Monitoring Drug Target Engagement in Cells and Tissues Using the Cellular Thermal Shift Assay

Abstract

The efficacy of therapeutics is dependent on a drug binding to its cognate target. Optimization of target engagement by drugs in cells is often challenging, because drug binding cannot be monitored inside cells. We have developed a method for evaluating drug binding to target proteins in cells and tissue samples. This cellular thermal shift assay (CETSA) is based on the biophysical principle of ligand-induced thermal stabilization of target proteins. Using this assay, we validated drug binding for a set of important clinical targets and monitored processes of drug transport and activation, off-target effects and drug resistance in cancer cell lines, as well as drug distribution in tissues. CETSA is likely to become a valuable tool for the validation and optimization of drug target engagement.

Axitinib blocks Wnt/β-catenin signaling and directs asymmetric cell division in cancer

“…In addition, axitinib at 10 μM significantly increased the stability of SHPRH in an intact living cell-based cellular thermal shift assay (CETSA) assay (Fig. 3C) (34), supporting the physical interaction of axitinib with SHPRH in vivo.…”

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Plant Chemical Genetics: From Phenotype-Based Screens to Synthetic Biology

“…Notably, biochemical approaches, such as affinity purification, or label-free approaches, such as the cellular thermal shift assay (CETSA; Martinez Molina et al, 2013), are well suited to identify direct targets, especially at a proteome-wide level. Numerous examples of affinity purifications exist in mammalian systems (Ziegler et al, 2013).…”