Bcr-Abl directed compositions and uses for inhibiting Philadelphia chromosome stimulated cell growth

Abstract

Compositions comprising or encoding one or more peptides that inhibit the Bcr—Abl oncoprotein and that bind to molecules involved in Bcr—Abl function are disclosed. The peptides and polypeptides inhibit the growth of, and induce cell death of, Philadelphia chromosome-positive leukemia cells expressing the Bcr—Abl oncoprotein. Methods for treating leukemias (e.g., CML, ALL and AML), including autologous bone marrow transplant therapy, using the peptide and polypeptide compositions of the invention are also provided.

1 . A composition comprising a purified peptide or polypeptide, of between about 4 and about 500 amino acids in length, comprising a contiguous amino acid sequence from the Bcr—Abl protein that includes tyrosine 177, tyrosine 283 or tyrosine 360, which peptide or polypeptide becomes phosphorylated on tyrosine upon contact with Bcr—Abl.  
     
     
         2 . The composition of  claim 1 , wherein said composition comprises a peptide that includes tyrosine 177.  
     
     
         3 . The composition of  claim 2 , wherein said peptide comprises the sequence of SEQ ID NO:8.  
     
     
         4 . The composition of  claim 1 , wherein said composition comprises a peptide that includes tyrosine 283.  
     
     
         5 . The composition of  claim 4 , wherein said peptide comprises the sequence of SEQ ID NO:11.  
     
     
         6 . The composition of  claim 1 , wherein said composition comprises a peptide that includes tyrosine 360.  
     
     
         7 . The composition of  claim 6 , wherein said peptide comprises the sequence of SEQ ID NO: 10.  
     
     
         8 . The composition of  claim 6 , wherein said peptide comprises the sequence of SEQ ID NO:22.  
     
     
         9 . The composition of  claim 6 , wherein said peptide further comprises a phosphorylated Serine that corresponds to Serine 354.  
     
     
         10 . The composition of  claim 1 , wherein said composition comprises a first peptide that includes tyrosine 177 and a second peptide that includes tyrosine 283.  
     
     
         11 . The composition of  claim 1 , wherein said composition comprises a first peptide that includes tyrosine 177, a second peptide that includes tyrosine 283 and a third peptide that includes tyrosine 360.  
     
     
         12 . The composition of  claim 1 , wherein said composition comprises a single peptide that includes tyrosine 177 and tyrosine 283.  
     
     
         13 . The composition of  claim 1 , wherein said composition comprises a single peptide that includes tyrosine 177, tyrosine 283 and tyrosine 360.  
     
     
         14 . The composition of  claim 13 , wherein said peptide comprises the sequence of SEQ ID NO:28.  
     
     
         15 . The composition of  claim 1 , wherein said peptide is between about 10 and about 350 amino acids in length.  
     
     
         16 . The composition of  claim 15 , wherein said peptide is between about 10 and about 100 amino acids in length.  
     
     
         17 . The composition of  claim 16 , wherein said peptide is between about 10 and about 50 amino acids in length.  
     
     
         18 . The composition of any preceding claim, further comprising: 
 (a) a purified Shc-binding peptide that binds to an Abl SH3 binding protein-rich region of Shc;    (b) a purified Crkl-binding peptide that binds to a proline-rich Abl binding site on Crkl;    (c) a purified Ras Gap-binding peptide that binds to an SH2 domain of p120 Ras Gap;    (d) a purified Bcr-binding peptide or protein that binds to an N-terminal coiled-coil region of Bcr; or    (e) a purified Grb2-binding peptide or protein that binds to an N-terminal coiled-coil region of Bcr.    
     
     
         19 . The composition of  claim 18 , wherein said Ras Gap-binding peptide comprises the sequence of SEQ ID NO:11 or SEQ ID NO:12.  
     
     
         20 . The composition of  claim 18 , wherein said Bcr-binding peptide comprises the sequence of any one of SEQ ID NO:2 through SEQ ID NO:7.  
     
     
         21 . The composition of  claim 18 , wherein said Grb2-binding peptide comprises the sequence of SEQ ID NO: 13 or SEQ ID NO:8.  
     
     
         22 . The composition of any preceding claim, wherein said peptide or polypeptide is further associated with a liposome.  
     
     
         23 . The composition of any preceding claim, wherein said peptide or polypeptide is comprised in a pharmaceutically acceptable carrier.  
     
     
         24 . A composition according to any preceding claim, for use in enriching Philadelphia chromosome-negative cells in a mixture of cells containing Philadelphia chromosome-positive cells.  
     
     
         25 . An expression vector comprising a DNA sequence that expresses a peptide or polypeptide of between about 4 and about 500 amino acids in length that includes a contiguous amino acid sequence from the Bcr—Abl protein that includes tyrosine 177, tyrosine 283 or tyrosine 360, which peptide or polypeptide becomes phosphorylated on tyrosine upon contact with Bcr—Abl.  
     
     
         26 . The vector of  claim 25 , further defined as a retroviral vector.  
     
     
         27 . The vector of  claim 25 , further defined as an adenoviral vector.  
     
     
         28 . The vector of  claim 25 , further defined as a plasmid associated with a liposome.  
     
     
         29 . A vector according to any one of claims  25 - 28 , for use in enriching Philadelphia chromosome-negative cells in a mixture of cells containing Philadelphia chromosome-positive cells.  
     
     
         30 . A method for enriching Philadelphia chromosome-negative cells in a mixture of cells containing Philadelphia chromosome-positive cells, comprising contacting said cells with a composition in accordance with any one of claims  1  through  23  or a vector in accordance with any one of claims  25  through  28 , in an amount effective to enrich for Philadelphia chromosome-negative cells in said mixture.  
     
     
         31 . The method of  claim 30 , wherein said cells comprise bone marrow cells.  
     
     
         32 . The method of  claim 30 , wherein Philadelphia chromosome-negative cells are enriched relative to numbers naturally occurring in a bone marrow sample containing Philadelphia chromosome-positive cells.  
     
     
         33 . A method of purging a bone marrow sample of Philadelphia chromosome-positive cells, comprising contacting a bone marrow sample that contains Philadelphia chromosome-positive cells with a composition in accordance with any one of claims  1  through  23  or a vector in accordance with any one of claims  25  through  28 , in an amount effective to reduce the numbers of Philadelphia chromosome-positive cells in said bone marrow sample.  
     
     
         34 . The method of  claim 33 , wherein said bone marrow sample is obtained from a patient having or suspected of having CML, AML or ALL.  
     
     
         35 . A method of treating a patient with Philadelphia chromosome-positive leukemia, comprising treating a bone marrow sample of said patient with a composition in accordance with any one of claims  1  through  23  or a vector in accordance with any one of claims  25  through  28  in an amount effective to prepare an essentially leukemia cell-free autologous bone marrow sample and administering said treated sample to said patient.  
     
     
         36 . A method of treating a patient with Philadelphia chromosome-positive leukemia, comprising obtaining a bone marrow sample from said patient, contacting said sample ex vivo with a composition in accordance with any one of claims  1  through  23  or a vector in accordance with any one of claims  25  through  28 , in an amount effective and for a period of time sufficient to purge Philadelphia chromosome-positive cells from said sample and re-administering said purged sample to said patient.  
     
     
         37 . A method of treating a patient with Philadelphia chromosome-positive leukemia, comprising administering to said patient a therapeutically effective amount of a composition in accordance with any one of claims  1  through  23  or a vector in accordance with any one of claims  25  through  28 .  
     
     
         38 . Use of a composition according to any one of claims  1  through  23  or a vector according to any one of claims  25  through  28  in the manufacture of a medicament for treating an animal with leukemia associated with Philadelphia chromosome-positive cells, wherein the medicament is administered to a tissue sample removed from said animal in an amount effective to enrich for Philadelphia chromosome-negative cells in said tissue sample and wherein the treated tissue sample is re-administered to said animal.  
     
     
         39 . Use of a composition according to any one of claims  1  through  23  or a vector according to any one of claims  25  through  28  in the manufacture of a medicament for treating an animal with leukemia associated with Philadelphia chromosome-positive cells, wherein the medicament is administered to an animal with leukemia in an amount effective to enrich for Philadelphia chromosome-negative cells in said animal.