Cancer is a widespread disease characterized by unregulated cell growth and evasion of apoptotic stimuli. Most conventional therapies lack cancer cell specificity and induce cell death indirectly through cellular damage in a p53-dependent manner. A direct and selective p53-independent cancer therapy is the application of Tumor necrosis factor-Related Apoptosis-Inducing Ligand (TRAIL). Recombinant human TRAIL (rhTRAIL) can induce apoptosis in a broad range of transformed human cells while showing minimal toxicity towards non-malignant cells. However, some cancers are resistant to rhTRAIL-induced apoptosis as a direct result of overexpressed antiapoptotic proteins. One key driver mediating the expression of such antiapoptotic proteins is Casein Kinase 2 (CK2). CK2 is upregulated in the nucleus of all cancer cells and promotes the expression of inhibitors of apoptosis resulting in resistance to cell death. Therefore, by inhibiting CK2, TRAIL-resistant cells may become more sensitive to TRAIL-induced apoptosis. We utilized cotreatment with Ellagic Acid (EA), a natural CK2 inhibitor, and TRAIL against the malignant melanoma cell line A375, which is resistant to TRAIL. Our in vitro analysis showed that TRAIL and EA, as single-agents, can inhibit cell growth in a dose-dependent manner, and the combination treatment of TRAIL plus EA synergistically enhanced the cytostatic effects of the single-agent treatments. We show that TRAIL reduces the viability of A375 cells; whereas, EA although it can inhibit growth, cannot reduce the viability of A375 as determined by Trypan Blue exclusion assays. In combination treatments, the cytotoxic effects of TRAIL were not enhanced. Annexin-V Apoptosis assays and Western Blot analysis show that TRAIL can induce apoptosis in A375 cells characterized by the formation of Annexin-V positive cells, PARP fragmentation and the activation of caspase 3. Additionally, TRAIL initiated both the extrinsic and intrinsic pathways of apoptosis marked by caspase 8 activation and the mitochondrial release of cytochrome c, respectively. EA treatment alone did not induce apoptosis and did not induce formation of Annexin-V positive cells or PARP fragmentation. Combination treatments show that EA does not potentiate TRAIL-induced apoptosis. In vivo, TRAIL-treated nude mice bearing A375 xenografts exhibited ∼80% tumor growth inhibition and a prolonged survival rate compared to the untreated control group. EA was not able to inhibit the growth of established A375 tumors and did not augment the in vivo antiproliferative effects of TRAIL. These data demonstrate that EA can enhance the cytostatic but not the cytotoxic or proapoptotic effects of TRAIL in vitro. In vivo, TRAIL can inhibit the growth of established tumors as a single-agent but is not augmented by cotreatment with EA.
Citation Format: Katherine Turner, Daniel Lindner, Michael Kalafatis. Effects of TRAIL and ellagic acid on malignant melanoma A375 in vitro and in vivo. [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 2289. doi:10.1158/1538-7445.AM2014-2289