Ubiquitin-Specific Protease 6 mRNA Lipid Nanoparticles Ignite Antitumor Immunity and Suppress Tumorigenesis in Ewing Sarcoma.

Ewing sarcoma is an aggressive pediatric cancer that has remained refractory to current therapeutics. Immunotherapy has been unsuccessful in Ewing sarcoma, largely due to poor understanding of how its immune tumor microenvironment is regulated. We recently demonstrated that ubiquitin-specific protease 6 (USP6) can remodel the Ewing sarcoma immune landscape to engender an antitumorigenic tumor microenvironment. USP6 expression in Ewing sarcoma cells enhances surface expression of immunostimulatory ligands and receptors and induces production of multiple chemokines, driving recruitment and activation of tumor-suppressive immune lineages, including NK cells. We sought to harness this multifaceted immunostimulatory function into a novel therapeutic by delivering in vitro transcribed USP6 mRNA via ionizable lipid nanoparticles (LNP). Treatment of Ewing sarcoma cells with USP6 mRNA in vitro is capable of inducing the aforementioned antitumorigenic and immunostimulatory responses. In addition, USP6 mRNA-treated Ewing sarcoma cells elicit cytolytic activation of primary human CD8+ and CD4+ T lymphocytes and NK cells in vitro. Intratumoral delivery of USP6 mRNA LNPs suppresses growth of Ewing sarcoma xenografts, coincident with increased immune infiltration and activation. We further demonstrate that USP6 mRNA is capable of igniting an immunostimulatory program in other cancer types (including acute myeloid leukemia, melanoma, prostate cancer, head and neck cancer, and osteosarcoma) in vitro and suppressing acute myeloid leukemia xenograft growth in vivo. Treatment with USP6 mRNA LNPs was well-tolerated, with no observed gross toxicity. Together, these preclinical studies provide proof-of-concept for the immunogenic and antitumorigenic efficacy of USP6 mRNA LNPs and support its promise as a novel immunotherapeutic in diverse cancer types.