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Inhibition of indoleamine 2,3-dioxygenase activity by levo-1-methyl tryptophan blocks gamma interferon-induced Chlamydia trachomatis persistence in human epithelial cells.

ABSTRACT: Gamma interferon (IFN-?) induces expression of the tryptophan-catabolizing enzyme indoleamine 2,3-dioxygenase (IDO1) in human epithelial cells, the permissive cells for the obligate intracellular bacterium Chlamydia trachomatis. IDO1 depletes tryptophan by catabolizing it to kynurenine with consequences for C. trachomatis, which is a tryptophan auxotroph. In vitro studies reveal that tryptophan depletion can result in the formation of persistent (viable but noncultivable) chlamydial forms. Here, we tested the effects of the IDO1 inhibitor, levo-1-methyl-tryptophan (L-1MT), on IFN-?-induced C. trachomatis persistence. We found that addition of 0.2 mM L-1MT to IFN-?-exposed infected HeLa cell cultures restricted IDO1 activity at the mid-stage (20 h postinfection [hpi]) of the chlamydial developmental cycle. This delayed tryptophan depletion until the late stage (38 hpi) of the cycle. Parallel morphological and gene expression studies indicated a consequence of the delay was a block in the induction of C. trachomatis persistence by IFN-?. Furthermore, L-1MT addition allowed C. trachomatis to undergo secondary differentiation, albeit with limited productive multiplication of the bacterium. IFN-?-induced persistent infections in epithelial cells have been previously reported to be more resistant to doxycycline than normal productive infections in vitro. Pertinent to this observation, we found that L-1MT significantly improved the efficacy of doxycycline in clearing persistent C. trachomatis forms. It has been postulated that persistent forms of C. trachomatis may contribute to chronic chlamydial disease. Our findings suggest that IDO1 inhibitors such as L-1MT might provide a novel means to investigate, and potentially target, persistent chlamydial forms, particularly in conjunction with conventional therapeutics.

SUBMITTER: Ibana JA

PROVIDER: S-EPMC3257928 | biostudies-literature | 2011 Nov

REPOSITORIES: biostudies-literature

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Inhibition of indoleamine 2,3-dioxygenase activity by levo-1-methyl tryptophan blocks gamma interferon-induced Chlamydia trachomatis persistence in human epithelial cells.

Ibana Joyce A JA Belland Robert J RJ Zea Arnold H AH Schust Danny J DJ Nagamatsu Takeshi T AbdelRahman Yasser M YM Tate David J DJ Beatty Wandy L WL Aiyar Ashok A AA Quayle Alison J AJ

Infection and immunity 20110912 11

Gamma interferon (IFN-γ) induces expression of the tryptophan-catabolizing enzyme indoleamine 2,3-dioxygenase (IDO1) in human epithelial cells, the permissive cells for the obligate intracellular bacterium Chlamydia trachomatis. IDO1 depletes tryptophan by catabolizing it to kynurenine with consequences for C. trachomatis, which is a tryptophan auxotroph. In vitro studies reveal that tryptophan depletion can result in the formation of persistent (viable but noncultivable) chlamydial forms. Here, ...[more]

PMID: 21911470

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Project description:AimsNeutrophil granulocytes are the major cells involved in Chlamydia trachomatis (C. trachomatis)-mediated inflammation and histopathology. A key protein in human intracellular antichlamydial defense is the tryptophan-degrading enzyme indoleamine 2,3-dioxygenase (IDO) which limits the growth of the tryptophan auxotroph Chlamydia. Despite its importance, the role of IDO in the intracellular defense against Chlamydia in neutrophils is not well characterized.MethodsGlobal gene expression screen was used to evaluate the effect of C. trachomatis serovar D infection on the transcriptome of human neutrophil granulocytes. Tryptophan metabolite concentrations in the Chlamydia-infected and/or interferon-gamma (IFNG)-treated neutrophils were measured by ultra-high-performance liquid chromatography-tandem mass spectrometry (UHPLC-MS/MS).ResultsOur results indicate that the C. trachomatis infection had a major impact on neutrophil gene expression, inducing 1,295 genes and repressing 1,510 genes. A bioinformatics analysis revealed that important factors involved in the induction of neutrophil gene expression were the interferon-related transcription factors such as IRF1-5, IRF7-9, STAT2, ICSB, and ISGF3. One of the upregulated genes was ido1, a known infection- and interferon-induced host gene. The tryptophan-degrading activity of IDO1 was not induced significantly by Chlamydia infection alone, but the addition of IFNG greatly increased its activity. Despite the significant IDO activity in IFNG-treated cells, C. trachomatis growth was not affected by IFNG. This result was in contrast to what we observed in HeLa human cervical epithelial cells, where the IFNG-mediated inhibition of C. trachomatis growth was significant and the IFNG-induced IDO activity correlated with growth inhibition.ConclusionsIDO activity was not able to inhibit chlamydial growth in human neutrophils. Whether the IDO activity was not high enough for inhibition or other chlamydial growth-promoting host mechanisms were induced in the infected and interferon-treated neutrophils needs to be further investigated.

Project description:Kynurenine (Kyn), a metabolite of tryptophan (Trp), is a key regulator of mammal immune responses such as cancer immune tolerance. Indoleamine-2,3-dioxygenase (IDO) and tryptophan-2,3-dioxygenase (TDO) are main enzymes regulating the first and rate-limiting step of the Kyn pathway. To identify new small molecule inhibitors of TDO, we selected A172 glioblastoma cell line constitutively expressed TDO. Characterization of this cell line using kinase inhibitor library resulted in identification of MEK/ERK pathway-dependent TDO expression. After knowing the properties for TDO expression, we further proceeded to screen chemical library for TDO inhibitors. We previously determined that S-benzylisothiourea derivatives are enzymatic inhibitors of indoleamine 2,3-dioxygenase 1 (IDO1) and suggested that the isothiourea moiety could be an important pharmacophore for binding to heme. Based on this premise, we screened an in-house library composed of various isothiourea derivatives and identified a bisisothiourea derivative, PVZB3001, as an inhibitor of TDO. Interestingly, PVZB3001 also inhibited the enzymatic activity of IDO1 in both cell-based and cell-free assays but did not inhibit other heme enzymes. Molecular docking studies suggested the importance of isothiourea moieties at the ortho position of the phenyl ring for the inhibition of catalytic activity. PVZB3001 showed competitive inhibition against TDO, and this was supported by the docking simulation. PVZB3001 recovered natural killer (NK) cell viability and functions by inhibiting Kyn accumulation in conditioned medium of both IDO1- and TDO-expressing cells. Furthermore, oral administration of IDO1-overexpressing tumor-bearing mice with PVZB3001 significantly inhibited tumor growth. Thus, we identified a novel selective dual inhibitor of IDO1 and TDO using the Kyn production assay with a glioblastoma cell line. This inhibitor could be a useful pharmacological tool for modulating the Kyn pathway in a variety of experimental systems.

Project description:ObjectivesExtrahepatic tryptophan (Trp)-kynurenine (Kyn) metabolism via indoleamine 2,3-dioxygenase 1 (IDO1) induction was found to be associated with intrinsic immune regulation. However, the Trp-Kyn metabolism-associated immune regulation in dermatomyositis (DM) remains unknown. Therefore, we aimed to investigate the clinical relevance of the Trp-Kyn metabolism via IDO1 induction in DM.MethodsLiquid chromatography-mass spectrometry (HPLC-MS) was used to examine the serum Kyn and Trp concentrations in DM. In addition, we used X-tile software to determine the optimal cutoff value of the Kyn/Trp ratio, a surrogate marker for Trp-Kyn metabolism. Spearman analysis was performed to evaluate the association of Trp-Kyn metabolism with muscle enzymes and inflammatory markers.ResultsDM patients had significantly higher serum Kyn/Trp ratio (× 10-3) when compared with the healthy controls. The serum Kyn/Trp ratio was positively correlated with the levels of muscle enzymes and inflammatory markers. In addition, the serum Kyn/Trp ratio significantly decreased (36.89 (26.00-54.00) vs. 25.00 (18.00-37.00), P = 0.0006) after treatment. DM patients with high serum Kyn/Trp ratio had a significantly higher percentage of muscle weakness symptoms (62.5% vs. 20.0%, P = 0.019) and higher levels of LDH (316.0 (236.0-467.0) vs. 198.0 (144.0-256.0), P = 0.004) and AST (56.5 (35.0-92.2) vs. 23.0 (20.0-36.0), P = 0.002)) than those with low serum Kyn/Trp ratio. Multiple Cox regression analyses identified ln(Kyn/Trp) (HR 4.874, 95% CI 1.105-21.499, P = 0.036) as an independent prognostic predictor of mortality in DM.ConclusionsDM patients with enhanced Trp-Kyn metabolism at disease onset are characterized by more severe disease status and poor prognosis. Intrinsic immune regulation function via enhanced Trp-Kyn metabolism by IDO1 induction may be a potential therapeutic target in DM. Key Points • HPLC-MS identified increased serum Kyn/Trp ratio in DM patients, which positively correlated with levels of muscle enzymes and inflammatory markers and was downregulated upon treatment. • Cox regression analyses identified ln(Kyn/Trp) as an independent prognostic predictor of mortality in DM. • Monitoring intrinsic immune regulation function should be considered a potential therapeutic target in DM patients.

Dataset Information

Inhibition of indoleamine 2,3-dioxygenase activity by levo-1-methyl tryptophan blocks gamma interferon-induced Chlamydia trachomatis persistence in human epithelial cells.

Publications

Inhibition of indoleamine 2,3-dioxygenase activity by levo-1-methyl tryptophan blocks gamma interferon-induced Chlamydia trachomatis persistence in human epithelial cells.

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