Predicting the intrinsic membrane permeability by the solubility-diffusion model

Membranes, mainly built of phospholipid-bilayers, are the natural boundary for organisms to differentiate between exterior and interior environment–being an essential precondition for life. Due to their specific shape as a bilayer, membranes have a highly anisotropic structure including charged headgroups as well as an alkane-like hydrocarbon core. This affects the passive permeability of organic chemicals (neutral and ionic species) through membranes.

Apparent permeabilities through cell monolayers such as Caco-2 are considered to be an in-vitro gold standard for assessing the uptake efficiency of drugs. Yet, in contrast to planar lipid bilayer experiments that are well explained by the solubility diffusion model, intrinsic neutral membrane permeabilities extracted from Caco-2 experiments seemed not well predicted by the solubility diffusion model in the past.

Caco-2/MDCK model Mechanistic model of CACO-2/MDCK permeability, adapted from Bittermann and Goss 2017

We started our work on membrane permeability by setting up a model for the prediction of the passive permeability of organic chemicals through Caco-2 and MDCK monolayers based on an extensive literature review (Bittermann and Goss 2017) . The model was later refined to account for concentration-shift effects in the presence of pH differences between the apical and basolateral compartment and in the cytosol, that can lead to a decreased or increased apparent permeability through aqueous layers (Dahley et al. 2023).

Due to the limited overlap in existing experimental data between Caco-2/MDCK and planar lipid bilayers, we sought to enhance the overlap by generating our own permeability data in MDCK cells and black lipid membranes (BLM) (Dahley et al. 2022, 2023, 2024). Correlating MDCK and BLM data, we were able to extract a correlation between both permeabilities. This correlation not only shows that MDCK permeability is about 2 log units lower than BLM permeability, but allows for the in silico prediction of intrinsic Caco-2/MDCK permeability by the solubility diffusion model (Dahley et al. 2024). 

Correlation between experimental permeability measured in MDCK and BLM Correlation between experimental permeability measured in MDCK and BLM, taken from Dahley et al. 2024

This correlation also held true when predicting permeabilities extracted from literature, even for zwitterionic compounds (Ebert and Dahley 2024), but only after are thorough re-evaluation of literature data that were often limited by other effects than membrane permeability, such as the diffusion through the aqueous boundary layers (Ebert et al. 2024).

The model is built on the widespread assumption that ionic species do not passively permeate through membranes but can only take the paracellular route. While this assumption may hold in most cases, there are cases where passive ionic permeability through membranes cannot simply be neglected. Important examples are permanent ions or uncouplers (ionophores that move protons across lipid bilayers). For their up-take or effect, ionic permeability can be the limiting process.

Permeability data on organic ions are sparse in literature. Therefore, in cooperation with Prof. Peter Pohl (Institute of Biophysics, JKU Linz, Austria), we made our own electrophysiological measurements on planar lipid bilayers. We found anionic permeabilities ranging over more than 10 log units, and were able to establish a correlation to successfully predict anionic membrane permeability based on the solubility diffusion model (Ebert et al. 2018). This correlation also holds true for perfluoroalkylic acids (Ebert et al. 2020). These compounds exist mostly in their anionic form due to their low pKa. In cooperation with BIOVIA / Dassault Systèmes we also extended our predictions of membrane permeability to cationic organic compounds (Schwöbel et al. 2020).

While passive permeability through the membrane may not always be the limiting transport process, knowledge about passive membrane permeability can help assessing whether active transport is of relevance. Also, passive membrane permeability is a key to predicting uncoupling activity.

Our future focus will be possible size limitations for membrane permeability and the permeability through the blood brain barrier.

Related own publications:

Bittermann, Kai, and Kai Uwe Goss. 2017. “Predicting Apparent Passive Permeability of Caco-2 and MDCK Cell-Monolayers: A Mechanistic Model.” PLoS ONE 12 (12): 1–20.

Dahley, Carolin, Estella Dora Germaine Garessus, Andrea Ebert, and Kai Uwe Goss. 2022. “Impact of Cholesterol and Sphingomyelin on Intrinsic Membrane Permeability.” Biochimica et Biophysica Acta - Biomembranes 1864 (9): 183953.

Dahley, Carolin, Kai Uwe Goss, and Andrea Ebert. 2023. “Revisiting the PKa-Flux Method for Determining Intrinsic Membrane Permeability.” European Journal of Pharmaceutical Sciences 191 (July): 106592.

Dahley, Carolin, Tim Böckmann, Andrea Ebert, and Kai Uwe Goss. 2024. “Predicting the Intrinsic Membrane Permeability of Caco-2/MDCK Cells by the Solubility-Diffusion Model.” European Journal of Pharmaceutical Sciences 195 (January). 106720.

Ebert, A., and C Dahley. 2024. “Can Membrane Permeability of Zwitterionic Compounds Be Predicted by the Solubility Diffusion Model?” Eur. J. Pharm. Sci. 199 (August): 106819.

Ebert, Andrea, Flora Allendorf, Urs Berger, Kai Uwe Goss, and Nadin Ulrich. 2020. Membrane/Water Partitioning and Permeabilities of Perfluoroalkyl Acids and Four of Their Alternatives and the Effects on Toxicokinetic Behavior. Environmental Science & Technology. Vol. 54.

Ebert, Andrea, Carolin Dahley, and Kai Uwe Goss. 2024. “Pitfalls in Evaluating Permeability Experiments with Caco-2/MDCK Cell Monolayers.” European Journal of Pharmaceutical Sciences 194 (October 2023): 106699.

Ebert, Andrea, Christof Hannesschlaeger, Kai Uwe Goss, and Peter Pohl. 2018. “Passive Permeability of Planar Lipid Bilayers to Organic Anions.” Biophysj 115 (10): 1931–41.

Schwöbel, Johannes A.H., Andrea Ebert, Kai Bittermann, Uwe Huniar, Kai Uwe Goss, and Andreas Klamt. 2020. “COSMO Perm: Mechanistic Prediction of Passive Membrane Permeability for Neutral Compounds and Ions and Its PH Dependence.” Journal of Physical Chemistry B 124 (16): 3343–54.