13-Acetoxysarcocrassolide Exhibits Cytotoxic Activity Against Oral Cancer Cells Through the Interruption of the Keap1/Nrf2/p62/SQSTM1 Pathway: The Need to Move Beyond Classical Concepts

• Main Authors: Yi-Chang Liu, Bo-Rong Peng, Kai-Cheng Hsu, Mohamed El-Shazly, Shou-Ping Shih, Tony Eight Lin, Fu-Wen Kuo, Yi-Cheng Chou, Hung-Yu Lin, Mei-Chin Lu
• Format: Article
• Language: English
• Published: MDPI AG 2020-07-01
• Series: Marine Drugs
• Subjects: apoptosis; reactive oxygen species; 13-Acetoxysarcocrassolide; oral cancer; oxidative stress; Keap1/Nrf2/p62/SQSTM1
• Online Access: https://www.mdpi.com/1660-3397/18/8/382

13-Acetoxysarcocrassolide (13-AC), a marine cytotoxic product isolated from the alcyonacean coral<i> Lobophytum crassum</i>, exhibited potent antitumor and immunostimulant effects as reported in previous studies. However, the 13-AC antitumor mechanism of action against oral cancer cells remains unclear. The activity of 13-AC against Ca9-22 cancer cells was determined using MTT assay, flow cytometric analysis, immunofluorescence, immunoprecipitation, Western blotting, and siRNA. 13-AC induced apoptosis in oral cancer cells Ca9-22 through the disruption of mitochondrial membrane potential (MMP) and the stimulation of reactive oxygen species (ROS) generation. It increased the expression of apoptosis- and DNA damage-related proteins in a concentration- and time-dependent manner. It exerted potent antitumor effect against oral cancer cells, as demonstrated by the in vivo xenograft animal model. It significantly reduced the tumor volume (55.29%) and tumor weight (90.33%). The pretreatment of Ca9-22 cells with N-acetylcysteine (NAC) inhibited ROS production resulting in the attenuation of the cytotoxic activity of 13-AC. The induction of the Keap1-Nrf2 pathway and the promotion of p62/SQSTM1 were observed in Ca9-22 cells treated with 13-AC. The knockdown of p62 expression by siRNA transfection significantly attenuated the effect of 13-AC on the inhibition of cell viability. Our results indicate that 13-AC exerted its cytotoxic activity through the promotion of ROS generation and the suppression of the antioxidant enzyme activity. The apoptotic effect of 13-AC was found to be mediated through the interruption of the Keap1/Nrf2/p62/SQSTM1 pathway, suggesting its potential future application as an anticancer agent.