Effects of benzyl-, phenethyl-, and alpha-naphthyl isothiocyanates on P-glycoprotein- and MRP1-mediated transport

Abstract

The objective of this investigation was to evaluate the effects of two dietary isothiocyanates (ITCs), benzyl- (BITC) and phenethyl isothiocyanate (PEITC), and one synthetic ITC, alpha-naphthyl isothiocyanate (1-NITC), on the P-glycoprotein (P-gp)- and multidrug-resistance protein 1 (MRP1)-mediated efflux of daunomycin (DNM), determine whether PEITC is a substrate of P-gp and/or MRP1, and elucidate the mechanism(s) involved in the inhibition of transport. BITC, PEITC, and 1-NITC significantly increased the 2-h accumulation of DNM in MCF-7/ADR (P-gp overexpression), PANC-1 (MRP1 overexpression), and human colon adenocarcinoma Caco-2 cells (except for 1-NITC). The accumulation of (14)C-PEITC was not changed in Caco-2, human breast cancer MDA435/LCC6 and MDA435/LCC6MDR1 (P-gp overexpression) cells in the absence and presence of the P-gp inhibitor verapamil, but significantly increased with the MRP inhibitor MK571 in PANC-1 cells. The isocyanate and amine metabolites had no effect on DNM accumulation in any cell line. After 2- and 24-h ITC treatments, cellular concentrations of glutathione (GSH) in PANC-1 and Caco-2 cells were depleted by BITC and PEITC, but not by 1-NITC; glutathione-S-transferase activity exhibited small changes. Our results suggest that (1) BITC, PEITC, and 1-NITC inhibit the P-gp- and MRP1-mediated efflux of DNM; (2) PEITC and/or its conjugates do not represent P-gp substrates; (3) BITC and PEITC, but not 1-NITC, inhibit MRP1 through the depletion of intracellular GSH, which acts as a cosubstrate for DNM efflux via MRP1; and (4) PEITC and/or its conjugates are MRP1 substrates so binding interactions with DNM represent a second potential mechanism involved in MRP1 inhibition.