Figure 3 | Scientific Reports

Figure 3

From: Determining direct binders of the Androgen Receptor using a high-throughput Cellular Thermal Shift Assay

Figure 3

CETSA HT detects antagonism of agonist target engagement to derive apparent intracellular Ki. (a) ITDRFCETSA experiments performed for DHT in the presence of increasing concentrations of Enzalutamide. Enzalutamide reverses DHT-mediated thermal stabilisation of AR in a concentration dependent manner, showing competitive antagonism of agonist target engagement. Data shown is the mean ± SD of n = 4 and is representative of five technical repeats, summarised in (c). (b) Data from (a) re-plotted as a Schild plot showing fold change in agonist ITDRFCETSA EC50 at indicated antagonist concentration. Data is representative of five technical repeats. (c) Intracellular AR antagonist Ki determined by CETSA HT for a range of AR antagonists. Values shown are the geometric mean ± SD of ≥3 technical repeats, each n = 4. Representative raw data is reported in Supplementary Fig. S8. (d) Application of CETSA HT to determine intracellular Ki is not significantly affected by the temperature of the heat shock. The plotted data is the mean ± SD of three technical repeats, each n = 4. Representative raw data is reported in Supplementary Fig. S9.