CETSA-based target engagement of taxanes as biomarkers for efficacy and resistance

Abstract

The use of taxanes has for decades been crucial for treatment of several cancers. A major limitation of these therapies is inherent or acquired drug resistance. A key to improved outcome of taxane-based therapies is to develop tools to predict and monitor drug efficacy and resistance in the clinical setting allowing for treatment and dose stratification for individual patients. To assess treatment efficacy up to the level of drug target engagement, we have established several formats of tubulin-specific Cellular Thermal Shift Assays (CETSAs). This technique was evaluated in breast and prostate cancer models and in a cohort of breast cancer patients. Here we show that taxanes induce significant CETSA shifts in cell lines as well as in animal models including patient-derived xenograft (PDX) models. Furthermore, isothermal dose response CETSA measurements allowed for drugs to be rapidly ranked according to their reported potency. Using multidrug resistant cancer cell lines and taxane-resistant PDX models we demonstrate that CETSA can identify taxane resistance up to the level of target engagement. An imaging-based CETSA format was also established, which in principle allows for taxane target engagement to be accessed in specific cell types in complex cell mixtures. Using a highly sensitive implementation of CETSA, we measured target engagement in fine needle aspirates from breast cancer patients, revealing a range of different sensitivities. Together, our data support that CETSA is a robust tool for assessing taxane target engagement in preclinical models and clinical material and therefore should be evaluated as a prognostic tool during taxane-based therapies.

Metalloprotease Gp63 targeting novel glycoside exhibits potential antileishmanial activity

“…This was in accordance with previous reports suggesting role of Gp63 in promastigote multiplication [46] Further to assess strong interactions between a drug and its protein target in a physiologically relevant cellular environment we performed CETSA. Previous studies showed that CETSA has been performed in complex protein samples and in live cells displaying an altered melting or aggregation behaviour upon exposure to increased temperature [47]. CETSA has also been employed to identify Plasmodium falciparum protein targets in parasite lysates [48].…”

Recent advances in proteome‐wide label‐free target deconvolution for bioactive small molecules

“…The applications of CETSA extend well beyond the cellular level. Very encouragingly, the Western‐blotting‐ and imaging‐based CETSA experiments have been successfully applied to assess drug–target engagement activities directly in mice and human tumor biopsies 22,87,88 . More recently, a systematic study of proteome‐wide MS‐CETSA profiling in vivo was reported by the Cellzome group 56 .…”

Currently Available Strategies for Target Identification of Bioactive Natural Products

“…In the same year, Vasaturo’s group used a combination of proteomics, CETSA and classical biochemical techniques to demonstrate that the interaction of oridonin with NCL can effectively modulate the activity of heat shock protein 70 (HSP70) ( Vasaturo et al, 2018 ). In 2019, Anette et al established a multi-group microtubule protein-specific CETSA technique to reveal the anticancer activity of paclitaxel, which binds to the β-microtubulin on the luminal side ( Langeback et al, 2019 ). Guo’s group extracted a derivative from Aspergillus flavus that was selectively toxic to phosphoglycerate dehydrogenase (PHGDH)-dependent cancer cells.…”