Diagnostic and Therapeutic Targets for Leukemia

Abstract

The present invention relates to methods of identifying candidate compounds for the treatment of leukemia and diagnostic methods based on histone methylation and HoxA5 promoter activity.

1 - 13 . (canceled) 
     
     
         14 . A method of identifying a candidate compound for the prevention or treatment of leukemia, the method comprising:
 contacting a CALM-AF10 fusion protein with a test compound under conditions sufficient for binding of the test compound to the CALM-AF10 fusion protein; and   detecting binding between the test compound and the CALM-AF10 fusion protein,   wherein binding of the test compound to the CALM-AF10 fusion protein indicates that the test compound is a candidate compound for the prevention or treatment of leukemia.   
     
     
         15 - 17 . (canceled) 
     
     
         18 . The method of  claim 14 , wherein the leukemia is T cell acute lymphoid leukemia (T-ALL) or acute myeloid leukemia subtype M0/1 (AML-M0/1). 
     
     
         19 . A method of identifying a candidate compound for the prevention or treatment of leukemia, the method comprising:
 contacting a nucleic acid comprising a HoxA5 promoter region with a CALM polypeptide or CALM-AF10 fusion protein in the presence of a test compound under conditions sufficient for binding of the CALM polypeptide or CALM-AF10 fusion protein to the HoxA5 promoter; and   detecting interaction of the CALM polypeptide or CALM-AF10 fusion protein with the HoxA5 promoter,   wherein a reduction in interaction between the CALM polypeptide or CALM-AF10 fusion protein with the HoxA5 promoter in the presence of the test compound as compared with the level of interaction in the absence of the test compound indicates that the test compound is a candidate compound for the prevention or treatment of leukemia.   
     
     
         20 . The method of  claim 19 , wherein the nucleic acid further comprises exon 1 or exon 2 of the HoxA5 gene. 
     
     
         21 . The method of  claim 19 , wherein the nucleic acid further comprises the intron between exons 1 and 2 of the HoxA5 gene. 
     
     
         22 . (canceled) 
     
     
         23 . The method of  claim 19 , wherein CALM polypeptide or CALM-5 AF10 fusion protein interaction with the HoxA5 promoter is evaluated by determining binding of the CALM polypeptide or CALM-AF10 fusion protein to the HoxA5 promoter. 
     
     
         24 . The method of  claim 19 , wherein CALM or CALM-AF10 interaction with the HoxA5 promoter is evaluated by determining histone H3 lysine 79 (H3-K79) methylation of HoxA5. 
     
     
         25 . The method of  claim 19 , wherein CALM or CALM-AF10 interaction with the HoxA5 promoter is evaluated by determining HoxA5 promoter activity. 
     
     
         26 - 31 . (canceled) 
     
     
         32 . The method of  claim 19 , wherein the leukemia is T cell acute lymphoid leukemia (T-ALL) or acute myeloid leukemia subtype M0/1 (AML-M0/1). 
     
     
         33 . A method of identifying a candidate compound for the prevention or treatment of leukemia, the method comprising:
 contacting a nucleic acid comprising a HoxA5 promoter region with a DOT1L polypeptide and a CALM-AF10 fusion protein in the presence of a test compound under conditions sufficient for binding of the DOT1L polypeptide and the CALM-AF10 fusion protein to form a complex and for the complex to bind to the HoxA5 promoter; and   detecting interaction of the DOT1L/CALM-AF10 complex with the HoxA5 promoter,   wherein a reduction in interaction of the DOT1L/CALM-AF10 complex with the HoxA5 promoter in the presence of the test compound as compared with the level of interaction in the absence of the test compound indicates that the test compound is a candidate compound for the prevention or treatment of leukemia.   
     
     
         34 . The method of  claim 33 , wherein the nucleic acid further comprises exon 1 or exon 2 of the HoxA5 gene. 
     
     
         35 . The method of  claim 33 , wherein the nucleic acid further comprises the intron between exons 1 and 2 of the HoxA5 gene. 
     
     
         36 . (canceled) 
     
     
         37 . The method of  claim 33 , wherein interaction of the DOT1L/CALM-AF10 complex with the HoxA5 promoter is evaluated by determining binding of the DOT1L/CALM-AF10 complex to the HoxA5 promoter. 
     
     
         38 . The method of  claim 33 , wherein interaction of the DOT1L/CALM-AF10 complex with the HoxA5 promoter is evaluated by determining histone H3 lysine 79 (H3-K79) methylation of the HoxA5 promoter. 
     
     
         39 . The method of  claim 33 , wherein interaction of the DOT1L/CALM-AF10 complex with the HoxA5 promoter is evaluated by determining HoxA5 promoter activity. 
     
     
         40 - 45 . (canceled) 
     
     
         46 . The method of  claim 33 , wherein the leukemia is T cell acute lymphoid leukemia (T-ALL) or acute myeloid leukemia subtype M0/1 (AML-M0/1). 
     
     
         47 . A method of identifying a candidate compound for the prevention or treatment of leukemia, the method comprising:
 contacting a nucleic acid comprising a HoxA5 promoter region or any other portion of the HoxA5 gene with a test compound under conditions sufficient for the test compound to bind to the HoxA5 promoter region or the any other portion of the HoxA5 gene; and   detecting binding between the test compound and the HoxA5 promoter or the any other portion of the HoxA5 gene,   wherein binding between the test compound and the HoxA5 promoter or any other portion of the HoxA5 gene indicates that the test compound is a candidate compound for the prevention or treatment of leukemia.   
     
     
         48 - 49 . (canceled) 
     
     
         50 . The method of  claim 47 , wherein the leukemia is T cell acute lymphoid leukemia (T-ALL) or acute myeloid leukemia subtype M0/1 (AML-M0/1). 
     
     
         51 . A method of identifying a candidate compound for the prevention or treatment of leukemia, the method comprising:
 contacting a nucleic acid comprising a HoxA5 promoter region with a test compound under conditions sufficient for HoxA5 promoter activity; and   detecting HoxA5 promoter activity,   wherein a reduction in HoxA5 promoter activity in the presence of the test compound as compared with the level of HoxA5 promoter activity in the absence of the test compound indicates that the test compound is a candidate compound for the prevention or treatment of leukemia.   
     
     
         52 . The method of  claim 51 , wherein the nucleic acid further comprises the HoxA5 exon 1 or the HoxA5 exon 2. 
     
     
         53 . The method of  claim 51 , wherein the nucleic acid further comprises the HoxA5 intron between exon 1 and exon 2. 
     
     
         54 - 58 . (canceled) 
     
     
         59 . The method of  claim 51 , wherein the leukemia is T cell acute lymphoid leukemia (T-ALL) or acute myeloid leukemia subtype M0/1 (AML-M0/1). 
     
     
         60 - 68 . (canceled) 
     
     
         69 . A method of diagnosing whether a subject has or is at risk for developing leukemia or determining the prognosis for the course of the disease, the method comprising determining HoxA5 promoter activity in the subject. 
     
     
         70 . (canceled) 
     
     
         71 . A method of treating a subject afflicted with or at risk for leukemia comprising administering an effective amount of an inhibitory nucleic acid or an antibody directed against HoxA5, CALM, AF10, or CALM-AF10. 
     
     
         72 . The method of  claim 71 , wherein the inhibitory nucleic acid is a shRNA. 
     
     
         73 . The method of  claim 72 , wherein the shRNA is directed against CALM-AF10. 
     
     
         74 . The method of  claim 73 , wherein the shRNA comprises the nucleic acid sequence of SEQ ID NO: 3. 
     
     
         75 . The method of  claim 71 , wherein the leukemia is T cell acute lymphoid leukemia (T-ALL) or acute myeloid leukemia subtype M0/1 (AML-M0/1). 
     
     
         76 . An inhibitory nucleic acid directed against CALM-AF10. 
     
     
         77 . The inhibitory nucleic acid of  claim 76 , wherein the nucleic acid is a shRNA. 
     
     
         78 . The inhibitory nucleic acid of  claim 77 , wherein the shRNA comprises the nucleic acid of SEQ ID NO: 3.