Biochemical mechanism of Caffeic Acid Phenylethyl Ester (CAPE) selective toxicity towards melanoma cell lines

Abstract

In the current work, we investigated the in-vitro biochemical mechanism of caffeic acid phenylethyl ester (CAPE) toxicity and eight hydroxycinnamic/caffeic acid derivatives in-vitro, using tyrosinase enzyme as a molecular target in human SK-MEL-28 melanoma cells. Enzymatic reaction models using tyrosinase/O2 and HRP/H2O2 were used to delineate the role of one- and two-electron oxidation. Ascorbic acid (AA), NADH and GSH depletion were used as markers of quinone formation and oxidative stress in CAPE induced toxicity in melanoma cells. Ethylenediamine, an o-quinone trap, prevented the formation of o-quinone and oxidations of AA and NADH mediated by tyrosinase bioactivation of CAPE. The IC50 of CAPE towards SK-MEL-28 melanoma cells was 15μM. Dicoumarol, a diaphorase inhibitor, and 1-bromoheptane, a GSH depleting agent, increased CAPE’s toxicity towards SK-MEL-28 cells indicating quinone formation played an important role in CAPE induced cell toxicity. Cyclosporin-A and trifluoperazine, inhibitors of the mitochondrial membrane permeability transition pore (PTP), prevented CAPE toxicity towards melanoma cells. We further investigated the role of tyrosinase in CAPE toxicity in the presence of a shRNA plasmid, targeting tyrosinase mRNA. Results from tyrosinase shRNA experiments showed that CAPE led to negligible anti-proliferative effect, apoptotic cell death and ROS formation in shRNA plasmid treated cells. Furthermore, it was also found that CAPE selectively caused escalation in the ROS formation and intracellular GSH (ICG) depletion in melanocytic human SK-MEL-28 cells which express functional tyrosinase. In contrast, CAPE did not lead to ROS formation and ICG depletion in amelanotic C32 melanoma cells, which do not express functional tyrosinase. These findings suggest that tyrosinase plays a major role in CAPE’s selective toxicity towards melanocytic melanoma cell lines. Our findings suggest that the mechanisms of CAPE toxicity in SK-MEL-28 melanoma cells mediated by tyrosinase bioactivation of CAPE included quinone formation, ROS formation, intracellular GSH depletion and induced mitochondrial toxicity.

Phenylpropanoids in radioregulation: double edged sword

“…In addition, CAPE (2) showed a radiosensitization effect on A549 lung cancer cells, which suggested the possibility that treatment with CAPE (2) can result in enhancement of local control of lung cancer by radiotherapy without normal lung damage in vivo (Chen et al, 2004). Other studies have demonstrated that CAPE (2) not only showed itself as a strong activator of ROS generation possibly due to GSH depletion and reduction of mitochondrial membrane potential, but also had a radiosensitizing capability for IR co-treatment through inhibition of IR-induced NF-κB activation, resulting in sensitivity to IR and enhanced IR-induced apoptosis in several cancer cells (Chen et al, 2005;Lee et al, 2008;Kudugunti et al, 2010).…”

Existence of G-quadruplex structures in promoter region of oncogenes confirmed by G-quadruplex DNA cross-linking strategy

“…A gradual dosage-dependent inhibition of cell proliferation was observed upon 6 days treatment with compound 1 . Then DNA damage by compound 1 in the B16-F1 cells was characterized through alkaline comet assay3536. For negative control experiment, HeLa cells were used and nearly no DNA damage was observed in the even treated with higher concentrations of compound 1 (SI Figure S16a–f), which supported the tyrosinase-stimulated cross-linking mechanism in vivo .…”

The Ethanol Extract of Polish Propolis Exhibits Anti-Proliferative and/or Pro-Apoptotic Effect on HCT 116 Colon Cancer and Me45 Malignant Melanoma Cells In Vitro Conditions.

“…The usefulness of propolis in therapy of neoplastic diseases is connected predominantly with its ability to induce apoptosis and anti-proliferative activities. In numerous in vitro research, propolis exhibited proapoptotic activities on various types of neoplastic cells such as in the cases of: laryngeal cancer, lung carcinoma, pancreatic cancer, thyroid neoplasm, colorectal cancer, breast cancer, prostate cancer and malignant glioma [24,25].…”