Proteasome inhibitor interacts synergistically with autophagy inhibitor to suppress proliferation and induce apoptosis in hepatocellular carcinoma

Cancer. 2012 Nov 15;118(22):5560-71. doi: 10.1002/cncr.27586. Epub 2012 Apr 19.

Abstract

Background: The ubiquitin-proteasome system and autophagy-lysosome system are 2 major protein degradation pathways in eukaryotic cells, which are tightly linked to cancer. Proteasome inhibitors have been approved in clinical use against hematologic malignancies, but their application in solid tumors is uncertain. Moreover, the role of autophagy after proteasome inhibition is controversial.

Methods: Two proteasome inhibitors, 2 autophagy inhibitors, and 3 hepatocellular carcinoma (HCC) cell lines were investigated in the current study. In vitro, cell proliferation was evaluated by 3-(4,5-dimethylthiazol-2-yl)-2,5-diphenyltetrazolium bromide (MTT) assay, cell apoptosis was evaluated by flow cytometry analysis of annexin-V/propidium iodide staining, and autophagy was evaluated by green fluorescent protein-light chain 3 (GFP-LC3) redistribution and LC3 Western blot analysis. In vivo, Ki-67 staining was used to detect cell proliferation, terminal deoxynucleotidyl transferase-mediated deoxyuridine triphosphate nick-end labeling (TUNEL) staining was used to detect apoptosis, and electron microscopy and p62 immunohistochemical staining were used to detect autophagy.

Results: Proteasome inhibitors suppressed proliferation, induced apoptosis, and activated autophagy in HCC cell lines in vitro, and autophagy exerted a protective role after proteasome inhibition. In vivo, anticancer effects of bortezomib on the MHCC-97H orthotopic model (human HCC cells) were different from the effects observed on the Huh-7 subcutaneous model (human HCC cells). The autophagy inhibitor chloroquine interacted synergistically with bortezomib to suppress proliferation and induce apoptosis in both tumor models.

Conclusions: The current results indicated that simultaneous targeting of the proteasome and autophagy pathways may represent a promising method for HCC treatment.

Publication types

  • Research Support, Non-U.S. Gov't

MeSH terms

  • Adenine / analogs & derivatives
  • Adenine / pharmacology
  • Amino Acid Chloromethyl Ketones / pharmacology
  • Antineoplastic Agents / pharmacology
  • Apoptosis Regulatory Proteins / metabolism
  • Apoptosis*
  • Autophagy / drug effects
  • Beclin-1
  • Boronic Acids / pharmacology
  • Bortezomib
  • Carcinoma, Hepatocellular / drug therapy*
  • Carcinoma, Hepatocellular / pathology*
  • Cell Line, Tumor
  • Cell Proliferation / drug effects*
  • Chloroquine / pharmacology
  • Cysteine Proteinase Inhibitors / pharmacology
  • Humans
  • Leupeptins / pharmacology
  • Membrane Proteins / metabolism
  • Microtubule-Associated Proteins / metabolism
  • Proteasome Endopeptidase Complex / metabolism
  • Proteasome Inhibitors / pharmacology*
  • Pyrazines / pharmacology

Substances

  • Amino Acid Chloromethyl Ketones
  • Antineoplastic Agents
  • Apoptosis Regulatory Proteins
  • BECN1 protein, human
  • Beclin-1
  • Boronic Acids
  • Cysteine Proteinase Inhibitors
  • Leupeptins
  • Membrane Proteins
  • Microtubule-Associated Proteins
  • Proteasome Inhibitors
  • Pyrazines
  • benzyloxycarbonylvalyl-alanyl-aspartyl fluoromethyl ketone
  • light chain 3, human
  • 3-methyladenine
  • Bortezomib
  • Chloroquine
  • Proteasome Endopeptidase Complex
  • Adenine
  • benzyloxycarbonylleucyl-leucyl-leucine aldehyde