Brain tissue binding of drugs: evaluation and validation of solid supported porcine brain membrane vesicles (TRANSIL) as a novel high-throughput method.
Estimating the unbound fraction of drugs in brain has become essential for the evaluation and interpretation of the pharmacokinetics and pharmacodynamics of new central nervous system drug candidates. Dialysis-based methods are considered to be accurate for estimating the fraction unbound in brain; however, these techniques are hampered by a low throughput. In this study, we present a novel, matrix-free, high-throughput method for estimating the unbound fraction, based on a sample pooling approach combining the TRANSIL brain absorption assay with liquid chromatography-mass spectrometry. The base measurement of the TRANSIL approach is the affinity to brain membranes, and this method is used directly to predict the free fraction in brain. The method was evaluated by comparing the free fraction of drugs in brain [f(u,brain) (%)] obtained using the TRANSIL brain absorption assay and equilibrium dialysis methods for a test set of 65 drugs (27 marketed and 38 proprietary drugs). A good correlation (r(2) > 0.93) of f(u,brain) (%) between the TRANSIL brain absorption assay and equilibrium dialysis was observed. Moreover, we compared the lipid composition of rat and porcine brain and analyzed the influence of the brain albumin content on brain tissue binding measurement. The comparison of the lipid composition indicated only minor differences between rat and porcine brain, and albumin appears to have a low impact on brain tissue binding measurements. The TRANSIL brain absorption assay with sample pooling methodology not only significantly reduces the biological matrix required but also increases the throughput, compared with the conventional dialysis methods.
GlaxoSmithKline Neurosciences Centre of Excellence for Drug Discovery, Verona, Italy. raffaele.longhi@aptuit.com
Raffaele
R
Longhi
Silvia
S
Corbioli
Stefano
S
Fontana
Federicia
F
Vinco
Simone
S
Braggio
Lydia
L
Helmdach
Jürgen
J
Schiller
Hinnerk
H
Boriss
D000042:Absorption; D000418:Albumins / Q000378:metabolism; D000818:Animals; D001921:Brain / Q000378:metabolism*; D001923:Brain Chemistry; D002462:Cell Membrane / Q000378:metabolism*; D002851:Chromatography, High Pressure Liquid; D004353:Drug Evaluation, Preclinical / Q000295:instrumentation / Q000379:methods*; D066298:In Vitro Techniques; D008055:Lipids / Q000032:analysis; D008297:Male; D017551:Microdialysis; D004364:Pharmaceutical Preparations / Q000378:metabolism*; D011485:Protein Binding; D051381:Rats; D019032:Spectrometry, Mass, Matrix-Assisted Laser Desorption-Ionization; D013552:Swine; D014018:Tissue Distribution
D000418:Albumins; D008055:Lipids; D004364:Pharmaceutical Preparations