Category: Automation and High-Throughput Technologies
Affinity of a therapeutic molecule binding to a target protein is one of the primary characteristics that delimits therapeutic potency in vivo and in vitro. It is critical to develop robust and reliable methods to study binding kinetics and equilibrium of therapeutic candidates to protein targets resident or associated with the cell membrane given their preponderance as therapeutic drug targets. Unlike soluble proteins with well-defined protein concentration values, membrane protein expression is often heterogeneous and sensitive to culture conditions for the expressing cells. In addition to unknown membrane protein concentration, therapeutic protein binding to targets located on the cell surface can induce target internalization and/or signaling. Target mediated activities can concomitantly affect the concentration of antibody/antigen complex that is measured in cell binding assays for affinity determinations. To overcome some of these difficulties, we established an automated cell membrane equilibrium binding assay platform with integration of FACS and KinExA systems. Automation enabled sample preparation of four candidate therapeutic molecules for determination of their solution equilibrium affinity for the same target simultaneously. FACS analysis measures target protein density on the cell surface prior to membrane preparation, while KinExA quantifies free antibody concentration at equilibrium of antibody binding with cell membrane for Kd determination. This presentation describes the development, application, and automation of solution based, label free binding analysis for membrane proteins, such as CD20, resident in their natural environments.
Qing Chen– Principal Scientist, Amgen, Oxnard, CA
Joined Amgen in 1998, I have obtained deep understanding of protein therapeutic discovery and development through participation and leading of various research projects. I led cross functional teams to advance 3 projects achieving PST status (AMG 777, AMG 434, and AMG 842). With expertise in binding kinetic/equilibrium analysis, I established Biomolecular Interaction Analysis (BMI) platform to study protein-protein interactions using complementary Biochemical and Biophysical technologies, such as SPR, BLI, and KinExA. Leading BMI groups at ATO (2000 – current) and AWA (2011-2014), I have supported advancement of almost all large molecule programs from ST/PT to RPT and PST. I was also involved in development, regulatory filing and IP protection of molecules, such as Prolia/Xgeva, Vectibix, Repatha, Romosozumab, AMG 334, and Biosimilars.