#METABOLOMICS WORKBENCH jgiafaglione_20230912_153152 DATATRACK_ID:4310 STUDY_ID:ST002859 ANALYSIS_ID:AN004689 PROJECT_ID:PR001788
VERSION             	1
CREATED_ON             	September 13, 2023, 3:30 pm
#PROJECT
PR:PROJECT_TITLE                 	MYC is a regulator of AR inhibition-induced metabolic requirements in prostate
PR:PROJECT_TITLE                 	cancer
PR:PROJECT_SUMMARY               	Advanced prostate cancers are treated with therapies targeting the androgen
PR:PROJECT_SUMMARY               	receptor (AR) signaling pathway. While many tumors initially respond to AR
PR:PROJECT_SUMMARY               	inhibition, nearly all develop resistance. It is critical to understand how
PR:PROJECT_SUMMARY               	prostate tumor cells respond to AR inhibition in order to exploit
PR:PROJECT_SUMMARY               	therapy-induced phenotypes prior to the outgrowth of treatment-resistant
PR:PROJECT_SUMMARY               	disease. Here, we comprehensively characterize the effect of AR blockade on
PR:PROJECT_SUMMARY               	prostate cancer metabolism using transcriptomics, metabolomics and bioenergetics
PR:PROJECT_SUMMARY               	approaches. The metabolic response to AR inhibition is defined by reduced
PR:PROJECT_SUMMARY               	glycolysis, robust elongation of mitochondria, and increased reliance on
PR:PROJECT_SUMMARY               	mitochondrial oxidative metabolism. We establish DRP1 activity and MYC signaling
PR:PROJECT_SUMMARY               	as mediators of AR blockade-induced metabolic phenotypes. Rescuing DRP1
PR:PROJECT_SUMMARY               	phosphorylation after AR inhibition restores mitochondrial fission, while
PR:PROJECT_SUMMARY               	rescuing MYC restores glycolytic activity and prevents sensitivity to complex I
PR:PROJECT_SUMMARY               	inhibition. Our study provides new insight into the regulation of
PR:PROJECT_SUMMARY               	treatment-induced metabolic phenotypes and vulnerabilities in prostate cancer.
PR:INSTITUTE                     	UCLA
PR:LABORATORY                    	Goldstein Lab
PR:LAST_NAME                     	Giafaglione
PR:FIRST_NAME                    	Jenna
PR:ADDRESS                       	610 Charles E Young Dr S, Los Angeles, CA, 90024, USA
PR:EMAIL                         	jgiafaglione@g.ucla.edu
PR:PHONE                         	408-728-3065
#STUDY
ST:STUDY_TITLE                   	Glucose tracing in vehicle- and Enzalutamide-treated 16D prostate cancer cells
ST:STUDY_SUMMARY                 	In this study, we sought to determine how Enzalutamide treatment alters
ST:STUDY_SUMMARY                 	metabolism in 16D prostate cancer cells.
ST:INSTITUTE                     	UCLA
ST:LABORATORY                    	Goldstein Lab
ST:LAST_NAME                     	Giafaglione
ST:FIRST_NAME                    	Jenna
ST:ADDRESS                       	610 Charles E Young Dr S, Los Angeles, CA, 90024, USA
ST:EMAIL                         	jgiafaglione@g.ucla.edu
ST:PHONE                         	408-728-3065
#SUBJECT
SU:SUBJECT_TYPE                  	Cultured cells
SU:SUBJECT_SPECIES               	Homo sapiens
SU:TAXONOMY_ID                   	9606
#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           	-	Naive-1	Treatment:Vehicle (DMSO)	RAW_FILE_NAME=Naive-1.D
SUBJECT_SAMPLE_FACTORS           	-	Naive-2	Treatment:Vehicle (DMSO)	RAW_FILE_NAME=Naive-2.D
SUBJECT_SAMPLE_FACTORS           	-	Naive-3	Treatment:Vehicle (DMSO)	RAW_FILE_NAME=Naive-3.D
SUBJECT_SAMPLE_FACTORS           	-	LTEnza-1	Treatment:10uM Enzalutamide	RAW_FILE_NAME=LTEnza-1.D
SUBJECT_SAMPLE_FACTORS           	-	LTEnza-2	Treatment:10uM Enzalutamide	RAW_FILE_NAME=LTEnza-2.D
SUBJECT_SAMPLE_FACTORS           	-	LTEnza-3	Treatment:10uM Enzalutamide	RAW_FILE_NAME=LTEnza-3.D
#COLLECTION
CO:COLLECTION_SUMMARY            	16D prostate cancer cells were cultured with vehicle or Enzalutamide for >2
CO:COLLECTION_SUMMARY            	months.
CO:SAMPLE_TYPE                   	Cultured cells
#TREATMENT
TR:TREATMENT_SUMMARY             	Vehicle- and Enzalutamide-treated 16D cells were cultured in U13C-glucose
TR:TREATMENT_SUMMARY             	containing media for 24 hours.
#SAMPLEPREP
SP:SAMPLEPREP_SUMMARY            	Cell plates were placed on ice and quickly washed with ice-cold 0.9% (w/v) NaCl.
SP:SAMPLEPREP_SUMMARY            	Cells were immediately treated with 500 μL of ice-cold MeOH and 200 μL water
SP:SAMPLEPREP_SUMMARY            	containing 1 μg of the internal standard norvaline. Cells were then scraped and
SP:SAMPLEPREP_SUMMARY            	placed in 1.5 mL centrifuge tubes kept on ice. Next, 500 μL of chloroform was
SP:SAMPLEPREP_SUMMARY            	added, after which samples were vortexed for 1 minute and then spun at 10,000g
SP:SAMPLEPREP_SUMMARY            	for 5 minutes at 4C. The aqueous layer was transferred to a GC/MS sample vial
SP:SAMPLEPREP_SUMMARY            	and dried overnight using a refrigerated CentriVap.
#CHROMATOGRAPHY
CH:CHROMATOGRAPHY_TYPE           	GC
CH:INSTRUMENT_NAME               	Agilent 7890A
CH:COLUMN_NAME                   	Agilent DB5-MS (30m x 0.25mm, 0.25um)
CH:SOLVENT_A                     	N/A (GC-MS)
CH:SOLVENT_B                     	N/A (GC-MS)
CH:FLOW_GRADIENT                 	N/A (GC-MS)
CH:FLOW_RATE                     	1 ml/min
CH:COLUMN_TEMPERATURE            	320
#ANALYSIS
AN:ANALYSIS_TYPE                 	MS
#MS
MS:INSTRUMENT_NAME               	Agilent 5977B
MS:INSTRUMENT_TYPE               	GC-TOF
MS:MS_TYPE                       	EI
MS:ION_MODE                      	POSITIVE
MS:MS_COMMENTS                   	Data was analyzed using Agilent MassHunter software. Stable isotope tracing data
MS:MS_COMMENTS                   	was corrected for natural abundance of heavy isotopes with FluxFix software
MS:MS_COMMENTS                   	using a reference set of unlabeled metabolite standards2.
#MS_METABOLITE_DATA
MS_METABOLITE_DATA:UNITS	abundance
MS_METABOLITE_DATA_START
Samples	LTEnza-1	LTEnza-2	LTEnza-3	Naive-1	Naive-2	Naive-3
Factors	Treatment:10uM Enzalutamide	Treatment:10uM Enzalutamide	Treatment:10uM Enzalutamide	Treatment:Vehicle (DMSO)	Treatment:Vehicle (DMSO)	Treatment:Vehicle (DMSO)
a-Ketoglutarate(M0)	4817	5590	5590	7260	11349	7052
a-Ketoglutarate(M1)	1871	2147	2142	2252	3580	2191
a-Ketoglutarate(M2)	2737	3056	3140	3476	5676	3407
a-Ketoglutarate(M3)	1169	1290	1360	1359	2219	1316
a-Ketoglutarate(M4)	1028	1056	1077	1407	2513	1370
a-Ketoglutarate(M5)	367	379	396	540	955	524
a-Ketoglutarate(M6)	295	266	252	410	825	399
Alanine260(M0)	107078	116383	124676	134414	194701	143788
Alanine260(M1)	26711	29408	31162	35597	51341	38451
Alanine260(M2)	16733	18275	19584	28969	41522	31270
Alanine260(M3)	93374	102326	114773	260684	367279	277440
Alanine260(M4)	18411	20210	22646	51788	73180	55104
Asparagine(M0)	138694	161375	171544	258307	359721	255412
Asparagine(M1)	51958	60489	64388	95639	133582	94495
Asparagine(M2)	27466	31829	34169	50354	70674	49942
Asparagine(M3)	8332	9660	10421	15355	21528	15204
Asparagine(M4)	2594	2969	3253	4774	6668	4692
Asparagine(M5)	656	757	845	1225	1721	1207
Aspartate418(M0)	37420	44921	41319	64308	99647	71398
Aspartate418(M1)	18244	21677	20053	27699	43335	30817
Aspartate418(M2)	16103	18961	17670	27766	43879	31367
Aspartate418(M3)	8330	9676	9242	17333	27316	19098
Aspartate418(M4)	3475	4025	3907	7553	11965	8383
Aspartate418(M5)	1091	1253	1210	2516	3985	2774
Citrate459(M0)	89386	98150	92028	93311	129957	90867
Citrate459(M1)	43634	47900	45226	42689	59637	41523
Citrate459(M2)	58936	64868	62839	94541	130632	91258
Citrate459(M3)	28958	31848	31246	44417	61513	43037
Citrate459(M4)	18281	19942	19889	34791	47864	33527
Citrate459(M5)	8078	8786	8963	18361	25056	17608
Citrate459(M6)	3024	3279	3378	7597	10265	7244
Citrate459(M7)	944	990	1051	2530	3381	2393
Citrate591(M0)	16929	18649	17468	17793	25083	17326
Citrate591(M1)	10150	11301	10592	10200	14406	9950
Citrate591(M2)	12521	13845	13298	19288	26931	18554
Citrate591(M3)	6997	7712	7492	10665	14953	10302
Citrate591(M4)	4348	4788	4735	7984	11104	7671
Citrate591(M5)	2069	2277	2276	4401	6114	4254
Citrate591(M6)	835	910	927	1970	2753	1907
Citrate591(M7)	291	303	319	732	1019	703
Cysteine(M0)	13246	23228	18265	13915	18516	13325
Cysteine(M1)	4610	8080	6392	4831	6457	4615
Cysteine(M2)	2698	4752	3734	2828	3776	2710
Cysteine(M3)	694	1214	968	741	987	704
Cysteine(M4)	204	360	287	223	295	211
Fumarate(M0)	10101	10705	10688	22613	41051	27917
Fumarate(M1)	3938	4106	4105	7127	13060	8857
Fumarate(M2)	4072	4150	4169	8174	15333	10259
Fumarate(M3)	1881	1921	1999	4712	8808	5887
Fumarate(M4)	878	887	934	1922	3567	2392
Fumarate(M5)	213	218	234	507	948	631
GABA(M0)	1159	158	120	4608	459	6009
GABA(M1)	62	29	59	1139	229	76
GABA(M2)	75	94	93	66	80	36
GABA(M3)	13	17	15	323	48	23
GABA(M4)	36	6	2	64	111	60
GABA(M5)	6	13	21	21	78	7
Glutamate432(M0)	487392	546968	522964	515849	671889	528426
Glutamate432(M1)	244872	280644	272333	216119	285079	222165
Glutamate432(M2)	325967	368186	364399	284563	389275	297313
Glutamate432(M3)	165478	185849	186258	131736	182176	138502
Glutamate432(M4)	99790	111241	113290	88411	122590	92797
Glutamate432(M5)	40725	45339	46847	39595	54778	41306
Glutamate432(M6)	13213	14644	15154	13247	18385	13799
Isoleu-274(M0)	17736	20387	20210	17111	30016	21782
Isoleucine274(M0)	15533	17341	18047	16131	28662	20929
Isoleucine274(M1)	3642	4429	4626	4356	6855	4978
Isoleucine274(M2)	1540	1709	1780	1654	2826	2078
Isoleucine274(M3)	267	262	265	257	477	321
Isoleucine274(M4)	49	36	55	2	78	63
Isoleucine274(M5)	1	0	7	2	10	0
Isoleucine274(M6)	1		2	1		3
Isoleucine274(M7)	9	7		14	9	1
Itaconate(M0)	86	89	87	69	97	67
Itaconate(M1)	29	23	29	11	34	26
Itaconate(M2)	10	174	14	12	14	14
Itaconate(M3)	7	138	157	124	179	124
Itaconate(M4)	18434	19676	21296	20997	29644	19267
Itaconate(M5)	4817	5121	5515	5491	7678	4958
Itaconate(M6)	1395	1466	1561	1527	2131	1386
KIC(M0)	2783	2843	3055	3948	6151	4561
KIV(M0)						
KMV(M0)	2273	1970	2100	4214	6030	4755
Lactate261(M0)	23688	24258	27929	82318	116387	86593
Lactate261(M1)	5966	6136	7024	21274	30294	22496
Lactate261(M2)	4253	4373	5074	19866	28040	21004
Lactate261(M3)	27536	27953	33733	196348	273753	207054
Lactate262(M4)	5311	5433	6583	38027	53342	40111
Malate(M0)	19128	21708	21680	59404	93726	66766
Malate(M1)	9783	10987	11033	25220	40266	28592
Malate(M2)	9987	11078	11325	27018	43481	30583
Malate(M3)	5278	5810	6019	16507	26614	18608
Malate(M4)	2381	2651	2734	7457	11959	8398
Malate(M5)	755	832	865	2501	4028	2785
Methionine320(M0)	5439	6383	6380	4989	8659	5835
Methionine320(M1)	1418	1675	1668	1333	2319	1570
Methionine320(M2)	792	940	940	746	1277	869
Methionine320(M3)	165	198	199	169	284	201
Methionine320(M4)	125	143	147	163	345	235
Methionine320(M5)	15	2	17	34	59	41
Methionine320(M6)	10	7	8	17	33	23
N-AcetylAspartate(M0)	1191	2742	106924	760	39393	22455
N-AcetylAspartate(M1)		30408	1417	197	11123	6677
N-AcetylAspartate(M2)		518	33802	12257	19880	11994
N-AcetylAspartate(M3)	7		11656	4111	6703	143
N-AcetylAspartate(M4)			161		4486	91
N-AcetylAspartate(M5)		42	2176	8	1736	1038
Norvaline 260(M0)	385852	424772	451174	308201	464995	351452
Norvaline 260(M1)	95309	105312	111921	76017	115498	87009
Norvaline 260(M2)	37437	41322	44217	29769	45403	34170
Norvaline 260(M3)	5842	6361	6768	4602	6943	5220
Norvaline 288(M0)	322235	355395	373873	256958	392856	295952
Norvaline 288(M1)	81034	89452	94472	64654	99188	74427
Norvaline 288(M2)	32132	35508	37400	25603	39370	29486
Norvaline 288(M3)	5223	5768	6074	4177	6410	4784
Ornithine474(M0)	44	42	44	24	29	26
Ornithine474(M1)	141	142	144	81	112	75
Ornithine474(M2)						
Ornithine474(M3)	47	51	49	31	31	29
Ornithine474(M4)	17	16	15	10	10	9
Ornithine474(M5)	18	25	22	4	6	3
Ornithine474(M6)	13	16	15	2	2	1
Pyruvate(M0)	41826	44348	44545	62023	75730	58373
Pyruvate(M1)	5713	5986	7954	11198	11251	8451
Pyruvate(M2)	4194	4626	4885	10104	12102	9566
Pyruvate(M3)	27410	31627	33381	115858	133796	108709
Pyruvate(M4)	2531	3039	3130	10854	12396	9767
Serine390(M0)	198505	230108	229948	178738	257218	188361
Serine390(M1)	104745	121483	121875	125780	182001	132903
Serine390(M2)	64903	75017	75538	86481	125666	91676
Serine390(M3)	40832	47993	48150	119594	171973	124300
Serine390(M4)	12749	15005	15043	38092	54827	39472
Succinate(M0)	6074	6017	7436	9479	14487	9239
Succinate(M1)	1942	1945	2325	2590	4093	2670
Succinate(M2)	1839	1828	2203	2660	4718	3041
Succinate(M3)	720	713	867	1080	1971	1294
Succinate(M4)	356	369	466	531	986	668
Succinate(M5)	78	83	101	122	225	153
Threonine404(M0)	11248	12626	14573	20703	32352	20982
Threonine404(M1)	3974	4422	5104	7284	11345	7355
Threonine404(M2)	1859	2094	2411	3413	5343	3462
Threonine404(M3)	434	482	549	801	1241	796
Threonine404(M4)	102	114	128	190	289	193
Threonine404(M5)	18	22	25	35	49	34
Urea(M0)	104108	115021	119406	115088	215387	143522
Urea(M1)	21583	24062	24901	23740	45000	30221
Urea(M2)						
Valine288(M0)	6117	6806	6712	5872	10488	7363
Valine288(M1)	1552	1713	1714	1491	2656	1881
Valine288(M2)	599	669	666	586	1033	737
Valine288(M3)	93	120	111	91	175	121
Valine288(M4)	10	16	20	14	35	21
Valine288(M5)	8	1	1	7	12	0
Valine288(M6)	5	0			2	1
MS_METABOLITE_DATA_END
#METABOLITES
METABOLITES_START
metabolite_name
a-Ketoglutarate(M0)
a-Ketoglutarate(M1)
a-Ketoglutarate(M2)
a-Ketoglutarate(M3)
a-Ketoglutarate(M4)
a-Ketoglutarate(M5)
a-Ketoglutarate(M6)
Alanine260(M0)
Alanine260(M1)
Alanine260(M2)
Alanine260(M3)
Alanine260(M4)
Asparagine(M0)
Asparagine(M1)
Asparagine(M2)
Asparagine(M3)
Asparagine(M4)
Asparagine(M5)
Aspartate418(M0)
Aspartate418(M1)
Aspartate418(M2)
Aspartate418(M3)
Aspartate418(M4)
Aspartate418(M5)
Citrate459(M0)
Citrate459(M1)
Citrate459(M2)
Citrate459(M3)
Citrate459(M4)
Citrate459(M5)
Citrate459(M6)
Citrate459(M7)
Citrate591(M0)
Citrate591(M1)
Citrate591(M2)
Citrate591(M3)
Citrate591(M4)
Citrate591(M5)
Citrate591(M6)
Citrate591(M7)
Cysteine(M0)
Cysteine(M1)
Cysteine(M2)
Cysteine(M3)
Cysteine(M4)
Fumarate(M0)
Fumarate(M1)
Fumarate(M2)
Fumarate(M3)
Fumarate(M4)
Fumarate(M5)
GABA(M0)
GABA(M1)
GABA(M2)
GABA(M3)
GABA(M4)
GABA(M5)
Glutamate432(M0)
Glutamate432(M1)
Glutamate432(M2)
Glutamate432(M3)
Glutamate432(M4)
Glutamate432(M5)
Glutamate432(M6)
Isoleu-274(M0)
Isoleucine274(M0)
Isoleucine274(M1)
Isoleucine274(M2)
Isoleucine274(M3)
Isoleucine274(M4)
Isoleucine274(M5)
Isoleucine274(M6)
Isoleucine274(M7)
Itaconate(M0)
Itaconate(M1)
Itaconate(M2)
Itaconate(M3)
Itaconate(M4)
Itaconate(M5)
Itaconate(M6)
KIC(M0)
KIV(M0)
KMV(M0)
Lactate261(M0)
Lactate261(M1)
Lactate261(M2)
Lactate261(M3)
Lactate262(M4)
Malate(M0)
Malate(M1)
Malate(M2)
Malate(M3)
Malate(M4)
Malate(M5)
Methionine320(M0)
Methionine320(M1)
Methionine320(M2)
Methionine320(M3)
Methionine320(M4)
Methionine320(M5)
Methionine320(M6)
N-AcetylAspartate(M0)
N-AcetylAspartate(M1)
N-AcetylAspartate(M2)
N-AcetylAspartate(M3)
N-AcetylAspartate(M4)
N-AcetylAspartate(M5)
Norvaline 260(M0)
Norvaline 260(M1)
Norvaline 260(M2)
Norvaline 260(M3)
Norvaline 288(M0)
Norvaline 288(M1)
Norvaline 288(M2)
Norvaline 288(M3)
Ornithine474(M0)
Ornithine474(M1)
Ornithine474(M2)
Ornithine474(M3)
Ornithine474(M4)
Ornithine474(M5)
Ornithine474(M6)
Pyruvate(M0)
Pyruvate(M1)
Pyruvate(M2)
Pyruvate(M3)
Pyruvate(M4)
Serine390(M0)
Serine390(M1)
Serine390(M2)
Serine390(M3)
Serine390(M4)
Succinate(M0)
Succinate(M1)
Succinate(M2)
Succinate(M3)
Succinate(M4)
Succinate(M5)
Threonine404(M0)
Threonine404(M1)
Threonine404(M2)
Threonine404(M3)
Threonine404(M4)
Threonine404(M5)
Urea(M0)
Urea(M1)
Urea(M2)
Valine288(M0)
Valine288(M1)
Valine288(M2)
Valine288(M3)
Valine288(M4)
Valine288(M5)
Valine288(M6)
METABOLITES_END
#END