Search

Background Intrahepatic cholangiocarcinoma (iCCA) is a highly malignant tumor characterized by an intensive desmoplastic reaction due to the exaggerated presence of the extracellular (ECM) matrix components. Liver fibroblasts close to the tumor, activated by transforming growth factor (TGF)-β1 and expressing high levels of α-smooth muscle actin (α-SMA), become cancer-associated fibroblasts (CAFs). CAFs are deputed to produce and secrete ECM components and crosstalk with cancer cells favoring tumor progression and resistance to therapy. Overexpression of Notch signaling is implicated in CCA development and growth. The study aimed to determine the effectiveness of the Notch inhibitor, Crenigacestat, on the surrounding microenvironment of iCCA. Methods We investigated Crenigacestat’s effectiveness in a PDX model of iCCA and human primary culture of CAFs isolated from patients with iCCA. Results In silico analysis of transcriptomic profiling from PDX iCCA tissues treated with Crenigacestat highlighted “liver fibrosis” as one of the most modulated pathways. In the iCCA PDX model, Crenigacestat treatment significantly (p p p p p Conclusions Notch signaling inhibition reduces the peritumoral desmoplastic reaction in iCCA, blocking the TGF-β1 canonical pathway.

Background & Aims Mounting evidence implicates the Hippo downstream effectors Yes-associated protein (YAP) and transcriptional co-activator with PDZ-binding motif (TAZ) in hepatocellular carcinoma (HCC). We investigated the functional contribution of YAP and/or TAZ to c-MYC-induced liver tumor development. Methods The requirement for YAP and/or TAZ in c-Myc-driven hepatocarcinogenesis was analyzed using conditional Yap, Taz, and Yap;Taz knockout (KO) mice. An hepatocyte-specific inducible TTR-CreERT2 KO system was applied to evaluate the role of YAP and TAZ during tumor progression. Expression patterns of YAP, TAZ, c-MYC, and BCL2L12 were analyzed in human HCC samples. Results We found that the Hippo cascade is inactivated in c-Myc-induced mouse HCC. Intriguingly, TAZ mRNA levels and activation status correlated with c-MYC activity in human and mouse HCC, but YAP mRNA levels did not. We demonstrated that TAZ is a direct transcriptional target of c-MYC. In c-Myc induced murine HCCs, ablation of Taz, but not Yap, completely prevented tumor development. Mechanistically, TAZ was required to avoid c-Myc-induced hepatocyte apoptosis during tumor initiation. The anti-apoptotic BCL2L12 gene was identified as a novel target regulated specifically by YAP/TAZ, whose silencing strongly suppressed c-Myc-driven murine hepatocarcinogenesis. In c-Myc murine HCC lesions, conditional knockout of Taz, but not Yap, led to tumor regression, supporting the requirement of TAZ for c-Myc-driven HCC progression. Conclusions TAZ is a pivotal player at the crossroad between the c-MYC and Hippo pathways in HCC. Targeting TAZ might be beneficial for the treatment of patients with HCC and c-MYC activation. Lay summary The identification of novel treatment targets and approaches for patients with hepatocellular carcinoma is crucial to improve survival outcomes. We identified TAZ as a transcriptional target of c-MYC which plays a critical role in c-MYC-dependent hepatocarcinogenesis. TAZ could potentially be targeted for the treatment of patients with c-MYC-driven hepatocellular carcinoma.

Intrahepatic cholangiocarcinoma (iCCA) is a deadly disease with rising incidence and few treatment options. An altered expression and/or activation of NOTCH1–3 receptors has been shown to play a role in iCCA development and progression. In this study, we established a new CCA patient-derived xenograft model, which was validated by immunohistochemistry and transcriptomic analysis. The effects of Notch pathway suppression by the Crenigacestat (LY3039478)-specific inhibitor were evaluated in human iCCA cell lines and the PDX model. In vitro, LY3039478 significantly reduced Notch pathway components, including NICD1 and HES1, but not the other Notch receptors, in a panel of five different iCCA cell lines. In the PDX model, LY3039478 significantly inhibited the Notch pathway and tumor growth to the same extent as gemcitabine. Furthermore, gene expression analysis of iCCA mouse tissues treated with LY3039478 revealed a downregulation of VEGFA, HES1, and MMP13 genes. In the same tissues, DLL4 and CD31 co-localized, and their expression was significantly inhibited in the treated mice, as it happened in the case of MMP13. In an in vitro angiogenesis model, LY3039478 inhibited vessel formation, which was restored by the addition of MMP13. Finally, RNA-sequencing expression data of iCCA patients and matched surrounding normal liver tissues downloaded from the GEO database demonstrated that NOTCH1, HES1, MMP13, DLL4, and VEGFA genes were significantly upregulated in tumors compared with adjacent nontumorous tissues. These data were confirmed by our group, using an independent cohort of iCCA specimens. Conclusion: We have developed and validated a new iCCA PDX model to test in vivo the activity of LY3039478, demonstrating its inhibitory role in Notch-dependent angiogenesis. Thus, the present data provide new knowledge on Notch signaling in iCCA, and support the inhibition of the Notch cascade as a promising strategy for the treatment of this disease.

Hepatocellular carcinoma (HCC) is the major type of primary liver cancer. Genomic studies have revealed that HCC is a heterogeneous disease with multiple subtypes. BRG1, encoded by the SMARCA4 gene, is a key component of SWI/SNF chromatin-remodeling complexes. Based on TCGA studies, somatic mutations of SMARCA4 occur in ~3% of human HCC samples. Additional studies suggest that BRG1 is overexpressed in human HCC specimens and may promote HCC growth and invasion. However, the precise functional roles of BRG1 in HCC remain poorly delineated. Here, we analyzed BRG1 in human HCC samples as well as in mouse models. We found that BRG1 is overexpressed in most of human HCC samples, especially in those associated with poorer prognosis. BRG1 expression levels positively correlate with cell cycle and negatively with metabolic pathways in the Cancer Genome Atlas (TCGA) human HCC data set. In a murine HCC model induced by c-MYC overexpression, ablation of the Brg1 gene completely repressed HCC formation. In striking contrast, however, we discovered that concomitant deletion of Brg1 and overexpression of c-Met or mutant NRas (NRASV12) triggered HCC formation in mice. Altogether, the present data indicate that BRG1 possesses both oncogenic and tumor-suppressing roles depending on the oncogenic stimuli during hepatocarcinogenesis.

Background: Hepatocellular carcinoma (HCC) is the most common primary liver cancer histotype, characterized by high biological aggressiveness and scarce treatment options. Recently, we have established a clinically relevant murine HCC model by co-expressing activated forms of v-akt murine thymoma viral oncogene homolog (AKT) and oncogene c-mesenchymal-epithelial transition (c-Met) proto-oncogenes in the mouse liver via hydrodynamic tail vein injection (AKT/c-MET mice). Tumor cells from these mice demonstrated high activity of the AKT/ mammalian target of rapamycin (mTOR) and Ras/ Mitogen-activated protein kinase (MAPK) signaling cascades, two pathways frequently co-induced in human HCC. Methods: Here, we investigated the therapeutic efficacy of sorafenib, regorafenib, the MEK inhibitor PD901 as well as the pan-mTOR inhibitor MLN0128 in the AKT/c-Met preclinical HCC model. Results: In these mice, neither sorafenib nor regorafenib demonstrated any efficacy. In contrast, administration of PD901 inhibited cell cycle progression of HCC cells in vitro. Combined PD901 and MLN0128 administration resulted in a pronounced growth constraint of HCC cell lines. In vivo, treatment with PD901 or MLN0128 alone moderately slowed HCC growth in AKT/c-MET mice. Importantly, the simultaneous administration of the two drugs led to a stable disease with limited tumor progression in mice. Mechanistically, combined mitogen-activated extracellular signal-regulated kinase (MEK) and mTOR inhibition resulted in a stronger cell cycle inhibition and growth arrest both in vitro and in vivo. Conclusions: Our study indicates that combination of MEK and mTOR inhibitors might represent an effective therapeutic approach against human HCC.