Background and Aim: The transcription factor NF-κB is constitutively activated in many human cancers, and induces the expression of multiple genes including COX-2, VEGF, and IAP. Therefore, the blocking of NF-κB activity may be associated with the suppression of angiogenesis, and induction of apoptosis. It is also reported that NF-κB is further activated by chemotherapy in some cancer cell lines and NF-κB activation is one of mechanisms by which tumors are induced to become resistant to chemotherapy. Thus, it is expected that the blocking of NF-κB could attenuate the tumor growth, and enhance the anti-tumor effect of chemotherapeutic compounds. In this study, we examined the relevance of NF-κB in the resistance of pancreatic carcinoma cell lines against for gemcitabine. We and other groups reported that hyperthermia-induced Hsp70 could inhibit Ikk, resulting in the inhibition of NF-κB activation. Therefore, we speculated that the activated NF-κB in pancreatic cell line might be inhibited by hyperthermia, resulting in the enhancement of gemcitabine-induced cytotoxicity. Methods: We used human pancreatic carcinoma cell lines, AsPC-1 and MIAPaCa-2. AsPC-1 and MIAPaCa-2 were treated with various concentration (0, 5, 10, 20 and 30μM) of gemcitabine for 24 hours. Hyperthermia (43 degrees Celsius 1hr) was combined with gemcitabine at the various timing. The effect of gemcitabine and heat treatment on cell survival was determined by WST-8 assay. The status of NF-κB in carcinoma cells exposed to gemcitabine was investigated by EMSA and immunocytochemistry. We analyzed apoptosis and necrosis in AsPC-1 and MIAPaCa-2 by flow cytometry. Furthermore the levels of Hsp70, cyclin D1, caspase-3, VEGF in each treatment group were detected by Western bloting.Results:1) A significant cytotoxicity was observed with gemcitabine. 2) Gemcitabine activated NF-κB binding activity in both cell lines. 3) Hyperthermia inhibited the gemcitabine-induced activation of NF-κB. 4) Hyperthermia enhanced the cytotoxicity of gemcitabine especially when hyperthermia was performed 24hr before the treatment of gemcitabine.5) The levels of Hsp70 was increased by hyperthermia. Gemcitabine did not affect the protein level of Hsp70. The levels of pre-caspase-3 were decreased by hyperthermia combined with gemcitabine.Conclusions: Hyperthermia inhibited gemcitabine-induced activation of NF-κB, resulting in the enhancement of the cytotoxicity of gemcitabine. It is important to investigate the schedule-dependency of hyperthermia combined with gemcitabine.

98th AACR Annual Meeting-- Apr 14-18, 2007; Los Angeles, CA