Artepillin C (3,5-diprenyl-4-hydroxycinnamic acid) sensitizes LNCaP prostate cancer cells to TRAIL-induced apoptosis

Naturally occurring phenolic compounds have been shown to sensitize prostatecancer cells to tumour necrosis factor-related apoptosis-inducing ligand (TRAIL)-inducedapoptosis. TRAIL is a potent stimulator of apoptosis in cancer cells and an importantimmune effector molecule in the surveillance and elimination of developing tumours.However, many cancer cells are resistant to TRAIL-mediated death. In this study,we aimed to determine the mechanisms by which TRAIL resistance can be overcomein prostate cancer cells by 3,5-diprenyl-4-hydroxycinnamic acid (artepillin C).Artepillin C is a bioactive component of Brazilian green propolis that possessesantitumour and chemopreventive activities. TRAIL-resistant LNCaP prostate cancercells were treated with TRAIL and artepillin C. Cytotoxicity was measured by MTTand lactate dehydrogenase (LDH) assays. Apoptosis was detected using Annexin V-FITCstaining by flow cytometry and fluorescence microscopy. Death receptor (DR) (TRAIL-R1/DR4and TRAIL-R2/DR5) expression was analyzed using flow cytometry. Mitochondrialmembrane potential (∆ψm) was evaluated using DePsipher staining by fluorescencemicro-scopy. The inhibition of NF-κB (p65) activation was confirmed with the ELISA-basedTransAM NF-κB kit. Caspase-8 and caspase-3 activities were determined by colorimetricprotease assays. The results showed that artepillin C sensitized the TRAIL-resistantLNCaP cells by engaging the extrinsic (receptor-mediated) and intrinsic (mitochondrial)apoptotic pathways. Artepillin C increased the expression of TRAIL-R2 and decreasedthe activity of NF-κB. Co-treatment with TRAIL and artepillin C induced the significantactivation of caspase-8 and caspase-3, as well as the disruption of ∆ψm. Thesefindings show that prostate cancer cells can be sensitized to TRAIL-mediated immunopreventionby artepillin C and confirm the role of phenolic compounds in prostate cancerimmunochemoprevention.