In vivo xenograft mouse models

All in vivo efficacy studies were approved by the Institutional Animal Care and Use Committee (IACUC) of the University of Kansas Medical Center (KUMC) (IACUC# 2017–2387). Four-week old female athymic nude mice (Foxnl^nu/Foxnl^nu) were purchased from The Jackson Laboratory (Bar Harbor, ME) and inoculated subcutaneously in the right flanks with a suspension of TC-71 cells (1 × 10⁶) or CHLA-258 cells (5 × 10⁶) mixed with an equal volume of ice-cold Matrigel (BD Biosciences). After tumors were established, mice were randomized into two treatment groups and treated as follows: 1) vehicle control; equivalent dose of entinostat vehicle, oral gavage, once daily and 2) entinostat; 25 mg/kg of entinostat in 2% DMSO and 30% PEG300 (pH 2.1–2.5), oral gavage, once daily. Tumor volume and body weight were measured at least twice per week. Tumor volumes were measured with calipers and calculated using the following formula: volume (mm³) = length × (width)²/2. Mice were humanely euthanized and gross necropsies were performed when tumor volumes exceeded 4,000 mm³ (TC-71 study) or after 36 days of treatment (CHLA-258 study).