PrPSc -interacting molecules and uses thereof

Abstract

Peptides are disclosed which specifically bind to PrP Sc . Methods of identifying these peptides by sequence-independent and/or sequence-dependent screening assays are also disclosed. Various applications of the method and of the peptides identified are described, including their use for identifying drugs for treating prion diseases.

We claim:  
     
         1 . A method for identifying a peptide that binds to PrP Sc  or a fragment thereof, said method comprising the steps of: 
 (a) contacting a peptide of about 200 or fewer amino acids with a PrP Sc  polypeptide or fragment thereof under conditions that allow for complex formation between said peptide and PrP Sc  or fragment thereof; and    (b) detecting said complex, wherein the presence of the complex identifies said peptide as one which selectively binds to PrP Sc  or a fragment thereof.    
     
     
         2 . The method of  claim 1 , wherein said peptide is a fragment of PrP.  
     
     
         3 . The method of  claim 1 , wherein said peptide is about 100 to 150 amino acids in length.  
     
     
         4 . The method of  claim 1 , wherein said peptide is about 50 to 100 amino acids in length.  
     
     
         5 . The method of  claim 1 , wherein said peptide is about 25 to 50 amino acids in length.  
     
     
         6 . The method of  claim 1 , wherein said peptide is about 9 to 25 amino acids in length.  
     
     
         7 . The method of  claim 1 , wherein said peptide is a tripeptide.  
     
     
         8 . The method of  claim 1 , wherein said peptide is a tetrapeptide.  
     
     
         9 . The method of  claim 1 , wherein said peptide is a pentapeptide.  
     
     
         10 . The method of  claim 1 , wherein said peptide is a hexapeptide.  
     
     
         11 . The method of  claim 1 , wherein said peptide is a heptapeptide.  
     
     
         12 . The method of  claim 1 , wherein said peptide is an octapeptide.  
     
     
         13 . The method of  claim 1 , wherein said peptide is a nonapeptide.  
     
     
         14 . The method of  claim 1 , wherein said peptide is a decapeptide.  
     
     
         15 . The method of  claim 1 , wherein said peptide includes a YYX or YSA motif.  
     
     
         16 . The method of  claim 15 , wherein said YYX or YSA motif is repeated in said peptide.  
     
     
         17 . The method of  claim 16 , wherein said YYX or YSA motif is tandemly repeated in said peptide.  
     
     
         18 . The method of  claim 15 , wherein said YYX motif is selected from the group consisting of YYR, YYD, YYA, and YYQ.  
     
     
         19 . The method of  claim 15 , wherein said motif includes the amino acids YSA.  
     
     
         20 . The method of  claim 1 , wherein said peptide includes an YYXXYYXYY (SEQ ID NO: 1; where X is any amino acid) motif.  
     
     
         21 . The method of  claim 1 , wherein said peptide is coupled to a scaffolding agent.  
     
     
         22 . The method of  claim 21 , wherein said scaffolding agent is 4-map or 8-map.  
     
     
         23 . The method of  claim 1 , wherein said peptide is covalently coupled to a detectable-agent, solid support, or carrier.  
     
     
         24 . The method of  claim 1 , wherein said complex is detected using ELISA, RIA, western blotting, immunoprecipitation, fluorescence polarization or flow cytometry.  
     
     
         25 . A method for detecting a PrP Sc  in a biological sample, said method comprising the steps of: 
 (a) contacting said biological sample with a peptide of about 200 or fewer amino acids which selectively binds to PrP Sc  or a fragment thereof under conditions that allow for complex formation between said peptide and a PrP Sc  polypeptide or fragment thereof, and    (b) detecting said complex as an indication that said PrP Sc  is present in said biological sample.    
     
     
         26 . The method of  claim 25 , wherein said peptide does not substantially bind PrP C .  
     
     
         27 . The method of  claim 25 , wherein said peptide is 9 to 20 amino acids in length.  
     
     
         28 . The method of  claim 25 , wherein said peptide comprises a YYX or YSA motif.  
     
     
         29 . The method of  claim 28 , wherein said YYX motif is selected from the group consisting of YYR, YYD, YYA, and YYQ.  
     
     
         30 . The method of  claim 25 , wherein said peptide includes a YSA.  
     
     
         31 . The method of  claim 26 , wherein said peptide includes a YYXXYYXYY (where X is any amino acid) motif.  
     
     
         32 . The method of  claim 25 , wherein said biological sample comprises a tissue or cell, a tissue or cell extract, a bodily fluid, or a biopsy.  
     
     
         33 . The method of  claim 25 , wherein said PrP Sc  is from a human, a livestock species, or a pet species.  
     
     
         34 . The method of  claim 28 , wherein said peptide comprising said YYX motif is covalently coupled to a detectable-label.  
     
     
         35 . The method of  claim 28 , wherein said peptide comprising said YYX or YSA motif is covalently coupled to a solid support or carrier.  
     
     
         36 . The method of  claim 25 , wherein said complex is detected using an ELISA, RIA, western blotting, immunoprecipitation, fluorescence polarization, or flow cytometry.  
     
     
         37 . The method of  claim 25 , wherein said PrP Sc  in said biological sample is amplified PrP Sc .  
     
     
         38 . A method for diagnosing a prion disease in a mammal, said method comprising the steps of: 
 (a) contacting a biological sample of said mammal with a peptide of about 200 or fewer amino acids which selectively binds to PrP Sc  or a fragment thereof under conditions that allow for complex formation between said peptide and a PrP Sc  polypeptide or fragment thereof; and    (b) detecting said complex, which if present indicates a prion disease in said mammal.    
     
     
         39 . The method of  claim 38 , wherein said prion disease is selected from the group consisting of variant Creutzfeldt-Jakob Disease, bovine spongiform encephalopathy, scrapie, transmissible spongiform encephalopathy, and chronic wasting disease.  
     
     
         40 . The method of  claim 38 , wherein said peptide does not substantially bind  
     
     
         41 . The method of  claim 38 , wherein said peptide comprises a YYX or YSA motif.  
     
     
         42 . The method of  claim 41 , wherein said YYX motif is selected from the group consisting of YYR, YYD, YYA, and YYQ.  
     
     
         43 . The method of  claim 38 , wherein said peptide includes a YSA motif or a YAR motif.  
     
     
         44 . The method of  claim 38 , wherein said biological sample comprises a tissue or cell, a tissue or cell extract, a bodily fluid, or a biopsy.  
     
     
         45 . The method of  claim 38 , wherein said PrP Sc  is from a human, a livestock species, or a pet species.  
     
     
         46 . The method of  claim 41 , wherein said peptide comprising said YYX motif is covalently coupled to a detectable-agent.  
     
     
         47 . The method of  claim 41 , wherein said peptide comprising said YYX motif is covalently coupled to a solid substrate.  
     
     
         48 . The method of  claim 38 , wherein said complex is detected using an ELISA, RIA, western blotting, immunoprecipitation, fluorescence polarization or flow cytometry.  
     
     
         49 . A method for treating or preventing a prion disease in a mammal, comprising administering to said mammal an effective amount of a peptide of about 200 or fewer amino acids which selectively binds to PrP Sc  or a fragment thereof in a pharmaceutically-acceptable carrier and under conditions that allow for complex formation between said peptide and a PrP Sc  polypeptide or fragment thereof.  
     
     
         50 . The method of  claim 49 , wherein said prion disease is selected from the group consisting of variant Creutzfeldt-Jakob Disease, bovine spongiform encephalopathy, and scrapie.  
     
     
         51 . The method of  claim 49 , wherein said peptide does not substantially bind PrP C .  
     
     
         52 . The method of  claim 49 , wherein said peptide comprises a YYX or a YSA motif.  
     
     
         53 . The method of  claim 52 , wherein said YYX motif is selected from the group consisting of YYR, YYD, YYA, and YYQ.  
     
     
         54 . The method of  claim 52 , wherein said peptide includes a YSA motif.  
     
     
         55 . A method of inhibiting PrP Sc  in a biological sample, said method comprising: 
 treating the biological sample with a peptide of about 200 or fewer amino acids which selectively binds to PrP Sc  or a fragment thereof under conditions that allow for complex formation between said peptide and a PrP Sc  polypeptide or fragment thereof and for a period of time sufficient to permit the formation of a complex comprising said peptide and a PrP Sc .    
     
     
         56 . The method of  claim 55 , wherein said biological sample is a bodily fluid, a tissue or organ.  
     
     
         57 . The method of  claim 55 , wherein said biological sample is perfused with said peptide.  
     
     
         58 . The method of  claim 56 , wherein said peptide does not substantially bind PrP C .  
     
     
         59 . A method for decontaminating PrP Sc  from a biological sample, said method comprising the steps of: 
 (a) treating the biological sample with a peptide of about 200 or fewer amino acids which selectively binds to PrP Sc  or a fragment thereof under conditions that allow for complex formation between said peptide and a PrP Sc  polypeptide or fragment thereof and for a period of time sufficient to permit the formation of complex comprising said peptide and PrP Sc ; and    (b ) recovering said complex from said biological sample.    
     
     
         60 . The method of  claim 59 , wherein said biological sample is a tissue, bodily fluid, or organ.  
     
     
         61 . The method of  claim 59 , wherein said biological sample is perfused with said peptide.  
     
     
         62 . The method of  claim 59 , wherein said peptide does not substantially bind PrP C .  
     
     
         63 . The method of  claim 59 , wherein said peptide comprises a YYX or YSA motif.  
     
     
         64 . The method of  claim 63 , wherein said YYX motif is selected from the group consisting of YYR, YYD, YYA, and YYQ.  
     
     
         65 . The method of  claim 59 , wherein said peptide includes a YSA motif.  
     
     
         66 . A method for identifying an agent for the treatment of a prion disease, said method comprising the steps of: 
 (a) combining a peptide of about 200 or fewer amino acids which selectively binds to PrP Sc  or a fragment thereof, PrP Sc , and an agent under conditions allowing for complex formation of said peptide and said PrP Sc ; and    (b) determining whether complex formation is increased or decreased in comparison to complex formation in the absence of said agent, thereby identifying an agent for treating said prion diseases.    
     
     
         67 . The method of  claim 66 , wherein said prion disease is selected from the group consisting of variant Creutzfeldt-Jakob Disease, bovine spongiform encephalopathy, and scrapie.  
     
     
         68 . The method of  claim 66 , wherein said peptide does not substantially bind  
     
     
         69 . The method of  claim 66 , wherein said peptide comprises a YYX or YSA motif.  
     
     
         70 . The method of  claim 69 , wherein said YYX motif is selected from the group consisting of YYR, YYD, YYA, and YYQ.  
     
     
         71 . The method of  claim 66 , wherein said peptide includes a YSA motif.  
     
     
         72 . The method of  claim 66 , wherein said prion disease affects a human, a livestock species, or a pet species.  
     
     
         73 . The method of  claim 66 , wherein said prion disease affects a human, bovine, sheep, or goat.  
     
     
         74 . The method of  claim 66 , wherein said agent increases complex formation.  
     
     
         75 . The method of  claim 66 , wherein said agent decreases complex formation.  
     
     
         76 . An apparatus for detecting PrP Sc  in a biological sample, the apparatus comprising: a peptide-fixed portion where a peptide for trapping an amount of an analyte in a sample is present and in a predetermined amount.  
     
     
         77 . The apparatus of  claim 76 , wherein said peptide comprises a YYX, YYR, YYD, or YYQ amino acid sequence, said peptide having antigenicity as a PrP Sc .  
     
     
         78 . The apparatus of  claim 76 , wherein said peptide is composed of 18 or fewer amino acids.  
     
     
         79 . The apparatus of  claim 76 , wherein said peptide is composed of 12 or fewer amino acids.  
     
     
         80 . The apparatus of  claim 76 , wherein said peptide is the nonapeptide having the amino acid sequence YYRRYYRYY (SEQ ID NO: 2).  
     
     
         81 . The apparatus of  claim 77 , wherein said peptide is composed of 8 or fewer amino acids.  
     
     
         82 . The apparatus of  claim 77 , wherein said peptide is composed of 5 or fewer amino acids.  
     
     
         83 . The apparatus of  claim 77 , wherein said peptide is the tripeptide having the amino acid sequence YYR.  
     
     
         84 . The apparatus of  claim 77 , wherein said analyte is a bodily fluid.  
     
     
         85 . A method for detecting an anti-PrP Sc  antibody in a biological sample, said method comprising the steps of: 
 (a) contacting said biological sample with a peptide of about 200 or fewer amino acids which selectively binds to PrP Sc  or a fragment thereof under conditions that allow for complex formation between said peptide and anti-PrP Sc  antibody; and    (b) detecting said complexes as an indication that anti-PrP Sc  antibody is present in said biological sample.