Anti-angiogenetische Therapie zur Behandlung vom humanen Pankreaskarzinom nach orthotoper Implantation in die Nacktmaus durch Blockade von NFκB mit Hilfe des Proteasom Inhibitor PS-341

Abstract

PS-341, a boronate inhibitor of the multisubunit protease complex, blocks the degradation of regulatory proteins involved in cell cycle regulation and cellular survival. The proteasome is required for the activation of NF-КB via the degradation of the inhibitory protein IКB. NF-КB triggers survival pathways in a variety of cells via the upregulation of inhibitors of apoptosis. Recent reports indicate that NF-КB is constitutively activated in pancreatic carcinoma. Treatment of pancreatic tumors implanted orthotopically in nude mice with PS-341 showed a decrease in angiogenic endpoints. We therefore investigated the effect of PS-341 on NF-КB-mediated survival and VEGF production in human pancreatic cancer cell lines. Human pancreatic cell lines Mia-PaCall and L3.6pl were injected orthotopically in nude mice. After 14 days the tumors were treated bi-weekly with various doses of PS-341 for up to 4 weeks. The tumors were then harvested and evaluated for apoptosis, proliferation, NF-КB activity, and angiogenic markers. In vitro analysis of NF-КB activity was done using gel shift techniques, and dual luciferase assays using a NF-КB consensus sequence and a full length VEGF promoter. Treatment of established tumors resulted in the marked decrease in viability and proliferation. In addition, there was a decrease in NF-КB activity, VEGF production, and microvessel density in the treated tumors. In vitro analysis demonstrated a marked decrease in NF-КB activity via gel shift and NF-КB driven luciferase. PS-341 also decreased the production of VEGF in the cell lines as measured by full-length VEGF promoter in front of a luciferase construct. The effects of PS- 341 mimicked transient transfection of the super repressor of NF-КB, IКB, with serine to alanine mutations at the regulatory residues. PS-341 is an effective therapy against human pancreatic carcinoma. These tumors apparently rely on NF-КB mediated survival pathways. Treatment reduces proliferation and increases tumor cell apoptosis via the blockade of NF-КB activation. The reduction in NF-КB activity reduces the production of VEGF, which acts as a survival factor for tumor endothelial cells resulting in a reduction of tumor vascularity.