Membrane permeability and P-glycoprotein (Pgp) can be limiting factors for blood-brain barrier penetration. The objectives of this study were to determine whether there are differences in the in vitro permeability, Pgp substrate profiles, and physicochemical properties of drugs for central nervous system (CNS) and non-CNS indications, and whether these differences are useful criteria in selecting compounds for drug development. Apparent permeability (P app ) and Pgp substrate profiles for 93 CNS (n ϭ 48) and non-CNS (n ϭ 45) drugs were determined by monolayer efflux. Calcein-AM inhibition assays were used to supplement the efflux results. The CNS set (2 of 48, 4.2%) had a 7-fold lower incidence of passive permeability values Ͻ150 nm/s compared with the non-CNS set (13 of 45, 28.9%). The majority of drugs (72.0%, 67 of 93) were not Pgp substrates; however, 49.5% (46 of 93) were positive in the calcein-AM assay when tested at 100 M. The CNS drug set (n ϭ 7 of 48, 14.6%) had a 3-fold lower incidence of Pgp-mediated efflux than the non-CNS drug set (n ϭ 19 of 45, 42.2%). Analysis of 18 physicochemical properties revealed that the CNS drug set had fewer hydrogen bond donors, fewer positive charges, greater lipophilicity, lower polar surface area, and reduced flexibility compared with the non-CNS group (p Ͻ 0.05), properties that enhance membrane permeability. This study on a large, diverse set of marketed compounds clearly demonstrates that permeability, Pgp-mediated efflux, and certain physicochemical properties are factors that differentiate CNS and non-CNS drugs. For CNS delivery, a drug should ideally have an in vitro passive permeability Ͼ150 nm/s and not be a good (B 3 A/A 3 B ratio Ͻ2.5) Pgp substrate.The delivery of a new drug candidate to the central nervous system (CNS) can be a significant challenge during drug development. Often, the CNS distribution of a drug is poor because of exclusion at the blood-brain barrier (BBB) (Abbott and Romero, 1996;Pardridge, 1997). The BBB is composed of a single layer of endothelial cells connected by tight junctions. Brain microvascular endothelial cells lack fenestrations, have few pinocytotic vesicles, and express a variety of metabolic enzymes and membrane efflux transporters, such as P-glycoprotein (Pgp) (Rubin and Staddon, 1999; Kusuhara and Sugiyama, 2001a,b). These features make the BBB a formidable barrier that drugs must overcome to reach the brain parenchyma.Early assessment of the ability of a drug candidate to penetrate the CNS is critical during the drug discovery selection process, especially for therapeutic indications that require delivery to a CNS site of action. Equally important is the ability to design drugs for non-CNS indications that have minimal brain penetration to avoid undesirable CNS side effects. Over the past several years, academia and industry have invested significant effort in the development and implementation of lead optimization screens, including in vitro assays and computational models to evaluate CNS penetration.A number o...

ABSTRACT:Because modulation of P-glycoprotein (Pgp) through inhibition or induction can lead to drug-drug interactions by altering intestinal, central nervous system, renal, or biliary efflux, it is anticipated that information regarding the potential interaction of drug candidates with Pgp will be a future regulatory expectation. Therefore, to be able to utilize in vitro Pgp inhibition findings to guide clinical drug interaction studies, the utility of five probe substrates (calcein-AM, colchicine, digoxin, prazosin, and vinblastine) was evaluated by inhibiting their Pgp-mediated transport across multidrug resistance-1-transfected Madin-Darby canine kidney cell type II monolayers with 20 diverse drugs having various degrees of Pgp interaction (e.g., efflux ratio, ATPase, and calcein-AM inhibition). Overall, the rank order of inhibition was generally similar with IC 50 values typically within 3-to 5-fold of each other. However, several notable differences in the IC 50 values were observed. Digoxin and prazosin were the most sensitive probes (e.g., lowest IC 50 values), followed by colchicine, vinblastine, and calcein-AM. Inclusion of other considerations such as a large dynamic range, commercially available radiolabel, and a clinically meaningful probe makes digoxin an attractive probe substrate. Therefore, it is recommended that digoxin be considered as the standard in vitro probe to investigate the inhibition profiles of new drug candidates. Furthermore, this study shows that it may not be necessary to generate IC 50 values with multiple probe substrates for Pgp as is currently done for cytochrome P450 3A4. Finally, a strategy integrating results from in vitro assays (efflux, inhibition, and ATPase) is provided to further guide clinical interaction studies.P-glycoprotein (Pgp) is a member of the ATP-binding cassette superfamily of transport proteins and is expressed in numerous tissues such as the luminal membrane of the small intestine and blood-brain barrier, and the apical membranes of excretory organs such as liver and kidney (Ayrton and Morgan, 2001). Pgp has broad substrate recognition, which can affect the pharmacokinetics, efficacy, safety, and target organ specificity of drugs. As a result, drug-drug interactions resulting from inhibition or induction of Pgp are a recognized clinical concern (Englund et al., 2004;Balayssac et al., 2005) recently highlighted in the Food and Drug Administration (FDA) concept paper "Drug Interaction Studies-Study Design, Data Analysis, and Implications for Dosing and Labeling" (FDA, 2004).Despite many years of investigation, considerable uncertainty remains about the number of drug binding sites within Pgp and their mutual relationships. It is postulated that the transmembrane regions of Pgp form a large binding pocket (Sharom et al., 1998;Lugo and Sharom, 2005) composed of amino acid residues from multiple transmembrane segments Clarke, 2001, 2002). Recent experiments investigating drug binding (Martin et al., 2000), fluorescent dye uptake (Shapiro and Ling, 1997;Lugo ...

Dolutegravir (DTG; S/GSK1349572) is a potent HIV-1 integrase inhibitor with a distinct resistance profile and a once-daily dose regimen that does not require pharmacokinetic boosting. This work investigated the in vitro drug transport and metabolism of DTG and assessed the potential for clinical drug-drug interactions. DTG is a substrate for the efflux transporters P-glycoprotein (Pgp) and human breast cancer resistance protein (BCRP). Its high intrinsic membrane permeability limits the impact these transporters have on DTG's intestinal absorption. UDP-glucuronosyltransferase (UGT) 1A1 is the main enzyme responsible for the metabolism of DTG in vivo, with cytochrome P450 (P450) 3A4 being a notable pathway and UGT1A3 and UGT1A9 being only minor pathways. DTG demonstrated little or no inhibition (IC 50 values > 30 mM) in vitro of the transporters Pgp, BCRP, multidrug resistance protein 2, organic anion transporting polypeptide 1B1/3, organic cation transporter (OCT) 1, or the drug metabolizing enzymes CYP1A2, 2A6, 2B6, 2C8, 2C9, 2C19, 2D6, 3A4, UGT1A1, or 2B7. Further, DTG did not induce CYP1A2, 2B6, or 3A4 mRNA in vitro using human hepatocytes. DTG does inhibit the renal OCT2 (IC 50 = 1.9 mM) transporter, which provides a mechanistic basis for the mild increases in serum creatinine observed in clinical studies. These in vitro studies demonstrate a low propensity for DTG to be a perpetrator of clinical drug interactions and provide a basis for predicting when other drugs could result in a drug interaction with DTG.

Drug transporters can have a significant impact on the absorption, distribution, metabolism, excretion, and toxicity of a drug (Borst and Oude Elferink, 2002). Transporters can be effective barriers to drug exposure, be the rate determining step in the uptake and/or excretion of a compound or metabolite, and be a cause of drug-drug interactions. Drug efflux transport systems are the most extensively studied family of drug transporters owing to the phenomenon of multidrug resistance, in which cancer cells become cross-resistant to multiple cytotoxic anticancer agents after treatment with only a single drug (Borst and Oude Elferink, 2002;Lepper et al., 2005). Drugs that are substrates for efflux transporters are pumped out of the cell to result in decreased intracellular accumulation, permitting cancer cells to survive lethal doses of cytotoxic drugs. This phenomenon has been characterized in a number of cancers including breast cancer.Breast cancer is the most prevalent cancer in the world, affecting more than 4 million women (Parkin et al., 2005). Each year, breast cancer is diagnosed in more than 1 million women, and 450,000 die of this disease. Breast cancer is frequently associated with increased expression and activation of the epidermal growth factor receptor (EGFR; also known as ErbB), a family of transmembrane tyrosine kinase receptors (Rowinsky, 2004;. These Article, publication date, and citation information can be found at