Xav 939

IL8, ICAM1, MIF, CXCL12, EGF and bFGF recombinant protein were from PeproTech (Rocky Hill, America). Tamoxifen and Insulin were purchased from Sigma-Aldrich (Darmstadt, Germany). The Wnt/β-catenin pathway inhibitor XAV939 and activator CHIR99021 were from TargetMol (Massachusetts, America). The CXCR1 and CXCR2 antagonist Repertaxin was obtained from MedChem Express (America). The Annexin V-FITC Apoptosis Detection Kit (Cat#APOAF-50TST) was obtained from Sigma-Aldrich (Darmstadt, Germany). The ALDEFLUOR™ Kit (Cat#01700) was obtained from Stem Cell (Canada). ChIP Assay Kit (Cat#P2078) was obtained from Beyotime (Shanghai, China). Proteome Profiler Human Cytokine Array Kit (Cat#ARY005B) was purchased from R&D System (California, USA). Human IL-8/NAP-1 Platinum ELISA Kit (Cat#BMS204/3) was purchased from eBioscience (California, USA).

The TβRI inhibitor LY2157299 (Interchim, LSK040) was added to the cells at a final concentration of 10 µM. The SMAD3 inhibitor (E)-SIS3 (MedChemExpress, HY-13013) was used at a final concentration of 10 µM. Cells were treated with 0.5 µM PD184352 (Sigma Aldrich, PZ0181), a MEK inhibitor, 10 µM SP600125 (Sigma Aldrich, S5567), a JNK inhibitor, 10 µM SB203580 (Sigma Aldrich, S8307), a p38 inhibitor and 0.1 µM wortmannin (Sigma Aldrich, W1628), a PI3K inhibitor. All treatments with the inhibitors were performed 1 h prior the addition of TGFβ1. The GSK3 inhibitor CHIR-99021 (TargetMol, T2310) and the tankyrase inhibitor XAV-939 (TargetMol, T1878) were used at 10 µM final concentration. The SMARCA2/4 bromodomain inhibitor PFI-3 (MedChemExpress, HY-12409) and the allosteric dual SMARCA2 and brahma-related gene 1 (BRG1)/SMARCA4 ATPase activity inhibitor BRM/BRG1 ATP Inhibitor-1 (MedChemExpress, HY-119374) were administrated to cells at 10 and 5 µM final concentrations, respectively. Dimethyl sulfoxide (DMSO) was used as a vehicle control treatment.

HCC cells were seeded into a 96-well plate at the density of 2000 cells per well, followed by the treatment of 3-HAA (Sigma, Cat: 148776), XAV-939 (TargetMol, Cat: T1878), and/or sorafenib (Meilunbio, Cat: MB1226) at appropriate doses as indicated elsewhere. CCK8 assays were performed in triplicates as instructed by the manufacturer (Dojindo, Tabaru, Japan). Absorbance was measured at 450 nm using a microplate reader and cell viability was normalized to control and the mean of at least three independent experiments was calculated.