B16-F10 and B16-OVA melanoma, MC38 colon carcinoma or 4 T1 breast carcinoma cells were injected subcutaneously (5 × 105–106) into the right flank of 8- to 10-week-old female C57BL/6 or BALB/c (6–11 mice/group) on day 0. Tumors were measured twice per week with calipers and the volume calculated (length x width2/2). When tumors reached a volume of 80–100 mm3 (day 0) mice were randomized into different groups of treatment according to the experiment. Poly I:C or BO-112 formulation (2.5 mg/Kg, 100 μl), was administered by intratumoral injection twice per week for three weeks (six doses in total). The control group received intratumoral injections of 5% glucose (BO-112 vehicle, identical volume) or PEI (identical amount per dose as present in each dose of BO-112). Survival was monitored daily, and tumors were measured twice per week until the animals died or the tumor volume reached the maximum allowed size. The in vitro cytotoxicity of BO-112 in mouse and human cell lines was continuously assessed by measuring electric impedance in an xCELLigence machine (ACEA). Tumor cells (1.5-2 × 105) were seeded on specific 8-well plates to measure electric impedance. After 4-5 h, BO-112 or Poly I:C (Sigma) was added to culture media at identical concentrations in a final volume of 200 μL per well. PEI (Polyplus-transfection®) was added to culture media at the same concentrations as it is present in BO-112 formulation. Electric impedance was measured every five minutes for 48 h. In vitro BO-112 cytotoxicity was also assessed.