A Novel Potent Anticancer Compound Optimized from a Natural Oridonin Scaffold Induces Apoptosis and Cell Cycle Arrest through the Mitochondrial Pathway

Abstract

The cytotoxicity of the natural ent-kaurene diterpenoid, oridonin, has been extensively studied. However, the application of oridonin for cancer therapy was hampered primarily by its moderate potency. In this study, a series of oridonin A-ring modified analogues, and their derivatives bearing various substituents on 14-OH position, were designed, synthesized, and evaluated for anticancer efficacy. Some of the derivatives were significantly more potent than oridonin against both drug-sensitive and drug-resistant cancer cells. The most potent compound, 13p, was 200-fold more efficacious than oridonin in MCF-7 cancer cells. Furthermore, 13p induced apoptosis and cell cycle arrest at the G2/M phase. A decrease in mitochondrial membrane potential and an increase in Bax/Bcl-2 ratio, accompanied by activated caspase-3 cleavage, were observed in MCF-7 cells after treatment with 13p, suggesting that the mitochondrial pathway was involved in the 13p-mediated apoptosis. Moreover, 13p significantly inhibited tumor growth in mouse xenograft models and had no observable toxic effect.

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“…The IC50 values were calculated according to the dose-dependent curves. All the tests were repeated in at least three independent experiments [38][39][40].…”

“…The collected cells were fixed by adding 70% ethanol at 4 o C overnight and incubated for 30 min in PBS containing 100 μL RNase A and 400 μL of propidium iodide. Analysis of the cell DNA content was performed with the system software (Cell Quest, BD Biosciences, USA) [38][39][40].…”

“…Then 5 μL Annexin V-APC and 5 μL 7-AAD were added successively and the mixture was incubated for 15 min under dark conditions at 25 C. Apoptosis was analyzed using a FACS Calibur flow cytometer (BectoneDickinson, San Jose, CA, USA) [38][39][40].…”

“…The bound antibodies were detected by horseradish peroxidase (HRP) conjugated second antibodies and visualized using an enhanced chemiluminescent reagent. The relative levels of each signaling event to control GAPDH were determined by densimetric scanning [20,29,40].…”

“…The mice were sacrificed, and the tumors were excised and weighed. The inhibition rate was calculated as follows: Tumor inhibitory ratio (%) = (1-average tumor weight of treated group/average tumor weight of control group) × 100% [40,41].…”