High-resolution and high-throughput protocols for measuring drug/human serum albumin interactions using BIACORE

Area of Science:

• Pharmacology
• Biochemistry
• Analytical Chemistry

Background:

• Characterizing chemical compound binding to human serum albumin (HSA) is crucial for drug development.
• Understanding drug-HSA interactions aids in predicting drug efficacy and safety.
• Warfarin serves as a model compound to validate new binding assay technologies.

Purpose of the Study:

• To validate BIACORE Surface Plasmon Resonance (SPR) biosensors for determining drug-HSA binding constants.
• To establish a high-throughput protocol for assessing drug-HSA interactions.
• To demonstrate the reliability and applicability of SPR technology in drug discovery.

Main Methods:

• Utilized BIACORE SPR biosensors with immobilized HSA to study warfarin binding.
• Determined binding constants (affinity and dissociation rates) for warfarin-HSA interactions.
• Developed and applied a high-throughput protocol using low compound concentrations and simultaneous data fitting for 10 test compounds.

Main Results:

• SPR biosensor measurements for warfarin-HSA binding were highly reproducible.
• Determined binding affinity (K(d) = 3.7 ± 1.2 µM) and dissociation rate (k(d) = 1.2 s⁻¹) for warfarin matched previous findings.
• SPR-determined % bound values correlated well with solution-based methods, validating the SPR protocol.

Conclusions:

• BIACORE SPR biosensor technology reliably characterizes drug-HSA interactions, even for small molecules.
• The developed high-throughput protocol enables efficient evaluation of drug candidates.
• SPR technology offers a sensitive, automated method with minimal sample requirements for drug discovery.