“ Sub-toxic doses of acetaminophen induce intestinal P-glycoprotein expression ans activity

GHANEM C., ARIAS A., VILLANUEVA S., NOVAK A., FILIA MF., DELLICARPINI G., MOTTINO A., RUBIO MC

Buenos Aires- Argentina

Simposio; International Symposium on drug transport and metabolism; 2009

IUPHAR-SAFE

Sub-toxic doses of acetaminophen induce intestinal P-glycoprotein expression and activity Carolina I. Ghanem1,2, Agostina Arias3, Silvina Villanueva3, Analía Novak2, María F. Filia1, Griselda Dellicarpini2, Aldo D. Mottino3 and Modesto C. Rubio1 1 Instituto de Investigaciones Farmacológicas. FFYB-CONICET-UBA. Argentina 2 Cátedra de Fisiopatología. FFYB-UBA. Argentina 3 Instituto de Fisiología Experimental, FBIOyF-CONICET-UNR. Argentina The well known analgesic-antipyretic acetaminophen (APAP) is one of the most sold over the counter drugs and is usually co-administered with other therapeutic agents. Previously, we have demonstrated that a toxic dose of APAP (1 g/kg i.p.) induced liver P-glycoprotein (P-gp) expression (1). Moreover, sub-toxic doses of the drug (0.2; 0.3 and 0.6 g/kg i.p., three consecutive days) produced a marked up-regulation of the basolateral transporter Mrp3 relative to apical Mrp2 in the same organ, affecting APAP pharmacokinetics (2) and toxicity (3). Our following aim was to evaluate the effect of sub-toxic doses of APAP on intestinal P-gp expression in rats and their possible interaction with other drugs that are substrates of this transporter. The sub-toxic treatment increased P-gp expression in duodenum and ileum in treated rats compared to controls (240 and 160%, respectively, P<0.05). The increased expression of P-gp in the treated group, with conservation of its physiological localization (apical lumen of the intestine), was confirmed by confocal immunofluorescence microscopy. To test if the described protein induction modifies the activity of P-gp in vitro, we evaluated the secretion of rhodamine 123 (R123), a P-gp substrate (4), using intestinal everted sacs in the presence and absence of verapamil (V), a known P-gp inhibitor. The cumulative R123 secretion was 44% higher in APAP-treated rats compared to controls (P<0.05). In the presence of verapamil, R123 secretion decreased P-gp activity in both groups compared to their own activity in the absence of the inhibitor (78 and 72% in APAP and control groups, respectively). We then studied if sub-toxic APAP treatment modified the in vivo absorption of digoxin, a typical substrate used to study P-gp activity (5). A dose of 0.2 mg/kg of digoxin, including traces of [3H] digoxin, was incorporated into the duodenum, and portal blood was sampled up to 30 min. At the end of the study, the portal digoxin concentration was 67% lower in the APAP-treated group compared to controls (expressed as % of the dose/ml; P<0.05). Finally, we studied the effect of 12 different concentrations of APAP (0 to 40 mM) on viability of LS174T cells, a human intestinal cell line, to select a sub-toxic dose. LS174T cells were exposed to the sub-toxic dose (5 mM) of APAP for 48 hs. Subsequently, P-gp expression and activity were evaluated. The cells treated with APAP expressed 60% more protein and showed 80% more activity than control cells. In conclusion, sub-toxic doses of APAP induced intestinal P-gp expression and activity in rats. This increased expression could modify the pharmacokinetics of other drugs, substrates of P-gp that are administered orally. Consistently, preliminary studies have shown induction of P-gp in a human intestinal cell line exposed to sub-toxic concentrations of APAP. (1)               Ghanem et. al. 2004. Biochem Pharmacol 68: 791-798. (2)               Ghanem et. al. 2005. JPET 315: 987–995. (3)               Ghanem et. al. 2009. Biochem Pharmacol 77: 1621–1628. (4)               Efferth  et. al. 1989. Anticancer Res 9: 1633-1637. (5)               Fromm et. al. 1999. Circulation 99: 552–557.