Long-term ethanol exposure causes human liver cancer cells to become resistant to mitomycin C treatment through the inactivation of bad-mediated apoptosis.
The aim of this study was to test whether long-term ethanol consumption confers therapeutic resistance to human liver cancer patients infected with hepatitis B virus (HBV). Chronic ethanol-treated cells were established by consecutively culturing a human hepatocellular carcinoma cell line, Hep 3B, which contains integrated HBV sequences, for 20-40 passages with or without 10 mM ethanol (designated as E20-E40 and C20-C40, respectively). Flow cytometry analysis demonstrated that a growth promoting effect of long-term ethanol treatment was induced in the E40 cells through preferential acceleration of S-phase in these cells. Lower protein expression levels of p16, p21/Cip1, and p27/Kip1 were detected in the ethanol-treated E40 cells. We further demonstrated that long-term ethanol-treated E40 cells develop drug resistance in response to mitomycin C (MMC) treatment (>8 microM). Immunoblot analysis revealed that caspase-8-mediated mitochondrial apoptotic signals (such as Bad) were inactivated in the MMC-resistant E40 cells. Immunoprecipitation experiments demonstrated that the sequestration of phosphorylated Bad (Ser-112) through its binding with 14-3-3 was detected more profoundly in the MMC-resistant E40 cells. Next, we examined the therapeutic efficacy of MMC (10 mg MMC/kg body weight, three times per week) in severe combined immunodeficient (SCID) mice bearing E40- and C40-xenografted tumors. Significant reductions (>3-fold) in tumor growth were detected in MMC-treated C40-xenografted mice. In vivo and in vitro studies demonstrated that AKT- and extracellular signal-regulated kinase (ERK)-mediated survival factors inhibited the Bad-induced mitochondrial apoptotic signals that were involved in E40 tumor cells and that conferred resistance to MMC.
Huang
Ching-Shui
CS
Department of Surgery, Cathay General Hospital, and Graduate Institute of Medical Sciences, Taipei Medical University, Taipei, Taiwan.
Lee
Yi-Ru
YR
Chen
Ching-Shyang
CS
Tu
Shih-Hsin
SH
Wang
Ying-Jan
YJ
Lee
Chia-Hwa
CH
Chen
Li-Ching
LC
Chang
Hui-Wen
HW
Chang
Chien-Hsi
CH
Chih-Ming
Su
S
Wu
Chih-Hsiung
CH
Ho
Yuan-Soon
YS
D017209:Apoptosis / Q000187:drug effects; D053181:Caspase 8 / Q000378:metabolism; D000431:Ethanol / Q000378:metabolism / Q000494:pharmacology; D005434:Flow Cytometry; D006801:Humans; D008113:Liver Neoplasms / Q000188:drug therapy / Q000378:metabolism; D008928:Mitochondria / Q000378:metabolism; D016685:Mitomycin / Q000378:metabolism / Q000494:pharmacology* / Q000627:therapeutic use*; D013267:Stilbenes
D013267:Stilbenes; C006711:benzoylamido-4'-aminostilbene-2,2'-disulfonate; D000431:Ethanol; D016685:Mitomycin; D053181:Caspase 8