Background The CAR T cell therapy initiated as a treatment for CD19⁺ malignant B cell has rapidly expanded to many other cancers including solid tumors. Among multiple key factors to improve efficacy while minimizing adverse effects, it is critical to understand the mechanisms of T cell exhaustion and harness it. Upregulation of TIGIT is a hallmark of exhausted T cells in non-responders among non-Hodgkin’s lymphoma (NHL) patients under CD19⁺ CAR T therapy.¹ However, we have not paid attention to PBMC that can (in)directly interact with CAR⁺T cells and its synergistic role in the treatment result. Furthermore, it is desirable to identify transcriptional factors (TFs) facilitating the unexpected T cell status. Ultimately, we can target them with TIGIT for better outcomes.

Methods We evaluate 27 scRNA-seq data for CAR⁺T (n=12) and its matched PBMC (n=15) in two sequential time points after the infusion from 8 relapsed/refractory NHL patients; 6 responders (R) and 2 non-responders (NR).¹

Results TIGIT is upregulated in post-infusion PBMC samples. The TIGIT is overexpressed across CD8+T, NK, and most significantly Treg population. DEG analysis reveals that more NR cells activate TIGIT whereas more R cells activate CD226. Furthermore, NECTIN2 is mostly expressed in a monocyte subpopulation dominated by the NR group. CellphoneDB analysis³ demonstrates that KLRC1 in an inflammatory T population in CAR+T interacts with HLA-E detected across PBMC cells (potentially originated from malignant B cells). These suggest that more prevalent immune checkpoints in NR trigger immune evasion.^4–7 Next, we investigate how the dysfunctional T state was acquired. A gene set analysis uncovers that it is due to chronic antigen exposure (CAE).² NK-like T cell populations resemble dysfunctional tumor-infiltrated lymphocytes (TILs) under CAE. Among monocyte populations, the NR-dominant monocytes upregulate TFs represented into CAE TILs. A comprehensive TF analysis reveals that TSC22D3, ZFP36, and DUSP1 known for anti-inflammatory response⁸ are strikingly overexpressed in the NR group. A higher interaction of TSC22D3 with AP-1 proteins results in a lack of AP-1 to form a binding with NFAT required for T cell effectors. Even worse, NR patients upregulate NR4A2 expression. The gene potentially interferes with the binding of NFAT/AP-1 and remodels chromatin accessibility to facilitate T cell exhaustion.⁹

Conclusions In conclusion, dysregulated TSC22D3 and NR4A2 compromise the effector function of CAR⁺T, hijack AP-1 proteins, and impede eradicating the antigens, and thus CAR⁺T cells transit to exhausted T cells represented by overexpressed TIGIT.

References

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