Novel ubiquitin ligases as therapeutic targets

Abstract

The present invention relates to the discovery, identification and characterization of nucleotides that encode novel substrate-targeting subunits of ubiquitin ligases. The invention encompasses nucleotides encoding novel substrate-targeting subunits of ubiquitin ligases: FBP1, FBP2, FBP3, FBP4, FBP5, FBP6, FBP7, FBP8, FBP9, FBP10, FBP11, FBP12, FBP13, FBP14, FBP15, FBP16, FBP17, FBP18, FBP19, FBP20, FBP21, FBP22, FBP23, FBP24, and FBP25, transgenic mice, knock-out mice, host cell expression systems and proteins encoded by the nucleotides of the present invention. The present invention relates to screening assays that use the novel substrate-targeting subunits to identify potential therapeutic agents such as small molecules, compounds or derivatives and analogues of the novel ubiquitin ligases which modulate activity of the novel ubiquitin ligases for the treatment of proliferative and differentiative disorders, such as cancer, major opportunistic infections, immune disorders, certain cardiovascular diseases, and inflammatory disorders. The invention further encompasses therapeutic protocols and pharmaceutical compositions designed to target ubiquitin ligases and their substrates for the treatment of proliferative disorders.

1 - 49 . (canceled)  
     
     
         50 . An isolated nucleic acid molecule comprising a nucleotide sequence which encodes a polypeptide comprising the amino acid sequence of SEQ ID NO: 10.  
     
     
         51 . An isolated nucleic acid molecule which encodes an F-box polypeptide, or a fragment thereof, said nucleic acid molecule comprising a nucleotide sequence of SEQ ID NO: 9.  
     
     
         52 . An isolated nucleic acid sequence derived from a mammalian genome that: 
 a) hybridizes under highly stringent conditions to the nucleotide sequence of SEQ ID NO: 9; and    b) encodes a gene product which contains an F-box motif and binds to Skp 1.    
     
     
         53 . A nucleotide vector containing the nucleotide sequence of  claim 50 ,  51  or  52 .  
     
     
         54 . An expression vector containing the nucleotide sequence of  claim 50 ,  51  or  52 , in operative association with a nucleotide regulatory sequence that controls expression of the nucleotide sequence in a host cell.  
     
     
         55 . A genetically engineered host cell that contains the nucleotide sequence of  claim 50 ,  51  or  52 , in operative association with a nucleotide regulatory sequence that controls expression of the nucleotide sequence in the host cell.  
     
     
         56 . A transgenic animal having cells which harbor a transgene comprising the nucleic acid of  claim 50 ,  51  or  52 .  
     
     
         57 . An animal inactivated in the locus comprising the nucleotide sequence of  claim 50 ,  51  or  52 .  
     
     
         58 . An isolated F-box polypeptide having the amino acid sequence of SEQ ID NO: 10.  
     
     
         59 . An antibody that immunospecifically binds the polypeptide of  claim 58 .  
     
     
         60 . A method of diagnosing proliferative and differentiative related disorders comprising measuring FBP5 gene expression in a patient sample.  
     
     
         61 . A method for screening compounds useful for the treatment of proliferative and differentiative disorders comprising contacting a compound with a cell expressing an F-box polypeptide having the amino acid sequence of SEQ ID NO: 10, or a fragment thereof, and its substrate, and detecting a change in the F-box polypeptide activity.  
     
     
         62 . The method of  claim 61  wherein the change in the F-box polypeptide activity is detected by detecting a change in the interaction of the F-box polypeptide with one or more polypeptides.  
     
     
         63 . The method of  claim 62  in which one of the one or more polypeptides is the substrate of the F-box polypeptide.  
     
     
         64 . The method of  claim 62  in which at least one of the one or more polypeptides is a component of the ubiquitin pathway.  
     
     
         65 . The method of  claim 62  in which one of the one or more polypeptides is Skp1.  
     
     
         66 . The method of  claim 61  wherein the change in the F-box polypeptide activity is detected by detecting a change in the ubiquitination or degradation of the substrate.  
     
     
         67 . A method for screening compounds useful for the treatment of proliferative and differentiative disorders comprising contacting a compound with a cell or a cell extract expressing Fbp5 and one or more Fbp5 substrates, and detecting a change in the activity of Fbp5.  
     
     
         68 . The method of  claim 67  wherein the change in the activity of Fbp5 is detected by detecting a change in the interaction of Fbp5 with one of the one or more Fbp5 substrates.  
     
     
         69 . The method of  claim 67  wherein the change in the activity of Fbp5 is detected by detecting a change in the ubiquitination or degradation of one of the one or more Fbp5 substrates.  
     
     
         70 . A method for treating a proliferative or differentiative disorder in a mammal comprising administering to the mammal a compound to the mammal that modulates the synthesis, expression or activity of the FBP5 gene or gene product so that symptoms of the disorder are ameliorated.  
     
     
         71 . The method of  claim 70  in which the disorder is breast cancer.  
     
     
         72 . The method of  claim 70  in which the disorder is ovarian cancer.  
     
     
         73 . The method of  claim 70  in which the disorder is prostate cancer.  
     
     
         74 . The method of  claim 70  in which the disorder is small cell lung carcinoma.