Most conventional cancer therapies are only modestly effective against CRPCa. Therefore, the development of new and effective therapies to treat early-stage prostate cancer (PCa) to prevent progression of PCa to CRPCa is warranted. Our collaborator Dr. Wilding and his coworkers'basic science investigation of probable mechanism(s) of PCa progression led to a class of natural cancer causative agents, cellular reactive oxygen species (ROS) such as superoxide, hydroxyl radical, hydrogen peroxide, etc. Their published and strong preliminary data show that in human PCa cells, excess cellular ROS activate the transcription factor NF-kappa B that prevents apoptosis and drives PCa cell proliferation in the absence of androgen. It is proposed that targeted inhibitors o PCa specific ROS generation pathway(s) will be more effective than are chemical and/or dietary anti-oxidants in preventing PCa recurrence and progression to CRPCa. Our published data show androgen induces spermidine/spermine acetyl transferase (SSAT) that initiates a spermidine and spermine oxidation pathway to generate copious amounts of ROS specifically in PCa cells that are naturally rich in spermine and spermidine. As JunD protein expression is induced by androgen specifically in PCa cells, it is hypothesized that JunD complexes with the activated androgen receptor (AR) to induce SSAT gene expression and ROS production in PCa cells. We and others have also shown that NF-kappa B can induce SSAT expression, thus setting up a feed-forward loop for SSAT activation, ROS production and PCa progression. Therefore, it is proposed that specific inhibitor(s) of JunD-AR complex should block SSAT induction, ROS production, NF-kappa B activation and stop the autocrine feed-forward loop that helps CRPCa progression. A high-throughput screen using Gaussia luciferase enzyme reconstitution assay for in situ protein-protein interaction, thus far, identified seven inhibitorsthat can block JunD-AR interaction. Two of these compounds blocked androgen-induced ROS generation in androgen-dependent PCa cells and showed growth inhibitory effects against both androgen- dependent and androgen-independent human PCa cells at sub-micromolar to low micromolar levels. Using computer-aided molecular docking to determine the binding site(s) of the inhibitor, we propose to design analogs of the two lead compounds that should more efficiently block androgen-induced ROS generation and cell growth.
Our Specific Aims are: 1) To develop models for small molecule interaction with JunD- AR complex using in silico docking and design and synthesize analogs of small molecule inhibitors of JunD- AR interaction;2) To identify promising agents from their relative abilities to disrupt JunD-AR interaction and their efficacies in reducing intracellular ROS and inhibit cultured human PCa cell growth.;3) To identify the lead drug candidate from the studies in Aim 2 and test its ability to inhibit human PCa growth in nude mouse xenografts and the same in a transgenic mouse model developing spontaneous PCa.
Advanced hormone refractory metastatic prostate cancer (CRPCa) is the second leading cause of cancer deaths among US men. Most conventional cancer therapies are only modestly effective against CRPCa. Therefore, the development of new and effective therapies to prevent prostate cancer (PCa) recurrence and/or progression to CRPCa is warranted. Our collaborator Dr. Wilding and his coworkers'investigation of probable mechanism(s) of PCa recurrence and progression led to a class of natural cancer causative agents, cellular reactive oxygen species (ROS). ROS levels are generally high in PCa cells as compared to their normal counterpart. Male hormone testosterone induces copious amounts of ROS generation specifically in PCa cells. Recent discoveries show that the main source of the prostatic ROS is oxidation of spermine and spermidine that is found in very high level in the prostate. It has also been demonstrated that activated androgen receptor binds with a protein JunD to induce the spermine/spermidine oxidation. Therefore, it is proposed that specific inhibitor(s) of JunD-AR complex should inhibit spermine/spermidine oxidation and should thus block ROS production in PCa cells and prevent CRPCa proliferation. We used a high throughput screen to identify inhibitors of JunD-AR interaction and identified several compounds that can inhibit JunD-AR interaction. One of these compounds blocked androgen-induced ROS generation in androgen-dependent PCa cells and showed growth inhibitory effects against both androgen-dependent and androgen-independent human PCa cells. Colby has patented these compounds for preclinical and clinical development for treatment of early-stage progressing PCa patients. Using computer-aided molecular design to determine the binding site(s) of the inhibitor, here, we propose to develop new analogs of the lead compound that should more efficiently block androgen-induced ROS generation and cell growth. The most active agent, thus identified, can be further developed for preclinical and clinical testing to prevent progression of early-stage androgen-dependent PCa to CRPCa.
