#METABOLOMICS WORKBENCH fernandezlab_20191213_081922_mwtab.txt DATATRACK_ID:1869 STUDY_ID:ST001290 ANALYSIS_ID:AN002143 PROJECT_ID:PR000872 VERSION 1 CREATED_ON December 17, 2019, 1:42 pm #PROJECT PR:PROJECT_TITLE Lipidome Signatures of Metastasis in a Transgenic Mouse Model of Sonic Hedgehog PR:PROJECT_TITLE Medulloblastoma. PR:PROJECT_TYPE Untargeted metabolomics PR:PROJECT_SUMMARY Medulloblastoma (MB), the most common malignant pediatric brain tumor, has high PR:PROJECT_SUMMARY propensity to metastasize. Currently, the standard treatment for MB patients PR:PROJECT_SUMMARY includes radiation therapy administered to the entire brain and spine for the PR:PROJECT_SUMMARY purpose of treating or preventing against metastasis. Due to this aggressive PR:PROJECT_SUMMARY treatment, the majority of long-term survivors will be left with permanent and PR:PROJECT_SUMMARY debilitating neurocognitive impairments and, for the 30-40% patients that fail PR:PROJECT_SUMMARY to respond to radiation, all will relapse with terminal metastatic disease. An PR:PROJECT_SUMMARY understanding of the underlying biology that drives MB metastasis is lacking, PR:PROJECT_SUMMARY and is critically needed in order to develop targeted therapeutics for its PR:PROJECT_SUMMARY prevention. To examine the metastatic biology of sonic hedgehog (SHH) MB, the PR:PROJECT_SUMMARY human MB subgroup with the worst clinical outcome, we first generated a robust PR:PROJECT_SUMMARY SmoA1-Math-GFP mouse model that reliably reproduces human SHH MB whereby PR:PROJECT_SUMMARY metastases can be visualized under fluorescence microscopy. Lipidome alterations PR:PROJECT_SUMMARY associated with metastasis were then investigated by applying Ultra-Performance PR:PROJECT_SUMMARY Liquid Chromatography Mass Spectrometry (UPLC-MS) under both positive and PR:PROJECT_SUMMARY negative ionization modes to primary tumor samples collected from mice with PR:PROJECT_SUMMARY (n=18) and without (n=7) metastasis. This study provides the first insights into PR:PROJECT_SUMMARY dysregulations of lipid metabolism associated with SHH MB metastatic PR:PROJECT_SUMMARY progression, and thus serves as a guide toward novel targeted therapies. PR:INSTITUTE Georgia Institute of Technology PR:DEPARTMENT Chemistry PR:LABORATORY Fernández PR:LAST_NAME Huang PR:FIRST_NAME Danning PR:ADDRESS 901 Atlantic Dr NE, Atlanta, GA, 30332, USA PR:EMAIL dhuang74@gatech.edu PR:PHONE 404-512-7523 #STUDY ST:STUDY_TITLE Lipidome Signatures of Metastasis in a Transgenic Mouse Model of Sonic Hedgehog ST:STUDY_TITLE Medulloblastoma. ST:STUDY_TYPE Untargeted metabolomics ST:STUDY_SUMMARY Metabolic alternations were investigated by applying Ultra Performance Liquid ST:STUDY_SUMMARY Chromatography Mass Spectrometry (UPLC-MS) to mice brain tissue samples ST:STUDY_SUMMARY collected from SmoA1-Math-GFP mice with (n=18) and without (n=7) metastasis. All ST:STUDY_SUMMARY samples were analyzed using reverse phase (RP) UPLC-MS analysis in positive and ST:STUDY_SUMMARY negative ion modes. ST:INSTITUTE Georgia Institute of Technology ST:DEPARTMENT Chemistry ST:LABORATORY Fernández ST:LAST_NAME Huang ST:FIRST_NAME Danning ST:ADDRESS 901 Atlantic Dr NE ST:EMAIL dhuang74@gatech.edu ST:PHONE 4045127523 ST:NUM_GROUPS 2 ST:TOTAL_SUBJECTS 25 ST:NUM_MALES 5 ST:NUM_FEMALES 20 #SUBJECT SU:SUBJECT_TYPE Mammal SU:SUBJECT_SPECIES Mus musculus SU:TAXONOMY_ID 10090 SU:GENOTYPE_STRAIN SmoA1-Math-GFP SU:AGE_OR_AGE_RANGE 12-36 weeks SU:GENDER Male and female #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 - SG215 Mice strain:SmoA1-Math-GFP | Group:PT DOB=7/20/14; Sex=F; Age (weeks)=26; RAW_FILE_NAME=SG215.raw SUBJECT_SAMPLE_FACTORS - SG218 Mice strain:SmoA1-Math-GFP | Group:PT DOB=7/20/14; Sex=F; Age (weeks)=26; RAW_FILE_NAME=SG218.raw SUBJECT_SAMPLE_FACTORS - SG174 Mice strain:SmoA1-Math-GFP | Group:PT DOB=5/13/14; Sex=M; Age (weeks)=30; RAW_FILE_NAME=SG174.raw SUBJECT_SAMPLE_FACTORS - SG221 Mice strain:SmoA1-Math-GFP | Group:PT DOB=7/21/14; Sex=F; Age (weeks)=13; RAW_FILE_NAME=SG221.raw SUBJECT_SAMPLE_FACTORS - SG223 Mice strain:SmoA1-Math-GFP | Group:PT DOB=7/21/14; Sex=F; Age (weeks)=14; RAW_FILE_NAME=SG223.raw SUBJECT_SAMPLE_FACTORS - SG203 Mice strain:SmoA1-Math-GFP | Group:PT DOB=6/15/14; Sex=F; Age (weeks)=22; RAW_FILE_NAME=SG203.raw SUBJECT_SAMPLE_FACTORS - SG175 Mice strain:SmoA1-Math-GFP | Group:PT DOB=3/12/14; Sex=F; Age (weeks)=36; RAW_FILE_NAME=SG175.raw SUBJECT_SAMPLE_FACTORS - SG204 Mice strain:SmoA1-Math-GFP | Group:PT DOB=6/15/14; Sex=F; Age (weeks)=23; RAW_FILE_NAME=SG204.raw SUBJECT_SAMPLE_FACTORS - SG217 Mice strain:SmoA1-Math-GFP | Group:PT DOB=7/21/14; Sex=F; Age (weeks)=20; RAW_FILE_NAME=SG217.raw SUBJECT_SAMPLE_FACTORS - SGTA11 Mice strain:SmoA1-Math-GFP | Group:PT DOB=7/10/14; Sex=F; Age (weeks)=22; RAW_FILE_NAME=SGTA11.raw SUBJECT_SAMPLE_FACTORS - SGTA9 Mice strain:SmoA1-Math-GFP | Group:PT DOB=4/28/14; Sex=F; Age (weeks)=27; RAW_FILE_NAME=SGTA9.raw SUBJECT_SAMPLE_FACTORS - SGTA10 Mice strain:SmoA1-Math-GFP | Group:PT DOB=6/10/14; Sex=F; Age (weeks)=21; RAW_FILE_NAME=SGTA10.raw SUBJECT_SAMPLE_FACTORS - SGTA4 Mice strain:SmoA1-Math-GFP | Group:PT DOB=2/1/14; Sex=F; Age (weeks)=18; RAW_FILE_NAME=SGTA4.raw SUBJECT_SAMPLE_FACTORS - SGTA3 Mice strain:SmoA1-Math-GFP | Group:PT DOB=10/16/13; Sex=F; Age (weeks)=34; RAW_FILE_NAME=SGTA3.raw SUBJECT_SAMPLE_FACTORS - SG105 Mice strain:SmoA1-Math-GFP | Group:PT DOB=9/23/14; Sex=M; Age (weeks)=29; RAW_FILE_NAME=SG105.raw SUBJECT_SAMPLE_FACTORS - SG138 Mice strain:SmoA1-Math-GFP | Group:PT DOB=11/2/13; Sex=F; Age (weeks)=21; RAW_FILE_NAME=SG138.raw SUBJECT_SAMPLE_FACTORS - SG113 Mice strain:SmoA1-Math-GFP | Group:PT DOB=9/10/13; Sex=M; Age (weeks)=34; RAW_FILE_NAME=SG113.raw SUBJECT_SAMPLE_FACTORS - CASE40 Mice strain:SmoA1-Math-GFP | Group:PT DOB=11/20/14; Sex=F; Age (weeks)=18; RAW_FILE_NAME=CASE40.raw SUBJECT_SAMPLE_FACTORS - SGTA13 Mice strain:SmoA1-Math-GFP | Group:PTMT DOB=4/28/14; Sex=F; Age (weeks)=34; RAW_FILE_NAME=SGTA13.raw SUBJECT_SAMPLE_FACTORS - SG228 Mice strain:SmoA1-Math-GFP | Group:PTMT DOB=7/20/14; Sex=M; Age (weeks)=26; RAW_FILE_NAME=SG228.raw SUBJECT_SAMPLE_FACTORS - SGTA12 Mice strain:SmoA1-Math-GFP | Group:PTMT DOB=6/10/14; Sex=F; Age (weeks)=26; RAW_FILE_NAME=SGTA12.raw SUBJECT_SAMPLE_FACTORS - SGTA Mice strain:SmoA1-Math-GFP | Group:PTMT DOB=10/16/13; Sex=F; Age (weeks)=29; RAW_FILE_NAME=SGTA.raw SUBJECT_SAMPLE_FACTORS - SG191 Mice strain:SmoA1-Math-GFP | Group:PTMT DOB=5/13/14; Sex=F; Age (weeks)=13; RAW_FILE_NAME=SG191.raw SUBJECT_SAMPLE_FACTORS - CASE41 Mice strain:SmoA1-Math-GFP | Group:PTMT DOB=8/18/14; Sex=M; Age (weeks)=32; RAW_FILE_NAME=CASE41.raw SUBJECT_SAMPLE_FACTORS - SG222 Mice strain:SmoA1-Math-GFP | Group:PTMT DOB=7/20/14; Sex=F; Age (weeks)=12; RAW_FILE_NAME=SG222.raw #COLLECTION CO:COLLECTION_SUMMARY Mice brain and spinal cord were dissected from each mouse and fluorescence CO:COLLECTION_SUMMARY microscopy was used to visualize the tumors. Mice having a GFP signal in the CO:COLLECTION_SUMMARY cerebellum only were designated as having a primary tumor without metastasis (PT CO:COLLECTION_SUMMARY group, n=18). Mice having a strong GFP signal in the cerebellum and positive GFP CO:COLLECTION_SUMMARY signal in the spinal cord were designated as having a primary tumor with CO:COLLECTION_SUMMARY metastasis (PTMT group, n=7). With aid of GFP visualization, the primary tumor CO:COLLECTION_SUMMARY in each mouse brain was cut from the cerebellum, and immediately frozen using CO:COLLECTION_SUMMARY liquid nitrogen, and stored at -80 °C. CO:SAMPLE_TYPE Brain CO:STORAGE_CONDITIONS -80℃ #TREATMENT TR:TREATMENT_SUMMARY SmoA1-Math1-GFP mice were generated by crossing SmoA1 mice with Math1-driven GFP TR:TREATMENT_SUMMARY reporter mice. All mice were kept under the same condition and typically TR:TREATMENT_SUMMARY developed detectable MB tumors by MR imaging and clinical symptoms around 23 TR:TREATMENT_SUMMARY weeks. #SAMPLEPREP SP:SAMPLEPREP_SUMMARY Frozen mice brain tissue samples were cryo-pulverized with liquid nitrogen and SP:SAMPLEPREP_SUMMARY weighed. 2-propanol (20 uL/mg) was added to the cryo-pulverized tissue samples SP:SAMPLEPREP_SUMMARY to precipitate proteins. Samples were vortex-mixed for 1 min, and centrifuged at SP:SAMPLEPREP_SUMMARY 13,000 rpm for 7 min. Following centrifugation, 100 uL of supernatant was SP:SAMPLEPREP_SUMMARY transferred to an autosampler vial and analyzed with UPLC-MS. A blank sample, SP:SAMPLEPREP_SUMMARY prepared with 100 uL 2-propanol, underwent the same process as the murine tumor SP:SAMPLEPREP_SUMMARY samples. A pooled quality control (QC) sample was prepared by mixing a 10 uL SP:SAMPLEPREP_SUMMARY aliquot of supernatant of each sample. #CHROMATOGRAPHY CH:CHROMATOGRAPHY_TYPE Reversed phase CH:INSTRUMENT_NAME Thermo Dionex Ultimate 3000 CH:COLUMN_NAME Waters Acquity BEH C18 (50 x 2.1mm, 1.7 um) CH:COLUMN_TEMPERATURE 55 CH:SOLVENT_A water/acetonitrile (40:60 v/v), 10 mM ammonium formate and 0.1% formic acid CH:SOLVENT_B acetonitrile/isopropanol (10:90 v/v), 10 mM ammonium formate and 0.1% formic CH:SOLVENT_B acid CH:SAMPLE_INJECTION 5μL #ANALYSIS AN:ANALYSIS_TYPE MS #MS MS:INSTRUMENT_NAME Thermo Q Exactive HF hybrid Orbitrap MS:INSTRUMENT_TYPE Orbitrap MS:MS_TYPE ESI MS:ION_MODE POSITIVE MS:MS_COMMENTS Samples were analyzed in positive and negative ion modes for RP chromatography. MS:MS_COMMENTS Spectral features were extracted from the raw data using Progenesis QI v2.1 MS:MS_COMMENTS software. MS:CAPILLARY_TEMPERATURE 275 °C MS:SPRAY_VOLTAGE 3.0 kV MS:MS_RESULTS_FILE ST001290_AN002143_Results.txt UNITS:Normalized Peak Abundance Has m/z:Yes Has RT:Yes RT units:Minutes #END