Simulating the influence of plasma protein on measured receptor affinity in biochemical assays reveals the utility of Schild analysis for estimating compound affinity for plasma proteins

• Published in: British journal of pharmacology 2015-11, Vol.172 (21), p.5037-5049
• Main Authors: Blakeley, D, Sykes, D A, Ensor, P, Bertran, E, Aston, P J, Charlton, S J
• Format: Article
• Language: English
• Subjects: Blood Proteins - metabolism; Ligands; Mathematical models; Models, Theoretical; Pharmaceutical Preparations - metabolism; Plasma; Protein Binding; Proteins; Receptors, Drug - metabolism; Research Paper; Research Papers
• ISSN: 0007-1188; 1476-5381
• DOI: 10.1111/bph.13263

Background and Purpose Plasma protein binding (PPB) influences the free fraction of drug available to bind to its target and is therefore an important consideration in drug discovery. While traditional methods for assessing PPB (e.g. rapid equilibrium dialysis) are suitable for comparing compounds with relatively weak PPB, they are not able to accurately discriminate between highly bound compounds (typically >99.5%). The aim of the present work was to use mathematical modelling to explore the potential utility of receptor binding and cellular functional assays to estimate the affinity of compounds for plasma proteins. Plasma proteins are routinely added to in vitro assays, so a secondary goal was to investigate the effect of plasma proteins on observed ligand–receptor interactions. Experimental Approach Using the principle of conservation of mass and the law of mass action, a cubic equation was derived describing the ligand–receptor complex [LR] in the presence of plasma protein at equilibrium. Key Results The model demonstrates the profound influence of PPB on in vitro assays and identifies the utility of Schild analysis, which is usually applied to determine receptor‐antagonist affinities, for calculating affinity at plasma proteins (termed KP). We have also extended this analysis to functional effects using operational modelling and demonstrate that these approaches can also be applied to cell‐based assay systems. Conclusions and Implications These mathematical models can potentially be used in conjunction with experimental data to estimate drug‐plasma protein affinities in the earliest phases of drug discovery programmes.