Synergism Between IL21 and Anti-PD-1 Combination Therapy is Underpinned by the Coordinated Reprogramming of the Immune Cellular Network in the Tumor Microenvironment

Abstract

T cell-stimulating cytokines and immune checkpoint inhibitors (ICIs) are an ideal combination for increasing response rates of cancer immunotherapy. However, the results of clinical trials have not been satisfying. It is important to understand the mechanism of synergy between these two therapeutic modalities. Here, through integrated analysis of multiple single-cell RNA sequencing (scRNAseq) datasets of human tumor-infiltrating immune cells, we demonstrate that IL21 is produced by tumor-associated T follicular helper (Tfh) cells and hyperactivated/exhausted CXCL13+CD4+ T cells in the human tumor microenvironment (TME). In the mouse model, the hyperactivated/exhausted CD4+ T cell-derived IL21 enhances the helper function of CD4+ T cells that boost CD8+ T cell-mediated immune responses during PD-1 blockade immunotherapy. In addition, we demonstrated that IL21’s anti-tumor activity did not require T-cell trafficking. Using scRNAseq analysis of the whole tumor-infiltrating immune cells, we demonstrated that IL21 treatment in combination with anti-PD-1 blockade synergistically drives tumor antigen-specific CD8+ T cells to undergo clonal expansion and differentiate towards the hyperactive/exhausted functional state in the TME. In addition, IL21 treatment and anti-PD-1 blockade synergistically promote dendritic cell (DC) activation and maturation to mature DC (mDC) as well as monocyte to macrophage 1 (M1) differentiation in the TME. Furthermore, the combined treatment reprograms the immune cellular network by reshaping cell-cell communication in the TME. Our study establishes unique mechanisms of synergy between IL21 and PD-1-based ICI in the TME through the coordinated promotion of type 1 immune responses.

Enhanced abscopal anti-tumor response via a triple combination of thermal ablation, IL-21, and PD-1 inhibition therapy

“…Considering the pivotal role of CD8 + T cells in augmenting MWA's anti-tumor efficacy through IL-21, we categorized CD8 + T cells into distinct subtypes, including effector, cycling, IFN-induced, exhausted, and stem-like (Fig. 4 A–E) following the classification method outlined in our previously published articles [ 17 ]. We noted a pronounced presence of effector CD8 + T cells within the TME.…”

“…Further investigation is required to explore whether tumor-specific T cells migrate through the lymph nodes into the contralateral TME. We injected mice with FTY720, an S1P receptor inhibitor that blocks T cells trafficking [ 17 ], 2 days before MWA. Blocking lymphocyte efflux counteracted the anti-tumor effect of MWA combined with IL-21 (Fig.…”

“…Simultaneously, the expression of molecules associated with antigen presentation on the surface of DC1 cells increases, the expression of the activating and costimulatory molecules CD86 and PD-L1 was also elevated. Recent research results also indicate that the combination of IL-21 and anti-PD-1 mAbs produces similar effects [ 16 , 17 ]. The anti-tumor mechanism produced by ablation includes the uptake of antigens by DCs and their migration into the TDLN to activate T cells, and the spread of tumor antigen-specific T cells to the contralateral tumor through the TDLN [ 32 ].…”

“…IL-21 is produced by CXCL13 + CD4 + T cells in human colon cancer, liver cancer, and non-small cell lung cancer. This cell population highly expresses molecules such as PD-1 and IFN-γ, and actively responds to PD-L1 blockade treatment [ 17 , 30 ]. These results suggest that the expression of IL-21 in tumor tissues promotes anti-tumor immune responses.…”

“…Under in vitro conditions, IL-21 suppresses Tregs expansion by inhibiting Foxp3 expression [ 11 ], and mice treated with IL-21 did not cause side effects such as in vivo toxicity of vascular leak syndrome even at high concentrations [ 12 , 13 ]. The anti-tumor effects of IL-21 have been extensively studied in various preclinical mouse tumor models, combining IL-21 and immune checkpoint inhibitors exerts more effective anti-tumor effects [ 14 – 17 ]. In addition, recombinant IL-21 has been tested as an anti-tumor agent in various clinical trials, achieving good results in colorectal cancer and metastatic renal cell carcinoma [ 18 , 19 ].…”