Abstract
3,5-dihydroxy Q1 -4-ethyl-trans-stilbene (DETS) is a natural stilbene, which was first identified as bioactive bacterial secondary metabolite isolated from Bacillus cereus associated with a rhabditid entomopathogenic nematode. The present study was intended to investigate the antioxidant and anticancer activity of this compound in vitro. Antioxidant activity was investigated by assaying DPPH free radical scavenging, superoxide radical-(O2..) scavenging, hydroxyl radical scavenging and metal chelating activity, which proved that the compound is a powerful antioxidant. The metal chelating activity of DETS was higher than butylated hydroxyanisol (BHA) and gallic acid, two well-known antioxidants. As the molecule exhibited strong antioxidant potential, it was further evaluated for cytotoxic activity toward five cancer cells of various origins. Since the compound has a strong structural similarity with resveratrol (trans- 3,4,5-trihydroxystilbene), a well-studied chemopreventive polyphenolic antioxidant, its anticancer activity was compared with that of resveratrol. Among the five cancer cells studied, the compound showed maximum cytotoxicity toward the human melanoma cell line, [A375, IC50: 24.01 μM] followed by cervical [HeLa-46.17 μM], colon [SW480- 47.28 μM], liver [HepG2- 69.56 μM] and breast [MCF-7- 84.31 μM] cancer cells. A375 was much more sensitive to DETS compared to the non-melanoma cell line, A431, in which the IC50 of the compound was more than double (49.60 μM). In the present study, the anticancer activity of DETS against melanoma was confirmed by various apoptosis assays. We also observed that DETS, like resveratrol, down-regulates the expression status of major molecules contributing to melanoma progression, such as BRAF, β-catenin and Brn-2, all of which converge in MITF-M, the master regulator of melanoma signaling. The regulatory role of MITF-M in DETS-induced cytotoxicity in melanoma cells was confirmed by comparing the cytotoxicity of DETS in A375 cells (IC50-24.01 μM), with that in SK-MEL-2 (IC50-67.6 μM), another melanoma cells which highly over-express MITF-M. The compound arrests the cells at S-G2 transition state of the cell cycle, as resveratrol. Our results indicate that DETS is a powerful antioxidant, having anticancer efficacy comparable with that of resveratrol, and is a potential candidate to be explored by in vivo studies and in-depth mechanistic evaluation. To our knowledge, this is the first report on the antioxidant and anticancer properties of DETS.
Highlights
Microbial secondary metabolites have received considerable attention as they exhibit significant antibiotic and cytotoxic activities (Bérdy, 2012) and are excellent antioxidants (Saurav and Kannabiran, 2012)
Dulbecco’s modified Eagle’s medium (DMEM) was obtained from Life Technologies (Grand Island, NY, USA); Fetal bovine serum (FBS) from PAN Biotech (GmbH, Aidenbach, Germany); Brn-2 (POU domain, class3, transcription factor 2) and MITF-M (Microphthalmia-associated transcription factor) were purchased from Abcam (Cambridge, UK). β-actin and caspases were purchased from Cell Signaling (Beverly, MA, USA) and antibodies against poly ADP-ribose polymerase (PARP) β-catenin, BRAF(serine/threonine-protein kinase B-Raf) and Annexin V apoptosis detection kit was from Santa Cruz Biotechnology (Santa Cruz, CA, USA)
The DETS was found to be a potent scavenger of superoxide radical generated in riboflavin-NBT-light system in vitro
Summary
Microbial secondary metabolites have received considerable attention as they exhibit significant antibiotic and cytotoxic activities (Bérdy, 2012) and are excellent antioxidants (Saurav and Kannabiran, 2012). Several compounds isolated from microbes and plants are rich sources of antioxidants and have been shown to possess excellent anticancer potential (Anto et al, 1995; Abdel-Fattah et al, 2012; Ramasubburayan et al, 2015) Though synthetic antioxidants such as BHA and BHT have been shown to protect the human body from oxidative damage, there has been a great concern regarding their toxicity and carcinogenic side effects (Hwang et al, 2013). Melanoma is often characterized by resistance to cytotoxic drugs that contributes to the high morbidity and mortality rates in patients worldwide This emphasizes the importance of discovering new compounds that are both safe and effective against melanoma. We have conducted a detailed investigation of the antioxidant potential of this compound and have explored in vitro, the relevance of evaluating it as an anticancer agent against malignant melanoma, as our preliminary observations indicate that melanoma cells are highly sensitive to this compound
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