Vincent J. Wacher

Differentiation of absorption and first‐pass gut and hepatic metabolism in humans: Studies with cyclosporine

The low and variable bioavailability of cyclosporine has been attributed to poor absorption. However, recent studies have suggested that intestinal first‐pass metabolism exerts a significant effect on bioavailability. We describe theory and methods to differentiate the contribution from oral absorption and intestinal and hepatic metabolism to overall cyclosporine bioavailability. Analysis of data from previous studies in our laboratories shows that in the absence of intestinal metabolism, cyclosporine absorption from its presently available dosage form averages at least 65% ± 12% in healthy volunteers and 77% ± 19% in kidney transplant patients. Analysis also suggests that the extraction ratio for cyclosporine in the gut is approximately twice the hepatic extraction and that cyclosporine absorption does not present a problem, with an average of 86% of the drug absorbed intact from its commercially available product in healthy volunteers. The boundary condition analysis described should have broad application in the differentiation of factors responsible for poor bioavailability.

Active secretion and enterocytic drug metabolism barriers to drug absorption

Intestinal phase I metabolism and active extrusion of absorbed drug have only recently been recognized as major determinants of oral drug bioavailability. Both CYP3A4, the major phase I drug metabolizing enzyme in humans, and the multidrug efflux pump, P-glycoprotein (P-gp), are present at high levels in the villus enterocytes of the small intestine, the primary site of absorption for orally administered drugs. Moreover, these proteins are induced by many of the same compounds and demonstrate a broad overlap in substrate and inhibitor specificities, suggesting that they act as a concerted barrier to drug absorption. Clinical studies have demonstrated that inhibition of CYP3A4-mediated intestinal metabolism can significantly improve the oral bioavailability of a wide range of drugs. Intestinal P-gp is a major route of elimination for both orally and intravenously administered anticancer drugs in animal models, and experiements with the Caco-2 cell line have provided strong evidence that inhibition of intestinal P-gp is another means by which oral drug bioavailability could be enhanced.

The effects of ketoconazole on the intestinal metabolism and bioavailability of cyclosporine.

The pharmacokinetics of cyclosporine were studied in the blood of five normal healthy volunteers (two men and three women) after each received oral and intravenous cyclosporine alone and with concomitant oral ketoconazole. Administration of ketoconazole caused a significant decrease in intravenous cyclosporine clearance (0.18 ± 0.05 L/kg/hr versus 0.32 ± 0.09 L/hr/kg) and a significant increase in cyclosporine oral bioavailability (56.4% ± 11.7% versus 22.4% ± 4.8%) compared with values before ketoconazole administration. Steady‐state volume of distribution for intravenously administered cyclosporine was unchanged (1.26 ± 0.44 L/kg versus 1.10 ± 0.27 L/kg). Hepatic bioavailability (1 — hepatic extraction ratio) calculated for intravenous cyclosporine increased by 11% in the presence of ketoconazole (86.3% ± 3.7% versus 75.2% ± 6.6% without ketoconazole), which accounts for only one third of the observed increase in cyclosporine oral bioavailability. Because it is unlikely that ketoconazole had a significant effect on either cyclosporine absorption or hepatic blood flow, the increase in cyclosporine bioavailability observed in this study is most likely explained by inhibition of gastrointestinal cytochrome P450 enzymes.

Intestinal MDR transport proteins and P-450 enzymes as barriers to oral drug delivery

Cytochrome P-450 3A4 (CYP3A4), the major phase I drug metabolizing enzyme in humans, and the multidrug efflux pump, MDR or P-glycoprotein (P-gp), are present at high levels in the villus tip enterocytes of the small intestine, the primary site of absorption for orally administered drugs. These proteins are induced or inhibited by many of the same compounds and demonstrate a broad overlap in substrate and inhibitor specificities, suggesting that they act as a concerted barrier to drug absorption. A series of studies from our laboratory of cyclosporine and tacrolimus in humans and a novel cysteine protease inhibitor in rats, dosed concomitantly with inhibitors and inducers of CYP3A4 and P-gp, suggest that gut extraction can be modeled using measures of intestinal metabolism and absorption rate, the latter reflecting changes in P-gp. Results evaluating a preliminary model applied to the CYP3A substrate drugs midazolam, indinavir, saquinavir, and rifabutin suggest that the model may be useful for predicting in vivo intestinal metabolism from in vitro data.

Grapefruit juice activates P-glycoprotein-mediated drug transport.

AbstractPurpose. Grapefruit juice (GJ) is known to increase the oral bioavailability of many CYP3A-substrates by inhibiting intestinal phase-I metabolism. However, the magnitude of AUC increase is often insignificant and highly variable. Since we earlier suggested that CYP3A and P-glycoprotein (P-gp) form a concerted barrier to drug absorption, we investigated the role of P-gp in GJ-drug interactions. Methods. The transcellular bidirectional flux of drugs that are (i) CYP3A-and/or P-gp substrates (Vinblastine, Cyclosporine, Digoxin, Fexofenadine, Losartan) or that are (ii) primary CYP3A-substrates (Felodipine, Nifedipine) was evaluated across MDCK-MDR1 cell monolayers with or without GJ, verifying monolayer integrity at all times. Results. While both apical-to-basal (A-B) and basal-to-apical (B-A) fluxes of all CYP3A/P-gp substrates tested were increased in the presence of GJ, the resulting net efflux (B-A/A-B) was in all cases significantly greater with GJ than control (Vin, 28.0 vs. 5.1; CsA, 9.9 vs. 2.8; Dig, 22. 9 vs. 14.7, Fex, 22.3 vs. 11.1, Los, 39.6 vs. 26). In contrast, no such GJ flux effect was observed with Pel and Nif, substrates of CYP3A only (2 vs. 1.7 and 1.2 vs. 1.3). Conclusions. GJ significantly activates P-gp-mediated efflux of drugs that are substrates of P-gp, potentially partially counteracting the CYP3A-inhibitory effects of GJ.

Active secretion and enterocytic drug metabolism barriers to drug absorption1PII of original article: S0169-409X(96)003304. The article was originally published in Advanced Drug Delivery Reviews 20 (1996) 99–112.1

Intestinal phase I metabolism and active extrusion of absorbed drug have only recently been recognized as major determinants of oral drug bioavailability. Both CYP3A4, the major phase I drug metabolizing enzyme in humans, and the multidrug efflux pump, P-glycoprotein (P-gp), are present at high levels in the villus enterocytes of the small intestine, the primary site of absorption for orally administered drugs. Moreover, these proteins are induced by many of the same compounds and demonstrate a broad overlap in substrate and inhibitor specificities, suggesting that they act as a concerted barrier to drug absorption. Clinical studies have demonstrated that inhibition of CYP3A4-mediated intestinal metabolism can significantly improve the oral bioavailability of a wide range of drugs. Intestinal P-gp is a major route of elimination for both orally and intravenously administered anticancer drugs in animal models, and experiments with the Caco-2 cell line have provided strong evidence that inhibition of intestinal P-gp is another means by which oral drug bioavailability could be enhanced.

Intestinal drug metabolism and antitransport processes: A potential paradigm shift in oral drug delivery

Poor oral bioavailability for many drugs is generally attributed to poor solubility in the gastrointestinal fluids, poor gut membrane permeability and/or extensive hepatic first-pass elimination. Recently, however, it has been recognized that cytochrome P-450 3A mediated drug metabolism in the intestine, and P-glycoprotein counter-transport processes may also contribute significantly to poor drug bioavailability. We have shown that cyclosporin, a highly lipid soluble, large molecular weight compound does not appear to have absorption problems, with approximately 86% of the present commercial formulation being absorbed intact in healthy volunteers. Rather, the low bioavailability of cyclosporin results from extensive metabolic extraction in the gut which approaches 60%. Recently, a strong overlap has been identified between substrates for gut metabolism by cytochrome P-450 3A and gut counter-transport by P-glycoprotein, suggesting that these processes may work together to limit the bioavailability of a large number of drug substances. Recognition of this potential for metabolism and counter-transport process in the gut leads to a new perspective on improving drug bioavailability that differs from the traditional physico-chemical approach.

Grapefruit juice exerts stimulatory effects on P-glycoprotein

Clinical Pharmacology & Therapeutics (1999) 65, 205–205; doi:

Pharmacokinetics of Cyclosporine Pre- and Post-Liver Transplantation

Cyclosporine (CyA) is an immunosuppressant metabolized primarily by the liver and small intestine. The pharmacokinetics (PK) of CyA were studied in 6 patients prior to and 1 to 3 months after liver transplantation (tx). Sixteen blood samples were collected over 24 hours following a 2–3 mg/kg intravenous dose of CyA. PK parameters, presented as mean ± SD, were estimated using noncompartmental techniques. Pre‐tx AUCs (14,540 ± 5200 μg•h/L) were found to be significantly higher than during the post‐tx phase (8120 ± 2870 μg•h/L, p = 0.04). CyA clearance values were lower pre‐tx as compared to post‐tx (0.21 ± 0.06 L/h/kg vs. 0.38 ± 0.14 L/h/kg, respectively). There was no change in volume of distribution. End‐stage liver disease can markedly decrease hepatic clearance of CyA relative to patients with stable hepatic function post—liver tx. The degree of impairment in clearance is not consistent or predictable based on liver function tests.