Infiltrating macrophages are a key component of inflammation during tumorigenesis, but the direct evidence of such linkage remains unclear. We report here that persistent co-culturing of immortalized prostate epithelial cells with macrophages, without adding any carcinogens, induces prostate tumorigenesis, and that induction involves the alteration of signaling of macrophage androgen receptor (AR)-inflammatory chemokine CCL4-STAT3 activation as well as epithelial-to-mesenchymal transition (EMT) and down-regulation of p53/PTEN tumor suppressors. In vivo studies further showed that PTEN+/− mice lacking macrophage AR developed far fewer prostatic intraepithelial neoplasia (PIN) lesions, supporting an in vivo role for macrophage AR during prostate tumorigenesis. CCL4 neutralizing antibody effectively blocked macrophage-induced prostate tumorigenic signaling, and targeting AR via an AR degradation enhancer, ASC-J9®, reduced CCL4 expression and xenografted tumor growth in vivo. Importantly, CCL4 upregulation was associated with increased Snail expression and down-regulation of p53/PTEN in high-grade PIN and prostate cancer. Together, our results identify the AR-CCL4-STAT3 axis as key regulators during prostate tumor initiation and highlight the important roles of infiltrating macrophages and inflammatory cytokines for the prostate tumorigenesis.

Early studies suggested androgen receptor (AR) splice variants might contribute to the progression of prostate cancer (PCa) into castration resistance. However, the therapeutic strategy to target these AR splice variants still remains unresolved. Through tissue survey of tumors from the same patients before and after castration resistance, we found that the expression of AR3, a major AR splice variant that lacks the AR ligand-binding domain, was substantially increased after castration resistance development. The currently used antiandrogen, Casodex, showed little growth suppression in CWR22Rv1 cells. Importantly, we found that AR degradation enhancer ASC-J9 could degrade both full-length (fAR) and AR3 in CWR22Rv1 cells as well as in C4-2 and C81 cells with addition of AR3. The consequences of such degradation of both fAR and AR3 might then result in the inhibition of AR transcriptional activity and cell growth in vitro. More importantly, suppression of AR3 specifically by short-hairpin AR3 or degradation of AR3 by ASC-J9 resulted in suppression of AR transcriptional activity and cell growth in CWR22Rv1-fARKD (fAR knockdown) cells in which DHT failed to induce, suggesting the importance of targeting AR3. Finally, we demonstrated the in vivo therapeutic effects of ASC-J9 by showing the inhibition of PCa growth using the xenografted model of CWR22Rv1 cells orthotopically implanted into castrated nude mice with undetectable serum testosterone. These results suggested that targeting both fAR- and AR3-mediated PCa growth by ASC-J9 may represent the novel therapeutic approach to suppress castration-resistant PCa. Successful clinical trials targeting both fAR and AR3 may help us to battle castration-resistant PCa in the future.