xav939 and Liver-Neoplasms

AXIN1 mutations are observed in 8-10% of hepatocellular carcinomas (HCCs) and originally were considered to support tumor growth by aberrantly enhancing β-catenin signaling. This view has however been challenged by reports showing neither a clear nuclear β-catenin accumulation nor clearly enhanced expression of β-catenin target genes. Here, using nine HCC lines, we show that AXIN1 mutation or siRNA mediated knockdown contributes to enhanced β-catenin signaling in all AXIN1-mutant and non-mutant lines, also confirmed by reduced signaling in AXIN1-repaired SNU449 cells. Both AXIN1 and AXIN2 work synergistically to control β-catenin signaling. While in the AXIN1-mutant lines, AXIN2 is solely responsible for keeping signaling in check, in the non-mutant lines both AXIN proteins contribute to β-catenin regulation to varying levels. The AXIN proteins have gained substantial interest in cancer research for a second reason. Their activity in the β-catenin destruction complex can be increased by tankyrase inhibitors, which thus may serve as a therapeutic option to reduce the growth of β-catenin-dependent cancers. At concentrations that inhibit tankyrase activity, some lines (e.g. HepG2, SNU398) were clearly affected in colony formation, but in most cases apparently independent from effects on β-catenin signaling. Overall, our analyses show that AXIN1 inactivation leads to enhanced β-catenin signaling in HCC cell lines, questioning the strong statements that have been made in this regard. Enhancing AXIN activity by tankyrase monotherapy provides however no effective treatment to affect their growth exclusively through reducing β-catenin signaling.

Topics: Axin Protein; beta Catenin; Carcinoma, Hepatocellular; Cell Line, Tumor; Heterocyclic Compounds, 3-Ring; Humans; Liver Neoplasms; Tankyrases; Tumor Stem Cell Assay; Wnt Signaling Pathway

Astrotactin 1 (ASTN1) is known to serve a physiological role in neuronal migration; however its role in liver cancer remains to be determined. In the present study, ASTN1 levels were lower in liver cancer tissues compared with those in matching normal tissue. ASTN1 levels were negatively associated with microscopic vascular invasion, advanced clinical stage and a less favorable prognosis in patients with hepatocellular carcinoma (HCC). Furthermore, ASTN1 overexpression in a liver cancer cell line reduced the migratory and invasive capacity of the cells. Based on bioinformatics analysis, ASTN1 levels were negatively associated with the Wnt signaling pathway. In addition, ASTN1 downregulated the protein expression levels of β‑catenin, T‑cell factor (TCF)1, TCF4, Jun proto‑oncogene (C‑jun), Myc proto‑oncogene (C‑myc), cyclooxygenase‑2 (COX2), metalloproteinase (MMP)2, MMP9 and vascular endothelial growth factor (VEGF) protein levels, indicative of suppression of Wnt signaling. Furthermore, XAV939‑induced Wnt signaling suppression reversed the ASTN1‑mediated inhibition of invasion and migration in cells. Overexpression of ASTN1 in xenografts reduced cancer development as well as Wnt signaling. TIMER analysis showed that ASTN1 expression was negatively correlated with B cell, macrophage and neutrophil infiltrating levels in HCC. Together, the results of the present study showed that ASTN1 reduced the migratory and invasive capacity of liver cancer cells, potentially served as a candidate biomarker for diagnosis and prediction of the prognosis of HCC, and was associated with immune infiltration. Understanding the underlying mechanisms of action of ASTN1 may facilitate the development of novel strategies for prevention and treatment of liver cancer.

Topics: Adult; Aged; beta Catenin; Carcinoma, Hepatocellular; Cell Movement; Cell Proliferation; Female; Gene Expression Regulation, Neoplastic; Genes, myc; Hepatocyte Nuclear Factor 1-alpha; Heterocyclic Compounds, 3-Ring; Humans; JNK Mitogen-Activated Protein Kinases; Liver Neoplasms; Male; Matrix Metalloproteinase 2; Matrix Metalloproteinase 9; Middle Aged; Neoplasm Invasiveness; Nerve Tissue Proteins; Receptors, Cell Surface; Transcription Factor 4; Wnt Signaling Pathway; Young Adult

Previous data indicate that Tankyrase inhibitors exert anti-growth functions in many cancer cell lines due to their ability to inactivate the YAP protooncogene. In the present manuscript, we investigated the effect of Tankyrase inhibitors on the growth of hepatocellular carcinoma (HCC) cell lines and the molecular mechanisms involved. For this purpose, we performed cell proliferation assay by colony-forming ability in seven human HCC cells subjected to XAV-939 and G007-LK Tankyrase inhibitors. Noticeably, the two Tankyrase inhibitors suppressed the HCC cell growth in a dose-dependent manner. Furthermore, we found that Tankyrase inhibitors synergized with MEK and AKT inhibitors to suppress HCC cell proliferation. At the molecular level, Tankyrase inhibitors significantly decreased YAP protein levels, reduced the expression of YAP target genes, and inhibited YAP/TEAD luciferase reporter activity. In addition, Tankyrase inhibitors administration was accompanied by upregulation of Angiomotin-like 1 (AMOTL1) and Angiomotin-like 2 (AMOTL2) proteins, two major negative regulators of YAP. Altogether, the present data indicate that XAV-939 and G007-LK Tankyrase inhibitors could suppress proliferation of hepatocellular carcinoma cells and downregulate YAP/TAZ by stabilizing AMOTL1 and AMOTL2 proteins, thus representing new potential anticancer drugs against hepatocellular carcinoma.

Topics: Adaptor Proteins, Signal Transducing; Angiomotins; Antineoplastic Agents; Apoptosis; Blotting, Western; Carcinoma, Hepatocellular; Carrier Proteins; Cell Line, Tumor; Cell Proliferation; Dose-Response Relationship, Drug; Heterocyclic Compounds, 3-Ring; Humans; Liver Neoplasms; Membrane Proteins; Phosphoproteins; Real-Time Polymerase Chain Reaction; Sulfones; Tankyrases; Transcription Factors; Triazoles; YAP-Signaling Proteins

Deregulated WNT/β-catenin signaling contributes to the development of a subgroup of hepatocellular carcinoma (HCC), the second leading cause of cancer deaths worldwide. Within this pathway, the tankyrase enzymes (TNKS1 and TNKS2) degrade AXIN and thereby enhance β-catenin activity. We evaluate TNKS enzymes as potential therapeutic targets in HCC, and the anti-tumor efficacy of tankyrase inhibitors (XAV939, and its novel nitro-substituted derivative WXL-8) in HCC cells. Using semi-quantitative RT-PCR, we found significantly elevated levels of TNKS1/2 mRNA in tumor liver tissues compared to adjacent non-tumor livers, at protein levels only TNKS1 is increased. In HepG2, Huh7cells, siRNA-mediated knockdown suppression of endogenous TNKS1 and TNKS2 reduced cell proliferation, together with decreased nuclear β-catenin levels. XAV939 and WXL-8 inhibited cell proliferation and colony formation in HepG2, Huh7, and Hep40 cells (p < 0.05), with stabilization of AXIN1 and AXIN2, and decreased β-catenin protein levels. XAV939 and WXL-8 also attenuated rhWNT3A-induced TOPflash luciferase reporter activity in HCC cells, indicating reduced β-catenin transcriptional activity, consistent with decreased nuclear β-catenin levels. In vivo, intra-tumor injections of XAV939 or WXL-8 significantly inhibited the growth of subcutaneous HepG2 xenografts (P < 0.05). We suggest that tankyrase inhibition is a potential therapeutic approach for treating a subgroup HCC with aberrant WNT/β-catenin signaling pathway.

Topics: Animals; Axin Protein; beta Catenin; Carcinoma, Hepatocellular; Cell Proliferation; Dose-Response Relationship, Drug; Gene Expression Regulation, Neoplastic; Genes, Reporter; Hep G2 Cells; Heterocyclic Compounds, 3-Ring; Humans; Liver Neoplasms; Male; Mice, Inbred NOD; Mice, SCID; Molecular Targeted Therapy; Poly(ADP-ribose) Polymerase Inhibitors; Protein Stability; RNA Interference; Tankyrases; Time Factors; Transfection; Tumor Burden; Wnt Signaling Pathway; Wnt3A Protein; Xenograft Model Antitumor Assays