Arginine metabolism is a key parameter of tumor immunology and availability of this specific amino acid for both cancer and immune cells is a potentially decisive factor for tumor growth versus cancer elimination. A recurrent finding in cancer patients is the depletion of the amino acid arginine in the tumor micromilieu. This arginine consumption is due to the enzyme arginase, which can be expressed by tumor cells themselves or by tumor-invading myeloid cells (myeloid-derived suppressor cells, MDSC) of the patient’s immune system. We have first described the specific and constitutive expression of arginase I in human neutrophil granulocytes. Upon liberation of the enzyme and consecutive arginine depletion, human T lymphocytes and NK cells are severely suppressed in their effector functions. MDSC-induced arginine depletion has emerged as a key tumor immune escape mechanism and inhibition of this pathway is associated with tumor control or eradication in a variety of murine tumor models. Arginine availability, on the other hand, is also necessary for cancer cell proliferation and tumor growth. Arginine depletion is therefore emerging as a novel metabolically-oriented treatment strategy that aims to deprive cancer cells of this crucial nutrient.
Our laboratory therefore works on (1) a better understanding of intracellular arginine metabolism and various strategies to inhibit MDSC-mediated tumor immune escape, (2) analyses of the membrane transport proteins, which are necessary to provide arginine to immune and tumor cells and (3) novel therapeutic strategies to maximize arginine deprivation as efficient anti-tumor effector pathway.
Project 1: Molecular T cell immunotherapy and inhibition of tumor immune escape mechanisms (TP06 of CRC1292)
Yagmur Bülbül, Anke Werner
T cells can be retrovirally equipped with T cell receptors (TCRs) that are able to recognise tumour-associated antigens (TAA) with high affinity and thereby redirected to kill tumour cells in vitro and in vivo. A heterogenous population of immature or activated myeloid cells (myeloid-derived suppressor cells, MDSC) has emerged as one of the key effector populations of tumour immune escape, which potentially inhibits antitumoural effectiveness of TAA reactive T cells. This project aims to improve conventional T cell-based tumour immunotherapy by analysis and suppression of MDSC-mediated tumour immune escape mechanisms. For this purpose, we will use adoptive transfer of T cells redirected with HLA.A*02:01-restricted TCRs of defined specificities (p53, MDM2, XBP-1) in both a syngeneic mouse tumour model and a NSG-A2 xenograft model. MDSC frequency in the circulation and in the tumour micromilieu as well as MDSC-associated tumour immune escape mechanisms (arginase and downstream products of arginine metabolism, reactive oxygen and nitrogen species) and suppressive properties will be analysed in vitro and in vivo. In parallel, T cell expansion, phenotype and effector functions will be studied and correlated with the MDSC data. MDSC-mediated immunosuppressive pathways will then be specifically targeted by (a) pharmacological inhibitors for arginase, COX-2 or nitric oxide synthase, (b) cytokine receptor blocking antibodies, (c) inhibition of the generation or expansion of MDSC or (d) maturation / differentiation of MDSC into effective antigen presenting cells. T cell proliferation and effector functions will be further boosted by modulating arginine availability via supplementation of citrulline, a non-toxic precursor amino acid for cellular arginine biosynthesis. In summary, our novel combined approach has the potential to synergistically enhance the efficacy of anti-tumour immunotherapy as a prerequisite for its successful translational implementation in clinical practice as powerful anti-cancer treatment.
This is project TP06 (together with Prof. Dr. Matthias Theobald) within the novel CRC1292 (“Targeting convergent mechanisms of inefficient immunity in tumors and chronic infections”; start: 01/2018; https://sfb1292.uni-mainz.de/; http://www.uni-mainz.de/presse/aktuell/3502_DEU_HTML.php).
Funding 2018-2021: German Science Foundation (DFG)
Project 2: Regulation and function of arginine transport in human immune and tumor cells
Anke Werner, Niklas Wildenberg
Since arginine availability is crucial for full T cell activation, we analyse how this amino acid is transported into human T lymphocytes and other immune cells. While the complexity and diversity of Cationic Amino Acid Transporters (CATs) and Heteromeric Amino Acid Transporters (HATs) is well appreciated in various mammalian tissues, practically nothing is known about the expression and regulation of potential arginine transport proteins in primary human immune cells as well as tumor entities. We have joined forces with the group of Prof. Dr. Ellen Closs (Institute of Pharmacology) here in Mainz in order to bring light into this black box. We recently demonstrated that the human cationic amino acid transporter-1 (hCAT-1) is necessary for arginine import and efficient human T cell activation. The identification of the relevant arginine transport proteins in human immune cells will not only lead to a better understanding of the regulation of adaptive immune responses but will also potentially identify novel target structures for immunosuppressive pharmacological interference with immune cell function. Also, targeting non-redundant arginine import proteins in cancer cells is a novel, potentially highly promising anti-tumor treatment strategy.
Funding 2012-2015: German Science Foundation (DFG; MU 1547/4-1)
Funding 2016-2018: Stiftung Rheinland-Pfalz für Innovation
Project 3: Induction of arginine deficiency as novel metabolic treatment strategy for multiple myeloma
Björn Jacobi, Johannes Windschmitt
Multiple myeloma is the second most prevalent hematological cancer. The disease is due to malignant expansion of antibody-secreting plasma cells, is characterised by a high degree of morbidity (e.g. anemia, bone fractures and pain, kidney failure, infections) and is still largely incurable. Due their high immunoglobulin synthesis and secretion, myeloma cells are especially dependent on a proper protein metabolism with degradation and disposal of misfolded proteins. To cope with misfolded proteins, myeloma cells rely on a cellular stress response program (Unfolded Protein Response, UPR), which preserves cellular viability under physiological conditions but induces cell death upon prolonged or exaggerated induction via misfolded intracellular proteins. Since availability of arginine is necessary for unimpaired protein synthesis, we are currently analysing the role of therapeutic arginine deprivation as potential UPR inducer. This strategy is also tested in combination with (i) the anti-myeloma drug class of proteasome inhibitors, which interfere with proper protein degradation and (ii) the arginine analogue canavanine, which by itself can induce massive protein misfolding due to incorporation into nascent protein chains instead of arginine. We also characterize the arginine transporters in myeloma cells as potential novel target structures for pharmacological interference with arginine supply for these cancer cells.
Funding 2016-2018: Stiftung Rheinland-Pfalz für Innovation