2-3-bis(2-hydroxyethylsulfanyl)-(1-4)naphthoquinone and Breast-Neoplasms

Mitogen-activated protein kinase phosphatase (MKP)-1 is a dual-specificity phosphatase that negatively regulates the activity of mitogen-activated kinases and that is overexpressed in human tumors. Contemporary studies suggest that induction of MKP-1 during chemotherapy may limit the efficacy of clinically used antineoplastic agents. Thus, MKP-1 is a rational target to enhance anticancer drug activity, but suitable small-molecule inhibitors of MKP-1 are currently unavailable. Here, we have used a high-content, multiparameter fluorescence-based chemical complementation assay for MKP activity in intact mammalian cells to evaluate the cellular MKP-1 and MKP-3 inhibitory activities of four previously described, quinone-based, dual-specificity phosphatase inhibitors, that is, NSC 672121, NSC 95397, DA-3003-1 (NSC 663284), and JUN-1111. All compounds induced formation of reactive oxygen species in mammalian cells, but only one (NSC 95397) inhibited cellular MKP-1 and MKP-3 with an IC(50) of 13 mumol/L. Chemical induction of MKP-1 by dexamethasone protected cells from paclitaxel-induced apoptosis but had no effect on NSC 95397. NSC 95397 phenocopied the effects of MKP-1 small inhibitory RNA by reversing the cytoprotective effects of dexamethasone in paclitaxel-treated cells. Isobologram analysis revealed synergism between paclitaxel and NSC 95397 only in the presence of dexamethasone. The data show the power of a well-defined cellular assay for identifying cell-active inhibitors of MKPs and support the hypothesis that small-molecule inhibitors of MKP-1 may be useful as antineoplastic agents under conditions of high MKP-1 expression.

Topics: Antineoplastic Combined Chemotherapy Protocols; Breast Neoplasms; Cell Survival; Cytoprotection; Dexamethasone; Drug Evaluation, Preclinical; Drug Resistance, Neoplasm; Drug Synergism; Dual Specificity Phosphatase 1; Dual Specificity Phosphatase 6; HeLa Cells; Humans; Mitogen-Activated Protein Kinase Phosphatases; Models, Biological; Naphthoquinones; Paclitaxel; Quinones; Reactive Oxygen Species; Tumor Cells, Cultured

Cdc25 phosphatases are important in cell cycle control and activate cyclin-dependent kinases (Cdk). Efforts are currently under way to synthesize specific small-molecule Cdc25 inhibitors that might have anticancer properties. NSC 95397, a protein tyrosine phosphatase antagonist from the National Cancer Institute library, was reported to be a potent Cdc25 inhibitor. We have synthesized two hydroxyl derivatives of NSC 95397, monohydroxyl-NSC 95397 and dihydroxyl-NSC 95397, which both have enhanced activity for inhibiting Cdc25s. The new analogues, especially dihydroxyl-NSC 95397, potently inhibited the growth of human hepatoma and breast cancer cells in vitro. They influenced two signaling pathways. The dual phosphorylation of extracellular signal-regulated kinase 1/2 (ERK1/2) was induced, likely due to inhibition of the ERK phosphatase activity in Hep 3B cell lysate but not the dual specificity ERK phosphatase MKP-1. They also inhibited Cdc25 enzymatic activities and induced tyrosine phosphorylation of the Cdc25 target Cdks. Addition of hydroxyl groups to the naphthoquinone ring thus enhanced the potency of NSC 95397. These two new compounds may be useful probes for the biological functions of Cdc25s and have the potential for disrupting the cell cycle of growing tumor cells.

Topics: Antineoplastic Agents; Biotinylation; Blotting, Western; Breast Neoplasms; Carcinoma, Hepatocellular; cdc25 Phosphatases; Cell Cycle Proteins; Cell Line, Tumor; Cell Proliferation; Cell-Free System; DNA; Dual Specificity Phosphatase 1; Flavonoids; Flow Cytometry; Humans; Immediate-Early Proteins; Immunoprecipitation; Mitogen-Activated Protein Kinase 1; Mitogen-Activated Protein Kinase 3; Naphthoquinones; Phosphoprotein Phosphatases; Phosphorylation; Protein Phosphatase 1; Protein Tyrosine Phosphatases; Signal Transduction; Time Factors; Transfection; Tyrosine