Abstract 443: Mechanisms of IL-1 production and release in pancreatic cancer cells

Abstract

Abstract Pancreatic Ductal AdenoCarcinoma (PDAC) is a very aggressive disease, characterized by desmoplasia and Th2-inflammation. Th2-inflammation correlates with poor prognosis and it is driven by the release of the Thymic Stromal LymphoPoietin (TSLP) from Cancer Associated Fibroblasts (CAFs) activated by tumor-derived pro-inflammatory cytokines. The aims of our study are: i) to verify which tumor-derived cytokines are responsible for CAFs activation, ii) to understand the stimuli/mechanisms of tumor-derived cytokine production in pancreatic cancer cells. Analyzing a series of 9 PDAC cell lines by both Real-time PCR and immunofluorescence, we found heterogeneous expression of IL-1α, IL-1β and TNF-α. Furthermore, ELISA tests revealed that some PDAC cell lines could either actively secrete or release during necrosis these pro-inflammatory cytokines. Next, we treated CAFs obtained from surgical patient samples with rhIL-1α, rhIL-1β, rhTNF-α and PDAC cell supernatants. Interestingly, we observed an induction in TSLP secretion that was significantly reduced by anti-IL1α and Anakinra (the recombinant form of IL-1Ra), suggesting a predominant role of the IL-1/IL-1R axis. It is known that IL-1α and TLRs ligands can activate NF-kB that in turn induces the synthesis of proIL-1β that is cleaved to matureIL-1β by caspase-1, upon inflammasome activation. In line with this, we performed Western Blot analysis and we observed constitutive activation of both NF-kB and inflammasome. We also demonstrated that IL-1α is able to increase IL-1β production in PDAC cells, with an autocrine loop. Furthermore, treating THP1 cells (a monocytic cell line that secrete IL-1β only upon stimulation) with PDAC cell supernatants we found that both IL-1-expressing and non-IL-1-expressing cells can release inflammasome-activating factors. Since pancreatic cancer is highly hypoxic, we cultured PDAC cell lines in hypoxic conditions and we demonstrated that prolonged severe hypoxia, in combination with starvation, induces an increase in both IL-1α and IL-1β secretion. In conclusion, our study demonstrates that PDAC cells can constitutively express IL-1α, IL-1β and TNF-α, thus inducing TSLP secretion from CAFs and eventually Th2-inflammation. PDAC cells secrete inflammasome-activating factors that sustain cytokine production with an autocrine loop, and that could induce cytokine release from infiltrating macrophages. Finally, the hypoxic environment of pancreatic cancer can further increase IL-1α and IL-1β secretion. Our findings support the development of clinical studies combining the use of Anakinra with conventional chemotherapy, in order to interfere with the IL-1/IL-1R axis and therefore Th2-inflammation, thus improving the prognosis of pancreatic cancer patients. Citation Format: Emanuela Brunetto, Lucia De Monte, Silvia Heltai, Maria Pia Protti. Mechanisms of IL-1 production and release in pancreatic cancer cells. [abstract]. In: Proceedings of the 106th Annual Meeting of the American Association for Cancer Research; 2015 Apr 18-22; Philadelphia, PA. Philadelphia (PA): AACR; Cancer Res 2015;75(15 Suppl):Abstract nr 443. doi:10.1158/1538-7445.AM2015-443