Cytotoxicity of Tumor Antigen Specific Human T Cells Is Unimpaired by Arginine Depletion

Markus Munder,Hartmut Goldschmidt,Pascale Kropf,Anthony D Ho,Yvonne Samstag,Guido Wabnitz,Diana Knies,Nadja Feldmeyer,Michael Hundemer,Ralf-Holger Voss,Roland Conradi,Sergej Medenhoff,Ingrid Müller,Melanie Engelhardt,Matthias Theobald,Claudia Luckner-Minden

doi:10.1371/journal.pone.0063521

Abstract

Tumor-growth is often associated with the expansion of myeloid derived suppressor cells that lead to local or systemic arginine depletion via the enzyme arginase. It is generally assumed that this arginine deficiency induces a global shut-down of T cell activation with ensuing tumor immune escape. While the impact of arginine depletion on polyclonal T cell proliferation and cytokine secretion is well documented, its influence on chemotaxis, cytotoxicity and antigen specific activation of human T cells has not been demonstrated so far. We show here that chemotaxis and early calcium signaling of human T cells are unimpaired in the absence of arginine. We then analyzed CD8+ T cell activation in a tumor peptide as well as a viral peptide antigen specific system: (i) CD8+ T cells with specificity against the MART-1aa26–35*A27L tumor antigen expanded with in vitro generated dendritic cells, and (ii) clonal CMV pp65aa495–503 specific T cells and T cells retrovirally transduced with a CMV pp65aa495–503 specific T cell receptor were analyzed. Our data demonstrate that human CD8+ T cell antigen specific cytotoxicity and perforin secretion are completely preserved in the absence of arginine, while antigen specific proliferation as well as IFN-γ and granzyme B secretion are severely compromised. These novel results highlight the complexity of antigen specific T cell activation and demonstrate that human T cells can preserve important activation-induced effector functions in the context of arginine deficiency.

Highlights

The fate of a growing tumor is based on the proliferative capacity of the cancer cell itself but rather dictated by the complex interplay of various invading cell types, most prominently antitumoral and regulatory immune cells
We report here that human T cell chemotaxis, early calcium signaling and MART-1aa26–35*A27L specific CD8+ T cell mediated cytotoxicity are uncompromised in the absence of arginine while interferon-gamma (IFN-c) and granzyme B secretion are suppressed when tumor antigen specific T cells were restimulated with the cognate peptide under arginine-limiting conditions
Our study highlights the following two novel aspects: (i) human T cell functions were analyzed in an antigen specific context and suppression of IFN-c secretion, granzyme B mobilization and proliferation were recapitulated; (ii) we show that important human T cell functions – chemotaxis, calcium signaling, cytotoxicity – are completely preserved in the absence of arginine

Summary

Introduction

The fate of a growing tumor is based on the proliferative capacity of the cancer cell itself but rather dictated by the complex interplay of various invading cell types, most prominently antitumoral and regulatory immune cells. The endogenous or therapy-induced antitumoral immune attack is often inhibited by tumor immune escape mechanisms [1], [2]. -called myeloid-derived suppressor cells (MDSC) inhibit effectively antitumoral adaptive immune responses mainly by the production of reactive oxygen intermediates and by the expression of the arginine-metabolizing enzymes nitric oxide synthase and arginase [3,4]. Murine and human MDSC have been shown to express the hepatic isoform arginase I constitutively or inducibly [6]. In human T lymphocytes, the absence of arginine induces a downregulation of the signal transducing T cell receptor-associated f chain [8,9], impairs dephosphorylation of the actin-binding protein cofilin [10] and inhibits progression through the cell cycle via induction of a G0–G1 arrest [11]

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Results

Conclusion