Local delivery of IL-12 mRNA and indoximod prodrug potentiates antitumor immunity by increasing T cell effector function

The administration of recombinant cytokines, particularly interleukin-12 (IL-12), holds promising clinical potential for treating various cancers. Sustained intratumoral delivery of IL-12 can restore tumor resident CD8⁺ effector T cells and induce the priming of antitumor CD8⁺ effector T cells. However, these CD8⁺ T cell-dependent anticancer efficacy is usually transient and accompanies the activation of immune suppressive CD4⁺Foxp3⁺ T regulatory cells. The underlying mechanism of T regulatory cell activation in IL-12 therapy is the upregulation of IFNγ dependent indoleamine 2,3-dioxygenase (IDO) expression. Due to this negative feedback, the combinatorial use of drugs should be considered to enhance the efficacy of IL-12-mediated therapy. Herein, we designed a lipid nanoparticle (LNP) system which can successfully deliver an IDO inhibitor indoximod (IND) and IL-12 encoding mRNA. In order to increase the loading efficiency, the IND prodrug was synthesized by conjugating IND with cholesterol by ester linkage. Optimized IND prodrug encapsulating LNP successfully transfected tumor cells and macrophages, resulting in the secretion of IL-12 cytokine. With IL-12 transfection, macrophages upregulated T cell co-stimulation factor and released TNFα cytokine, indicating that the tumor microenvironment could be changed from cold tumor to hot tumor for immunotherapy. Also, by the elevated secretion of IL-12 cytokine, T cells release high levels of IFNγ, which is a central role in IL-12-mediated immunotherapy. This co-delivery system presents a promising strategy to overcome the limitations of single IL-12-mediated therapy by simultaneously promoting antitumor immune responses and inhibiting immunosuppressive mechanisms, thereby enhancing the overall efficacy of cancer immunotherapy.