Characterization of the<i>in vivo</i>immune network of IDO, tryptophan metabolism, PD-L1, and<i>CTLA-4</i>in circulating immune cells in melanoma

Abstract

In melanoma, both the induction of immunosuppression by tumor cells and the inflammatory antitumor response can induce an upregulation of counter-regulatory mechanisms such as indoleamine 2,3-dioxygenase (IDO), programmed death-ligand 1 (PD-L1) and CTLA-4⁺ regulatory T-cells (Tregs) in the tumor microenvironment. Even though these immunosuppressive mediators are targets for immunotherapy, research investigating their expression in the peripheral blood is lacking. We therefore, performed flow cytometry on PBMCs of stage I-IV melanoma patients. IDO expression was detected in plasmacytoid dendritic cells (pDC) and monocytic myeloid-derived suppressor cells (mMDSC), and increased in advanced disease stage (<i>p</i> = 0.027). Tryptophan breakdown confirmed the functional activity of IDO and was linked with increased PD-L1+ cytotoxic T-cells (<i>p</i> = 0.009), relative lymphopenia (<i>p</i> = 0.036), and a higher mDC/pDC ratio (<i>p</i> = 0.002). High levels of circulating PD-L1+ cytotoxic T-cells were associated with increased <i>CTLA-4</i> expression by Tregs (<i>p</i> = 0.005) and MDSC levels (<i>p</i> = 0.033). This illustrates that counter-regulatory immune mechanisms in melanoma should be considered as one interrelated signaling network. Moreover, both increased PD-L1+ T-cells and <i>CTLA-4</i> expression in Tregs conferred a negative prognosis, indicating their <i>in vivo</i> relevance. Remarkably, circulating <i>CTLA-4</i>, IDO, and pDC levels were altered according to prior invasion of the sentinel lymph node and IDO expression in the sentinel was associated with more IDO+ PBMCs. We conclude that the expression of IDO, PD-L1, and <i>CTLA-4</i> in the peripheral blood of melanoma patients is strongly interconnected, associated with advanced disease and negative outcome, independent of disease stage. Combination treatments targeting several of these markers are therefore likely to exert a synergistic response.