Apparent receptor-mediated activation of Ca2+-dependent conductive Cl−transport by shark-derived polyaminosterols

Abstract

The shark liver antimicrobial polyaminosterol squalamine is an angiogenesis inhibitor under clinical investigation as an anti-cancer agent and as a treatment for the choroidal neovascularization associated with macular degeneration of the retina. The related polyaminosterol MSI-1436 is an appetite suppressant that decreases systemic insulin resistance. However, the mechanisms of action of these polyaminosterols are unknown. We report effects of MSI-1436 on Xenopus oocytes consistent with the existence of a receptor for polyaminosterols. MSI-1436 activates bidirectional, trans-chloride-independent Cl-flux in Xenopus oocytes. At least part of this DIDS-sensitive Cl−flux is conductive, as measured using two-electrode voltage-clamp and on-cell patch-clamp techniques. MSI-1436 also elevates cytosolic Ca2+concentration ([Ca2+]) and increases bidirectional45Ca2+flux. Activation of Cl−flux and elevation of cytosolic [Ca2+] by MSI-1436 both are accelerated by lowering bath Ca2+and are not acutely inhibited by extracellular EGTA. Elevation of cytosolic [Ca2+] by MSI-1436 requires heparin-sensitive intracellular Ca2+stores. Although injected EGTA abolishes the increased conductive Cl−flux, that Cl−flux is not dependent on heparin-sensitive stores. In low-bath Ca2+conditions, several structurally related polyaminosterols act as strong agonists or weak agonists of conductive Cl−flux in oocytes. Weak agonist polyaminosterols antagonize the strong agonist, MSI-1436, but upon addition of the conductive Cl−transport inhibitor DIDS, they are converted into strong agonists. Together, these properties operationally define a polyaminosterol receptor at or near the surface of the Xenopus oocyte, provide an initial description of receptor signaling, and suggest routes toward further understanding of a novel class of appetite suppressants and angiogenesis inhibitors.