Since the protein targets of any given compound hit from a phenotypic screen are initially unknown, one major challenge with phenotypic–based drug discovery is the need for target identification or deconvolution. Choosing the most efficient and reliable method for target deconvolution from the outset can minimize risks to your drug discovery pipeline and therefore help protect your investments.
Combining CETSA® with a mass spectrometric (MS) protein quantification readout makes it possible to measure not only the direct binding of a compound to various proteins in the cellular environment but also to assess subsequent downstream consequences of the initial interactions. Therefore, CETSA® Explore allows for unbiased proteome-wide identification of direct interactions with individual proteins, as well as effects on cellular pathways. Being unbiased and hypothesis-free, CETSA® Explore can be used for target deconvolution studies for hit compounds from phenotypic screens. Since CETSA® can be performed in any sample matrix, the profiling and deconvolution can be conducted directly in disease relevant systems, such as those used for the phenotypic screen
There are several examples in the public domain where CETSA® Explore has been used for target deconvolution. Kitawaga et al. reported on the use of CETSA® Explore for understanding the cellular targets and signaling pathways of hit compounds from a phenotypic screen. The experiments provided strong evidence that PIP4Ks are the pharmacological protein targets of these novel compound hits. In another report, CETSA® Explore was used to investigate the targets of two antimalarial drugs. The study effectively identified purine nucleoside phosphorylase as the target of quinine and mefloquine and provided critical knowledge for the understanding of the resistance mechanisms of these structurally related quinoline drugs.
Mayumi Kitagawa, Pei-Ju Liao, Kyung Hee Lee, Jasmine Wong, See Cheng Shang, et al. Dual blockade of the lipid kinase PIP4Ks and mitotic pathways leads to cancer-selective lethality. (2017) Nature Communications. DOI: 10.1038/s41467-017-02287-5
Dziekan JM, Yu H, Chen D, Dai L, et al. Identifying purine nucleoside phosphorylase as the target of quinine using cellular thermal shift assay. (2019) Sci Transl Med. DOI: 10.1126/scitranslmed.aau3174
Friman T. Mass spectrometry-based Cellular Thermal Shift Assay (CETSA®) for target deconvolution in phenotypic drug discovery. (2020) Bioorganic & Medicinal Chemistry. DOI: https://doi.org/10.1016/j.bmc.2019.115174