Antiprotease strategy in combination with gemcitabine as a putative novel therapeutic approach for pancreatic cancer treatment

Introduction: gabexate mesylate (gm), a protease inhibitor, has been shown to exert a significant antitumoral activity in pancreatic cancer (PC) cells. Aim: To evaluate whether pre-treatment with gm could improve PC cells response toward gemcitabine (gem). Methods: Studies have been performed on PanC-1, SW1990 PC cells and ea.hy926 endothelial cells. Treatment consisted of gm 50 µg/ml for 24 hrs, followed by 24 hrs of treatment with gem 6 µg/ml; 24 hrs of gm 50 µg/ml followed by 24 hrs in culture medium; 24 hrs of gem 6 µg/ml followed by 24 hrs in culture medium. aspects studied included: cell viability (mTT assay), cell invasiveness and migration (Boyden chambers chemoinvasion and chemotaxis assay), angiogenesis (endothelial tube formation assay), nf-kB activation (nf-kB luciferase assay, Western Blotting,), vegf and il-8 levels (eliSa), mmP-2 and mmP-9 activity (gelatin zymography), Ras signalling (Western Blotting). results: gm significantly enhanced gem anti-invasive and anti-angiogenic efficacy by inhibition of gem-induced nf-kB activation (P< 0.001). This phenomenon occurred by the block of ikBalpha degradation that, in turn, prevented Rela/p65 nuclear translocation (P<0.001) and resulted in down-regulation of the nf-kB products mmP-2 and mmP-9, vegf and il-8 (P< 0.001). drugs in combination also inhibited eRK1/2 and aKT activation, two pathways targeting nf-kB, by increased expression of their endogenous inhibitors RKiP and PTen, respectively. gm did not enhance the gem action on cell viability after 24 hrs. Conclusion: These in vitro data indicate that gm pretreatment significantly improves gem anti-invasive and anti-angiogenic efficacy in PC cells, in part by nf-kB inhibition.