Protein kinase c zeta as a drug target for arthritis and other inflammatory diseases

Abstract

The present invention is based on the discovery that ζPKC expression is increased in the tissues of arthritis patients as compared to normal individuals. Accordingly, the present invention provides methods of diagnosing, prognosing, and monitoring the course of arthritis in a patient based on increased ζPKC gene expression in arthritic tissue. The present invention further provides compounds that inhibit the expression of ζPKC for use as remedies in the treatment of arthritis, including, but not limited to, inhibitory polynucleotides and polypeptides, small molecules, and peptide inhibitors. In addition, the present invention provides pharmaceutical formulations and routes of administration for such remedies, as well as methods for assessing their efficacy.

1 . A method for use in the diagnosis of arthritis in a subject comprising the steps of:
 detecting a test amount of a ζPKC gene product in a sample from the subject; and   comparing the test amount with a normal amount of the ζPKC gene product in a control sample,   whereby a finding that the test amount is greater than the normal amount provides a positive indication in the diagnosis of arthritis.   
     
     
         2 . The method of  claim 1 , wherein the sample comprises chondrocytes. 
     
     
         3 . The method of  claim 1 , wherein the ζPKC gene product comprises RNA or cDNA. 
     
     
         4 . The method of  claim 1 , wherein the ζPKC gene product is ζPKC polypeptide. 
     
     
         5 . A method for use in the prognosis of arthritis in a subject comprising the steps of:
 detecting a test amount of a ζPKC gene product in a sample from the subject; and   comparing the test amount with prognostic amounts of the ζPKC gene product in control samples,   whereby a comparison of the test amount with the prognostic amounts provides an indication of the prognosis of arthritis.   
     
     
         6 . The method of  claim 5 , wherein the sample comprises chondrocytes. 
     
     
         7 . The method of  claim 5 , wherein the ζPKC gene product comprises RNA or cDNA. 
     
     
         8 . The method of  claim 5 , wherein the ζPKC gene product is ζPKC polypeptide. 
     
     
         9 . A method for use in monitoring the course of arthritis in a subject comprising the steps of:
 detecting a first test amount of a ζPKC gene product in a sample from the subject at a first time;   detecting a second test amount of the ζPKC gene product in a sample from the subject at a second, later time; and   comparing the first test amount and the second test amount,   whereby an increase in the amount of the ζPKC gene product in the second test amount as compared with the first test amount indicates progression of arthritis, and   whereby a decrease in the amount of the PKC gene product in the second test amount as compared with the first test amount indicates remission of arthritis.   
     
     
         10 . The method of  claim 9 , wherein the sample comprises chondrocytes. 
     
     
         11 . The method of  claim 9 , wherein the PKC gene product comprises RNA or cDNA. 
     
     
         12 . The method of  claim 9 , wherein the ζPKC gene product is ζPKC polypeptide. 
     
     
         13 . A method for assessing the efficacy of a treatment for arthritis in a subject comprising the steps of:
 detecting a first test amount of a ζPKC gene product in a sample from the subject prior to treatment;   detecting a second test amount of the ζPKC gene product in a sample from the subject after treatment; and   comparing the first test amount and the second test amount,   whereby a decrease in the amount of the ζPKC gene product in the second test amount as compared with the first test amount indicates that the treatment for arthritis is efficacious.   
     
     
         14 . The method of  claim 13 , wherein the sample comprises chondrocytes. 
     
     
         15 . The method of  claim 13 , wherein the ζPKC gene product comprises RNA or cDNA. 
     
     
         16 . The method of  claim 13 , wherein the ζPKC gene product is ζPKC polypeptide. 
     
     
         17 . A method of screening for a compound capable of inhibiting arthritis in a subject comprising the steps of:
 providing a first sample and a second sample containing equivalent amounts of ζPKC;   contacting the first sample with the compound; and   determining whether the activity of ζPKC in the first sample is decreased relative to the activity of ζPKC in the second sample not contacted with the compound,   whereby a decrease in the activity of ζPKC in the first sample as compared with the second sample indicates that the compound inhibits arthritis in the subject.   
     
     
         18 . The method of  claim 17 , wherein the compound inhibits the activity of PKC in chondrocytes. 
     
     
         19 . The method of  claim 17 , wherein the compound is a small molecule. 
     
     
         20 . The method of  claim 17 , wherein the activity of ζPKC is determined by use of an enzymatic protein kinase assay. 
     
     
         21 . The method of  claim 17 , wherein the activity of ζPKC is determined by use of a chondrocyte pellet assay. 
     
     
         22 . The method of  claim 17 , wherein the activity of ζPKC is determined by use of an assay measuring proteoglycan degradation. 
     
     
         23 . The method of  claim 17 , wherein the activity of ζPKC is determined by use of an assay measuring NF-κB activity. 
     
     
         24 . A method of screening for a compound capable of inhibiting arthritis in a subject comprising the steps of:
 providing a first sample and a second sample containing equivalent amounts of cells that express ζPKC;   contacting the first sample with the compound; and   determining whether the expression of ζPKC gene product in the first sample is decreased relative to the expression of ζPKC gene product in the second sample not contacted with the compound,   whereby a decrease in the expression of ζPKC gene product in the first sample as compared with the second sample indicates that the compound inhibits arthritis in the subject.   
     
     
         25 . The method of  claim 24 , wherein the compound inhibits the expression of PKC gene product in chondrocytes. 
     
     
         26 . The method of  claim 24 , wherein the compound is a small molecule. 
     
     
         27 . The method of  claim 24 , wherein the expression of ζPKC gene product is determined by use of an enzymatic protein kinase assay. 
     
     
         28 . The method of  claim 24 , wherein the expression of ζPKC gene product is determined by use of a chondrocyte pellet assay. 
     
     
         29 . The method of  claim 24 , wherein the expression of ζPKC gene product is determined by use of an assay measuring proteoglycan degradation. 
     
     
         30 . The method of  claim 24 , wherein the expression of ζPKC gene product is determined by use of an assay measuring NF-κB activity. 
     
     
         31 . A method for the treatment of arthritis in a subject comprising administering to the subject a compound that inhibits the activity of ζPKC in the subject. 
     
     
         32 . The method of  claim 31 , wherein the compound inhibits the activity of PKC in chondrocytes. 
     
     
         33 . The method of  claim 31 , wherein the compound is an antisense polynucleotide. 
     
     
         34 . The method of  claim 31 , wherein the compound is a small molecule. 
     
     
         35 . The method of  claim 31 , wherein the compound is a siRNA molecule. 
     
     
         36 . The method of  claim 35 , wherein the siRNA molecule is selected from the group consisting of siRNA molecules shown in  FIG. 1 . 
     
     
         37 . A method for the treatment of arthritis in a subject comprising administering to the subject a compound that inhibits the expression of ζPKC in the subject. 
     
     
         38 . The method of  claim 37 , wherein the compound inhibits the expression of ζPKC in chondrocytes. 
     
     
         39 . The method of  claim 37 , wherein the compound is an antisense polynucleotide. 
     
     
         40 . The method of  claim 37 , wherein the compound is a small molecule. 
     
     
         41 . The method of  claim 37 , wherein the compound is a siRNA molecule. 
     
     
         42 . The method of  claim 41 , wherein the siRNA molecule is selected from the group consisting of siRNA molecules shown in  FIG. 1 . 
     
     
         43 . A siRNA molecule that inhibits the expression or activity of ζPKC. 
     
     
         44 . The siRNA molecule of  claim 43 , wherein the siRNA molecule is selected from the group consisting of siRNA molecules shown in  FIG. 1 .