M410, A Combretastatin A4 Analogue, Disrupts Microtubules And Inhibits Hif-1 Alpha In Human Breast Cancer Cells

Hypoxia-inducible factor-1 (HIF-1) is a primary transcriptional factor that targets a series of genes participating in angiogenesis and cell proliferation. HIF-1 is a heterodimer consisting of a constitutively-expressed HIF-1 beta subunit and an oxygen-regulated HIF-1 alpha subunit. Overexpression of HIF-1 alpha has been found in various types of cancer. Targeting HIF-1 alpha may be a novel approach to cancer therapy. Previous findings showed that a newly synthesized compound (Z)-3,4',5-trimethoxylstilbene-3'-0-phosphate disodium (M410), an analogue of the microtubule-targeting agent, combretastatin A4, inhibited the polymerization of bovine brain tubulin and induced mitotic arrest. The aim of the present study was to determine the mechanism of M410 destabilizes microtubules and inhibits HIF-1 alpha in breast cancer cells. We performed 3-(4,5-dimethylthiazol-2-yl)-2,5-diphenyltetrazolium bromide (MTT) assay, immunofluorescence and confocal microscopy, ELISA assay, transient transfections and reporter gene assay, immunoblot analysis and isolation and analysis of RNA to evaluate the mechanisms of M410 on breast cancer. SPSS 17.0 was used to analyze the data. The results showed that the growth of breast cancer cells was inhibited in a dose-dependent manner. MDA-MB-231 was the most sensitive, with a 50% growth inhibition (GI50) of 111.4 +/- 2.2 nM. HIF-1 alpha expression was clearly reduced following M410 treatment in a dose-dependent manner. M410 downregulated the nuclear accumulation of HIF-1 alpha, and the strong correlation between disruption of the microtubule cytoskeleton and the inhibition of HIF-1 alpha expression was independent of mitotic arrest. Furthermore, M410 inhibited HIF-1 alpha at the post-transcriptional level and inhibited the vascular endothelial growth factor (VEGF) at the transcription level. M410 downregulated HIF-1 alpha expression in a proteasome-independent manner. In conclusion, M410 depolymerized microtubules and downregulated HIF-1 alpha protein levels in a proteasome-independent manner and reduced the mRNA of HIF-1-targeted genes in the MDA-MB-231 breast cancer cell line.