Anti-tumor activity of F8-LIGHT was tested in two syngeneic murine models of cancer. CT26 and WEHI-164 tumors were implanted in the right flank of BALB/c mice by subcutaneous injection of 3 × 106 and 2.5 × 106 cells, respectively. Treatments were started when tumors reached a volume of about 100 mm3. The indicated dose of immunocytokines or the corresponding volume of saline, were administered by intravenous injection in the lateral tail vein, every other day for a total of three injections. The immune-checkpoint inhibitor anti-PD-1 (BioXCell, clone 29 F.1A12) was administered in the same way, at alternate days. Mice were inspected daily. Body weight was monitored, tumor was measured with a caliper and tumor size was determined using the formula: ½ x (major diameter) x (minor diameter)2. Mice with ulcerated hemorrhagic tumor, or with tumor bigger than 1500 mm3 where euthanized.

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