A Shift in Thinking: Cellular Thermal Shift Assay-Enabled Drug Discovery

Abstract

A decade has passed since the cellular thermal shift assay (CETSA) was introduced to the drug discovery community. Over the years, the method has guided numerous projects by providing insights about, for example, target engagement, lead generation, target identification, lead optimization, and preclinical profiling. With this Microperspective, we intend to highlight recently published applications of CETSA and how the data generated can enable efficient decision-making and prioritization throughout the drug discovery and development value chain.

Modular Nanotransporters Delivering Biologically Active Molecules to the Surface of Mitochondria

“…The interaction of MNT 1 with Keap1 in AML-12 cells was also studied using cellular thermal shift assays (CETSA) [ 34 , 35 , 36 , 37 , 38 ]. Using this method, a Keap1 melting curve in its natural protein environment was obtained in AML-12 cells ( Figure 4 , blue curve).…”

“…In summary, in AML-12 cells transiently transformed with Keap1-hrGFP, MNT 1 is able to interact with Keap1. In order to understand whether MNT 1 is able to interact with native intracellular Keap1 cellular thermal shift assays (CETSA) were used [ 34 , 35 , 36 , 37 , 38 ]. Using this method, so-called protein melting curves are obtained, which depend on whether the protein forms a complex with other molecules or not.…”

“…The cellular thermal shift assay (CETSA) [ 34 , 35 , 36 , 37 , 38 ] was performed in the following way. Cells from 25 cm 2 cultural flask were trypsinized and pelleted (200× g , 5 min).…”

Quantitative target engagement of RIPK1 in human whole blood via the cellular thermal shift assay for potential pre-clinical and clinical applications

“…In drug discovery, understanding and quantifying small molecule target engagement is crucial for the characterization of preclinical compounds and clinical drug candidates. Target engagement helps to validate the mechanism of action of lead molecules, determine pharmacokinetic-pharmacodynamic relationships in animal models and trials, and evaluate and mitigate potential off-targets (1,2).…”