Fine-Tuning of Optimal TCR Signaling in Tumor-Redirected CD8 T Cells by Distinct TCR Affinity-Mediated Mechanisms

Abstract

Redirecting CD8 T cell immunity with self/tumor-specific affinity-matured T cell receptors (TCRs) is a promising approach for clinical adoptive T cell therapy, with the aim to improve treatment efficacy. Despite numerous functional-based studies, little is known about the characteristics of TCR signaling (i.e., intensity, duration, and amplification) and the regulatory mechanisms underlying optimal therapeutic T cell responses. Using a panel of human SUP-T1 and primary CD8 T cells engineered with incremental affinity TCRs against the cancer-testis antigen NY-ESO-1, we found that upon activation, T cells with optimal-affinity TCRs generated intense and sustained proximal (CD3ζ, LCK) signals associated with distal (ERK1/2) amplification-gain and increased function. In contrast, in T cells with very high affinity TCRs, signal initiation was rapid and strong yet only transient, resulting in poor MAPK activation and low proliferation potential even at high antigen stimulation dose. Under resting conditions, the levels of surface TCR/CD3ε, CD8β, and CD28 expression and of CD3ζ phosphorylation were significantly reduced in those hyporesponsive cells, suggesting the presence of TCR affinity-related activation thresholds. We also show that SHP phosphatases were involved along the TCR affinity gradient, but displayed spatially distinct regulatory roles. While PTPN6/SHP-1 phosphatase activity controlled TCR signaling initiation and subsequent amplification by counteracting CD3ζ and ERK1/2 phosphorylation, PTPN11/SHP-2 augmented MAPK activation without affecting proximal TCR signaling. Together, our findings indicate that optimal TCR signaling can be finely tuned by TCR affinity-dependent SHP-1 and SHP-2 activity, and this may readily be determined at the TCR/CD3 complex level. We propose that these TCR affinity-associated regulations represent potential protective mechanisms preventing high affinity TCR-mediated autoimmune diseases.

Revisiting the PD-1 pathway

“…Although SHP-1 mediates inhibitory signals ( 78 ), SHP-2 is considered a positive regulator of T cell activation ( 79 – 81 ) and may act by dephosphorylating inhibitory sites of positive regulators. Both phosphatases are involved in the formation of the TCR signalosome ( 82 ), and their phosphatase activities are regulated by phosphorylation/dephosphorylation as well as by oxidation/reduction reactions ( 83 – 85 ). Since PD-1 expression is induced only after activation and participates in TCR signaling microclusters upon binding to PD-L1 ( 63 ), it is reasonable to speculate that PD-1 molecules are clustered and stabilized by PD-L1 interactions around TCR molecules, serving as “switches” of positive signalosome conformations to negative ones.…”

Long-term surviving cancer patients as a source of therapeutic TCR

“…TCR affinity increased beyond the physiological range comes with the risk of creating novel molecules with unpredictable binding specificities to structurally related peptide sequences from different antigens, creating off-target toxicity [8,9]. Moreover, TCR variants were tested and the existence of a TCR affinity-related activation threshold was demonstrated both in vitro [14] and in vivo [15]. Here the authors of the two studies proposed that this threshold marked the limit between epitope recognition and autoimmunity.…”

T‐cell receptor affinity in the age of cancer immunotherapy

“…Using the OT‐1 APL system, low‐affinity–primed memory cells produce more IL‐2 and fully differentiated memory cells that are CD45RB HI 68 . Using specific T cells with low and high affinity for the NY‐ESO‐1 TAA peptide complexed with HLA‐A2, the high‐affinity interactions resulted in transient signal activation leading to poor MAPK production and low proliferation ability 69 . Taken together, these data suggest that low‐affinity T cells have a separate, but necessary role in T‐cell function (summarized in Figure 2).…”