In vitro activity: UT-155 is a potent and selective androgen receptor (AR) degraders (SARD) that markedly reduce the activity of wild-type and splice variant isoforms of AR at submicromolar doses. Three SARDs (UT-69, UT-155, and (R)-UT-155) bind the amino-terminal transcriptional activation domain AF-1, which has not been targeted for degradation previously, with two of these SARD (UT-69 and UT-155) also binding the carboxy-terminal ligand binding domain. Despite different mechanisms of action, all three SARDs degraded wild-type AR and inhibited AR function, exhibiting greater inhibitory potency than the approved AR antagonists. Collectively, our results introduce a new candidate class of next-generation therapeutics to manage advanced prostate cancer. Androgen receptor (AR) mediates the growth of prostate cancer throughout its course of development, including in abnormal splice variants (AR-SV)-driven advanced stage castration-resistant disease. AR stabilization by androgens makes it distinct from other steroid receptors, which are typically ubiquitinated and degraded by proteasomes after ligand binding. Thus, targeting AR in advanced prostate cancer requires the development of agents that can sustainably degrade variant isoforms for effective therapy.

Kinase Assay: UT-155 is a selective androgen receptor (AR) antagonist, with a Ki of 267 nM for AR-RBD. UT-69 and UT-155 both compete for binding to the LBD and also reduce AR protein levels at 24 hours comparable to the observed decrease in transcriptional activity. To determine whether the reduction in expression was required to inhibit AR activity or whether the competitive displacement of androgen from the LBD is sufficient to inhibit transcriptional activity, we evaluated the effect of the SARDs on the pre-mRNA of NDRG1 and MT2A genes that are rapidly induced by hormones.

Cell Assay: Cells were plated at varying densities and in different serum-containing medium depending on the cell line in 96-well plates, treated as indicated in the figures, and viability measured using sulforhodamine B (SRB) or cell-titer glo assay (Promega, Madison, WI).