Characterization of P-glycoprotein Mediated Transport of K02, a Novel Vinylsulfone Peptidomimetic Cysteine Protease Inhibitor, Across MDR1-MDCK and Caco-2 Cell Monolayers

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Here we characterized the transport properties of morpholine-urea-phenylalanine- homophenylalanine-vinylsulfone-phenyl (K02), a newly developed peptidomimetic cysteine protease inhibitor, across monolayers of P-gp-expressed MDR1 transfected MDCK cells (MDR1-MDCK) and Caco-2 cells.


MDR1-MDCK, MDCK and Caco-2 cells, grown to confluence on Transwell insert membranes, were used to investigate transcellular transport of [14C]-K02.


The basolateral to apical (B-A) flux of 10 μM [I4C]-K02 across MDR1-MDCK cells was markedly greater than its apical to basolateral (A-B) flux (ratio = 39). This specific B-A transport was temperature dependent and saturable, with an apparent Michaelis-Menten constant and maximum velocity of 69.1 ± 19.5 μM and 148.9 ± 16.3 pmol/min/cm2, respectively. This B-A flux was significantly inhibited by cyclosporine (IC50 = 17.1 ± 0.7 μM), vinblastine (IC50 = 75.9 ± 13.0 μM) and verapamil (IC50 = 236 ± 63 μM). In Caco-2 cell monolayers, the B-A flux was reduced about 50% compared to that in MDR1-MDCK and the A-B flux was increased about 8-fold. The apparent Michaelis-Menten constant and maximum velocity values for the B-A transport were 71.8 ± 45.9 μM and 35.3 ± 9.0 pmol/min/cm2. This B-A flux was also significantly inhibited by P-gp substrates/ inhibitors. Western blots showed that the P-gp expression in MDR1-MDCK cells was about 10-fold that in Caco-2 cells.


K02 is transported by P-gp in both MDR1-MDCK and Caco-2 cells, and the in vitro interactions between K02 and various P-gp substrates may provide strategies to overcome the bioavailability barrier by intestinal P-gp.

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