Investigation of the marine compound spongistatin 1 links the inhibition of PKCα translocation to nonmitotic effects of tubulin antagonism in angiogenesis

Abstract

The aims of the study were to meet the demand of new tubulin antagonists with fewer side effects by characterizing the antiangiogenic properties of the experimental compound spongistatin 1, and to elucidate nonmitotic mechanisms by which tubulin antagonists inhibit angiogenesis. Although tubulin-inhibiting drugs and their antiangiogenic properties have been investigated for a long time, surprisingly little is known about their underlying mechanisms of action. Antiangiogenic effects of spongistatin 1 were investigated in endothelial cells in vitro, including functional cell-based assays, live-cell imaging, and a kinome array, and in the mouse cornea pocket assay in vivo. Spongistatin 1 inhibited angiogenesis at nanomolar concentrations (IC(50): cytotoxicity>50 nM, proliferation 100 pM, migration 1.0 nM, tube formation 1.0 nM, chemotaxis 1.0 nM, aortic ring sprouting 500 pM, neovascularization in vivo 10 microg/kg). Further, a kinome array and validating data showed that spongistatin 1 inhibits the phosphorylation activity of protein kinase Calpha (PKCalpha), an essential kinase in angiogenesis, and its translocation to the membrane. Thus, we conclude that PKCalpha might be an important target for the antiangiogenic effects of tubulin antagonism. In addition, the data from the kinase array suggest that different tubulin antagonists might have individual intracellular actions.

Flavopiridol Protects Against Inflammation by Attenuating Leukocyte-Endothelial Interaction via Inhibition of Cyclin-Dependent Kinase 9

“…10 Briefly, HUVECs were treated either with tumor necrosis factor-␣ (TNF-␣; 10 ng/mL) for 15 minutes or with flavopiridol (100 nmol/L) for 30 minutes before TNF-␣. Native protein lysates were generated by the M-PER buffer (Pierce, Rockford, Ill).…”

Marine Sponge Derived Antiangiogenic Compounds

“…As in cancer cells, probably the most relevant effect of this compound on endothelial cells is the inhibition of cell proliferation, which is exerted at non-toxic concentrations. The antiangiogenic effect of spongistatin 1 seems to be due to nonmitotic effects and have been proposed to hint toward an as yet unknown mechanism of tubulin antagonism during angiogenesis: the inhibition of protein kinase Cα (PKCα) translocation [157].…”

Marine-Derived Angiogenesis Inhibitors for Cancer Therapy

“…Some inhibitors could prominently inhibit angiogenesis at much lower concentrations/doses than cytotoxic ones [21,22], suggesting the involvement of additional mechanism(s) apart from their microtubule inhibition. Actually, spongistatin 1 could inhibit the enzymatic activity and intracellular translocation of PKCα that is an essential kinase in angiogenesis [21].…”