Enhancing the Generation of Eomeshi CD8+ T Cells Augments the Efficacy of OX40- and CTLA-4-Targeted Immunotherapy

Abstract

CTLA-4 blockade in combination with an agonist OX40-specific monoclonal antibody synergizes to augment antitumor immunity through enhanced T-cell effector function, leading to increased survival in preclinical cancer models. We have shown previously that anti-OX40/anti-CTLA-4 combination therapy synergistically enhances the expression of Eomesodermin (Eomes) in CD8⁺ T cells. Eomes is a critical transcription factor for the differentiation and memory function of CD8⁺ T cells. We hypothesized that Eomes^hiCD8⁺ T cells were necessary for anti-OX40/anti-CTLA-4 immunotherapy efficacy and that further enhancement of this population would improve tumor-free survival. Indeed, CD8⁺ T cell-specific deletion of Eomes abrogated the efficacy of anti-OX40/anti-CTLA-4 therapy. We also found that anti-OX40/anti-CTLA-4-induced Eomes^hiCD8⁺ T cells expressed lower levels of checkpoint receptors (PD1, Tim-3, and Lag-3) and higher levels of effector cytokines (IFNγ and TNFα) than their Eomes^lo counterparts. Eomes expression is negatively regulated in T cells through interleukin-2-inducible T-cell kinase (ITK) signaling. We investigated the impact of modulating ITK signaling with ibrutinib, an FDA-approved tyrosine kinase inhibitor, and found that anti-OX40/anti-CTLA-4/ibrutinib therapy further enhanced CD8⁺ T cell-specific Eomes expression, leading to enhanced tumor regression and improved survival, both of which were associated with increased T-cell effector function across multiple tumor models. Taken together, these data demonstrate the potential of anti-OX40/anti-CTLA-4/ibrutinib as a triple therapy to improve the efficacy of immunotherapy.