Disulfiram eradicates tumor-initiating hepatocellular carcinoma cells in ROS-p38 MAPK pathway-dependent and -independent manners.
Tumor-initiating cells (TICs) play a central role in tumor development, metastasis, and recurrence. In the present study, we investigated the effect of disulfiram (DSF), an inhibitor of aldehyde dehydrogenase, toward tumor-initiating hepatocellular carcinoma (HCC) cells. DSF treatment suppressed the anchorage-independent sphere formation of both HCC cells. Flow cytometric analyses showed that DSF but not 5-fluorouracil (5-FU) drastically reduces the number of tumor-initiating HCC cells. The sphere formation assays of epithelial cell adhesion molecule (EpCAM)(+) HCC cells co-treated with p38-specific inhibitor revealed that DSF suppresses self-renewal capability mainly through the activation of reactive oxygen species (ROS)-p38 MAPK pathway. Microarray experiments also revealed the enrichment of the gene set involved in p38 MAPK signaling in EpCAM(+) cells treated with DSF but not 5-FU. In addition, DSF appeared to downregulate Glypican 3 (GPC3) in a manner independent of ROS-p38 MAPK pathway. GPC3 was co-expressed with EpCAM in HCC cell lines and primary HCC cells and GPC3-knockdown reduced the number of EpCAM(+) cells by compromising their self-renewal capability and inducing the apoptosis. These results indicate that DSF impaired the tumorigenicity of tumor-initiating HCC cells through activation of ROS-p38 pathway and in part through the downregulation of GPC3. DSF might be a promising therapeutic agent for the eradication of tumor-initiating HCC cells.
Department of Gastroenterology and Nephrology, Graduate School of Medicine, Chiba University, Chiba, Japan ; Department of Cellular and Molecular Medicine, Graduate School of Medicine, Chiba University, Chiba, Japan.
Tetsuhiro
T
Chiba
Department of Gastroenterology and Nephrology, Graduate School of Medicine, Chiba University, Chiba, Japan ; Department of Cellular and Molecular Medicine, Graduate School of Medicine, Chiba University, Chiba, Japan.
Eiichiro
E
Suzuki
Department of Gastroenterology and Nephrology, Graduate School of Medicine, Chiba University, Chiba, Japan ; Department of Cellular and Molecular Medicine, Graduate School of Medicine, Chiba University, Chiba, Japan.
Kaori
K
Yuki
Institute of Liver Studies, King's College Hospital, Denmark Hill, London, United Kingdom.
Yoh
Y
Zen
Department of Cellular and Molecular Medicine, Graduate School of Medicine, Chiba University, Chiba, Japan.
Motohiko
M
Oshima
Department of Cellular and Molecular Medicine, Graduate School of Medicine, Chiba University, Chiba, Japan.
Satoru
S
Miyagi
Department of Cellular and Molecular Medicine, Graduate School of Medicine, Chiba University, Chiba, Japan.
Atsunori
A
Saraya
Department of Cellular and Molecular Medicine, Graduate School of Medicine, Chiba University, Chiba, Japan.
Shuhei
S
Koide
Department of Gastroenterology and Nephrology, Graduate School of Medicine, Chiba University, Chiba, Japan.
Tenyu
T
Motoyama
Department of Gastroenterology and Nephrology, Graduate School of Medicine, Chiba University, Chiba, Japan.
Sadahisa
S
Ogasawara
Department of Gastroenterology and Nephrology, Graduate School of Medicine, Chiba University, Chiba, Japan.
Yoshihiko
Y
Ooka
Department of Gastroenterology and Nephrology, Graduate School of Medicine, Chiba University, Chiba, Japan.
Akinobu
A
Tawada
Division of Cancer Immunotherapy, Research Center for Innovative Oncology, National Cancer Center Hospital East, Kashiwa, Japan.
Tetsuya
T
Nakatsura
Department of Gastroenterology, Graduate School of Medicine, Kanazawa University, Kanazawa, Japan.
Takehiro
T
Hayashi
Department of Gastroenterology, Graduate School of Medicine, Kanazawa University, Kanazawa, Japan.
Taro
T
Yamashita
Department of Gastroenterology, Graduate School of Medicine, Kanazawa University, Kanazawa, Japan.
Syuichi
S
Kaneko
Department of General Surgery, Graduate School of Medicine, Chiba University, Chiba, Japan.
Masaru
M
Miyazaki
Department of Cellular and Molecular Medicine, Graduate School of Medicine, Chiba University, Chiba, Japan.
Atsushi
A
Iwama
Department of Gastroenterology and Nephrology, Graduate School of Medicine, Chiba University, Chiba, Japan.
Osamu
O
Yokosuka
D000444:Aldehyde Dehydrogenase / Q000378:metabolism; D000818:Animals; D000951:Antigens, Neoplasm / Q000378:metabolism; D063646:Carcinogenesis / Q000187:drug effects / Q000473:pathology; D006528:Carcinoma, Hepatocellular / Q000188:drug therapy / Q000201:enzymology* / Q000235:genetics / Q000473:pathology; D015815:Cell Adhesion Molecules / Q000378:metabolism; D002452:Cell Count; D045744:Cell Line, Tumor; D049109:Cell Proliferation / Q000187:drug effects; D004221:Disulfiram / Q000494:pharmacology* / Q000627:therapeutic use; D004789:Enzyme Activation / Q000187:drug effects; D000071858:Epithelial Cell Adhesion Molecule; D005434:Flow Cytometry; D020869:Gene Expression Profiling; D015972:Gene Expression Regulation, Neoplastic / Q000187:drug effects; D053673:Glypicans / Q000235:genetics / Q000378:metabolism; D042541:Intracellular Space / Q000187:drug effects / Q000378:metabolism; D008113:Liver Neoplasms / Q000188:drug therapy / Q000201:enzymology* / Q000235:genetics / Q000473:pathology; D020935:MAP Kinase Signaling System / Q000187:drug effects*; D051379:Mice; D016513:Mice, SCID; D014411:Neoplastic Stem Cells / Q000187:drug effects / Q000201:enzymology / Q000473:pathology*; D017382:Reactive Oxygen Species / Q000378:metabolism*; D018874:Spheroids, Cellular / Q000187:drug effects / Q000473:pathology; D023041:Xenograft Model Antitumor Assays; D048051:p38 Mitogen-Activated Protein Kinases / Q000378:metabolism*
D000951:Antigens, Neoplasm; D015815:Cell Adhesion Molecules; D000071858:Epithelial Cell Adhesion Molecule; D053673:Glypicans; D017382:Reactive Oxygen Species; D000444:Aldehyde Dehydrogenase; D048051:p38 Mitogen-Activated Protein Kinases; D004221:Disulfiram