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

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Science  05 Jul 2013:
Vol. 341, Issue 6141, pp. 84-87
DOI: 10.1126/science.1233606

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Drug Targeting

Drug efficacy depends on the extent of binding to a cellular target (often a protein) with adverse effects caused by excessive target binding or by off-target binding. Engagement of a target protein inside cells is influenced by the effective drug concentration and by factors that regulate the protein conformation, making it difficult to predict efficacy based on in vitro affinity studies. Martinez Molina et al. (p. 84) took advantage of the shift in protein thermal stability caused by drug binding to directly monitor target protein-drug interactions in cells. The method was used to monitor drug target engagement in cancer cells and in mouse livers and kidneys.


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.

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