The Androgen receptor (AR) transcription pathway is known to be a key biological process in the growth of prostate cancer. To specifically interrupt this biological process without side effects, coactivator binding inhibitors (CBIs) have been developed which are designed to target a unique binding site called activation function 2 (AF-2) which is located in the AR ligand-binding domain (LBD).
In this thesis, research was carried out to synthesize compounds which are able to block this protein-protein interface (PPI) and to develop a binding assay to quantify their effectiveness as blocking agents.
In the synthetic work, to obtain a model compound for use as a positive control during the development of the CBI binding assay, a six-step synthesis of a known pyrimidine-based CBI with a‘FXXLF’ motif was completed. The final product was characterized spectroscopically and shown to be identical to that reported in the literature. In addition, to target another interesting binding site, binding function 3 (BF-3), several analogs of an indoline-indole based lead compound were designed and were successfully synthesized. An investigation into the oxidation of these to give indole-indole control compounds was also conducted. These BF-3 antagonists are ready for screening in the cell-based assay to produce inhibition data.
For the biology part, a luciferase reporter (LR) assay and a fluorescence polarization (FP) assay were performed on both the AF-2 and BF-3 targeted model compounds. These two assays generated good inhibition data against the AR transcriptional activity and binding towards rat AR-LDB, respectively. The optimization of the FP assay was also successfully achieved with respect to the buffer components and the protocol.
Finally, having successfully recorded the activities of the mentioned model compounds in the FP assay, an alpha-helix mimetic CBI which had been designed and synthesized previously in the Spivey research group was tested using the FP assay platform and shown to produce good binding data towards AR. It gave a micromolar level IC50, 18.72 ± 3.94 μM. This promising result provides a strong impetus for further investigation of this type of compound for the modulation of AR activity.