Targeting Proteins for Degradation: Characterizing PROTAC Kinetics and Mode of Action Using Live-Cell Assays
Summary
A new generation of heterobifunctional small molecules, termed PROTACs, offer significant potential for new drug discovery strategies by targeting specific proteins for removal from the cell. Characterizing and optimizing PROTACs for degradation efficacy represents a significant challenge, particularly in understanding the individual processes and potential failure points that control the resulting degradation.
In this webinar, Dr. Riching presents luminescence-based and energy transfer technologies for live-cell, kinetic characterization of PROTAC compound mechanism of action. To quantify key degradation parameters, including rate, recovery, degradation max (Dmax) and DC50, she will show how CRISPR-Cas9 genetic tagging of target proteins with HiBiT enables extended kinetic monitoring of endogenous target protein levels after PROTAC treatment. Using BET family proteins and PROTACs as a case study, she will demonstrate how to measure key cellular mechanisms that affect PROTAC functionality, including permeability, PROTAC compound binding affinity to both target and E3 ligase in lytic and live-cell formats, kinetics of E3 ternary complex formation, target ubiquitination and recruitment to the proteasome.
Speaker
Krristin M. Riching, PhD
Senior Research Scientist
Promega Corporation
Kristin Riching received her Ph.D. from the University of Wisconsin–Madison, where she studied the structural and mechanical properties of collagen fibers and their effects on breast cancer cell migration in invasive ductal carcinoma. She then worked as a postdoctoral researcher at Promega studying ligand-induced protein interactions within the ErbB signaling network and their roles in cancer progression. She has been at Promega for four years and is currently a Senior Scientist developing approaches to monitor functional mechanisms of proteasome-mediated protein degradation.