Sw872

Eribulin was kindly provided by Eisai Inc. (Tokyo, Japan) and Sapporo Medical University for in vitro and in vivo use, respectively. MK-2206 was obtained from ChemScene (Monmouth, NJ, USA). Pazopanib was obtained from AdooQ BioScience (Manassas, VA, USA). Stock solutions of these reagents were generated by dissolving the powder in 100% dimethyl sulfoxide (DMSO; Sigma–Aldrich St. Louis, MO, USA) at 10 mM. HT1080, SK-LMS-1 and SW872 cell lines were purchased from ATCC (Manassas, VA, USA). Both cell lines were maintained in DMEM (Sigma–Aldrich) containing 10% fetal bovine serum, 1% penicillin–streptomycin and 2 μM L-glutamine.

Human fibrosarcoma (FS; HS 93.T), leiomyosarcoma (LMS; HS5.T), liposarcoma (LPS; SW872), and rhabdomyosarcoma (RMS; RD) cell lines were obtained from ATCC (Manassas, VA, USA) and cultured in DMEM with 10% FBS, 2 mM L-glutamine, 100 U/mL penicillin, and 100 mg/mL streptomycin at 37 °C in a humidified 5% CO₂ atmosphere.

The human liposarcoma cell lines SW872 (an undifferentiated liposarcoma, ATCC catalog number: HTB-92) and SW982 (another undifferentiated liposarcoma as evaluated by histopathology, ATCC catalog number: HTB-93) were purchased from the ATCC (Rockville, MD). These cell lines were cultured in RPMI 1640 (Invitrogen, Carlsbad, CA) supplemented with 10% FBS, 100 units/ml penicillin and 100 μg/ml streptomycin (Invitrogen). Cells were incubated at 37°C in a 5% CO₂-95% air atmosphere and passaged when near confluent monolayers were achieved using Trypsin-EDTA solution. Cells were free of Mycoplasma contamination, as tested by the MycoAlert Mycoplasma Detection Kit from Cambrex (Rockland, ME).

We obtained HDF (human dermal fibroblast cell line) (Cat #, 106-05a) from CELL APPLICATIONS, INC. (San Diego, CA, USA). We purchased MG63 (human osteosarcoma), HT1080 (human fibrosarcoma), SW872 (human liposarcoma), SW1353 (human chondrosarcoma), hTERT-HME1 (human mammary epithelial cell line) (Cat #, CRL-4010), and NuLi-1 (human bronchial epithelial cell line) (Cat #, CRL-4011) from ATCC (Manassas, VA, USA).
HDF cells were maintained in basal medium with a growth supplement (Cat #, 115-485, 116-GS, CELL APPLICATIONS, INC., San Diego, CA, USA) at 37 °C in a humidified atmosphere containing 5% CO₂. All sarcoma cell lines, hTERT-HME1, and NuLi-1 cells were cultured in Dulbecco’s Modified Eagle Medium (D-MEM) (Thermo Fisher Scientific, Waltham, MA, USA) containing 10% fetal bovine serum (Thermo Fisher Scientific) and 1% penicillin–streptomycin–glutamine mixed solution (nacalai tesque, Kyoto, Japan) under the same conditions (37 °C, 5% CO₂). The hTERT-HME1 and NuLi-1 cells were immortalized by introducing the catalytic subunit of human telomerase (hTERT) or E6/E7 and hTERT, respectively.

MDA-MB-231 (Sigma Aldrich, Vienna, Austria), HaCat (ATCC, US), MCF10A (LGC Promochem, US), A375 (ATCC, US), SW-872 (ATCC, US), 93T449 (ATCC, US), SW1353 (CLS, Germany) and juvenile fibroblasts fresh established from foreskin samples were obtained from Division of Biomedical Research (BMF), Medical University of Graz, Austria. HeLa (ATCC®, Guernsey, UK), fibroblast, HaCat, SW1353 and A375 cells were cultured in DMEM supplemented with 2 mM glutamine, 1% PS (100 U/mL penicillin, 100 μg/mL streptomycin) and 10% fetal bovine serum (FBS). MCF10A were cultured in DMEM with single quot kit suppl. Gr, 5% Horse Serum, 20 ng/mL hEGF, 0.5 μg/ml hydrocortison, 100 ng/ml choleratoxin, 10 μg/ml insulin and 2 mM glutamine. MDA-MB-231 were maintained in DMEM Hams F12 with 10% FBS, 2 mM glutamine and 1% PS. SW872 were cultured in DMEM Hams F12 supplemented with 5% FBS, 2 mM glutamine and 1% PS. 93T449 were cultured with RPMI-1640 with 10% FBS, 2 mM glutamine, 10mM 4-(2-hydroxyethyl)-1-piperazineethanesulfonic acid (HEPES), 1mM sodium pyruvate and 1% PS.
Cells were maintained in a humidified incubator at 37°C with 5% CO₂. HeLa cells were treated for up to 3 days with AdOx (40 μM), MS023 (10 μM), GSK715 (2 μM), GSK591 (1 μM) and DMSO, before cell extracts were prepared.