Abstract B057: The CD122-biased immunostimulatory cytokine NKTR-214 combined with checkpoint blockade leads to mobilization of antitumor immunity and synergistic activity

Background: NKTR-214 is a novel immunostimulatory cytokine which provides a sustained and biased activation signal through the heterodimeric IL-2R receptor (IL2Rbg or CD122), resulting in significant expansion of CD8+ T and NK cells relative to regulatory T cells in the tumor microenvironment. As a single agent in preclinical tumor models, NKTR-214 significantly increases the intratumoral CD8T/Treg ratio (>400). Here we examine activity and pharmacodynamic effects of NKTR-214 alone and combined with immune checkpoint blockade in both a murine tumor model and in a human in vitro co-culture system. Methods: Mice bearing subcutaneous CT26 colon tumors were treated with single agent NKTR-214 (q9d), murine anti-CTLA-4 or anti-PD-1 (twice-weekly), or combinations of these agents. Serum cytokines were assessed by multiplex ELISA (Quansys). T cell clonality and infiltration were assessed in tumors 7 days after treatment initiation, with TCR V and J beta usage determined by ImmunoSEQ (Adaptive Biotechnologies). Effects on human cytokine and chemokine expression were examined by the BioMAP Combo ELECT (DiscoverX) in vitro system utilizing human fibroblasts or endothelial cells, PBMCs and either HT29 colon adenocarcinoma or HT1299 NSCLC cell lines. 1-PEG-IL2, the most active species of the NKTR-214 prodrug, or Keytruda (anti-PD-1) was added over a range of concentrations alone or in combination, with biomarkers assessed 48 hours later. Results: In tumor-bearing mice, NKTR-214 led to increases in serum IFNg as well as increases in the chemokine MCP-1 (CCL2), a chemoattractant of CD4 T, CD8 T and NK cells. Single-agent NKTR-214 led to significant tumor growth inhibition and 10% tumor free animals, while single-agents anti-CTLA-4 or anti-PD-1 lacked efficacy. Combination of NKTR-214 with checkpoint blockade was synergistic and led to greater tumor-free animals (67% in combination with anti-CTLA-4, 90% with anti-PD-1). NKTR-214 combined with anti-PD-1 was also more efficacious than the combination of anti-CTLA-4 and anti-PD-1. TCR repertoire analysis demonstrated superior increases in TIL clonality and infiltration with NKTR-214 compared to either anti-CTLA-4 or anti-PD-1 alone. NKTR-214 combined with either mode of checkpoint inhibition led to significant increases in both parameters, with the greatest effect occurring when combined with anti-PD-1. In the human BioMAP system, NKTR-214 active species led to significant increases in granzyme B, IFNg, IL-6, IL-17A and TNFa