Sorafenib is the only drug approved for the treatment of hepatocellular carcinoma (HCC). The bioenergetic propensity of cancer cells has been correlated to anticancer drug resistance, but such correlation is unclear in sorafenib resistance of HCC. Six sorafenib-naive HCC cell lines and one sorafenib-resistant HCC cell line (Huh-7R; derived from sorafenib-sensitive Huh-7) were used. The bioenergetic propensity was calculated by measurement of lactate in the presence or absence of oligomycin. Dichloroacetate (DCA), a pyruvate dehydrogenase kinase (PDK) inhibitor, and siRNA of hexokinase 2 (HK2) were used to target relevant pathways of cancer metabolism. Cell viability, mitochondrial membrane potential, and sub-G1 fraction were measured for in vitro efficacy. Reactive oxygen species (ROS), adenosine triphosphate (ATP) and glucose uptake were also measured. A subcutaneous xenograft mouse model was used for in vivo efficacy. The bioenergetic propensity for using glycolysis correlated with decreased sorafenib sensitivity (R²=0.9067, among sorafenib-naive cell lines; P=0.003, compared between Huh-7 and Huh-7 R). DCA reduced lactate production and increased ROS and ATP, indicating activation of oxidative phosphorylation (OXPHOS). DCA markedly sensitised sorafenib-resistant HCC cells to sorafenib-induced apoptosis (sub-G1 (combination vs sorafenib): Hep3B, 65.4±8.4% vs 13±2.9%; Huh-7 R, 25.3± 5.7% vs 4.3±1.5%; each P<0.0001), whereas siRNA of HK2 did not. Sorafenib (10 mg kg⁻¹ per day) plus DCA (100 mg kg⁻¹ per day) also resulted in superior tumour regression than sorafenib alone in mice (tumour size: −87% vs −36%, P<0.001). The bioenergetic propensity is a potentially useful predictive biomarker of sorafenib sensitivity, and activation of OXPHOS by PDK inhibitors may overcome sorafenib resistance of HCC.

Background Regorafenib and other multikinase inhibitors may enhance antitumor efficacy of anti-program cell death-1 (anti-PD1) therapy in hepatocellular carcinoma (HCC). Its immunomodulatory effects, besides anti-angiogenesis, were not clearly defined. Methods In vivo antitumor efficacy was tested in multiple syngeneic liver cancer models. Murine bone marrow–derived macrophages (BMDMs) were tested in vitro for modulation of polarization by regorafenib and activation of cocultured T cells. Markers of M1/M2 polarization were measured by quantitative reverse transcription PCR (RT-PCR), arginase activity, flow cytometry, and ELISA. Knockdown of p38 kinase and downstream Creb1/Klf4 signaling on macrophage polarization were confirmed by using knockdown of the upstream MAPK14 kinase, chemical p38 kinase inhibitor, and chromatin immunoprecipitation. Results Regorafenib (5 mg/kg/day, corresponding to about half of human clinical dosage) inhibited tumor growth and angiogenesis in vivo similarly to DC-101 (anti-VEGFR2 antibody) but produced higher T cell activation and M1 macrophage polarization, increased the ratio of M1/M2 polarized BMDMs and proliferation/activation of cocultured T cells in vitro, indicating angiogenesis-independent immunomodulatory effects. Suppression of p38 kinase phosphorylation and downstream Creb1/Klf4 activity in BMDMs by regorafenib reversed M2 polarization. Regorafenib enhanced antitumor efficacy of adoptively transferred antigen-specific T cells. Synergistic antitumor efficacy between regorafenib and anti-PD1 was associated with multiple immune-related pathways in the tumor microenvironment. Conclusion Regorafenib may enhance antitumor immunity through modulation of macrophage polarization, independent of its anti-angiogenic effects. Optimization of regorafenib dosage for rational design of combination therapy regimen may improve the therapeutic index in the clinic.

Immune checkpoint inhibitors (ICIs), including antibodies that target programmed cell death protein 1 (PD-1), programmed death-ligand 1 (PD-L1), or cytotoxic T lymphocyte antigen 4 (CTLA4), represent some of the most important breakthroughs in new drug development for oncology therapy from the past decade. CXC chemokine ligand 13 (CXCL13) exclusively binds CXC chemokine receptor type 5 (CXCR5), which plays a critical role in immune cell recruitment and activation and the regulation of the adaptive immune response. CXCL13 is a key molecular determinant of the formation of tertiary lymphoid structures (TLSs), which are organized aggregates of T, B, and dendritic cells that participate in the adaptive antitumor immune response. CXCL13 may also serve as a prognostic and predictive factor, and the role played by CXCL13 in some ICI-responsive tumor types has gained intense interest. This review discusses how CXCL13/CXCR5 signaling modulates cancer and immune cells to promote lymphocyte infiltration, activation by tumor antigens, and differentiation to increase the antitumor immune response. We also summarize recent preclinical and clinical evidence regarding the ICI-therapeutic implications of targeting the CXCL13/CXCR5 axis and discuss the potential role of this signaling pathway in cancer immunotherapy.

Markers that could accurately predict responses to the general kinase inhibitor sorafenib are needed to better leverage its clinical applications. In this study, we examined a hypothesized role in the drug response for the growth arrest DNA damage-inducible gene 45β (GADD45β), which is commonly underexpressed in hepatocellular carcinoma (HCC) where sorafenib may offer an important new therapeutic option. The anticancer activity of sorafenib-induced GADD45β expression was tested in a panel of HCC cell lines and xenograft models. We found that GADD45β mRNA and protein expression were induced relatively more prominently in HCC cells that were biologically sensitive to sorafenib treatment. GADD45β induction was not found after treatment with either the mitogen-activated protein kinase–extracellular signal-regulated kinase (ERK) kinase (MEK) inhibitor U0126 or the Raf inhibitor ZM336372, suggesting that GADD45β induction by sorafenib was independent of Raf/MEK/ERK signaling activity. However, c-Jun NH₂-terminal kinase (JNK) kinase activation occurred preferentially in sorafenib-sensitive cells. Small interfering RNA–mediated knockdown of GADD45βor JNK kinase limited the proapoptotic effects of sorafenib in sorafenib-sensitive cells. We defined the −339/−267 region in the GADD45β promoter containing activator protein-1 and SP1-binding sites as a crucial region for GADD45β induction by sorafenib. Together, our findings suggest that GADD45β induction contributes to sorafenib-induced apoptosis in HCC cells, prompting further studies to validate its potential value in predicting sorafenib efficacy. Cancer Res; 70(22); 9309–18. ©2010 AACR.

Multikinase inhibitors with antiangiogenic properties used to be standard therapy for patients with advanced hepatocellular carcinoma (HCC). Recently, several antiangiogenic agents (lenvatinib, cabozantinib, and ramucirumab) have demonstrated antitumor activity for advanced HCC in randomized controlled trials. However, the landscape of drug development for HCC may change dramatically with the advent of immune checkpoint inhibitor therapy, particularly the anti–programmed cell death-1 (anti-PD1) agents. In addition, early-phase clinical trials of combination of anti–PD-1 and antiangiogenic agents have shown very promising anti-tumor activity in patients with advanced HCC. Therefore, the critical research questions at present are whether this combination strategy will be the next generation of standard therapy and which antiangiogenic agents will be the optimal partner for the combination. All of the 4 multikinase inhibitors for HCC (sorafenib, regorafenib, lenvatinib, and cabozantinib) have been reported to have immune modulatory effects. The authors systematically reviewed the pre-clinical evidence of their immune modulatory effects to explore whether these effects were mediated by angiogenesis inhibition or by other “off-target” effects on the tumor microenvironment. Studies of sorafenib comprised the majority (58 of the 71) of the research articles reviewed. Potentially beneficial effects on anti-tumor immunity may result from increased M1 polarization of macrophages and stimulation of CD8 T cell function. On the other hand, high dosage of the kinase inhibitors in pre-clinical models and hypoxia associated with angiogenesis may contribute to immune suppression in the tumor microenvironment. Sorafenib and other multikinase inhibitors may promote anti-tumor immunity through modulation of multiple immune cell types as well as the tumor microenvironment. The optimal immune modulatory dosage should be defined to facilitate design of future combination regimens. Part of this study has been presented at the 9^th Asia Pacific Primary Liver Cancer Expert Meeting. July 6–8, 2018, Seoul, Korea. ^* Yu-Yang Lin and Ching-Ting Tan contributed equally to this work. ^§ Dr. Da-Liang Ou is a National Taiwan University YongLin Scholar since 2018.

Aberrant histone methylation is a frequent event during tumor development and progression. KMT1E (also known as SETDB1) is a histone H3K9 methyltransferase that contributes to epigenetic silencing of both oncogenes and tumor suppressor genes in cancer cells. In this report, we demonstrate that KMT1E acts as a metastasis suppressor that is strongly downregulated in highly metastatic lung cancer cells. Restoring KMT1E expression in this setting suppressed filopodia formation, migration, and invasive behavior. Conversely, loss of KMT1E in lung cancer cells with limited metastatic potential promoted migration in vitro and restored metastatic prowess in vivo. Mechanistic investigations indicated that KMT1E cooperates with the TGFβ-regulated complex SMAD2/3 to repress metastasis through ANXA2. Together, our findings defined an essential role for the KMT1E/SMAD2/3 repressor complex in TGFβ-mediated lung cancer metastasis. Cancer Res; 74(24); 7333–43. ©2014 AACR.