The anti-tumor effect of Curcuma phaeocaulis Cyclepeptide in human hepatoma HepG2 cells

Background: The Curcuma phaeocaulis has been used in traditional Chinese medicine to treat liver diseases through thousands of years. Extracts of Curcuma phaeocaulis possess many valuable pharmacological activities including antioxidant, anti-inflammatory and anti-cancer effects, however, the underlying mechanism of its anti-tumour activity is unclear. We therefore set up experiments to isolate Curcuma phaeocaulis peptides (CCPs) from fresh Curcuma phaeocaulis and evaluate the anti-tumour effects of CCPs in human hepatoma cells HepG2 cells. Methods: Curcuma phaeocaulis peptides (CCPs) were isolated from fresh Curcuma phaeocaulis rhizomes, and purified by Reversed Phase-High Performance Liquid Chromatography. Liver cancer cell lines HuH7, HepG2 cells and human liver cell line HL-7702 cells were used throughout the experiment. The HepG2 cells were treated with the CPP-1 (0 μM, 11.4 μM, 57.0 μM and 285 μM) for 24 h and cell apoptosis was evaluated by flow cytometry. Pro-apoptotic factors (eg. Bax, Bid) and anti-apoptotic factors (eg. Bcl-2, Bcl-xL) were invested in cells treated with CCP. Caspase activation was examined by detecting the protein expressions of Caspase 3, Caspase 9, Cleaved caspase 3, Cleaved caspase 9 and Cleaved PARP using western blot. The intracellular ROS level was detected by DCF fluorescence analysis method reported previously. Furthermore, the mitochondrial transmembrane potential (ΔΨm) was assessed using the fluorescent indicator JC-1 Results: CPP-1 [Cycle-(Leu-His-Ile-Trp)] performed most anti-proliferative activity and inhibited the proliferation of HepG2 cells in a concentration and time-dependent manner. CCP-1-induced a significant mitochondrial transmembrane potential (Δψm) collapse, and cytochrome c release. CPP-1 induced HepG2 cell apoptosis accompanied with the Caspases activation, p53/MDM2 status regulation and Bcl-2/Bax ratio down-regulation. Conclusions: Our data support that the cyclepeptides from Curcuma phaeocaulis have significant in vitro anti-proliferative activity in liver cancer HepG2 cells. Our experiment further confirm that the CPP-1 induces apoptosis in HepG2 cells through the Caspase-dependent pathway, in part by regulating the p53/MDM2 status and Bcl-2 family members expression. Our results further indicate that CPP-1 could be a valuable drug candidate for effective human liver cancer therapy