#METABOLOMICS WORKBENCH TEXT OUTPUT STUDY_ID:	ANALYSIS_ID:	
VERSION             	1
CREATED_ON          	2017-07-14
#PROJECT
PR:PROJECT_TITLE    	Stimulated Raman Immunohistochemistry
PR:PROJECT_TYPE     	MS analysis
PR:PROJECT_SUMMARY  	Profiling of WT, IDH1 R132H, WT + 2HG, Mut + AG!5198
PR:INSTITUTE        	University of Michigan
PR:DEPARTMENT       	Neursurgery
PR:LABORATORY       	Orringer Lab
PR:LAST_NAME        	Orringer
PR:FIRST_NAME       	Daniel
PR:ADDRESS          	Ann Arbor, MI
PR:EMAIL            	dorringe@umich.edu
PR:PHONE            	734-647-9222
PR:FUNDING_SOURCE   	
PR:PROJECT_COMMENTS 	
PR:PUBLICATIONS     	
#STUDY
ST:STUDY_TITLE      	Metabolomic profiling of IDH1 mutation
ST:STUDY_TYPE       	MS analysis
ST:STUDY_SUMMARY    	We are interested in using both spontaneous and stimulated Raman microscopy to visualize these metabolomic changes as spectral alterations. We have two isogneic cell lines of normal human astrocytes differing only by a point mutation in the IDH-1 gene. We will work with the metabolomics core to elucidate the changes in central metabolism and lipid synthesys in an effort to determine the precise biochemical alterations underlying observed spectral differences. We wil then use a selective inhibitor of the IDH1 R132H to demonstrate to attempt to return TCA metabolome and lipidome to WT phenotype. Lastly, we will use a cell-permeablized variant of 2HG (2R-octyl-alpha-hydroxyglutarate) to recaptitulate the R132H mutant phenotype in wild-type cells, providing strong evidence that 2HG accumulation uderlies the metabolomic (and thus, spectral) changes observed.
ST:INSTITUTE        	University of Michigan
ST:DEPARTMENT       	Biomedical Research Core Facilities
ST:LABORATORY       	Metabolomics core
ST:LAST_NAME        	Kachman
ST:FIRST_NAME       	Maureen
ST:ADDRESS          	6300 Brehm Tower, 1000 Wall Street, Ann Arbor, MI 48105-5714
ST:EMAIL            	mkachman@med.umich.edu
ST:PHONE            	(734) 232-8175
ST:SUBMIT_DATE      	11/10/2016 02:54:55 PM
ST:NUM_GROUPS       	2
ST:TOTAL_SUBJECTS   	3
ST:STUDY_COMMENTS   	The IDH-1 R132H mutation in low grade gliomas preceptitates oncogenesis through a neomorphic enzyme function: the reduction of alpha-ketoglutarate (aKG) to 2-hydroxygutarate (2HG). 2HG accumukates to extremely high (~100mM) concentrations in IDH1 mutant cells and has substantial, predictable metabolic effects, including inhibition of a-KG dependent dioxygenases and hypermethylation of histones and chromatin.
#SUBJECT_SAMPLE_FACTORS:          	SUBJECT(optional)[tab]SAMPLE[tab]FACTORS(NAME:VALUE pairs separated by |)[tab]Additional sample data
SUBJECT_SAMPLE_FACTORS            	SU0016656	S00026530	IDH1_MUTATION_STATUS:WT	
SUBJECT_SAMPLE_FACTORS            	SU0016657	S00026531	IDH1_MUTATION_STATUS:WT	
SUBJECT_SAMPLE_FACTORS            	SU0016658	S00026532	IDH1_MUTATION_STATUS:WT	
SUBJECT_SAMPLE_FACTORS            	SU0016656	S00026533	IDH1_MUTATION_STATUS:WT	
SUBJECT_SAMPLE_FACTORS            	SU0016657	S00026534	IDH1_MUTATION_STATUS:WT	
SUBJECT_SAMPLE_FACTORS            	SU0016658	S00026535	IDH1_MUTATION_STATUS:WT	
SUBJECT_SAMPLE_FACTORS            	SU0016656	S00026536	IDH1_MUTATION_STATUS:WT	
SUBJECT_SAMPLE_FACTORS            	SU0016657	S00026537	IDH1_MUTATION_STATUS:WT	
SUBJECT_SAMPLE_FACTORS            	SU0016658	S00026538	IDH1_MUTATION_STATUS:WT	
SUBJECT_SAMPLE_FACTORS            	SU0016656	S00026539	IDH1_MUTATION_STATUS:R132H Mutant	
SUBJECT_SAMPLE_FACTORS            	SU0016657	S00026540	IDH1_MUTATION_STATUS:R132H Mutant	
SUBJECT_SAMPLE_FACTORS            	SU0016658	S00026541	IDH1_MUTATION_STATUS:R132H Mutant	
SUBJECT_SAMPLE_FACTORS            	SU0016656	S00026542	IDH1_MUTATION_STATUS:R132H Mutant	
SUBJECT_SAMPLE_FACTORS            	SU0016657	S00026543	IDH1_MUTATION_STATUS:R132H Mutant	
SUBJECT_SAMPLE_FACTORS            	SU0016658	S00026544	IDH1_MUTATION_STATUS:R132H Mutant	
SUBJECT_SAMPLE_FACTORS            	SU0016656	S00026545	IDH1_MUTATION_STATUS:R132H Mutant	
SUBJECT_SAMPLE_FACTORS            	SU0016657	S00026546	IDH1_MUTATION_STATUS:R132H Mutant	
SUBJECT_SAMPLE_FACTORS            	SU0016658	S00026547	IDH1_MUTATION_STATUS:R132H Mutant	
#SUBJECT
SU:SUBJECT_TYPE                   	HUMAN
SU:SUBJECT_SPECIES                	Homo sapiens
SU:TAXONOMY_ID                    	9606
#COLLECTION
CO:COLLECTION_SUMMARY             	-
#TREATMENT
TR:TREATMENT_SUMMARY              	-
#SAMPLEPREP
SP:SAMPLEPREP_SUMMARY             	-
SP:SAMPLEPREP_PROTOCOL_FILENAME   	A037-2HG cells updated-20160721.pdf
#CHROMATOGRAPHY
CH:CHROMATOGRAPHY_TYPE            	C18
CH:INSTRUMENT_NAME                	6490A QQQ LC/MS (Agilent)
CH:COLUMN_NAME                    	ACQUITY UPLC HSS T3, 100A, 1.8mkm, 2.1mm X 50mm (Waters Corporation)
CH:METHODS_FILENAME               	2016-03-16-2HG-6490
#ANALYSIS
AN:ANALYSIS_TYPE                  	MRM
AN:INSTRUMENT_NAME                	6490A QQQ LC/MS (Agilent)
AN:ANALYSIS_PROTOCOL_FILE         	A037-2HG cells updated-20160721.pdf
AN:ACQUISITION_PARAMETERS_FILE    	2016-03-16-2HG-6490
AN:PROCESSING_PARAMETERS_FILE     	-
#MS
MS:INSTRUMENT_TYPE                	QQQ
MS:MS_TYPE                        	
MS:ION_MODE                       	NEGATIVE
#MS_METABOLITE_DATA
MS_METABOLITE_DATA:UNITS	uM/ng protein
MS_METABOLITE_DATA_START
Sample	S00026530	S00026531	S00026532	S00026533	S00026534	S00026535	S00026536	S00026537	S00026538	S00026539	S00026540	S00026541	S00026542	S00026543	S00026544	S00026545	S00026546	S00026547
Factors	IDH1_MUTATION_STATUS:WT	IDH1_MUTATION_STATUS:WT	IDH1_MUTATION_STATUS:WT	IDH1_MUTATION_STATUS:WT	IDH1_MUTATION_STATUS:WT	IDH1_MUTATION_STATUS:WT	IDH1_MUTATION_STATUS:WT	IDH1_MUTATION_STATUS:WT	IDH1_MUTATION_STATUS:WT	IDH1_MUTATION_STATUS:R132H Mutant	IDH1_MUTATION_STATUS:R132H Mutant	IDH1_MUTATION_STATUS:R132H Mutant	IDH1_MUTATION_STATUS:R132H Mutant	IDH1_MUTATION_STATUS:R132H Mutant	IDH1_MUTATION_STATUS:R132H Mutant	IDH1_MUTATION_STATUS:R132H Mutant	IDH1_MUTATION_STATUS:R132H Mutant	IDH1_MUTATION_STATUS:R132H Mutant
(2R)-2-HYDROXYPENTANEDIOIC ACID	4.071306714	3.098099464	4.703858735	4.798879275	3.086003529	4.044164366	2.321686412	2.332264382	2.737507717	43.54915156	38.63892668	24.66584537	38.18522914	42.39913148	41.66765007	34.76693065	33.5109968	32.91367874
(2S)-2-HYDROXYPENTANEDIOIC ACID	7.295459461	5.89119898	5.925238318	6.187509603	6.209511497	5.454204664	5.148028685	5.1653177	5.473715581	6.299511709	4.89508229	3.822237079	4.885550348	6.014301899	4.834213193	4.660787068	4.835995756	4.299657892
MS_METABOLITE_DATA_END
#METABOLITES
METABOLITES_START
metabolite_name	moverz_quant	ri	ri_type	pubchem_id	inchi_key	kegg_id	other_id	other_id_type
(2R)-2-HYDROXYPENTANEDIOIC ACID				439391				
(2S)-2-HYDROXYPENTANEDIOIC ACID				439939				
METABOLITES_END
