Syngeneic murine colorectal cancer (CRC) and breast cancer cells were treated with temozolomide (TMZ) and/or cisplatin (CDDP) at clinically relevant concentrations in a pulsatile schedule (2 days treatment followed by drug washout and cell passaging) for a total of 12in vitrotreatment cycles (priming phase). Drug concentrations were as follows: 25 M for TMZ and 1.5 M for CDDP. Additionally, CRC cells were also treated with the standard-of-care cytotoxic agents: 2 M for 5-fluorouracil (5FU), 5 M for oxaliplatin (Oxa), and 35 nM for SN38 (the main active metabolite of irinotecan). Primed cells were kept in drug-free medium for two passages and then injected into immunodeficient and immunocompetent mice to study the extent of immunological control induced by treatment with chemotherapeutic agents (editing phase). Whole exome sequencing (WES) was performed at baseline (T0), at the end of thein vitropriming phase (T1) and on established tumors at the end of thein vivochallenge phase (T2).
