Abstract 2054: WTX-124 is a novel IL-2 pro-drug that is conditionally activated in tumors and drives anti-tumor immunity by activating tumor infiltrating CD8+ T cells

Abstract Despite the success of immune checkpoint inhibitors (ICI), clinical responses to ICI treatment are still limited to a fraction of patients within specific indications, demonstrating a significant unmet clinical need for novel immunotherapies. In cancer, preclinical and clinical studies have demonstrated the promise of cytokine therapy to stimulate anti-tumor immunity. One of these key cytokines, interleukin-2 (IL-2), is approved for clinical use in metastatic melanoma and renal cell carcinoma. Unfortunately, IL-2 has a poor pharmacokinetic (PK) profile and is linked to serious toxicities, such as vascular leak syndrome (VLS), which limit its clinical utility. To address these limitations, we designed WTX-124, an IL-2 pro-drug (IL-2 INDUKINE™ protein) that takes advantage of dysregulated protease activity in the tumor microenvironment (TME) to selectively release active IL-2 in the tumor after systemic administration. Peripheral inactivation is achieved by linking the cytokine to an inactivation domain using a tumor protease-sensitive linker. WTX-124 is also engineered with a half-life extension element to improve its PK profile and maintain longer exposure in the tumor. Once WTX-124 reaches the tumor, tumor-associated proteases cleave the linkers and release fully active IL-2. Treatment with WTX-124 resulted in the complete rejection of MC38 tumors in a manner dependent on proteolytic processing of the INDUKINE™ protein, as a non-cleavable version of WTX-124 lacked anti-tumor activity. In addition, tumor rejection was dependent on CD8+ T cells, as CD8+ T cell depletion during WTX-124 treatment resulted in a loss of tumor control. Unlike free IL-2, WTX-124 was well tolerated and did not induce VLS in mice, demonstrating the efficacy of the inactivation domain at restricting peripheral toxicity. Due to its enhanced anti-tumor activity and decreased toxicity profile, the therapeutic window of WTX-124 was greatly improved compared to that of native IL-2. In addition, a WTX-124 variant using attenuated (non-α) IL-2 was not active in our efficacy model. Mechanistically, WTX-124 treatment drove the expansion and activation of tumor infiltrating CD8+ T cells and NK cells. Furthermore, WTX-124 treatment resulted in selective activation of the tumor infiltrating CD8+ T cells, with limited evidence of peripheral T cell activation in the blood, spleen, and lymph nodes of tumor bearing animals. In a less immunogenic model, B16F10, WTX-124 also induced CD8+ T cell and NK cell activation and revealed combinatorial anti-tumor activity with αPD-1 therapy. Lastly, in a sample set of over 100 primary human tumor samples, WTX-124 was broadly processed across many indications and disease stages, while remaining intact in the presence of healthy human primary cells from various tissues. Together, these data support the clinical development of WTX-124. Citation Format: Christopher J. Nirschl, Heather R. Brodkin, Daniel J. Hicklin, Nesreen Ismail, Kristin Morris, Andres Salmeron, Cynthia Seidel-Dugan, Philipp Steiner, Zoe Steuert, Jenna M. Sullivan, William M. Winston. WTX-124 is a novel IL-2 pro-drug that is conditionally activated in tumors and drives anti-tumor immunity by activating tumor infiltrating CD8+ T cells [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2022; 2022 Apr 8-13. Philadelphia (PA): AACR; Cancer Res 2022;82(12_Suppl):Abstract nr 2054.