To establish orthotopic models, either 66cl4-shMYC-NC/1/3 or 4T1-MYC-vec/oe cells in 50 µL of DMEM medium were injected into the fourth mammary gland of 6-week-old female BALB/c or NSG mice (66cl4: 1×106 cells; 4T1: 5×105 cells). For CD8+ T cell depletion experiments, 6-week-old female BALB/c mice were treated with rat IgG2b (BioXCell, clone LTF-2, 100 µg injected intraperitoneally on day 1, followed by 50 µg on day 3 and 50 µg weekly for the remainder of the experiment) or anti-mouse CD8a (BioXCell, clone 2.43, 100 µg injected intraperitoneally on day 1, followed by 50 µg on day 3 and 50 µg weekly for the remainder of the experiment). For in vivo combination therapy, mice were treated with diluent or decitabine alone (S1200, Selleck, 0.8 mg/kg, intraperitoneal injection, every day) or in combination with isotype IgG (BioXcell, clone 2A3, 100 µg injected intraperitoneally, on days 3, 7, 10, 14) or anti-PD-1 antibody (BioXcell, clone RMP1-14, 100 µg injected intraperitoneally, on days 3, 7, 10, 14).
Tumor growth was measured 2–3 times per week using calipers. The primary tumors were harvested for analysis once they reached a 3–5 week time point or a volume of 1 or 2 cm3 using the formula: volume = (width2 ×length)/2. Mice were humanely sacrificed after measurement. For experimental accuracy, these data were also included.
Tumor growth was measured 2–3 times per week using calipers. The primary tumors were harvested for analysis once they reached a 3–5 week time point or a volume of 1 or 2 cm3 using the formula: volume = (width2 ×length)/2. Mice were humanely sacrificed after measurement. For experimental accuracy, these data were also included.