A High-Throughput Cell-Based Method to Predict the Unbound Drug Fraction in the Brain

ABSTRACT: Optimization of drug efficacy in the brain requires understanding of the local exposure to unbound drug at the site of action. This relies on measurements of the unbound drug fraction ( f u,brain), which currently requires access to brain tissue. Here, we present a novel methodology using homogenates of cultured cells for rapid estimation of f u,brain. In our setup, drug binding to human embryonic kidney cell (HEK293) homogenate was measured in a small-scale dialysis apparatus. To increase throughput, we combined drugs into cassettes for simultaneous measurement of multiple compounds. Our method estimated f u,brain with an average error of 1.9-fold. We propose that our simple method can be used as an inexpensive, easily available and high-throughput alternative to brain tissues excised from laboratory animals. Thereby, estimates of unbound drug exposure can now be implemented at a much earlier stage of the drug discovery process, when molecular property changes are easier to make