Curcumol simultaneously induces both apoptosis and autophagy in human nasopharyngeal carcinoma cells.

Autophagy is usually considered as a protective mechanism against cell death, and in the meantime, leads to cell injury even apoptosis. Apoptosis and autophagy are very closely connected and may cooperate, coexist, or antagonize each other on progressive occurrence of cell death triggered by natural compounds. Therefore, the interplay between the two modes of death is essential for the overall fate of cancer cells. Our previous study revealed that curcumol induced apoptosis in nasopharyngeal carcinoma (NPC) cells. Recently, curcumol was found to induce autophagy in cancer cells. However, whether curcumol can induce NPC cells autophagy and the effects of autophagy on apoptosis remain elusive. In this study, we found that curcumol induced autophagy through AMPK/mTOR pathway in CNE-2 cells. Moreover, inhibiting autophagy by autophagy inhibitor 3-methyladenine (3-MA) or apoptosis inhibitor z-VAD-fmk significantly increased proliferation while attenuated apoptosis and autophagy compared with the curcumol 212 µM group. In contrast, combining curcumol with autophagy agonist rapamycin and apoptosis inducer MG132 synergized the apoptotic and autophagic effect of curcumol. Taken together, our study demonstrates that curcumol promotes autophagy in NPC via AMPK/mTOR pathway, induces autophagy enhances the activity of curcumol in NPC cells; the combination of autophagy inducer and curcumol can be a new therapeutic strategy for NPC.

Curcumol inhibits EBV-positive Nasopharyngeal carcinoma migration and invasion by targeting nucleolin.

Metastasis is a major cause of recurrence and death in patients with EBV-positive Nasopharyngeal carcinoma (NPC). Previous reports documented that curcumol has both anti-cancer and anti-viral effects, but there is little literature systematically addressing the mechanism of curcumol in EBV-positive tumors. Previously we found that nucelolin (NCL) is a target protein of curcumol in CNE2 cells, an EBV-negative NPC, and in this experiment, we reported a critical role for NCL in promoting migration and invasion of C666-1 cells, an EBV-positive NPC, and found that the expression of NCL determined the level of curcumol's efficacy. Mechanistically, NCL interacted with Epstein-Barr Virus Nuclear Antigen 1 (EBNA1) to activate VEGFA/VEGFR1/PI3K/AKT signaling pathway, which in turn promoted NPC cell invasion and metastasis. Moreover, further study showed that the differential expression of NCL and curcumol intervention only had a regulatory effect on the nuclear accumulation of VEGFR1, which strengthened the anti-cancer effect of curcumol mediated through NCL. Our findings indicated that curcumol exerted anti EBV-positive NPC invasion and metastasis by downregulating EBNA1 and inhibiting VEGFA/VEGFR1/PI3K/AKT signaling by targeting NCL, which provides a novel pharmacological basis for curcumol's clinical use in treating patients with EBV-positive NPC.

Curcumol induces cell cycle arrest and apoptosis by inhibiting IGF-1R/PI3K/Akt signaling pathway in human nasopharyngeal carcinoma CNE-2 cells.

Curcumol has been proved to possess antitumor effects in vivo and in vitro in several cancers. Previously, we have found that curcumol induced apoptosis in CNE-2 cells, but its underlying mechanism has not yet been studied well. Recently, our team clarified that curcumol inhibited colorectal cancer cells' growth partially through insulin-like growth factor 1 receptor (IGF-1R) pathway. Given the key importance of IGF-1R pathway in tumorigenesis, we want to explore whether curcumol effects on nasopharyngeal carcinoma (NPC) cells relates to IGF-1R and its downstream pathway inactivation. In this study, we found that curcumol inhibited IGF-1R and p-Akt expression in a dose- and time-dependent way. In addition, it also regulated their downstream GSK-3ß's activity in CNE-2 cells, which further triggering alterations in the expression of cycle- and apoptosis-related molecules, and then leading to G0/G1-phase arrest and apoptosis. Moreover, curcumol's effect on CNE-2 cells was partly eliminated by IGF-1R's agonist IGF-1. In conclusion, our findings indicated that the inhibitory effect of curcumol on proliferation of NPC cells is related to the inhibition of IGF-1R and its downstream PI3K/Akt/GSK-3ß pathway.

Identification and validation nucleolin as a target of curcumol in nasopharyngeal carcinoma cells.

Identification of the specific protein target(s) of a drug is a critical step in unraveling its mechanisms of action (MOA) in many natural products. Curcumol, isolated from well known Chinese medicinal plant Curcuma zedoary, has been shown to possess multiple biological activities. It can inhibit nasopharyngeal carcinoma (NPC) proliferation and induce apoptosis, but its target protein(s) in NPC cells remains unclear. In this study, we employed a mass spectrometry-based chemical proteomics approach reveal the possible protein targets of curcumol in NPC cells. Cellular thermal shift assay (CETSA), molecular docking and cell-based assay was used to validate the binding interactions. Chemical proteomics capturing uncovered that NCL is a target of curcumol in NPC cells, Molecular docking showed that curcumol bound to NCL with an -7.8 kcal/mol binding free energy. Cell function analysis found that curcumol's treatment leads to a degradation of NCL in NPC cells, and it showed slight effects on NP69 cells. In conclusion, our results providing evidences that NCL is a target protein of curcumol. We revealed that the anti-cancer effects of curcumol in NPC cells are mediated, at least in part, by NCL inhibition. SIGNIFICANCE: Many natural products showed high bioactivity, while their mechanisms of action (MOA) are very poor or completely missed. Understanding the MOA of natural drugs can thoroughly exploit their therapeutic potential and minimize their adverse side effects. Identification of the specific protein target(s) of a drug is a critical step in unraveling its MOA. Compound-centric chemical proteomics is a classic chemical proteomics approach which integrates chemical synthesis with cell biology and mass spectrometry (MS) to identify protein targets of natural products determine the drug mechanism of action, describe its toxicity, and figure out the possible cause of off-target. It is an affinity-based chemical proteomics method to identify small molecule-protein interactions through affinity chromatography approach coupled with mass spectrometry, has been conventionally used to identify target proteins and has yielded good results. Curcumol, has shown effective inhibition on Nasopharyngeal Carcinoma (NPC) Cells, interacted with NCL and then initiated the anti-tumor biological effect. This research demonstrated the effectiveness of chemical proteomics approaches in natural drugs molecular target identification, revealing and understanding of the novel mechanism of actions of curcumol is crucial for cancer prevention and treatment in nasopharynx cancer.

Curcumol induces cell cycle arrest in colon cancer cells via reactive oxygen species and Akt/ GSK3ß/cyclin D1 pathway.

ETHNOPHARMACOLOGICAL RELEVANCE: Curcuma kwangsiensis S. G. Lee & C. F. Liang (Guangxi ezhu, in Chinese) belongs to the Zingiberaceae family, has been used as a traditionally Chinese medicine nearly 2000 year. Curcumol is one of the guaiane-type sesquiterpenoid hemiketal isolated from medicine plant Curcuma kwangsiensis S. G. Lee & C. F. Liang, which has been reported possesses anti-cancer effects. Our previous study found that the most contribution to inhibit nasopharyngeal carcinoma cell growth was curcumol. AIM OF THE STUDY: To assess the effect of curcumol on cell cycle arrest against human colon cancer cells (CRC) cells (LoVo and SW480) and explore its mechanism in vitro and in vivo. MATERIALS AND METHODS: Curcumol was dissolved in absolute ethyl alcohol. The concentration of absolute ethyl alcohol in the control group or in experimental samples was always 1/500 (v/v) of the final medium volume. LoVo and SW480 cells were treated with different concentrations of curcumol (0, 53, 106, 212 and 424µM). And then the cell cycle of each group was examined by flow cytometry. The protein levels of PI3K, p-Akt, cyclin D1, cyclin E, CDK2, CDK4 and GSK3ß were determined by Western blot. The mRNA expression of PI3K, Akt, cyclin D1, CDK4, P27, p21, and P16 in the treated cells were analyzed by real-time RT-PCR. In addition, the antitumor activity of curcumol was evaluated in nude mice bearing orthotopic tumor implants. RESULTS: Curcumol induced cell cycle arrest in G1/S phase. RT-qPCR and Western blot data showed that curcumol enhanced the expression of GSK3ß, P27, p21 and P16, and decreased the levels of PI3K, phosphorylated Akt (p-Akt), cyclin D1, CDK4, cyclin E and CDK2. Furthermore, curcumol induced reactive oxygen species (ROS) generation in LoVo cells, and ROS scavenger N-acetylcysteine (NAC) significantly reversed curcumol-induced cell growth inhibition. Besides, curcumol also prevented the growth of human colon cancer cells xenografts in nude mouse, accompanied by the reduction of PI3K, Akt, cyclin D1, CDK4, cycln E and significant increase of GSK3ß. CONCLUSIONS: Curcumol caused cell cycle arrest at the G0/G1 phase by ROS production and Akt/ GSK3ß/cyclin D1 pathways inactivation, indicating the potential of curcumol in the prevention of colon cancer carcinogenesis.

Constituents of Gnetum montanum.

A new stilbene, gnetifolin M (1), was isolated from the lianas of Gnetum montanum, together with seven known compounds, resveratrol (2), gnetol (3), 4', 5,7-trihydroxy-3'-methoxyflavone, beta-sitosterol, daucosterol, ursolic acid, and tetracosanoic acid. The structure of 1 was determined to be 2-(5'-methoxy-3'-hydroxyphenyl)-4-hydroxybenzofuran on the basis of spectroscopic evidence.