Combined treatment with denbinobin and Fas ligand has a synergistic cytotoxic effect in human pancreatic adenocarcinoma BxPC-3 cells.
OBJECTIVE
Human pancreatic carcinoma is a highly malignant cancer. Previous studies have shown that the decoy receptor 3 (DcR3) for Fas ligand (FasL) plays significant roles in tumour progression and immune suppression. In the present study, we evaluated the anti-cancer activity of a natural compound, denbinobin (5-hydroxy-3,7-dimethoxy-1,4-phenanthraquinone), through decreasing DcR3 levels in human pancreatic adenocarcinoma cell lines.


METHODS
We used immunoprecipitation and ELISA assays to examine DcR3 levels, and used FACS to determine the percentage of cells with a sub-G1 DNA content.


RESULTS
AsPC-1 and BxPC-3 human pancreatic cancer cells express high levels of DcR3. Denbinobin concentration-dependently decreased DcR3 levels in BxPC-3 cells. MTT and flow cytometry assays indicated that BxPC-3 was FasL-resistant because high concentrations (100 ng.mL(-1)) of soluble FasL did not inhibit cell growth. However, combinations of denbinobin (3 micromol.L(-1)) with lower concentrations of soluble FasL (10, 30 and 50 ng.mL(-1)) or membrane-bound FasL, were synergistic on cell growth inhibition and apoptosis. Exogenous excess DcR3 reversed this synergistic effect. We observed no significant increase in the levels of surface Fas, cleaved forms of caspase-8, -3, -9, Bax, Bid, Bcl-xL, cytochrome c or mitochondrial membrane potentials following denbinobin treatment. However, denbinobin treatment increased the levels of apoptosis-inducing factor.


CONCLUSIONS
Denbinobin and FasL trigger a synergistic cytotoxic effect in human pancreatic adenocarcinoma cells. Denbinobin mediated a decrease in levels of DcR3, which played a major role in this synergistic effect, and also increased caspase-independent apoptosis, via apoptosis-inducing factor.
Yang
C R
CR
School of Pharmacy, College of Medicine, National Taiwan University, Taipei, Taiwan. cryang@ntu.edu.tw
Guh
J H
JH
Teng
C M
CM
Chen
C C
CC
Chen
P H
PH
D000230:Adenocarcinoma / Q000378:metabolism / Q000473:pathology*; D000880:Anthraquinones / Q000494:pharmacology*; D000972:Antineoplastic Agents, Phytogenic / Q000494:pharmacology*; D017209:Apoptosis; D051033:Apoptosis Inducing Factor / Q000378:metabolism; D045744:Cell Line, Tumor; D002470:Cell Survival / Q000187:drug effects; D004357:Drug Synergism; D053222:Fas Ligand Protein / Q000494:pharmacology*; D006801:Humans; D010190:Pancreatic Neoplasms / Q000378:metabolism / Q000473:pathology*; D010616:Phenanthrenes / Q000494:pharmacology*; D053323:Receptors, Tumor Necrosis Factor, Member 6b / Q000096:biosynthesis
D000880:Anthraquinones; D000972:Antineoplastic Agents, Phytogenic; D051033:Apoptosis Inducing Factor; D053222:Fas Ligand Protein; D010616:Phenanthrenes; D053323:Receptors, Tumor Necrosis Factor, Member 6b; C505911:TNFRSF6B protein, human; C436544:denbinobin