Combined effects of SN-38 and pentoxifylline on the induction of apoptosis through the activation of CPP-32 in pancreatic adenocarcinoma cell lines

Background. Pentoxifylline (PENT) is a theophylline derivative that enhances cytotoxic effects against tumor cells pretreated with antitumor agents. It has also been reported that chemotherapy can induce apoptosis in some carcinoma cells. We investigated the effects of PENT on human pancreatic adenocarcinoma cells pretreated with SN-38, an active form of CPT-11 (a camptothecin analogue) and we also examined the participation of CPP-32, a member of the interleukin 1β-converting enzyme (ICE) family proteases, in chemotherapeutic agent-induced apoptosis. Methods. Human pancreatic adenocarcinoma cells (PK-1, PK-8) were cultured in RPMI 1640. Lethal effects were examined by MTT assay; DNA fragmentation was analyzed by agarose gel electrophoresis; and Western blot analysis was performed with anti-CPP-32 monoclonal antibody. Results. Pretreatment with SN-38 followed by PENT increased the cytotoxic effect compared with that seen for treatment with SN-38 alone. Isobologram analysis of the IC₅₀ value revealed that PENT had supra-additive effects when administed after SN-38, but not when administered prior to or simultaneously with SN-38. Agarose gel electrophoresis showed typical DNA ladders in the DNA of cells treated with SN-38 and PENT. The acridine orange (AO) staining method was used to observe the morphological changes characteristic of apoptosis. Western blot analysis verified that activation of CPP-32 accompanied the development of apoptosis. In addition, SN-38-induced apoptosis was prevented by pretreatment with Ac-DEVD-CHO (DEVD), an inhibitor of CPP-32. Conclusions. These results indicate that the antitumor activity of SN-38 is attributable to apoptosis through the activation of CPP-32, and that combined treatment with PENT enhances the induction of apoptosis by SN-38. Accordingly, the use of PENT may provide a combined modality treatment for pancreatic cancer. Received: August 15, 1997 / Accepted: November 27, 1998