Combined PD-L1/TGFβ blockade attenuates T-cell exhaustion

Introduction

Cancer Immunotherapy holds great promise, but patients with excluded tumours (CD8 T cells accumulate at the tumour–stroma boundary) show increased resistance to check-point blockade compared to patients with inflamed tumours (CD8 T cells infiltrate the tumour parenchyma).

Chronic antigen exposure in cancer and other diseases induces T cells to enter an exhausted/dysfunctional state characterized by gradual loss of effector functions and increased levels of co-inhibitory receptors.

In this study, Castiglioni et al, used a multi-omics approach to unravel how PD-L1 and TGFβ affect the CD8 T cell phenotypes and impact the tumour micro environment (TME).

Main Points

• The authors have previously reported that TGFβ hampers response to anti-PD-L1 treatment in a CD8 T cell-dependent manner.
• Using a syngeneic immune-excluded EMT6 breast tumor model, the authors found that combining therapeutic anti-PD-L1 with anti-TGFβ treatment (neutralizing all three active TGFβ isoforms) promoted tumor regression and survival compared to control and single agent.
• Single cell RNA sequencing (scRNA-sec) of intratumoral CD8 T cells isolated from day 7 tumors identified eleven clusters among the 25,063 sorted TCRb⁺ T cells.
• An in-depth analysis of the non-naïve CD8 T cell compartment revealed eight transcriptional clusters, which were characterized by comparing their gene expression profiles to known transcriptional markers of T cell function.
• Based on this analysis the presence of stem cell-like (T_SCL ) CD8 T cells (PD-1^lowLAG3⁻TIM3⁻) and progenitor-exhausted (T_PEX) CD8 T cells (PD-1^hiLAG3⁺TIM3⁻) in the tumors were confirmed using flow cytometry.
• Comparing the abundance of different CD8 T cell subsets between treatment conditions showed that dual blockade of TGFβ and PD-L1 increased the abundance of T_SCL at the expense of T_PEX CD8 T cells.
• Furthermore, combined PD-L1 andTGFβ blockade allowed for T_SCL expansion in the tumor.
• Adding a neutralizing IFNγ antibody to anti-PD-L1/TGFβ treatment completely inhibited the anti-tumor response caused by the combination, demonstrating the IFNγ dependency of the dual blockade.
• In addition, when IFNGR1 was knocked out in EMT6 tumor cells, they retained their ability to form tumors in vivo, but the anti-tumor efficacy of the anti-PD-L1/TGFβ combination decreased, supporting the importance of the IFNγ response in inducing tumor rejection in combination treatment.

Concluding Remarks

In summary, dual blockade successfully converted the EMT6 immune-excluded phenotype into an inflamed tumor. T_SCL cells expanded in tumors following combination therapy, underlining their potential to fuel the anti-tumor response. Furthermore, they may give rise to a high IFNγ-expressing T effector cell pool and TME interferon licensing, while the relative abundance of T_PEX is decreased.

Epistem Services

Epistem offers several pre-clinical human cancer models (including breast) to assess the efficacy of cancer immunotherapies as single agents or in combination. Furthermore, there is a routine deployment of multi-modal assessment studies including scRNA-seq of a wide variety of tissues and cells utilising a range of expertise to delineate cellular compartmentalisation and function. To aid in the assessment we have a 15 colour Novocyte flow cytometer, which gives the opportunity to evaluate treatment effects and elucidate the function of specific immune cell types ex vivo.