Engineered Variants Of Neo-2/15 Potently Expand Car-T Cells And Promote Antitumor Activity In Lymphoma And Solid Tumor Mouse Models

Chimeric antigen receptor (CAR) T cells have achieved remarkable success as therapies for B cell malignancies, but failure might occur due to insufficient CAR-T cell expansion. Furthermore, CAR-T cells have been less effective in treating epithelial cancers, which is believed to be due to poor CAR-T cell accumulation at the tumor site. Interleukin-2 (IL-2) is a potent signaling molecule that induces T cell proliferation and survival. Therefore, it has been suggested that IL-2 co-administration might be effective in promoting CAR-T expansion, persistence, and tumor infiltration. Natural IL-2 acts by binding to the trimeric IL-2 receptor complex consisting of the IL-2Rα, IL-2Rβ and common-γ subunits. Although the IL-2Rα subunit is not directly involved in the signaling cascade, when present, it greatly increases the binding (and signaling) potency of IL-2. An obstacle to applying IL-2 to CAR-T therapy is the expression of IL-2Rα on many target-cells, such as immunosuppressive regulatory T cells (Treg) and endothelial cells. Consequently, high doses of IL-2 cause severe toxicities, including vascular leak syndrome, while low doses are therapeutically suboptimal due to preferential expansion of Treg cells. In addition, as a therapeutic, IL-2 has poor pharmacokinetics, necessitating frequent dosing or continuous infusion. Neoleukin-2/15 (Neo-2/15), a recently reported de novo IL-2 and IL-15 mimetic, effectively agonizes the IL-2 receptor IL-2Rβγ and has no dependency (or interaction) with IL-2Rα. Therefore, Neo-2/15 can activate T cells with high potency regardless of IL-2Rα expression. Here, we studied the combination of CAR-T cells with two reengineered versions of Neo-2/15. In the first strategy, NL-201, a half-life-extended version of Neo-2/15, was dosed weekly (300μg/kg) in a B cell lymphoma xenograft model. NL-201 induced potent CD19 CAR-T cell expansion and prolonged mice survival. NL-201 also induced potent ROR1 CAR-T cell expansion in a solid tumor syngeneic model of mice engrafted with the mutant K-ras/p53-/- tumor line overexpressing hROR1. NL-201 significantly increased the frequency of CAR-T cells at the tumor site compared to IL-2 and also dramatically increased the CD8:Treg ratio in the tumor. In the second approach, Neo-2/15 was targeted to engineered CAR-T cells by fusing the molecule to a DARPin domain specific for a transduction marker co-expressed with the CAR. In a B cell tumor xenograft model, targeted Neo-2/15 effectively increased CAR-T cell expansion and prolonged survival compared to treatment with CAR-T cells and non-targeted Neo-2/15. Together, these results show that engineered versions of Neo-2/15 induce robust expansion of CAR-T cells and can enhance their antitumor activity. Citation Format: Isabel Leung, Megan Templeton, Alfredo Rubio, Carl Walkey, Daniel-Adriano Silva, Stanley Riddell. Engineered variants of Neo-2/15 potently expand CAR-T cells and promote antitumor activity in lymphoma and solid tumor mouse models [abstract]. In: Proceedings of the Annual Meeting of the American Association for Cancer Research 2020; 2020 Apr 27-28 and Jun 22-24. Philadelphia (PA): AACR; Cancer Res 2020;80(16 Suppl):Abstract nr 2222.