Marked IDO2 expression and activity related to autophagy and apoptosis in brain tissue of fatal tuberculous meningitis.

In about 1% of tuberculosis (TB) patients, Mycobacterium tuberculosis (M. tuberculosis) can disseminate to the meninges, causing tuberculous meningitis (TBM) with mortality rate up to 60%. Chronic granulomatous inflammation (non-necrotizing and necrotizing) in the brain is the histological hallmark of TBM. The tryptophan-catabolizing enzyme indoleamine 2,3-dioxygenase 1 (IDO1) and the generated kynurenine metabolites exert major effector functions relevant to TB granuloma functioning. Here we have assessed immunohistochemically IDO1 expression and activity and its effector function and that of its isoform, IDO2, in post-mortem brain tissue of patients that demised with neurotuberculosis. We also related these findings to brain tissue of fatal/severe COVID-19. In this study, IDO1 and IDO2 were abundantly expressed and active in tuberculoid granulomas and were associated with the presence of M. tuberculosis as well as markers of autophagy and apoptosis. Like in fatal/severe COVID-19, IDO2 was also prominent in specific brain regions, such as the inferior olivary nucleus of medulla oblongata and cerebellum, but not associated with granulomas or with M. tuberculosis. Spatially associated apoptosis was observed in TBM, whereas in fatal COVID-19 autophagy dominated. Together, our findings highlight IDO2 as a potentially relevant effector enzyme in TBM, which may relate to the symptomology of TBM.

Mass Balance of the Indoleamine 2,3-Dioxygenase Inhibitor Navoximod (GDC-0919) in Rats and Dogs: Unexpected Cyanide Release from Imidazo[5,1-a]isoindole and Species Differences in Glucuronidation.

Navoximod (GDC-0919) is a small molecule inhibitor of indoleamine 2,3-dioxygenase 1 (IDO1) developed to reduce T cell immunosuppression associated with cancer. This study describes the absorption, metabolism, and excretion (AME) of navoximod in rats and dogs after a single oral dose of [14C]-navoximod. An unexpected thiocyanate metabolite M1 and a chiral inversion metabolite M51 were captured as the major circulating metabolites in rats, accounting for 30% and 18% of 0-24 hours exposure, respectively. These two metabolites combined had much lower systemic exposure in dogs and humans (<6% and <1%). The novel cyanide release is proposed to occur via 4,5-epoxidation on the fused imidazole ring, leading to ring opening and rearrangement along with the release of cyanide. The decyanated metabolites were identified and confirmed by synthetic standards, which supported the proposed mechanism. In dogs, glucuronidation to M19 was the major clearance mechanism, representing 59% of the dose in the bile of bile duct-cannulated (BDC) dogs and 19% of the dose in the urine of intact dogs. Additionally, M19 also represented 52% of drug related exposure in circulation in dogs. In comparison, in humans, navoximod was mainly cleared through glucuronidation to M28 and excreted in urine (60% of the dose). The differences in the metabolism and elimination observed in vivo were qualitatively recapitulated in vitro with liver microsomes, suspended hepatocytes, and cocultured primary hepatocytes. The striking species differences in regioselective glucuronidation is likely explained by the species differences in UGT1A9, which was mainly responsible for M28 formation in humans. SIGNIFICANCE STATEMENT: The results from this study demonstrated significant species differences in metabolism (especially glucuronidation) and elimination of navoximod among rats, dogs, and humans. The study also illustrated the mechanism of a novel cyanide release metabolism from the fused imidazo[5,1-a]isoindole ring. Such biotransformation should be kept in mind when working with imidazole-containing new chemical entities in drug discovery and development.

Label-free tumor cell screening based on IDO1-mediated tryptophan metabolism at single cell level.

Indoleamine 2,3-dioxygenase 1 (IDO1) plays a critical role in inflammatory and immunometabolism programming through catalyzing the oxidation of tryptophan (Trp) into downstream N-formylkynurenine. IDO1 is typically up-regulated in malignant tumors, making it a potential biomarker for cancer diagnosis. Here we show an effective strategy for tumor cell detection by integrating IDO1 activity assay with single cell-encapsulated droplets on a microfluidic platform for high-throughput bioanalysis. Mixed cells, as well as other cofactors, are encapsulated in individual droplets, which act as dynamic microreactors for IDO1-catalyzed oxidation of Trp. After pico-injection of a biosensing ensemble consisting of the macrocycle cucurbit [8]uril (Q8) and a fluorescent guest, rapid and robust screening of tumor cells by fluorescence signal is achieved in a few minutes reporting to Trp depletion, expanding the scope of conventional antibody-based detection of protein biomarkers. The results represent the first example of quantifying IDO1 enzymatic activity at the single cell level with a high-throughput performance, therefore promising warning signs and early diagnosis of tumor cells.

Assessment of interferon gamma and indoleamine 2,3-dioxygenase 1 analysis during respiratory syncytial virus infection in infants in Italy: an observational case-control study.

OBJECTIVES: The aim of this study was to measure interferon gamma (IFN-γ) and indoleamine 2,3-dioxygenase 1 (IDO1) values in the White blood cells of infants during respiratory tract infections and to compare these with healthy age-matched controls. DESIGN: This was a prospective, observational case-control study conducted in 2019-2020. SETTING: The study took place at Regina Margherita Children's Hospital, Turin, Italy. PARTICIPANTS: The study comprised 63 infants, including 26 patients hospitalised for bronchiolitis due to a respiratory syncytial virus (RSV) infection and 37 age-matched controls. The inclusion criteria included a positive RSV test for an infant with bronchiolitis. METHODS: We collected peripheral blood and measured the relative quantification of messenger RNA (mRNA) expression of IFN-γ and IDO1 with TaqMan real-time PCR amplification. The data were collected on the first day of admission. RESULTS: The mean age of the 26 patients with RSV bronchiolitis (53.8% female) was 85 (9-346) days when they were admitted to the hospital. Their mean gestational age at birth was 38 weeks and their mean birth weight was 3100 (2780-3730) g. The expression of IFN-γ was significantly reduced in patients with bronchiolitis RSV compared with healthy controls (p=0.0132). However, there was no significant difference between the two groups when the IDO1 mRNA expression values in their WCC were measured (p=0.0642). CONCLUSION: Our findings did not clarify whether IDO1 expression was related to the early stage of the disease or to the young age of the infants. The data provide evidence that IFN-γ was significantly reduced in infants with bronchiolitis due to RSV, compared with age-matched healthy controls, but the IDO1 was not different. New investigations that focus on subjects infected with RSV at different stages of infancy would help to clarify whether IDO1 expression can be related to age.

Indoleamine 2,3-dioxygenase (IDO)-1 and IDO-2 activity and severe course of COVID-19.

COVID-19 is a pandemic with high morbidity and mortality. In an autopsy cohort of COVID-19 patients, we found extensive accumulation of the tryptophan degradation products 3-hydroxy-anthranilic acid and quinolinic acid in the lungs, heart, and brain. This was not related to the expression of the tryptophan-catabolizing indoleamine 2,3-dioxygenase (IDO)-1, but rather to that of its isoform IDO-2, which otherwise is expressed rarely. Bioavailability of tryptophan is an absolute requirement for proper cell functioning and synthesis of hormones, whereas its degradation products can cause cell death. Markers of apoptosis and severe cellular stress were associated with IDO-2 expression in large areas of lung and heart tissue, whereas affected areas in brain were more restricted. Analyses of tissue, cerebrospinal fluid, and sequential plasma samples indicate early initiation of the kynurenine/aryl-hydrocarbon receptor/IDO-2 axis as a positive feedback loop, potentially leading to severe COVID-19 pathology. © 2021 The Authors. The Journal of Pathology published by John Wiley & Sons, Ltd on behalf of The Pathological Society of Great Britain and Ireland.

Characterization of the temporal, cell-specific and interferon-inducible patterns of indoleamine 2,3 dioxygenase 1 (IDO1) expression in the human placenta across gestation.

INTRODUCTION: The human placenta performs multiple functions necessary for successful pregnancy, but the metabolic pathways and molecular mechanisms responsible for regulating placental development and functions remain incompletely understood. Catabolism of the essential amino acid tryptophan has numerous critical roles in normal physiology, including inflammation. The kynurenine pathway, which accounts for ∼90% of tryptophan breakdown, is mediated by indoleamine 2,3 dioxygenase 1 (IDO1) in the placenta. In pregnant mice, alterations of IDO1 activity or expression result in fetal resorption and a preeclampsia-like phenotype. Decreased IDO1 expression at the maternal-fetal interface has also been linked to preeclampsia, in utero growth restriction and recurrent miscarriage in humans. These collective observations suggest essential role(s) for IDO1 in maintaining healthy pregnancy. Despite these important roles, the precise temporal, cell-specific and inflammatory cytokine-mediated patterns of IDO1 expression in the human placenta have not been thoroughly characterized across gestation. METHODS: Western blot and whole mount immunofluorescence (WMIF) were utilized to characterize and quantify basal and interferon (IFN)-inducible IDO1 expression in 1st trimester (7-13 weeks), 2nd trimester (14-22 weeks) and term (39-41 weeks) placental villi. RESULTS: IDO1 expression is activated in the human placenta between the 13th and 14th weeks of pregnancy, increases through the 2nd trimester and remains elevated at term. Constitutive IDO1 expression is restricted to placental endothelial cells. Interestingly, different types of IFNs have distinct effects on IDO1 expression in the human placenta. DISCUSSION: Our collective results are consistent with potential role(s) for IDO1 in the regulation of vascular functions in placental villi.

Early reduction in PD-L1 expression predicts faster treatment response in human cutaneous leishmaniasis.

Cutaneous leishmaniasis (CL) is caused by Leishmania donovani in Sri Lanka. Pentavalent antimonials (e.g., sodium stibogluconate [SSG]) remain first-line drugs for CL with no new effective treatments emerging. We studied whole blood and lesion transcriptomes from Sri Lankan patients with CL at presentation and during SSG treatment. From lesions but not whole blood, we identified differential expression of immune-related genes, including immune checkpoint molecules, after onset of treatment. Using spatial profiling and RNA-FISH, we confirmed reduced expression of programmed death-ligand 1 (PD-L1) and indoleamine 2,3-dioxygenase 1 (IDO1) proteins on treatment in lesions of a second validation cohort and further demonstrated significantly higher expression of these checkpoint molecules on parasite-infected compared with noninfected lesional CD68+ monocytes and macrophages. Crucially, early reduction in PD-L1 but not IDO1 expression was predictive of rate of clinical cure (HR = 4.88) and occurred in parallel with reduction in parasite load. Our data support a model whereby the initial anti-leishmanial activity of antimonial drugs alleviates checkpoint inhibition on T cells, facilitating immune-drug synergism and clinical cure. Our findings demonstrate that PD-L1 expression can be used as a predictor of rapidity of clinical response to SSG treatment in Sri Lanka and support further evaluation of PD-L1 as a host-directed therapeutic in leishmaniasis.

Prognostic significance of T cells, PD-L1 immune checkpoint and tumour associated macrophages in clear cell carcinoma of the ovary.

OBJECTIVE: To define the pre-treatment tumour immune landscape of clear cell carcinoma of the ovary (CCOC). METHODS: We investigated the infiltration profiles of selected immune cell populations and immune checkpoint proteins that have been previously shown to have prognostic relevance in high grade serous carcinoma of the ovary to determine their association with clinical outcomes in CCOC patients. Using multiplex immunohistochemistry, we evaluated the density of CD3+, FoxP3+, CD8+ T cells, CD20+ B cells and expression of PD-1, PD-L1 and IDO1 immune checkpoints in a cohort of 162 CCOC tumour specimens on a tissue microarray. RESULTS: Increased infiltration of CD3+ CD8- (helper T) cells, CD8+ (cytotoxic T) cells, and CD68+ macrophages significantly associated with shorter disease-free survival, recurrence-free survival and overall survival. Importantly, higher expression of PD-L1 and IDO-1 immune checkpoints was associated with better clinical outcomes. CONCLUSION: Findings from our study are foundational towards the development of immune classifiers and biomarkers of response to immune checkpoint blockade therapy in CCOC.

Expression of B7-H4 and IDO1 is associated with drug resistance and poor prognosis in high-grade serous ovarian carcinomas.

High-grade serous ovarian carcinoma (HGSC) is the most lethal gynecologic malignancy. While immune checkpoint inhibitors against PD-L1 and CTLA-4 have shown significant effects in multiple tumor types, the response rate to single-agent immune checkpoint inhibitors is low in HGSC. Alternative biomarkers and targets must be identified to guide patient selection and new therapeutic strategies in HGSC. Here, we aim to investigate the clinical significance of novel immune modulators, including B7-H4, IDO1, Tim3, IL6, and IL-8, in patients with HGSC. A total of 48 patients with HGSCs, comprising 24 cases that were sensitive and 24 that were resistant to standard paclitaxel and carboplatin chemotherapy, were selected for our initial analysis. A NanoString assay including 33 immune-related genes was used to compare the expression of different immune regulatory molecules in the sensitive and resistant groups. Differentially expressed proteins were verified using multiplex immunohistochemical staining on tissue arrays of 202 patients with HGSCs who underwent primary surgery at MDACC. We analyzed the expression levels of immune checkpoints and compared expression profiles with clinicopathologic features including response, progression-free survival, and overall survival. HGSC tumors resistant to therapy expressed higher levels of B7-H4 (69.3%), IDO1 (71.8%), Tim3 (89.1%), and inflammatory factors IL-6 and IL-8, and expressed higher Tim3 in stromal components. High expression of B7-H4 and IDO1 was associated with significantly lower overall survival and progression-free survival. B7-H4 and IDO1 were co-expressed in 49.1% of studied cases. A panel of immunomodulatory proteins including B7-H4, IDO1, Tim3, IL-6, and IL-8 are expressed at high levels in HGSCs. These modulators represent novel targets to enhance immunotherapy in patients with HGSCs.

MenSCs exert a supportive role in establishing a pregnancy-friendly microenvironment by inhibiting TH17 polarization.

OBJECTIVES: Uncontrolled TH17 differentiation has been suggested to play a role in the pathogenesis of pregnancy loss. We recently showed that menstrual blood stromal/stem cells (MenSCs) alter functional features of natural killer cells. Here, we hypothesized that MenSCs could modulate differentiation of TH17 cells. METHOD: MenSCs were collected from 18 apparently healthy women and characterized. Bone marrow mesenchymal stem cells (BMSCs) served as a control. TH17 polarization and proliferation of purified T CD4+ cells were assessed by flow cytometry in a well-defined co-culture system containing T CD4+ cells and MenSCs or BMSCs. Indoleamine 2,3-Dioxygenase (IDO) activity was evaluated in MenSC and BMSC culture supernatants by a colorimetric assay. The impact of MenSCs on expression of transcription factors, RORC, T-bet, Gata3, NRP-1 and Helios were studied by qPCR. RESULTS: MenSCs significantly inhibited TH17 differentiation (p = 0.0383) and percentage of the cells co-expressing IL-17 and IFN-γ (p = 0.0023). PGE2 blockade significantly reduced percentage and proliferation of T CD4+IL-17+ (p = 0.003, p = 0.0018), T CD4+ IFN-γ+ (p = 0.002, p = 0.0022) and T CD4+IL-17+ IFN-γ+ (p = 0.004, p = 0.02) cells. MenSCs produced a considerable activity of IDO (p = 0.0002), induced a significant rise in the Treg frequency (p = 0.0091) and a sharp increase in TH17/Tregs ratio (p = 0.0022). MenSCs increased expression of NRP1 (p = 0.001), while downregulated expression of RORC in T cells (p = 0.001). CONCLUSION: Our results suggest a supportive role for MenSCs in establishing a pregnancy-friendly microenvironment in the uterus and put forth the idea that inherent abnormalities of MenSCs may be a basis for dysregulated endometrial immune network leading to pregnancy loss.

Reliable chromatographic assay for measuring of indoleamine 2,3-dioxygenase 1 (IDO1) activity in human cancer cells.

The kynurenine pathway is the major tryptophan degradation routes generating bioactive compounds important in physiology and diseases. Depending on cell type it is initiated enzymatically by tryptophan-2,3-dioxygenase (TDO) or indoleamine-2,3-dioxygenase 1 and 2 (IDO1 and IDO2) to yield N-formylkynurenine as the precursor of further metabolites. Herein, we describe an accurate high-pressure liquid chromatography coupled with a diode array detector (HPLC-DAD) method to serve for IDO1 activity determination in human cancer cells cultured in vitro. Enzymatic activity was expressed as the rate of ʟ-kynurenine generation by 1 mg of proteins obtained from cancer cells. Our approach shows the limit of detection and limit of quantification at 12.9 and 43.0 nM Kyn, respectively. Applicability of this method was demonstrated in different cells (ovarian and breast cancer)exposed to various conditions and has successfully passed the validation process. This approach presents a useful model to study the role of kynurenine pathway in cancer biology.

IDO1 Expression in Melanoma Metastases Is Low and Associated With Improved Overall Survival.

Indoleamine 2-3 dioxygenase 1 (IDO1) expression may contribute to immunologic escape by melanoma metastases. However, a recent clinical trial failed to identify any clinical benefits of IDO1 inhibition in patients with unresectable metastatic melanoma, and prior characterizations of IDO1 expression have predominately studied primary lesions and local metastases, generating uncertainty regarding IDO1 expression in distant metastases. We hypothesized that IDO1 expression in such lesions would be low and correlated with decreased overall survival (OS). Metastases from patients (n=96) with stage IIIb to IV melanoma underwent tissue microarray construction and immunohistochemical staining for IDO1. Th1-related gene expression was determined quantitatively. Associations between OS and IDO1 expression were assessed with multivariate models. Of 96 metastatic lesions, 28% were IDOpos, and 85% exhibited IDO1 expression in <10% of tumor cells. IDOpos lesions were associated with improved OS (28.9 vs. 10.5 mo, P=0.02) and expression of Th1-related genes. OS was not associated with IDO1 expression in a multivariate analysis of all patients; however, IDO1 expression (hazard ratio=0.25, P=0.01) and intratumoral CD8+ T-cell density (hazard ratio=0.99, P<0.01) were correlated with OS in patients who underwent metastasectomy with curative-intent. IDOpos metastases were less likely to recur after metastasectomy (54% vs. 16%, P=0.01). IDO1 expression was low in melanoma metastases and correlated with OS after metastasectomy with curative-intent. Intratumoral CD8+ T cells and Th1-related genes were correlated with IDO1 expression, as was tumor recurrence. These suggest that IDO1 expression may be a marker of immunologic tumor control, and may inform participant selection in future trials of IDO1 inhibitors.

PD­L1 and IDO­1 expression in undifferentiated pleomorphic sarcoma: The associations with tumor infiltrating lymphocytes, dMMR and HLA class I.

The prognosis of undifferentiated pleomorphic sarcoma (UPS) is generally unfavorable. Recently, clinical trials such as SARC028 demonstrated the utility of cancer immunotherapy for soft tissue sarcomas. The aim of the present study was to assess the expression of PD­L1 and IDO­1 as prognostic factors and therapeutic targets. A total of 52 primary UPS cases were retrieved and two UPS cell lines were utilized for supplementary analysis. Immunohistochemical staining of anti­PD­L1 (28­8), IDO­1, CD8, CD4, CD3, HLA class I, MSH2, MSH6, MLH1 and PMS2 was carried out. Immunohistochemically, 19 of 52 (36.5%) cases showed PD­L1 expression at least focally (≥1%) and 5 of 52 (9.62%) showed strong PD­L1 expression (≥50%). Overall, 25 of 52 (48.1%) cases expressed IDO­1 (≥1%). Two tumors were evaluated as having deficient mismatch repair and six tumors as having the loss of HLA class I. PD­L1 expression (≥1%) was significantly related to the infiltration of CD8­ and CD3­positive lymphocytes, but strong PD­L1 expression (≥50%) did not present a significant relationship with tumor­infiltrating lymphocytes. IDO­1 expression was also associated with CD8­, CD4­, and CD3­positive lymphocytes. In vitro, both PD­L1 and IDO­1 were induced by IFN­Î³ stimulation. In survival analysis, strong PD­L1 expression (≥50%) was a significant poor prognostic factor, while IDO­1 expression (≥1%) was a favorable one. In conclusion, UPS was shown to frequently express PD­L1 and IDO­1. It was suggested that PD­L1 expression (≥50%) and IDO­1 expression are poor and favorable prognostic factors of UPS patients, respectively.

Effect of IFN-γ on the kynurenine/tryptophan ratio in monolayer-cultured keratinocytes and a 3D reconstructed human epidermis model.

BACKGROUND: Interferon-gamma (IFN-γ) represents a potent inducer for keratinocyte inflammatory and immune activation in vitro. Since tryptophan (trp) conversion to kynurenine (kyn) is involved in inflammation, the topical kyn/trp ratio may serve as a biomarker of skin inflammation. However, the trp metabolism in keratinocytes exposed to IFN-γ is not yet fully understood. OBJECTIVE: The aim of this study was to establish a human epidermis model in order to quantify cytokine and kyn/trp secretion from IFN-γ stimulated cells and tissues. Moreover, to compare the cell response of 2D-cultured keratinocytes and the 3D epidermis model. METHODS: Polycarbonate filters were used on which primary keratinocytes could attach and stratify in order to form the typical layers of reconstructed human epidermis (RHE). After IFN-γ treatment, secretion of kyn/trp was measured by high performance liquid chromatography. Gene and protein expression of indoleamine 2,3-dioxygenase 1 (IDO) was analyzed with real-time PCR and immunohistochemistry. The secretion of cytokines was quantified with ELISA. RESULTS: Trp catabolism to kyn was significantly increased (P < 0.01) in the 2D culture in response to IFN-γ treatment. Before kyn secretion, IDO was strongly upregulated (P < 0.001). IFN-γ treatment also increased the secretion of IL-6 and IL-8 from the keratinocytes. In the RHE, IDO was upregulated by IFN-γ, and kyn secretion could be detected. Interestingly, IDO expression was only present in the basal cells of the RHE. CONCLUSION: Our results suggest that IFN-γ acts as an inducer of trp degradation preferentially in undifferentiated keratinocytes, indicated by the IDO expression in the basal layer of the RHE.

Indoleamine 2,3-Dioxygenase-1 Expression in Adrenocortical Carcinoma.

BACKGROUND: Indoleamine 2,3-dioxygenase 1 (IDO-1) is overexpressed in many human carcinomas and a successful target for therapy in mouse models. Prognosis of patients with advanced adrenocortical carcinoma (ACC) is poor due to the lack of effective treatments, and new therapies are therefore needed. Herein, we investigate whether IDO-1 is expressed in human ACC tissues. METHODS: 53 tissue samples from patients with ACC, adrenal adenoma (AA), adrenocortical tumors (ACTs), and normal adrenal were identified. Immunohistochemistry was performed on formalin-fixed, paraffin-embedded slides for IDO-1. Samples were scored for cytoplasmic staining as per intensity and the percent of positive cells and for stromal staining by percent of positive cells. Tumor characteristics, PD-L1, PDL-2, and CD-8+ T-lymphocyte expression were also determined. RESULTS: Samples from 32 ACC, 3 ACT, 15 AA, and 3 normal adrenal were analyzed. IDO-1 was expressed in tumor tissue in 22 of 32 ACC samples, compared with 8 of 15 AA sample (P = 0.344). IDO-1 expression was significantly increased in stromal tissue of ACC samples (16 of 33), compared with AA samples (0 of 15) (P = 0.001). IDO-1 expression in ACC and AA samples was associated with PD-L2 expression (P = 0.034). IDO-1 expression in ACC stromal tissue was associated with CD8+ T-lymphocyte infiltration (P = 0.028). CONCLUSIONS: IDO-1 is expressed in a majority of ACC samples. Its expression in tumor tissue is associated with PD-L2 expression, and expression in stroma is associated with CD8+ cell infiltration. IDO-1 inhibition, alone or in combination with PD-1 inhibition, could therefore be an interesting target in treatment of ACC.

IDO Targeting in Sarcoma: Biological and Clinical Implications.

Sarcomas are heterogeneous malignant mesenchymal neoplasms with limited sensitivity to immunotherapy. We recently demonstrated an increase in Kynurenine Pathway (KP) activity in the plasma of sarcoma patients treated with pembrolizumab. While the KP has already been described to favor immune escape through the degradation of L-Tryptophan and production of metabolites including L-Kynurenine, Indoleamine 2,3 dioxygenase (IDO1), a first rate-limiting enzyme of the KP, still represents an attractive therapeutic target, and its blockade had not yet been investigated in sarcomas. Using immunohistochemistry, IDO1 and CD8, expression profiles were addressed within 203 cases of human sarcomas. At a preclinical level, we investigated the modulation of the KP upon PDL1 blockade in a syngeneic model of sarcoma through mRNA quantification of key KP enzymes within the tumor. Furthermore, in order to evaluate the possible anti-tumor effect of IDO blockade in combination with PDL1 blockade, an innovative IDO inhibitor (GDC-0919) was used. Its effect was first assessed on Kynurenine to Tryptophan ratio at plasmatic level and also within the tumor. Following GDC-0919 treatment, alone or in combination with anti-PDL1 antibody, tumor growth, immune cell infiltration, and gene expression profiling were measured. IDO1 expression was observed in 39.1% of human sarcoma cases and was significantly higher in tumors with high CD8 infiltration. In the pre-clinical setting, blockade of PDL1 led to a strong anti-tumor effect and was associated with an intratumoral inflammatory cytokines signature driven by Ifng but also with a modulation of the KP enzymes including Ido1 and Ido2. IDO1 inhibition using GDC-0919 resulted in (i) a significant decrease of plasmatic Kynurenine to Tryptophan ratio and in (ii) a decrease of tumoral Kynurenine. However, GDC-0919 used alone or combined with anti-PDL1, did not show anti-tumoral activity and did not affect the tumor immune cell infiltrate. In order to elucidate the mechanism(s) underlying the lack of effect of GDC-0919, we analyzed the gene expression profile of intratumoral biopsies. Interestingly, we have found that GDC-0919 induced a downregulation of the expression of pvr and granzymes, and an upregulation of inhba and Dtx4 suggesting a potential role of the IDO pathway in the control of NK function.

Indoleamine 2,3-dioxygenase and trophoblast invasion in caesarean scar pregnancy: Implications for the aetiopathogenesis of placenta accreta spectrum.

Immunohistochemical localisation of indoleamine 2,3-dioxygenase was studied in order to better understand the pathophysiology of placenta accreta spectrum. In the decidua staining for indoleamine 2,3-dioxygenase was found in the glandular epithelium with some additional positive cells. Extravillous cytotrophoblast invasion was present in the myometrium which was not covered by the decidual tissue whereas myometrial invasion of cytotrophoblasts was absent where this tissue lay deep to decidua. These results suggest that indoleamine 2,3-dioxygenase expression in the decidua may normally control trophoblast invasion and absence of its expression where decidua is absent may be involved in the pathogenesis of the over-invaded placenta.

Presence of the protein indoleamine 2,3-dioxygenase (IDO) in the maternal-fetal interface of the yolk sac placenta of blue shark, Prionace glauca.

Indoleamine 2 3-dioxygenase (IDO) is a protein usually described in mammals, which, among other functions, participates in the maternal-fetal tolerance process. The blue-shark, Prionace glauca (Linnaeus, 1758) is a viviparous placentary species in which the yolk sac develops during the pregnancy, turning into a placenta for matrotrophic nutrition of the embryo. The purpose of this study was to investigate the expression of IDO in the P. glauca maternal-fetal interface along three gestation phases and describe its distribution and the meaning of its presence. The results showed IDO labelling during the yolk sac/placenta development in the ectoderm on the three development phases and in the endoderm at the two first phases. In the uterine epithelium, IDO was observed in the last two phases. These interface tissues are major contact areas between the mother and the semiallogeneic conceptus and this relation could induce an immunological response against the fetus. Therefore, the presence of IDO may indicate that it could have a similar role in the mechanism of maternal-fetal tolerance in the P. glauca placental interface, as described in eutherian mammals.

The local immune phenotype influences prognosis in patients with nodal-positive rectal cancer after neoadjuvant chemoradiation.

INDROCUTION: The local immune contexture in patients with locally advanced rectal cancer (LARC) has important prognostic value after neoadjuvant chemoradiation and surgical resection. In this study, we examined the prognostic role of Indoleamine-2,3-Dioxygenase (IDO1) and infiltrating cytotoxic T lymphocytes (CD8+) according to the nodal stage of LARC patients. MASTERIAL AND METHODS: Expression of IDO1 and CD8 was evaluated through immunohistochemistry in 106 archival tumour tissue samples from patients following neoadjuvant chemoradiation and radical resection. The average infiltration of IDO1+ and CD8+ cells was calculated and expressed as total scores as previously described. Kaplan-Meier curves were used to describe overall and disease-free survival. RESULTS: In nodal-positive tumours (N+), IDO-positivity was associated with a reduced disease-free survival (DFS) (p = 0.063) and CD8-positivity with an impaired OS (p = 0.024). Patients with a N+ LARC and a high total IDO1 score showed a clear advantage regarding five-year disease-free survival rates compared with patients with a low total IDO1 score (N+ 5y-DFS IDO1 high: 66.7% vs IDO low: 19%). We also detected better 5-years-OS rates in N+ LARC with a high total CD8 score (N+ 5y-OS CD8 high: 83.3% vs CD8 low: 32.3%). These survival benefits were not evident in patients with N-tumours. CONCLUSION: Analysis of the local CD8 and IDO1 expression influences prognosis in nodal-positive LARC patients after multimodal therapy and may be a helpful tool in specifying individual adjuvant treatment strategies according to different immune profiles.

Displacement Induced Off-On Fluorescent Biosensor Targeting IDO1 Activity in Live Cells.

We show how the macrocyclic host cucurbit[8]uril (CB[8]) and a fluorescent dye form a biosensing ensemble while its cavity simultaneously traps tryptophan, the upstream substrate of IDO1 enzymes, therefore providing a label-free method to monitor the activity of IDO1 in real time. Incubation of malignant HeLa and HepG2 cells overexpressing IDO1 with the associative biosensor resulted in its spontaneous uptake and a fluorescence switch-on response in situ, which can be traced to the displacement of tryptophan from CB[8] upon IDO1-catalyzed oxidation. The results, for the first time, establish a supramolecular sensing concept for the detection of intracellular enzymatic activity in live cells, thus allowing direct cell-based analysis and inhibitor screening compatible with commercial instruments including microplate reader, fluorescent microscopy, and flow cytometry.