title: High performance affinity chromatography (HPAC) as a high-throughput screening tool in drug discovery to study drug-plasma protein interactions.
abstract: Drug-plasma protein binding is an important parameter that, together with other physicochemical properties such as lipophilicity and pK(a), greatly influences drug absorption, distribution, metabolism, and excretion (ADME). Therefore, it is important for pharmaceutical companies to develop a rapid screening assay to examine plasma protein binding during the early stages of the drug discovery process. Human serum albumin (HSA) and α(1)-acid glycoprotein (AGP) are the most important plasma proteins that are capable of binding drugs. In this work, an automated and high-throughput (<3 min/compound) strategy was developed using high performance affinity chromatography (HPAC) with commercial HSA and AGP columns to evaluate drug-plasma protein interactions for drug screening. A generic gradient was used throughout the study to separate drugs that were weakly and tightly bound to HSA and AGP. To accelerate the analysis time, the system was calibrated in a single run by pooling reference compounds without overloading the column. For both HSA and AGP studies, the developed methods were successfully transferred from HPAC-UV to HPAC-MS with single quadrupole MS detection and ammonium acetate, pH 7.0 as a volatile mobile phase. The MS detection enhanced the sensitivity, selectivity, and throughput of the method by pooling unknown compounds. For HSA analyses, the binding percentages obtained using HPAC were well correlated with the binding percentages from the literature. This method was also able to rank compounds based on their affinity for HSA. Concerning the AGP analyses, the quality of the correlation between the binding percentages obtained in HPAC and those from the literature was weaker. However, the method was able to classify compounds into weak, medium, and strong binders and rank compounds based on their affinity for AGP.
authors.affiliation: School of Pharmaceutical Sciences, University of Geneva, University of Lausanne, 30, Quai Ernest-Ansermet, 1211 Geneva 4, Switzerland.
authors.forename: Karine
authors.initials: K
authors.lastname: Vuignier
authors.forename: Davy
authors.initials: D
authors.lastname: Guillarme
authors.forename: Jean-Luc
authors.initials: JL
authors.lastname: Veuthey
authors.forename: Pierre-Alain
authors.initials: PA
authors.lastname: Carrupt
authors.forename: Julie
authors.initials: J
authors.lastname: Schappler
mesh_terms: D001798:Blood Proteins / Q000032:analysis* / Q000378:metabolism; D002846:Chromatography, Affinity / Q000379:methods* / Q000592:standards; D055808:Drug Discovery / Q000379:methods*; D004347:Drug Interactions / Q000502:physiology*; D057166:High-Throughput Screening Assays / Q000379:methods* / Q000592:standards; D006801:Humans; D004364:Pharmaceutical Preparations / Q000097:blood*; D011485:Protein Binding / Q000502:physiology
chemical_list: D001798:Blood Proteins; D004364:Pharmaceutical Preparations