Daiad-2/15: A conditionally active, targeted, hyperpotent IL-2/IL-15 immune agonist is safe and effective in vivo

Although high-dose interleukin-2 (IL-2) has been approved for the treatment of certain cancers since 1992, widespread use has been limited due to systemic toxicity, including cytokine release syndrome and vascular leak syndrome. Versions of IL-2 with reduced potency have been studied, but these variants have reduced efficacy as well as toxicity. In this report, we describe Daiad-2/15, a computationally designed, highly potent IL-2/IL-15 agonist that is delivered specifically to tumor-targeting T cells without systemic immune activation. Using our Daiad TM platform technology, a logic gate was engineered by splitting Neo-2/15 into two separate pieces. The individual split Neo-2/15 domains are unable to bind or activate the IL-2 receptor complex. By fusing each portion of the cytokine mimetic to a targeting domain, two distinct modules are created, which together constitute Daiad-2/15. In vitro , Daiad-2/15 activates only T cells that express the targeted antigen, with a 10,000-fold lower threshold for signaling compared to antigen-negative T cells. By targeting two different antigens, the Daiad-2/15 acts as an effective ‘AND' gate, only activating T cells that express both antigens. Furthermore, Daiad-2/15 does not trans-activate target-negative cells, even when present in an admixed culture. The Daiad architecture is compatible with both full length and single-domain antibodies and leads to antigen-specific T cell expansion in vitro . We designed Daiad-2/15 to activate exhausted T cells in the tumor microenvironment using nanobodies specific for PD1 and LAG3. We evaluated the tolerability and antitumor activity of Daiad-2/15 in a stringent syngeneic tumor model, CT26. Individual checkpoint-specific Daiad-2/15 modules had no impact on tumor growth or weight loss, while the wild-type IL-2 and the parental Neo2/15 cytokine mimetic were toxic in vivo . Treating mice with both Daiad-2/15 modules, specific for PD1and LAG3, resulted in significantly improved antitumor activity compared to the two nanobodies alone, with no signs of systemic toxicity. Together, our data suggest that Daiad-2/15 could potentiate the systemic delivery of a highly potent immune agonist to specific immune cells in the tumor microenvironment and draining lymph nodes with a wide therapeutic index. Additional preclinical studies are underway, and clinical testing of Daiad-2/15 is planned.