Fig. 3

Primary tumor treatment with AGI-134 causes tumor regression, activates complement and FSL distribution in tumors. a Glycolipid detection in B16-F10 tumors: FSL-Fluorescein was used as surrogate molecule for AGI-134 visualization in tumors. 1 × 10⁶ B16-F10 cells were grafted onto immunized α1,3GT^−/− mice on both flanks. Five days later, the two tumors on each mouse were treated with 100 μL of 1 mg/mL FSL-fluorescein on one flank and with 100 μL PBS on the other flank. The following day, the tumors were excised and frozen in OCT compound. The tumors were sectioned and labeled with DAPI. Pictures in the GFP and DAPI channels for FSL and tumor cell nucleus DNA visualization were taken using ×4–×40 objectives (×10 example pictures are shown). The pictures show representative data of DAPI and GFP channel picture overlays for a vehicle- and a fluorescein-lipid treated tumor from the same mouse. b In complement activation experiments, B16-F10 tumors were treated by intratumoral injection of vehicle (PBS) or 1 mg AGI-134 on Day 6 post B16-F10 cell grafting. 2.5 h after treatment, tumors were excised, homogenized and the complement factor C5a measured by ELISA. Each symbol represents the total C5a in the tumor homogenate of each mouse, median C5a values are indicated by the bars. Differences between the PBS vs. AGI-134 treatment groups were assessed by Mann–Whitney test (**p < 0.003). c In primary tumor regression experiments, back transformed least square geometric means for PBS and AGI-134 treatments across the timepoints were calculated and fold-reduction in geometric means ± 95% CI plotted, (*p < 0.05, n = 13)