Document resource
Background The development of IL-2 analogs for cancer immunotherapy has primarily focused on reducing systemic toxicity by limiting peripheral regulatory T cell (T reg) expansion. To achieve this, several IL-2 analogs have been engineered to eliminate binding to IL-2Rα (CD25). However, they have failed to demonstrate sufficient safety and efficacy in clinical trials. Given the intrinsic role of Tregs in maintaining immune homeostasis and regulating excessive immune activation, Tregs may function as a critical safeguard during IL-2 therapy. Here, we investigated the critical role of CD25 binding within HM16390 in promoting peripheral Treg expansion and reducing systemic toxicity.Methods Male C57BL/6 mice were pretreated with either anti-CD25 antibody (for Treg depletion) or isotype control. One week later, mice received a single subcutaneous (SC) administration of HM16390 at a near maximum tolerated dose or a non-alpha variant at its LD 100. Circulating immune cells and lung pathology were analyzed on days 1, 3, and 7. Cynomolgus monkeys received both compounds via stepwise SC dosing every three weeks, with peripheral immune profiling and toxicological assessments.Results HM16390 induced comparable CD8 + T cell expansion in both Treg-depleted and isotype control mice; however, peripheral Treg dynamics differed substantially. By day 3 post-treatment, Treg increased to 23.4% in isotype controls whereas it remained suppressed at 8.8% in Treg-depleted mice (37.6% of the isotype control), confirming the effective establishment of the Treg-depletion model. Despite being administered at the same dose, HM16390 elicited distinct pulmonary pathologies depending on Treg status. In Treg-depleted mice, HM16390 treatment resulted in a significantly elevated lung wet-to-dry ratio (6.3 ± 0.9), compared to the isotype control (5.2 ± 0.2, within the normal range), and progression to moderate-grade inflammation, indicative of transient but pronounced pulmonary edema. Notably, these adverse effects were comparable to those observed in isotype control mice treated with the non-alpha variant (6.6 ± 0.7). Similarly, in cynomolgus monkeys, the non-alpha variant induced a rapid surge in CD8+ T cells without accompanying Treg modulation, which was associated with poor tolerability and lethality at high doses. In contrast, HM16390 induced a dose-dependent increase in Tregs, which appeared to support a gradual and sustained expansion of CD8+ T cells with improved tolerability.Conclusions Unlike non-alpha variants, HM16390 preserves T reg activation while expanding CD8+ T cells, leading to improved safety and tolerability in both murine and primate models. These findings support fine-tuned CD25 binding as a promising approach to enhance the therapeutic index of IL-2-based cancer immunotherapy.