Rapamycin sirolimus

Rapamycin (sirolimus) (LC Laboratories) was dissolved in MeOH. Homoharringtonine was purchased from Sequoia Research Products Limited. Hoechst 33342 (Life Technologies) was used at 2 μg/mL in PBS. Antibodies: HA (1:1000; Covance), GLUT1 and HIF-2α (1:1000; Novus Biologicals), Bcl-xL and Caspase 9 (1:1000; Cell Signaling Technology), and α-Tubulin (1:5000; Sigma Aldrich).

Human osteosarcoma (OS) cell lines (HOS-MNNG and 143B) human rhabdomyosarcoma (RMS) cell lines (RD, alveolar RMS and Rh30, embryonal RMS) and the human breast cancer cell line MCF7 were obtained from the American Type Culture Collection (ATCC). Cells were grown in DMEM supplemented with 10% fetal bovine serum, penicillin, streptomycin, L-glutamine and incubated at 37°C with 5% CO₂ and 100% humidity. FK-506 (tacrolimus, LC Laboratories), CCI-779 (temsirolimus, LC Laboratories), rapamycin (sirolimus, LC Laboratories), Torin1 (kind gift from Dr. Nathanael S. Gray, Dana-Farber Cancer Institute and David M. Sabatini, Whitehead Institute/MIT/HHMI), thapsigargin (#T-9033, Sigma-Aldrich Corporation) and AZD-8055 (kind gift from Astra Zeneca Corporation).

Rapamycin (Sirolimus®) was purchased from LC Laboratories (MA). PD98059, an ERK inhibitor, was purchased from Sigma Aldrich (St. Louis, MO). ApoAlert™ DNA Fragmentation Assay kit was purchased from BD Biosciences, Palo Alto, CA. DAPI was bought from Vector Laboratories, Inc. CA. Antibodies for phospho-serine⁴⁷³ AKT, AKT, p-S6, S6, p-ERK1/2, ERK1/2, p-P38, P38, phospho-tyrosine⁷⁰⁵-STAT3, STAT3, Bcl-2, and Bax were purchased from Cell Signaling Technology. SOD-2, ferritin heavy chain, and GAPDH-HRP were purchased from Santa Cruz Biotechnology.