LAUSR

Background Previously, we demonstrated that radiation therapy (RT) combined with Bempegaldesleukin (BEMPEG;NKTR-214), a first-in-class CD122-preferential IL-2 pathway agonist, led to enhanced anti-tumor efficacy through a T cell-dependent mechanism. However, we observed only modest systemic responses to BEMPEG/RT across several murine tumor models. Therefore, we explored alternative approaches to improve systemic tumor-specific immunity. We evaluated whether intratumoral NKTR-262, a polymer-modified toll-like receptor (TLR) 7/8 agonist, combined with systemic BEMPEG treatment resulted in improved tumor-specific immunity and survival compared to BEMPEG combined with RT. We hypothesized that BEMPEG/NKTR-262 immunotherapy would promote synergistic activation of local immunostimulatory innate immune responses followed by systemic adaptive immunity to significantly improve tumor regression and overall survival. Methods Tumor-bearing mice (CT26; EMT6) received BEMPEG (0.8 mg/kg; iv), RT (12 Gy x 1), and/or intratumoral NKTR-262 (0.5 mg/kg). Flow cytometry was used to evaluate CD4+ and CD8+ T cell activation status in the blood and/or tumor (7 days post-treatment) and NK cell activity in the tumor (1, 3 days post-treatment). The contribution of specific immune subsets was determined by depletion of CD4+, CD8+, or NK cells. CD8+ T cell activity was determined in vitro by tracking apoptosis in an Incucyte assay. Data are representative of 1–2 independent experiments (n=5–14/group) and statistical significance was determined by 1-way ANOVA (p-value cut-off of 0.05). Results BEMPEG/NKTR-262 resulted in significantly improved survival compared to BEMPEG/RT. BEMPEG/NKTR-262 efficacy was NK and CD8+ T cell-dependent, while BEMPEG/RT primarily relied on CD8+ T cells. Response to BEMPEG/NKTR-262 was characterized by a significant expansion of activated CD8+ T cells (GzmA+; Ki-67+; ICOS+; PD-1+) in the blood, which correlated with reduced tumor size (p Conclusions Combining BEMPEG with NKTR-262 lead to an early and robust NK cell expansion not observed in the BEMPEG/RT combination. The improved tumor regression and survival was dependent on the NKTR-262 driven expansion of NK cells. A clinical trial of BEMPEG/NKTR-262 for patients with metastatic solid tumors is in progress (NCT03435640).