Figure 3
Irinotecan inhibits HIF1A protein accumulation and angiogenesis in colon tumor xenografts. Western blot analysis of HIF1A protein accumulation in human colon tumor xenografts is shown in panel A. Tumors from mice treated with irinotecan (T) show a complete inhibition of HIF1A protein accumulation compared with control tumors from untreated mice (C). The inhibition of HIF1A protein accumulation is reversed in xenografts that have started to grow again (R) after the end of the treatment period. Results were confirmed in two independent experiments. Panel B shows the detection of intratumor hypoxia in human colon tumor xenografts of mice treated with irinotecan. Mice were injected intraperitoneally with 100 mg/kg pimonidazole 1 h before euthanasia. Pimonidazole adducts in hypoxic areas of the tumors were identified using hypoxyprobe monoclonal antibodies on paraffin-embedded sections. Bars represent 250 µm. Results were confirmed in two independent experiments. Panel C shows histological examinations of human colon tumor xenografts in mice treated with irinotecan. H&E and trichrome staining show a decrease in tumor cellularity and an increase in extracellular matrix deposition, respectively, in tumors from irinotecan-treated mice (bars represent 2.5 mm). Cellular proliferation was evaluated with Ki-67 labeling within clusters of tumor cells (bars represent 500 µm). CD31 staining shows well-formed vessels (arrows) in the control tumors of untreated mice, whereas the vascular network is completely disorganized in the tumors of irinotecan-treated mice. This effect is reversed in tumors that have started to grow again after the end of the treatment period (bars represent 1 mm). Results were confirmed in two independent experiments. Panel D shows the quantification of the impact of irinotecan treatment on cell proliferation as assessed by Ki-67 staining. The mean values for the average percentage of Ki-67-positive cells were calculated along with SEM. No significant difference was observed in tumor cells from control (C), irinotecan-treated (T) and regrowth (R) samples.