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Assaying pharmacodynamic endpoints with targeted therapy: Flavopiridol and 17AAG induced dephosphorylation of histone H1.5 in acute myeloid leukemia
Author(s) -
Wang Liwen,
Harshman Sean W.,
Liu Shujun,
Ren Chen,
Xu Hua,
Sallans Larry,
Grever Michael,
Byrd John C.,
Marcucci Guido,
Freitas Michael A.
Publication year - 2010
Publication title -
proteomics
Language(s) - English
Resource type - Journals
SCImago Journal Rank - 1.26
H-Index - 167
eISSN - 1615-9861
pISSN - 1615-9853
DOI - 10.1002/pmic.201000080
Subject(s) - phosphoproteomics , phosphorylation , cancer research , dephosphorylation , biology , histone deacetylase , kinase , histone deacetylase inhibitor , histone h1 , histone , protein phosphorylation , biochemistry , protein kinase a , phosphatase , gene
Histone H1 is commonly used to assay kinase activity in vitro . As many promising targeted therapies affect kinase activity of specific enzymes involved in cancer transformation, H1 phosphorylation can serve as potential pharmacodynamic marker for drug activity within the cell. In this study we utilized a phosphoproteomic workflow to characterize histone H1 phosphorylation changes associated with two targeted therapies in the Kasumi‐1 acute myeloid leukemia cell line. The phosphoproteomic workflow was first validated with standard casein phosphoproteins and then applied to the direct analysis of histone H1 from Kasumi‐1 nuclear lysates. Ten H1 phosphorylation sites were identified on the H1 variants, H1.2, H1.3, H1.4, H1.5 and H1.x. LC MS profiling of intact H1s demonstrated global dephosphorylation of H1.5 associated with therapy by the cyclin‐dependent kinase inhibitor, flavopiridol and the Heat Shock Protein 90 inhibitor, 17‐(Allylamino)‐17‐demethoxygeldanamycin. In contrast, independent treatments with a nucleotide analog, proteosome inhibitor and histone deacetylase inhibitor did not exhibit decreased H1.5 phosphorylation. The data presented herein demonstrate that potential of histones to assess the cellular response of reagents that have direct and indirect effects on kinase activity that alters histone phosphorylation. As such, this approach may be a highly informative marker for response to targeted therapies influencing histone phosphorylation.