The role of thioredoxin reductase and glutathione reductase in plumbagin-induced, reactive oxygen species-mediated apoptosis in cancer cell lines.
Plumbagin is a secondary metabolite that was first identified in the Plumbago genus of plants. It is a naphthoquinone compound with anti-atherosclerosis, anticancer, anti-inflammatory, antimicrobial, contraceptive, cardiotonic, immunosuppressive, and neuroprotective activities. However, the mechanisms of plumbagin's activities are largely unknown. In this study, we examined the effect of plumbagin on HepG2 hepatocellular carcinoma cells as well as LLC lung cancer cells, SiHa cervical carcinoma cells. Plumbagin significantly decreased HepG2 cell viability in a dose-dependent manner. Additionally, treatment with plumbagin significantly increased the Bax/Bcl-2 ratio and caspase-3/7 activity. Using the similarity ensemble approach (SEA)-a state-of-the-art cheminformatic technique-we identified two previously unknown cellular targets of plumbagin: thioredoxin reductase (TrxR) and glutathione reductase (GR). This was then confirmed using protein- and cell-based assays. We found that plumbagin was directly reduced by TrxR, and that this reduction was inhibited by the TrxR inhibitor, sodium aurothiomalate (ATM). Plumbagin also decreased the activity of GR. Plumbagin, and the GR inhibitor sodium arsenite all increased intracellular reactive oxygen species (ROS) levels and this increase was significantly attenuated by pretreatment with the ROS scavenger N-acetyl-cysteine (NAC) in HepG2 cells. Plumbagin increased TrxR-1 and heme oxygenase (HO)-1 expression and pretreatment with NAC significantly attenuated the plumbagin-induced increase of TrxR-1 and HO-1 expression in HepG2 cells, LLC cells and SiHa cells. Pretreatment with NAC significantly prevented the plumbagin-induced decrease in cell viability in these cell types. In conclusion, plumbagin exerted its anticancer effect by directly interacting with TrxR and GR, and thus increasing intracellular ROS levels.
College of Pharmacy, Research Institute of Pharmaceutical Sciences, Kyungpook National University, Daegu, Republic of Korea.
Geun Hye
GH
Hwang
Department of Veterinary Physiology, College of Veterinary Medicine, Seoul National University, Seoul, Republic of Korea.
Jung Min
JM
Ryu
College of Pharmacy, Research Institute of Pharmaceutical Sciences, Kyungpook National University, Daegu, Republic of Korea.
Yu Jin
YJ
Jeon
College of Pharmacy, Research Institute of Pharmaceutical Sciences, Kyungpook National University, Daegu, Republic of Korea.
Joonhyeok
J
Choi
Department of Veterinary Physiology, College of Veterinary Medicine, Seoul National University, Seoul, Republic of Korea.
Ho Jae
HJ
Han
College of Pharmacy, Research Institute of Pharmaceutical Sciences, Kyungpook National University, Daegu, Republic of Korea.
You-Mie
YM
Lee
College of Pharmacy, Research Institute of Pharmaceutical Sciences, Kyungpook National University, Daegu, Republic of Korea.
Sangkyu
S
Lee
College of Pharmacy, Research Institute of Pharmaceutical Sciences, Kyungpook National University, Daegu, Republic of Korea.
Jong-Sup
JS
Bae
College of Pharmacy, Research Institute of Pharmaceutical Sciences, Kyungpook National University, Daegu, Republic of Korea.
Jong-Wha
JW
Jung
Department of Biology Education, College of Education, Pusan National University, Busan, Republic of Korea.
Woochul
W
Chang
Department of Immunobiology, Yale University School of Medicine, New Haven, CT, USA.
Lark Kyun
LK
Kim
College of Pharmacy, Research Institute of Pharmaceutical Sciences, Kyungpook National University, Daegu, Republic of Korea. Electronic address: jjee@knu.ac.kr.
Jun-Goo
JG
Jee
College of Pharmacy, Research Institute of Pharmaceutical Sciences, Kyungpook National University, Daegu, Republic of Korea. Electronic address: vetmedic@knu.ac.kr.
Min Young
MY
Lee
D000972:Antineoplastic Agents, Phytogenic / Q000737:chemistry / Q000494:pharmacology*; D017209:Apoptosis / Q000187:drug effects / Q000502:physiology*; D045744:Cell Line, Tumor; D004305:Dose-Response Relationship, Drug; D005980:Glutathione Reductase / Q000502:physiology*; D056945:Hep G2 Cells; D006801:Humans; D009285:Naphthoquinones / Q000737:chemistry / Q000494:pharmacology*; D017382:Reactive Oxygen Species / Q000378:metabolism*; D013880:Thioredoxin-Disulfide Reductase / Q000502:physiology*
D000972:Antineoplastic Agents, Phytogenic; D009285:Naphthoquinones; D017382:Reactive Oxygen Species; D005980:Glutathione Reductase; D013880:Thioredoxin-Disulfide Reductase; C014758:plumbagin
Anticancer; Glutathione reductase; Plumbagin; Reactive oxygen species; Thioredoxin reductase