Autophagy-related cell death by pan-histone deacetylase inhibition in liver cancer
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Pietro Di Fazio1, Petra Waldegger2, Samir Jabari3, Susanne Lingelbach4, Roberta Montalbano1, Matthias Ocker5,8, Emily P. Slater1, Detlef K. Bartsch1, Romana Illig6, Daniel Neureiter6, Thaddeus T. Wissniowski7
1Department of Visceral, Thoracic and Vascular Surgery, Philipps University of Marburg, Marburg, Germany
2Institute for Biomedical Aging Research, University of Innsbruck, Rennweg, Innsbruck, Austria
3Institute for Anatomy I, University of Erlangen-Nurnberg, Erlangen, Germany
4Department of Urology, Philipps University of Marburg, Marburg, Germany
5Institute for Surgical Research, Philipps University of Marburg, Marburg, Germany
6Institute of Pathology, Paracelsus Medical University/Salzburger Landeskliniken (SALK), Salzburg, Austria
7Department of Gastroenterology and Endocrinology, Philipps University of Marburg, Marburg, Germany
8Experimental Medicine Oncology, Bayer Pharma AG, Berlin Germany
Pietro Di Fazio, email: email@example.com
Keywords: autophagic cell death, liver cancer, pan-deacetylase inhibitor, cancer therapy, autophagosomes
Received: September 24, 2015 Accepted: March 18, 2016 Published: April 05, 2016
Autophagy is a homeostatic, catabolic degradation process and cell fate essential regulatory mechanism. Protracted autophagy triggers cell death; its aberrant function is responsible for several malignancies. Panobinostat, a potent pan-deacetylase inhibitor, causes endoplasmic reticulum stress-induced cell death. The aim of this study was to investigate the role of autophagy in deacetylase inhibitor-triggered liver cancer cell death.
HepG2 (p53wt) and Hep3B (p53 null) liver cancer cell lines were exposed to panobinostat. RT-qPCR and western blot confirmed autophagic factor modulation. Immuno-fluorescence, -precipitation and -histochemistry as well as transmission electron microscopy verified autophagosome formation. The cytotoxicity of panobinostat and autophagy modulators was detected using a real time cell viability assay.
Panobinostat induced autophagy-related factor expression and aggregation. Map1LC3B and Beclin1 were significantly over-expressed in HepG2 xenografts in nude mice treated with panobinostat for 4 weeks. Subcellular distribution of Beclin1 increased with the appearance of autophagosomes-like aggregates. Cytosolic loss of p53, in HepG2, and p73, in Hep3B cells, and a corresponding gain of their nuclear level, together with modulation of DRAM1, were observed. Autophagosome aggregation was visible after 6 h of treatment. Treatment of cells stably expressing GFP-RFPtag Map1LC3B resulted in aggregation and a fluorescence switch, thus confirming autophagosome formation and maturation. Tamoxifen, an inducer of autophagy, caused only a block in cell proliferation; but in combination with panobinostat it resulted in cell death.
Autophagy triggers cell demise in liver cancer. Its modulation by the combination of tamoxifen and panobinostat could be a new option for palliative treatment of hepatocellular carcinoma.
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