Abstract

Hypoxia-inducible factor 1 (HIF-1) plays a critical role for tumor adaptation to hypoxia and promotes angiogenesis. Antitumor imidazoacridinone C-1311 is a DNA reactive topoisomerase II and FLT3 receptor tyrosine kinase inhibitor selected for phase II clinical trials for breast cancer. Here, we demonstrate the new mechanism of C-1311 action involving HIF-1a, vascular endothelial growth factor (VEGF) and angiogenesis as additional targets. In a cell-free system, C-1311 prevented HIF-1a binding to oligonucleotide encompassing a canonical hypoxia-responsive element (HRE), but did not disrupt the formation of protein-DNA complex through direct interference with HIF-1a. In human colon cancer HCT116 cells, C-1311 was found to down-regulate HIF-1a mRNA and protein level, which prevented hypoxic activation of VEGF gene, resulting in decreased VEGF protein expression. Importantly, C-1311 blocked hypoxia-induced VEGF protein production irrespective of the p53 tumor suppressor status (wt versus lack of p53). To a lesser extent, C-1311 also reduced basal VEGF mRNA and protein level under normoxia. In murine melanoma B16/F10 cells,C-1311 only slightly inhibited HIF-1a mRNA accumulation but vitally decreased VEGF mRNA and protein expression. Consistent with HIF-1a and VEGF inhibition/depletion observed in vitro, C-1311 significantly affected angiogenesis in vivo. C-1311 at a single dose (40 mg/kg), inhibited bFGF stimulated angiogenesis by 70%, in terms of vascularization of Matrigel plugs in mice. Compared with single administration, continuous low C-1311 dosing (8 mg/kg/day for 5 days), gave more profound angiogenesis inhibition (80%), suggesting that this effect is schedule-dependent. Antiangiogenic activity of C-1311 was further confirmed in experiments, where angiogenesis was stimulated by murine melanoma B16/F10 cells encapsulated in alginate beads and implanted into mice. C-1311 at 8 mg/kg, when given for 9 days, completely abrogated vascularization of alginate implants. Even at lower doses of 4-6 mg/kg, C-1311 decreased implants' angiogenesis by 40-60%, respectively, indicating that its inhibitory action on tumor angiogenesis is dose-dependent. In summary, this study provides the first evidence, that antitumor effect of C-1311 involve both interference with HIF-1a/HRE transactivating activity and downregulation of HIF-1a mRNA and protein accumulation, resulting in decreased hypoxia-induced VEGF expression. Importantly, C-1311-driven VEGF depletion leads to a significant reduction of in vivo angiogenesis.

Authors (5)