ABSTRACT:Thirty-two structurally diverse drugs used for the treatment of various conditions of the central nervous system (CNS), along with two active metabolites, and eight non-CNS drugs were measured in brain, plasma, and cerebrospinal fluid in the P-glycoprotein (P-gp) knockout mouse model after subcutaneous administration, and the data were compared with corresponding data obtained in wild-type mice. Total brain-to-plasma (B/P) ratios for the CNS agents ranged from 0.060 to 24. Of the 34 CNS-active agents, only 7 demonstrated B/P area under the plasma concentration curve ratios between P-gp knockout and wild-type mice that did not differ significantly from unity. Most of the remaining drugs demonstrated 1.1-to 2.6-fold greater B/P ratios in P-gp knockout mice versus wild-type mice. Three, risperidone, its active metabolite 9-hydroxyrisperidone, and metoclopramide, showed marked differences in B/P ratios between knockout and wild-type mice (6.6-to 17-fold). Differences in B/P ratios and cerebrospinal fluid/ plasma ratios between wild-type and knockout animals were correlated. Through the use of this model, it appears that most CNSactive agents demonstrate at least some P-gp-mediated transport that can affect brain concentrations. However, the impact for the majority of agents is probably minor. The example of risperidone illustrates that even good P-gp substrates can still be clinically useful CNS-active agents. However, for such agents, unbound plasma concentrations may need to be greater than values projected using receptor affinity data to achieve adequate receptor occupancy for effect.Active transport mechanisms as determinants of drug absorption, distribution, and clearance have been the focus of considerable research effort over the past decade. Of the numerous transporter proteins recently investigated, the one for which the greatest amount of knowledge exists is P-glycoprotein (MDR1). Originally described as a transporter involved in imparting drug resistance to tumor cells, P-glycoprotein has been demonstrated to be important in reducing absorption of drugs from the intestinal lumen, in active secretion of drugs into urine and bile, and in extrusion of drugs from vital organs such as the brain and reproductive tissues (Troutman et al., 2002). As such, P-glycoprotein-mediated transport has become an important issue in the discovery and development of new drugs. For example, new compounds that are promising with regard to target receptor/ enzyme activity can be severely hampered in their ability to elicit pharmacological effects in vivo should they be good substrates for P-glycoprotein, especially if the route of administration is intended to be oral or the target tissues is one rich in P-glycoprotein activity. Furthermore, the potential for drug-drug interactions arises in the event that the P-glycoprotein substrate is coadministered with another agent that can inhibit P-glycoprotein.Several models have been developed to assess drugs as P-glycoprotein substrates. In vitro models have included the Caco...

The ATP-driven drug export pump, P-glycoprotein, is a primary gatekeeper of the blood-brain barrier and a major impediment to central nervous system (CNS) pharmacotherapy. Reducing P-glycoprotein activity dramatically increases penetration of many therapeutic drugs into the CNS. Previous studies in rat showed that brain capillary P-glycoprotein was transcriptionally up-regulated by the pregnane X receptor (PXR), a xenobioticactivated nuclear receptor. Here we used a transgenic mouse expressing human PXR (hPXR) to determine the consequences of increased blood-brain barrier P-glycoprotein activity. P-glycoprotein expression and transport activity in brain capillaries from transgenic mice was significantly increased when capillaries were exposed to the hPXR ligands, rifampin and hyperforin, in vitro and when the mice were dosed with rifampin in vivo. Plasma rifampin levels in induced mice were comparable with literature values for patients. We also administered methadone, a CNS-acting, P-glycoprotein substrate, to control and rifampin-induced transgenic mice and measured the drug's antinociceptive effect. In rifampin-induced mice, the methadone effect was reduced by approximately 70%, even though plasma methadone levels were similar to those found in transgenic controls not exposed to rifampin. Thus, hPXR activation in vivo increased P-glycoprotein activity and tightened the blood-brain barrier to methadone, reducing the drug's CNS efficacy. This is the first demonstration of the ability of bloodbrain barrier PXR to alter the efficacy of a CNS-acting drug.

This study was conducted to assess the utility of unbound brain EC 50 (EC 50,u ) as a measure of in vivo potency for centrally active drugs. Seven -opioid agonists (alfentanil, fentanyl, loperamide, methadone, meperidine, morphine, and sufentanil) were selected as model central nervous system drugs because they elicit a readily measurable central effect (antinociception) and their clinical pharmacokinetics/pharmacodynamics are well understood. Mice received an equipotent subcutaneous dose of one of the model opioids. The time course of antinociception and the serum and brain concentrations were determined. A pharmacokinetic/pharmacodynamic model was used to estimate relevant parameters. In vitro measures of opioid binding affinity (K i ) and functional activity [EC 50 for agonist stimulated guanosine 5Ј-O-(3-[35 S]thio)triphosphate binding] and relevant clinical parameters were obtained to construct in vitro-to-preclinical and preclinical-to-clinical correlations. The strongest in vitro-to-in vivo correlation was observed between K i and unbound brain EC 50,u (r 2 ϳ 0.8). A strong correlation between mouse serum and human plasma EC 50 was observed (r 2 ϭ 0.949); the correlation was improved when corrected for protein binding (r 2 ϭ 0.995). Clinical equipotent i.v. dose was only moderately related to K i . However, estimates of ED 50 and EC 50 (total serum, unbound serum, total brain, and unbound brain) were significant predictors of clinical equipotent i.v. dose; the best correlation was observed for brain EC 50,u (r 2 ϭ 0.982). For each opioid, brain equilibration half-life in mice was almost identical to the plasma effect-site equilibration half-life measured clinically. These results indicate that the mouse is a good model for opioid human brain disposition and clinical pharmacology and that superior in vitro-to-preclinical and preclinicalto-clinical correlations can be achieved with relevant unbound concentrations.In drug discovery, in vitro assays and preclinical animal studies are widely used to assess compound potency and to identify compound(s) that may have a desirable clinical response. Several options for assessing compound potency are available, including in vitro (receptor binding or functional assays) and in vivo (animal studies to determine dose-response or concentration-response relationships) protocols. In vitro binding and functional assays, by nature, are designed to estimate the intrinsic affinity or potency at the receptor of interest, whereas in vivo experiments take into account the full spectrum of pharmacokinetic and pharmacodynamic processes that ultimately determine biological response.Ideally, in vitro potency would translate to or predict in vivo potency. Often, this is the case, and significant correlations between in vivo ED 50 or EC 50 and in vitro potency have been established for a variety of therapeutic targets (Leysen et al., 1983;Visser et al., 2003). However, when there is no correlation between in vivo and in vitro potency measures, the validity of the in vitro as...

ABSTRACT:Previous studies have indicated that P-glycoprotein (P-gp) attenuates the central nervous system penetration and central activity of some opioids. The impact of P-gp-mediated efflux on the disposition and efficacy of the synthetic opioid alfentanil currently is unknown. In this study, P-gp-competent [mdr1a(؉/؉)] and P-gpdeficient [mdr1a(؊/؊)] mice were used to investigate the impact of P-gp-mediated efflux on the systemic pharmacokinetics, brain disposition, and central activity of alfentanil. Equipotent doses of alfentanil were administered to mdr1a(؉/؉) and mdr1a(؊/؊) mice (0.2 and 0.067 mg/kg, respectively), and the time course of brain and serum concentrations as well as antinociception were determined. A pharmacokinetic-pharmacodynamic (PK-PD) model was fit to the data and used to assess the impact of P-gp on parameters associated with alfentanil disposition and action. The mdr1a(؉/؉) mice were less sensitive to alfentanil than mdr1a(؊/؊) mice, requiring a 3-fold higher dose to produce similar antinociception. PK-PD modeling revealed no differences in alfentanil systemic pharmacokinetics between P-gp expressers and nonexpressers. However, the steady-state brain-to-serum concentration ratio (K p,brain,ss ) was ϳ3-fold lower in mdr1a(؉/؉) mice compared with mdr1a(؊/؊) mice (0.19 ؎ 0.01 versus 0.54 ؎ 0.04, respectively). Consistent with the ϳ3-fold lower K p,brain,ss , the antinociception versus serum concentration relationship in mdr1a(؉/؉) mice was shifted ϳ3-fold rightward compared with mdr1a(؊/؊) mice. However, there was no difference in the antinociception versus brain concentration relationship, or in the brain tissue EC 50 (11 ؎ 1.8 versus 9.2 ؎ 1.7 ng/g), between mdr1a(؉/؉) and mdr1a(؊/؊) mice. These results indicate that alfentanil is an in vivo P-gp substrate and are consistent with the hypothesis that P-gp-mediated efflux attenuates antinociception by reducing alfentanil K p,brain,ss .