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Indoleamine 2,3-dioxygenase 1 (IDO1) catalyzing the conversion of tryptophan (Trp) to kynurenine (Kyn) in kynurenine pathway (KP) is involved in the immunosuppression in pancreatic cancer (PC), but the value of IDO1 as an independent prognostic marker for PC is uncertain. Moreover, the correlation between tryptophan 2,3-dioxygenase (TDO), an isozyme of IDO1, and PC is largely unknown. Using TCGA database, the correlation between IDO1 and/or TDO expression and PC patients' survival was analyzed. The expressions of IDO1 and TDO in PC cells and PC mice were examined. The effects of IDO1, TDO or dual inhibition on IDO1 and TDO effector pathway (Aryl hydrocarbon receptor, AhR) and on migration and invasion of PC cells were investigated. The block effect of IDO1/TDO dual inhibitor RY103 on KP was evaluated. The preclinical efficacy of RY103 and its immunomodulatory effect on KPIC orthotopic PC mice and Pan02 tumor-bearing mice were explored. Results showed that IDO1/TDO co-expression is an independent prognostic marker for PC. RY103 can significantly block KP and target Kyn-AhR pathway to blunt the migration and invasion of PC cells, exhibit preclinical efficacy and ameliorate IDO1/TDO-mediated immunosuppression in PC mice., (Copyright © 2021 Elsevier B.V. All rights reserved.)

Some methane-oxidizing bacteria use the ribosomally synthesized, posttranslationally modified natural product methanobactin (Mbn) to acquire copper for their primary metabolic enzyme, particulate methane monooxygenase. The operons encoding the machinery to biosynthesize and transport Mbns typically include genes for two proteins, MbnH and MbnP, which are also found as a pair in other genomic contexts related to copper homeostasis. While the MbnH protein, a member of the bacterial diheme cytochrome c peroxidase (bC c P)/MauG superfamily, has been characterized, the structure and function of MbnP, the relationship between the two proteins, and their role in copper homeostasis remain unclear. Biochemical characterization of MbnP from the methanotroph Methylosinus trichosporium OB3b now reveals that MbnP binds a single copper ion, present in the +1 oxidation state, with high affinity. Copper binding to MbnP in vivo is dependent on oxidation of the first tryptophan in a conserved WxW motif to a kynurenine, a transformation that occurs through an interaction of MbnH with MbnP. The 2.04-Å-resolution crystal structure of MbnP reveals a unique fold and an unusual copper-binding site involving a histidine, a methionine, a solvent ligand, and the kynurenine. Although the kynurenine residue may not serve as a Cu I primary-sphere ligand, being positioned ∼2.9 Å away from the Cu I ion, its presence is required for copper binding. Genomic neighborhood analysis indicates that MbnP proteins, and by extension kynurenine-containing copper sites, are widespread and may play diverse roles in microbial copper homeostasis., Competing Interests: The authors declare no competing interest.

Through structural modification of an oxalamide derived chemotype, a novel class of highly potent, orally bioavailable IDO1-specific inhibitors was identified. Representative compound 18 inhibited human IDO1 with IC 50 values of 3.9 nM and 52 nM in a cellular and human whole blood assay, respectively. In vitro assessment of the ADME properties of 18 demonstrated very high metabolic stability. Pharmacokinetic profiling in mice showed a significantly reduced clearance compared to the oxalamides. In a mouse pharmacodynamic model 18 nearly completely suppressed lipopolysaccharide-induced kynurenine production. Hepatocyte data of 18 suggest the human clearance to be in a similar range to linrodostat (1)., (Copyright © 2020 Elsevier Ltd. All rights reserved.)

The bromodomain and extra terminal domain (BET) family members, including BRD2, BRD3, and BRD4, act as epigenetic readers to regulate gene expression. Indoleamine 2,3-dioxygenase 1 (IDO1) is an enzyme that participates in tumor immune escape primarily by catalyzing tryptophan to L-kynurenine. Here, we report that IDO1 is a new target gene of the BET family. RNA profiling showed that compound 9, a new BET inhibitor, reduced IDO1 mRNA up to seven times in Ty-82 cells. IDO1 differentially expressed in tumor cells and its expression could be induced with interferon gamma (IFN-γ). BET inhibitors (ABBV-075, JQ1, and OTX015) inhibited both constitutive and IFN-γ-inducible expression of IDO1. Similarly, reduction of BRD2, BRD3, or BRD4 decreased IDO1 expression. All these BET family members bound to the IDO1 promoter via the acetylated histone H3. JQ1 led to their release and reduced enrichment of RNA polymerase II (Pol II) on the promoter. IFN-γ increased the binding of BRD2, BRD3, BRD4, and Pol II on the IDO1 promoter by increasing the acetylation of histone H3, which could be prevented by JQ1 partially or even completely. Furthermore, both JQ1 and OTX015 decreased the production of L-kynurenine. The combination of BET inhibitors with the IDO1 inhibitor further reduced L-kynurenine, though only marginally. Importantly, the BET inhibitor ABBV-075 significantly inhibited the growth of human Ty-82 xenografts in nude mice and reduced both protein and mRNA levels of IDO1 in the xenografts. This finding lays a basis for the potential combination of BET inhibitors and IDO1 inhibitors for the treatment of IDO1-expressing cancers.

l-4-Chlorokynurenine (l-4-Cl-Kyn) is a neuropharmaceutical drug candidate that is in development for the treatment of major depressive disorder. Recently, this amino acid was naturally found as a residue in the lipopeptide antibiotic taromycin. Herein, we report the unprecedented conversion of l-tryptophan into l-4-Cl-Kyn catalyzed by four enzymes in the taromycin biosynthetic pathway from the marine bacterium Saccharomonospora sp. CNQ-490. We used genetic, biochemical, structural, and analytical techniques to establish l-4-Cl-Kyn biosynthesis, which is initiated by the flavin-dependent tryptophan chlorinase Tar14 and its flavin reductase partner Tar15. This work revealed the first tryptophan 2,3-dioxygenase (Tar13) and kynurenine formamidase (Tar16) enzymes that are selective for chlorinated substrates. The substrate scope of Tar13, Tar14, and Tar16 was examined and revealed intriguing promiscuity, thereby opening doors for the targeted engineering of these enzymes as useful biocatalysts., (© 2019 Wiley-VCH Verlag GmbH & Co. KGaA, Weinheim.)

Tryptophan-2,3-dioxygenase (TDO2), a rate-limiting enzyme in the tryptophan catabolism pathway, is induced in triple-negative breast cancer (TNBC) by inflammatory signals and anchorage-independent conditions. TNBCs express extremely low levels of the miR-200 family compared with estrogen receptor-positive (ER + ) breast cancer. In normal epithelial cells and ER + breast cancers and cell lines, high levels of the family member miR-200c serve to target and repress genes involved in epithelial-to-mesenchymal transition (EMT). To identify mechanism(s) that permit TNBC to express TDO2 and other proteins not expressed in the more well-differentiated ER + breast cancers, miRNA-200c was restored in TNBC cell lines. The data demonstrate that miR-200c targeted TDO2 directly resulting in reduced production of the immunosuppressive metabolite kynurenine. Furthermore, in addition to reversing a classic EMT signature, miR-200c repressed many genes encoding immunosuppressive factors including CD274/CD273, HMOX-1 , and GDF15 . Restoration of miR-200c revealed a mechanism, whereby TNBC hijacks a gene expression program reminiscent of that used by trophoblasts to suppress the maternal immune system to ensure fetal tolerance during pregnancy. IMPLICATIONS: Knowledge of the regulation of tumor-derived immunosuppressive factors will facilitate development of novel therapeutic strategies that complement current immunotherapy to reduce mortality for patients with TNBC., (©2018 American Association for Cancer Research.)

Kynurenine is biosynthesised from tryptophan catalysed by indoleamine 2,3-dioxygenase (IDO). The abrogation of kynurenine production is considered a promising therapeutic target for immunological cancer treatment. In the course of our IDO inhibitor programme, formal cyclisation of the isothiourea moiety of the IDO inhibitor 1 afforded the 5-Cl-benzimidazole derivative 2b-6, which inhibited both recombinant human IDO (rhIDO) activity and cellular kynurenine production. Further derivatisation of 2b-6 provided the potent inhibitor of cellular kynurenine production 2i (IC 50  = 0.34 µM), which unexpectedly exerted little effect on the enzymatic activity of rhIDO. Elucidation of the mechanism of action revealed that compound 2i suppresses IDO expression at the protein level by inhibiting STAT1 expression in IFN-γ-treated A431 cells. The kynurenine-production inhibitor 2i is expected to be a promising starting point for a novel approach to immunological cancer treatment., (Copyright © 2018 Elsevier Ltd. All rights reserved.)

Our antiviral arsenal to fight influenza viruses is limited and we need novel anti-flu drugs. Recently, cellular drug targets came into focus and omics analysis were instrumental to suggest candidate factors. In this issue of The FEBS Journal, Kainov and colleagues used transcriptome data to investigate virus-induced changes in tryptophan metabolism that may serve as immunomodulatory approach against viruses., (© 2017 Federation of European Biochemical Societies.)

Tryptophan 2,3-dioxygenase (TDO) and indoleamine 2,3-dioxygenase (IDO) play a central role in tryptophan metabolism and are involved in many cellular and disease processes. Here we report the crystal structure of human TDO (hTDO) in a ternary complex with the substrates L-Trp and O 2 and in a binary complex with the product N-formylkynurenine (NFK), defining for the first time the binding modes of both substrates and the product of this enzyme. The structure indicates that the dioxygenation reaction is initiated by a direct attack of O 2 on the C 2 atom of the L-Trp indole ring. The structure also reveals an exo binding site for L-Trp, located ~42 Å from the active site and formed by residues conserved among tryptophan-auxotrophic TDOs. Biochemical and cellular studies indicate that Trp binding at this exo site does not affect enzyme catalysis but instead it retards the degradation of hTDO through the ubiquitin-dependent proteasomal pathway. This exo site may therefore provide a novel L-Trp-mediated regulation mechanism for cellular degradation of hTDO, which may have important implications in human diseases.

Obesity is an increasingly urgent global problem, yet, little is known about its causes and less is known how obesity can be effectively treated. We showed previously that the aryl hydrocarbon receptor (AHR) plays a role in the regulation of body mass in mice fed Western diet. The AHR is a ligand-activated nuclear receptor that regulates genes involved in a number of biological pathways, including xenobiotic metabolism and T cell polarization. This study was an investigation into whether inhibition of the AHR prevents Western diet-based obesity. Male C57Bl/6J mice were fed control and Western diets with and without the AHR antagonist α-naphthoflavone or CH-223191, and a mouse hepatocyte cell line was used to delineate relevant cellular pathways. Studies are presented showing that the AHR antagonists α-naphthoflavone and CH-223191 significantly reduce obesity and adiposity and ameliorates liver steatosis in male C57Bl/6J mice fed a Western diet. Mice deficient in the tryptophan metabolizing enzyme indoleamine 2,3-dioxygenase 1 (IDO1) were also resistant to obesity. Using an AHR-directed, luciferase-expressing mouse hepatocyte cell line, we show that the transforming growth factor β1 (TGFβ1) signaling pathway via PI3K and NF-κB and the toll-like receptor 2/4 (TLR2/4) signaling pathway stimulated by oxidized low-density lipoproteins via NF-κB, each induce luciferase expression; however, TLR2/4 signaling was significantly reduced by inhibition of IDO1. At physiological levels, kynurenine but not kynurenic acid (both tryptophan metabolites and known AHR agonists) activated AHR-directed luciferase expression. We propose a hepatocyte-based model, in which kynurenine production is increased by enhanced IDO1 activity stimulated by TGFβ1 and TLR2/4 signaling, via PI3K and NF-κB, to perpetuate a cycle of AHR activation to cause obesity; and inhibition of the AHR, in turn, blocks the cycle's output to prevent obesity. The AHR with its broad ligand binding specificity is a promising candidate for a potentially simple therapeutic approach for the prevention and treatment of obesity and associated complications., (Copyright © 2016 Elsevier Inc. All rights reserved.)

Indoleamine 2,3-dioxygenase (IDO), a tryptophan-catabolizing intracellular enzyme of the L-kynurenine pathway, causes preneoplastic cells and tumor cells to escape the immune system by inducing immune tolerance; this mechanism might be associated with the development and progression of human malignancies. In the present study, we investigated the role of IDO in diethylnitrosamine (DEN)-induced hepatocarcinogenesis by using IDO-knockout (KO) mice. To induce hepatocellular carcinoma (HCC), hepatic adenoma, and preneoplastic hepatocellular lesions termed foci of cellular alteration (FCA), male IDO-wild-type (WT) and IDO-KO mice with a C57BL/6J background received a single intraperitoneal injection of DEN at 2 weeks of age. The mice were sacrificed to evaluate the development of FCA and hepatocellular neoplasms. HCC overexpressed IDO and L-kynurenine compared to surrounding normal tissue in the DEN-treated IDO-WT mice. The number and cell proliferative activity of FCAs, and the incidence and multiplicity of HCC were significantly greater in the IDO-WT than in the IDO-KO mice. The expression levels of the IDO protein, of L-kynurenine, and of IFN-γ, COX-2, TNF-α, and Foxp3 mRNA were also significantly increased in the DEN-induced hepatic tumors that developed in the IDO-WT mice. The mRNA expression levels of CD8, perforin and granzyme B were markedly increased in hepatic tumors developed in IDO-KO mice. Moreover, Foxp3-positive inflammatory cells had infiltrated into the livers of DEN-treated IDO-WT mice, whereas fewer cells had infiltrated into the livers of IDO-KO mice. Induction of IDO and elevation of L-kynurenine might play a critical role in both the early and late phase of liver carcinogenesis. Our findings suggest that inhibition of IDO might offer a promising strategy for the prevention of liver cancer.

A key link between amino acid catabolism and immune regulation in cancer is the augmented tryptophan (Trp) catabolism through the kynurenine pathway (KP), a metabolic route induced by interferon-γ (IFN-γ) and related to poor prognosis in melanomas. Besides its role in cancer, IFN-γ plays a key role in the control of pigmentation homeostasis. Here we measured KP metabolites in human melanoma lines and skin melanocytes and fibroblasts in response to IFN-γ. In general, IFN-γ affected KP in skin cells more than in melanoma cells, supporting IFN-γ roles in skin physiology and that of stromal cells in modulating the tumor microenvironment.

Batrachochytrium dendrobatidis is a fungal pathogen in the phylum Chytridiomycota that causes the skin disease chytridiomycosis. Chytridiomycosis is considered an emerging infectious disease linked to worldwide amphibian declines and extinctions. Although amphibians have well-developed immune defenses, clearance of this pathogen from the skin is often impaired. Previously, we showed that the adaptive immune system is involved in the control of the pathogen, but B. dendrobatidis releases factors that inhibit in vitro and in vivo lymphocyte responses and induce lymphocyte apoptosis. Little is known about the nature of the inhibitory factors released by this fungus. Here, we describe the isolation and characterization of three fungal metabolites produced by B. dendrobatidis but not by the closely related nonpathogenic chytrid Homolaphlyctis polyrhiza. These metabolites are methylthioadenosine (MTA), tryptophan, and an oxidized product of tryptophan, kynurenine (Kyn). Independently, both MTA and Kyn inhibit the survival and proliferation of amphibian lymphocytes and the Jurkat human T cell leukemia cell line. However, working together, they become effective at much lower concentrations. We hypothesize that B. dendrobatidis can adapt its metabolism to release products that alter the local environment in the skin to inhibit immunity and enhance the survival of the pathogen., (Copyright © 2015, American Society for Microbiology. All Rights Reserved.)

Ethnopharmacological Relevance: Danzhi Xiaoyao San (DXS) is a canonical Chinese medicine formula from Principles of Internal Medicine, which was written during the Ming dynasty. This formula is approved and commercialized for use in the prevention and treatment of affective disorders. This study is aimed to investigate the hypothesis that DXS treats depressive-like behavior by shifting the balance of the kynurenine (Kyn)/serotonin (5-HT) pathway toward the 5-HT pathway through the downregulation of hippocampal indoleamine 2,3-dioxygenase (IDO)., Materials and Methods: Chemical fingerprints of gardenoside, paeoniflorin, ferulic acid, paeonol, and ligustilide in standard extraction were used as the material bases of DXS. Rats with depressive-like behavior induced by chronic unpredictable mild stress (CUMS) were randomly divided into four groups, namely the control, model, DXS, and fluoxetine groups. Cytokines, IDO, and tryptophan (Trp) catabolites were analyzed by enzyme-linked immunosorbent assay, western blot, and liquid chromatography-electrospray ionization tandem mass spectrometry, respectively., Results: DXS significantly increased crossing grid numbers, sucrose consumption, and body weight. This treatment significantly decreased the serum levels of tumor necrosis factor-α and interleukin 6 (IL-6). However, DXS elicited no significant effects on IL-1β, IL-2, and interferon γ. DXS downregulated the activity of IDO and subsequent production of Kyn in the hippocampus. This treatment upregulated the hippocampal contents of Trp and 5-HT but did not influence 5-HT turnover., Conclusions: DXS exhibited antidepressant-like effects on rats exposed to CUMS. DXS reduced IDO activity to shift the balance of the Kyn/5-HT pathway toward the 5-HT pathway., (Copyright © 2014 Elsevier Ireland Ltd. All rights reserved.)

The kynurenine pathway (KP) is the principal route of L-tryptophan (TRP) catabolism leading to the production of kynurenine (KYN), the neuroprotectants, kynurenic acid (KYNA) and picolinic acid (PIC), the excitotoxin, quinolinic acid (QUIN) and the essential pyridine nucleotide, nicotinamide adenine dinucleotide (NAD(+)). The enzymes indoleamine 2,3-dioxygenase-1 (IDO-1), indoleamine 2,3-dioxygenase-2 (IDO-2) and tryptophan 2,3-dioxygenase (TDO-2) initiate the first step of the KP. IDO-1 and TDO-2 induction in tumors are crucial mechanisms implicated to play pivotal roles in suppressing anti-tumor immunity. Here, we report the first comprehensive characterisation of the KP in 1) cultured human glioma cells and 2) plasma from patients with glioblastoma (GBM). Our data revealed that interferon-gamma (IFN-γ) stimulation significantly potentiated the expression of the KP enzymes, IDO-1 IDO-2, kynureninase (KYNU), kynurenine hydroxylase (KMO) and significantly down-regulated 2-amino-3-carboxymuconate semialdehyde decarboxylase (ACMSD) and kynurenine aminotransferase-I (KAT-I) expression in cultured human glioma cells. This significantly increased KP activity but significantly lowered the KYNA/KYN neuroprotective ratio in human cultured glioma cells. KP activation (KYN/TRP) was significantly higher, whereas the concentrations of the neuroreactive KP metabolites TRP, KYNA, QUIN and PIC and the KYNA/KYN ratio were significantly lower in GBM patient plasma (n = 18) compared to controls. These results provide further evidence for the involvement of the KP in glioma pathophysiology and highlight a potential role of KP products as novel and highly attractive therapeutic targets to evaluate for the treatment of brain tumors, aimed at restoring anti-tumor immunity and reducing the capacity for malignant cells to produce NAD(+), which is necessary for energy production and DNA repair.

Indoleamine 2,3-dioxygenase (IDO) is a rate-limiting enzyme in tryptophan catabolism that plays an important role in the induction of immune tolerance. Its role in graft-versus-tumor effect after allogeneic stem cell transplantation (allo-SCT) remains unclear. Using a murine graft-versus-tumor model of reduced-intensity allo-HSCT followed by donor leukocyte infusion (DLI), we examined the role of IDO inhibition. Two stereoisomers of 1-methyl tryptophan (1-MT), a small-molecule inhibitor of IDO, reduced the growth of inoculated tumor in the mice that received DLI and had higher expression of IDO1 and IFNγ. However, L-1MT, but not D-1MT, mitigated tumor growth in mice that did not receive DLI and did not express IDO1 and IFNγ. Accordingly, both stereoisomers reduced plasma kynurenine concentrations early after DLI and enhanced in vitro cytotoxic lymphocyte function after allogeneic mixed lymphocyte reaction. Furthermore, L-1MT was more efficient in causing direct cytotoxic effects than D-1MT. Our results suggest that IDO inhibition can benefit anti-tumor therapy in the setting of reduced-intensity allo-SCT using DLI., (Copyright © 2014 ISEH - International Society for Experimental Hematology. Published by Elsevier Inc. All rights reserved.)

Pseudomonas aeruginosa is responsible for persistent infections in cystic fibrosis patients, suggesting an ability to circumvent innate immune defenses. This bacterium uses the kynurenine pathway to catabolize tryptophan. Interestingly, many host cells also produce kynurenine, which is known to control immune system homeostasis. We showed that most strains of P. aeruginosa isolated from cystic fibrosis patients produce a high level of kynurenine. Moreover, a strong transcriptional activation of kynA (the first gene involved in the kynurenine pathway) was observed upon contact with immune cells and particularly with neutrophils. In addition, using coculture of human neutrophils with various strains of P. aeruginosa producing no (ΔkynA) or a high level of kynurenine (ΔkynU or ΔkynA pkynA), we demonstrated that kynurenine promotes bacterial survival. In addition, increasing the amount kynurenine inhibits reactive oxygen species production by activated neutrophils, as evaluated by chemiluminescence with luminol or isoluminol or SOD-sensitive cytochrome c reduction assay. This inhibition is due neither to a phagocytosis defect nor to direct NADPH oxidase inhibition. Indeed, kynurenine has no effect on oxygen consumption by neutrophils activated by PMA or opsonized zymosan. Using in vitro reactive oxygen species-producing systems, we showed that kynurenine scavenges hydrogen peroxide and, to a lesser extent, superoxide. Kynurenine׳s scavenging effect occurs mainly intracellularly after bacterial stimulation, probably in the phagosome. In conclusion, the kynurenine pathway allows P. aeruginosa to circumvent the innate immune response by scavenging neutrophil reactive oxygen species production., (Copyright © 2014 Elsevier Inc. All rights reserved.)

During early brain development, N-methyl-d-aspartate (NMDA) receptors are involved in cell migration, neuritogenesis, axon guidance and synapse formation, but the mechanisms which regulate NMDA receptor density and function remain unclear. The kynurenine pathway of tryptophan metabolism includes an agonist (quinolinic acid) and an antagonist (kynurenic acid) at NMDA receptors and we have previously shown that inhibition of the pathway using the kynurenine-3-monoxygenase inhibitor Ro61-8048 in late gestation produces rapid changes in protein expression in the embryos and effects on synaptic transmission lasting until postnatal day 21 (P21). The present study sought to determine whether any of these effects are maintained into adulthood. After prenatal injections of Ro61-8048 the litter was allowed to develop to P60 when some offspring were euthanized and the brains removed for examination. Analysis of protein expression by Western blotting revealed significantly reduced expression of the GluN2A subunit (32%) and the morphogenetic protein sonic hedgehog (31%), with a 29% increase in the expression of doublecortin, a protein associated with neurogenesis. No changes were seen in mRNA abundance using quantitative real-time polymerase chain reaction. Neuronal excitability was normal in the CA1 region of hippocampal slices but paired-pulse stimulation revealed less inhibition at short interpulse intervals. The amount of long-term potentiation was decreased by 49% in treated pups and recovery after low-frequency stimulation was delayed. The results not only strengthen the view that basal, constitutive kynurenine metabolism is involved in normal brain development, but also show that changes induced prenatally can affect the brains of adult offspring and those changes are quite different from those seen previously at weaning (P21). Those changes may be mediated by altered expression of NMDAR subunits and sonic hedgehog., (Copyright © 2013 IBRO. Published by Elsevier Ltd. All rights reserved.)

Kynurenine monooxygenase (KMO) catalyzes the conversion of kynurenine to 3-hydroxykynurenine. Modulation of KMO activity has been implicated in several neurodegenerative diseases, including Huntington disease. Our goal is to develop potent and selective small-molecule KMO inhibitors with suitable pharmacokinetic characteristics for in vivo proof-of-concept studies and subsequent clinical development. We developed a comprehensive panel of biochemical and cell-based assays that use liquid chromatography/tandem mass spectrometry to quantify unlabeled kynurenine and 3-hydroxykynurenine. We describe assays to measure KMO inhibition in cell and tissue extracts, as well as cellular assays including heterologous cell lines and primary rat microglia and human peripheral blood mononuclear cells.

Indoleamine 2,3-dioxygenase 1 (IDO1) metabolizes L-tryptophan to kynurenines (KYN), inducing T-cell suppression either directly or by altering antigen-presenting-cell function. Cyclooxygenase (COX)-2, the rate-limiting enzyme in the synthesis of prostaglandins, is over-expressed by several tumours. We aimed at determining whether COX-2 inhibitors down-regulate the IFN-g-induced expression of IDO1 in acute myeloid leukaemia (AML) cells. IFN-γ at 100 ng/mL up-regulated COX-2 and IDO1 in HL-60 AML cells, both at mRNA and protein level. The increased COX-2 and IDO1 expression correlated with heightened production of prostaglandin (PG)E₂ and kynurenines, respectively. Nimesulide, a preferential COX-2 inhibitor, down-regulated IDO1 mRNA/protein and attenuated kynurenine synthesis, suggesting that overall IDO inhibition resulted both from reduced IDO1 gene transcription and from inhibited IDO1 catalytic activity. From a functional standpoint, IFN-g-challenged HL-60 cells promoted the in vitro conversion of allogeneic CD4⁺CD25⁻ T cells into bona fide CD4⁺CD25⁺FoxP3⁺ regulatory T cells, an effect that was significantly reduced by treatment of IFN-γ-activated HL-60 cells with nimesulide. Overall, these data point to COX-2 inhibition as a potential strategy to be pursued with the aim at circumventing leukaemia-induced, IDO-mediated immune dysfunction.

NAD(+) is synthesized from tryptophan either via the kynurenine (de novo) pathway or via the salvage pathway by reutilizing intermediates such as nicotinic acid or nicotinamide ribose. Quinolinic acid is an intermediate in the kynurenine pathway. We have discovered that the budding yeast Saccharomyces cerevisiae secretes quinolinic acid into the medium and also utilizes extracellular quinolinic acid as a novel NAD(+) precursor. We provide evidence that extracellular quinolinic acid enters the cell via Tna1, a high-affinity nicotinic acid permease, and thereby helps to increase the intracellular concentration of NAD(+). Transcription of genes involved in the kynurenine pathway and Tna1 was increased, responding to a low intracellular NAD(+) concentration, in cells bearing mutations of these genes; this transcriptional induction was suppressed by supplementation with quinolinic acid or nicotinic acid. Our data thus shed new light on the significance of quinolinic acid, which had previously been recognized only as an intermediate in the kynurenine pathway.

IDO2 is a newly discovered enzyme with 43 % similarity to classical IDO (IDO1) protein and shares the same critical catalytic residues. IDO1 catalyzes the initial and rate-limiting step in the degradation of tryptophan and is a key enzyme in mediating tumor immune tolerance via arrest of T cell proliferation. The role of IDO2 in human T cell immunity remains controversial. Here, we demonstrate that similar to IDO1, IDO2 also degrades tryptophan into kynurenine and is inhibited more efficiently by Levo-1-methyl tryptophan (L-1MT), an IDO1 competitive inhibitor, than by dextro-methyl tryptophan (D-1MT). Although IDO2 enzyme activity is weaker than IDO1, it is less sensitive to 1-MT inhibition than IDO1. Moreover, our results indicate that human CD4(+) and CD8(+) T cell proliferation was inhibited by IDO2, but both L-1MT and D-1MT could not reverse IDO2-mediated arrest of cell proliferation, even at high concentrations. These data indicate that IDO2 is an inhibitory mechanism in human T cell proliferation and support efforts to develop more effective IDO1 and IDO2 inhibitors in order to overcome IDO-mediated immune tolerance.

Kynurenic acid (KYNA), an antagonist of the α7 nicotinic acetylcholine receptor and the N-methyl-D-aspartate receptor, and 3-hydroxykynurenine (3-HK), a generator of reactive oxygen species, are neuroactive metabolites of the kynurenine pathway of tryptophan degradation. In the mammalian brain as elsewhere, both compounds derive from a common bioprecursor, L-kynurenine (L-KYN). Recent studies in rats demonstrated that D-kynurenine (D-KYN), a metabolite of the bacterial amino acid D-tryptophan, can also function as a bioprecursor of brain KYNA. We now investigated the conversion of systemically administered D-KYN to KYNA in mice and also explored the possible production of 3-HK in the same animals. Thirty min after an injection of D-KYN or L-KYN (30 mg/kg, i.p.), newly produced KYNA and 3-HK were recovered from plasma, liver, forebrain and cerebellum in all cases. Using a new chiral separation method, 3-HK produced from D-KYN was positively identified as D-3-HK. L-KYN was the more effective precursor of KYNA in all tissues and also exceeded D-KYN as a precursor of brain 3-HK. In contrast, D-KYN was more potent as a precursor of 3-HK in the liver. The production of both KYNA and 3-HK from D-KYN was rapid in all tissues, peaking at 15-30 min following a systemic injection of D-KYN. These results show that biosynthetic routes other than those classically ascribed to L-KYN can account for the synthesis of both KYNA and 3-HK in vivo. This new insight may be of significant physiological or pathological relevance., (Copyright © 2012 Elsevier B.V. All rights reserved.)

Background: The kynurenic acid (KYNA) hypothesis for schizophrenia is partly based on studies showing increased brain levels of KYNA in patients. KYNA is an endogenous metabolite of tryptophan (TRP) produced in astrocytes and antagonizes N-methyl-D-aspartate and α7* nicotinic receptors., Methods: The formation of KYNA is determined by the availability of substrate, and hence, we analyzed KYNA and its precursors, kynurenine (KYN) and TRP, in the cerebrospinal fluid (CSF) of patients with schizophrenia. CSF from male patients with schizophrenia on olanzapine treatment (n = 16) was compared with healthy male volunteers (n = 29)., Results: KYN and KYNA concentrations were higher in patients with schizophrenia (60.7 ± 4.37 nM and 2.03 ± 0.23 nM, respectively) compared with healthy volunteers (28.6 ± 1.44 nM and 1.36 ± 0.08 nM, respectively), whereas TRP did not differ between the groups. In all subjects, KYN positively correlated to KYNA., Conclusion: Our results demonstrate increased levels of CSF KYN and KYNA in patients with schizophrenia and further support the hypothesis that KYNA is involved in the pathophysiology of schizophrenia.

Kynurenine (Kyn), a metabolite of tryptophan (Trp), is known to be a key regulator of human immune responses including cancer immune tolerance. Therefore, abrogation of Kyn production from cancer cells by small molecules may be a promising approach to anticancer therapy. Indeed, several small molecule inhibitors of indoleamine 2,3-dioxygenase (IDO), a rate-limiting enzyme in the catabolism of Trp to Kyn, exert antitumor effects in animal models. We screened our chemical libraries using a cell-based Kyn production assay to identify a new type of small molecules that regulate Kyn production, and for the first time identified a benzenesulfonamide derivative (compound 1) as a hit with the ability to inhibit Kyn production in interferon-γ (IFN-γ)-stimulated A431 and HeLa cells. Unlike the previously identified S-benzylisothiourea derivative, compound 2, compound 1 had little effect on the enzymatic activity of recombinant human IDO in vitro but suppressed the expression of IDO at the mRNA level in cells. Furthermore, compound 1 suppressed STAT1-dependent transcriptional activity and DNA binding, whereas no decrement in either the expression or phosphorylation level of STAT1 was observed. The inhibition of IDO expression by several benzenesulfonamide derivatives is associated with the suppression of STAT1. Thus, compound 1 and its analogs might be useful for analyzing the regulation of IDO activation, and STAT1-targeting could be an alternative to the IDO-directed approach for the regulation of Kyn levels by small molecules in the tumor microenvironment., (Copyright © 2012 Elsevier Inc. All rights reserved.)

Increased inflammation and reduced neurogenesis have been associated with the pathophysiology of major depression. Here, we show for the first time how IL-1β, a pro-inflammatory cytokine shown to be increased in depressed patients, decreases neurogenesis in human hippocampal progenitor cells. IL-1β was detrimental to neurogenesis, as shown by a decrease in the number of doublecortin-positive neuroblasts (-28%), and mature, microtubule-associated protein-2-positive neurons (-36%). Analysis of the enzymes that regulate the kynurenine pathway showed that IL-1β induced an upregulation of transcripts for indolamine-2,3-dioxygenase (IDO), kynurenine 3-monooxygenase (KMO), and kynureninase (42-, 12- and 30-fold increase, respectively, under differentiating conditions), the enzymes involved in the neurotoxic arm of the kynurenine pathway. Moreover, treatment with IL-1β resulted in an increase in kynurenine, the catabolic product of IDO-induced tryptophan metabolism. Interestingly, co-treatment with the KMO inhibitor Ro 61-8048 reversed the detrimental effects of IL-1β on neurogenesis. These observations indicate that IL-1β has a critical role in regulating neurogenesis whereas affecting the availability of tryptophan and the production of enzymes conducive to toxic metabolites. Our results suggest that inhibition of the kynurenine pathway may provide a new therapy to revert inflammatory-induced reduction in neurogenesis.

Although tuberculosis (TB) causes more deaths than any other pathogen, most infected individuals harbor the pathogen without signs of disease. We explored the metabolome of >400 small molecules in serum of uninfected individuals, latently infected healthy individuals and patients with active TB. We identified changes in amino acid, lipid and nucleotide metabolism pathways, providing evidence for anti-inflammatory metabolomic changes in TB. Metabolic profiles indicate increased activity of indoleamine 2,3 dioxygenase 1 (IDO1), decreased phospholipase activity, increased abundance of adenosine metabolism products, as well as indicators of fibrotic lesions in active disease as compared to latent infection. Consistent with our predictions, we experimentally demonstrate TB-induced IDO1 activity. Furthermore, we demonstrate a link between metabolic profiles and cytokine signaling. Finally, we show that 20 metabolites are sufficient for robust discrimination of TB patients from healthy individuals. Our results provide specific insights into the biology of TB and pave the way for the rational development of metabolic biomarkers for TB.

Objectives: To determine whether tryptophan metabolism to kynurenine contributes to the direct regulation of vascular tone in human septic shock., Background: Indoleamine 2,3-dioxygenase 1 is an inducible enzyme that converts tryptophan to kynurenine and shares functional similarities with inducible nitric oxide synthase. Recently, kynurenine has been identified as an endothelium-derived relaxing factor produced during inflammation, raising the possibility that this novel pathway may contribute to hypotension in human sepsis., Design: Prospective, matched, single-center, cohort study., Settings: Intensive care unit of a tertiary teaching hospital matched to control subjects from the general medical ward and healthy volunteers., Subjects: Patients (n = 16) with septic shock had indoleamine 2,3-dioxygenase activity assessed as the kynurenine-to-tryptophan ratio, and the severity of hypotension was determined by their inotrope requirements. Healthy and blood pressure-matched nonseptic control subjects were also studied., Interventions: None., Measurements and Main Results: Tissues from septic and control patients were stained for the presence of indoleamine 2,3-dioxygenase 1. Indoleamine 2,3-dioxygenase activity increased up to ninefold in patients with septic shock and was significantly higher than in the two control groups (p < .01). Indoleamine 2,3-dioxygenase activity was strongly correlated with inotrope requirements (p < .001). Indoleamine 2,3-dioxygenase protein was expressed in inflamed cardiac tissue as well as in endothelial cells of resistance vessels in hearts and kidneys from subjects who died from sepsis., Conclusions: : Indoleamine 2,3-dioxygenase 1 is expressed in resistance vessels in human sepsis and Indoleamine 2,3-dioxygenase activity correlates with hypotension in human septic shock. Indoleamine 2,3-dioxygenase 1 is thus a potential novel contributor to hypotension in sepsis.

Kynurenine is a small molecule derived from tryptophan when this amino acid is metabolised via the kynurenine pathway. The biological activity of kynurenine and its metabolites (kynurenines) is well recognised. Therefore, understanding the regulation of the subsequent biochemical reactions is essential for the design of therapeutic strategies which aim to interfere with the kynurenine pathway. However, kynurenine concentration in the body may not only be determined by the efficiency of kynurenine synthesis but also by the rate of kynurenine clearance. In this review, current knowledge about the mechanisms of kynurenine production and routes of its clearance is presented. In addition, the involvement of kynurenine and its metabolites in the biology of different T cell subsets (including Th17 cells and regulatory T cells) and neuronal cells is discussed.

Background: Inflammation has been linked to depression and suicide risk. One inflammatory process that has been minimally investigated in this regard is cytokine-stimulated production of kynurenine (KYN) from tryptophan (TRP). Recent data suggest that KYN increases in cerebrospinal fluid (CSF) are associated with depressive symptoms secondary to immune activation. KYN may alter dopaminergic and glutamatergic tone, thereby contributing to increased arousal, agitation and impulsivity - important risk factors in suicide. We hypothesized that patients with major depressive disorder (MDD) and a history of suicide attempt would have higher levels of KYN than depressed nonattempters, who in turn would have higher levels than healthy volunteers., Methods: Plasma KYN, TRP, and neopterin were assayed by high performance liquid chromatography in three groups: healthy volunteers (n=31) and patients with MDD with (n=14) and without (n=16) history of suicide attempt. Analysis of variance tested for group differences in KYN levels., Results: KYN levels differed across groups (F=4.03, df=(2,58), and p=0.023): a priori planned contrasts showed that KYN was higher in the MDD suicide attempter subgroup compared with MDD non-attempters (t=2.105, df=58, and p=0.040), who did not differ from healthy volunteers (t=0.418, df=58, and p=0.677). In post hoc testing, KYN but not TRP was associated with attempt status, and only suicide attempters exhibited a positive correlation of the cytokine activation marker neopterin with the KYN:TRP ratio, suggesting that KYN production may be influenced by inflammatory processes among suicide attempters., Conclusion: These preliminary results suggest that KYN and related molecular pathways may be implicated in the pathophysiology of suicidal behavior., (Copyright © 2011. Published by Elsevier Inc.)

Indoleamine 2,3-dioxygenase (IDO) is the first and rate-limiting enzyme of tryptophan catabolism through the kynurenine pathway. Intriguingly, IDO is constitutively and highly expressed in the mammalian epididymis in contrast to most other tissues where IDO is induced by proinflammatory cytokines, such as interferons. To gain insight into the role of IDO in the physiology of the mammalian epididymis, we studied both wild type and Ido1(-/-)-deficient mice. In the caput epididymis of Ido1(-/-) animals, the lack of IDO activity was not compensated by other tryptophan-catabolizing enzymes and led to the loss of kynurenine production. The absence of IDO generated an inflammatory state in the caput epididymis as revealed by an increased accumulation of various inflammation markers. The absence of IDO also increased the tryptophan content of the caput epididymis and generated a parallel increase in caput epididymal protein content as a consequence of deficient proteasomal activity. Surprisingly, the lack of IDO expression had no noticeable impact on overall male fertility but did induce highly significant increases in both the number and the percentage of abnormal spermatozoa. These changes coincided with a significant decrease in white blood cell count in epididymal fluid compared with wild type mice. These data provide support for IDO playing a hitherto unsuspected role in sperm quality control in the epididymis involving the ubiquitination of defective spermatozoa and their subsequent removal.

Quinomycin antibiotics, represented by echinomycin, are an important class of antitumor antibiotics. We have recently succeeded in identification of biosynthetic gene clusters of echinomycin and SW-163D, and have achieved heterologous production of echinomycin in Escherichia coli. In addition, we have engineered echinomycin non-ribosomal peptide synthetase to generate echinomycin derivatives. However, the biosynthetic pathways of intercalative chromophores quinoxaline-2-carboxylic acid (QXC) and 3-hydroxyquinaldic acid (HQA), which are important for biological activity, were not fully elucidated. Here, we report experiments involving incorporation of a putative advanced precursor, (2S, 3R)-[6'-(2)H]-3-hydroxy-L-kynurenine, and functional analysis of the enzymes Swb1 and Swb2 responsible for late-stage biosynthesis of HQA. On the basis of these experimental results, we propose biosynthetic pathways for both QXC and HQA through the common intermediate 3-hydroxy-L-kynurenine.

Indoleamine 2,3-dioxygenase (IDO), an enzyme expressed in many cell types, catalyses degradation of tryptophan (Trp) to kynurenine (Kyn) and may exert immunosuppressive functions, mediated mainly by kynurenines. Therefore, increased Kyn concentrations would be expected to protect allografts from rejection. We conducted this study to examine whether Kyn has predictive value for kidney graft outcome. End-stage renal disease patients (n = 210) demonstrated an increased Kyn/Trp ratio compared with healthy controls (n = 30). Both Kyn and Trp levels were significantly higher in patients who subsequently developed acute rejection than in patients who did not (p < 0.001 and p < 0.001, respectively). Furthermore, pretransplantation Kyn and Trp plasma concentrations were significantly different in patients who went on to develop acute rejection (high values) or acute tubular necrosis (low values) (p = 0.007 and p = 0.021, respectively). After transplantation Kyn levels decreased. Approximately 3 days before biopsy-confirmed rejection, Kyn was significantly increased in patients with rejection compared with those without rejection (p < 0.001). Contrary to expectation, high Kyn plasma levels before transplantation were not predictive of low rejection risk. Although informative in overall terms, at the present stage, Kyn levels do not allow the concise risk differentiation of individual patients., (Copyright © 2010 American Society for Histocompatibility and Immunogenetics. Published by Elsevier Inc. All rights reserved.)

Control of blood vessel tone is central to vascular homeostasis. Here we show that metabolism of tryptophan to kynurenine by indoleamine 2,3-dioxygenase (Ido) expressed in endothelial cells contributes to arterial vessel relaxation and the control of blood pressure. Infection of mice with malarial parasites (Plasmodium berghei) or induction of endotoxemia in mice led to endothelial expression of Ido, decreased plasma tryptophan concentration, increased kynurenine concentration and hypotension. Pharmacological inhibition of Ido increased blood pressure in systemically inflamed mice but not in mice deficient in Ido or interferon-gamma, which is required for Ido induction. Both tryptophan and kynurenine dilated preconstricted porcine coronary arteries; the dilating effect of tryptophan required the presence of active Ido and an intact endothelium, whereas the effect of kynurenine was endothelium independent. The arterial relaxation induced by kynurenine was mediated by activation of the adenylate and soluble guanylate cyclase pathways. Kynurenine administration decreased blood pressure in a dose-dependent manner in spontaneously hypertensive rats. Our results identify tryptophan metabolism by Ido as a new pathway contributing to the regulation of vascular tone.

Mesenchymal stem cells (MSC) display unique suppressive properties on T-cell immunity, thus representing an attractive vehicle for the treatment of conditions associated with harmful T-cell responses such as organ-specific autoimmunity and graft-versus-host disease. Toll-like receptors (TLR) are primarily expressed on antigen-presenting cells and recognize conserved pathogen-derived components. Ligation of TLR activates multiple innate and adaptive immune response pathways to eliminate and protect against invading pathogens. In this work, we show that TLR expressed on human bone marrow-derived MSC enhanced the immunosuppressive phenotype of MSC. Immunosuppression mediated by TLR was dependent on the production of immunosuppressive kynurenines by the tryptophan-degrading enzyme indoleamine-2,3-dioxygenase-1 (IDO1). Induction of IDO1 by TLR involved an autocrine interferon (IFN)-beta signaling loop, which was dependent on protein kinase R (PKR), but independent of IFN-gamma. These data define a new role for TLR in MSC immunobiology, which is to augment the immunosuppressive properties of MSC in the absence of IFN-gamma rather than inducing proinflammatory immune response pathways. PKR and IFN-beta play a central, previously unidentified role in orchestrating the production of immunosuppressive kynurenines by MSC.

Background: Recently the L-tryptophan (Trp) metabolites such as L-kynurenine(Kyn), L-kinurenic acid, quinolinic acid (QA) and picolinic acid (PA) have been shown physiologically important in central nervous and immune system, and various enzyme activities concerning their production were reported to be affected by insulin-dependent diabetes mellitus. However, the states of these metabolites in diabetes have not been clarified enough yet., Aim of Study: The present study was performed to make clear the states of the productions of L-Kyn, QA, PA and nicotinamide (Nam) in vitro in the hepatocytes prepared from streptozotocin (STZ)-induced diabetic rats using [5-3H]L-Trp., Methods: The diabetic model rats were made by STZ injection (60 mg/kg) and the hepatocytes isolated from the rats were incubated with [5-3H]L-Trp. The amounts of metabolites derived from L-Trp were determined by the isotope-dilution methods., Results: The alpha-amino-beta-carboxymuconate-epsilon-semiarldehyde decarboxylase (ACMSD) mRNA level in the diabetic group was greatly higher than that in the control group. In the STZ-induced diabetes group, the amount of [5-3H]L-Trp converted to tritiated water, L-Kyn or QA were found to be more than 3 times of that in the control group, respectively. The produced amounts of PA and Nam were not significantly different between the diabetic and the control groups., Conclusions: It is suggested that STZ-diabetes mellitus causes augmentations of both L-Kyn and QA generations but not those of PA and Nam in liver, indicating the possibility that the immune and neuronal systems of insulin dependent diabetes mellitus would be influenced by the increased amounts of LKyn and QA but not by those of PA and Nam.

Objective: Surgical results have shown the superiority of human heart valve and vascular allografts over artificial prostheses when used for the treatment of infectious cardiovascular diseases. However, the mechanism of infection resistance in these allografts has not been determined. In this study the contribution of the inflammatory response after allogeneic transplantation to the antimicrobial mechanism was assessed, focusing on the induction of indoleamine 2,3-dioxygenase, a tryptophan-metabolizing enzyme., Methods: Aortic transplantation was performed with inbred rats, and aortic allografts, isografts, and control grafts were obtained for the following analyses. The extent of inflammatory-related and indoleamine 2,3-dioxygenase gene expression was measured by means of quantitative reverse transcriptase-polymerase chain reaction, and tryptophan metabolite production in the graft was measured by means of liquid chromatographic/tandem mass spectrometric analysis. The bacteriostatic effect of each graft and tryptophan metabolites was determined by using the methicillin-resistant Staphylococcus aureus proliferation assay., Results: The inflammatory response, including interferon gamma, tumor necrosis factor alpha, and indoleamine 2,3-dioxygenase gene expression, was significant in the allografts but minimal in the isografts and control grafts. Methicillin-resistant S. aureus proliferation was remarkably suppressed when cultured with the allografts but not with the control grafts. Among tryptophan metabolites, the bacteriostatic effect against methicillin-resistant S. aureus was remarkable with 3-hydroxykynurenine, with a minimum inhibitory concentration of 32 mg/L. The 3-hydroxykynurenine level in the allografts was 9-fold greater than that in the control grafts., Conclusion: The bacteriostatic effect of the allografts was acquired by inducing indoleamine 2,3-dioxygenase, which resulted in local production of 3-hydroxykynurenine as an antimicrobial agent. This is the first report to document a mechanism of the allograft's infection-resistant property against methicillin-resistant S. aureus growth.

Indoleamine 2,3-dioxygenase (IDO) is an heme-containing enzyme involved in the regulation of important immunological responses and neurological processes. The enzyme catalyzes the oxidative cleavage of the pyrrole ring of the indole nucleus of tryptophan (Trp) to yield N-formylkynurenine, that is the initial and rate limiting step of the kynurenine pathway. Some indole derivatives have been reported to act as effectors of the enzyme by enhancing its catalytic activity. On the basis of the recent availability of the crystal structure of IDO, in this work we investigate substrate recognition and enhancer binding to IDO using molecular docking experiments. In addition, conformational transitions of IDO in response to substrate and enhancer binding are studied using coarse graining simulations with the program FIRST. The results enable us to identify (i) the binding site of enhancer modulators; (ii) the motion of an electrostatic gate that regulates the access of the substrate to the catalytic site of the enzyme; (iii) the movement of the anchoring region of a hairpin loop that may assist the shuttle of substrates/products to/from the catalytic site of IDO. These data, combined with available site-directed mutagenesis experiments, reveal that conformational transitions of IDO in response to substrate and enhancer binding are controlled by distinct combination of two conformational states (open and close) of the above structural motifs. On this basis, a molecular mechanism regarding substrate recognition and activity enhancement by indole derivatives is proposed.

The metabolism of the amino acid L-tryptophan is a highly regulated physiological process leading to the generation of several neuroactive compounds within the central nervous system. These include the aminergic neurotransmitter serotonin (5-hydroxytryptamine, 5-HT), products of the kynurenine pathway of tryptophan metabolism (including 3-hydroxykynurenine, 3-hydroxyanthranilic acid, quinolinic acid and kynurenic acid), the neurohormone melatonin, several neuroactive kynuramine metabolites of melatonin, and the trace amine tryptamine. The integral role of central serotonergic systems in the modulation of physiology and behaviour has been well documented since the first description of serotonergic neurons in the brain some 40 years ago. However, while the significance of the peripheral kynurenine pathway of tryptophan metabolism has also been recognised for several decades, it has only recently been appreciated that the synthesis of kynurenines within the central nervous system has important consequences for physiology and behaviour. Altered kynurenine metabolism has been implicated in the pathophysiology of conditions such as acquired immunodeficiency syndrome (AIDS)-related dementia, Huntington's disease and Alzheimer's disease. In this review we discuss the molecular mechanisms involved in regulating the metabolism of tryptophan and consider the medical implications associated with dysregulation of both serotonergic and kynurenine pathways of tryptophan metabolism.

Dendritic cell (DC) tryptophan catabolism has emerged in recent years as a major mechanism of peripheral tolerance. However, there are features of this mechanism, initiated by IDO, that are still unclear, including the role of enzymes that are downstream of IDO in the kynurenine pathway and the role of the associated production of kynurenines. In this study, we provide evidence that 1) murine DCs express all enzymes necessary for synthesis of the downstream product of tryptophan breakdown, quinolinate; 2) IFN-gamma enhances transcriptional expression of all of these enzymes, although posttranslational inactivation of IDO may prevent metabolic steps that are subsequent and consequent to IDO; 3) overcoming the IDO-dependent blockade by provision of a downstream quinolinate precursor activates the pathway and leads to the onset of suppressive properties; and 4) tolerogenic DCs can confer suppressive ability on otherwise immunogenic DCs across a Transwell in an IDO-dependent fashion. Altogether, these data indicate that kynurenine pathway enzymes downstream of IDO can initiate tolerogenesis by DCs independently of tryptophan deprivation. The paracrine production of kynurenines might be one mechanism used by IDO-competent cells to convert DCs lacking functional IDO to a tolerogenic phenotype within an IFN-gamma-rich environment.

Indoleamine 2,3-dioxygenase (IDO) catabolizes tryptophan to kynurenine. In the immune system, the reduction in tryptophan and increase in kynurenine act to suppress T-cell function. In the nervous system, kynurenine can be further metabolized to quinolinic acid, which can be neurotoxic. IDO is known to be expressed by microglia and its levels are upregulated by interferon-gamma (IFNgamma). We report here that IDO immunoreactivity is also localized in neurons, and that IDO is upregulated by IFNgamma in neurons of the hippocampus. Thus, neuronal IDO could contribute to the vulnerability of neurons to inflammatory conditions.

In mammalian cells, the essential amino acid tryptophan is degraded primarily by the kynurenine pathway, a cascade of enzymatic steps containing several biologically active compounds. Metabolites of this pathway, collectively termed 'kynurenines', have been shown to be involved in many diverse physiological and pathological processes. In particular, fluctuations in the levels of kynurenines have discrete effects on the nervous and immune systems. A considerable number of pharmacological tools have recently become available to probe the kynurenine pathway experimentally. Some of these 'kynurenergic' agents can be envisioned to be of therapeutic value, especially in the treatment of diseases that are associated with impaired kynurenine pathway metabolism.

Objective: Several somatic illnesses are associated with psychiatric comorbidity. Evidence is provided that availability of the essential amino acid tryptophan, which is the precursor of serotonin, may cause this phenomenon., Methods: We performed a database search to find relevant articles published between 1966 and 2002. For our search strategy, we combined several diseases from the categories hormonal, gastrointestinal, and inflammatory with the search terms "tryptophan" and "serotonin.", Results: The catabolism of tryptophan is stimulated under the influence of stress, hormones and inflammation by the induction of the enzymes tryptophan pyrrolase (in the liver) and IDO (ubiquitous). Because of the reduction in blood levels of tryptophan under these circumstances the formation of cerebral serotonin is decreased., Conclusions: It is argued that the coupling of peripheral tryptophan levels and cerebral serotonin levels has physiological significance. The clinical implications and therapeutic consequences of changes in tryptophan and consequently serotonin metabolism are discussed.

Cytotoxic T lymphocyte-associated antigen 4 (CTLA-4) plays a critical role in peripheral tolerance. However, regulatory pathways initiated by the interactions of CTLA-4 with B7 counterligands expressed on antigen-presenting cells are not completely understood. We show here that long-term survival of pancreatic islet allografts induced by the soluble fusion protein CTLA-4-immunoglobulin (CTLA-4-Ig) is contingent upon effective tryptophan catabolism in the host. In vitro, we show that CTLA-4-Ig regulates cytokine-dependent tryptophan catabolism in B7-expressing dendritic cells. These data suggest that modulation of tryptophan catabolism is a means by which CTLA-4 functions in vivo and that CTLA-4 acts as a ligand for B7 receptor molecules that transduce intracellular signals.

Tryptophan (TRP) metabolism via the kynurenine pathway leads to formations of neuroactive substances like kynurenine (KYN) and 3-hydroxykynurenine (3-HK), which may be involved in the pathogenesis of several human brain diseases. 3-Hydroxykynurenine especially is known to have strong neurotoxic properties. The generation of reactive oxygen species (ROS) leads to neuronal cell death with apoptotic features. Because the chronic renal insufficiency (CRI) results in disturbances in the functioning of the central nervous system (CNS), it is conceivable that the metabolism of some kynurenines may be altered and could play an important role in uremic encephalopathy. The levels of TRP, KYN and 3-HK were measured in the plasma and in different brain regions of uremic rats. The total plasma concentration of TRP as well as in all the studied brain samples was significantly diminished during uremia. Surprisingly, the level of KYN and 3-HK were elevated both in the plasma and different brain regions of CRI animals. KYN concentrations were approximately two times higher in the cerebellum, midbrain and cortex compared to the control group. The changes of 3-HK levels were more pronounced in the striatum and medulla than in other structures. This data suggests that CRI results in deep disturbances on the kynurenine pathway in CNS, which could be responsible for neurological abnormalities seen in uremia.