Grp78 sirna

DT-010 (purity > 98%, Figure 1A) were synthesized at Jinan University, China (Scheme 1). TMP and DSS (Figure 1A) were purchased from Shanghai Banghai Chemical Company (China) and Xi’an Honson Biotechnology (China), respectively. The Dulbecco’s Modified Eagle’s Medium (DMEM), penicillin-streptomycin and fetal bovine serum (FBS) used in this study were obtained from Life Technologies (USA). 3-(4, 5-dimethylthiazol-2-yl)-2, 5-diphenyltetrazoliumbromide (MTT) was purchased from Sigma-Aldrich (USA). GRP78 siRNA was provided by Santa Cruz Biotechnology (USA). An XF Glycolysis Stress Test Kit was purchased from Seahorse Bioscience (USA). Lactate assay kit was purchased from BioVision (USA).
Scheme 1 Synthesis of DT-010. Reagents and conditions: (a) KMnO₄, 45°C, overnight, 50%; (b) CH₃CH₂OH, EDCI, DMAP, r.t., 84%; (c) C₃H₅MgBr, THF, 0°C to r.t., 34%; (d) Ac₂O, HClO₄, r.t., 3 h, 36%; (e) C₂Cl₂O₂, DMF, CH₂Cl₂; (f) n-C₄H₉Li, THF, 0°C to r.t., 41%; (g) Na₂CO₃, CH₃OH, H₂O, 82%.

JNK inhibitor SP600125 (SP), eIF2α phosphatase enzymes inhibitor salubrinal (Sal) and mTOR inhibitor rapamycin (Rap) were purchased from Tocris Bioscience (Bristol, UK). p70S6K inhibitor PF-4708671 (PF) was purchased from Selleck Chemicals (Houston, TX, USA). AP-1 inhibitor curcumin, cell counting kit-8 (CCK8) and ER stress inducer tunicamycin (Tun) were purchased from Sigma (Lyon, France). The eIF4E/eIF4G interaction inhibitor 4EGI-1, mTOR siRNA, GFP siRNA, JNK siRNA, GRP78 siRNA, ATF4 siRNA and antibodies against GRP78, eIF2α and β-actin were purchased from Santa Cruz Biotechnology (Heidelberg, Germany). Antibodies against phospho-eIF2α (Ser-51), phospho-p70S6K (Thr-389), phospho-mTOR (Ser-2448), phospho-Raptor (Ser-863), phospho-c-Jun (Ser-73), phospho-JNK (Thr-183/Tyr-185), phospho-4E-BP1 (Thr-37/46), p70S6K, mTOR, Raptor, c-Jun, JNK, 4E-BP1 and ATF4 were purchased from Cell Signaling Technology (Danvers, MA, USA).