotype A having an amino acid sequence of SEQ ID NO:8 or a canine immunoglobulin heavy chain constant domain of isotype D having an amino acid sequence of SEQ ID NO:11, comprising:
 (i) subjecting a source mixture comprising target immunoglobulins or fusion proteins, or antigen-binding fragments thereof, to adhere affinity chromatography; and 
 (ii) subjecting the source mixture to anion exchange chromatography. 
 
     
     
         67 . An antibody, fusion protein, or binding fragment produced by the method of  claim 65 . 
     
     
         68 . An antibody, fusion protein, or binding fragment produced by the method of  claim 66 .