Design and Characterization of a Protein Superagonist of IL-15 Fused with IL-15Rα and A High-Affinity T Cell Receptor

摘要

To avoid high systemic doses, strategies involving antigen-specific delivery of cytokine via linked antibodies or antibody fragments have been used. Targeting cancer-associated peptides presented by major histocompatibility complex (MHC) molecules (pepMHC) increases the number of potential target antigens and takes advantage of cross-presentation on tumor stroma and in draining lymph nodes. Here, we use a soluble, high-affinity single-chain T cell receptor Vα-Vβ (scTv), to deliver cytokines to intracellular tumor-associated antigens presented as pepMHC. As typical wild-type T cell receptors (TCRs) exhibit low affinity (K_d = 1—100 pM or more), we used an engineered TCR, m33, that binds its antigenic peptide SIYRYYGL (S1Y) bound to the murine class I major histocompatability complex protein H2-K~b (SIY/K~b) with nanomolar affinity (K_d = 30 nM). We generated constructs consisting of m33 scTv fused to murine interleukin 2 (IL-2), interleukin 15 (IL-15), or IL-15/IL-15Rα (IL-15 linked to IL-15Rα. sushi domain, called "superfusion"). The fusions were purified with good yields and bound specifically to SIY/K~B with high affinity. Proper cytokine folding and binding were confirmed, and the fusions were capable of stimulating proliferation of cytokine-dependent cells, both when added directly and when presented in trans, bound to cells with the target pepMHC. The m33 superfusion was particularly potent and stable and represents a promising design for targeted antitumor immunomodulation.