RT Journal Article SR Electronic T1 Development of a novel system for estimating human intestinal absorption using Caco-2 cells in the absence of esterase activity JF Drug Metabolism and Disposition JO Drug Metab Dispos FD American Society for Pharmacology and Experimental Therapeutics SP dmd.109.029413 DO 10.1124/dmd.109.029413 A1 Kayoko Ohura A1 Hisae Sakamoto A1 Shin-ichi Ninomiya A1 Teruko Imai YR 2009 UL http://dmd.aspetjournals.org/content/early/2009/11/18/dmd.109.029413.abstract AB Both mRNA and protein levels of the carboxylesterase (CES) isozymes, hCE1 and hCE2, in Caco-2 cells increase in a time-dependent manner, but hCE1 levels are always higher than those of hCE2. In human small intestine, however, the picture is reversed, with hCE2 the predominant isozyme. Drugs hydrolyzed by hCE1 not by hCE2 can be hydrolyzed in Caco-2 cells, but barely hydrolyzed in human small intestine. The results in Caco-2 cells can be misleading as a predictor of what will happen in human small intestine. In the present study, we proposed a novel method for predicting the absorption of prodrugs in the absence of CES-mediated hydrolysis in Caco-2 cells. The specific inhibition against CES was achieved using bis-p-nitrophenyl phosphate (BNPP). The optimal concentration of BNPP was determined at 200 µM by measuring the transport and hydrolysis of O-butyryl-propranolol (butyryl-PL) as a probe. BNPP concentrations of more than 200 µM inhibited 86% of hydrolysis of butyryl-PL, resulted in increase of its apparent permeability. Treatment with 200 µM BNPP did not affect paracellular transport, passive diffusion, or carrier-mediated transport. Furthermore, the proposed evaluation system was tested for ethyl fexofenadine (ethyl-FXD), which is a superior substrate for hCE1 but a poor one for hCE2. CES-mediated hydrolysis of ethyl-FXD was 94% inhibited by 200 µM BNPP, and ethyl-FXD was passively transported as an intact prodrug. From above observation, the novel evaluation system is effective for the prediction of human intestinal absorption of ester-type prodrugs.The American Society for Pharmacology and Experimental Therapeutics