Unraveling the Rules of Passive Permeation Through the Blood-Brain Barrier

Failure to cross the Blood-Brain Barrier (BBB) is a major attrition factor in drug development for brain disorders affecting millions of people in Europe [1]. Most xenobiotics are not recognized by active transporters and therefore passive permeation is a significant route, if not the most important, for their permeation through the BBB under non-disruptive conditions [2]. The establishment of Quantitative Structure Property Relations (QSPRs) for passive permeation is of major impact in drug development as it allows the selection of drug candidates with better in vivo efficacy at early stages.
The current paradigm of passive permeation through biomembranes, based on the Overton rule, does not allow a quantitative description and has limited predictive value [3]. These limitations derive from unsuitable predictors for solute association with and diffusion through biomembranes and little knowledge of the specificity of membrane efflux proteins such as P-glycoprotein (P-gp). This lack of QSPRs lays on the fact that permeation is the result of several steps (partition, diffusion and P-gp efflux), each dependent on distinct structural properties of the permeating solute. In contrast to current approaches, in this project we will establish QSPRs for each of the relevant steps in the permeation process. Each step depends on well defined structural parameters and good correlations are anticipated (preliminary data in the attachments). Binding to blood components also plays a significant role in the observed permeation [4] and this will also be evaluated in this project. The global permeation will then be calculated using mechanistic models based on the predicted parameters for each step [4].
This project includes computational studies, as well as experimental studies using model systems (liposomes with a lipid composition representative of endothelial membranes) and a novel tissue-engineered BBB mimic. The model systems represent a major improvement when compared with the usually considered methodologies where the rate of passive permeation is predicted from the partition of the solute to homogeneous non-polar fluids [2]. The use of liposomes permits focusing on passive pathways while maintaining the structural, dynamic and electrostatic properties of the natural barrier.
The information available in the literature for the relevant parameters in the permeation of small molecules through lipid bilayers (partition, insertion/desorption and translocation) and association with blood components will first be collected to build a manually curated database (Task1). This will be complemented by new experimental results and with Molecular Dynamics (MD) simulations when the information available for a given of compounds is not considered sufficient (Task2). Widely applied feature selection and dimension reduction techniques [5] will be employed to determine the most important parameters for each step in the permeation process, which will then be used to derive QSPRs.
QSPRs will also be attempted for the intrinsic activity of P-gp. This will be achieved through consideration of the effect of P-gp on the overall drug’s rate of permeation through defect free liposomes including reconstituted P-gp. The distinct pathways for permeation (passive and P-gp efflux) will be included in a mechanistic model [4] allowing the quantitative evaluation of P-gp activity. The molecular determinants of the drugs interaction with P-gp will be assessed experimentally and from MD simulations. (Tasks 3 and 4)
A computer application will be developed (Task5) to predict the ability of drugs to passively permeate across tight endothelia, based on the structure of the solute and on the QSPRs identified for each step. The predictive value of this application will be tested and validated using a confluent cell monolayer of human endothelial cells grown in conditions so as to develop all characteristics of the in vivo system, including the glycocalyx (Task6).
The results from this study will produce a substantial impact in pharmacology and medicinal chemistry, and thus on therapeutics and public health. The integrated knowledge here obtained will allow a rational design of drugs targeted to the brain as well as reduce secondary effects associated with the treatment of non-brain targeted diseases.

1. Gustavsson, A., ... and J. Olesen, (2011), "Cost of disorders of the brain in Europe 2010", Eur. Neuropsychopharmacol., 21, 718
2. Smith, D., P. Artursson, A. Avdeef, L. Di, G. F. Ecker, B. Faller, J. B. Houston, M. Kansy, E. H. Kerns, S. D. Kramer, H. Lennernas, H. van de Waterbeemd, K. Sugano and B. Testa, (2014), "Passive Lipoidal Diffusion and Carrier-Mediated Cell Uptake Are Both Important Mechanisms of Membrane Permeation in Drug Disposition", Molecular Pharmaceutics, 11, 1727
3. Gleeson, M. P., A. Hersey and S. Hannongbua, (2011), "In-Silico ADME Models: A General Assessment of their Utility in Drug Discovery Applications", Current Topics in Medicinal Chemistry, 11, 358
4. Filipe, H. A. L., A. Salvador, J. M. Silvestre, W. L. C. Vaz and M. J. Moreno, (2014), "Beyond Overton's Rule: Quantitative Modeling of Passive Permeation through Tight Cell Monolayers", Molecular Pharmaceutics, 11, 3696
5. Mensch, J., J. Oyarzabal, C. Mackie and P. Augustijns, (2009), "In vivo, in vitro and in silico methods for small molecule transfer across the BBB", J. Pharm. Sci., 98, 4429