Abstract

Successful drug discovery depends on the compound being therapeutic: it must bind to the right target and be both effective and safe. To achieve this, a targeted drug must be present at the site of action and occupy the intended target with high specificity and selectivity. 

The Cellular thermal shift assay, CETSA ®, is a label-free method that measures compound target engagement in live cells. Mass spectrometry-based CETSA offers unbiased proteome wide analysis of a compound’s effect on thousands of proteins.

Since drug treatment affects the primary cellular target(s) and have downstream pathway effects, the MS format makes it possible to understand the mode of action for a particular drug by following the global changes in protein stabilities that compound treatment causes. By comparing results from different cell matrices, various doses and incubation lengths makes it possible to understand key parameters such as cellular uptake, target specificity and discovery off-target effects.

Here Pelago Bioscience present two examples of how CETSA MS has enabled identification of primary hits and novel biomarkers as well as confirmed results from alternative phenotypic deconvolution programs. The results suggest that CETSA MS can be used not only for primary target engagement measurements, but also to reveal the wider biological impact of a drug in a physiologically relevant system.