Cyclin dependent kinase binding compounds

Abstract

The present invention identifies substances having the property of binding to cyclin dependent kinase (cdk) comprising: (i) a peptide including amino acid residues 84 to 103 of full length p16 protein, or an active portion or derivative thereof; or (ii) a functional mimetic of the fragment. active portion or derivative; the substance excludes full length p16. p15. p18 and p19 proteins. These substances are useful in tumour suppression by inhibiting the phosphorylation of Rb protein. Also described herein is the resolution of the amino acid motifs responsible for binding cdks, an FLD motif. corresponding to amino acid residues 90 to 92 of full length p16 protein. and an LVVL motif, corresponding to amino acid residues 94 to 97 of full length p16 protein. The substances disclosed herein can be used in the treatment of hyperproliferative disorders and to screen and design molecules having the similar properties.

1 . A substance having the property of binding to cyclin dependent kinase (cdk) comprising: 
 (i) a peptide including amino acid residues 84 to 103 of full length p16 protein, or an active portion or derivative thereof; or,    (ii) a functional mimetic of the fragment, active portion or derivative;    wherein the substance excludes full length p16, p15, p18 and p19 proteins.    
     
     
         2 . The substance of  claim 1  wherein the cyclin dependent kinase is cdk4 or cdk6.  
     
     
         3 . The substance of  claim 1  or  claim 2  wherein binding of the substance to the cyclin dependent kinase causes inhibition of pRb phosphorylation.  
     
     
         4 . The substance of any one of  claims 1  to  3  wherein the peptide includes amino acid residues 89 to 97 of full length p16 protein.  
     
     
         5 . The substance of any one of the preceding claims wherein the peptide fragment includes the amino acid motifs FLD, corresponding to amino acid residues 90 to 92 of full length p16 protein, and/or LVVL, corresponding to amino acid residues 94 to 97 of full length p16 protein.  
     
     
         6 . The substance of  claim 5  wherein the peptide fragment includes the amino acid motif FLDxLVVL, wherein x one or more amino acids.  
     
     
         7 . The substance of any one of the preceding claims wherein the aspartic acid residue in the peptide fragment at the position corresponding to position 92 of full length p16 protein is substituted for a hydrophobic amino acid residue.  
     
     
         8 . The substance of  claim 8  wherein the hydrophobic amino acid residue is alanine.  
     
     
         9 . The substance of any one of the preceding claims derived from p16, p15, p18 or p19 protein.  
     
     
         10 . The substance of any one of the preceding claims coupled to a carrier molecule so that the substance can be delivered to cells.  
     
     
         11 . The substance of  claim 10  wherein the carrier molecule comprises a peptide derived from an Ancennapedia homeodomain.  
     
     
         12 . The substance of any one of the preceding claims wherein the peptide fragment is coupled to a stabilising molecule.  
     
     
         13 . A pharmaceutical composition comprising one or more of the substances of any one of the preceding claims, in combination with a physiologically acceptable carrier.  
     
     
         14 . A substance of any one of  claims 1  to  12  for use in a method of medical treatment.  
     
     
         15 . The use of a substance of any one of  claims 1  to  12  in the preparation of a medicament for the treatment of a hyperproliferative disorder.  
     
     
         16 . The use of  claim 15  wherein the hyperproliferative disorder is cancer, psoriasis or arteriogenesis.  
     
     
         17 . Nucleic acid encoding the substances of any one of  claims 1  to  12 .  
     
     
         18 . A vector incorporating the nucleic acid of  claim 17  operably linked to expression control sequences.  
     
     
         19 . The use of a substance of any one of  claims 1  to  12  in a method of in screening for (i) compounds having the one or more of the biological activities of the substances described above or (ii) compounds which are binding partners of one of the substances.  
     
     
         20 . A method of identifying compounds which compete with a substance of any one of  claims 1  to  12 , the method comprising: 
 (a) binding a predetermined quantity of the substance which is detectably labelled to a cyclin dependent kinase (cdk);  
 (b) adding a candidate compound; and,  
 (c) determining the amount of the labelled compound that remains bound to thecdk or which becomes displaced by the candidate compound.  
 
     
     
         21 . A method of identifying mimetics of a substance of any one of  claims 1  to  12 , the method comprising: 
 (a) immobilising one or more candidate compounds on a solid substrate;  
 (b) exposing the substrate to a labelled cyclin dependent kinase (cdk);  
 (c) selecting the candidate compounds that bind to cdk.  
 
     
     
         22 . The method of  claim 20  or  claim 21  wherein the cdks are produced in reticulocyte lysates.  
     
     
         23 . The method of any one of  claims 20  to  22  wherein the cell dependent kinase is cdk4 or cdk6.  
     
     
         24 . The method of any one of  claims 20  to  23 , further comprising testing the candidate compound for the property of inhibiting pRb phosphorylation and/or testing the compound for the property of inhibiting the entry of cells into the S-phase.  
     
     
         25 . The method of any one of  claims 20  to  24  wherein the candidate compounds are selected from a synthetic combinatorial library.  
     
     
         26 . The use of a fragment of p16 protein including the amino acid motifs FLD, corresponding to amino acid residues 90 to 92 of full length p16 protein, and/or LVVL, corresponding to amino acid residues 94 to 97 of full length p16 protein in the design of an organic compound which is modelled to resemble the three dimensional structure of said amino acid motifs, the organic compound having the properties of binding to cyclin dependent kinase and/or inhibiting pRb phosphorylation.