Abstract 159: Macrophage Migration Inhibitory Factor (MIF) enhances proliferation and expression of tumorigenic cytokines in a bladder cancer cell line

Abstract

Abstract Background: MIF is a proinflammatory cytokine with regulatory properties over tumor suppressor proteins involved in bladder cancer. We have previously reported that the absence of MIF in transgenic mice or the use of a small molecule oral inhibitor of MIF (CPSI-1306, Cytokine PharmaSciences, Inc, King of Prussia, PA) leads to decreased angiogenesis and invasive potential in the BBN mouse model of bladder cancer. The objective of this study was to evaluate potential pathways of MIF activity in a cell culture model. Methods: High grade, metastatic HTB-5 bladder cancer cells were treated with rhMIF 1-100 ng/ml (CPSI). Measurements included cell counts, proliferation by 3H-thymidine incorporation (TdR), phosphorylated and total ERK by Western analysis and mRNA expression of TNF-α, IL-6, VEGF and CD74 (putative receptor for MIF) by real time PCR. CPSI-1306 (0.5-500 nM) was tested for its ability to decrease ERK phosphorylation. Results: MIF 100 ng/ml increased cell counts 2-fold (p<0.01) on culture days 2-4. MIF increased TdR, normalized to cell numbers, 30% (p<0.01). MIF (1-100 ng/ml) stimulated ERK phosphorylation in a dose dependent manner, which was abrogated by the addition of CPSI-1306 at doses above 5 nM. A specific inhibitor of ERK (PD98059, 50 μm) blocked the MIF stimulated increase in cell number. MIF treatment for 24 hrs increased mRNA expression of the agiogenic factor VEGF and the cytokine TNF-α 2-fold (p<0.01). IL-6 mRNA was unchanged. mRNA levels for the putative MIF receptor, CD74, increased 1.3 fold with treatment (p=0.01). Conclusion: MIF increased ERK phosphorylation, which was blocked by CPSI-1306, a small molecule inhibitor of MIF. MIF also increased cell proliferation via an ERK pathway. These data, along with the increased cytokine expression, provide a potential explanation for the decreased angiogenesis and invasion seen in our in vivo murine models using global knockout of MIF or CPSI-1306. This study is the first to begin to elucidate mechanistic pathways involved in MIFs role in bladder cancer and possible pathways involved in CPSI-1306 inhibition. Funding Source: ACS MRSG-08-270-01-CCE, NIH RO1DK48361 Citation Format: {Authors}. {Abstract title} [abstract]. In: Proceedings of the 103rd Annual Meeting of the American Association for Cancer Research; 2012 Mar 31-Apr 4; Chicago, IL. Philadelphia (PA): AACR; Cancer Res 2012;72(8 Suppl):Abstract nr 159. doi:1538-7445.AM2012-159