Journal Article

A Novel MMP-2 Inhibitor 3-azidowithaferin A (3-azidoWA) Abrogates Cancer Cell Invasion and Angiogenesis by Modulating Extracellular Par-4

Khan, Sheema, Amin, Hina, Yousuf, Khalid, Mukherjee, Debaraj, Goswami, Anindya, Rah, Bilal, Jamwal, Gayatri

2012

Overview

Withaferin A, which is a naturally derived steroidal lactone, has been found to prevent angiogenesis and metastasis in diverse tumor models. It has also been recognized by different groups for prominent anti-carcinogenic roles. However, in spite of these studies on withanolides, their detailed anti-metastatic mechanism of action remained unknown. The current study has poised to address the machinery involved in invasion regulation by stable derivative of Withaferin A, 3-azido Withaferin A (3-azidoWA) in human cervical HeLa and prostate PC-3 cells. Sub-toxic concentration of 3-azidowithaferin A (3-azido WA) inhibited cancer cell motility and invasion in wound healing and Boyden chamber invasion by suppressing MMP-2 activity in gelatin zymography and its expression has proved to be a major obstacle in chemo-sensitivity. We have uncovered a novel mechanism of 3-azidoWA induced extracellular pro-apoptotic candidate tumor suppressor Par-4 protein stimulation in conditioned media and also noticed a concomitant marked reduction in pAkt and pERK signaling by immunoblot analysis. Furthermore, our zymography results suggest 3-azidoWA induced MMP-2 inhibition was mediated through secretory Par-4. The inhibition of apoptosis by 3-azidoWA could not restore MMP-2 gelatinase activity. In addition to this, our in vivo animal experiments data showed 3-azidoWA abrogated neovascularisation in dose dependent manner in mouse Matrigel plug assay. For this report, we found that 3-azidoWA suppressed motility and invasion of HeLa and PC-3 cells in MMP-2 dependent manner. Our in vitro result strongly suggests that sub-toxic doses of 3-azidoWA enhanced the secretion of extracellular Par-4 that abolished secretory MMP-2 expression and activity. Depletion of secretory Par-4 restored MMP-2 expression and invasion capability of HeLa and PC-3 cells. Further, our findings implied that 3-azidoWA attenuated internal phospho-ERK and phospho-Akt expression in a dose dependent manner might play a key role in inhibition of mouse angiogenesis by 3-azidoWA.

• Publisher: Public Library of Science,Public Library of Science (PLoS)
• Subject: AKT protein / Analysis / Angiogenesis / Animal experimentation / Animal models / Animal research / Animals / Antineoplastic Agents - chemical synthesis / Antineoplastic Agents - pharmacology / Apoptosis / Apoptosis - drug effects / Azides - chemical synthesis / Azides - pharmacology / Biocompatibility / Biological Assay / Biology / Boyden chamber / Cancer / Cancer metastasis / Caprolactone / Carcinogens / Cell culture / Cell Line, Tumor / Cell Movement - drug effects / Cervix / Chemotherapy / Clinical trials / Collagen / Conditioning / Drug Combinations / Extracellular Signal-Regulated MAP Kinases - genetics / Extracellular Signal-Regulated MAP Kinases - metabolism / Gelatin / Gelatinase / Gelatinase A / Gene Expression Regulation, Neoplastic / Humans / Inhibition / Integrative medicine / Kinases / Laboratories / Laminin / Machinery and equipment / Matrix Metalloproteinase 2 - genetics / Matrix Metalloproteinase 2 - metabolism / Matrix Metalloproteinase Inhibitors - chemical synthesis / Matrix Metalloproteinase Inhibitors - pharmacology / Medicine / Metastases / Metastasis / Mice / Mice, Inbred C57BL / Motility / Natural products / Neoplasm Invasiveness - prevention & control / Neovascularization / Neovascularization, Pathologic - prevention & control / Penicillin / Pharmacology / Phosphorylation / Prostate apoptosis response 4 protein / Prostate cancer / Proteoglycans / Proto-Oncogene Proteins c-akt - genetics / Proto-Oncogene Proteins c-akt - metabolism / Receptors, Thrombin - antagonists & inhibitors / Receptors, Thrombin - genetics / Receptors, Thrombin - metabolism / Signal Transduction / Signaling / Tumor suppressor genes / Withanolides - chemical synthesis / Withanolides - pharmacology / Wound healing