The processing of intracellular reactive oxygen species (ROS) by nuclear factor erythroid‐derived 2‐like 2 (Nrf2) and NADPH quinone oxidoreductase 1 (Nqo1) is important for tumor metastasis. However, ...the clinical and biological significance of Nrf2/Nqo1 expression in hepatocellular carcinoma (HCC) remains unclear. We aimed to clarify the clinical importance of Nrf2/Nqo1 expression in HCC and evaluate the association of Nrf2/Nqo1 expression with HCC metastasis. We also evaluated the impact of Nqo1 modulation on HCC metastatic potential. We used spheroids derived from HCC cell lines. In anchorage‐independent culture, HCC cells showed increased ROS, leading to the upregulation of Nrf2/Nqo1. Futile stimulation of Nqo1 by β‐lapachone induces excessive oxidative stress and dramatically increased anoikis sensitivity, finally diminishing the spheroid formation ability, which was far stronger than depletion of Nqo1. We analyzed 117 cases of primary HCC who underwent curative resection. Overexpression of Nrf2/Nqo1 in primary HCC was associated with tumor size, high α‐fetoprotein, and des‐γ‐carboxy‐prothrombin levels. Overexpression of Nrf2/Nqo1 was also associated with multiple intrahepatic recurrences (P = .0073) and was an independent risk factor for poor prognosis (P = .0031). NADPH quinone oxidoreductase 1 plays an important role in anchorage‐independent survival, which is essential for survival for circulation and distant metastasis of HCC cells. These results suggest that targeting Nqo1 activity could be a potential strategy for HCC adjuvant therapy.
Downregulation of NADPH quinone oxidoreductase 1 (Nqo1) in suspended culture promoted intracellular reactive oxygen species production and induced anoikis, resulting in attenuated spheroid formation. Activation of Nqo1 by β‐lapachone showed a similar effect on anoikis sensitivity and diminished spheroid formation. Overexpression of Nrf2/Nqo1 was associated with intrahepatic recurrence and was an independent risk factor for poor prognosis.
The tyrosine kinase inhibitor lenvatinib is used to treat advanced hepatocellular carcinoma (HCC). Ferroptosis is a type of cell death characterized by the iron‐dependent accumulation of lethal lipid ...reactive oxygen species (ROS). Nuclear factor erythroid‐derived 2‐like 2 (Nrf2) protects HCC cells against ferroptosis. However, the mechanism of lenvatinib‐induced cytotoxicity and the relationships between lenvatinib resistance and Nrf2 are unclear. Thus, we investigated the relationship between lenvatinib and ferroptosis and clarified the involvement of Nrf2 in lenvatinib‐induced cytotoxicity. Cell viability, lipid ROS levels, and protein expression were measured using Hep3B and HuH7 cells treated with lenvatinib or erastin. We examined these variables after silencing fibroblast growth factor receptor‐4 (FGFR4) or Nrf2 and overexpressing‐Nrf2. We immunohistochemically evaluated FGFR4 expression in recurrent lesions after resection and clarified the relationship between FGFR4 expression and lenvatinib efficacy. Lenvatinib suppressed system Xc− (xCT) and glutathione peroxidase 4 (GPX4) expression. Inhibition of the cystine import activity of xCT and GPX4 resulted in the accumulation of lipid ROS. Silencing‐FGFR4 suppressed xCT and GPX4 expression and increased lipid ROS levels. Nrf2‐silenced HCC cells displayed sensitivity to lenvatinib and high lipid ROS levels. In contrast, Nrf2‐overexpressing HCC cells displayed resistance to lenvatinib and low lipid ROS levels. The efficacy of lenvatinib was significantly lower in recurrent HCC lesions with low‐FGFR4 expression than in those with high‐FGFR4 expression. Patients with FGFR4‐positive HCC displayed significantly longer progression‐free survival than those with FGFR4‐negative HCC. Lenvatinib induced ferroptosis by inhibiting FGFR4. Nrf2 is involved in the sensitivity of HCC to lenvatinib.
Regulation of ferroptosis by fibroblast growth factor receptor 4 (FGFR4) and nuclear factor erythroid‐derived 2‐like 2 (Nrf2) in hepatocellular carcinoma GPX4, glutathione peroxidase 4.
Abstract
Comprehensive analysis of clinical samples has recently identified molecular and immunological classification of hepatocellular carcinoma (HCC), and the
CTNNB1
(β-catenin)-mutated subtype ...exhibits distinctive characteristics of immunosuppressive tumor microenvironment. For clarifying the molecular mechanisms, we first established human and mouse HCC cells with exon 3 skipping of β-catenin, which promoted nuclear translocation and activated the Wnt/β-catenin signaling pathway, by using newly developed multiplex CRISPR/Cas9-based genome engineering system. Gene set enrichment analysis indicated downregulation of immune-associated gene sets in the HCC cells with activated β-catenin signaling. Comparative analysis of gene expression profiles between HCC cells harboring wild-type and exon 3 skipping β-catenin elucidated that the expression levels of four cytokines were commonly decreased in human and mouse β-catenin-mutated HCC cells. Public exome and transcriptome data of 373 human HCC samples showed significant downregulation of two candidate cytokine genes,
CCL20
and
CXCL2
, in HCC tumors with β-catenin hotspot mutations. T cell killing assays and immunohistochemical analysis of grafted tumor tissues demonstrated that the mouse
Ctnnb1
Δex3
HCC cells evaded immunosurveillance. Taken together, this study discovered that cytokine controlled by β-catenin signaling activation could contribute to immune evasion, and provided novel insights into cancer immunotherapy for the β-catenin-mutated HCC subtype.
The development of hepatocellular carcinoma (HCC) is a multistep process with a complex interaction of various genetic backgrounds and the tumor microenvironment. In addition to the development of ...rational approaches to epidemiologic research, early detection, and diagnosis, considerable progress has been made in systemic treatment with molecular‐targeted agents for patients with advanced HCC. Moreover, encouraging reports of recent clinical trials of combination therapy with immune‐checkpoint inhibitors (ICIs) has raised high hopes. Each HCC is the result of a unique combination of somatic alterations, including genetic, epigenetic, transcriptomic, and metabolic events, leading to conclusive tumoral heterogeneity. Recent advances in comprehensive genetic analysis have accelerated molecular classification and defined subtypes with specific characteristics, including immune‐associated molecular profiles reflecting the immune reactivity in the tumor. In considering the development of therapeutic strategies in combination with immunotherapy, proper interpretation of molecular pathological characterization could lead to effective therapeutic deployment and enable individualization of the management of HCC. Here, we review distinctive molecular alterations in the subtype classification of HCC, current therapies, and representative clinical trials with alternative immune‐combination approaches from a molecular pathological point.
Highlight
Advances in molecular pathology have shifted the paradigm of hepatocellular carcinoma treatment. Comprehensive gene stratification analysis advances our understanding of molecular pathology, ultimately leading to precision medicine. Kabashima and colleagues reviewed our current knowledge from a perspective of molecular pathology, including the characterization of sub‐classifications, molecular targeted therapies and immunological therapies.
Aim
The present study investigated the effect of sarcopenia on short‐ and long‐term surgical outcomes and identified potential prognostic factors for hepatocellular carcinoma (HCC) following ...hepatectomy among patients 70 years of age and older.
Methods
Patient data were retrospectively collected for 296 consecutive patients who underwent hepatectomy for HCC with curative intent. Patients were assigned to two groups according to age (younger than 70 years, and 70 years and older), and the presence of sarcopenia. The clinicopathological, surgical outcome, and long‐term survival data were analyzed.
Results
Sarcopenia was present in 112 of 296 (37.8%) patients with HCC, and 35% of patients aged 70 years and older. Elderly patients had significantly lower serum albumin levels, prognostic nutrition index, percentage of liver cirrhosis, and histological intrahepatic metastasis compared with patients younger than 70 years. Overall survival and disease‐free survival rates in patients with sarcopenia correlated with significantly poor prognosis in the group aged 70 years and older. Multivariate analysis revealed that sarcopenia was predictive of an unfavorable prognosis.
Conclusion
This retrospective analysis revealed that sarcopenia was predictive of worse overall survival and recurrence‐free survival after hepatectomy in patients 70 years of age and older with HCC.
Approximately 2% of healthy persons are infected with human pegivirus (HPgV). HPgV is transmitted via vertical, sexual, and blood‐borne routes. Recently, the association of HPgV infection with the ...risk of lymphoma was reported. Here, we examined the prevalence of chronic HPgV infection in liver transplantation (LT) recipients and patients with hepatectomy and the influence of HPgV infection after LT on clinical and perioperative factors. We enrolled 313 LT recipients and 187 patients with hepatectomy who received care at the Kyusyu University Hospital between May 1997 and September 2017. Of the 313 recipients and 187 patients enrolled in this study, 44 recipients (14.1%) and 2 patients (1.1%) had HPgV viremia, respectively. There was no significant association between HPgV infection and LT outcomes. Interestingly, one recipient was infected with HPgV during the peritransplant period, which was likely transmitted via blood transfusion because HPgV RNA was detected from the blood bag transfused to the recipient during LT. We reviewed the available literature on the prevalence HPgV infections in other organ‐transplanted patients and whether they impacted clinical outcomes. They also had the higher prevalence of HPgV infection, while it appears to be of low or no consequences. In addition, HPgV infection induced the upregulation of interferon‐stimulated gene (ISG) expression in peripheral blood mononuclear cells. LT recipients had higher HPgV viremia compared to patients with hepatectomy. Although HPgV infection was not associated with LT‐related outcomes, it induced ISG expression in recipients.
Highlight
HPgV infection induced the upregulation of interferon‐stimulated gene (ISG) expression in peripheral blood mononuclear cells.
Aim
Liver transplantation (LT) is the only curative therapy for decompensated liver cirrhosis. For recipients of living donor LT (LDLT), restoration of liver function after transplantation is highly ...dependent on liver regenerative capacity, which requires large amounts of intracellular energy. Mitochondrial metabolism provides a stable supply of adenosine 5′‐triphosphate (ATP) for liver regeneration. Mitophagy is a selective process in which damaged, non‐functional mitochondria are degraded and replaced with new functional mitochondria. We investigated the relationship between expression of Syntaxin17 (STX17), a key protein in mitophagy regulation, in donor livers and graft survival.
Methods
We examined STX17 expression in grafts from 143 LDLT donors who underwent right lobe resection and investigated the relationship between STX17 expression and graft function. We investigated the correlations among STX17 expression, mitochondrial membrane potential and cell proliferation, using a STX17‐knockdown hepatocyte cell line.
Results
Recipients transplanted with low STX17‐expression grafts had significantly lower graft survival rates than recipients transplanted with high STX17‐expression grafts (88.9% vs. 100%, p < 0.01). Multivariate analysis showed that low STX17 expression (HR: 10.7, CI: 1.29–88.0, p < 0.05) and the absence of splenectomy (HR: 6.27, CI: 1.59–24.8, p < 0.01) were independent predictive factors for small‐for‐size graft syndrome, which is the severe complication in LDLT. In the vitro experiments, the percentage of depolarized damaged mitochondria was increased in the STX17‐knockdown hepatocyte cell line, suggesting decreased mitophagy and ATP synthesis. Cell proliferation was significantly decreased in the STX17‐knockdown hepatocyte cell line.
Conclusion
STX17 contributes to mitophagy and maintenance of mitochondrial function in hepatocytes and may be a predictor of graft dysfunction in LDLT patients.
The effects of oxidative stress on various carcinomas were reported in previous studies, but those in intrahepatic cholangiocarcinoma (ICC) have not been fully elucidated. The purpose of this study ...was, thus, to reveal the effects of oxidative DNA damage and repair enzymes on ICC.
The levels of 8-hydroxydeoxyguanosine (8-OHdG) and 8-OHdG DNA glycosylase (OGG1) were immunohistochemically evaluated in specimens resected from 63 patients with ICC.
Low OGG1 expression was related to tumour depth T4 (p=0.04), venous invasion (p=0.0005), lymphatic vessel invasion (p=0.03), and perineural invasion (p=0.03). Compared to the high-OGG1-expression group, patients with low OGG1 expression had a significantly poorer prognosis (overall survival: p=0.04, recurrence-free survival: p=0.02). Unlike for OGG1, the expression levels of 8-OHdG showed no association with prognosis.
Oxidative DNA damage and DNA repair enzymes may be closely related to ICC progression.
Introduction
Programmed cell death ligand 1 (PD-L1) expression in tumor tissues is measured as a predictor of the therapeutic efficacy of immune checkpoint inhibitors (ICIs) in many cancer types. ...PD-L1 expression is evaluated by immunohistochemical staining using 3,3´-diaminobenzidine (DAB) chronogenesis (IHC-DAB); however, quantitative and reproducibility issues remain. We focused on a highly sensitive quantitative immunohistochemical method using phosphor-integrated dots (PIDs), which are fluorescent nanoparticles, and evaluated PD-L1 expression between the PID method and conventional DAB method.
Methods
In total, 155 patients with metastatic or recurrent cancer treated with ICIs were enrolled from four university hospitals. Tumor tissue specimens collected before treatment were subjected to immunohistochemical staining with both the PID and conventional DAB methods to evaluate PD-L1 protein expression.
Results
PD-L1 expression assessed using the PID and DAB methods was positively correlated. We quantified PD-L1 expression using the PID method and calculated PD-L1 PID scores. The PID score was significantly higher in the responder group than in the non-responder group. Survival analysis demonstrated that PD-L1 expression evaluated using the IHC-DAB method was not associated with progression-free survival (PFS) or overall survival (OS). Yet, PFS and OS were strikingly prolonged in the high PD-L1 PID score group.
Conclusion
Quantification of PD-L1 expression as a PID score was more effective in predicting the treatment efficacy and prognosis of patients with cancer treated with ICIs. The quantitative evaluation of PD-L1 expression using the PID method is a novel strategy for protein detection. It is highly significant that the PID method was able to identify a group of patients with a favorable prognosis who could not be identified by the conventional DAB method.
Aim
Our aim was to evaluate the clinical outcomes of telaprevir (TVR)‐ or simeprevir (SMV)‐based triple therapy for recurrent hepatitis C after living donor liver transplantation.
Methods
Twenty‐six ...patients received antiviral therapy, consisting of either TVR (n = 12) or SMV (n = 14) in combination with pegylated interferon and ribavirin, plus cyclosporin.
Results
More patients had a dose reduction of the direct‐acting agent (36.3% vs 0.0%, P = 0.02) or required blood transfusion for anemia (58.3% vs 7.1%, P < 0.01) in the TVR group. The cyclosporin trough/dose ratio increased significantly from week 0 to week 4 in the TVR group (1.6 ± 0.4 to 5.1 ± 2.0, P < 0.01), but not in the SMV group (1.2 ± 0.3 to 1.3 ± 0.2, P = 0.68). The 24‐week cumulative viral clearance rate was 91.7% and 85.7% in the TVR and in SMV groups, respectively. The early viral response and sustained viral response rates were 91.7% and 83.3%, respectively, in the TVR group, compared with 85.7% and 64.3%, respectively, in the SMV group. Interferon‐mediated graft dysfunction occurred in four and five patients in the TVR and SMV groups, respectively; two patients were treated by oral steroids, five by steroid pulse and two by thymoglobulin, resulting in viral breakthrough in one case.
Conclusion
SMV‐based triple therapy was associated with fewer adverse events and drug interactions with cyclosporin, and possibly less antiviral properties to TVR. Interferon‐mediated graft dysfunction is a significant clinical problem that warrants particular caution following living donor liver transplantation.
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