A classical approach to treating prostate cancer uses antagonist ligands - so-called anti-androgens, such as bicalutamide - which block gene transcription through binding to a lipophilic pocket at the ligand binding domain of the androgen receptor (AR). An alternative strategy has been developed using compounds which directly target the surface charge clamp by mimicking the coactivator's highly conserved alpha-helical motif. Thus, to gain additional knowledge about the AR-coactivator interaction, the use of natural miniproteins as a source of novel AR-coactivator inhibitors incorporating the FXXLF motif was explored. Their stable well-defined alpha-helical secondary structures make miniproteins ideal candidates for development into AR-coactivator inhibitors. Therefore, starting from two potent miniprotein scaffold structures, identified from previous work, systematic point mutations aimed at improving AR affinity were introduced using solid-phase peptide synthesis (SPPS). Structure-activity relationship studies were performed, from which a number of high affinity inhibitors, typically in the low micromolar-to-high nanomolar range, with a ten-fold gain in potency compared with the reference compounds, were identified, thus highlighting the high potential for these scaffolds.