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...