title: PM-20, a novel inhibitor of Cdc25A, induces extracellular signal-regulated kinase 1/2 phosphorylation and inhibits hepatocellular carcinoma growth in vitro and in vivo.
abstract: We have synthesized several new phenyl maleimide compounds, which are potent growth inhibitors of several human tumor cell lines. Among these, PM-20 was the most potent with an IC50 of 700 nmol/L for Hep3B human hepatoma cell growth. Two other derivatives, PM-26 and PM-38, did not inhibit Hep3B cell growth even at 100 micromol/L. Interestingly, under identical experimental conditions, PM-20 inhibited DNA synthesis of primary cultures of normal hepatocytes at a 10-fold higher concentration than that needed to inhibit the DNA synthesis of the Hep3B hepatoma cells. PM-20 affected two cellular signaling pathways in Hep3B cells: Cdc25 phosphatase and extracellular signal-regulated kinase (ERK) 1/2. It competitively inhibited the activity of Cdc25 (preferentially Cdc25A) by binding to the active site, likely through the catalytic cysteine, but did not inhibit PTP1B, CD45, or MKP-1 phosphatases. As a result of its action, tyrosine phosphorylation of the cellular Cdc25A substrates Cdk2 and Cdk4 was induced. It also induced strong and persistent phosphorylation of the Cdc25A substrate ERK1/2. Hep3B cell lysates were found to contain ERK2 phosphatase(s) activity, which was inhibited by the actions of PM-20. However, activity of exogenous dual-specificity ERK2 phosphatase MKP1 was not inhibited. Induction of ERK1/2 phosphorylation correlated with the potency of growth inhibition in tumor cell lines and inhibition of ERK1/2 phosphorylation by the mitogen-activated protein kinase (MAPK)/ERK kinase 1/2 inhibitor U0126 or overexpression of the cdc25A gene in Hep3B cells antagonized the growth inhibitory actions of PM-20. Growth of transplantable rat hepatoma cells in vivo was also inhibited by PM-20 action with a concomitant induction of pERK in the tumors. The mechanism(s) of growth inhibition of Hep3B hepatoma cells by the phenyl maleimide PM-20 involves prolonged ERK1/2 phosphorylation, likely resulting from inhibition of the ERK phosphatase Cdc25A. PM-20 thus represents a novel class of tumor growth inhibitor that inhibits mainly Cdc25A, is dependent on ERK activation, and has a considerable margin of selectivity for tumor cells compared with normal cells.
authors.affiliation: Liver Cancer Center, Starzl Transplantation Institute, University of Pittsburgh, 200 Lothrop Street, Pittsburgh, PA 15260, USA.
authors.forename: Siddhartha
authors.initials: S
authors.lastname: Kar
authors.forename: Meifang
authors.initials: M
authors.lastname: Wang
authors.forename: Wei
authors.initials: W
authors.lastname: Yao
authors.forename: Christopher J
authors.initials: CJ
authors.lastname: Michejda
authors.forename: Brian I
authors.initials: BI
authors.lastname: Carr
mesh_terms: D000818:Animals; D006528:Carcinoma, Hepatocellular / Q000188:drug therapy* / Q000201:enzymology / Q000473:pathology; D004789:Enzyme Activation / Q000187:drug effects; D004791:Enzyme Inhibitors / Q000627:therapeutic use*; D006801:Humans; D066298:In Vitro Techniques; D008113:Liver Neoplasms / Q000188:drug therapy* / Q000201:enzymology / Q000473:pathology; D008301:Maleimides / Q000737:chemistry / Q000627:therapeutic use*; D019950:Mitogen-Activated Protein Kinase 1 / Q000378:metabolism*; D048052:Mitogen-Activated Protein Kinase 3 / Q000378:metabolism*; D020928:Mitogen-Activated Protein Kinases / Q000378:metabolism; D010766:Phosphorylation / Q000187:drug effects; D051381:Rats; D014407:Tumor Cells, Cultured; D020687:cdc25 Phosphatases / Q000037:antagonists & inhibitors* / Q000378:metabolism
chemical_list: D004791:Enzyme Inhibitors; D008301:Maleimides; C043592:maleimide; D019950:Mitogen-Activated Protein Kinase 1; D048052:Mitogen-Activated Protein Kinase 3; D020928:Mitogen-Activated Protein Kinases; C517004:CDC25A protein, human; C517007:Cdc25a protein, rat; D020687:cdc25 Phosphatases