Potent Inhibition of Macropinocytosis by Niclosamide in Cancer Cells: A Novel Mechanism for the Anticancer Efficacy for the Antihelminthic

SIMPLE SUMMARY: Niclosamide, a drug used to treat tapeworm infection, has been shown to have potential as an anticancer drug. The mechanism proposed for this action is the drug-induced inhibition of multiple signaling pathways. Here we uncover an additional, but novel and hitherto unknown, mechanism for the anticancer efficacy of this antihelminthic. Niclosamide forces cancer cells to nutrient starvation by blocking macropinocytosis, SLC38A5-mediated amino acid entry and H(+)-coupled nutrient transport pathways. Macropinocytosis is a process by which cells take up extracellular fluid along with the components present in the fluid, and this process is stimulated by an alkaline pH in the cytoplasm. SLC38A5 is an amino acid transporter that is stimulated by an outwardly directed H(+) gradient. The function of niclosamide as a H(+) channel is responsible for the newly discovered actions of this drug reported here. The drug also inhibits SLC38A5 by direct interaction with the transporter. ABSTRACT: Niclosamide, a drug used to treat tapeworm infection, possesses anticancer effects by interfering with multiple signaling pathways. Niclosamide also causes intracellular acidification. We have recently discovered that the amino acid transporter SLC38A5, an amino acid-dependent Na(+)/H(+) exchanger, activates macropinocytosis in cancer cells via amino acid-induced intracellular alkalinization. Therefore, we asked whether niclosamide will block basal and SLC38A5-mediated macropinocytosis via intracellular acidification. We monitored macropinocytosis in pancreatic and breast cancer cells using TMR-dextran and the function of SLC38A5 by measuring Li(+)-stimulated serine uptake. The peptide transporter activity was measured by the uptake of glycylsarcosine. Treatment of the cancer cells with niclosamide caused intracellular acidification. The drug blocked basal and serine-induced macropinocytosis with differential potency, with an EC(50) of ~5 μM for the former and ~0.4 μM for the latter. The increased potency for amino acid-mediated macropinocytosis is due to direct inhibition of SLC38A5 by niclosamide in addition to the ability of the drug to cause intracellular acidification. The drug also inhibited the activity of the H(+)-coupled peptide transporter. We conclude that niclosamide induces nutrient starvation in cancer cells by blocking macropinocytosis, SLC38A5 and the peptide transporter. These studies uncover novel, hitherto unknown, mechanisms for the anticancer efficacy of this antihelminthic.