SHP-1 phosphatase activity counteracts increased T cell receptor affinity
Michael Hebeisen, … , Daniel E. Speiser, Nathalie Rufer
Michael Hebeisen, … , Daniel E. Speiser, Nathalie Rufer
Published February 8, 2013
Citation Information: J Clin Invest. 2013;123(3):1044-1056. https://doi.org/10.1172/JCI65325.
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Research Article Immunology

SHP-1 phosphatase activity counteracts increased T cell receptor affinity

Abstract

Anti-self/tumor T cell function can be improved by increasing TCR-peptide MHC (pMHC) affinity within physiological limits, but paradoxically further increases (Kd < 1 μM) lead to drastic functional declines. Using human CD8+ T cells engineered with TCRs of incremental affinity for the tumor antigen HLA-A2/NY-ESO-1, we investigated the molecular mechanisms underlying this high-affinity–associated loss of function. As compared with cells expressing TCR affinities generating optimal function (Kd = 5 to 1 μM), those with supraphysiological affinity (Kd = 1 μM to 15 nM) showed impaired gene expression, signaling, and surface expression of activatory/costimulatory receptors. Preferential expression of the inhibitory receptor programmed cell death-1 (PD-1) was limited to T cells with the highest TCR affinity, correlating with full functional recovery upon PD-1 ligand 1 (PD-L1) blockade. In contrast, upregulation of the Src homology 2 domain-containing phosphatase 1 (SHP-1/PTPN6) was broad, with gradually enhanced expression in CD8+ T cells with increasing TCR affinities. Consequently, pharmacological inhibition of SHP-1 with sodium stibogluconate augmented the function of all engineered T cells, and this correlated with the TCR affinity–dependent levels of SHP-1. These data highlight an unexpected and global role of SHP-1 in regulating CD8+ T cell activation and responsiveness and support the development of therapies inhibiting protein tyrosine phosphatases to enhance T cell–mediated immunity.

Authors

Michael Hebeisen, Lukas Baitsch, Danilo Presotto, Petra Baumgaertner, Pedro Romero, Olivier Michielin, Daniel E. Speiser, Nathalie Rufer

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Figure 6

Levels of SHP-1 protein and of miR-155 and miR-181a expression in CD8+ T cells engineered with TCRs of incremental affinities.

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Levels of SHP-1 protein and of miR-155 and miR-181a expression in CD8+ T...
(A) Unstimulated at baseline (0 hours) and log2 fold change (0–6 hour difference) expression levels of SHP1 transcripts as detected in microarray analysis. (B and C) TCR-transduced CD8+ T cells (B) and SUP-T1 cells (C) were stimulated with 10 μg/ml A2/NY-ESO-1157–165 multimers for the indicated time points and assessed for SHP-1 phosphorylation and total SHP-1 levels by Western blotting. Actin (B) or α-tubulin (C) expression levels were used as loading controls between samples. Data are each representative of 3 independent experiments. For CD8+ transduced T cell samples, all lines were run on the same gel, but were noncontiguous. TCR-untransduced SUP-T1 cells are shown as empty controls (Ø). (D) To allow direct comparison between the engineered SUP-T1 cells, intensity of SHP-1 phosphorylation and of total SHP-1 levels was quantified and normalized to α-tubulin (n = 3 independent experiments). (E) Expression levels of miR-181a and miR-155 were determined by qRT-PCR in TCR-transduced CD8+ T cells following stimulation with 0.1 μg/ml A2/NY-ESO-1157–165 multimers at the indicated time points. Data represent relative expression compared with RNU48 control and are representative of 3 independent experiments. Pri-miR-155 (BIC) mRNA expression values (inset) were retrieved from the microarray analysis.