The understanding of the tumor microenvironment (TME) has been expanding in recent years in the context of interactions among different cell types, through direct cell–cell communication as well as through soluble factors. It has become evident that the development of a successful antitumor response depends on several TME factors. In this context, the number, type, and subsets of immune cells, as well as the functionality, memory, and exhaustion state of leukocytes are key factors of the TME. Both the presence and functionality of immune cells, in particular T cells, are regulated by cellular and soluble factors of the TME. In this regard, one fundamental reason for failure of antitumor responses is hijacked immune cells, which contribute to the immunosuppressive TME in multiple ways. Specifically, reactive oxygen species (ROS), metabolites, and anti-inflammatory cytokines have central roles in generating an immunosuppressive TME. In this review, we focused on recent developments in the immune cell constituents of the TME, and the micromilieu control of antitumor responses. Furthermore, we highlighted the current challenges of T cell-based immunotherapies and potential future strategies to consider for strengthening their effectiveness.

The actin cytoskeleton of eukaryotic cells is a dynamic, fibrous network that is regulated by the concerted action of actin-binding proteins (ABPs). In particular, rapid polarization of cells in response to internal and external stimuli is fundamental to cell migration and invasion. Various isoforms of ABPs in different tissues equip cells with variable degrees of migratory and adhesive capacities. In addition, regulation of ABPs by posttranslational modifications (PTM) is pivotal to the rapid responsiveness of cells. In this context, phosphorylation of ABPs and its functional consequences have been studied extensively. However, the study of reduction/oxidation (redox) modifications of oxidation-sensitive cysteine and methionine residues of actin, ABPs, adhesion molecules, and signaling proteins regulating actin cytoskeletal dynamics has only recently emerged as a field. The relevance of such protein oxidations to cellular physiology and pathophysiology has remained largely elusive. Importantly, studying protein oxidation spatiotemporally can provide novel insights into localized redox regulation of cellular functions. In this review, we focus on the redox regulation of the actin cytoskeleton, its challenges, and recently developed tools to study its physiological and pathophysiological consequences.

The activity and function of T-cells are influenced by the intra- and extracellular redox milieu. Oxidative stress induces hypo responsiveness of untransformed T-cells. Vice versa increased glutathione (GSH) levels or decreased levels of reactive oxygen species (ROS) prime T-cell metabolism for inflammation, e.g., in rheumatoid arthritis. Therefore, balancing the T-cell redox milieu may represent a promising new option for therapeutic immune modulation. Here we show that sulforaphane (SFN), a compound derived from plants of the Brassicaceae family, e.g., broccoli, induces a pro-oxidative state in untransformed human T-cells of healthy donors or RA patients. This manifested as an increase of intracellular ROS and a marked decrease of GSH. Consistently, increased global cysteine sulfenylation was detected. Importantly, a major target for SFN-mediated protein oxidation was STAT3, a transcription factor involved in the regulation of T_H17-related genes. Accordingly, SFN significantly inhibited the activation of untransformed human T-cells derived from healthy donors or RA patients, and downregulated the expression of the transcription factor RORγt, and the T_H17-related cytokines IL-17A, IL-17F, and IL-22, which play a major role within the pathophysiology of many chronic inflammatory/autoimmune diseases. The inhibitory effects of SFN could be abolished by exogenously supplied GSH and by the GSH replenishing antioxidant N-acetylcysteine (NAC). Together, our study provides mechanistic insights into the mode of action of the natural substance SFN. It specifically exerts T_H17 prone immunosuppressive effects on untransformed human T-cells by decreasing GSH and accumulation of ROS. Thus, SFN may offer novel clinical options for the treatment of T_H17 related chronic inflammatory/autoimmune diseases such as rheumatoid arthritis.

The interplay between keratinocytes and immune cells, especially T cells, plays an important role in the pathogenesis of chronic inflammatory skin diseases. During psoriasis, keratinocytes attract T cells by releasing chemokines, while skin-infiltrating self-reactive T cells secrete proinflammatory cytokines, e.g., IFNγ and IL-17A, that cause epidermal hyperplasia. Similarly, in chronic graft-versus-host disease, allogenic IFNγ-producing Th1/Tc1 and IL-17-producing Th17/Tc17 cells are recruited by keratinocyte-derived chemokines and accumulate in the skin. However, whether keratinocytes act as nonprofessional antigen-presenting cells to directly activate naive human T cells in the epidermis remains unknown. Here, we demonstrate that under proinflammatory conditions, primary human keratinocytes indeed activate naive human T cells. This activation required cell contact and costimulatory signaling via CD58/CD2 and CD54/LFA-1. Naive T cells costimulated by keratinocytes selectively differentiated into Th1 and Th17 cells. In particular, keratinocyte-initiated Th1 differentiation was dependent on costimulation through CD58/CD2. The latter molecule initiated STAT1 signaling and IFNγ production in T cells. Costimulation of T cells by keratinocytes resulting in Th1 and Th17 differentiation represents a new explanation for the local enrichment of Th1 and Th17 cells in the skin of patients with a chronic inflammatory skin disease. Consequently, local interference with T cell–keratinocyte interactions may represent a novel strategy for the treatment of Th1 and Th17 cell-driven skin diseases.

The death receptor CD95 is expressed in every cancer cell, thus providing a promising tool to target cancer. Activation of CD95 can, however, lead to apoptosis or proliferation. Yet the molecular determinants of CD95's mode of action remain unclear. Here, we identify an optimal distance between CD95Ligand molecules that enables specific clustering of receptor-ligand pairs, leading to efficient CD95 activation. Surprisingly, efficient CD95 activation leads to apoptosis in cancer cells in vitro and increased tumor growth in vivo. We show that allowing a 3D aggregation of cancer cells in vitro switches the apoptotic response to proliferation. Indeed, we demonstrate that the absence or presence of cell-cell contacts dictates the cell response to CD95. Cell contacts increase global levels of phosphorylated tyrosines, including CD95's tyrosine. A tyrosine-to-alanine CD95 mutant blocks proliferation in cells in contact. Our study sheds light into the regulatory mechanism of CD95 activation that can be further explored for anti-cancer therapies.

Several antitumor therapies work by increasing reactive oxygen species (ROS) within the tumor micromilieu. Here, we reveal that L-plastin (LPL), an established tumor marker, is reversibly regulated by ROS-induced thiol oxidation on Cys101, which forms a disulfide bridge with Cys42. LPL reduction is mediated by the Thioredoxin1 (TRX1) system, as shown by TRX1 trapping, TRX1 knockdown and blockade of Thioredoxin1 reductase (TRXR1) with auranofin. LPL oxidation diminishes its actin-bundling capacity. Ratiometric imaging using an LPL-roGFP-Orp1 fusion protein and a dimedone-based proximity ligation assay (PLA) reveal that LPL oxidation occurs primarily in actin-based cellular extrusions and strongly inhibits cell spreading and filopodial extension formation in tumor cells. This effect is accompanied by decreased tumor cell migration, invasion and extracellular matrix (ECM) degradation. Since LPL oxidation occurs following treatment of tumors with auranofin or γ-irradiation, it may be a molecular mechanism contributing to the effectiveness of tumor treatment with redox-altering therapies.