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The amountof ILDR2 peptides in kidney and testis samples derived fromhuman, monkey and mouse.

The immunoglobulin-like domain containing receptor 2 (ILDR2), a type I transmembrane protein belonging to the B7 family of immunomodulatory receptors, has been described to induce an immunosuppressive effect on T-cell responses. Besides its expression in several nonlymphoid tissue types, we found that ILDR2 was also expressed in fibroblastic reticular cells (FRC) in the stromal part of the lymph node. These immunoregulatory cells were located in the T-cell zone and were essential for the recruitment of naïve T cells and activated dendritic cells to the lymph nodes. Previously, it has been shown that an ILDR2-Fc fusion protein exhibits immunomodulatory effects in several models of autoimmune diseases, such as multiple sclerosis, rheumatoid arthritis, and type I diabetes. Herein, we report the generation and characterization of a human/mouse/monkey cross-reactive anti-ILDR2 hIgG2 antibody, BAY 1905254, developed to block the immunosuppressive activity of ILDR2 for cancer immunotherapy. BAY 1905254 was shown to promote T-cell activation in vitro and enhance antigen-specific T-cell proliferation and cytotoxicity in vivo in mice. BAY 1905254 also showed potent efficacy in various syngeneic mouse cancer models, and the efficacy was found to correlate with increasing mutational load in the cancer models used. Additive or even synergistic antitumor effects were observed when BAY 1905254 was administered in combination with anti–PD-L1, an immunogenic cell death–inducing chemotherapeutic, or with tumor antigen immunization. Taken together, our data showed that BAY 1905254 is a potential drug candidate for cancer immunotherapy, supporting its further evaluation.

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The dynamics and phenotypes of intratumoral myeloid cells during tumor progression are poorly understood. Here we define myeloid cellular states in gliomas by longitudinal single-cell profiling and demonstrate their strict control by the tumor genotype: in isocitrate dehydrogenase (IDH)-mutant tumors, differentiation of infiltrating myeloid cells is blocked, resulting in an immature phenotype. In late-stage gliomas, monocyte-derived macrophages drive tolerogenic alignment of the microenvironment, thus preventing T cell response. We define the IDH-dependent tumor education of infiltrating macrophages to be causally related to a complex re-orchestration of tryptophan metabolism, resulting in activation of the aryl hydrocarbon receptor. We further show that the altered metabolism of IDH-mutant gliomas maintains this axis in bystander cells and that pharmacological inhibition of tryptophan metabolism can reverse immunosuppression. In conclusion, we provide evidence of a glioma genotype-dependent intratumoral network of resident and recruited myeloid cells and identify tryptophan metabolism as a target for immunotherapy of IDH-mutant tumors. Platten and colleagues find that tryptophan metabolism by myeloid cells contributes to immunosuppressive microenvironment uniquely in IDH-mutant gliomas, which can be overcome by inhibiting this pathway in murine tumor models.

Introduction: Many patients do not benefit from immunotherapies targeting immune checkpoints such as PD(L)-1 due to a variety of resistance mechanisms. The AhR pathway is downstream of the Trp-IDO/TDO-Kyn axis. High levels of immune-suppressive AhR-activating ligands, such as kynurenine derived from IDO1/TDO2-expressing tumors, have been implicated as a potential resistance mechanism and are associated with poor responses to PD-1 therapies. Compared with inhibition of IDO1/TDO2 alone, direct AhR inhibition can block the activation of this transcription factor and may better counteract immune suppression. BAY2416964 is a novel, oral AhR inhibitor currently in a Phase I clinical trial in patients with solid tumors. Preclinically, BAY2416964 can block the activation of AhR by kynurenine and relieve its immune-suppressive effects, thereby restoring anti-tumor T-cell activity, reducing the level of inhibitory myeloid-derived suppressor cells and regulatory T cells, and improving the effectiveness of PD-1 blockade. We have analyzed PK and biomarker data from the ongoing monotherapy dose-escalation study to explore a potential optimal dose and schedule for effective AhR inhibition. Methods: PK data were intensely sampled from patients in 10 different groups with different dosing regimens (once and twice daily) and various food-intake scenarios. AhR downstream target gene expression after ex vivo kynurenic acid stimulation of patients’ peripheral blood mononuclear cells (PBMCs) was assessed. A population PK (popPK) model was developed and calibrated to characterize the PK of BAY2416964 across treatment groups. This model accounts for the non-linear relationship between dose and bioavailability as well as the effect of food intake. Results: The preliminary popPK model identified a non-linear relationship between the BAY2416964 dose and its (relative) bioavailability which was dependent on the respective food condition. The popPK model was able to describe the clinical BAY2416964 plasma exposures from all dose groups, including the effect of food intake. Ex vivo analysis of PBMCs showed inhibition of kynurenic acid-induced AhR downstream gene expression (such as CYP1A1, CYP1B1), suggesting effective in vivo target engagement in the doses tested. Conclusion: This modeling-based PK analysis along with target engagement in peripheral blood informed the posology to be tested in the dose-expansion part of the ongoing clinical trial. Citation Format: Kyriakos Papadopoulos, Michael Cecchini, Juanita S. Lopez, Dirk Jäger, Ki Young Chung, Michael Platten, Florian Prinz, Yuko Ishii, David Schaer, Ilona Gutcher, Gabriele Leder, Radost Pencheva, Thomas Wagener, Christian Scheerans, Carsten Zieschang, Andrea Wagner, Spyros Stamatelos, Ecaterina Dumbrava. Preliminary analysis of pharmacokinetic (PK) and target engagement biomarkers from a first in human phase 1 study of immunomodulatory aryl hydrocarbon receptor (AhR) inhibitor BAY2416964 [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2023; Part 2 (Clinical Trials and Late-Breaking Research); 2023 Apr 14-19; Orlando, FL. Philadelphia (PA): AACR; Cancer Res 2023;83(8_Suppl):Abstract nr CT276.

Re-constitution of anti-tumor T-cell responses by clinically-approved immune checkpoint inhibitors (ICIs) targeting CTLA4 or PD-1/PD-L1 represents a breakthrough cancer therapy. Nevertheless, a substantial number of patients do not benefit from these new therapeutic modalities chiefly due to local immunosuppression in the tumor microenvironment. In addition, the long circulation time of ICIs restricts options to modify dosing regimens for management of adverse effects. Oral small molecule inhibitors as next generation immune-oncology agents may - in contrast to antibodies - allow targeting of intracellular targets for a defined duration of time. This will permit a fine-tuning of efficacy versus tolerability in single-agent treatment as well as in combination with approved ICIs. The overexpression of indole dioxygenase (IDO1) and tryptophan dioxygenase (TDO2) by many tumors results in increased metabolism of tryptophan (TRP) into kynurenine (KYN), which induces immunosuppression via activation of the aryl hydrocarbon receptor (AhR). Inhibition of AhR was proposed to restore T-cell function and induce tumor rejection. However, it was expected that identification of selective lead candidates for AhR inhibition would be challenging due to the known affinity of the AhR ligand binding site for polyaromatic ligands such as 2,3,7,8-tetrachlorodibenzodioxine (TCDD). A library of 4 million compounds was screened in a cell-based HTS campaign. A thorough hit reduction process was performed based on stringent filter parameters for lead-likeness. This process delivered a hit set of significant chemical diversity. Out of several compound classes with drug-like properties, 1,3-diaryl-pyrazin-6-one-5-carboxylic amides were selected as a preferred lead series. A comprehensive SAR exploration, including in vitro mechanistic and functional validation, was performed. Lead optimization was strongly emphasized on improving lipophilicity efficiency (LLE) to balance potency with a viable PK and CYP450 interaction profile. Several candidates suitable for in vivo profiling were identified and BAY-218 was advanced to in-depth pharmacodynamic and pharmacokinetic in vivo assessments. BAY-218 showed mono-therapeutic efficacy that was comparable to ICI treatment and further therapeutic improvement was achieved by combination with an anti-PD-L1 antibody. We were able to characterize 1,3-diaryl-pyrazin-6-one-5-carboxylic amides as a new and unprecedented class of AhR inhibitors as well as identify the key substitutions that contribute to the overall compound profile. Citation Format: Norbert Schmees, Ilona Gutcher, Ulrike Roehn, Horst Irlbacher, Detlef Stoeckigt, Benjamin Bader, Christina Kober, Lars Roese, Rafael Carretero, Iris Oezen, Ludiwg Zorn, Michael Platten, Ingo Volker Hartung, Bertolt Kreft, Hilmar Weinmann. Identification of BAY-218, a potent and selective small-molecule AhR inhibitor, as a new modality to counteract tumor immunosuppression [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2019; 2019 Mar 29-Apr 3; Atlanta, GA. Philadelphia (PA): AACR; Cancer Res 2019;79(13 Suppl):Abstract nr 4454.

Tumor cells co-opt multiple pathways in order to evade attack by infiltrating immune cells. One such mechanism is the upregulation of indole-2,3-dioxygenase (IDO1) and/or tryptophan-2,3-dioxygenase (TDO2), both of which are first-step, rate-limiting enzymes degrading tryptophan to the immunosuppressive metabolites kynurenine (KYN) and kynurenic acid (KA). KYN and KA bind and activate the aryl hydrocarbon receptor (AhR), which is expressed in many cell types and is well known for its immunosuppressive effects. Targeting of the AhR with an inhibitor may therefore provide a novel immunotherapeutic approach for enhancing anti-tumoral immune responses and treating cancer. Here we describe the identification and functional immune characterization of BAY-218, a novel, selective and potent AhR small molecule inhibitor. Mechanistically, BAY-218 inhibited AhR nuclear translocation, dioxin response element (DRE)-luciferase reporter expression and AhR-regulated target gene expression induced by both exogenous and endogenous AhR ligands. In vitro, BAY-218 rescued TNFα production from KA-suppressed LPS-treated primary human monocytes. Furthermore, BAY-218 enhanced T cell cytokine production in a human mixed lymphocyte reaction (MLR) and a mouse antigen-specific bone-marrow-derived dendritic cell (BMDC)-OT-I T cell co-culture. In the MLR, BAY-218 increased anti-PD1 antibody-mediated IL-2 and IFNγ secretion, while an IDO inhibitor did not, indicating that BAY-218 is able to block AhR activation mediated by ligands outside of the IDO-KYN pathway. In vivo, BAY-218 enhanced anti-tumoral immune responses and reduced tumor growth in the syngeneic mouse tumor models CT26 and B16-OVA. FACS analysis of leukocytes infiltrating B16-OVA tumors demonstrated that administration of BAY-218 increased the frequency of tumor-infiltrating CD8+ T cells and NK cells while decreasing GR1+ myeloid cells and CD206+M2 macrophages. Furthermore, BAY-218 enhanced therapeutic efficacy of an anti-PD-L1 antibody in the CT26 model. In summary, AhR inhibition with BAY-218 stimulates pro-inflammatory monocyte and T cell responses in vitro and drives anti-tumor immune responses, resulting in decreased tumor growth, in vivo. Thus, inhibiting AhR represents a novel immunotherapeutic approach for blocking AhR-mediated tumor-associated immunosuppression. Citation Format: Ilona Gutcher, Christina Kober, Lars Roese, Julian Roewe, Norbert Schmees, Florian Prinz, Matyas Gorjanacz, Ulrike Roehn, Benjamin Bader, Horst Irlbacher, Detlef Stoeckigt, Rafael Carretero, Katharina Sahm, Iris Oezen, Hilmar Weinmann, Ingo V. Hartung, Bertolt Kreft, Michael Platten. Blocking tumor-associated immune suppression with BAY-218, a novel, selective aryl hydrocarbon receptor (AhR) inhibitor [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2019; 2019 Mar 29-Apr 3; Atlanta, GA. Philadelphia (PA): AACR; Cancer Res 2019;79(13 Suppl):Abstract nr 1288.

SummaryTGF-β1 is a regulatory cytokine that has an important role in controlling T cell differentiation. T cell-produced TGF-β1 acts on T cells to promote Th17 cell differentiation and the development of experimental autoimmune encephalomyelitis (EAE). However, the exact TGF-β1-producing T cell subset required for Th17 cell generation and its cellular mechanism of action remain unknown. Here we showed that deletion of the Tgfb1 gene from activated T cells and Treg cells, but not Treg cells alone, abrogated Th17 cell differentiation, resulting in almost complete protection from EAE. Furthermore, differentiation of T cells both in vitro and in vivo demonstrated that TGF-β1 was highly expressed by Th17 cells and acted in a predominantly autocrine manner to maintain Th17 cells in vivo. These findings reveal an essential role for activated T cell-produced TGF-β1 in promoting the differentiation of Th17 cells and controlling inflammatory diseases.

Summary T helper 17 (Th17) cell development is driven by cytokines including transforming growth factor-β (TGF-β), interleukin-6 (IL-6), IL-1, and IL-23. Regulatory T (Treg) cells can provide the TGF-β in vitro, but their role in vivo remains unclear, particularly because Treg cells inhibit inflammation in many models of Th17 cell-associated autoimmunity. We used mice expressing Diphtheria toxin receptor under control of the Foxp3 promoter to deplete Foxp3 + Treg cells in adult mice during in vivo Th17 cell priming. Treg cell depletion resulted in a reduced frequency of antigen-specific IL-17 producers in draining lymph nodes and blood, correlating with reduced inflammatory skin responses. In contrast, Treg cells did not promote IL-17 secretion after initial activation stages. Treg cell production of TGF-β was not required for Th17 cell promotion, and neither was suppression of Th1 cell-associated cytokines. Rather, regulation of IL-2 availability and resultant signaling through CD25 by Treg cells was found to play an important role.

ILDR2 is a type I transmembrane protein belonging to the B7 family of immunomodulatory receptors with 98 % sequence identity between the extracellular domains of human and murine orthologues. The exact physiological role of ILDR2 is still unclear, but it has been implicated in the development of type 2 diabetes and the formation of tri-cellular junctions. Our detailed analyses reveal that ILDR2 is expressed in kidney, testis, liver, and lymph node fibroblastic reticular cells (FRC). The latter is particularly interesting since FRC belong to the CD45- stromal population, a specialized cell subset located in the T cell zone. These cells are essential for recruitment of naïve T cells and activated dendritic cells to the lymph node and have been reported to exhibit immuno-regulatory properties. Due to its structural similarity to members of the B7 family and due to its expression in FRCs, we speculated that ILDR2 might play a role e.g. in T cell priming and the initiation of antigen-specific T cell responses. In line with this, we and our collaborators from Compugen were able to show that an ILDR2 -Fc fusion protein is able to a) bind to activated (but not naïve) T cells, b) suppress TCR-stimulated cytokine secretion and c) exhibit immunomodulatory effects in several models of autoimmune diseases, i.e. multiple sclerosis, rheumatoid arthritis, and type 1 diabetes. As a consequence, we set out to generate antibodies against this novel immuno-oncology target and describe here for the first time the characterization of BAY 1905254, a human/mouse cross-reactive IgG2 antibody blocking the immunosuppressive activity of ILDR2. BAY 1905254 specifically binds to ILDR2 but not to the closely related family members ILDR1 and ILDR3/LSR. It enhances antigen-specific T cell proliferation in vivo in an ovalbumin vaccination model with OT-I transgenic T cells. BAY 1905254 exhibits anti-tumor activity in different syngeneic mouse models with efficacy correlating with increasing mutational load. Furthermore, additive/synergistic anti-tumor effects can be observed in combination with either an anti-PD-L1 Ab, an immunogenic cell death inducing chemotherapeutic (Docetaxel) or tumor antigen immunization. Ex vivo analysis reveal that BAY 1905254 treatment results in increased intra-tumoral IFN-y levels and enhanced infiltration of CD45+ cells, e.g. of CD8α+ dendritic cells (DCs). Interestingly, this DC population is known to play a crucial role in cross-presentation, thus, in the initiation of CD8+ T cell responses. In summary, BAY 1905254 is a function-blocking Ab able to counteract the immuno-suppressive function of the newly discovered immuno-oncology target ILDR2. The Ab exhibits in vivo efficacy both in monotherapy as well as in combination with various other approaches, and it is planned to advance BAY 1905254 into FiM trials in 2018. Citation Format: Julia Huetter, Uwe Gritzan, Ilona Gutcher, Sven Golfier, Wolf-Dietrich Doecke, Merlin Verena Luetke-Eversloh, Helge Roider, John Hunter, Andrew Pow, Spencer Liang, Zurit Levine, Ofer Levy, Ilan Vaaknin, Bertolt Kreft, Lars Roese. Discovery and preclinical characterization of BAY 1905254 a novel immune checkpoint inhibitor for cancer immunotherapy targeting the immunoglobulin-like domain containing receptor 2 (ILDR2) [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2018; 2018 Apr 14-18; Chicago, IL. Philadelphia (PA): AACR; Cancer Res 2018;78(13 Suppl):Abstract nr 2778.

SummaryThe action of type I interferons in the central nervous system (CNS) during autoimmunity is largely unknown. Here, we demonstrate elevated interferon beta concentrations in the CNS, but not blood, of mice with experimental autoimmune encephalomyelitis (EAE), a model for CNS autoimmunity. Furthermore, mice devoid of the broadly expressed type I IFN receptor (IFNAR) developed exacerbated clinical disease accompanied by a markedly higher inflammation, demyelination, and lethality without shifting the T helper 17 (Th17) or Th1 cell immune response. Whereas adoptive transfer of encephalitogenic T cells led to enhanced disease in Ifnar1−/− mice, newly created conditional mice with B or T lymphocyte-specific IFNAR ablation showed normal EAE. The engagement of IFNAR on neuroectodermal CNS cells had no protective effect. In contrast, absence of IFNAR on myeloid cells led to severe disease with an enhanced effector phase and increased lethality, indicating a distinct protective function of type I IFNs during autoimmune inflammation of the CNS.

T cell–mediated autoimmune diseases such as multiple sclerosis and rheumatoid arthritis are driven by autoaggressive Th cells. The pathogenicity of such Th cells has, in the past, been considered to be dictated by their cytokine polarization profile. The polarization of such effector T cells relies critically upon the actions of cytokines secreted by APCs. While Th1 polarization has long been associated with the pathogenesis of autoimmune diseases, recent data obtained in gene-targeted mice and the discovery of Th17 cell involvement in autoimmunity conflict with this hypothesis. In light of these recent developments, we discuss in this review the actions of APC-derived cytokines and their emerging roles in T cell polarization in the context of autoimmune inflammatory responses.

The inhibitory effects of the novel Kv1.5 channel blocker, S9947 (2'-(benzyloxycarbonylaminomethyl)biphenyl-2-carboxylic acid 2-(2-pyridyl)ethylamide), on cloned human Kv1.5 (hKv1.5), expressed in both Xenopus oocytes and Chinese hamster ovary (CHO) cells, and on native cardiac ultrarapid delayed rectifier potassium currents (IKur) in rat (ventricle myocytes) and human (atrial myocytes) were investigated. The influence of S9947 on the action potential was examined in rat ventricular myocytes. Using the two-electrode voltage-clamp technique in Xenopus oocytes and the patch-clamp technique (whole cell configuration) in CHO cells, hKv1.5 was inhibited by S9947 with IC50 values of 0.65 microM and 0.42 microM, respectively. In addition, inhibition of human Kv4.3 (hKv4.3) and HERG by 10 microM S9947 was low (approximately 20%) and absent, respectively. Using the patch-clamp technique in the whole cell configuration, IKur currents in rat ventricular (rIKur) cardiomyocytes and human atrial (hIKur) cardiomyocytes were inhibited by S9947 with IC50 values of 0.96 microM and 0.07 microM, respectively. In contrast, rat cardiac inward rectifier current (rIK1) and rat (rIto) and human (hIto) cardiac transient outward currents were only inhibited by approximately 20% with 10 microM S9947. In rat cardiomyocytes, using the patch-clamp technique, action potential duration was increased by S9947 in a concentration-dependent (0.3-10 microM) and rate-independent manner. The data show that S9947 suppresses both cloned (Kv1.5) and native (IKur) cardiac potassium currents. Furthermore, S9947 prolongs rat action potential in a rate-independent manner.

In this study, we analysed the inhibitory potency, blocking characteristics and putative binding sites of three structurally distinct Kv1.5 channel inhibitors on cloned human Kv1.5 channels. Obtained IC(50) values for S9947, MSD-D and ICAGEN-4 were 0.7 microM, 0.5 microM, and 1.6 microM, respectively. The Hill-coefficients were close to 1 for S9947 and approximately 2 for MSD-D and ICAGEN-4. All three compounds inhibited Kv1.5 channels preferentially in the open state, with Kv1.5 block displaying positive frequency dependence, but no clear voltage and potassium dependence. In contrast to slow on- and off-rates of apparent binding of MSD-D and ICAGEN-4, S9947 had fast on- and off-rates resulting in faster adaptation to changes in pulse frequency. Utilizing Alanine-scanning and in silico modeling we suggest binding of the compounds to the central cavity with crucial residues Ile508 and Val512 in the S6-segment. Residue Thr480 located at the base of the selectivity filter is important for ICAGEN-4 and S9947 inhibition, but less so for MSD-D binding. Our docking models suggest that the innermost potassium ion in the selectivity filter may form a tertiary complex with oxygens of S9947 and ICAGEN-4 and residue Thr480. This binding component is absent in the MSD-D block. As S9947 and ICAGEN-4 show faster block with proceeding channel openings, formation of this tertiary complex may increasingly stabilise binding of S9947 and ICAGEN-4, thereby explaining open channel block kinetics of these compounds.

T helper type 1 (T(H)1) lymphocytes are considered to be the main pathogenic cell type responsible for organ-specific autoimmune inflammation. As interleukin 18 (IL-18) is a cofactor with IL-12 in promoting T(H)1 cell development, we examined the function of IL-18 and its receptor, IL-18R, in autoimmune central nervous system inflammation. Similar to IL-12-deficient mice, IL-18-deficient mice were susceptible to experimental autoimmune encephalomyelitis. In contrast, IL-18R alpha-deficient mice were resistant to experimental autoimmune encephalomyelitis, indicating involvement of an IL-18R alpha ligand other than IL-18 with encephalitogenic properties. Moreover, engagement of IL-18R alpha on antigen-presenting cells was required for the generation of pathogenic IL-17-producing T helper cells. Thus, IL-18 and T(H)1 cells are dispensable, whereas IL-18R alpha and IL-17-producing T helper cells are required, for autoimmune central nervous system inflammation.

Inflammatory diseases of the CNS, such as MS and its animal model EAE, are characterized by infiltration of activated lymphocytes and phagocytes into the CNS. Within the CNS, activation of resident cells initiates an inflammatory cascade, leading to tissue destruction, demyelination, and neurologic deficit. TLRs recognize microbes and are pivotal mediators of innate immunity. Within the CNS, augmented TLR expression during EAE is observed, even in the absence of any apparent microbial involvement. To determine the functional relevance of this phenomenon during sterile autoimmunity, we studied the role of different TLRs as well as their common signaling adaptor MyD88 in the development of EAE. We found that MyD88–/– mice were completely EAE resistant. Surprisingly, this protection is partly due to engagement of the CpG receptor TLR9. Restricting the MyD88 or TLR9 mutation to host radio-resistant cells, including the cells within the CNS, revealed that engagement of radio-resistant cells modulated the disease course and histopathological changes. Our data clearly demonstrate that both TLR9 and MyD88 are essential modulators of the autoimmune process during the effector phase of disease and suggest that endogenous “danger signals” modulate the disease pathogenesis.

Th17 development is driven by multiple cytokines, including TGF-β. Regulatory T cells (Tregs) promote Th17 development through production of TGF-β in vitro. However their role in vivo remains unclear, particularly since Tregs inhibit inflammation in many models of autoimmunity where Th17 cells are active. Using mice expressing Diphtheria toxin receptor under control of the Foxp3 promoter, Foxp3+ Tregs were depleted in adult mice during in vivo Th17 priming. Treg depletion resulted in a reduced frequency of antigen-specific IL-17 producers in draining lymph nodes and blood, correlating with reduced inflammatory skin responses. In contrast, Tregs did not promote IL-17 after initial activation stages. Treg production of TGF-β was not required for Th17 promotion, neither was suppression of Th1-associated cytokines. Rather, regulation of IL-2 availability and resultant signaling through CD25 and pSTAT5 by Tregs was found to play an important role.

BACKGROUND: IDH1-mutated gliomas are associated with less abundant and phenotypically skewed innate and adaptive immune cell infiltrates compared to IDH1 wild-type tumors. Despite this, the most frequent mutation - IDH1R132H - represents a clonal shared neoantigen and mutations in IDH are associated with a more favorable prognosis. While the tumor cell-intrinsic consequences of the oncometabolite R-2-hydroxyglutarate (R-2-HG) accumulating in IDH1-mutated gliomas as a result of a neomorphic enzymatic function, are well-characterized, potential direct paracrine effects of R-2-HG influencing the glioma immune microenvironment remain incompletely understood. AIM: This study aimed at characterizing the impact of the oncometabolite R-2-HG on the innate immune microenvironment of IDH1-mutated gliomas. METHODS AND RESULTS: By means of comprehensive analyses of expression datasets from human gliomas and syngeneic murine tumor models as well as transporter studies we demonstrate that R-2-HG is imported by both microglia and macrophages via SLC family transporters and suppresses their function in a paracrine manner. Functional analyses of microglia and macrophages indicate an R-2-HG-driven induction of tolerogenicity as evidenced by accumulation of IL10 and TGFβ and suppression of MHC-II expression, which results in impaired activation of antigen-specific T cells and activation of immune checkpoint molecules. Multi-level signature profiling of human tumor-infiltrating as well as primary immune cells was complemented by reporter gene assays and pathway analyses and revealed that R-2-HG activates the cytosolic transcription factor aryl hydrocarbon receptor (AHR), a key immunomodulatory target of immunosuppressive tryptophan metabolism. By means of knockout models, the observed immunosuppressive phenotype was shown to be AHR-dependent. Functional relevance of R-2-HG-mediated, AHR-driven impairment of myeloid cell immunity was demonstrated in vivo by pharmacological AHR inhibition, increasing the efficacy of checkpoint blockade. CONCLUSION: R-2-HG impairs antitumor immunity in IDH1-mutated gliomas by activating the AHR in innate immune cells, thus suppressing the innate immune microenvironment by compromising antigen presentation and activation of antigen-specific T cells. This, together with recent findings on inhibitory effects on T cell immunity, represents a novel mechanism of immune evasion of an immunogenic driver mutation and opens a novel therapeutic approach to IDH1-mutated gliomas.