title: Gambogic acid synergistically potentiates cisplatin-induced apoptosis in non-small-cell lung cancer through suppressing NF-κB and MAPK/HO-1 signalling.
abstract: BACKGROUND
  Gambogic acid (GA) has been reported to have potent anticancer activity and is authorised to be tested in phase II clinical trials for treatment of non-small-cell lung cancer (NSCLC). The present study aims to investigate whether GA would be synergistic with cisplatin (CDDP) against the NSCLC.
  
  
  METHODS
  1-(4,5-Dimethylthiazol-2-yl)-3,5-diphenylformazan (MTT), combination index (CI) isobologram, western blot, quantitative PCR, flow cytometry, electrophoretic mobility shift assay, xenograft tumour models and terminal deoxynucleotide transferase-mediated dUTP nick-end labelling analysis were used in this study.
  
  
  RESULTS
  The cell viability results showed that sequential CDDP-GA treatment resulted in a strong synergistic action in A549, NCI-H460, and NCI-H1299 cell lines, whereas the reverse sequence and simultaneous treatments led to a slight synergistic or additive action. Increased sub-G1 phase cells and enhanced PARP cleavage demonstrated that the sequence of CDDP-GA treatment markedly increased apoptosis in comparison with other treatments. Furthermore, the sequential combination could enhance the activation of caspase-3, -8, and 9, increase the expression of Fas and Bax, and decrease the expression of Bcl-2, survivin and X-inhibitor of apoptosis protein (X-IAP) in A549 and NCI-H460 cell lines. In addition, increased apoptosis was correlated with enhanced reactive oxygen species generation. Importantly, it was found that, followed by CDDP treatment, GA could inhibit NF-κB and mitogen-activated protein kinase (MAPK)/heme oxygenase-1 (HO-1) signalling pathways, which have been validated to reduce ROS release and confer CDDP resistance. The roles of NF-κB and MAPK pathways were further confirmed by using specific inhibitors, which significantly increased ROS release and apoptosis induced by the sequential combination of CDDP and GA. Moreover, our results indicated that the combination of CDDP and GA exerted increased antitumour effects on A549 xenograft models through inhibiting NF-κB, HO-1, and subsequently inducing apoptosis.
  
  
  CONCLUSIONS
  Gambogic acid sensitises lung cancer cells to CDDP in vitro and in vivo in NSCLC through inactivation of NF-κB and MAPK/HO-1 signalling pathways, providing a rationale for the combined use of CDDP and GA in lung cancer chemotherapy.
authors.affiliation: 1] Department of Pharmacology, Shenyang Pharmaceutical University, Shenyang 110016, People's Republic of China [2] Jiangsu Kanion Pharmaceutical Co. Ltd, Lianyungang 222001, People's Republic of China.
authors.forename: L-H
authors.initials: LH
authors.lastname: Wang
authors.affiliation: Department of Pharmacology, Shenyang Pharmaceutical University, Shenyang 110016, People's Republic of China.
authors.forename: Y
authors.initials: Y
authors.lastname: Li
authors.affiliation: Department of Pharmacology, Shenyang Pharmaceutical University, Shenyang 110016, People's Republic of China.
authors.forename: S-N
authors.initials: SN
authors.lastname: Yang
authors.affiliation: Department of Pharmacology, Shenyang Pharmaceutical University, Shenyang 110016, People's Republic of China.
authors.forename: F-Y
authors.initials: FY
authors.lastname: Wang
authors.affiliation: Department of Pharmacology, Shenyang Pharmaceutical University, Shenyang 110016, People's Republic of China.
authors.forename: Y
authors.initials: Y
authors.lastname: Hou
authors.affiliation: Department of Pharmacology, Shenyang Pharmaceutical University, Shenyang 110016, People's Republic of China.
authors.forename: W
authors.initials: W
authors.lastname: Cui
authors.affiliation: Department of Pharmacology, Shenyang Pharmaceutical University, Shenyang 110016, People's Republic of China.
authors.forename: K
authors.initials: K
authors.lastname: Chen
authors.affiliation: Department of Pharmacology, Shenyang Pharmaceutical University, Shenyang 110016, People's Republic of China.
authors.forename: Q
authors.initials: Q
authors.lastname: Cao
authors.affiliation: Department of Pharmacology, Shenyang Pharmaceutical University, Shenyang 110016, People's Republic of China.
authors.forename: S
authors.initials: S
authors.lastname: Wang
authors.affiliation: Department of Pharmacology, Shenyang Pharmaceutical University, Shenyang 110016, People's Republic of China.
authors.forename: T-Y
authors.initials: TY
authors.lastname: Zhang
authors.affiliation: Jiangsu Kanion Pharmaceutical Co. Ltd, Lianyungang 222001, People's Republic of China.
authors.forename: Z-Z
authors.initials: ZZ
authors.lastname: Wang
authors.affiliation: Jiangsu Kanion Pharmaceutical Co. Ltd, Lianyungang 222001, People's Republic of China.
authors.forename: W
authors.initials: W
authors.lastname: Xiao
authors.affiliation: Department of Pharmacology, Shenyang Pharmaceutical University, Shenyang 110016, People's Republic of China.
authors.forename: J-Y
authors.initials: JY
authors.lastname: Yang
authors.affiliation: Department of Pharmacology, Shenyang Pharmaceutical University, Shenyang 110016, People's Republic of China.
authors.forename: C-F
authors.initials: CF
authors.lastname: Wu
mesh_terms: D000971:Antineoplastic Combined Chemotherapy Protocols / Q000494:pharmacology; D017209:Apoptosis / Q000187:drug effects / Q000235:genetics; D051017:Apoptosis Regulatory Proteins / Q000235:genetics / Q000378:metabolism; D002289:Carcinoma, Non-Small-Cell Lung / Q000188:drug therapy* / Q000201:enzymology / Q000235:genetics / Q000378:metabolism; D020169:Caspases / Q000235:genetics / Q000378:metabolism; D045744:Cell Line, Tumor; D002470:Cell Survival / Q000187:drug effects / Q000235:genetics; D002945:Cisplatin / Q000008:administration & dosage / Q000494:pharmacology*; D015536:Down-Regulation / Q000187:drug effects; D004357:Drug Synergism; D051547:Heme Oxygenase-1 / Q000037:antagonists & inhibitors* / Q000235:genetics / Q000378:metabolism; D006801:Humans; D008175:Lung Neoplasms / Q000188:drug therapy* / Q000201:enzymology / Q000235:genetics / Q000378:metabolism; D020935:MAP Kinase Signaling System / Q000187:drug effects*; D020928:Mitogen-Activated Protein Kinases / Q000037:antagonists & inhibitors / Q000235:genetics / Q000378:metabolism; D016328:NF-kappa B / Q000037:antagonists & inhibitors* / Q000235:genetics / Q000378:metabolism; D017382:Reactive Oxygen Species / Q000378:metabolism; D015398:Signal Transduction / Q000187:drug effects; D044004:Xanthones / Q000008:administration & dosage / Q000494:pharmacology*
chemical_list: D051017:Apoptosis Regulatory Proteins; D016328:NF-kappa B; D017382:Reactive Oxygen Species; D044004:Xanthones; C052659:gambogic acid; C496541:HMOX1 protein, human; D051547:Heme Oxygenase-1; D020928:Mitogen-Activated Protein Kinases; D020169:Caspases; D002945:Cisplatin