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"PROJECT":{"PROJECT_TITLE":"Trisomy 21 activates the kynurenine pathway via increased dosage of interferon receptors","PROJECT_SUMMARY":"Trisomy 21 (T21) causes Down syndrome (DS), affecting immune and neurological function by unknown mechanisms. We report here a large metabolomics study of plasma and cerebrospinal fluid showing that people with DS produce elevated levels of kynurenine and quinolinic acid, two tryptophan catabolites with potent immunosuppressive and neurotoxic properties, respectively. We demonstrate that immune cells of people with DS overexpress IDO1, the rate-limiting enzyme in the kynurenine pathway (KP) and a known interferon (IFN)-stimulated gene. Furthermore, we show positive correlations among levels of IFN-inducible cytokines and KP dysregulation. Using metabolic tracing assays, we determine that IFN stimulation causes IDO1 overexpression and kynurenine overproduction in cells with T21, dependent on overexpression of IFN receptors encoded on chromosome 21. Finally, we show a mouse model of DS carrying triplication of the IFN receptors exhibits KP dysregulation. Altogether, these results reveal a mechanism by which T21 could drive immunosuppression and neurotoxicity in DS.","INSTITUTE":"University of Colorado Denver","LAST_NAME":"Culp-Hill","FIRST_NAME":"Rachel","ADDRESS":"12801 E 17th Ave L18-9403D, Aurora, Colorado, 80045, USA","EMAIL":"rachel.hill@cuanschutz.edu","PHONE":"303-724-5798"},

"STUDY":{"STUDY_TITLE":"Global Metabolic Analysis Trisomy 21 - Cohort 3, Plasma","STUDY_SUMMARY":"Global metabolic analysis of plasma from individuals with and without trisomy 21.","INSTITUTE":"University of Colorado Denver","LAST_NAME":"Culp-Hill","FIRST_NAME":"Rachel","ADDRESS":"12801 E 17th Ave L18-9403D, Aurora, Colorado, 80045, USA","EMAIL":"rachel.hill@cuanschutz.edu","PHONE":"303-724-5798"},

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],
"COLLECTION":{"COLLECTION_SUMMARY":"All human subjects in this study were consented according to Colorado Multiple Institutional Review Board (COMIRB)-approved protocols. Written informed consent was obtained from parents or guardians of participants under the age of 18, and assent was obtained from participants over the age of 7 who were cognitively able to assent. Deidentified plasma samples for Cohort 1 were obtained from the Translational Nexus Clinical Data Registry and Biobank (University of Colorado Anschutz Medical Campus, COMIRB 08-1276). Additional plasma and WBC samples were obtained through the Crnic’s Institute Human Trisome Project (University of Colorado Anschutz Medical Campus, COMIRB 15-2170, www.trisome.org). Plasma was collected in Vacutainer tubes (EDTA–purple capped or Lithium heparin–light green capped) and stored at -80°C. Participant medical history was collected by the respective biobanks.","SAMPLE_TYPE":"Blood (plasma)"},

"TREATMENT":{"TREATMENT_SUMMARY":"N/A"},

"SAMPLEPREP":{"SAMPLEPREP_SUMMARY":"For plasma analyses, a volume of 20μL of was extracted in 480μL of ice-cold methanol:acetonitrile:water (5:3:2). For CSF analyses, a volume of 20μL was extracted in 180μL and for cell-based experiments, 2 million cells were extracted in 1mL of the same ice-cold lysis solution. Subsequently, these solutions were vortexed for 30 minutes at 4°C. Insoluble proteins were pelleted by centrifugation (10 minutes at 4°C and 12,000 g) and supernatants were collected and stored at -80°C until analysis. For quantitative analysis of kynurenine pathway (KP) metabolites, supernatants were spun in a Speedvac until dry and resuspended in 0.1% formic acid in water as previously described (PMID: 30213797, 30143553)."},

"CHROMATOGRAPHY":{"CHROMATOGRAPHY_SUMMARY":"UHPLC-MS metabolomics analyses were performed using a Vanquish UHPLC system coupled online to a Q Exactive mass spectrometer (Thermo Fisher, Bremen, Germany). 20uL of supernatant was injected for plasma and CSF samples, and 10uL of supernatant was injected for cell samples. Samples were resolved over a Kinetex C18 column (2.1x150 mm, 1.7μm; Phenomenex, Torrance, CA, USA) at 45°C using a 5-minute gradient at 450μL/minute from 5-95% B (A: water/0.1% formic acid; B: acetonitrile/0.1% formic acid) for positive mode. To monitor possible technical variability, aliquots of each of the individual samples were combined to make technical replicates, which were run after every 15 samples. Additionally, in each experiment, several lysis solution aliquots were run as blanks for artifact identification.","CHROMATOGRAPHY_TYPE":"Reversed phase","INSTRUMENT_NAME":"Thermo Vanquish","COLUMN_NAME":"Phenomenex Kinetex C18 (150 x 2.1mm, 2.6 um)","FLOW_GRADIENT":"5-95% B","FLOW_RATE":"450uL/min","COLUMN_TEMPERATURE":"45","METHODS_FILENAME":"5MMpos_method.pdf","SOLVENT_A":"water, 0.1% formic acid","SOLVENT_B":"acetonitrile, 0.1% formic acid","ANALYTICAL_TIME":"5min"},

"ANALYSIS":{"ANALYSIS_TYPE":"MS","LABORATORY_NAME":"D'Alessandro Lab","OPERATOR_NAME":"Rachel Culp-Hill"},

"MS":{"INSTRUMENT_NAME":"Thermo Q Exactive Orbitrap","INSTRUMENT_TYPE":"Orbitrap","MS_TYPE":"ESI","ION_MODE":"POSITIVE","MS_COMMENTS":"The Q Exactive mass spectrometer (Thermo Fisher Scientific, San Jose, CA, USA) was operated in positive ion mode using electrospray ionization, scanning in Full MS mode (1μscan) from 65 to 975 m/z at 70,000 resolution, with 4 kV spray voltage, 45 sheath gas, 15 auxiliary gas. MS analysis and data elaboration were performed as described74,75. Calibration was performed prior to analysis using the PierceTM Positive Ion Calibration Solution (Thermo Fisher Scientific)."},

"MS_METABOLITE_DATA":{
"Units":"relative abundance",

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m/z":"153.0391","medRt":"0.585"},{"Metabolite":"Allantoate","KEGG ID":"C00499","HMDB ID":"","PubChem ID":"","quantitated m/z":"177.0607","medRt":"0.611"},{"Metabolite":"5-Hydroxyisourate","KEGG ID":"C11821","HMDB ID":"","PubChem ID":"","quantitated m/z":"185.0318","medRt":"0.680"},{"Metabolite":"2',3'-Cyclic CMP","KEGG ID":"C02354","HMDB ID":"","PubChem ID":"","quantitated m/z":"306.0494","medRt":"0.687"},{"Metabolite":"5-6-Dihydrothymine","KEGG ID":"C00906","HMDB ID":"","PubChem ID":"","quantitated m/z":"129.0659","medRt":"0.678"},{"Metabolite":"Pyridoxamine 5'-phosphate","KEGG ID":"C00647","HMDB ID":"","PubChem ID":"","quantitated m/z":"249.0607","medRt":"0.683"},{"Metabolite":"5-Oxoproline","KEGG ID":"C01879","HMDB ID":"","PubChem ID":"","quantitated m/z":"130.05","medRt":"0.690"},{"Metabolite":"Cys-Gly","KEGG ID":"C01419","HMDB ID":"","PubChem ID":"","quantitated m/z":"179.0484","medRt":"0.678"},{"Metabolite":"L-Homocysteine","KEGG ID":"C00155","HMDB ID":"","PubChem ID":"","quantitated m/z":"136.0428","medRt":"0.685"},{"Metabolite":"Dimethylglycine","KEGG ID":"C01026","HMDB ID":"","PubChem ID":"","quantitated m/z":"104.0711","medRt":"0.691"},{"Metabolite":"Phosphoserine","KEGG ID":"C02532","HMDB ID":"","PubChem ID":"","quantitated m/z":"186.0183","medRt":"0.680"},{"Metabolite":"Ornithine","KEGG ID":"C01602","HMDB ID":"","PubChem ID":"","quantitated m/z":"133.0971","medRt":"0.608"},{"Metabolite":"L-Citrulline","KEGG ID":"C00327","HMDB ID":"","PubChem ID":"","quantitated m/z":"176.103","medRt":"0.689"},{"Metabolite":"Cadaverine","KEGG ID":"C01672","HMDB ID":"","PubChem ID":"","quantitated m/z":"103.1233","medRt":"0.601"},{"Metabolite":"Spermidine","KEGG ID":"C00315","HMDB ID":"","PubChem ID":"","quantitated m/z":"146.1649","medRt":"0.612"},{"Metabolite":"Spermine","KEGG ID":"C00750","HMDB ID":"","PubChem ID":"","quantitated m/z":"203.2227","medRt":"0.615"},{"Metabolite":"Carnosine","KEGG ID":"C00386","HMDB ID":"","PubChem ID":"","quantitated m/z":"227.1152","medRt":"0.650"},{"Metabolite":"Creatine","KEGG ID":"C00300","HMDB ID":"","PubChem ID":"","quantitated m/z":"132.0768","medRt":"0.677"},{"Metabolite":"Creatinine","KEGG ID":"C00791","HMDB ID":"","PubChem ID":"","quantitated m/z":"114.0664","medRt":"0.685"},{"Metabolite":"Pantothenol","KEGG ID":"C00864","HMDB ID":"","PubChem ID":"","quantitated m/z":"220.1174","medRt":"1.668"},{"Metabolite":"Taurine","KEGG ID":"C00245","HMDB ID":"","PubChem ID":"","quantitated m/z":"126.0221","medRt":"0.722"},{"Metabolite":"Indole","KEGG ID":"C00463","HMDB ID":"","PubChem ID":"","quantitated m/z":"118.0654","medRt":"1.727"},{"Metabolite":"Indole-3-acetate","KEGG ID":"C00954","HMDB ID":"","PubChem ID":"","quantitated m/z":"176.0704","medRt":"1.897"},{"Metabolite":"Indoxyl","KEGG ID":"C05658","HMDB ID":"","PubChem ID":"","quantitated m/z":"134.0601","medRt":"1.850"},{"Metabolite":"g-Oxalo-crotonate","KEGG ID":"C03453","HMDB ID":"","PubChem ID":"","quantitated m/z":"159.0275","medRt":"0.684"},{"Metabolite":"L-Adrenaline","KEGG ID":"C00788","HMDB ID":"","PubChem ID":"","quantitated m/z":"184.0941","medRt":"0.667"},{"Metabolite":"N-Methylethanolamine phosphate","KEGG ID":"C01210","HMDB ID":"","PubChem ID":"","quantitated m/z":"156.0419","medRt":"0.668"},{"Metabolite":"Acetylcholine","KEGG ID":"C01996","HMDB ID":"","PubChem ID":"","quantitated m/z":"146.1173","medRt":"0.678"},{"Metabolite":"Choline","KEGG ID":"C00114","HMDB ID":"","PubChem ID":"","quantitated m/z":"104.1074","medRt":"0.666"},{"Metabolite":"Sphinganine 1-phosphate","KEGG ID":"C01120","HMDB ID":"","PubChem ID":"","quantitated m/z":"382.2707","medRt":"3.813"},{"Metabolite":"L-Carnitine","KEGG ID":"C00318","HMDB ID":"","PubChem ID":"","quantitated m/z":"162.1122","medRt":"0.672"},{"Metabolite":"Acetyl-carnitine","KEGG ID":"C02571","HMDB ID":"","PubChem ID":"","quantitated m/z":"204.123","medRt":"0.689"},{"Metabolite":"Propionyl-carnitine","KEGG ID":"C03017","HMDB ID":"","PubChem ID":"","quantitated m/z":"218.1386","medRt":"0.689"},{"Metabolite":"Butanoyl-l-carnitine","KEGG ID":"C02862","HMDB ID":"","PubChem ID":"","quantitated m/z":"232.1541","medRt":"0.690"},{"Metabolite":"Hydroxybutyrylcarnitine","KEGG ID":"","HMDB ID":"HMDB13127","PubChem ID":"","quantitated m/z":"248.1479","medRt":"1.750"},{"Metabolite":"Tiglylcarnitine","KEGG ID":"","HMDB ID":"HMDB02366","PubChem ID":"","quantitated m/z":"244.1541","medRt":"1.741"},{"Metabolite":"Hexanoyl-L-carnitine","KEGG ID":"","HMDB ID":"HMDB00756","PubChem ID":"","quantitated m/z":"260.1852","medRt":"1.800"},{"Metabolite":"L-octanoylcarnitine","KEGG ID":"C02838","HMDB ID":"","PubChem ID":"","quantitated m/z":"288.2162","medRt":"1.881"},{"Metabolite":"octenoyl-l-carnitine","KEGG ID":"","HMDB ID":"","PubChem ID":"70679121","quantitated m/z":"286.2007","medRt":"1.839"},{"Metabolite":"O-Decenoyl-L-carnitine","KEGG ID":"","HMDB ID":"","PubChem ID":"57357170","quantitated m/z":"314.2318","medRt":"1.937"},{"Metabolite":"O-dodecanoyl-carnitine","KEGG ID":"","HMDB ID":"HMDB02250","PubChem ID":"","quantitated m/z":"344.2786","medRt":"2.216"},{"Metabolite":"O-dodecenoyl-carnitine","KEGG ID":"","HMDB ID":"HMDB13326","PubChem ID":"","quantitated m/z":"342.2629","medRt":"2.062"},{"Metabolite":"O-tetradecanoyl-L-carnitine","KEGG ID":"","HMDB ID":"HMDB05066","PubChem ID":"","quantitated m/z":"372.31","medRt":"2.725"},{"Metabolite":"Bilirubin","KEGG ID":"C00486","HMDB ID":"","PubChem ID":"","quantitated m/z":"585.2703","medRt":"1.945"},{"Metabolite":"Methylenediurea","KEGG ID":"C06381","HMDB ID":"","PubChem ID":"","quantitated m/z":"133.0737","medRt":"0.677"},{"Metabolite":"N6-Methyl-L-lysine","KEGG ID":"C02728","HMDB ID":"","PubChem ID":"","quantitated m/z":"161.1283","medRt":"0.626"},{"Metabolite":"Ferric gluconate","KEGG ID":"C13511","HMDB ID":"","PubChem ID":"","quantitated m/z":"196.0604","medRt":"1.735"},{"Metabolite":"N-Amidino-L-aspartate","KEGG ID":"C03139","HMDB ID":"","PubChem ID":"","quantitated m/z":"176.0659","medRt":"0.674"},{"Metabolite":"N-Acyl-D-glutamate","KEGG ID":"C06379","HMDB ID":"","PubChem ID":"","quantitated m/z":"175.0476","medRt":"0.680"},{"Metabolite":"L-Kynurenine","KEGG ID":"C00328","HMDB ID":"","PubChem ID":"","quantitated m/z":"209.0917","medRt":"1.430"},{"Metabolite":"L-Formylkynurenine","KEGG ID":"C02700","HMDB ID":"","PubChem ID":"","quantitated m/z":"237.0862","medRt":"1.484"},{"Metabolite":"6-Hydroxykynurenic acid","KEGG ID":"C08480","HMDB ID":"","PubChem ID":"","quantitated m/z":"206.0473","medRt":"0.930"},{"Metabolite":"Quinolinic acid","KEGG ID":"C03722","HMDB ID":"","PubChem ID":"","quantitated m/z":"168.0289","medRt":"0.932"},{"Metabolite":"Anthranilate","KEGG ID":"C00108","HMDB ID":"","PubChem ID":"","quantitated m/z":"138.0547","medRt":"0.749"},{"Metabolite":"Picolinic acid","KEGG ID":"C10164","HMDB ID":"","PubChem ID":"","quantitated m/z":"124.0397","medRt":"0.601"}]
}

}