The transcription factor NF-kB has anti-apoptotic properties and may confer chemoresistance to cancer cells. Here, we describe human pancreatic carcinoma cell lines that di er in the responsiveness to the topoisomerase-2 inhibitors VP16 (20 mM) and doxorubicin (0.3 mM): Highly sensitive BxPC-3 and PT45-P1 cells, and Capan-1 and A818-4 cells that were almost resistant to both anti cancer drugs. VP16, but not doxorubicin, transiently induced NF-kB activity in all cell lines, whereas basal NF-kB binding was nearly undetectable in BxPc-3 and PT45-P1 cells, but rather high in Capan-1 and A818-4 cells, as demonstrated by gel-shift and luciferase assays. Treatment with various NF-kB inhibitors (Gliotoxin, MG132 and Sulfasalazine), or transfection with the IkBa super-repressor, strongly enhanced the apoptotic e ects of VP16 or doxorubicin on resistant Capan-1 and 818-4 cells. Our results indicate that under certain conditions the resistance of pancreatic carcinoma cells to chemotherapy is due to their constitutive NF-kB activity rather than the transient induction of NF-kB by some anti-cancer drugs. Blockade of basal NF-kB activity by well established drugs e ciently reduces chemoresistance of pancreatic cancer cells and o ers the potential for improved therapeutic strategies. Oncogene (2001) 20, 859 ± 868.
The early response gene IEX-1 is involved in the regulation of cellular growth and survival, and its expression is related to stress-, growth-and deathinducing signals. Addressing the role of IEX-1 in the promotion of apoptosis, we investigated the effect of IEX-1 on nuclear factor-jB (NF-jB) activation. Stably transfected HEK-293 cells conditionally overexpressing IEX-1 exhibit decreased levels of NF-jB activity, either basal or TNFa induced, as shown by gel-shift and luciferase reporter gene assay. Furthermore, activated p65 accumulated in the nuclei of 293 cells to a lower degree, if IEX-1 expression was increased. This inhibited NF-jB activation was preceded by an altered turnover of IjBa and phospho-IjBa. In addition, IEX-1 expression also inhibited the activity of the 26S-proteasome, as shown by a fluorometric proteasome assay. Conversely, disruption of IEX-1 expression in 293 cells by stable transfection with specific anti-IEX-1 hammerhead ribozymes increased NF-jB activity, and accelerated the degradation of IjBa. Along with these opposite effects of IEX-1 expression and IEX-1 disruption on NF-jB activation, the sensitivity of 293 cells towards various apoptotic stimuli also changed. In contrast to ribozymetransduced 293 cells that were significantly less sensitive to apoptosis, this sensitivity was enhanced if IEX-1 expression was increased. Our data suggest that IEX-1 -itself an NF-jB target gene -inhibits the activation of this transcription factor, and hereby may counteract the antiapoptotic potential of NF-jB.
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