#METABOLOMICS WORKBENCH juliehaines_20230221_121940 DATATRACK_ID:3758 STUDY_ID:ST002490 ANALYSIS_ID:AN004064 PROJECT_ID:PR001607
VERSION             	1
CREATED_ON             	February 21, 2023, 12:40 pm
#PROJECT
PR:PROJECT_TITLE                 	Simultaneous targeting of PD-1 and IL2Rβγ with radiation therapy to inhibit
PR:PROJECT_TITLE                 	pancreatic cancer growth and metastasis
PR:PROJECT_SUMMARY               	In pancreatic ductal adenocarcinoma (PDAC) patients, we show that response to
PR:PROJECT_SUMMARY               	radiation therapy (RT) is characterized by increased IL2Rβ and IL2Rγ and
PR:PROJECT_SUMMARY               	decreased ILR2α expression. The bispecific aPD1-IL2v is a PD-1-targeted IL-2
PR:PROJECT_SUMMARY               	variant (IL2v) immunocytokine with engineered IL-2 cis-targeted to PD-1 and
PR:PROJECT_SUMMARY               	abolished IL2Rα binding, which enhances tumor-antigen specific T cell
PR:PROJECT_SUMMARY               	activation while reducing regulatory T cell (Treg) suppression. Using aPD1-IL2v
PR:PROJECT_SUMMARY               	in orthotopic PDAC KPC-driven tumor models, we show marked improvement in local
PR:PROJECT_SUMMARY               	and metastatic survival along with profound increase in tumor-infiltrating
PR:PROJECT_SUMMARY               	polyfunctional CD8+ T cell subsets with a transcriptionally and metabolically
PR:PROJECT_SUMMARY               	active phenotype, and preferential activation of antigen-specific CD8+ T cells.
PR:PROJECT_SUMMARY               	In combination with single dose RT, aPD1-IL2v treatment results in a robust,
PR:PROJECT_SUMMARY               	durable expansion of polyfunctional CD8+ T cells, T cell stemness,
PR:PROJECT_SUMMARY               	tumor-specific memory immune response, natural killer (NK) cell activation, and
PR:PROJECT_SUMMARY               	decreased Tregs. These data show that aPD1-IL2v leads to profound local and
PR:PROJECT_SUMMARY               	distant response in PDAC.
PR:INSTITUTE                     	University of Colorado Denver
PR:LABORATORY                    	Lab of Angelo D'Alessandro in collaboration with lab of Sana Karam
PR:LAST_NAME                     	Haines
PR:FIRST_NAME                    	Julie
PR:ADDRESS                       	12801 E 17th Ave, Room 1303, Aurora, Colorado, 80045, USA
PR:EMAIL                         	julie.haines@cuanschutz.edu
PR:PHONE                         	3037243339
#STUDY
ST:STUDY_TITLE                   	Simultaneous targeting of PD-1 and IL2Rβγ with radiation therapy to inhibit
ST:STUDY_TITLE                   	pancreatic cancer growth and metastasis - human plasma metabolomics
ST:STUDY_SUMMARY                 	Patient plasma samples were collected as part of a Phase I radiation
ST:STUDY_SUMMARY                 	dose-escalation clinical trial (NCT02873598) before, during (6 hours post), and
ST:STUDY_SUMMARY                 	post (6 weeks) SBRT. Samples were collected and analyzed per COMIRB19–0328 for
ST:STUDY_SUMMARY                 	branched chain amino acids and kynurenine.
ST:INSTITUTE                     	University of Colorado Denver
ST:LAST_NAME                     	Haines
ST:FIRST_NAME                    	Julie
ST:ADDRESS                       	12801 E 17th Ave, Room 1303, Aurora, Colorado, 80045, USA
ST:EMAIL                         	julie.haines@cuanschutz.edu
ST:PHONE                         	3037243339
#SUBJECT
SU:SUBJECT_TYPE                  	Human
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           	-	1	Patient ID:C04 | Timepoint:Baseline	RAW_FILE_NAME=BW49-01_r18
SUBJECT_SAMPLE_FACTORS           	-	4	Patient ID:C05 | Timepoint:Baseline	RAW_FILE_NAME=BW49-02_r11
SUBJECT_SAMPLE_FACTORS           	-	7	Patient ID:C06 | Timepoint:Baseline	RAW_FILE_NAME=BW49-03_r5
SUBJECT_SAMPLE_FACTORS           	-	10	Patient ID:C08 | Timepoint:Baseline	RAW_FILE_NAME=BW49-04_r33
SUBJECT_SAMPLE_FACTORS           	-	13	Patient ID:C09 | Timepoint:Baseline	RAW_FILE_NAME=BW49-05_r9
SUBJECT_SAMPLE_FACTORS           	-	16	Patient ID:C10 | Timepoint:Baseline	RAW_FILE_NAME=BW49-06_r22
SUBJECT_SAMPLE_FACTORS           	-	19	Patient ID:C11 | Timepoint:Baseline	RAW_FILE_NAME=BW49-07_r31
SUBJECT_SAMPLE_FACTORS           	-	22	Patient ID:C13 | Timepoint:Baseline	RAW_FILE_NAME=BW49-08_r13
SUBJECT_SAMPLE_FACTORS           	-	25	Patient ID:C14 | Timepoint:Baseline	RAW_FILE_NAME=BW49-09_r35
SUBJECT_SAMPLE_FACTORS           	-	28	Patient ID:C15 | Timepoint:Baseline	RAW_FILE_NAME=BW49-10_r3
SUBJECT_SAMPLE_FACTORS           	-	31	Patient ID:C16 | Timepoint:Baseline	RAW_FILE_NAME=BW49-11_r1
SUBJECT_SAMPLE_FACTORS           	-	34	Patient ID:C17 | Timepoint:Baseline	RAW_FILE_NAME=BW49-12_r23
SUBJECT_SAMPLE_FACTORS           	-	2	Patient ID:C04 | Timepoint:During	RAW_FILE_NAME=BW49-13_r14
SUBJECT_SAMPLE_FACTORS           	-	5	Patient ID:C05 | Timepoint:During	RAW_FILE_NAME=BW49-14_r28
SUBJECT_SAMPLE_FACTORS           	-	8	Patient ID:C06 | Timepoint:During	RAW_FILE_NAME=BW49-15_r21
SUBJECT_SAMPLE_FACTORS           	-	11	Patient ID:C08 | Timepoint:During	RAW_FILE_NAME=BW49-16_r7
SUBJECT_SAMPLE_FACTORS           	-	14	Patient ID:C09 | Timepoint:During	RAW_FILE_NAME=BW49-17_r12
SUBJECT_SAMPLE_FACTORS           	-	17	Patient ID:C10 | Timepoint:During	RAW_FILE_NAME=BW49-18_r30
SUBJECT_SAMPLE_FACTORS           	-	20	Patient ID:C11 | Timepoint:During	RAW_FILE_NAME=BW49-19_r10
SUBJECT_SAMPLE_FACTORS           	-	23	Patient ID:C13 | Timepoint:During	RAW_FILE_NAME=BW49-20_r4
SUBJECT_SAMPLE_FACTORS           	-	26	Patient ID:C14 | Timepoint:During	RAW_FILE_NAME=BW49-21_r32
SUBJECT_SAMPLE_FACTORS           	-	29	Patient ID:C15 | Timepoint:During	RAW_FILE_NAME=BW49-22_r26
SUBJECT_SAMPLE_FACTORS           	-	32	Patient ID:C16 | Timepoint:During	RAW_FILE_NAME=BW49-23_r29
SUBJECT_SAMPLE_FACTORS           	-	35	Patient ID:C17 | Timepoint:During	RAW_FILE_NAME=BW49-24_r15
SUBJECT_SAMPLE_FACTORS           	-	3	Patient ID:C04 | Timepoint:Post	RAW_FILE_NAME=BW49-25_r6
SUBJECT_SAMPLE_FACTORS           	-	6	Patient ID:C05 | Timepoint:Post	RAW_FILE_NAME=BW49-26_r36
SUBJECT_SAMPLE_FACTORS           	-	9	Patient ID:C06 | Timepoint:Post	RAW_FILE_NAME=BW49-27_r20
SUBJECT_SAMPLE_FACTORS           	-	12	Patient ID:C08 | Timepoint:Post	RAW_FILE_NAME=BW49-28_r2
SUBJECT_SAMPLE_FACTORS           	-	15	Patient ID:C09 | Timepoint:Post	RAW_FILE_NAME=BW49-29_r19
SUBJECT_SAMPLE_FACTORS           	-	18	Patient ID:C10 | Timepoint:Post	RAW_FILE_NAME=BW49-30_r8
SUBJECT_SAMPLE_FACTORS           	-	21	Patient ID:C11 | Timepoint:Post	RAW_FILE_NAME=BW49-31_r24
SUBJECT_SAMPLE_FACTORS           	-	24	Patient ID:C13 | Timepoint:Post	RAW_FILE_NAME=BW49-32_r27
SUBJECT_SAMPLE_FACTORS           	-	27	Patient ID:C14 | Timepoint:Post	RAW_FILE_NAME=BW49-33_r25
SUBJECT_SAMPLE_FACTORS           	-	30	Patient ID:C15 | Timepoint:Post	RAW_FILE_NAME=BW49-34_r24
SUBJECT_SAMPLE_FACTORS           	-	33	Patient ID:C16 | Timepoint:Post	RAW_FILE_NAME=BW49-35_r17
#COLLECTION
CO:COLLECTION_SUMMARY            	Patient plasma samples were collected as part of a Phase I radiation
CO:COLLECTION_SUMMARY            	dose-escalation clinical trial (NCT02873598) before, during (6 hours post), and
CO:COLLECTION_SUMMARY            	post (6 weeks) SBRT. Samples were collected and analyzed per COMIRB19–0328.
CO:SAMPLE_TYPE                   	Blood (plasma)
#TREATMENT
TR:TREATMENT_SUMMARY             	All patients received neoadjuvant chemotherapy prior to SBRT. SBRT doses ranged
TR:TREATMENT_SUMMARY             	from 9Gy × 3 fractions to 11Gy ×3 fractions.
#SAMPLEPREP
SP:SAMPLEPREP_SUMMARY            	Plasma aliquots were thawed on ice then a 20 uL aliquot treated with 480 uL of
SP:SAMPLEPREP_SUMMARY            	cold 5:3:2 MeOH:acetonitrile:water. Samples were vortexed 30 min at 4 degrees C
SP:SAMPLEPREP_SUMMARY            	then supernatants clarified by centrifugation (10 min, 10,000 g, 4 degrees C)
SP:SAMPLEPREP_SUMMARY            	and transferred to autosampler vials.
SP:PROCESSING_STORAGE_CONDITIONS 	4℃
SP:EXTRACT_STORAGE               	-80℃
#CHROMATOGRAPHY
CH:CHROMATOGRAPHY_SUMMARY        	Positive C18
CH:CHROMATOGRAPHY_TYPE           	Reversed phase
CH:INSTRUMENT_NAME               	Thermo Vanquish
CH:COLUMN_NAME                   	Phenomenex Kinetex C18 2.1 x 150 mm, 1.7 um
CH:SOLVENT_A                     	100% water; 0.1% formic acid
CH:SOLVENT_B                     	100% acetonitrile; 0.1% formic acid
CH:FLOW_GRADIENT                 	0-0.5 min 5% B, 0.5-1.1 min 5-95% B, 1.1-2.75 min hold at 95% B, 2.75-3 min
CH:FLOW_GRADIENT                 	95-5% B, 3-5 min hold at 5% B.
CH:FLOW_RATE                     	450 uL/min
CH:COLUMN_TEMPERATURE            	45
CH:SAMPLE_INJECTION              	10 uL
#ANALYSIS
AN:ANALYSIS_TYPE                 	MS
#MS
MS:INSTRUMENT_NAME               	Thermo Q Exactive Orbitrap
MS:INSTRUMENT_TYPE               	Orbitrap
MS:MS_TYPE                       	ESI
MS:ION_MODE                      	POSITIVE
MS:MS_COMMENTS                   	Resolution 70,000, scan range 65-900 m/z, maximum injection time 200 ms,
MS:MS_COMMENTS                   	microscans 2, automatic gain control (AGC) 3 x 10^6 ions, source voltage 4.0 kV,
MS:MS_COMMENTS                   	capillary temperature 320 C, and sheath gas 45, auxiliary gas 15, and sweep gas
MS:MS_COMMENTS                   	0 (all nitrogen). Data converted to mzXML using RawConverter. Metabolites were
MS:MS_COMMENTS                   	annotated and integrated using Maven in conjunction with the KEGG database.
#MS_METABOLITE_DATA
MS_METABOLITE_DATA:UNITS	peak area
MS_METABOLITE_DATA_START
Samples	1	4	7	10	13	16	19	22	25	28	31	34	2	5	8	11	14	17	20	23	26	29	32	35	3	6	9	12	15	18	21	24	27	30	33
Factors	Patient ID:C04 | Timepoint:Baseline	Patient ID:C05 | Timepoint:Baseline	Patient ID:C06 | Timepoint:Baseline	Patient ID:C08 | Timepoint:Baseline	Patient ID:C09 | Timepoint:Baseline	Patient ID:C10 | Timepoint:Baseline	Patient ID:C11 | Timepoint:Baseline	Patient ID:C13 | Timepoint:Baseline	Patient ID:C14 | Timepoint:Baseline	Patient ID:C15 | Timepoint:Baseline	Patient ID:C16 | Timepoint:Baseline	Patient ID:C17 | Timepoint:Baseline	Patient ID:C04 | Timepoint:During	Patient ID:C05 | Timepoint:During	Patient ID:C06 | Timepoint:During	Patient ID:C08 | Timepoint:During	Patient ID:C09 | Timepoint:During	Patient ID:C10 | Timepoint:During	Patient ID:C11 | Timepoint:During	Patient ID:C13 | Timepoint:During	Patient ID:C14 | Timepoint:During	Patient ID:C15 | Timepoint:During	Patient ID:C16 | Timepoint:During	Patient ID:C17 | Timepoint:During	Patient ID:C04 | Timepoint:Post	Patient ID:C05 | Timepoint:Post	Patient ID:C06 | Timepoint:Post	Patient ID:C08 | Timepoint:Post	Patient ID:C09 | Timepoint:Post	Patient ID:C10 | Timepoint:Post	Patient ID:C11 | Timepoint:Post	Patient ID:C13 | Timepoint:Post	Patient ID:C14 | Timepoint:Post	Patient ID:C15 | Timepoint:Post	Patient ID:C16 | Timepoint:Post
L-leucine/isoleucine	27215180	28805310	27636740	34797230	28219400	43379600	42851890	36578500	34454600	19722300	19903230	21614110	39810820	34707530	23546040	33257360	25885160	21676690	49355150	53771920	51423520	34932040	18866020	34107500	30690000	46465420	40980990	35473880	49648100	42516120	21356560	57351630	41288030	25407170	70818020
L-valine	60218590	51431160	65579130	57297020	39206380	68792990	45958280	71277590	65534560	35423210	63660700	41626540	63525310	72474180	63148900	59207280	39259580	58136700	63855760	83846550	67293330	55375090	60518910	54950160	58044050	74934730	83326400	77739060	59787490	86874860	54153390	90059580	73366410	53977200	119463100
kynurenine	72157.52	142227.6	189133.3	186570.9	130459.6	198990.8	128218.6	160219.2	102085.6	122405.3	108144.5	313367.5	82897.82	190227.2	125268.2	194202.3	110696.3	113611	258809.7	130823.2	105283.6	170689.9	76949.13	249766.5	116543.8	149513.6	154770.3	237840.7	213216.8	186766.6	177919.4	169804.9	204573.2	128821.3	216468.3
MS_METABOLITE_DATA_END
#METABOLITES
METABOLITES_START
metabolite_name	KEGG ID	m/z	medRt
L-leucine/isoleucine	C00123	132.102	0.6674764
L-valine	C00183	118.0865	0.6581454
kynurenine	C00328	209.0919	0.6664889
METABOLITES_END
#END