#METABOLOMICS WORKBENCH JasonMiska_20231016_174741 DATATRACK_ID:4403 STUDY_ID:ST002935 ANALYSIS_ID:AN004814 PROJECT_ID:PR001826 VERSION 1 CREATED_ON October 17, 2023, 1:19 pm #PROJECT PR:PROJECT_TITLE Myeloid cell-derived creatine in the hypoxic niche promotes glioblastoma growth PR:PROJECT_TYPE LC-MS/MS PR:PROJECT_SUMMARY Glioblastoma (GBM) is a malignancy dominated by the infiltration of PR:PROJECT_SUMMARY tumor-associated myeloid cells (TAMCs). Examination of TAMC metabolic phenotypes PR:PROJECT_SUMMARY in mouse models and human GBM patients identified the de-novo creatine metabolic PR:PROJECT_SUMMARY pathway as a hallmark of TAMCs. Multi-omics analyses revealed that TAMCs PR:PROJECT_SUMMARY surround the hypoxic peri-necrotic regions of GBM and express the creatine PR:PROJECT_SUMMARY metabolic enzyme glycine amidinotransferase (GATM). Conversely, GBM cells PR:PROJECT_SUMMARY located within these same regions are uniquely specific in expressing the PR:PROJECT_SUMMARY creatine transporter (SLC6A8). Therefore, we hypothesized that TAMCs provide PR:PROJECT_SUMMARY creatine to tumors, promoting GBM progression. Isotopic tracing demonstrated PR:PROJECT_SUMMARY that TAMC-secreted creatine can be taken up by tumor cells. Creatine PR:PROJECT_SUMMARY supplementation protected tumors from hypoxia-induced stress which was abrogated PR:PROJECT_SUMMARY with genetic ablation or pharmacologic inhibition of SLC6A8. Lastly, inhibition PR:PROJECT_SUMMARY of creatine transport using the clinically relevant compound, RGX-202-01, PR:PROJECT_SUMMARY blunted tumor growth, and enhanced radiation therapy in-vivo. This work PR:PROJECT_SUMMARY highlights that myeloid-to-tumor transfer of creatine promotes tumor growth in PR:PROJECT_SUMMARY the hypoxic niche. PR:INSTITUTE Northwestern University, Feinberg School of Medicine PR:DEPARTMENT Neurological Surgery PR:LABORATORY Jason Miska PR:LAST_NAME Miska PR:FIRST_NAME Jason PR:ADDRESS 676 N St. Clair PR:EMAIL jason.miska@northwestern.edu PR:PHONE 8478678201 #STUDY ST:STUDY_TITLE Title: Myeloid cell-derived creatine in the hypoxic niche promotes glioblastoma ST:STUDY_TITLE growth ST:STUDY_TYPE PBMC vs Tumor CD163+ ST:STUDY_SUMMARY Glioblastoma (GBM) is a malignancy dominated by the infiltration of ST:STUDY_SUMMARY tumor-associated myeloid cells (TAMCs). Examination of TAMC metabolic phenotypes ST:STUDY_SUMMARY in mouse models and human GBM patients identified the de-novo creatine metabolic ST:STUDY_SUMMARY pathway as a hallmark of TAMCs. Multi-omics analyses revealed that TAMCs ST:STUDY_SUMMARY surround the hypoxic peri-necrotic regions of GBM and express the creatine ST:STUDY_SUMMARY metabolic enzyme glycine amidinotransferase (GATM). Conversely, GBM cells ST:STUDY_SUMMARY located within these same regions are uniquely specific in expressing the ST:STUDY_SUMMARY creatine transporter (SLC6A8). Therefore, we hypothesized that TAMCs provide ST:STUDY_SUMMARY creatine to tumors, promoting GBM progression. Isotopic tracing demonstrated ST:STUDY_SUMMARY that TAMC-secreted creatine can be taken up by tumor cells. Creatine ST:STUDY_SUMMARY supplementation protected tumors from hypoxia-induced stress which was abrogated ST:STUDY_SUMMARY with genetic ablation or pharmacologic inhibition of SLC6A8. Lastly, inhibition ST:STUDY_SUMMARY of creatine transport using the clinically relevant compound, RGX-202-01, ST:STUDY_SUMMARY blunted tumor growth, and enhanced radiation therapy in-vivo. This work ST:STUDY_SUMMARY highlights that myeloid-to-tumor transfer of creatine promotes tumor growth in ST:STUDY_SUMMARY the hypoxic niche. ST:INSTITUTE Northwestern University, Feinberg School of Medicine ST:DEPARTMENT Neurological Surgery ST:LABORATORY Jason Miska ST:LAST_NAME Miska ST:FIRST_NAME Jason ST:ADDRESS 676 N St. Clair ST:EMAIL jason.miska@northwestern.edu ST:PHONE 8478678201 ST:NUM_GROUPS 2 ST:TOTAL_SUBJECTS 5 #SUBJECT SU:SUBJECT_TYPE Human SU:SUBJECT_SPECIES Homo sapiens SU:TAXONOMY_ID 9606 #FACTORS #SUBJECT_SAMPLE_FACTORS: SUBJECT(optional)[tab]SAMPLE[tab]FACTORS(NAME:VALUE pairs separated by |)[tab]Raw file names and additional sample data SUBJECT_SAMPLE_FACTORS - Les-Jas-20190812-13 Treatment:PBMC RAW_FILE_NAME=Les-Jas-20190812-13 SUBJECT_SAMPLE_FACTORS - Les-Jas-20190812-14 Treatment:PBMC RAW_FILE_NAME=Les-Jas-20190812-14 SUBJECT_SAMPLE_FACTORS - Les-Jas-20190812-15 Treatment:PBMC RAW_FILE_NAME=Les-Jas-20190812-15 SUBJECT_SAMPLE_FACTORS - Les-Jas-20190812-16 Treatment:Tumor RAW_FILE_NAME=Les-Jas-20190812-16 SUBJECT_SAMPLE_FACTORS - Les-Jas-20190812-17 Treatment:Tumor RAW_FILE_NAME=Les-Jas-20190812-17 SUBJECT_SAMPLE_FACTORS - Les-Jas-20190812-18 Treatment:Tumor RAW_FILE_NAME=Les-Jas-20190812-18 SUBJECT_SAMPLE_FACTORS - Les-Jas-20200214-01 Treatment:Tumor RAW_FILE_NAME=Les-Jas-20200214-01 SUBJECT_SAMPLE_FACTORS - Les-Jas-20200214-02 Treatment:Tumor RAW_FILE_NAME=Les-Jas-20200214-02 SUBJECT_SAMPLE_FACTORS - Les-Jas-20200214-03 Treatment:PBMC RAW_FILE_NAME=Les-Jas-20200214-03 SUBJECT_SAMPLE_FACTORS - Les-Jas-20200214-04 Treatment:PBMC RAW_FILE_NAME=Les-Jas-20200214-04 #COLLECTION CO:COLLECTION_SUMMARY Magnetic bead isolation of cells To isolate specific populations of cells, CO:COLLECTION_SUMMARY single-cell suspensions as isolated above are preblocked with anti-CD16/32 for CO:COLLECTION_SUMMARY 15 min at 4°C. We then used the biotinylated anti-CD163 (all from Thermo Fisher CO:COLLECTION_SUMMARY Scientific) to label murine myeloid cells. Next, the cells were washed and then CO:COLLECTION_SUMMARY incubated with anti-biotin magnetic beads (Miltenyi Biotec) before performing CO:COLLECTION_SUMMARY manual positive selection using MS columns (Miltenyi Biotec). Purified cells CO:COLLECTION_SUMMARY were analyzed for all downstream metabolic analyses. CO:SAMPLE_TYPE Brain #TREATMENT TR:TREATMENT_SUMMARY Fresh cells isolated from PBMC or TIL of GBM patients #SAMPLEPREP SP:SAMPLEPREP_SUMMARY Isolated TAMC samples were dried using a SpeedVac. Acetonitrile (50%) was added SP:SAMPLEPREP_SUMMARY to the tube for reconstitution following overtaxing for 30 s. Sample solution SP:SAMPLEPREP_SUMMARY was then centrifuged for 15 min at 20,000g and 4°C. Supernatant was collected SP:SAMPLEPREP_SUMMARY for LC-MS analysis. #CHROMATOGRAPHY CH:CHROMATOGRAPHY_SUMMARY Samples were analyzed by high-performance LC (HPLC) and high-resolution MS and CH:CHROMATOGRAPHY_SUMMARY MS/MS (HPLC-MS/MS). The system consists of Thermo Q Exactive with an CH:CHROMATOGRAPHY_SUMMARY electrospray source and an UltiMate3000 (Thermo Fisher Scientific) series HPLC CH:CHROMATOGRAPHY_SUMMARY consisting of a binary pump, degasser, and autosampler outfitted with an XBridge CH:CHROMATOGRAPHY_SUMMARY Amide column (Waters; dimensions of 4.6 mm by 100 mm and a 3.5-μm particle CH:CHROMATOGRAPHY_SUMMARY size). The mobile phase A contained 95% water/5% acetonitrile (v/v), 20 mM CH:CHROMATOGRAPHY_SUMMARY ammonium hydroxide, and 20 mM ammonium acetate (pH 9.0); phase B was 100% CH:CHROMATOGRAPHY_SUMMARY acetonitrile. The gradient was performed as follows: 0 min, 15% A; 2.5 min, 30% CH:CHROMATOGRAPHY_SUMMARY A; 7 min, 43% A; 16 min, 62% A; 16.1 to 18 min, 75% A; and 18 to 25 min, 15% A CH:CHROMATOGRAPHY_SUMMARY with a flow rate of 400 µl/min. CH:CHROMATOGRAPHY_TYPE HILIC CH:INSTRUMENT_NAME Thermo Dionex Ultimate 3000 CH:COLUMN_NAME Water's Xbridge amide (100 x 3mm, 3.5 um) CH:SOLVENT_A 95% water/5% acetonitrile; 20 mM ammonium hydroxide; 20 mM ammonium acetate (pH CH:SOLVENT_A 9.0) CH:SOLVENT_B 100% acetonitrile CH:FLOW_GRADIENT 0 min, 15% A; 2.5 min, 30% A; 7 min, 43% A; 16 min, 62% A; 16.1 to 18 min, 75% CH:FLOW_GRADIENT A; and 18 to 25 min, 15% A CH:FLOW_RATE 400 μl/min CH:COLUMN_TEMPERATURE - #ANALYSIS AN:ANALYSIS_TYPE MS #MS MS:INSTRUMENT_NAME Thermo Q Exactive Plus Orbitrap MS:INSTRUMENT_TYPE Orbitrap MS:MS_TYPE ESI MS:ION_MODE UNSPECIFIED MS:MS_COMMENTS The capillary of the electrospray ionization source was set to 275°C, with MS:MS_COMMENTS sheath gas at 45 arbitrary units, auxiliary gas at 5 arbitrary units, and the MS:MS_COMMENTS spray voltage at 4.0 kV. In positive/negative polarity switching mode, a MS:MS_COMMENTS mass/charge ratio (m/z) scan range from 70 to 850 was chosen and MS1 data were MS:MS_COMMENTS collected at a resolution of 70,000. The automatic gain control target was set MS:MS_COMMENTS at 1 × 106, and the maximum injection time was 200 ms. The top five precursor MS:MS_COMMENTS ions were subsequently fragmented, in a data-dependent manner, using the MS:MS_COMMENTS higher-energy collisional dissociation cell set to 30% normalized collision MS:MS_COMMENTS energy in MS2 at a resolution power of 17,500. #MS_METABOLITE_DATA MS_METABOLITE_DATA:UNITS Peak Area MS_METABOLITE_DATA_START Samples Les-Jas-20190812-13 Les-Jas-20190812-14 Les-Jas-20190812-15 Les-Jas-20190812-16 Les-Jas-20190812-17 Les-Jas-20190812-18 Les-Jas-20200214-01 Les-Jas-20200214-02 Les-Jas-20200214-03 Les-Jas-20200214-04 Factors Treatment:PBMC Treatment:PBMC Treatment:PBMC Treatment:Tumor Treatment:Tumor Treatment:Tumor Treatment:Tumor Treatment:Tumor Treatment:PBMC Treatment:PBMC 1,3-bisphospho-D-glycerate 0 0 0 0 0 0 0 0 0 0 2-(Formamido)-N1-(5-phospho-D-ribosyl)acetamidine 0 0 0 0 0 0 0 0 0 0 2,3-bisphospho-D-glycerate 0 0 0 0 0 0 2190732.466 143113.0952 177631.9904 299048.3901 2/3-phosphoglycerate 46141.81004 53535.47435 54367.90792 49863.91701 520676.4536 0 11958211.65 2749000.17 3458662.387 3830196.175 2-HG 3371511.187 2002299.932 1322888.801 1435284.933 1056283.271 1694115.509 0 0 0 0 3,4-dihydroxymandelic acid 0 0 0 25016.94174 13712.69084 0 0 0 0 0 3,4-dihydroxyphenylpropanoate 0 0 0 0 0 0 0 0 0 0 3-hydroxypropanoic acid 0 0 0 0 0 0 60471344.28 29337300.69 24865805.21 71060908.42 3-methoxytyramine 0 0 0 0 0 23311.53874 34650.32982 35229.04505 27325.00029 20799.93014 3-phosphohydroxypyruvic acid 0 0 0 0 0 0 0 0 0 0 3-phospho-serine 0 0 0 0 0 0 52236.68162 0 25896.77836 17126.78827 3-ureidopropionic acid 16572.0674 11194.80957 0 0 0 0 15828.98507 34393.84684 10701.85039 13704.73516 4-phosphopantothenate 0 0 0 0 0 0 0 0 0 0 5,6-dihydroxyindole 0 0 0 0 0 0 0 17277.08803 0 14825.60444 5-methylcytosine 0 0 0 0 0 0 429750.4434 112065.4204 294842.6677 115919.0531 5-methyl-THF 0 0 0 0 0 0 0 0 0 0 5'-methylthioadenosine 0 38228.52188 0 0 0 0 0 1474186.72 904925.5263 1204594.402 6-aminocaproic acid 0 0 0 0 0 0 0 0 0 0 6-O-methylguanine 0 0 0 0 0 0 0 0 0 0 6-phosphogluconic acid 67173.05724 41381.73941 0 0 11961.94372 25886.32792 813720.0403 352696.007 177796.4493 110835.5411 6-phosphoglucono-D-lactone 0 0 0 0 0 0 0 0 0 0 acetoacetic acid 0 0 0 0 0 0 8598447.916 2963801.574 4270764.595 0 acetylcholine+ 525985.0506 2113260.722 512786.0744 531829.207 386068.1145 625718.435 5693489.716 410694.0319 16196826.88 3585035.656 aconitic acid 3447653.407 1603166.578 877352.5173 3457070.708 561718.8519 494046.6304 0 0 0 0 adenine 0 0 0 0 0 0 8511813.783 6783474.927 6601525.806 7192278.206 adenosine 0 0 0 806293.9725 338467.7621 0 3437707.891 1570796.634 1478714.808 1482643.213 adenylosuccinate 0 0 0 11705.87289 0 0 30228.8688 0 29535.17612 0 ADP 3862587.203 5131373.412 2905236.305 364848.6602 301023.3037 68382.91301 6996129.737 1256175.238 15636641.39 11008209.16 ADP-D-glucose 0 0 0 0 0 0 0 0 35988.18322 0 agmatine 0 0 8131.424967 0 0 0 40816.70392 123452.8898 76892.08042 22033.54762 AICA ribonucleotide 0 0 0 0 0 0 0 0 0 0 a-KG 1845817.49 2650537.152 2766964.172 2823133.072 2856794.129 1891019.252 97259.3872 741239.0028 390904.7979 166299.6301 alanine 0 0 0 0 0 0 12164162.8 5385016.809 34094149.34 9308821.164 allantoin 17820.17135 0 0 39094.88426 0 0 0 0 0 22384.46909 AMP 5741841.786 3281550.239 1756119.239 1725428.895 1571008.48 1161914.242 24423266.54 1123445.753 7260586.159 3620669.417 anandamide 0 0 0 0 0 0 0 0 0 0 arginine 7136162.07 6618648.374 10366292.45 11454738.42 6546520.121 10654233.29 28823375.38 2072247.193 10200906.21 7179568.377 asparagine 193270.1378 68467.55539 39334.90379 126613.1687 192946.3304 63179.43907 66681.4239 0 0 19229.10291 aspartic acid 1976164.218 1392226.147 718469.6162 2697245.004 93109.11124 363520.9973 10466254.05 2284224.841 6871295.495 4288521.582 ATP 1874745.528 2414806.957 1258319.752 17718.28102 8063.014312 0 2908680.225 975972.285 13886975.28 16489651.48 b-alanine 476776.5003 3846816.852 820069.4433 1173414.683 584774.7131 234669.0361 2195976.643 11468205.91 2258854.881 2000822.94 betaine 60555683.88 160132038.4 93543307.08 139106518.4 364118274.6 56091782.98 190860717 71845464.28 185936076.6 301801042 carbamoyl phosphate 0 13414.22863 0 0 0 29893.4712 31365.22927 153820.2493 44052.03115 111654.2985 carnitine+ 37235908 103478628.2 204487429.9 70115398.26 392093508 34917619.49 9678197.999 7995763.378 10498294.01 5149117.552 CDP 18333.84505 115352.3051 10144.51245 15190.18327 18406.57646 0 1457321.721 0 1592520.764 449739.566 CDP-choline 101445.7595 24083.49058 0 12449.32858 266859.5523 0 424680.3734 47504.80585 640287.3256 322917.0854 choline+ 25274188.67 58874558.31 41956961.73 68705905.69 56879377.25 47164913.68 101856893.8 49329212.45 136169974.5 33408612.02 citraconic acid 2500153.057 3225987.884 1275406.131 1087868.121 1195851.813 1879896.578 1931761.278 5352675.858 3890085.674 15128573.32 citramalic acid 5525230.323 3555049.019 1509034.658 2520915.934 961722.7007 4634427.939 3244839.603 2985410.634 913325.6444 1544659.402 citrate / citrate(iso) 210457977.6 233507948.1 178993508.7 113647954.9 72653035.13 95887444.09 357595451.7 220560767.4 210483859.3 186391672.1 citrulline 0 0 0 0 0 0 0 0 0 0 CMP 0 37659.17501 17579.94661 61664.14712 68539.0942 0 1014027.316 0 629977.5805 48802.93724 creatine 8790486.846 42027558.87 14157499.55 35866892.53 111129867.2 16136372.91 38877579.78 19408147.56 17558717.51 15977688.42 Creatinine 0 0 0 0 0 0 18326851.72 0 0 41087651.42 CTP 0 0 0 0 0 0 326378.4086 23191.19225 1225246.377 857139.8045 cystathionine 0 0 0 206162.8343 200418.767 0 2062425.316 713056.8945 9374.490579 9079.512197 cysteine 116417.9527 40983.71659 22362.9061 0 10927.85091 0 4784956.15 3212007.966 2024133.483 1432991.982 cystine 0 0 0 0 0 0 64225.49152 0 9461.044509 18653.9066 cytidine 0 0 0 0 0 0 0 0 0 0 cytosine 3783245.5 1190367.492 2346044.445 0 795033.6364 3765994.42 0 0 0 0 dADP 0 0 0 0 0 0 0 0 0 0 dAMP 0 0 0 0 0 0 0 0 0 0 D-arabinono-1,4-lactone 1493476.4 423658.5526 215167.2625 377385.7996 724258.6472 365133.428 1506901.818 301082.6452 1592240.448 992185.5591 dATP 0 0 0 0 0 0 0 0 0 0 dCDP 0 0 0 0 0 0 0 0 0 0 dCMP 0 0 0 0 0 0 0 0 0 0 dCTP 0 0 0 0 0 0 0 0 0 0 dehydroascorbic acid 182399.4801 188223.8749 45524.60832 0 132883.3019 0 18653846.75 2835139.237 12821135.13 10776658.07 deoxyguanosine 0 0 0 0 0 0 676512.9057 713929.1923 911826.9705 258961.9026 deoxyinosine 0 0 0 0 0 0 0 0 0 0 deoxyuridine 0 0 0 0 0 0 0 0 0 0 D-erythrose-4-phosphate 0 0 0 0 0 0 0 0 0 0 dGDP 796262.4437 606785.3496 38407.54924 0 0 29967.61636 0 0 1151363.166 1265217.119 dihydroorotate 22460.39098 0 15799.55933 0 0 0 0 0 0 0 DL-DOPA 0 0 0 0 0 0 0 0 0 0 DOPAC 22843.87178 0 0 0 0 0 0 28118.23788 0 18527.10495 DOPAL 0 0 0 0 0 0 69238.49036 364879.305 111901.6376 210434.3965 dopamine / octopamine 391404.2335 360998.8525 383970.3783 515909.4451 314232.8092 808590.9374 320693.4752 623087.8884 141255.6497 143836.8299 dopamine quinone 365670.2158 167421.1975 393314.993 349279.3555 91131.80801 245953.7807 696159.4171 855051.8093 810199.3118 973726.1167 DOPEG 0 0 0 0 0 0 0 0 0 0 DOPET 0 0 0 0 0 0 31565.90452 0 0 0 D-ribose-1/5-phosphate / D-ribulose 5-phosphate 623861.0327 24278.64932 0 725355.1511 13565.30716 0 1858553.864 317622.7198 375305.5548 79513.62561 D-sedoheptulose-7-phosphate 19087.87809 13645.29969 0 0 14615.47159 0 165267.5674 0 55759.54509 833541.7248 dTMP 0 0 0 0 0 0 0 0 0 0 dTTP 0 0 0 0 0 0 0 0 0 0 dUMP 0 0 0 0 0 0 0 0 0 0 dUTP 0 0 0 0 0 0 0 0 0 0 epinephrine 15597.5774 10236.82466 9426.063113 0 0 0 0 12103.42374 8579.016959 22238.32487 F-1,6/2,6-DP 2571180.859 1352245.864 821691.9969 497257.3688 191649.7884 146140.2757 23845516.7 10595637.03 6272058.267 5016796.517 F1P 0 0 0 0 0 0 0 0 0 0 F6P / G1P 195947.0164 54766.6984 0 0 402217.487 0 2507080.753 726631.0379 629255.7257 1983052.912 FAD 0 0 0 0 0 0 0 0 0 0 FMN 0 0 0 0 0 0 0 0 0 0 fumaric acid 2124997.102 2463825.76 2579432.287 3815766.647 2132681.988 3022127.544 0 524779.426 141485.8941 0 G6P 459248.6257 119823.7548 69517.70217 44470.73865 12436.02569 0 1900874.322 899813.5931 361517.5523 3628199.436 GABA 0 0 0 4189075.291 0 0 1728529.782 1078509.147 541050.4329 732334.0322 GalNAC/GlcNAC/ManNAC 53846.05618 118767.4131 38213.75465 63063.60421 12752.97392 83158.56461 198674.9339 53526.96539 85952.92061 47622.04491 GAP / DHAP 47734.70625 0 0 0 0 0 448310.5492 0 0 27837.35664 GDP 381263.5371 657842.849 321978.1542 133600.1476 97929.05212 0 5761891.853 238365.086 3615551.485 2394565.393 GDP-L-fucose 0 0 0 0 0 0 0 0 35988.18322 0 glucono-lactone 525529.1993 160359.8094 0 148518.1923 147838.6768 228840.4052 2457812.864 0 814855.5345 0 glutamine 3396768.257 7064071.273 5028450.773 4158542.275 2578380.359 3048578.3 2114525.794 849267.9999 5042975.474 1961078.707 glutaminic acid 7325868.27 18259400.45 7811013.121 13122806.41 3643265.156 5213475.737 10403620.15 2455616.998 17671114.16 9497886.935 glutathione 48130814.53 19569851.91 11193957.36 10644835.06 2092993.056 10797039.01 32259814.91 26853002.82 13600598.1 12724306.06 glutathione disulfide 2685068.218 1938438.07 1236638.746 916675.3673 548647.7529 379853.7014 5417581.592 2457989.09 1565597.293 1445137.023 glyceric acid 926075.0365 1370153.063 514923.8273 221205.5772 696670.7574 336743.5822 2243533.718 3198905.139 0 0 glycerol 2/3-phosphate 187743.5006 0 0 0 17540.2747 0 1872678.095 575257.1317 210727.686 128442.2564 glycine 0 0 0 0 2806765.386 0 2904543.111 837165.2488 3167555.717 1319546.085 glycylglycine 193270.1378 12606.8326 39334.90379 28941.90447 192946.3304 63179.43907 0 0 0 0 GMP 555060.9844 398503.3111 147390.8428 826158.516 652497.3593 73706.95315 2634343.843 87733.40369 805826.5514 337080.3903 GTP 104224.3453 128940.2174 12181.41754 0 0 0 926215.2535 43617.79014 2236031.585 2698347.06 guanidinoacetate 0 31380.30572 0 1060302.591 620755.6474 1170667.989 1139351.667 162194.3398 0 0 guanine 70974.78749 0 0 65272.83654 48279.60507 0 303656.1869 18362.90449 173149.0295 152878.7184 guanosine 0 0 0 18087.98143 0 0 0 0 0 0 gulonic-lactone 0 1353274.379 705314.3712 3229447.195 598597.6722 1191749.567 1764630.488 660249.4803 1960572.56 1516891.22 histamine 0 266827.9563 291709.0972 420726.7769 123278.4495 0 349417.0513 400176.0737 1163082.29 655130.2638 histidine 2898986.843 1483181.87 1236694.282 1973468.186 897591.3321 2626274.316 6609161.686 2735550.736 4872555.686 3499099.692 homocysteine 0 0 0 0 0 0 50345.5401 0 0 0 homoserine / theronine 3409309.58 1430187.086 1085271.484 2292418.987 940765.2228 1154483.073 10214694.23 3826717.35 7135181.895 5468152.879 homovanillic acid 0 0 0 0 80819.51311 0 34248.37511 20026.44165 0 0 hydroxyphenyllactic acid 23532.18498 0 0 0 71721.19528 0 146269.3795 0 25423.25259 0 hypoxanthine 1740994.23 10694.7006 68386.14597 2295474.765 435534.7617 211492.119 1060669.652 16765.62339 416404.051 317889.899 IDA 37793.08611 12819.53136 11255.66006 35395.81444 30342.3849 0 30337.80213 0 9201.587678 0 IDP 0 1251291.748 2035295.027 0 8434163.721 2175219.466 49399060.57 26953678.65 55755008.14 114413971.3 imidazole-4-acetate 189860.4767 94012.8553 106372.4856 54279.61859 48807.65496 133169.514 126499.671 41339.03839 162107.7348 135232.6224 IMP 1549334.243 489336.3738 448243.08 27507.22666 2569869.928 1725198.624 12373527.87 2610939.977 2232856.474 4628037.786 inosine 352347.0396 0 0 1589136.961 8026.571529 0 275741.3545 17489.57498 0 0 itaconic acid 2575742.622 2650570.816 2502693.623 1447790.316 1195851.813 1879896.578 111586.6809 1815221.514 730766.7928 52268.20286 kynurenine 0 14673.00986 0 0 0 0 36415.25866 0 0 29070.7409 lactic acid 237969026.2 111629072.9 233128903.7 173285160.9 67997423.32 313570090.6 497456153.6 90479298.5 340905466.6 482802343.5 L-arginino-succinate 0 0 0 11293.00708 0 0 163657.7851 0 33000.07684 0 leucine / leucine(iso) 0 0 0 0 3324933.848 0 114492796.7 61957904.93 57128871.49 60185099.58 levulinic acid 0 0 0 0 0 3393083.579 0 1896471.115 4189393.114 5682215.541 L-NMMA 14153.8743 24773.09272 32457.2806 0 28837.9918 55230.40794 495089.7157 12020.74245 18789.10141 29431.18211 lysine 0 0 0 1732075.831 0 0 20548971.76 815625.6967 4001958.73 3607818.757 maleic acid 0 1881354.538 1673616.35 820623.709 0 1287444.682 0 0 0 0 malic acid 122109568 56652839.11 40640802.85 51854778.17 21616185.03 31006318.77 37304804.24 26945279.14 21470582.65 12847478.1 methionine 1821290.88 1049054.027 924826.1868 992756.9836 920199.6704 974565.8162 9015803.943 2957024.055 4600465.803 3563893.491 methylglyoxal 0 0 0 0 0 0 0 0 0 0 methylmalonic acid 11047164.25 9461476.286 7482216.519 8519311.399 1281131.389 12571854.24 2136842.818 0 4381559.5 5681940.395 mevalonic acid 66506.70646 12958.40841 17246.78856 0 47240.24072 68614.43643 0 0 0 0 N1/N8-acetylspermidine 95522733.75 2647608.33 13948194.12 7325331.518 11898540.81 11856007.61 625078002.6 103300356.7 21658017.96 37554723.73 N-acetylaspartic acid 1591995.586 1020251.514 701612.1722 4303531.028 3614988.973 5629908.459 804175.8049 46418.26851 0 0 N-acetylaspartylglutamic acid 0 0 0 34644.04703 0 0 700859.4639 0 0 0 N-acetyl-b-alanine 0 440674.5467 2548142.227 5479398.331 1343447.612 1748174.915 3655760.23 298205.3502 2953779.586 5476272.876 N-acetyl-DL-alanine 0 2409884.587 1991565.737 5479398.331 1343447.612 637657.1915 2814847.232 90105.46823 3790252.231 1618453.759 N-acetyl-dopamine 82957.10708 62192.55594 22510.66073 18326.77751 35864.7035 32626.25846 0 23696.82307 0 19171.75361 N-acetylglucosamine 1/6-phosphate 71458.63276 226622.2015 41581.74023 23879.15573 57226.82519 0 262042.8341 0 567553.5945 400263.0868 NAD+ 86856.30391 24754.30663 37137.1817 39380.07685 18077.10508 0 1893949.872 164611.8952 2474108.248 998472.0062 NADH 0 0 0 0 0 0 0 0 0 0 NADP+ 0 0 0 12594.8757 0 0 1007595.032 29968.64475 704003.3481 345019.7476 NADPH 0 0 0 0 0 0 0 0 78308.57065 61105.84424 N-carbamoyl-L-aspartate 0 0 0 0 0 0 0 0 0 0 nicotinamide 1009705.799 1191993.55 856400.0351 1873242.808 1428458.879 1659380.27 6613197.054 2662734.364 3137188.73 4971992.153 N-methyltryptamine 0 0 0 0 0 0 0 14720.60316 0 14294.3613 NMN 0 0 0 0 0 0 11498.93317 0 8635.036058 0 norepinephrine / pyridoxine 0 14780.55853 11378.98804 0 10622.89103 0 419173.5554 236013.8114 347140.2298 165417.7867 ornithine 0 199468.8011 78068.28925 1134448.769 527792.9772 664979.2715 3249443.07 305582.4345 3625437.949 1507243.045 orotic acid 0 0 0 0 0 0 0 0 0 0 orotidylic acid 0 0 0 0 0 0 0 0 0 0 oxalacetic acid 22153.74507 0 0 0 0 0 0 0 0 0 pantothenic acid 0 0 16004.08042 0 0 30931.90322 2711489.339 54151.45823 900866.6386 1512470.156 phenethylamine 524397.3719 204388.6846 190937.5986 190748.9564 270078.1226 323069.2709 0 0 0 0 phenylalanine 0 0 0 876325.1088 1463593.423 0 18637993.68 6021565.376 8172735.116 12059019 phosphocreatine 500009.9396 49890.56472 42051.96597 2213561.674 2408794.251 989522.3053 8645691.636 2146505.804 246746.6305 148086.9698 phosphoenolpyruvate 52021.07224 148121.6749 161211.27 307309.1648 2110795.297 0 8243608.28 1327873.859 864097.1776 1429267.809 phosphorylcholine+ 14791942.16 3741002.753 3172188.379 2618522.797 3293930.106 1825213.887 72165066.22 17105762.15 37390748.69 25945640.78 proline 5308006.235 10746042.32 0 4773249.843 3245727.204 4982711.907 27983825.17 8993057.033 26355804.83 18286077.94 PRPP 0 0 0 0 0 0 0 0 0 0 pseudouridine 30009.25285 0 0 0 0 0 0 0 0 0 putrescine 21853867.37 2570876.91 1584051.489 1758837.89 2233691.829 2564901.727 34225947.87 4588764.268 1134158.723 1204694.775 pyridoxine 0 0 12049.91179 13716.26354 0 0 27257.14743 137085.1744 43123.60872 21647.91224 pyruvic acid 0 0 0 0 0 0 2709185.84 1808646.326 8413550.286 4635202.814 S-adenosyl-L-homocysteine 0 0 0 0 0 0 0 0 0 0 S-adenosyl-L-methionine+ 20854.43016 0 0 0 0 0 2129616.999 154765.2423 525971.5012 455912.596 S-adenosylmethioninamine+ 0 0 0 0 0 0 0 0 0 0 serine 0 0 588402.7019 923065.8919 746010.1146 424740.3705 390113.3678 1171651.905 649690.2508 79984.46271 serotonin 1017414.651 0 0 0 0 20644.58231 67812.98036 42672.71417 47027.40398 45899.90659 spermidine 132581082 65074049.93 52173717.9 71656094.87 30458862 112552088.7 123816286.2 414793102.2 344947551.5 437536790.8 spermine 30304639.63 20782738.42 19797589.9 28247192.23 12759260.97 40144896.42 14771453.18 19667805.58 12622376.86 12874781.22 succinic acid 0 9748546.522 8515921.428 8130749.378 8547091.833 17496082.22 3686710.856 4466301.175 3756951.158 2046657.55 taurine 41486436.08 74463304.38 45712943.58 20182583.7 27750324.58 3087461.991 181434183.5 9043205.056 443426528.1 316733831.6 thiamine+ 0 0 0 32526.33252 0 0 29293.28173 0 43281.98014 0 thymidine 24346.97384 13607.97067 13309.19289 0 34501.43599 0 0 0 20872.6775 0 thymine 0 0 0 0 401301.8003 0 1220121.292 0 0 590582.6529 tryptamine 0 13221.21429 11567.81558 0 0 0 42358.51722 0 11562.34614 76624.18909 tryptophan 22382.05194 306622.5812 194265.8725 179579.1237 152696.1782 175590.69 2831032.075 313429.3304 1677071.227 1834413.305 tyramine 0 0 0 0 137730.1689 0 175011.6499 373793.0746 0 736102.7753 tyrosine 378586.2861 454468.5149 309705.6278 512779.8796 606383.1171 451179.4054 5787929.25 609615.2915 3624406.799 2892029.674 UDP 258974.1905 1122005.285 782545.8385 291889.9416 372981.9883 0 4069630.189 0 2593531.642 2177067.429 UDP-D-glucose 67903.64289 367589.2542 126340.0173 29891.30965 71370.01357 0 396932.4199 0 734914.0218 694493.3311 UDP-N-acetyl-galactosamine/UDP-N-acetyl-glucosamine 147573.0383 96285.06796 127917.3347 0 0 0 2175344.531 153995.6075 2091390.318 1873565.584 UMP 1145789.473 748432.8357 190539.2348 314434.2947 343611.8345 31711.96282 1564433.071 0 481743.5582 184786.7826 uracil 199535.3643 0 0 0 0 0 580621.0002 499175.0827 0 150798.4133 urea 402407.6893 360899.7642 778729.3497 108767.867 313341.5712 504815.6261 507954.2973 730760.859 508873.6678 1016131.008 uric acid 0 0 0 0 0 0 971159.2661 0 311118.742 60342.34372 uridine 82206.13371 0 0 461980.0148 11110.45181 0 45795.4251 0 0 0 UTP 77139.00365 298186.4358 215461.2857 0 0 0 571751.3349 0 1468372.034 2333987.294 valine 0 0 4258216.617 0 2205658.159 0 39419251.38 1242645.523 2211698.645 23194397.11 xanthine 19003.07977 0 18711.79859 128763.2031 46686.54379 31738.82127 32601.20801 28086.93405 0 0 XMP 0 0 0 0 47418.2751 0 102092.0382 0 0 0 y-glutamylcysteine 0 0 0 0 0 0 0 16451.57371 0 0 MS_METABOLITE_DATA_END #METABOLITES METABOLITES_START metabolite_name 1,3-bisphospho-D-glycerate 2-(Formamido)-N1-(5-phospho-D-ribosyl)acetamidine 2,3-bisphospho-D-glycerate 2/3-phosphoglycerate 2-HG 3,4-dihydroxymandelic acid 3,4-dihydroxyphenylpropanoate 3-hydroxypropanoic acid 3-methoxytyramine 3-phosphohydroxypyruvic acid 3-phospho-serine 3-ureidopropionic acid 4-phosphopantothenate 5,6-dihydroxyindole 5-methylcytosine 5-methyl-THF 5'-methylthioadenosine 6-aminocaproic acid 6-O-methylguanine 6-phosphogluconic acid 6-phosphoglucono-D-lactone acetoacetic acid acetylcholine+ aconitic acid adenine adenosine adenylosuccinate ADP ADP-D-glucose agmatine AICA ribonucleotide a-KG alanine allantoin AMP anandamide arginine asparagine aspartic acid ATP b-alanine betaine carbamoyl phosphate carnitine+ CDP CDP-choline choline+ citraconic acid citramalic acid citrate / citrate(iso) citrulline CMP creatine Creatinine CTP cystathionine cysteine cystine cytidine cytosine dADP dAMP D-arabinono-1,4-lactone dATP dCDP dCMP dCTP dehydroascorbic acid deoxyguanosine deoxyinosine deoxyuridine D-erythrose-4-phosphate dGDP dihydroorotate DL-DOPA DOPAC DOPAL dopamine / octopamine dopamine quinone DOPEG DOPET D-ribose-1/5-phosphate / D-ribulose 5-phosphate D-sedoheptulose-7-phosphate dTMP dTTP dUMP dUTP epinephrine F-1,6/2,6-DP F1P F6P / G1P FAD FMN fumaric acid G6P GABA GalNAC/GlcNAC/ManNAC GAP / DHAP GDP GDP-L-fucose glucono-lactone glutamine glutaminic acid glutathione glutathione disulfide glyceric acid glycerol 2/3-phosphate glycine glycylglycine GMP GTP guanidinoacetate guanine guanosine gulonic-lactone histamine histidine homocysteine homoserine / theronine homovanillic acid hydroxyphenyllactic acid hypoxanthine IDA IDP imidazole-4-acetate IMP inosine itaconic acid kynurenine lactic acid L-arginino-succinate leucine / leucine(iso) levulinic acid L-NMMA lysine maleic acid malic acid methionine methylglyoxal methylmalonic acid mevalonic acid N1/N8-acetylspermidine N-acetylaspartic acid N-acetylaspartylglutamic acid N-acetyl-b-alanine N-acetyl-DL-alanine N-acetyl-dopamine N-acetylglucosamine 1/6-phosphate NAD+ NADH NADP+ NADPH N-carbamoyl-L-aspartate nicotinamide N-methyltryptamine NMN norepinephrine / pyridoxine ornithine orotic acid orotidylic acid oxalacetic acid pantothenic acid phenethylamine phenylalanine phosphocreatine phosphoenolpyruvate phosphorylcholine+ proline PRPP pseudouridine putrescine pyridoxine pyruvic acid S-adenosyl-L-homocysteine S-adenosyl-L-methionine+ S-adenosylmethioninamine+ serine serotonin spermidine spermine succinic acid taurine thiamine+ thymidine thymine tryptamine tryptophan tyramine tyrosine UDP UDP-D-glucose UDP-N-acetyl-galactosamine/UDP-N-acetyl-glucosamine UMP uracil urea uric acid uridine UTP valine xanthine XMP y-glutamylcysteine METABOLITES_END #END