The Blood-Brain Barrier and CNS Penetration: a Drug Discovery Point of View
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Bayer Schering Pharma, Research Pharmacokinetics, Germany
The examination and prediction of brain penetration remains a key challenge in the drug discovery and development of both CNS and non-CNS drugs.
Traditionally, the focus of the pharmacokinetic (PK) support in CNS drug discovery has been on increasing the blood-brain barrier (BBB) permeability in vitro and on elevating the brain/plasma ratio (Kp) in vivo. Unfortunately, however, neither of these parameters can be reliably linked to pharmacodynamic (PD) and efficacy readouts. While increasing BBB permeability may shorten the onset of drug action, an increase in the total amount in brain may not necessarily increase the relevant drug concentration at the pharmacological target site. Since the traditional Kp ratio is based on a crude homogenisation of brain tissue, it ignores the compartmentalisation of the brain and an increase favours non-specific binding to brain lipids rather than free drug levels which tend to correlate best with drug effects.
Successful drug discovery therefore depends on optimizing both the rate and the extent of CNS penetration in vivo (Fig 1.). A high rate of penetration results from high permeability as well as low binding to brain tissue. Measurement of whole-brain drug concentrations alone can be misleading and the extent of brain penetration is best assessed by determining the unbound drug concentration in the brain (i.e. the extracellular fluid levels) in relation to the free plasma concentration.
As the complex nature of the brain requires different compartments to be considered when trying to understand and improve new compounds, several complementary parameters need to be measured in vitro and in vivo and integrated into a coherent model of brain penetration and distribution. Since no one parameter alone can explain and predict CNS penetration, a new concept is currently emerging which acknowledges this complexity by integrating several aspects determining CNS penetration such as BBB permeability, unbound drug concentration in brain and intra-brain distribution.
1) BBB permeability and fraction unbound in vitro(Fig. 2 left). Despite three decades of experiences in culturing brain endothelial cells, there is still no generally satisfying in vitro model of the BBB available and most CNS discovery groups in industry use MDCK cells to screen their compounds for BBB permeability. Although MDCK cells are neither endothelial cells nor do they originate from brain, the high tightness of the monolayer produces permeability values which correlate well with in vivo brain permeation, especially when transfected with the MDR1 efflux pump which is highly active at the BBB in vivo. In addition to the permeability measurement, a so-called efflux ratio (ratio of the permeability of both directions) is determined allowing to identify and reject compounds with strong recognition by the MDR1 P-glycoprotein drug efflux pump.
The free fraction in brain can be studied in vitro by equilibrium dialysis of the compound between buffer and brain homogenate. The method is simple and can be performed in parallel with the estimation of the free fraction in plasma and is thus able to rapidly generate data on fu,brain and fu,plasma for a large number of compounds. As brain interstitial fluid (ISF) contains only a very low amount of protein, fu,brain can be used directly to estimate unbound levels of a compound in brain ISF. It is important to note that the free fraction in plasma cannot be taken as surrogate for the free fraction in brain since the lipid and protein composition of plasma and brain differ significantly, with plasma having twice as much protein while brain has 20-times more lipids. The implications on CNS penetration of the fraction unbound in blood and the fraction unbound in brain are illustrated in Fig. 3.
2) In vivo PK studies (Fig. 2 right). Once a compound has demonstrated favourable in vitro data, in vivo studies will follow to examine the behaviour of the compound in the whole animal. In order to be able to link the results with animal efficacy studies, the same species/strain should be taken. Besides the regular single dose PK study which determines the disposition PK and the oral PK of the compound (e.g. total blood clearance, volume of distribution at steady-state, AUC, half-life and oral bioavailability), the ‘classic’ total brain concentration vs. total plasma concentration ration (Kp) is determined either at several time points after ip, sc or po dosing, or after iv infusion at a single time point where near steady-state is assumed.
3) Integration and interpretation of data (Fig. 4). The resulting concentration data will now be expressed in several ways, acknowledging the lessons learned from the ‘free concept’ of brain penetration (Jeffrey & Summerfield, 2007; Hammarlund-Udenaes et al. 2008). First, the classic Kp is being calculated. Using fu,brain and fu,plasma data from the first tier, Kp is then transformed into the unbound Kp,uu. While the classic Kp value seems to be driven predominantly by non-specific binding to brain lipids and hence may just be an in vivo measure of lipophilicity, its unbound relative, Kp,uu is a measure of the extent of the distribution equilibrium of a compound between the unbound fractions in brain and in blood plasma. If the value is close to unity, passive diffusion across the BBB can be assumed (or any influx being offset by efflux). This interpretation may be further supported by the in vitro permeability and efflux ratio data from the MDR1-MCDK or Caco-2 assay. In case the distribution between plasma and brain is mainly determined by passive diffusion, free brain concentrations may be approximated directly from free plasma levels. The unbound volume of distribution, Vu,brain, is an indicator for the distribution behaviour of a compound within the brain and is irrespective of the extent of equilibrium between brain and plasma. Vu,brain is an apparent volume term which is calculated by dividing the total amount of drug in brain (corrected for the amount in the cerebral vasculature) by the unbound concentration of the drug in brain. Hence, Vu,brain does not describe a ‘real’ volume and is interpreted in analogy to the ‘classic’ Vss which is calculated from plasma concentration-time profiles obtained after iv dosing. Vu,brain indicates whether a compound distributes merely into ISF (Vu,brain ~0.2 ml/g brain), throughout the brain water space, i.e. into both ISF and ICF (~0.8 ml/g brain), or has a tendency to non-specifically bind to brain tissue (Vu,brain >0.8 ml/g brain).
Because permeability is a principal prerequisite for CNS penetration it is tempting to optimise compounds for a very high permeability applying rapid in vitro screens. However, a normal to good permeability may suffice well and chemistry efforts should rather be focused on improving fu,brain and hence Kp,uu and Cu,brain as these two parameters have a much greater impact on the desired pharmacological efficacy. Indeed, permeability may be of lesser importance during chronic administration schedules and even poor permeants are able to elicit significant pharmacological effects in human. Indeed, high specific binding at the pharmacological target in the CNS and greater free fractions in brain can counterbalance poor BBB permeation and/or extensive plasma protein binding. As a very general guidance for an optimal PK target profile of CNS drugs, Kp,uu should be close to 1, Papp >150 nm with little efflux, and fu,brain should be such as to allow the unbound concentrations in vivo to exceed the target affinity as determined in vitro. Kp may be recorded but should not be taken for decision making. Total CL and Vss should be kept low enough as to provide a high plasma exposure.
In summary, the complex structure of the CNS makes brain penetration a very complex feature which cannot be rationalised on the basis of any single parameter. Although total brain concentrations are still the most common measure of CNS exposure, it is now emerging that they are more of an indication of high non-specific binding to brain tissue rather than being pharmacologically relevant. Neither total brain levels nor BBB permeability can be taken without considering the binding capacity of the brain tissue when a link between exposure and efficacy is needed. The current paradigm of brain penetration therefore is changing towards a more compartmentalised view which allows to better rationalise the distribution of compounds within the brain and makes use of brain compartments which are more relevant pharmacological effects. The holistic concept of CNS penetration considers rate of permeation across the BBB, extent of brain penetration and the intra-brain distribution of a CNS drug as distinct but interrelated properties of a CNS drug which have to be determined by different in vitro and in vivo methods (Reichel 2009). Therefore, the integration of data from various assays and studies is becoming a central part of the evolving paradigm in order to develop quantitative relationships between dose, exposure and efficacy. By integrating these data and technologies into PK/PD modeling and simulations it will be possible to generate a working understanding of the pharmacokinetics and pharmacodynamics of potential drug candidates in the human CNS. Such an understanding will be paramount to define PK parameters which are favourable for the desired indication, to guide the preclinical development of the compound (e.g. dose selection for toxicity testing in higher species as well as first-in-man and therapeutic doses) and ultimately to increase the chances for a successful phase II study in human patients.
References
1. Jeffrey P, Summerfield SG (2007) Challenges for blood-brain barrier (BBB) screening. Xenobiotica 37(10-11):1135-1151
2. Hammarlund-Udenaes M, Fridén M, Syvänen S, Gupta A. (2008) On the rate and extent of drug delivery to the brain. Pharm. Res. 25(8):1737-1750
3. Reichel A. (2009) Addressing central nervous system (CNS) penetration in drug discovery: basics and implications of the evolving new concept. Chem. Biodivers. 6(11):2030-2049
Conference:
Pharmacology and Toxicology of the Blood-Brain Barrier: State of the Art, Needs for Future Research and Expected Benefits for the EU, Brussels, Belgium, 11 Feb - 12 Feb, 2010.
Presentation Type:
Oral Presentation
Topic:
Presentations
Citation:
Reichel
A
(2010). The Blood-Brain Barrier and CNS Penetration: a Drug Discovery Point of View.
Front. Pharmacol.
Conference Abstract:
Pharmacology and Toxicology of the Blood-Brain Barrier: State of the Art, Needs for Future Research and Expected Benefits for the EU.
doi: 10.3389/conf.fphar.2010.02.00028
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Received:
11 Mar 2010;
Published Online:
11 Mar 2010.
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Correspondence:
Andreas Reichel, Bayer Schering Pharma, Research Pharmacokinetics, Berlin, Germany, Andreas.Reichel@BayerHealthCare.com