Hepatocellular Carcinoma: Molecular Pathogenesis and Therapeutic Advances.

Hepatocellular carcinoma (HCC), the most common form of liver cancer, continues to be a serious medical problem with poor prognosis, without major therapeutic improvement for years and increasing incidence. Fortunately, advances in systemic treatment options are finally arriving for HCC patients. After a decade of sorafenib as a standard therapy for advanced HCC, several tyrosine kinase inhibitors (TKIs), antiangiogenic antibodies, and immune checkpoint inhibitors have reached the clinic. Although infections by hepatitis B virus and hepatitis C virus remain principal factors for HCC development, the rise of non- alcoholic steatohepatitis from diabetes mellitus or metabolic syndrome is impeding HCC decline. Knowledge of specific molecular mechanisms, based on the etiology and the HCC microenvironment that influence tumor growth and immune control, will be crucial for physician decision-making among a variety of drugs to prescribe. In addition, markers of treatment efficacy are needed to speed the movement of patients towards other potentially effective treatments. Consequently, research to provide scientific data for the evidence-based management of liver cancer is guaranteed in the coming years and discussed here.

Antioxidants Threaten Multikinase Inhibitor Efficacy against Liver Cancer by Blocking Mitochondrial Reactive Oxygen Species.

Sorafenib and regorafenib, multikinase inhibitors (MKIs) used as standard chemotherapeutic agents for hepatocellular carcinoma (HCC), generate reactive oxygen species (ROS) during cancer treatment. Antioxidant supplements are becoming popular additions to our diet, particularly glutathione derivatives and mitochondrial-directed compounds. To address their possible interference during HCC chemotherapy, we analyzed the effect of common antioxidants using hepatoma cell lines and tumor spheroids. In liver cancer cell lines, sorafenib and regorafenib induced mitochondrial ROS production and potent cell death after glutathione depletion. In contrast, cabozantinib only exhibited oxidative cell death in specific HCC cell lines. After sorafenib and regorafenib administration, antioxidants such as glutathione methyl ester and the superoxide scavenger MnTBAP decreased cell death and ROS production, precluding the MKI activity against hepatoma cells. Interestingly, sorafenib-induced mitochondrial damage caused PINK/Parkin-dependent mitophagy stimulation, altered by increased ROS production. Finally, in sorafenib-treated tumor spheroids, while ROS induction reduced tumor growth, antioxidant treatments favored tumor development. In conclusion, the anti-tumor activity of specific MKIs, such as regorafenib and sorafenib, is altered by the cellular redox status, suggesting that uncontrolled antioxidant intake during HCC treatment should be avoided or only endorsed to diminish chemotherapy-induced side effects, always under medical scrutiny.

Mitochondrial Glutathione: Recent Insights and Role in Disease.

Mitochondria are the main source of reactive oxygen species (ROS), most of them deriving from the mitochondrial respiratory chain. Among the numerous enzymatic and non-enzymatic antioxidant systems present in mitochondria, mitochondrial glutathione (mGSH) emerges as the main line of defense for maintaining the appropriate mitochondrial redox environment. mGSH's ability to act directly or as a co-factor in reactions catalyzed by other mitochondrial enzymes makes its presence essential to avoid or to repair oxidative modifications that can lead to mitochondrial dysfunction and subsequently to cell death. Since mitochondrial redox disorders play a central part in many diseases, harboring optimal levels of mGSH is vitally important. In this review, we will highlight the participation of mGSH as a contributor to disease progression in pathologies as diverse as Alzheimer's disease, alcoholic and non-alcoholic steatohepatitis, or diabetic nephropathy. Furthermore, the involvement of mitochondrial ROS in the signaling of new prescribed drugs and in other pathologies (or in other unmet medical needs, such as gender differences or coronavirus disease of 2019 (COVID-19) treatment) is still being revealed; guaranteeing that research on mGSH will be an interesting topic for years to come.

Regorafenib Alteration of the BCL-xL/MCL-1 Ratio Provides a Therapeutic Opportunity for BH3-Mimetics in Hepatocellular Carcinoma Models.

BACKGROUND: The multikinase inhibitor regorafenib, approved as second-line treatment for hepatocellular carcinoma (HCC) after sorafenib failure, may induce mitochondrial damage. BH3-mimetics, inhibitors of specific BCL-2 proteins, are valuable drugs in cancer therapy to amplify mitochondrial-dependent cell death. METHODS: In in vitro and in vivo HCC models, we tested regorafenib's effect on the BCL-2 network and the efficacy of BH3-mimetics on HCC treatment. RESULTS: In hepatoma cell lines and Hep3B liver spheroids, regorafenib cytotoxicity was potentiated by BCL-xL siRNA transfection or pharmacological inhibition (A-1331852), while BCL-2 antagonism had no effect. Mitochondrial outer membrane permeabilization, cytochrome c release, and caspase-3 activation mediated A-1331852/regorafenib-induced cell death. In a patient-derived xenograft (PDX) HCC model, BCL-xL inhibition stimulated regorafenib activity, drastically decreasing tumor growth. Moreover, regorafenib-resistant HepG2 cells displayed increased BCL-xL and reduced MCL-1 expression, while A-1331852 reinstated regorafenib efficacy in vitro and in a xenograft mouse model. Interestingly, BCL-xL levels, associated with poor prognosis in liver and colorectal cancer, and the BCL-xL/MCL-1 ratio were detected as being increased in HCC patients. CONCLUSION: Regorafenib primes tumor cells to BH3-mimetic-induced cell death, allowing BCL-xL inhibition with A-1331852 or other strategies based on BCL-xL degradation to enhance regorafenib efficacy, offering a novel approach for HCC treatment, particularly for tumors with an elevated BCL-xL/MCL-1 ratio.

A Functional Role of GAS6/TAM in Nonalcoholic Steatohepatitis Progression Implicates AXL as Therapeutic Target.

BACKGROUND AND AIMS: GAS6 signaling, through the TAM receptor tyrosine kinases AXL and MERTK, participates in chronic liver pathologies. Here, we addressed GAS6/TAM involvement in Non-Alcoholic SteatoHepatitis (NASH) development. METHODS: GAS6/TAM signaling was analyzed in cultured primary hepatocytes, hepatic stellate cells (HSC) and Kupffer cells (KCs). Axl-/-, Mertk-/- and wild-type C57BL/6 mice were fed with Chow, High Fat Choline-Deficient Methionine-Restricted (HFD) or methionine-choline-deficient (MCD) diet. HSC activation, liver inflammation and cytokine/chemokine production were measured by qPCR, mRNA Array analysis, western blotting and ELISA. GAS6, soluble AXL (sAXL) and MERTK (sMERTK) levels were analyzed in control individuals, steatotic and NASH patients. RESULTS: In primary mouse cultures, GAS6 or MERTK activation protected primary hepatocytes against lipid toxicity via AKT/STAT-3 signaling, while bemcentinib (small molecule AXL inhibitor BGB324) blocked AXL-induced fibrogenesis in primary HSCs and cytokine production in LPS-treated KCs. Accordingly; bemcentinib diminished liver inflammation and fibrosis in MCD- and HFD-fed mice. Upregulation of AXL and ADAM10/ADAM17 metalloproteinases increased sAXL in HFD-fed mice. Transcriptome profiling revealed major reduction in fibrotic- and inflammatory-related genes in HFD-fed mice after bemcentinib administration. HFD-fed Mertk-/- mice exhibited enhanced NASH, while Axl-/- mice were partially protected. In human serum, sAXL levels augmented even at initial stages, whereas GAS6 and sMERTK increased only in cirrhotic NASH patients. In agreement, sAXL increased in HFD-fed mice before fibrosis establishment, while bemcentinib prevented liver fibrosis/inflammation in early NASH. CONCLUSION: AXL signaling, increased in NASH patients, promotes fibrosis in HSCs and inflammation in KCs, while GAS6 protects cultured hepatocytes against lipotoxicity via MERTK. Bemcentinib, by blocking AXL signaling and increasing GAS6 levels, reduces experimental NASH, revealing AXL as an effective therapeutic target for clinical practice.

Antiapoptotic BCL-2 proteins determine sorafenib/regorafenib resistance and BH3-mimetic efficacy in hepatocellular carcinoma.

Sorafenib, systemic treatment for advanced hepatocellular carcinoma (HCC), and regorafenib, novel second line treatment after sorafenib failure, have efficacy limited by evasive mechanisms of acquired-drug resistance. BCL-2 proteins participate in the response to tyrosine kinase inhibitors; however, their role in HCC therapy with sorafenib/regorafenib remains uncertain. BH3-mimetic ABT-263 (navitoclax) enhanced sorafenib activity, inducing cell death via a mitochondrial caspase-dependent mechanism, after BCL-xL/BCL-2 inhibition. Sorafenib-resistant hepatoma cells (HepG2R and Hep3BR) exhibited altered mRNA expression of BCL-2 and other anti-apoptotic family members, such as MCL-1, priming drug-resistant cancer cells to death by BH3-mimetics. ABT-263 restored sorafenib efficacy in sorafenib-resistant cell lines and HCC mouse models. Moreover, in mice xenografts from patient-derived BCLC9 cells, better tumor response to sorafenib was associated to higher changes in the BCL-2 mRNA pattern. HCC non-treated patients displayed altered BCL-2, MCL-1 and BCL-xL mRNA levels respect to adjacent non-tumoral biopsies and an increased BCL-2/MCL-1 ratio, predictive of navitoclax efficacy. Moreover, regorafenib administration also modified the BCL-2/MCL-1 ratio and navitoclax sensitized hepatoma cells to regorafenib by a mitochondrial caspase-dependent mechanism. In conclusion, sorafenib/regorafenib response is determined by BCL-2 proteins, while increased BCL-2/MCL-1 ratio in HCC sensitizes drug resistant-tumors against ABT-263 co-administration. Thus, changes in the BCL-2 profile, altered in HCC patients, could help to follow-up sorafenib efficacy, allowing patient selection for combined therapy with BH3-mimetics or early switch them to second line therapy.