Cookies help us deliver our services. By using our services, you agree to our use of cookies. More information

Muntane 2017 MITOEAGLE Obergurgl

From Bioblast
Jordi Muntane
Integration of oxidative and nitrosative stress in the overall cell death signaling induced by Sorafenib in hepatoma cells.

Link: MitoEAGLE

Muntane J, Gonzalez R, Rodriguez-Hernandez MA, Navarro-Villaran E, Barcena JA, Cuadrado A, Padilla CA, Padillo FJ (2017)

Event: MitoEAGLE Obergurgl 2017

COST Action MitoEAGLE

Hepatocarcinoma is the third most common cause of death by cancer. Sorafenib is the uniquely accepted molecular targeted drug for the treatment of patients in the advanced stage. The present study evaluated the regulation of endoplasmic reticulum stress (ERS) signaling, autophagy, Nrf2 signaling, oxidative stress and cell death induced by Sorafenib in HCC cell lines.

HCC cell lines with different p53 genetic profile were used (HepG2, wild type; Hep3B, non-expression; and Huh7, mutated p53). We determined the effect of Sorafenib (0-100 μM) on different markers of ERS, Nrf2 signaling, oxidative/nitrosative stress, autophagy, cell proliferation and apoptosis. Also the role of ERS, JNK and autophagy was determined by different experimental interventions in Sorafenib-induced apoptosis. The therapeutic effect of Sorafenib was also assessed in vivo in tumors developed by subcutaneous implantation of HCC cells in athymic mice.

Sorafenib (10-100 μM) induced early (3-12 hours) mitochondrial hyperpolarization, ERS characterized by an increase of Ser51eIFα/eIFα, CHOP, IRE1α and sXBP1, but a decrease of ATF6 expression, overall temporally associated with the increase of the Thr183JNK/JNK and autophagy markers (Beclin, LC3I/LC3II and p62), reduction of Mcl-1 and Bcl-2, Ser473Akt/Akt, Thr308Akt/Akt and Ser2481mTOR. The process was followed by a reduction of the autophagic flux, with increase of Bim, Bak, caspase-3 activity and TUNEL, as well as reduction of cell proliferation (24 hours). The reduction of PERK and IRE1α signaling by siRNA downregulated autophagy and increased caspase-3 activity even at low Sorafenib concentration (10 nM) in HepG2. The inhibition of autophagy by 3-methyladenine and chloroquine potentiated Sorafenib-induced caspase-3 and cell proliferation inhibition in HepG2. The inhibition of Mcl-1 by A-1210477 increased Sorafenib-induced caspase-3. The inhibition of JNK by SP600125 potentiated the inhibition of cell proliferation, but not caspase-3, induced by Sorafenib in HepG2.

Sorafenib reduced dose-dependently nitric oxide, O2.- and H2O2 generation, and the overall pattern of S-nitrosylated, carbonylated and tyrosine nitrated proteins in Sorafenib-treated HepG2 cells. The posttranslational modification of cell death receptors altered apoptosis. In this sense, Sorafenib (10 µM) reduced S-nitrosylation of cell death receptors (CD95, TNF-R1 and Trail-R1) that correlated to a shift from caspase-8- to caspase-3-related apoptosis. Nrf2 signaling was also altered by the treatment. Sorafenib reduced luciferase activity in control, as well as VEGF- and PDGF-stimulated ARE-Luc-transfected HepG2 cells. This effect correlated to mRNA reduced expression of Nrf2-related genes (thioredoxin-1, thioredoxin reductase, hemoxigenase-1, NQO1, glutathione cysteine ligase and glutathione peroxidase). The reduction of Nrf2 signaling was related to an increased ratio of Ser9GSK3β/Tyr216GSK3β. The in vivo study confirmed the antitumoral properties (reduction tumor size, and pro-apoptotic, anti-proliferative, anti-angiogenic and anti-fibrotic effects) of Sorafenib in xenograft mice model.

The study showed that the early induction of ERS was related to the transient autophagy activation in Sorafenib-treated HepG2 cells. The inhibition of PERK and IRE1 signaling by siRNA reduced autophagy and increased apoptosis in Sorafenib-treated HepG2 cells. The later autophagy downregulation was associated with a reduction of cell proliferation and increased apoptosis in Sorafenib-treated HepG2 cells. Sorafenib reduced oxidative and nitrosative intracellular stress, S-nitrosylation of cell death receptors and caspase-8-related activity. Sorafenib reduced Nrf2-dependent signaling through regulation of GSK3β activity.


Bioblast editor: Kandolf G


Labels: MiParea: Pharmacology;toxicology  Pathology: Cancer  Stress:Cell death, Oxidative stress;RONS  Organism: Human  Tissue;cell: Liver 




Event: A1, Oral 


Affiliations

Muntané J(1,5), González R(1), Rodríguez-Hernández MA(1), Navarro-Villarán E(1), Bárcena JA(2), Cuadrado A(3), Padilla CA(2), Padillo FJ(4,5)
  1. Inst Biomedicine Seville (IBIS). IBiS/“Virgen del Rocío” Univ Hospital/CSIC/Univ Seville
  2. Dept Biochem Molecular Biol, Inst Maimonides Biomedical Research Córdoba (IMIBIC), Univ Córdoba
  3. Inst Biomedical Research “Alberto Sols”, Madrid
  4. Dept General Surgery. “Virgen del Rocío” Univ Hospital/IBiS/CSIC/Univ Seville
  5. CENTRO DE INVESTIGACIÓN BIOMÉDICA EN RED de Enfermedades Hepáticas y Digestivas (CIBERehd)
Spain.- [email protected]