Su11274

Gefitinib, SU11274, 17-DMAG, and 17-AAG were obtained from Selleckchem (Houston, TX, USA). All other chemicals were acquired from Sigma Chemical (St. Louis, MO, USA), unless otherwise indicated. Anti-EGFR, anti-total ERK, anti-phosphorylated (p)-ERK, anti-total AKT, and anti-p-AKT antibodies were obtained from Cell Signaling (Danvers, MA, USA). Anti-HSP70 and anti-HSP90 antibodies were obtained from Genetex (Irvine, CA, USA). All other antibodies were purchased from Santa Cruz Biotechnology (Dallas, TX, USA).

Human bone marrow mesenchymal stem cells (hBMSCs) were bought from Cyagen Biosciences Inc., (Goleta, CA, USA). Informed consent was obtained in accordance with the Declaration of Helsinki and the Institutional Review Board of the Southern Medical University (approval number SMU-2015123, 2016.01.10). Cells were cultured at 37 °C, 5% CO₂, in the Human Mesenchymal Stem Cell Basal Medium complemented with 10% of the qualified fetal bovine serum (FBS), 1% Penicillin-streptomycin and 1% Glutamine (all from Cyagen Biosciences Inc.). The potency of osteogenic, chondrogenic, and adipogenic differentiation has been confirmed before obtainment. Cells were infected with recombinant adenoviruses and the proliferation and osteogenic differentiation were assayed. In some experiments, cells were treated with signaling pathway inhibitors agents as indicated in the figure legends, including c-MET inhibitor SU11274 (10 μM), WNT inhibitor Wnt-C59 (2 μM), ERK1/2 inhibitor U0126 (10 μM), PI3K/AKT inhibitor LY294002 (10 μM) (all from Selleckchem, Houston, TX, USA), and DMSO (Sigma-Aldrich, St. Louis, MO, USA) was used as the negative control.

After trypsinization, 5 × 10⁵ transfected MKN28 cells were seeded on 6‐well plates for 12 hour. After 24 hours of serum deprivation, SU11274 (phosphorylated c‐Met (p‐c‐Met) inhibitor; Selleck Chemicals, Houston, TX, USA; 5 μmol/L), MK2206 (phosphorylated AKT (p‐AKT) inhibitor; Santa Cruz Biotechnology, Dallas, TX, USA; 5 μmol/L), and rapamycin (phosphorylated mTOR (p‐mTOR) inhibitor; Selleck Chemicals, Houston, TX, USA; 200 nmol/L) were added to the 6‐well plates and cocultured for 24 hours. Then, the cells were collected and probed by western blot assay.

PF-03084014 (Nirogacestat, #S8018), ARQ197 (Tivantinib, #S2753), EMD-1214063 (Tepotinib, # S7067), SU11274 (#S1080) were purchased from Selleckchem (www.selleckchem.com). BMS-777607 (CT-BMS777) and Cabozantinb (CT-XL184) were purchased from ChemieTek (Indianapolis, IN, USA). GSI-XII (Z-Ile-Leu-aldehyde, HY-12465) was purchased from MedChemExpress (Monmouth Junction, NJ, USA). All compound were dissolved in DMSO to a final concentration of 10 mmol/L.