Vincristine

Ba/F3 TEL-FGFR4, a murine hematopoietic cell line that expresses a constitutively active FGFR4 kinase domain [19 (link)], was cultured in RPMI-1640 medium supplemented with 10% fetal bovine serum (FBS), 2 mM L-glutamine, 1% penicillin/streptomycin, and 1.5 ug/mL puromycin. RMS559 is a FN RMS cell line with an FGFR4^V550L activating mutation; RH4 and SCMC are FP RMS cell lines with FGFR4 overexpression; and 7250 is a normal human fibroblast cell line. These cell lines were grown in DMEM medium supplemented with 10% FBS, 2 mM L-glutamine, and 1% penicillin/streptomycin. All cell lines were STR profiled and negative for mycoplasma. Futibatinib was obtained through a Cooperative Research and Development Agreement (CRADA) with Taiho Oncology (Princeton, NJ, USA). Irinotecan (752903, Pfizer, New York, NY, USA) and Vincristine (030906, Pfizer) were obtained through the NIH Division of Veterinary Resources Pharmacy.

Normal human smooth muscle (HA-VSMC), RMS (RH30, aRMS), (RD, eRMS) and rhabdoid cell line (A204) cells were procured from American Type Culture Collection (ATCC). While (CW9019, aRMS) and (SMS-CTR, eRMS) cells were gifted to JGP by Dr. Frederic G. Barr, Deputy Branch Chief and Senior Investigator Laboratory of Pathology, National Cancer Institute, USA. RMS cells were cultured in DMEM supplemented with 10% fetal bovine serum, 100 U/ml of penicillin, and 100 μg/ml of streptomycin at 37°C in a humidified atmosphere of 5% CO₂. HA-VSMC were grown in F-12K medium supplemented with 0.05 mg/ml ascorbic acid, 0.01 mg/ml insulin, 0.01 mg/ml transferrin, 10 ng/ml sodium selenite, 0.03 mg/ml endothelial cell growth supplement (ECGS); fetal bovine serum to a final concentration of 10%, HEPES to a final concentration of 10 mM, TES to a final concentration of 10 mM. GANT-61 was purchased from Cayman (MI, USA). Rapamycin was purchased from Fujian Kerui Pharmaceutical Co., Ltd (Fuzhou, Fujian, China). Temsirolimus and vincristine were obtained from Pfizer (NY, USA) and Hospira (IL, USA) respectively. Primers used in this study were obtained from Invitrogen (Supplementary Table S1-I and II). Primary antibodies were purchased as described in supplementary Table S2. Various treatments of these cells were performed when their confluence reached to about 70-80%.