Methods of use of the TACI/TACI-L interaction

Abstract

The invention discloses a novel interaction between a TNF receptor (TACI) and its interacting ligand (TACI-L). Also disclosed are methods of screening candidate molecules to determine potential antagonists and agonists of the TACI/TACI-L interaction. The use of the antagonists and agonists as therapeutics to treat autoimmune diseases, inflammation, and to inhibit graft vs. host rejections is further disclosed.

1 . A method of screening a test compound comprising the steps of:
 a. forming a composition comprising
 (i) a first isolated protein, comprising a polypeptide selected from the group consisting of:
 (a) the polypeptide of SEQ ID NO:2; 
 (b) a polypeptide comprising amino acids 2-166 of SEQ ID NO:2; 
 (c) a fragment of the polypeptide of SEQ ID NO:2; or 
 (d) a polypeptide encoded by a nucleic acid sequence that is at least 95% identical to SEQ ID NO: 1; 
 wherein said fragment of (i)(c) and said polypeptides of (i)(d) bind SEQ ID NO:4; 
 
 (ii) a second isolated protein, comprising a polypeptide selected from the group consisting of:
 (a) the polypeptide of SEQ ID NO:4; 
 (b) a polypeptide comprising amino acids 123-285 of SEQ ID NO:4; or 
 (c) a polypeptide comprising amino acids 73-285 of SEQ ID NO:4; 
 
 (iii) a test compound; and 
   b. assaying for the level of interaction of the protein of (i) and the protein of (ii), wherein the affinity constant for protein (i) and protein (ii) is from 1.53×10 −9  to 2.2×10 −9 ;   
       such that if the level obtained in step (b) differs from that obtained in the absence of the test compound, a test compound that affects the interaction of the protein of (i) and the protein of (ii) is identified. 
     
     
         2 . The method of  claim 1  wherein at least one of the proteins of (i) and the proteins of (ii) is labeled with a detectable moiety. 
     
     
         3 . The method of  claim 1  wherein both the proteins of (i) and (ii) are soluble. 
     
     
         4 . The method of  claim 3  wherein both the soluble protein of (i) and the soluble protein of (ii) are labeled with a detectable moiety. 
     
     
         5 . The method of  claim 1  wherein the test compound is an antibody. 
     
     
         6 . The method of  claim 5  wherein the antibody is a humanized antibody. 
     
     
         7 . The method of  claim 1  wherein the composition is formed by adding the test compound to the protein of (i) and the protein of (ii). 
     
     
         8 . The method of  claim 1  wherein step (b) comprises determining a dissociation constant of the interaction of the protein of (i) with the protein of (ii). 
     
     
         9 . The method of  claim 1  wherein step (b) comprises assessing activation of the protein of (i) in a cell. 
     
     
         10 . The method of  claim 9  wherein assessing activation of the protein of (i) in a cell is measured by calcium influx. 
     
     
         11 . The method of  claim 1  wherein the protein of (ii) is a polypeptide comprising amino acids 123-285 of SEQ ID NO:4 or a polypeptide comprising amino acids 73-285 of SEQ ID NO:4. 
     
     
         12 . The method of  claim 11  wherein the polypeptide comprising amino acids 123-285 of SEQ ID NO:4 or the polypeptide comprising amino acids 73-285 of SEQ ID NO:4 further comprises a leucine zipper domain. 
     
     
         13 . The method of  claim 1  wherein the protein of (i) is a polypeptide comprising amino acids 2-166 of SEQ ID NO:2. 
     
     
         14 . The method of  claim 13  wherein the polypeptide comprising amino acids 2-166 of SEQ ID NO:2 further comprises a Fc domain. 
     
     
         15 . A method of screening a test compound comprising the steps of:
 a. forming a composition comprising
 (i) an isolated protein selected from the group consisting of:
 (a) the polypeptide of SEQ ID NO:2; 
 (b) a polypeptide comprising amino acids 2-166 of SEQ ID NO:2; and 
 (c) a fragment of the polypeptide of SEQ ID NO:2; wherein said fragment binds SEQ ID NO:4; 
 
 (ii) the polypeptide of SEQ ID NO:4; and 
 (iii) a test compound; and 
   b. assaying for the level of interaction of the protein if (i) and the protein of (ii), wherein the affinity constant for protein (i) and protein (ii) is from 1.53×10 −9  to 2.2×10 −9 ;   
       such that if the level obtained in step (b) differs from that obtained in the absence of the test compound, a test compound that affects the interaction of the protein of (i) and the protein of (ii) is identified. 
     
     
         16 . A method of screening a test compound comprising the steps of:
 a. forming a composition comprising
 (i) the polypeptide of SEQ ID NO:2; 
 (ii) an isolated protein selected from the group consisting of:
 (a) the polypeptide of SEQ ID NO:4; 
 (b) a polypeptide comprising amino acids 123-285 of SEQ ID NO:4; 
 (c) a polypeptide comprising amino acids 73-285 of SEQ ID NO:4; and 
 (d) a fragment of the polypeptide of SEQ ID NO:4; wherein said fragment binds SEQ ID NO:2; and 
 
 (iii) a test compound; and 
   b. assaying for the level of interaction of the protein of (i) and the protein of (ii), wherein the affinity constant for protein (i) and protein (ii) is from 1.53×10 −9  to 2.2×10 −9 ;   
       such that if the level obtained in step (b) differs from that obtained in the absence of the test compound, a test compound that affects the interaction of the protein of (i) and the protein of (ii) is identified. 
     
     
         17 . A method of screening a test compound comprising the steps of:
 a. forming a composition comprising
 (i) the polypeptide of SEQ ID NO:2; 
 (ii) the polypeptide of SEQ ID NO:4; and 
 (iii) a test compound; and 
   b. assaying for the level of interaction of the polypeptide of (i) and the polypeptide of (ii), wherein the affinity constant for protein (i) and protein (ii) is from 1.53×10 −9  to 2.2×10 −9 ;   
       such that if the level obtained in step (b) differs from that obtained in the absence of the test compound, a test compound that affects the interaction of the protein of (i) and the protein of (ii) is identified. 
     
     
         18 . The method of  claim 11 , wherein the polypeptide comprising amino acids 123-285 of SEQ ID NO:4 or the polypeptide comprising amino acids 73-285 of SEQ ID NO:4 further comprises a Fc domain. 
     
     
         19 . The method of  claim 5 , wherein the antibody is human. 
     
     
         20 . The method of  claim 5 , wherein the antibody comprises a Fab fragment. 
     
     
         21 . The method of  claim 5 , wherein the antibody comprises a F(ab′) 2  fragment. 
     
     
         22 . A method of screening a test compound comprising the steps of:
 a. forming a composition comprising
 (i) a first isolated protein, comprising a polypeptide selected from the group consisting of:
 (a) the polypeptide of SEQ ID NO:2; or 
 (b) a polypeptide comprising amino acids 2-166 of SEQ ID NO:2; 
 
 (ii) a second isolated protein, comprising a polypeptide selected from the group consisting of:
 (a) the polypeptide of SEQ ID NO:4; 
 (b) a polypeptide comprising amino acids 123-285 of SEQ ID NO:4; 
 (c) a polypeptide comprising amino acids 73-285 of SEQ ID NO:4; 
 (d) a fragment of the polypeptide of SEQ ID NO:4; or 
 (e) a polypeptide encoded by a nucleic acid sequence that is at least 95% identical to SEQ ID NO:3; 
 
 wherein said fragment of (ii)(d) and said polypeptides of (ii)(e) bind SEQ ID NO:2; and 
 (iii) a test compound; and 
   b. assaying for the level of interaction of the protein of (i) and the protein of (ii), wherein the affinity constant for protein (i) and protein (ii) is from 1.53×10 −9  to 2.2×10 −9 ;   
       such that if the level obtained in step (b) differs from that obtained in the absence of the test compound, a test compound that affects the interaction of the protein of (i) and the protein of (ii) is identified.