ABSTRACT
Angiogenesis is important for tumor growth and metastasis. Hypoxia in tumors drives this angiogenic response by stabilizing Hypoxia Inducible Factors (HIF) and target genes like Vascular Endothelial Growth Factor (VEGF). HIF stability is regulated by Prolylhydroxylases (PHD)-mediated modification. Iron is an important cofactor in regulating the enzymatic activity of PHDs. Reducing intracellular iron, for instance, mimics hypoxia and induces a pro-angiogenic response. It is hypothesized that increasing the intracellular iron levels will have an opposite, anti-angiogenic effect. We tested this hypothesis by perturbing iron homeostasis in endothelial cells using a unique form of iron, Ferric Ammonium Citrate (FAC). FAC is a cell-permeable form of iron, which can passively enter into cells bypassing the transferrin receptor mediated uptake of transferrin-bound iron. Our studies show that FAC does not decrease the levels of HIF-1α and HIF-2α in endothelial cells but inhibits the autocrine stimulation of VEGF-Vascular Endothelial Growth Factor Receptor-2 (VEGFR-2) system by blocking receptor tyrosine kinase phosphorylation. FAC inhibits VEGF-induced endothelial cell proliferation, migration, tube formation and sprouting. Finally, systemic administration of FAC inhibits VEGF and tumor cell-induced angiogenesis in vivo. In conclusion, our studies show that cell-permeable iron attenuates VEGFR-2 mediated signaling and inhibits tumor angiogenesis.
Subject(s)
Carcinogenesis/metabolism , Endothelial Cells/physiology , Ferric Compounds/metabolism , Hypoxia/metabolism , Iron/metabolism , Neovascularization, Pathologic/metabolism , Quaternary Ammonium Compounds/metabolism , Vascular Endothelial Growth Factor Receptor-2/metabolism , Autocrine Communication , Basic Helix-Loop-Helix Transcription Factors/metabolism , Cell Membrane Permeability , Cell Proliferation , Human Umbilical Vein Endothelial Cells , Humans , Hypoxia-Inducible Factor 1, alpha Subunit/metabolism , Models, Immunological , Neoplasm Invasiveness , Prolyl Hydroxylases/metabolism , Receptor Protein-Tyrosine Kinases/metabolism , Signal Transduction , Vascular Endothelial Growth Factor A/metabolismABSTRACT
OBJECTIVE: Minnelide is a water-soluble pro-drug of triptolide, a natural product. The goal of this study was to evaluate the effectiveness of Minnelide on ovarian cancer growth in vitro and in vivo. METHODS: The effect of Minnelide on ovarian cancer cell proliferation was determined by real time electrical impedance measurements. Multiple mouse models with C200 and A2780 epithelial ovarian cancer cell lines were used to assess the efficacy of Minnelide in inhibiting ovarian cancer growth. RESULTS: Minnelide decreased cell viability of both platinum sensitive and resistant epithelial ovarian cancer cells in vitro. Minnelide with carboplatin showed additive effects in vitro. Minnelide monotherapy increased the survival of mice bearing established ovarian tumors. Minnelide, in combination with carboplatin and paclitaxel, improved overall survival of mice. CONCLUSIONS: Minnelide is a promising pro-drug for the treatment of ovarian cancer, especially when combined with standard chemotherapy.
Subject(s)
Antineoplastic Combined Chemotherapy Protocols/pharmacology , Apoptosis/drug effects , Cell Proliferation/drug effects , Neoplasms, Glandular and Epithelial/drug therapy , Organophosphates/pharmacology , Ovarian Neoplasms/drug therapy , Phenanthrenes/pharmacology , Prodrugs/pharmacology , Animals , Carboplatin/administration & dosage , Carcinoma, Ovarian Epithelial , Cell Line, Tumor , Cell Survival/drug effects , Diterpenes/pharmacology , Drug Resistance, Neoplasm/drug effects , Drug Screening Assays, Antitumor , Electric Impedance , Epoxy Compounds/pharmacology , Female , Humans , Mice , Mice, Nude , Neoplasms, Glandular and Epithelial/pathology , Ovarian Neoplasms/pathology , Paclitaxel/administration & dosage , Xenograft Model Antitumor AssaysABSTRACT
Hypoxia-driven changes in the tumor microenvironment facilitate cancer metastasis. In the present study, we investigated the regulatory cross talk between endocytic pathway, hypoxia, and tumor metastasis. Dynamin 2 (DNM2), a GTPase, is a critical mediator of endocytosis. Hypoxia decreased the levels of DNM2. DNM2 promoter has multiple hypoxia-inducible factor (HIF)-binding sites and genetic deletion of them relieved hypoxia-induced transcriptional suppression. Interestingly, DNM2 reciprocally regulated HIF. Inhibition of DNM2 GTPase activity and dominant-negative mutant of DNM2 showed a functional role for DNM2 in regulating HIF. Furthermore, the opposite strand of DNM2 gene encodes miR-199a, which is similarly reduced in cancer cells under hypoxia. miR-199a targets the 3'-UTR of HIF-1α and HIF-2α. Decreased miR-199a expression in hypoxia increased HIF levels. Exogenous expression of miR-199a decreased HIF, cell migration, and metastasis of ovarian cancer cells. miR-199a-mediated changes in HIF levels affected expression of the matrix-remodeling enzyme, lysyloxidase (LOX). LOX levels negatively correlated with progression-free survival in ovarian cancer patients. These results demonstrate a regulatory relationship between DNM2, miR-199a, and HIF, with implications in cancer metastasis.