Impact of carriers in oral absorption: Permeation across Caco-2 cells for the organic anions estrone-3-sulfate and glipizide.

Carriers may mediate the permeation across enterocytes for drug substances being organic anions. Carrier mediated permeation for the organic anions estrone-3-sulfate (ES) and glipizide across Caco-2 cells were investigated kinetically, and interactions on involved carriers evaluated. Initial uptakes (P(UP)) at apical and basolateral membranes, apparent permeabilities (P(APP)) and corresponding intracellular end-point accumulations (P(EPA)) of radioactive labeled compounds were studied. Possible effects of other anionic compounds were investigated. Apical P(UP) and absorptive P(APP) for ES were inhibited and its absorptive P(EPA) prevented in presence of the investigated organic anions and apical P(UP) was saturable with K(m) 23microM. Basolateral P(UP) and exsorptive P(APP) were inhibited, its exsorptive P(EPA) was prevented, and basolateral P(UP) and exsorptive P(APP) were saturable with K(m) 44microM and 38microM, respectively. BCRP inhibition affected both absorptive an exsorptive P(EPA) and P(APP) for ES. Glipizide apical P(UP) and absorptive P(APP) were not inhibitable. Basolateral P(UP) for glipizide was inhibitable, its P(EPA) prevented, and P(UP) was saturable with K(m) 56microM, but exsorptive P(APP) was not affected. Carrier mediated exsorption kinetics for ES are seen at both apical and basolateral membranes, resulting in predominant exsorption despite presence of absorptive carrier(s). Carrier mediated basolateral P(UP) for glipizide was observed, but glipizide P(APP) was not described by carrier kinetics. However, glipizide is affecting exsorption for ES, due to interactions on basolateral carrier. The study confirms that estrone-3-sulfate can be used to characterize anionic carrier kinetics. Furthermore it is suggested that estrone-3-sulfate may be used to identify compounds which may interact on anionic carriers.

Impact of transporters in oral absorption: a case study of in vitro and in vivo organic anion absorption.

A key determinant for oral bioavailability of a drug candidate is the intestinal epithelial permeation of the drug candidate. This intestinal permeation may be affected by interactions on membrane transporters expressed in the intestinal epithelial cells. The purpose of the present study was to investigate whether transporters were involved in the intestinal absorption of an organic anion A275 and to compare the impact of interactions related to transporters in the Caco-2 cell model versus the in vivo rat model of intestinal absorption. In both models, it was investigated whether intestinal permeation of A275 was concentration dependent and affected by inhibitors or competitive organic anions. Interactions related to transporters in intestinal permeation was clearly demonstrated in the Caco-2 cell model but was not directly evident for in vivo rat absorption. However, an observed biphasic in vivo absorption and a large intervariability between rats might mask a dose-dependent absorption of A275. To avoid these suggested interactions, a dose of at least 10 mg/kg, which saturates the intestinal transporters involved in A275 absorption, should be administered, but at doses below that the risk of such drug interactions should be taken into account.

Evaluation of cytochrome P450 activity in vitro, using dermal and hepatic microsomes from four species and two keratinocyte cell lines in culture.

The Cytochrome P450 (CYP450) enzymes are expressed in the skin, and despite a low activity, as compared to the hepatic counterpart, a role during transdermal delivery of a drug cannot be excluded. Additionally, the enzymes may play a role in local toxicity, and further knowledge of dermal CYP450 activity can contribute to elucidate this issue. To achieve this, a cocktail of six selective CYP450 probe substrates were incubated with dermal and hepatic microsomes isolated from mouse, rat, minipig and man. Different species were used to evaluate if a reliable substitute for human tissue was possible. Further, the hepatic microsomes were included in this study, to estimate if the hepatic CYP450 activity is predictive of dermal CYP450 activity. The CYP450 activity was determined in two keratinocyte cell lines as well, as this in vitro model is desirable due to the ease in handling, among other factors. Overall, the metabolism found in the dermal microsomes was very low, and major differences were observed between species. When comparing the activities in dermal and hepatic microsomes, the qualitative pattern was to some extent similar within species, but also a number of differences were observed. The CYP450 metabolic activity in the two keratinocyte cell lines was not comparable to metabolism in the human dermal microsomes.