Mv4 11

Human myeloid leukemia cells lines, TF-1a, TF-1, and MV4-11, were purchased from ATCC (Rockville, MD) and maintained in RPMI 1640 (TF-1a and TF-1) or IMDM (MV4-11) supplemented with 10% FBS without (TF-1a) or with GM-CSF (5 ng/mL, TF-1, MV4-11) at 37°C in humidified air containing 5% CO₂. All the media and sera were purchased from Life Technologies, Inc. (Gaithersburg, MD). During log-phase growth, the cells were collected and transferred to 6-well plates at a concentration of 10⁵ cells/mL. For differentiation study, PMA (Sigma Chemical Co., St. Louis, MO) was added to the cells and incubated for 48–72 h. For apoptosis assay, cells were treated with Bay 11-7085 (Calbiochem, La Jolla, CA) at different concentrations. The control cells were treated with PMA or Bay 11-7085 solvent. After the treatment for 24 and 48 hours, the cells were harvested into a 15 mL polypropylene centrifuge tube and spun down for 8 min at 600 RPM (80 g), after which the supernatant was discarded and the cells were resuspended in 0.5 mL of culture medium and 1-2 drops of the cell suspension were placed on a slide in the central area and moved around to form a thin and even film with a glass spreader. The glass spreader was made from glass transfer pipette over an alcohol burner for few seconds to minutes.

Human cell lines promyelocytic leukemia HL-60, acute lymphocytic leukemia lines Reh and RS4:11, acute myeloid leukemia MV-4-11, myelogenous leukemia lines K562 and KG-1a, epidermoid carcinoma A431, and Chinese hamster ovary (CHO) cell line were purchased from American Tissue Culture Collection (ATCC, Manassas, VA). The acute myeloid leukemia MOLM-14 line was purchased from the German Collection of Microorganisms and Cell Lines (DSMZ, Braunschweig Germany). The cell lines with the exception of A431, MV-4-11, and KG-1a, were cultured in RPMI-1640 Medium (ATCC) supplemented with 10% heat-inactivated fetal bovine serum (FBS). The A431 line was cultured in DMEM Medium (ATCC) supplemented with 10% heat inactivated FBS. The MV-4-11 cell line was cultured in IMDM Medium (ATCC) supplemented with 10% heat-inactivated FBS. The KG-1a line was cultured in IMDM Medium supplemented with 20% FBS. Where applicable, luciferase-expressing subclones were generated by stably transducing wild-type leukemia lines with lentiviral vector encoding firefly luciferase with or without GFP (Lentigen Technology, Inc., Gaithersburg, MD), followed by limiting dilution and selection of luciferase-positive clones.

Human AML cell lines MOLM13 (RRID:CVCL_2119), SET2 (RRID:CVCL_2187), SKM1 (RRID:CVCL_0098) and OCI-AML5 (RRID:CVCL_1620) were obtained from the DSMZ (Braunschweig, Germany). MV4-11 cells were obtained from the ATCC (Manassas, VA). All experiments with cell lines were performed within 6 months after thawing or obtaining from ATCC or DSMZ. The cell lines were also authenticated in the Characterized Cell Line Core Facility at M.D. Anderson Cancer Center, Houston TX. SKM1 and MOLM13 cells were cultured in RPMI media with 20% fetal bovine serum (FBS) and 1% penicillin/streptomycin and 1% non-essential amino acids. OCI-AML5 cells were cultured in MEM alpha media with 20% FBS and 1% penicillin/streptomycin and 1% non-essential amino acids. OCI-AML5 cultures were also supplemented with 10 ng/mL of GM-CSF (PeproTech, Rocky Hill, NJ). MV4-11 cells were cultured in ATCC-formulated Iscove's Modified Dulbecco's Medium (IMDM) with 20% FBS 1% penicillin/streptomycin and 1% non-essential amino acids. Luciferase-expressing MOLM13 cells were created by transducing cells with Luc-ZSGreen(GFP). pHIV-Luc-ZsGreen was a gift from Bryan Welm (Addgene plasmid #39196). Logarithmically growing, mycoplasma-negative cells were utilized for all experiments. Following drug treatments, cells were washed free of the drug(s) prior to the performance of the studies described.