Application and Cross-Validation of a High-Throughput SPR Method for Characterizing Covalent Binding Ligands

Abstract

We validate and present the application of a high-throughput surface plasmon resonance (SPR) method utilizing a regenerable sensor chip method for rapid determination of kinact/KI of irreversible covalent inhibitors. Using covalent MK2 and KRAS-G12C inhibitors as case studies, we demonstrated that kinact/KI results by SPR exhibit an excellent correlation with results by conventional methods, such as intact protein mass spectrometry and time-dependent enzymatic assays. In addition, the use of regenerable biotin chips greatly improves the throughput and reduces the cost. In summary, our studies highlight SPR as an efficient, accurate, and cost-effective method for determining kinact/KI, which offers distinctive advantages over traditional methods such as enzymatic and MS-based methods.