Gao, Jinming2024-06-072024-06-072022-052022-05May 2022https://hdl.handle.net/2152.5/10300Cytokines are secreted molecules that guide the immune system to respond correctly to various challenges. Among all cytokines, IL12 is perhaps the most powerful at polarizing the immune response into a Type 1, cell-mediated phenotype. Cell-mediated immunity plays a critical role in cancer immunoediting, allowing CTLs to recognize and kill aberrant cells. Because of this, IL12 has been tested in many different preclinical and clinical studies for its potential use as an anti-tumor therapeutic agent. However, IL12 also causes severe, dose-limiting systemic toxicity due to on-target, off-tumor activation of peripheral immune cells. Newer attempts at IL12-mediated delivery focus on restricting IL12 activity to within the tumor as much as possible, but they each have their own limitations. To address these problems, we developed a novel IL12 prodrug, pro-IL12, that is actively blocked until it is preferentially activated within the TME. We achieved this by using portions of the IL12 receptor attached with a flexible linker to sterically block the active site of IL12. The linker contains a substrate site that can be cleaved by tumor-specific proteinases, thereby releasing the blocker and activating the prodrug. Pro-IL12 successfully maintained anti-tumor efficacy with reduced toxicity compared to its non-prodrug counterpart. We determined that the mechanism of anti-tumor immunity was predominantly through pre-existing, intratumor CD8+ T cells that produce IFNγ after direct binding of the prodrug to cell surface IL12 receptor complexes. Pro-IL12 also worked in combination with TKI and ICB to achieve even more potent tumor control. In a follow up study, I propose that a higher dose of pro-IL12 might use distinct cellular and molecular mechanisms. Indeed, high dose pro-IL12 more effectively controls large tumors at the cost of reintroducing systemic toxicity. Mechanistically, this dose used a broader, T cell-dependent mechanism that was independent of IFNγ. Further analysis determined that IFNγ was responsible for all manifestations of toxicity and that IFNγ blockade given concurrently with pro-IL12 could limit toxicity with no effect on efficacy. Additionally, the absence of IFNγ signaling on T cells had no effect on their phenotype or ability to control the tumor. As a whole, these studies document the development of a next generation, IL12 immunotherapy for the treatment of solid tumors with an emphasis on its mechanisms of tumor control that are distinct from toxicity.application/pdfenCD8-Positive T-LymphocytesImmunotherapyInterleukin-12NeoplasmsProdrugsThesis2024-06-071438579251