#METABOLOMICS WORKBENCH STUDY_ID:ST000276 ANALYSIS_ID:AN000440 PROJECT_ID:PR000154 VERSION 1 CREATED_ON 01-15-2019 #PROJECT PR:PROJECT_TITLE Isocitrate dehydrogenase-1 and Glioma Studies PR:PROJECT_SUMMARY Dr. Stegh will define the role of a novel glioma oncoprotein, termed isocitrate PR:PROJECT_SUMMARY dehydrogenase-1 (IDH1), in driving progression and therapy resistance of PR:PROJECT_SUMMARY glioblastoma (GBM). Understanding the molecular basis of the therapy PR:PROJECT_SUMMARY refractoriness of GBM is one of the most important areas of glioma research. PR:PROJECT_SUMMARY IDH1 is a critical enzyme of the citric acid cycle (CAC) and is a master PR:PROJECT_SUMMARY regulator of metabolism. Building on his preliminary studies, Dr. Stegh will PR:PROJECT_SUMMARY molecularly characterize the precise mechanism, by which IDH1 protects glioma PR:PROJECT_SUMMARY cells from therapy-induced cell death using glioma cell and mouse models. To PR:PROJECT_SUMMARY target IDH1 signaling in GBM, he will leverage these model systems and PR:PROJECT_SUMMARY mechanistical knowledge to develop and preclinically characterize RNA PR:PROJECT_SUMMARY interference RNAi-based nanomaterials. He will generate RNAi-functionalized PR:PROJECT_SUMMARY spherical nucleic acids (SNAs) that neutralize IDH1 expression in established PR:PROJECT_SUMMARY gliomas. Due to the negative charge of the RNA backbone, however, siRNA PR:PROJECT_SUMMARY oligonucleotides have many downsides, such as they trigger auto-immune PR:PROJECT_SUMMARY responses, and cannot cross the blood-brain-barrier (BBB). In contrast, SNAs are PR:PROJECT_SUMMARY able to transverse cellular membranes, do not require the use of toxic auxiliary PR:PROJECT_SUMMARY reagents, and accumulate in cells and intracranial tumors very effectively. They PR:PROJECT_SUMMARY also exhibit extraordinary stability in physiological environments, cross the PR:PROJECT_SUMMARY BBB, are highly resistant to nuclease degradation, and thus, can move through PR:PROJECT_SUMMARY biological fluids and avoid being destroyed as “foreign materials.” Dr. PR:PROJECT_SUMMARY Stegh proposes to preclinically evaluate these IDH1-targeting nanoconjugates to PR:PROJECT_SUMMARY provide a fundamentally novel treatment option of patients diagnosed with GBM, PR:PROJECT_SUMMARY and will aid in successfully implementing RNAi-based therapies into PR:PROJECT_SUMMARY neuro-oncological practice. PR:INSTITUTE Northwestern University PR:DEPARTMENT Neurology PR:LABORATORY Stegh Lab PR:LAST_NAME Stegh PR:FIRST_NAME Alexander PR:ADDRESS Evanston, IL PR:EMAIL a-stegh@northwestern.edu PR:PHONE 312-503-2879 PR:DOI http://dx.doi.org/10.21228/M8688J #STUDY ST:STUDY_TITLE IDH1 and Glioma knockdown idh1 ST:STUDY_SUMMARY This study is a part of series performed for the same researcher through ST:STUDY_SUMMARY pilot/feasibility grant program, so the publication is relevant reference for ST:STUDY_SUMMARY other studies (ST000199)This specific experiment is a pilot study to compare the ST:STUDY_SUMMARY metabolism of cells transformed using empty vector against cells transformed ST:STUDY_SUMMARY with vector carrying short hairpin RNA (shRNA) targeted to silence isocitrate ST:STUDY_SUMMARY dehydrogenase-1 (IDH1) gene. ST:INSTITUTE University of Michigan ST:DEPARTMENT Neurology ST:LABORATORY Stegh Lab (Northwestern University) ST:LAST_NAME Calvert ST:FIRST_NAME Andrea ST:ADDRESS Evanston, IL ST:EMAIL a-calvert@u.northwestern.edu ST:PHONE 312-503-3134 #SUBJECT SU:SUBJECT_TYPE Human SU:SUBJECT_SPECIES Homo sapiens SU:TAXONOMY_ID 9606 SU:SPECIES_GROUP Human #SUBJECT_SAMPLE_FACTORS: SUBJECT(optional)[tab]SAMPLE[tab]FACTORS(NAME:VALUE pairs separated by |)[tab]Additional sample data SUBJECT_SAMPLE_FACTORS - S00014999 IDH1 Knockdown:No SUBJECT_SAMPLE_FACTORS - S00015000 IDH1 Knockdown:No SUBJECT_SAMPLE_FACTORS - S00015001 IDH1 Knockdown:No SUBJECT_SAMPLE_FACTORS - S00015002 IDH1 Knockdown:No SUBJECT_SAMPLE_FACTORS - S00015003 IDH1 Knockdown:No SUBJECT_SAMPLE_FACTORS - S00015004 IDH1 Knockdown:yes SUBJECT_SAMPLE_FACTORS - S00015005 IDH1 Knockdown:yes SUBJECT_SAMPLE_FACTORS - S00015006 IDH1 Knockdown:yes SUBJECT_SAMPLE_FACTORS - S00015007 IDH1 Knockdown:yes SUBJECT_SAMPLE_FACTORS - S00015008 IDH1 Knockdown:yes #COLLECTION CO:SAMPLE_TYPE Cells, Cultured CO:COLLECTION_SUMMARY - #TREATMENT TR:TREATMENT_SUMMARY - #SAMPLEPREP SP:SAMPLEPREP_PROTOCOL_FILENAME Gly-TCA-nucleotides_analysis_protocol-2015-03-09.docx SP:SAMPLEPREP_SUMMARY - #CHROMATOGRAPHY CH:METHODS_ID AQM020 CH:METHODS_FILENAME QTOF-002-HILIC-35min-1mm.m CH:INSTRUMENT_NAME Agilent 1260 CH:COLUMN_NAME Phenomenex Luna NH2 (150 x 1mm, 3um) CH:CHROMATOGRAPHY_TYPE HILIC #ANALYSIS AN:LABORATORY_NAME MRC2 (University of Michigan) AN:ANALYSIS_TYPE MS AN:ACQUISITION_PARAMETERS_FILE Column1_solv1_jetstream+_grad9.m AN:PROCESSING_PARAMETERS_FILE EX00317-MassHunterQuant-GlyTCA-DataAnalysis-LCMS-Method.m #MS MS:MS_COMMENTS - MS:INSTRUMENT_NAME Agilent 6530 QTOF MS:INSTRUMENT_TYPE QTOF MS:MS_TYPE ESI MS:ION_MODE POSITIVE #MS_METABOLITE_DATA MS_METABOLITE_DATA:UNITS pmol/µg protein MS_METABOLITE_DATA_START Samples S00014999 S00015000 S00015001 S00015002 S00015003 S00015004 S00015005 S00015006 S00015007 S00015008 Factors IDH1 Knockdown:No IDH1 Knockdown:No IDH1 Knockdown:No IDH1 Knockdown:No IDH1 Knockdown:No IDH1 Knockdown:yes IDH1 Knockdown:yes IDH1 Knockdown:yes IDH1 Knockdown:yes IDH1 Knockdown:yes 2 or 3-phosphoglycerate_2PG/3PG 15.2684 20.6190 23.4381 12.2102 15.2395 22.2005 15.0676 21.5047 15.1398 16.4459 6-Phosphogluconate 17.6862 23.7846 26.4129 13.7886 19.1897 27.0367 17.7068 27.2094 19.1007 21.8335 Acetyl-CoA 16.8594 22.4543 24.8554 13.1727 16.5836 22.1267 15.1568 22.5592 16.6330 17.4050 ADP 141.8273 180.8952 198.6299 129.0832 98.7574 205.7849 151.8220 236.9052 194.7472 181.0240 AMP 652.5372 829.6242 919.1071 448.6940 428.6916 749.0836 541.5650 735.6193 574.4925 463.0508 ATP 461.9507 470.3373 552.7637 479.1216 262.0738 694.2409 464.7696 870.7787 777.8531 794.8952 Citrate or Isocitrate 936.5463 949.5495 1230.5785 690.9270 821.0633 709.6282 465.9267 727.7756 626.5704 669.6705 erythrose 33.0442 28.7963 77.8063 24.1673 62.2370 7.6639 5.0417 9.2515 26.6524 Erythrose 4-phosphate 35.6925 29.9809 61.1633 29.5472 37.2907 42.0780 29.6363 55.4645 47.4340 45.8037 flavin adenine dinucleotide 7.4979 10.3560 12.1225 6.2196 7.6605 10.3921 7.1655 11.8516 8.7628 9.0587 Fructose 1_6-bisphosphate 69.3080 82.8799 98.7222 57.2519 45.6666 71.5205 60.1630 78.8434 60.9669 61.6814 Glucose 6-phosphate or fructose-6-phosphate 85.1880 75.1851 99.5117 60.8493 57.9876 91.3182 56.2837 114.4483 86.0812 92.1941 guanosine 5^-diphosphate 23.2924 45.0739 47.6427 28.1244 20.4777 44.3769 38.4507 69.3249 48.6709 52.9583 guanosine 5^-monophosphate 3.4553 5.3458 5.6254 3.0489 3.1901 4.5709 3.5829 5.8280 3.8469 3.7847 guanosine 5^-triphosphate 16.5563 26.3197 29.0029 17.4227 18.3531 29.8463 21.6192 38.5941 30.0206 34.9164 Malate 991.9140 907.7035 1279.6747 866.3719 698.6432 1111.6626 699.9722 944.3375 940.4108 902.7140 Nicotinamide Adenine Dinucleotide oxidized 90.7688 93.6636 121.4057 73.7462 63.4411 139.0454 86.6988 145.4194 112.1841 113.1869 nicotinamide adenine dinucleotide phosphate oxidized 4.3046 1.0805 4.2444 2.2002 3.7090 1.4878 5.8322 3.2584 3.2117 nicotinamide adenine dinucleotide phosphate reduced 26.7725 35.0579 36.4935 25.6258 24.2748 33.2817 27.0430 32.3769 31.7478 29.7391 Nicotinamide Adenine Dinucleotide reduced 13.8329 21.1491 23.3243 11.6355 15.8388 19.9616 13.2308 20.6977 13.9447 14.9794 Phosphoenolpyruvate 30.9780 45.3703 49.8261 24.6009 35.2011 46.0490 30.7370 30.9441 Ribose-5-phosphate or xylulose 5-phosphate 23.7059 14.9983 28.0342 15.1046 11.5529 28.3019 14.9329 24.1783 19.8986 18.4868 sedoheptulose 10.4741 12.7426 15.8358 5.6668 8.5453 10.3963 6.9639 11.3288 8.3560 10.3075 Sedoheptulose 7-phosphate 15.0706 17.8956 23.3110 13.3722 16.0837 23.7804 12.5196 28.5041 19.0734 19.9277 Sucrose 62.3013 85.6996 102.1007 54.5533 62.3990 100.4478 66.7923 89.4629 73.0057 79.7717 MS_METABOLITE_DATA_END #METABOLITES METABOLITES_START metabolite_name pubchem_id inchi_key kegg_id other_id other_id_type ri ri_type moverz_quant 2 or 3-phosphoglycerate_2PG/3PG 724 C00597 2PG/3PG UM_Target_ID 6-Phosphogluconate 91493 C00345 6PG UM_Target_ID Acetyl-CoA 444493 C00024 aCoA UM_Target_ID ADP 6022 C00008 ADP UM_Target_ID AMP 6083 C00020 AMP UM_Target_ID ATP 5957 C00002 ATP UM_Target_ID Citrate or Isocitrate 311 C00158 CIT/ICIT UM_Target_ID erythrose 94176 C02143 ERY UM_Target_ID Erythrose 4-phosphate 122357 C00279 E4P UM_Target_ID flavin adenine dinucleotide 643975 C00016 FAD UM_Target_ID Fructose 1,6-bisphosphate 172313 C00354 FBP UM_Target_ID Glucose 6-phosphate or fructose-6-phosphate 5958 C00092 G6P/F6P UM_Target_ID guanosine 5'-diphosphate 8977 C00035 GDP UM_Target_ID guanosine 5'-monophosphate 6804 C00144 GMP UM_Target_ID guanosine 5'-triphosphate 6830 C00044 GTP UM_Target_ID Malate 222656 C00149 MAL UM_Target_ID Nicotinamide Adenine Dinucleotide oxidized 10897651 C00003 NAD UM_Target_ID nicotinamide adenine dinucleotide phosphate oxidized 5886 C00006 NADP UM_Target_ID nicotinamide adenine dinucleotide phosphate reduced 5886 C00006 NADPH UM_Target_ID Nicotinamide Adenine Dinucleotide reduced 439153 C00004 NADH UM_Target_ID Phosphoenolpyruvate 1005 C00074 PEP UM_Target_ID Ribose-5-phosphate or xylulose 5-phosphate 77982 C00117 R5P/X5P UM_Target_ID sedoheptulose 102926 SED UM_Target_ID Sedoheptulose 7-phosphate 165007 C05382 S7P UM_Target_ID Sucrose 5988 C00089 SUC UM_Target_ID METABOLITES_END #END