C0621

The human TC cell lines gifted from Dr Electron Kebebew from Stanford University or purchased from University of Colorado Cancer Center or ATCC were used in this study: follicular TC - FTC133, papillary TC - BCPAP, TPC1; MTC - TT; anaplastic TC THJ16T, THJ29T, 8505C, OCUT2, KAT18, SW1736 and C643 (ATC); and HTC - XTC1. A rat pancreatic cell line, AR42J, with intrinsically high SSTR2 expression, was used as a positive control for SST analog uptake studies (13 (link)). The cells were authenticated by short tandem repeat (STR) analysis and contamination was excluded via IMPACT II PCR analysis.
The cells were grown in the standard media, supplemented with 10% FBS (ThermoFisher Scientific) for human cell lines and 20% for a rat AR42J cell line, as well as with 10 U/mL penicillin-streptomycin (Gibco) and 0.25 µg/mL amphotericin B (Gibco). For treatment (72h), valproic acid (VAC, Sigma Aldrich, PHR1061) was added to the culture medium at a final concentration of 2 mM or 4 mM, while tacedinaline (Sigma Aldrich, C0621) and decitabine (Sigma Aldrich, A3656) at final concentration of 500 ng/mL and 75 ng/mL, respectively which corresponds to their therapeutic serum concentrations (21 (link)).

The animal protocol for this study was approved by the NCI/CCR Animal Care and Use Committee (ASP SOP-001). QGP-1 or BON-1 cells (5×10⁶ cells) were injected subcutaneously into both flanks of five- to seven-week-old athymic nude male mice CAnN.Cg-Foxn1nu/Cr, (Jackson Laboratory). The resulting tumors were further subcutaneously implanted, to create homogenous tumors. Only tumors from generations 1 to 3 were utilized. Tumor size was monitored using a Vernier caliper, with tumor initiation size ranging from 5 to 7 mm. The mice were euthanized if they exhibited impeded movement, if there were signs of breathing difficulty or tumors exceeded 2 cm in diameter at any point in the study. Mice were randomly grouped into control and treatment groups and then treated via intraperitoneal (i.p.) daily injections with either VPA (300 mg/kg; P4543; Sigma Aldrich) dissolved in water, CI-994 (5mg, 7.5mg and 10mg/kg; C0621; Sigma Aldrich) dissolved in 0.1% DMSO, or 0.1% DMSO as control group. All interventions were done during the light cycle, and mice were not fasted before assessments.