Galunisertib ly2157299

PcTx1 (a potent and selective ASIC1α blocker, Psalmotoxin 1) and GN25 (a novel inhibitor of Snail‐p53 binding) were procured from ApexBio Technology (Houston, USA). MK2206 (a specific inhibitor of AKT), TWS119 (a specific inhibitor of GSK3β) and Galunisertib (LY2157299) (TGF‐β/Smad inhibitor) were obtained from Selleck Chemicals (Houston, TX, USA). C19 (an inhibitor of EMT) was obtained from Shanghai Dongcang Biological Technology Co., Ltd. (Shanghai, China). OXA and 5FU were obtained from Sigma‐Aldrich (St. Louis, USA). These reagents and chemicals were dissolved in dimethyl sulfoxide at −80℃ until use and then diluted to a suitable working concentration with RPMI‐1640 medium. TGF‐β1 was obtained from PeproTech (Rocky Hill, NJ). TRIzol reagent was bought from Invitrogen Corp. (Carlsbad, CA, USA). First Strand cDNA Synthesis Kit was acquired from Thermo Fisher Scientific (Waltham, MA, USA).

TNFα (210-TA), IL-1β (201-LB), IL-6 (206-IL) and IL-23 (1290-IL) were purchased from R&D systems. Galunisertib (LY2157299) was purchased from Selleckchem. 1,25(OH)₂D₃ (BML-DM200-0050) were from, Enzo Life Sciences, Inc., Ann Arbor, MI. Stock 1,25(OH)₂D₃ solution of 2.4 mM were diluted in >99.5% ethanol anhydrous. AhR agonist (FICZ, 5304) and AhR antagonist (CH-223191, 3858) were from TOCRIS Inc. FICZ and CH-223191 were solubilized in DMSO to make a 25 mM and a 100 mM stock solution, respectively. RORγt antagonist (SR-2211) was from TOCRIS Inc. SR-2211 (4869) was solubilized in DMSO to make a 10 mM stock solution. Recombinant human IL-21 (200-21) was from PeproTech and anti-IL-21 (NBP1-76740) was from Novus Biologicals.

The inhibitors used in this study were K02288 (Selleckchem, S7359, Houston, TX, USA), Galunisertib/LY2157299 (Selleckchem, S2230), RepSox (Sigma‐Aldrich/Merck, R0158), A‐83‐01 (MedChem Express, HY‐10432, Monmouth Junction, NJ, USA), Erlotinib (EGFRi, Selleckchem, S1023), and human ALK1 inhibitory antibody (R&D Systems, MAB3701). The hepsin inhibitory antibody (Ab25) was a kind gift from Dr. Kirchhofer, Genentech [10 (link)]. ZFH7116 provided by Dr. Janetka [12 (link)]. The antibodies used in this study were Anti‐sheep HRP (Upstate cell signaling solutions, #12‐342), anti‐rabbit HRP (Millipore, AP132P, Burlington, MA, USA), anti‐hepsin (R&D Systems, AF4776), anti‐GAPDH (CST#2118S), anti‐β‐tubulin (Abcam, ab6046, Cambridge, UK), anti‐pEGFR Y1068 (Abcam, ab40815), anti‐EGFR (Abcam, ab32077), anti‐pSmad2/3 (CST#8828), anti‐Smad2/3 (CST#8685), anti‐Smad2 (CST#5339), anti‐pH3 (Santa Cruz, sc‐8656, Dallas, TX, USA), total‐H3 (CST#9715), anti‐V5 (Invitrogen, #R96025), anti‐Ki67 (Abcam, ab15580), anti‐cleaved caspase 3 (CST#9661), anti‐F‐actin (phalloidin staining) (Thermo Fisher Scientific, A22283), anti‐pSmad1/5 (CST#9516) and anti‐ALK‐1 (R&D Systems, AF370).