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Peroxynitrite is involved in the mitochondrial dysfunction induced by Sorafenib in liver cancer cells
Patricia de la Cruz-Ojeda1, Elena Navarro-Villarán1, Marina Fuertes-Agudo2
1Institute of Biomedicine of Seville (IBiS), Hospital University "Virgen Del Rocío"/CSIC/University of Seville, Seville, Spain; Department of Medical Physiology and Biophysics, University of Seville, Seville, Spain; Biomedical Research Center for Hepatic and Digestive Diseases (CIBERehd), Madrid, Spain.
Background:
Sorafenib is a tyrosine kinase inhibitor (TKI) that belongs to the landscape of treatments for advanced stages of hepatocellular carcinoma (HCC). The induction of cell death and cell cycle arrest by Sorafenib has been associated with mitochondrial dysfunction in liver cancer cells. Our research aim was to decipher underlying oxidative and nitrosative stress induced by Sorafenib leading to mitochondrial dysfunction in liver cancer cells.
Methods:
MnTBAP, catalase and the scavenger of peroxynitrite FeTPPs were administered to Sorafenib (0-10 μM)-treated HepG2 cells. Oxygen consumption and glycolytic flux were determined in cultured cells. Mitochondrial complex activities were measured in mitochondrial fraction and cell lysates. The protein and mRNA expression of subunits of electron transport chain (ETC) were assessed by immunoblot and RNA-seq.
Results:
Sorafenib (10 μM) increased nitric oxide (NO) and superoxide anion (O2.-) leading to peroxynitrite generation, and drastically reduced oxygen consumption. Moreover, Sorafenib led to mitochondrial network disorganization and loss of membrane potential. The administration of FeTPPs influenced the recovery of mitochondrial network and oxygen consumption, as well as associated ATP production. Sorafenib downregulated the mRNA expression of all mitochondrial-encoded subunits of ETC and, at to a lesser extent, nuclear-encoded mitochondrial genes. The protein expression of complex I, complex III and complex IV was greatly affected by Sorafenib. Furthermore, Sorafenib diminished the activity of complex I in in-gel assays, whose expression and activity were restored by FeTPPs. However, Sorafenib did not affect the assembly of mitochondrial supercomplexes. Sorafenib altered glycolysis and reduced Krebs cycle intermediates and increased NAD/NADH ratio.
Conclusions:
The induction of cell death by Sorafenib was associated with peroxynitrite generation, which impacted the expression of ETC subunits and mitochondrial functionality in liver cancer cells.
Insights
Sorafenib causes liver cancer cell death by increasing oxidative stress and damaging mitochondria. Peroxynitrite generation disrupts electron transport chain function, impacting cell survival.
Area of Science:
- Biochemistry
- Cell Biology
- Oncology
Background:
- Sorafenib is a tyrosine kinase inhibitor (TKI) used for advanced hepatocellular carcinoma (HCC).
- Sorafenib-induced cell death is linked to mitochondrial dysfunction in liver cancer cells.
- Oxidative and nitrosative stress are implicated in Sorafenib's effects on mitochondria.
Purpose of the Study:
- To investigate the role of oxidative and nitrosative stress in Sorafenib-induced mitochondrial dysfunction in liver cancer cells.
- To elucidate the impact of Sorafenib on mitochondrial respiration and the electron transport chain (ETC).
Main Methods:
- HepG2 cells were treated with Sorafenib and agents like MnTBAP, catalase, and FeTPPs.
- Oxygen consumption, glycolytic flux, and mitochondrial complex activities were measured.
- Protein and mRNA expression of ETC subunits were analyzed using immunoblot and RNA-seq.
Main Results:
- Sorafenib (10 μM) elevated nitric oxide and superoxide, generating peroxynitrite, and decreased oxygen consumption.
- Mitochondrial network disorganization and loss of membrane potential were observed.
- FeTPPs treatment partially restored mitochondrial function and ATP production.
- Sorafenib downregulated mitochondrial and nuclear-encoded ETC subunits, affecting complex I, III, and IV protein expression and complex I activity.
- Glycolysis was altered, Krebs cycle intermediates reduced, and NAD/NADH ratio increased.
Conclusions:
- Sorafenib-induced cell death in liver cancer is associated with peroxynitrite generation.
- Peroxynitrite impacts ETC subunit expression and mitochondrial functionality.
- Targeting peroxynitrite may offer therapeutic strategies for Sorafenib-treated HCC.
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