Methods for identifying compounds for treatment of cell proliferative disorders associated with adaptor protein interactions

Abstract

The present invention relates to compositions and methods for the prevention and treatment of cell proliferative disorders wherein a protein tyrosine kinase or protein tyrosine phosphatase capable of complexing with a member of the SH2- and/or SH3-containing family of adaptor proteins is involved. This invention is based, in part, on the surprising discovery that the adaptor protein, GRB-2, binds the intracellular BCR-ABL tyrosine kinase product in vivo and is necessary for the activation of the oncogenic potential of the BCR/ABL product. The present invention further relates to protein tyrosine kinase/adaptor protein complexes and the uses of these complexes for the identification of agents capable of decreasing or inhibiting the interaction between the members of such complexes.

What is claimed is:  
     
         1 . A method for identifying a compound to be tested for an ability to modulate a cell proliferative disorder involving a protein tyrosine kinase polypeptide/adaptor polypeptide complex comprising: 
 (a) exposing at least one compound to a peptide comprising a functional portion of a member of the protein tyrosine kinase polypeptide/adaptor polypeptide complex for a time sufficient to allow binding of the compound to the functional portion of the member;    (b) removing non-bound compounds; and    (c) detecting the presence of the compound bound to the functional portion of the member of the protein tyrosine kinase polypeptide/adaptor polypeptide complex, thereby identifying a compound to be tested for an ability to modulate an cell proliferative disorder involving a protein tyrosine kinase polypeptide/adaptor polypeptide complex.    
     
     
         2 . A method for identifying a compound capable of modulating a cell proliferative disorder involving a protein tyrosine kinase polypeptide/adaptor polypeptide complex comprising, exposing the compound to a protein tyrosine kinase polypeptide/adaptor polypeptide complex for a time sufficient to allow disruption of the complex, and detecting the disruption of the protein tyrosine kinase polypeptide/adaptor polypeptide complex.  
     
     
         3 . The method of  claim 1  or  2  wherein the protein tyrosine kinase polypeptide of the protein tyrosine kinase polypeptide/adaptor polypeptide complex is a transmembrane, receptor protein tyrosine kinase polypeptide.  
     
     
         4 . The method of  claim 1  or  2  wherein the protein tyrosine kinase polypeptide of the protein tyrosine kinase polypeptide/adaptor polypeptide complex is an intracellular, cytoplasmic protein tyrosine kinase polypeptide.  
     
     
         5 . The method of  claim 4  wherein the intracellular, cytoplasmic protein tyrosine kinase polypeptide is a BCR/ABL intracellular, cytoplasmic protein tyrosine kinase polypeptide.  
     
     
         6 . The method of  claim 1  or  2  wherein the protein tyrosine kinase polypeptide of the protein tyrosine kinase polypeptide/adaptor polypeptide complex is an intracellular, nuclear protein tyrosine kinase polypeptide.  
     
     
         7 . The method of  claim 1  or  2  wherein the adaptor polypeptide of the protein tyrosine kinase polypeptide/adaptor polypeptide complex is a GRB polypeptide.  
     
     
         8 . The method of  claim 7  wherein the GRB polypeptide is a GRB-2 polypeptide.  
     
     
         9 . The method of  claim 7  wherein the GRB polypeptide is a GRB-1, GRB-4, GRB-7, or GRB-10 polypeptide.  
     
     
         10 . The method of  claim 1  or  2  wherein the protein tyrosine kinase polypeptide of the protein tyrosine kinase polypeptide/adaptor polypeptide complex is a BCR/ABL intracellular, cytoplasmic protein tyrosine kinase polypeptide and the adaptor polypeptide of the protein tyrosine kinase polypeptide/adaptor polypeptide complex is a GRB-2 polypeptide so that the protein tyrosine kinase polypeptide/adaptor polypeptide complex is a BCR/ABL polypeptide/adaptor polypeptide complex.  
     
     
         11 . The method of  claim 1  or  2  wherein the peptide comprising a functional portion of a member of the protein tyrosine kinase polypeptide/adaptor polypeptide comprises a peptide having at least 1 phosphorylated tyrosine amino acid residue.  
     
     
         12 . The method of  claim 1  or  2  wherein the peptide comprising a functional portion of a member of the protein tyrosine kinase polypeptide/adaptor polypeptide complex comprises a functional portion of a phosphorylation domain.  
     
     
         13 . The method of  claim 1  or  2  wherein the peptide comprising a functional portion of a member of the protein tyrosine kinase polypeptide/adaptor polypeptide complex comprises a functional portion of an SH2 domain.  
     
     
         14 . The method of  claim 1  or  2  wherein the peptide comprising a functional portion of a member of the protein tyrosine kinase polypeptide/adaptor polypeptide complex comprises a functional portion of an SH3 domain.  
     
     
         15 . The method of  claim 1  or  2  wherein the peptide comprising a functional portion of a member of the protein tyrosine kinase polypeptide/adaptor polypeptide complex comprises at least 4 consecutive amino acid residues of an SH2-binding domain.  
     
     
         16 . The method of  claim 1  or  2  wherein the peptide comprising a functional portion of a member of the protein tyrosine kinase polypeptide/adaptor polypeptide complex comprises a functional portion of an SH3-binding domain.  
     
     
         17 . The method of  claim 16  wherein the functional portion of an SH3-binding domain is at least 4 amino acid residues in length.  
     
     
         18 . The method of  claim 16  wherein the functional portion of an SH3-binding domain is at least 10 amino acid residues in length.  
     
     
         19 . The method of  claim 1  or  2  wherein the compound identified is further capable of disrupting a protein tyrosine kinase polypeptide/adaptor polypeptide complex.  
     
     
         20 . The method of  claim 1  or  2  wherein the compound capable of disrupting a protein tyrosine kinase polypeptide/adaptor polypeptide complex is further capable of modulating a cell proliferative disorder involving a protein tyrosine kinase polypeptide/adaptor polypeptide complex.  
     
     
         21 . The method of  claim 20  wherein the compound identified is further capable of disrupting a BCR/ABL polypeptide/GRB-2 polypeptide complex.  
     
     
         22 . The method of  claim 21  wherein the compound capable of disrupting a BCR/ABL/GRB-2 polypeptide/GRB-2 polypeptide complex is further capable of modulating an cell proliferative disorder involving a BCR/ABL polypeptide/GRB-2 polypeptide complex.  
     
     
         23 . The method of  claim 22  wherein the cell proliferative disorder is a chronic myelogenous leukemia.  
     
     
         24 . The method of  claim 22  wherein the cell proliferative disorder is an acute lymphocytic leukemia.  
     
     
         25 . The method of  claim 22  wherein the cell proliferative disorder is an acute myelogenous leukemia.  
     
     
         26 . A method for identifying a compound to be tested for an ability to modulate an cell proliferative disorder involving a protein tyrosine phosphatase polypeptide/adaptor polypeptide complex comprising: 
 (a) exposing at least one compound to a peptide comprising a functional portion of a member of the protein tyrosine phosphatase polypeptide/adaptor polypeptide complex for a time sufficient to allow binding of the compound to the functional portion of the member;    (b) removing non-bound compounds; and    (c) detecting the presence of the compound bound to the functional portion of the member of the protein tyrosine phosphatase polypeptide/adaptor polypeptide complex, thereby identifying a compound to be tested for an ability to modulate an cell proliferative disorder involving a protein tyrosine phosphatase polypeptide/adaptor polypeptide complex.    
     
     
         27 . A method for identifying a compound capable of modulating a cell proliferative disorder involving a protein tyrosine phosphatase polypeptide/adaptor polypeptide complex comprising, exposing the compound to a protein tyrosine phosphatase polypeptide/adaptor polypeptide complex for a time sufficient to allow disruption of the complex, and detecting the disruption of the protein tyrosine phosphatase polypeptide/adaptor polypeptide complex.  
     
     
         28 . The method of  claim 26  or  27  wherein the protein tyrosine phosphatase polypeptide of the protein tyrosine phosphatase polypeptide/adaptor polypeptide complex is a transmembrane, receptor protein tyrosine phosphatase polypeptide.  
     
     
         29 . The method of  claim 26  or  27  wherein the protein tyrosine phosphatase polypeptide of the protein tyrosine phosphatase polypeptide/adaptor polypeptide complex is an intracellular, cytoplasmic protein tyrosine phosphatase polypeptide.  
     
     
         30 . The method of  claim 26  or  27  wherein the adaptor polypeptide of the protein tyrosine phosphatase polypeptide/adaptor polypeptide complex is a GRB polypeptide.  
     
     
         31 . The method of  claim 30  wherein the GRB polypeptide is a GRB-2 polypeptide.  
     
     
         32 . The method of  claim 30  wherein the GRB polypeptide is a GRB-1, GRB-4, GRB-7, or GRB-10 polypeptide.  
     
     
         33 . The method of  claim 26  or  27  wherein the peptide comprising a functional portion of a member of the protein tyrosine phosphatase polypeptide/adaptor polypeptide complex comprises a peptide having at least 1 phosphorylated amino acid residue.  
     
     
         34 . The method of  claim 26  or  27  wherein the peptide comprising a functional portion of a member of the protein tyrosine phosphatase polypeptide/adaptor polypeptide complex comprises a functional portion of an SH2 domain.  
     
     
         35 . The method of  claim 26  or  27  wherein the peptide comprising a functional portion of a member of the protein tyrosine phosphatase polypeptide/adaptor polypeptide complex comprises a functional portion of an SH3 domain.  
     
     
         36 . The method of  claim 26  or  27  wherein the peptide comprising a functional portion of a member of the protein tyrosine phosphatase polypeptide/adaptor polypeptide complex comprises at least 4 consecutive amino acid residues of an SH2-binding domain.  
     
     
         37 . The method of  claim 26  or  27  wherein the peptide comprising a functional portion of a member of the protein tyrosine phosphatase polypeptide/adaptor polypeptide complex comprises a functional portion of an SH3-binding domain.  
     
     
         38 . The method of  claim 37  wherein the functional portion of an SH3-binding domain is at least 4 amino acid residues in length.  
     
     
         39 . The method of  claim 37  wherein the functional portion of an SH3-binding domain is at least 10 amino acid residues in length.  
     
     
         40 . A method for identifying a compound to be tested for an ability to modulate a cell proliferative disorder involving a transmembrane, receptor protein tyrosine kinase polypeptide/adaptor polypeptide complex comprising: 
 (a) contacting the compound to a cell capable of forming a receptor tyrosine kinase polypeptide/adaptor polypeptide complex for a time sufficient to allow binding of the compound to the receptor protein tyrosine kinase polypeptide of the receptor protein tyrosine kinase polypeptide/adaptor polypeptide complex;    (b) detecting the level of receptor protein tyrosine kinase polypeptide/adaptor polypeptide complex present in the cell of step (a);    (c) detecting the level of receptor protein tyrosine kinase polypeptide/adaptor polypeptide complex present in a cell of the type in step (a) that has not contacted the compound; and    (d) comparing the level of receptor protein tyrosine kinase polypeptide/adaptor polypeptide complex detected in step (b) to the level detected in step (c), so that if the level detected in step (c) is greater than the level detected in step (b), a compound to be tested for an ability to modulate an cell proliferative disorder involving a receptor protein tyrosine kinase polypeptide/adaptor polypeptide complex is identified.    
     
     
         41 . A method for identifying a compound to be tested for an ability to modulate an cell proliferative disorder involving a transmembrane, receptor protein tyrosine phosphatase polypeptide/adaptor polypeptide complex comprising: 
 (a) contacting the compound to a cell capable of forming a receptor tyrosine phosphatase polypeptide/adaptor polypeptide complex for a time sufficient to allow binding of the compound to the receptor protein tyrosine phosphatase polypeptide of the receptor protein tyrosine phosphatase polypeptide/adaptor polypeptide complex;    (b) detecting the level of receptor protein tyrosine phosphatase polypeptide/adaptor polypeptide complex present in the cell of step (a);    (c) detecting the level of receptor protein tyrosine phosphatase polypeptide/adaptor polypeptide complex present in a cell of the type in step (a) that has not contacted the compound; and    (d) comparing the level of receptor protein tyrosine phosphatase polypeptide/adaptor polypeptide complex detected in step (b) to the level detected in step (c), so that if the level detected in step (c) is greater than the level detected in step (b), a compound to be tested for an ability to modulate an cell proliferative disorder involving a receptor protein tyrosine phosphatase polypeptide/adaptor polypeptide complex has been identified.    
     
     
         42 . The method of  claim 40  or  41  wherein the adaptor polypeptide is a GRB polypeptide.  
     
     
         43 . The method of  claim 42  wherein the GRB polypeptide is a GRB-2 polypeptide.  
     
     
         44 . The method of  claim 42  wherein the GRB polypeptide is a GRB-1, GRB-4, GRB-7, or GRB-10 polypeptide.  
     
     
         45 . A method of modulating a cell proliferative disorder involving a protein tyrosine kinase polypeptide/adaptor polypeptide complex comprising, contacting a cell capable of forming a protein tyrosine kinase polypeptide/adaptor polypeptide complex with an amount of a compound sufficient to disrupt protein tyrosine kinase polypeptide/adaptor polypeptide complexes of the cell so that the cell proliferative disorder is modulated.  
     
     
         46 . A method of modulating an cell proliferative disorder involving a protein tyrosine kinase polypeptide/adaptor polypeptide complex in a mammal comprising, administering to the mammal an amount, of a compound sufficient to disrupt the protein tyrosine kinase polypeptide/adaptor polypeptide complex so that the cell proliferative disorder is modulated.  
     
     
         47 . The method of  claim 45  or  46  wherein the protein tyrosine kinase polypeptide of the protein tyrosine kinase polypeptide/adaptor polypeptide complex is a transmembrane, receptor protein tyrosine kinase polypeptide.  
     
     
         48 . The method of  claim 45  or  46  wherein the protein tyrosine kinase polypeptide of the protein tyrosine kinase polypeptide/adaptor polypeptide complex is an intracellular, cytoplasmic protein tyrosine kinase polypeptide.  
     
     
         49 . The method of  claim 48  wherein the intracellular, cytoplasmic protein tyrosine kinase polypeptide is a BCR/ABL intracellular, cytoplasmic protein tyrosine kinase polypeptide.  
     
     
         50 . The method of  claim 45  or  46  wherein the protein tyrosine kinase polypeptide of the protein tyrosine kinase polypeptide/adaptor polypeptide complex is an intracellular, nuclear protein tyrosine kinase polypeptide.  
     
     
         51 . The method of  claim 45  or  46  wherein the adaptor polypeptide of the protein tyrosine kinase polypeptide/adaptor polypeptide complex is a GRB polypeptide.  
     
     
         52 . The method of  claim 51  wherein the GRB polypeptide is a GRB-2 polypeptide.  
     
     
         53 . The method of  claim 51  wherein the GRB polypeptide is a GRB-1, GRB-4, GRB-7, or GRB-10 polypeptide.  
     
     
         54 . The method of  claim 45  or  46  wherein the protein tyrosine kinase polypeptide of the protein tyrosine kinase polypeptide/adaptor polypeptide complex is a BCR/ABL intracellular, cytoplasmic protein tyrosine kinase polypeptide and the adaptor polypeptide of the protein tyrosine kinase polypeptide/adaptor polypeptide complex is a GRB-2 polypeptide so that the protein tyrosine kinase polypeptide/adaptor polypeptide complex is a BCR/ABL polypeptide/adaptor polypeptide complex.  
     
     
         55 . The method of  claim 54  wherein the cell proliferative disorder modulated is a chronic myelogenous leukemia.  
     
     
         56 . The method of  claim 54  wherein the cell proliferative disorder modulated is an acute lymphocytic leukemia.  
     
     
         57 . The method of  claim 54  wherein the cell proliferative disorder modulated is an acute myelogenous leukemia.  
     
     
         58 . A method of modulating a cell proliferative disorder involving a protein tyrosine phosphatase polypeptide/adaptor polypeptide complex comprising, contacting a cell capable of forming a protein tyrosine phosphatase polypeptide/adaptor polypeptide complex with an amount of a compound sufficient to disrupt protein tyrosine phosphatase polypeptide/adaptor polypeptide complexes of the cell so that the cell proliferative disorder is modulated.  
     
     
         59 . A method of modulating a cell proliferative disorder involving a protein tyrosine phosphatase polypeptide/adaptor polypeptide complex in a mammal comprising, administering to the mammal an amount of a compound sufficient to disrupt protein tyrosine phosphatase polypeptide/adaptor polypeptide complex so that the cell proliferative disorder is modulated.  
     
     
         60 . The method of  claim 58  or  59  wherein the protein tyrosine phosphatase polypeptide of the protein tyrosine phosphatase polypeptide/adaptor polypeptide complex is a transmembrane, receptor protein tyrosine phosphatase polypeptide.  
     
     
         61 . The method of  claim 58  or  59  wherein the protein tyrosine phosphatase polypeptide of the protein tyrosine phosphatase polypeptide/adaptor polypeptide complex is an intracellular, cytoplasmic protein tyrosine phosphatase polypeptide.  
     
     
         62 . The method of  claim 58  or  59  wherein the adaptor polypeptide of the protein tyrosine phosphatase polypeptide/adaptor polypeptide complex is a GRB polypeptide.  
     
     
         63 . The method of  claim 62  wherein the GRB polypeptide is a GRB-2 polypeptide.  
     
     
         64 . The method of  claim 58  or  59  wherein the GRB polypeptide is a GRB-1, GRB-4, GRB-7, or GRB-10 polypeptide.  
     
     
         65 . An isolated BCR/ABL polypeptide/GRB-2 adaptor polypeptide complex.