Subcutaneous and orthotopic xenograft and syngeneic models were used to assess the effect of combinations on the in vivo efficacy of CEA-IL2v. Briefly, for the NSCLC xenograft lung model, female human CD16-transgenic SCID mice (Charles River Laboratories, Lyon, France) were inoculated with 3 × 106 A549 cells injected intravenously. For the colorectal liver metastases models, the CRC cell line LS174T was injected into the spleen (3 × 106 cells). For the gastric sc model, the N87 cells were injected subcutaneously (1 × 106 cells). For the breast orthotopic model, the cell line KPL-4 was injected into the mammary fat pad (5 × 106 cells). For the pancreatic syngeneic model, female CEA-transgenic C57BL/6-CEA mice (Charles River Laboratories, Lyon, France) were inoculated with 1 × 105 Panc02-CEA cells injected intrapancreatically. Mice were maintained under specific-pathogen-free conditions with daily cycles of 12 h light/darkness according to guidelines (GV-SOLAS; FELASA) and food and water were provided ad libitum. Continuous health monitoring was performed and the experimental study protocol was reviewed and approved by the Veterinary Department of Kanton Zurich.
Mice were randomized into different treatment groups and therapy started when evidence of tumor growth was visible in the target organ of killed scout animals (days indicated in figure legends). All treatments were administered IV. The termination criterion for sacrificing animals was sickness with locomotion impairment, and median OS was defined as the experimental day by which 50% of animals had been killed. Kaplan–Meier survival curves and the Pairwise Log-Rank test were used to compare survival between animals.