Cultivation and Characterization of Human Mucoepidermoid Carcinoma Cell Lines

University of Michigan Human Mucoepidermoid Carcinoma cell lines (UM-HMC-1, UM-HMC-3A, and UM-HMC-3B) (RRID:CVCL_Y473, RRID:CVCL_Y471, RRID:CVCL_Y472, respectively) were generated from surgical specimens (23 (link)). These cells were cultured in high glucose Dulbecco’s Modified Eagle’s Medium (Invitrogen; Carlsbad, CA, USA) supplemented with 10% fetal bovine serum (Atlanta Biologicals; Flowery Branch, GA, USA), 1% L-Glutamine (MilliporeSigma; Burlington, MA, USA), 1% Antibiotic-Antimycotic (MilliporeSigma), 400 ng/mL hydrocortisone (StemCell Technologies; Vancouver, BC, Canada), 20 ng/mL recombinant human epidermal growth factor (R&D Systems; Minneapolis, MN), and 5 μg/mL recombinant human insulin (Sigma-Aldrich; Burlington, MA, USA) at 37°C and 5% CO₂. Low passage primary human microvascular endothelial cells (HDMEC) (Lonza; Morristown, NJ, USA) were cultured in endothelial growth medium-2 for microvascular cells (Lonza). Small molecule inhibitors of MDM2-p53 interaction (i.e., MI-773, APG-115, and MI-1061) and MDM2 degrader (MD-224) were provided by Shaomeng Wang (University of Michigan) (22 (link), 24 (link), 25 (link)). MG132 (MilliporeSigma) was used to inhibit proteasomal degradation. PTC596 (MedChemExpress; Monmouth Junction, NJ), a small molecule inhibitor of Bmi-1, was used to evaluate the direct impact of Bmi-1 on MEC stemness.