gms | German Medical Science

Despite current chemotherapies pancreatic cancer steadfastly remains refractory to treatment. Here we tested a new approach of combining antiangiogenic and standard cytotoxic therapy in a mtastatic human pancreatic cancer nude mouse model.

Nude athymic mice were injected orthotopically with metastatic human L3.6pl cancer cells. Pancreatic tumors were allowed to become established for 8 days before initiation of rapamycin or gemcitabine treatment. Standard doses of rapamycin (1.5 mg/kg/d) and gemcitabine (100 mg/kg, 2x/week) were used in the first group of experiments, and all animals were sacrificed 28 days after tumor cell injection. Immunohistochemical analysis was performed from primary pancreatzic tumors for proliferation (Ki67), cell death (TUNEL), and apoptotic endothelial cells (CD31/TUNEL). To directly test the effect of rapamycin on tumor blood vessel flow dynamics, L3.6pl tumor cells were implanted into dorsal skin-fold chambers and vessels were examined by intravital microscopy on day 7. FACS analysis of HUVE cells was performed to detect annexin-V positive cells.

Following orthotopic tumor cell injection, rapamycin treatment alone reduced tumor volume 2-fold more than the standard gemcitabine therapy. Furthermore, when rapamycin and gemcitabine treatment were combined, tumors were growing to only 19% of the size observed with gemcitabine treatment alone. Interestingly, histologic analysis revealed tumor vessel endothelium detachment and thrombosis with rapamycin treatment. Angiogenesis observation in dorsal skin-fold chambers after rapamycin treatment directly illustrated unusually dilated tumor vessels highly susceptible to thrombosis. Furthermore, when we photodynamically promoted vascular thrombosis in tumors, blood flow was very rapidly blocked by thrombosis in rapamycin-treated mice, compared to controls as monitored by intravital microscopy. Furthermore, CD31/TUNEL staining of orthotopic tumors demonstrated apoptotic endothelial cells with rapamycin treatment, which was substantiated in vitro by increased annexin-V staining of rapamycin-treated human endothelial cells. In contrast, gemcitabine showed no antiangiogenic effects, but induced extensive tumor cell apoptosis in vivo, albeit, without concomitantly reducing cell proliferation.

Our data suggest rapamycin's antiangiogenic activity inhibits tumor expansion, thereby more positively balancing the potent cytotoxic effect of gemcitabine against tumor progression. Furthermore, our study provides the first evidence that tumor control achieved with rapamycin is related to tumor-vessel thrombosis associated with the death of endothelial cells. Rapamycin promotion of thrombosis preferentially in new pancreatic tumor vessels introduces a novel mechanism potentially contributing to its anticancer action.