Suppression Of C-Myc Oncogenic Signaling By Mk591, An Inhibitor Of Arachidonate 5-Lipoxygenase

Proceedings: AACR Annual Meeting 2014; April 5-9, 2014; San Diego, CA Background: Prostate cancer cells continuously generate 5(S)-HETE series of metabolites from arachidonic acid via 5-lipoxygenase (5-Lox) activity, and inhibition of 5-Lox blocks production of metabolites and triggers apoptosis both in androgen-sensitive as well as androgen-independent prostate cancer cells. This apoptosis is prevented by 5(S)-HETE and 5-oxoETE, but not by leukotrienes or metabolites of 12-Lox or 15-Lox, which is consistent with the idea that the 5-HETE series of metabolites are essential survival factors for prostate cancer cells. Thus, 5-Lox emerges as a novel molecular target for prostate cancer therapy. However, details of the downstream signaling mechanisms regulated by 5-Lox metabolites in prostate cancer cells are yet to be fully characterized, and the lack of suitable compounds with in vivo efficacy is hampering proper translational development to target 5-Lox. We addressed this problem by treating LNCaP human prostate cancer cells with MK591, a specific, third-generation inhibitor of 5-Lox activity, and analyzing the effects by gene-expression array, RT-PCR, and Western blot. Methods: Cells were treated with MK591 for eight hours and gene expression was analyzed by Illumina HT-12v4 whole genome gene-expression array. Expression of c-Myc was confirmed by RT-PCR and Western blot. Apoptosis was measured by annexin-V binding, PARP-cleavage, and detecting degradation of chromatin-DNA to nucleosomes. Transcriptional activation of c-Myc was analyzed by nuclear localization, DNA-binding, luciferase-reporter assays and expression of c-Myc-target genes. Results: We observed that MK591 strongly inhibits the expression of c-Myc oncogene in prostate cancer cells, as revealed by the gene expression array. Down-regulation of c-Myc by MK591 was confirmed by RT-PCR and Western blot as well as by the reduction in nuclear-localization and DNA-binding activities of c-Myc proteins. Treatment with MK591 dramatically decreased the Myc-driven E-box-luciferase reporter activity in prostate cancer cells which is effectively prevented by 5-oxoETE, a metabolic product of 5-Lox. Moreover, MK591 substantially reduced the expression of c-Myc target genes (e.g., cyclin D1, CDK4, survivin, Aurora kinase, Gemin4 and MCM10), and blocked the matrigel-invasion and soft-agar colony-formation by prostate cancer cells. Interestingly, while MK591 dramatically affected the Myc function in prostate cancer cells (where Myc is over-activated), it did not inhibit the basal c-Myc functions in non-cancer cells, such as human foreskin fibroblasts (HFF) which do not expresses 5-Lox. Conclusion: Our findings indicate that expression and oncogenic-function of c-Myc in prostate cancer cells are severely down-regulated by the 5-Lox inhibitor, MK591, and suggest that MK591 may be useful to treat advanced, aggressive prostate cancers which are frequently associated with over-activation of the c-Myc oncogene. Citation Format: Sivalokanathan Sarveswaran, Debrup Chakraborty, Jagadananda Ghosh. Suppression of c-Myc oncogenic signaling by MK591, an inhibitor of arachidonate 5-lipoxygenase. [abstract]. In: Proceedings of the 105th Annual Meeting of the American Association for Cancer Research; 2014 Apr 5-9; San Diego, CA. Philadelphia (PA): AACR; Cancer Res 2014;74(19 Suppl):Abstract nr 2702. doi:10.1158/1538-7445.AM2014-2702