Chemopreventive effect of a dual COX/LOX inhibitor licofelone in prostate cancer: Potential off-target activities

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Increased interest in blocking both cyclooxygenase (COX) and lipoxygenase (LOX) pathways by interfering with the production of both prostaglandins and the biosynthesis of leukotrienes (LTs) is emerging as a promising strategy for cancer chemoprevention. Licofelone, which specifically targets COX/LOX pathways, is under phase III clinical trials for treatment of osteoarthritis with promising initial results, has a broad spectrum of anticancer activities. Chemopreventive and therapeutic potentials of licofelone in the cancers of the colon and breast are reported in preclinical studies. In our earlier studies using cell culture models, we have demonstrated for the first time that licofelone significantly inhibited COX-2 and 5-LOX expression and prevented prostate cancer cell growth. However, in view of the inconsistency on the levels of COX-2 expression in prostate cancer, questions raised as to whether licofelone exerts anti-tumor effects via off-target activities. The detailed molecular mechanism underlying the anticancer effects of licofelone is yet to be investigated. In order to determine whether licofelone induces anti-tumor effects mediating potential off-target activities, we have carried out cell-based assays, including real-time PCR, immunofluorescence, and Western blot analysis using human (LNCaP, PC-3), rat (I-26) and mouse (59R) prostate cancer cells. Prostate cancer cells were treated with licofelone (10μM) up to 48 h to determine cell viability and induction of apoptosis. Using RNAi-mediated gene silencing techniques, we have downregulated the COX-2 and 5-LOX expression in prostate cancer cells in order to pharmacologically characterize the off-target effects of licofelone and to specifically determine whether it has impacted AKT and AR signaling pathways which contribute to prostate cancer progression. Our findings revealed that licofelone (10μM) inhibited cancer cell growth by enhancing apoptosis in both androgen-dependent and -independent prostate cancer cells. In addition, licofelone also induced apoptosis through activation of Apaf-1 and caspase-3. Semi-quantification revealed the appearance of early apoptotic cells (65%) at 48 h. Further, licofelone reduced the level of AKT and AR proteins. Interestingly, licofelone also inhibited PGE2/EP receptors, MMP proteins and vasculoendothelial growth factor (VEGF) (P<0.005). An in-depth analysis on the molecular targets indicated that licofelone has a remarkable effect on PPARγ activation. These results clearly demonstrated that licofelone has the ability to inhibit prostate cancer cell growth via effective apoptosis inducing mechanisms independent of COX/LOX pathways. Overall, our preliminary findings indicate that licofelone has multifaceted modes of action that facilitated prostate cancer cell growth inhibition (Supported partly by NIH/NCI CA106296-01, and CA107813-01 grants).