A Fusion of GMCSF and IL-21 Initiates.pdf

original article? The American Society of Gene Cell Therapy Molecular Therapy vol. 18 no. 7, 1293–1301 july 2010 1293 We hypothesized that fusing granulocyte-macrophage colony-stimulation factor (GMCSF) and interleukin (IL)-21 as a single bifunctional cytokine (hereafter GIFT-21) would lead to synergistic anticancer immune effects because of their respective roles in mediating inflammation. Mechanistic analysis of GIFT-21 found that it leads to IL-21Rα-dependent STAT3 hyperactivation while also contemporaneously behaving as a dominant- negative inhibitor of GMCSF-driven STAT5 activation. GIFT-21’s aberrant interactions with its cognate recep- tors on macrophages resulted in production of 30-fold greater amounts of IL-6, TNF-α, and MCP-1 when com- pared to controls. Furthermore, GIFT-21 treatment of primary B and T lymphocytes leads to STAT1-dependent apoptosis of IL-21Rα+ lymphocytes. B16 melanoma cells gene-enhanced to produce GIFT-21 were immune rejected by syngeneic C57Bl/6 mice comparable to the effect of IL-21 alone. However, a significant GIFT-21- driven survival advantage was seen when NOD-SCID mice were implanted with GIFT-21-secreting B16 cells, consistent with a meaningful role of macrophages in tumor rejection. Because GIFT-21 leads to apoptosis of IL-21Rα+ lymphocytes, we tested its cytolytic effect on IL-21Rα+ EL-4 lymphoma tumors implanted in C57Bl/6 mice and could demonstrate a significant increase in sur- vival. These data indicate that GIFT-21 is a novel IL-21Rα agonist that co-opts IL-21Rα-dependent signaling in a manner permissive for targeted cancer immunotherapy. Received 7 December 2009; accepted 2 March 2010; published online 13 April 2010. doi:10.1038/mt.2010.49 IntroductIon Cellular tumor vaccines can be generated by transfecting autologous or allogeneic tumor cells with cDNAs encoding for interleukins (ILs), cytokines, interferons, and molecules accessory to immune activation.1 The mechanism, by which the secre