Novel metalloprotease having aggrecanase activity

Abstract

This invention provides a novel metalloprotease having an aggrecanase activity which causes joint diseases, a gene coding for this metalloprotease, a promoter of the above metalloprotease, a method for screening a drug with the use of the above metalloprotease and a pharmaceutical composition for inhibiting degradation of proteoglycans, which comprises as the active ingredient a substance capable of inhibiting the aggrecanase activity of the above metalloprotease.

1 . A metalloprotease having an aggrecanase activity, which comprises an amino acid sequence of from the 213th position to the 583rd position of an amino acid sequence represented by SEQ ID NO:1 or which consists of an amino acid sequence of from the 213th position to the 583rd position of the amino acid sequence represented by SEQ ID NO:1 wherein from 1 to 10 amino acid residues are substituted, deleted and/or inserted.  
     
     
         2 . A metalloprotease having an aggrecanase activity, which comprises an amino acid sequence of from the 1st position to the 583rd position of an amino acid sequence represented by SEQ ID NO:1 or which consists of an amino acid sequence of from the 1st position to the 583rd position of the amino acid sequence represented by SEQ ID NO:1 wherein from 1 to 10 amino acid residues are substituted, deleted and/or inserted.  
     
     
         3 . A metalloprotease having an aggrecanase activity, which consists of an amino acid sequence represented by SEQ ID NO:1, an amino acid sequence of from the 1st position to the 687th position of an amino acid sequence represented by SEQ ID NO:1, an amino acid sequence of from the 1st position to the 583rd position of the amino acid sequence represented by SEQ ID NO:1, an amino acid sequence of from the 213th position to the 950th position of the amino acid sequence represented by SEQ ID NO:1, an amino acid sequence of from the 213th position to the 687th position of the amino acid sequence represented by SEQ ID NO:1 or an amino acid sequence of from the 213th position to the 583rd position of the amino acid sequence represented by SEQ ID NO:1, or any one of these sequences wherein from 1 to 10 amino acid residues are substituted, deleted and/or inserted.  
     
     
         4 . A gene which encodes an amino acid sequence of the metalloprotease having an aggrecanase activity described in any one of  claims 1  to  3 .  
     
     
         5 . A vector which comprises the gene described in  claim 4 .  
     
     
         6 . A host cell which comprises the vector described in  claim 5 .  
     
     
         7 . A method for producing the metalloprotease having an aggrecanase activity described in any one of  claims 1  to  3 , which comprises using the host cell described in  claim 6 .  
     
     
         8 . An antibody against the metalloprotease having an aggrecanase activity described in any one of  claims 1  to  3 .  
     
     
         9 . A method for screening a substance capable of inhibiting an aggrecanase activity of a metalloprotease, which comprises allowing the metalloprotease having an aggrecanase activity described in any one of  claims 1  to  3  to contact with a compound to be tested.  
     
     
         10 . A pharmaceutical composition for inhibiting degradation of proteoglycans, which comprises a substance capable of inhibiting the metalloprotease having an aggrecanase activity described in any one of  claims 1  to  3 , as an active ingredient.  
     
     
         11 . A gene represented by SEQ ID NO:24, 25, 26, 27, 28, 29, 30 or 31, or a gene represented by SEQ ID NO:24, 25, 26, 27, 28, 29, 30 or 31 wherein from 1 to 10 amino acid residues are substituted, deleted and/or inserted, which has a joint disease aggrecanase promoter activity.  
     
     
         12 . An antibody against a metalloprotease having aggrecanase activity, wherein said metalloprotease is selected from the group consisting of: 
 (a) a purified metalloprotease comprising an amino acid sequence represented by amino acids 213-583 of SEQ ID NO:1 or an amino acid sequence represented by amino acids 213-583 of SEQ ID NO:1 wherein from 1 to 10 amino acid residues are substituted, deleted and/or inserted, and wherein said metalloprotease has aggrecanase activity;    (b) a purified metalloprotease comprising an amino acid sequence represented by amino acids 1-583 of SEQ ID NO:1 or an amino acid sequence represented by amino acids 1-583 of SEQ ID NO:1 wherein from 1 to 10 amino acid residues are substituted, deleted and/or inserted, and wherein said metalloprotease has aggrecanase activity;    (c) a purified metalloprotease comprising an amino acid sequence selected from the group consisting of an amino acid sequence represented by amino acids 1-950 of SEQ ID NO:1, an amino acid sequence represented by amino acids 1-687 of SEQ ID NO:1, an amino acid sequence represented by amino acids 1-583 of SEQ ID NO:1, an amino acid sequence represented by amino acids 213-950 of SEQ ID NO:1, an amino acid sequence represented by amino acids 213-687 of SEQ ID NO:1, an amino acid sequence represented by amino acids 213-583 of SEQ ID NO:1, and any one of said sequences wherein from 1 to 10 amino acid residues are substituted, deleted and/or inserted, and wherein said metalloprotease has aggrecanase activity; and    (d) a purified metalloprotease comprising an amino acid sequence that has 90% or more sequence homology with the amino acid sequence set forth in SEQ ID NO:1, wherein said metalloprotease has aggrecanase activity.    
     
     
         13 . A method for screening a substance capable of inhibiting an aggrecanse activity of a metalloprotease, which comprises allowing the metalloprotease to contact a compound to be tested, wherein the metalloprotease is selected from the group consisting of: 
 (a) a purified metalloprotease comprising an amino acid sequence represented by amino acids 213-583 of SEQ ID NO:1 or an amino acid sequence represented by amino acids 213-583 of SEQ ID NO:1 wherein from 1 to 10 amino acid residues are substituted, deleted and/or inserted, and wherein said metalloprotease has aggrecanase activity;    (b) a purified metalloprotease comprising an amino acid sequence represented by amino acids 1-583 of SEQ ID NO:1 or an amino acid sequence represented by amino acids 1-583 of SEQ ID NO:1 wherein from 1 to 10 amino acid residues are substituted, deleted and/or inserted, and wherein said metalloprotease has aggrecanase activity;    (c) a purified metalloprotease comprising an amino acid sequence selected from the group consisting of an amino acid sequence represented by amino acids 1-950 of SEQ ID NO:1, an amino acid sequence represented by amino acids 1-687 of SEQ ID NO:1, an amino acid sequence represented by amino acids 1-583 of SEQ ID NO:1, an amino acid sequence represented by amino acids 213-950 of SEQ ID NO:1, an amino acid sequence represented by amino acids 213-687 of SEQ ID NO:1, an amino acid sequence represented by amino acids 213-583 of SEQ ID NO:1, and any one of said sequences wherein from 1 to 10 amino acid residues are substituted, deleted and/or inserted, and wherein said metalloprotease has aggrecanase activity; and    (d) a purified metalloprotease comprising an amino acid sequence that has 90% or more sequence homology with the amino acid sequence set forth in SEQ ID NO:1, wherein said metalloprotease has aggrecanase activity.    
     
     
         14 . A pharmaceutical composition for inhibiting degradation of proteoglycans, which comprises a substance capable of inhibiting a metalloprotease selected from the group consisting of: 
 (a) a purified metalloprotease comprising an amino acid sequence represented by amino acids 213-583 of SEQ ID NO:1 or an amino acid sequence represented by amino acids 213-583 of SEQ ID NO:1 wherein from 1 to 10 amino acid residues are substituted, deleted and/or inserted, and wherein said metalloprotease has aggrecanase activity;    (b) a purified metalloprotease comprising an amino acid sequence represented by amino acids 1-583 of SEQ ID NO:1 or an amino acid sequence represented by amino acids 1-583 of SEQ ID NO:1 wherein from 1 to 10 amino acid residues are substituted, deleted and/or inserted, and wherein said metalloprotease has aggrecanase activity;    (c) a purified metalloprotease comprising an amino acid sequence selected from the group consisting of an amino acid sequence represented by amino acids 1-950 of SEQ ID NO:1, an amino acid sequence represented by amino acids 1-687 of SEQ ID NO:1, an amino acid sequence represented by amino acids 1-583 of SEQ ID NO:1, an amino acid sequence represented by amino acids 213-950 of SEQ ID NO:1, an amino acid sequence represented by amino acids 213-687 of SEQ ID NO:1, an amino acid sequence represented by amino acids 213-583 of SEQ ID NO:1, and any one of said sequences wherein from 1 to 10 amino acid residues are substituted, deleted and/or inserted, and wherein said metalloprotease has aggrecanase activity; and    (d) a purified metalloprotease comprising an amino acid sequence that has 90% or more sequence homology with the amino acid sequence set forth in SEQ ID NO:1, wherein said metalloprotease has aggrecanase activity.    
     
     
         15 . A method of treating a joint disease, comprising administering to a patient in need of treatment a compound obtainable by the method of  claim 13 , thereby treating a joint disease.  
     
     
         16 . A polynucleotide represented by SEQ ID NO:24, 25, 26, 27, 28, 29, 30 or 31, or a polynucleotide represented by SEQ ID NO: 24, 25, 26, 27, 28, 29, 30 or 31 and wherein from 1 to 10 nucleotide residues are substituted, deleted and/or inserted, wherein said polynucleotide has a joint disease aggrecanase promoter activity.  
     
     
         17 . A method for screening a substance capable of inhibiting aggrecanase promoter activity, which comprises allowing a cell transformed with the polynucleotide described in  claim 16  to contact a compound to be tested.