Inhibition of EGFR signaling augments oridonin-induced apoptosis in human laryngeal cancer cells via enhancing oxidative stress coincident with activation of both the intrinsic and extrinsic apoptotic pathways.
Oridonin, a bioactive diterpenoid isolated from Rabdosia rubescens, has been reported to have anti-tumor effects, while the epidermal growth factor receptor (EGFR) signal pathway has been reported to play a vital role in the biological progression of several tumors and to be a target for therapeutic intervention. In this work, we show that inhibition of EGFR with tyrphostin AG1478 enhances oridonin-induced cell death in human laryngeal cancer cells HEp-2, a cell line characterized by EGFR gene amplification. The enhanced apoptotic effect correlates with high expression and activation of Bax, FADD, caspase-8 as well as caspase-3 and decreased protein levels of Bcl(2) and SIRT1, suggesting that both the extrinsic and intrinsic apoptosis pathways are involved in the apoptotic processes. However, treatment with oridonin and AG1478 greatly enhances nuclear translocation of apoptosis inducing factor (AIF) without caspase-9 activation, indicating that the apoptosis occurs via a caspase-9-independent mitochondrial pathway. Here, it is the active form of caspase-8 but not caspase-9 that activates downstream effector caspase-3, resulting in the cleavage of critical cellular proteins and apoptosis. Furthermore, the combined use of AG1478 and oridonin augments the production of reactive oxygen species (ROS). Incubation of cells with N-Acetylcysteine (NAC) attenuates the apoptosis and the mitochondrial membrane potential (Deltapsim) disruption induced by the combination of oridonin and AG1478, which indicates that ROS plays a pivotal role in cell death. In conclusion, targeting EGFR combined with other conventional pro-apoptotic drugs should be a potentially very effective anti-neoplastic therapy for laryngeal cancer.
Kang
Ning
N
China-Japan Research Institute of Medical and Pharmaceutical Sciences, Shenyang Pharmaceutical University, 103 Wenhua Road, Shenyang, 110016, PR China. kangndd@163.com
Zhang
Jing-Hai
JH
Qiu
Feng
F
Tashiro
Shin-Ichi
S
Onodera
Satoshi
S
Ikejima
Takashi
T
D000971:Antineoplastic Combined Chemotherapy Protocols / Q000494:pharmacology*; D017209:Apoptosis / Q000187:drug effects; D051033:Apoptosis Inducing Factor / Q000378:metabolism; D002294:Carcinoma, Squamous Cell / Q000188:drug therapy* / Q000378:metabolism / Q000473:pathology; D045744:Cell Line, Tumor; D045786:Diterpenes, Kaurane / Q000494:pharmacology*; D004357:Drug Synergism; D004791:Enzyme Inhibitors / Q000494:pharmacology; D066246:ErbB Receptors / Q000037:antagonists & inhibitors* / Q000378:metabolism; D053200:Fas-Associated Death Domain Protein / Q000378:metabolism; D006801:Humans; D007822:Laryngeal Neoplasms / Q000188:drug therapy* / Q000378:metabolism / Q000473:pathology; D053078:Membrane Potential, Mitochondrial / Q000187:drug effects; D018384:Oxidative Stress / Q000187:drug effects; D011065:Poly(ADP-ribose) Polymerases / Q000378:metabolism; D019253:Proto-Oncogene Proteins c-bcl-2 / Q000378:metabolism; D011799:Quinazolines; D017382:Reactive Oxygen Species / Q000378:metabolism; D015398:Signal Transduction / Q000187:drug effects; D056564:Sirtuin 1 / Q000037:antagonists & inhibitors / Q000378:metabolism; D020032:Tyrphostins / Q000494:pharmacology*; D051028:bcl-2-Associated X Protein / Q000378:metabolism
C494829:AIFM1 protein, human; D051033:Apoptosis Inducing Factor; C494826:BAX protein, human; D045786:Diterpenes, Kaurane; D004791:Enzyme Inhibitors; C505533:FADD protein, human; D053200:Fas-Associated Death Domain Protein; D019253:Proto-Oncogene Proteins c-bcl-2; D011799:Quinazolines; D017382:Reactive Oxygen Species; D020032:Tyrphostins; D051028:bcl-2-Associated X Protein; C011959:oridonin; C101044:RTKI cpd; D011065:Poly(ADP-ribose) Polymerases; D066246:ErbB Receptors; C447939:SIRT1 protein, human; D056564:Sirtuin 1