Multi-faceted immunostimulatory therapy for Ewing Sarcoma (MITES)
Ewing sarcoma (ES) is the second most pediatric common bone cancer (1,2). While patients with localized disease have a five-year survival rate of ~75%, this plummets to 20% for metastatic and recurrent disease. Immunotherapy has had limited success in ES, in large part because it is considered immunologically “cold,” with few tumor infiltrating T lymphocytes (TILs) (3-5). However, TILs actually vary significantly, and high levels portend greatly improved survival (6,7). TIL abundance correlated with the interferon (IFN)g-inducible chemokines CXCL9/CXCL10 and CCL5 (6). These immune features (i.e. IFNg response, CXCL9/10 production, and high CD8+ TIL recruitment) define a “hot” TME, and predict both improved overall survival and response to checkpoint inhibitors across many malignancies (8,9). However, the tumor-intrinsic factors that confer this favorable immune phenotype, in ES or any other cancer, are largely unknown, and their identification represents a window of opportunity for therapeutic intervention to improve patient outcome. We identify USP6 as a potent hot TME agonist with pleiotropic immunostimulatory effects in ES (10). In vitro, USP6: a) directly induces an IFN gene signature (including CXCL9/10 and CCL5); b) synergistically magnifies expression of these and other chemokines in response to ectopic IFNs; c) and directly stimulates cytolytic activity of natural killer (NK) cells. In vivo, USP6 inhibits ES tumor growth in nude mice, and enhances infiltration/activation of NK cells and antigen-presenting cells (APCs). Furthermore, in ES patients USP6 expression is associated with significantly improved survival, IFN signature, CXCL10 production, and infiltration of CD8+ T cells and NK cells. We hypothesize that USP6 improves ES patient outcome due to its multi-faceted immunostimulatory functions, which engender a hot TME with resultant activation of multiple cytolytic immune lineages that effect tumor cell elimination. Our goal is to harness the multi-faceted immune-igniting properties of USP6 into a novel immunotherapeutic by delivering USP6 mRNA using lipid nanoparticles (LNPs) customized for in vivo delivery into ES cells. Preliminary data strongly support the feasibility of this approach, and indicate that USP6 levels can dictate the degree of immune infiltration. Herein, we will establish a pre-clinical mouse model to test the efficacy of USP6 LNPs in inhibiting both primary tumor growth as well as disseminated disease, and validate whether anti-tumorigenic effects arise through immune activation, particularly NK cells. If successful, USP6 LNPs could be readily used in other solid tumors, greatly broadening their impact. Furthermore, we predict that USP6 LNPs would serve not only as an effective monotherapy, but we posit that by modulating the immune tumor microenvironment (TME), they could greatly enhance the efficacy and utility of existing immunotherapies, such as CAR-T cells and immune checkpoint inhibitors.