No abstract

The hypoxic environment of tumors dictates the phenotype of local myeloid-derived suppressor cells (MDSCs) via HIF-1a expression; hypoxia converts splenic MDSCs from specific into nonspecific suppressors.

Myeloid suppressor cells with high arginase activity are found in tumors and spleen of mice with colon and lung cancer. These cells, described as macrophages or immature dendritic cells, deplete arginine and impair T cell proliferation and cytokine production. Although arginase activity has been described in cancer patients, it is thought to originate from tumor cells metabolizing arginine to ornithine needed to sustain rapid cell proliferation. The goal of this study was to determine whether myeloid suppressor cells producing high arginase existed in renal cell carcinoma patients. Peripheral blood mononuclear cells from 123 patients with metastatic renal cell carcinoma, prior to treatment, were found to have a significantly increased arginase activity. These patients had a markedly decreased cytokine production and expressed low levels of T cell receptor CD3zeta chain. Cell separation studies showed that the increased arginase activity was limited to a specific subset of CD11b+, CD14-, CD15+ cells with a polymorphonuclear granulocyte morphology and markers, instead of macrophages or dendritic cells described in mouse models. Furthermore, these patients had low levels of arginine and high levels of ornithine in plasma. Depletion of the CD11b+, CD14- myeloid suppressor cells reestablished T cell proliferation and CD3zeta chain expression. These results showed, for the first time, the existence of suppressor myeloid cells producing arginase in human cancer patients. In addition, it supports the concept that blocking arginase may be an important step in the success of immunotherapy.

IntroductionL-arginine (L-Arg) is a nonessential amino acid that plays a central role in regulating the immune response. 1 In mammalian cells, L-Arg can be catabolized by 4 enzymatic pathways, namely nitric oxide synthase, arginases I and II, arginine:glycine amidinotransferase, and arginine decarboxylase. L-Arg is profoundly reduced in cancer patients, 2 following liver transplantation, 3 or in severe trauma 4 by an increased production of arginase I. This results in a decreased T-cell proliferation and an impaired T-cell function. This effect can be reversed in trauma by the enteral or parenteral supplementation of We demonstrated that activated T cells cultured in medium without L-Arg or cocultured with myeloidderived suppressor cells (MDSCs) isolated from tumors and producing arginase I have a decreased proliferation, a low expression of T-cell receptor CD3 chain, and an impaired production of cytokines. 2,6,7 However, the mechanisms by which L-Arg starvation blocks T-cell proliferation have not been determined.Signaling through the T-cell receptor, as shown by calcium flux and tyrosine phosphorylation, was not affected for the first 12 hours of culture in the absence of L-Arg and therefore could not completely explain the low proliferation of T cells. 8,9 Furthermore, certain T-cell functions such as up-regulation of IL-2 receptor alpha and production of IL-2 were maintained even in the absence of L-Arg. 8,9 Therefore, we explored whether changes in proteins regulating cell cycle could explain the loss of proliferation in T cells cultured without L-Arg. Cyclin-dependent kinase 4 (cdk4) and cyclin-dependent kinase 6 (cdk6) associate with the D-type cyclins, including cyclin D3, to regulate the progression through early G 1 and into the S phase of cell cycle. This regulation requires inactivation of cyclin D/cdk complex inhibitors and phosphorylation of the Rb protein family. Phosphorylation of Rb by cyclin/cdk complexes induces the subsequent release and nuclear translocation of E2F transcription factors, inducing the expression of genes that promote cell-cycle progression into late G 1 and S phases. 10 The effects of amino acid starvation have been well studied in yeast and some tumor cell lines; however, their role in regulating cell cycle in T cells is unknown. The results shown here demonstrate that L-Arg depletion selectively impairs the expression of cyclin D3 and cdk4, blocking the downstream signaling. GCN2, a kinase involved in amino acid starvation, plays a central role in regulating the cell-cycle arrest induced by L-Arg starvation. These results may provide a new understanding of the impairment of the immune response in various diseases where myeloid-derived suppressor cells, producing high levels of arginase, deplete L-Arg. Materials and methods Cells, cultures, and chemicalsHuman peripheral blood mononuclear cells were obtained from healthy donor buffy coats. T cells were purified using human T-cell enrichment columns (R&D systems, Minneapolis, MN), following the vendor's recommendations. T-ce...