Summary of Study ST003819
This data is available at the NIH Common Fund's National Metabolomics Data Repository (NMDR) website, the Metabolomics Workbench, https://www.metabolomicsworkbench.org, where it has been assigned Project ID PR002390. The data can be accessed directly via it's Project DOI: 10.21228/M8Q250 This work is supported by NIH grant, U2C- DK119886.
See: https://www.metabolomicsworkbench.org/about/howtocite.php
This study contains a large results data set and is not available in the mwTab file. It is only available for download via FTP as data file(s) here.
| Study ID | ST003819 |
| Study Title | Polar metabolite profiling of SLC31A1 knockout cells compared to intergenic sgRNA containing cells |
| Study Summary | To investigate the effects of SLC31A1 knockout on steady state metabolite levels, SEM (acute lymphoblastic leukaemia cell line) cells containing either a SLC31A1-targeting sgRNA (two different guides, guide #2 or guide #4) or a intergenic -targetting sgRNA (i.e. a control sgRNA, "Intergenic1") and Cas9 were FACS-purified (Fluorescence Activated Cell Sorting) based on SLC31A1 expression. Cells were then cultured in RPMI-1640 containing 10% FBS and penicillin and streptomycin in 6-well plates, and then collected for metabolomics. SLC31A1 knockout cells were found to exhibit profound electron transport chain inhibition, resulting in accumulation of carbamoyl aspartic acid/ureidosuccinic acid, and depletion of aspartate, as well as broad changes in nucleotide mono, di and triphosphate levels. |
| Institute | Boston Childrens Hospital |
| Last Name | Wong |
| First Name | Alan |
| Address | 300 Longwood Avenue |
| alan.wong@childrens.harvard.edu | |
| Phone | (617) 355-7433 |
| Submit Date | 2024-10-07 |
| Num Groups | 4 |
| Raw Data Available | Yes |
| Raw Data File Type(s) | mzML, raw(Thermo) |
| Analysis Type Detail | LC-MS |
| Release Date | 2025-12-22 |
| Release Version | 1 |
Select appropriate tab below to view additional metadata details:
Project:
| Project ID: | PR002390 |
| Project DOI: | doi: 10.21228/M8Q250 |
| Project Title: | In vivo CRISPR screen identifies copper metabolism as a vulnerability in acute lymphoblastic leukemia |
| Project Summary: | The nutrient-sparse cerebrospinal fluid (CSF) poses a significant challenge to spreading cancer cells. Despite this challenge, leukemia often spreads to the CSF and represents a significant clinical complication. To uncover nutritional dependencies of leukemia cells in the CSF that could be targeted therapeutically, we conducted an in vivo targeted CRISPR screen in a xenograft model of leukemia. We found that SLC31A1, the primary cell surface copper importer, is a genetic dependency of leukemia in both the central nervous system as well as in the hematopoietic organs. Perturbation of copper metabolism leads to complex IV deficiency, perturbed nucleotide metabolism and slowed leukemia cell proliferation. Furthermore, nutritional copper depletion reduced cancer progression in cell line based and patient-derived xenograft models of leukemia. Copper thus appears to be an actionable micronutrient in leukemia. |
| Institute: | Boston Children's Hospital |
| Department: | Pathology |
| Laboratory: | Naama Kanarek |
| Last Name: | Wong |
| First Name: | Alan |
| Address: | 300 Longwood Avenue, Boston, MA, 02115, USA |
| Email: | alan.wong@childrens.harvard.edu |
| Phone: | (617) 355-7433 |
| Funding Source: | NCI 1R01CA282477-01A1 |
Subject:
| Subject ID: | SU003953 |
| Subject Type: | Cultured cells |
| Subject Species: | Homo sapiens |
| Taxonomy ID: | 9606 |
| Cell Strain Details: | SEM leukemia cells |
| Subject ID: | SU004575 |
| Subject Type: | Cultured cells |
| Subject Species: | Homo sapiens |
| Taxonomy ID: | 9606 |
| Cell Strain Details: | SEM leukemia cells |
Factors:
Subject type: Cultured cells; Subject species: Homo sapiens (Factor headings shown in green)
| mb_sample_id | local_sample_id | gRNA | |
|---|---|---|---|
| SA418828 | 20240119_QE2_HILIC_SLC31A1ko_metabolomics_AYW1058 | Intergenic1 | |
| SA418829 | 20240119_QE2_HILIC_SLC31A1ko_metabolomics_AYW1059 | Intergenic1 | |
| SA418830 | 20240119_QE2_HILIC_SLC31A1ko_metabolomics_AYW1060 | Intergenic1 | |
| SA418831 | 20240119_QE2_HILIC_SLC31A1ko_metabolomics_AYW1064 | Intergenic1 | |
| SA418832 | 20240119_QE2_HILIC_SLC31A1ko_metabolomics_AYW1065 | Intergenic1 | |
| SA418833 | 20240119_QE2_HILIC_SLC31A1ko_metabolomics_AYW1066 | Intergenic1 | |
| SA418834 | 20240119_QE2_HILIC_SLC31A1ko_metabolomics_AYW1061 | sgSLC31A1-#2 | |
| SA418835 | 20240119_QE2_HILIC_SLC31A1ko_metabolomics_AYW1062 | sgSLC31A1-#2 | |
| SA418836 | 20240119_QE2_HILIC_SLC31A1ko_metabolomics_AYW1063 | sgSLC31A1-#2 | |
| SA418837 | 20240119_QE2_HILIC_SLC31A1ko_metabolomics_AYW1067 | sgSLC31A1-#4 | |
| SA418838 | 20240119_QE2_HILIC_SLC31A1ko_metabolomics_AYW1068 | sgSLC31A1-#4 | |
| SA418839 | 20240119_QE2_HILIC_SLC31A1ko_metabolomics_AYW1069 | sgSLC31A1-#4 | |
| SA522274 | 20240119_QE2_HILIC_SLC31A1ko_metabolomics_AYW1058 | SEM leukemia cells | Intergenic1 |
| SA522275 | 20240119_QE2_HILIC_SLC31A1ko_metabolomics_AYW1059 | SEM leukemia cells | Intergenic1 |
| SA522276 | 20240119_QE2_HILIC_SLC31A1ko_metabolomics_AYW1060 | SEM leukemia cells | Intergenic1 |
| SA522277 | 20240119_QE2_HILIC_SLC31A1ko_metabolomics_AYW1064 | SEM leukemia cells | Intergenic1 |
| SA522278 | 20240119_QE2_HILIC_SLC31A1ko_metabolomics_AYW1065 | SEM leukemia cells | Intergenic1 |
| SA522279 | 20240119_QE2_HILIC_SLC31A1ko_metabolomics_AYW1066 | SEM leukemia cells | Intergenic1 |
| SA522280 | 20240119_QE2_HILIC_SLC31A1ko_metabolomics_AYW1061 | SEM leukemia cells | sgSLC31A1-#2 |
| SA522281 | 20240119_QE2_HILIC_SLC31A1ko_metabolomics_AYW1062 | SEM leukemia cells | sgSLC31A1-#2 |
| SA522282 | 20240119_QE2_HILIC_SLC31A1ko_metabolomics_AYW1063 | SEM leukemia cells | sgSLC31A1-#2 |
| SA522283 | 20240119_QE2_HILIC_SLC31A1ko_metabolomics_AYW1067 | SEM leukemia cells | sgSLC31A1-#4 |
| SA522284 | 20240119_QE2_HILIC_SLC31A1ko_metabolomics_AYW1068 | SEM leukemia cells | sgSLC31A1-#4 |
| SA522285 | 20240119_QE2_HILIC_SLC31A1ko_metabolomics_AYW1069 | SEM leukemia cells | sgSLC31A1-#4 |
| Showing results 1 to 24 of 24 |
Collection:
| Collection ID: | CO003946 |
| Collection Summary: | One million cells from culture were collected via centrifugation for 20 seconds at 18,000xG at 4C, washed once with ice-cold 0.9% NaCl, and collected via centrifugation for 20 seconds at 18,000xG at 4C. Cells were cultured in RPMI-1640 with 10% FBS and penicillin/streptomycin in a 37C incubator with 5% CO2. |
| Sample Type: | Leukemia cells |
| Collection ID: | CO004568 |
| Collection Summary: | One million cells from culture were collected via centrifugation for 20 seconds at 18,000xG at 4°C, washed once with ice-cold 0.9% NaCl, and collected via centrifugation for 20 seconds at 18,000xG at 4°C. Cells were cultured in RPMI-1640 with 10% FBS and penicillin/streptomycin in a 37°C incubator with 5% CO2. |
| Sample Type: | Leukemia cells |
Treatment:
| Treatment ID: | TR003962 |
| Treatment Summary: | Cells harboring control or SLC31A1-targetting guideRNAs were treated with doxycycline at 2ug/mL for 3 days, then FACS purified for SLC31A1-hi (in the case of control guideRNA) or SLC31A1-lo (in the case of the SLC31A1-targetting guideRNA). After sorting, cells were expanded in vitro. |
| Treatment ID: | TR004584 |
| Treatment Summary: | Cells harboring control or SLC31A1-targetting guideRNAs were treated with doxycycline at 2ug/mL for 3 days, then FACS purified for SLC31A1-hi (in the case of control guideRNA) or SLC31A1-lo (in the case of the SLC31A1-targetting guideRNA). After sorting, cells were expanded in vitro. |
Sample Preparation:
| Sampleprep ID: | SP003959 |
| Sampleprep Summary: | Cell pellet was resuspended in 400uL of 100% LC-MS grade methanol supplemented with isotopically-labelled amino acid standards [Cambridge Isotope Laboratories, MSK-A2-1.2], aminopterin, and reduced glutathione standard [Cambridge Isotope Laboratories, CNLM-6245-10]). with repeated pipetting up and down and vortexing for 10 seconds. Then, 100uL of LCMS-grade water containing 125 mM Ammonium Acetate, 10 mM Na-Ascorbate, and 7.9 mg/mL 5,5-dithio-bis-(2-nitrobenzoic acid (Ellman's reagent) was added, and the sample vortex for another 10 seconds. Samples were then centrifuged for 10 minutes at 18,000 g to pellet cell debris. The supernatant was transferred to a new tube and dried on ice using a liquid nitrogen dryer. |
| Sampleprep ID: | SP004581 |
| Sampleprep Summary: | Cell pellet was resuspended in 400uL of 100% LC-MS grade methanol supplemented with isotopically-labelled amino acid standards [Cambridge Isotope Laboratories, MSK-A2-1.2], aminopterin, and reduced glutathione standard [Cambridge Isotope Laboratories, CNLM-6245-10]). with repeated pipetting up and down and vortexing for 10 seconds. Then, 100uL of LCMS-grade water containing 125 mM Ammonium Acetate, 10 mM Na-Ascorbate, and 7.9 mg/mL 5,5-dithio-bis-(2-nitrobenzoic acid (Ellman's reagent) was added, and the sample vortex for another 10 seconds. Samples were then centrifuged for 10 minutes at 18,000 g to pellet cell debris. The supernatant was transferred to a new tube and dried on ice using a liquid nitrogen dryer. |
Combined analysis:
| Analysis ID | AN006279 | AN007387 |
|---|---|---|
| Chromatography ID | CH004762 | CH005595 |
| MS ID | MS005981 | MS007080 |
| Analysis type | MS | MS |
| Chromatography type | HILIC | HILIC |
| Chromatography system | Thermo Vanquish | Thermo Vanquish |
| Column | Merck SeQuant ZIC-HILIC (150 x 2.1mm,5um) | Merck SeQuant ZIC-HILIC (150 x 2.1 mm, 5 μm) |
| MS Type | ESI | ESI |
| MS instrument type | Orbitrap | Orbitrap |
| MS instrument name | Thermo Q Exactive Orbitrap | Thermo Q Exactive Orbitrap |
| Ion Mode | UNSPECIFIED | UNSPECIFIED |
| Units | Normalized peak area | Normalized peak area |
Chromatography:
| Chromatography ID: | CH004762 |
| Chromatography Summary: | 2 μL of each sample was injected into a ZIC-pHILIC 150 x 2.1 mm (5 μm particle size) column (EMD Millipore) operated on a Vanquish™ Flex UHPLC system (Thermo Fisher Scientific). Chromatographic separation was achieved using the following conditions: buffer A was acetonitrile; buffer B was 20 mM ammonium carbonate, 0.1% ammonium hydroxide in water; resulting pH is around 9 without pH adjustment. Gradient conditions used were: 0-20 min: linear gradient from 20% to 80% B; 20-24 min: hold at 80% B; 24-24.1 min: from 80% to 20% B; 24.1-32 min: hold at 20% B at 0.150 mL/min flow rate. The column oven and autosampler tray were held at 25 °C and 4 °C, respectively. |
| Instrument Name: | Thermo Vanquish |
| Column Name: | Merck SeQuant ZIC-HILIC (150 x 2.1mm,5um) |
| Column Temperature: | 25 |
| Flow Gradient: | Gradient conditions were as follows: 0-20min: linear gradient from 20 to 80% B; 20–20.5 min: from 80 to 20% B; 20.5–28 min: hold at 20% B. |
| Flow Rate: | 0.15 mL/min |
| Solvent A: | 100% acetonitrile |
| Solvent B: | 100% water; 20mM ammonium carbonate; 0.1% ammonium hydroxide |
| Chromatography Type: | HILIC |
| Chromatography ID: | CH005595 |
| Chromatography Summary: | 2 μL of each sample was injected into a ZIC-pHILIC 150 x 2.1 mm (5 μm particle size) column (EMD Millipore) operated on a Vanquish™ Flex UHPLC system (Thermo Fisher Scientific). Chromatographic separation was achieved using the following conditions: buffer A was acetonitrile; buffer B was 20 mM ammonium carbonate, 0.1% ammonium hydroxide in water; resulting pH is around 9 without pH adjustment. Gradient conditions used were: 0-20 min: linear gradient from 20% to 80% B; 20-24 min: hold at 80% B; 24-24.1 min: from 80% to 20% B; 24.1-32 min: hold at 20% B at 0.150 mL/min flow rate. The column oven and autosampler tray were held at 25°C and 4°C, respectively. |
| Instrument Name: | Thermo Vanquish |
| Column Name: | Merck SeQuant ZIC-HILIC (150 x 2.1 mm, 5 μm) |
| Column Temperature: | 25°C |
| Flow Gradient: | Gradient conditions were as follows: 0-20min: linear gradient from 20 to 80% B; 20–20.5 min: from 80 to 20% B; 20.5–28 min: hold at 20% B. |
| Flow Rate: | 0.15 mL/min |
| Solvent A: | 100% Acetonitrile |
| Solvent B: | 100% Water; 20mM ammonium carbonate; 0.1% ammonium hydroxide |
| Chromatography Type: | HILIC |
MS:
| MS ID: | MS005981 |
| Analysis ID: | AN006279 |
| Instrument Name: | Thermo Q Exactive Orbitrap |
| Instrument Type: | Orbitrap |
| MS Type: | ESI |
| MS Comments: | MS data acquisition was performed using a QExactive benchtop orbitrap mass spectrometer equipped with an Ion Max source and a HESI II probe (Thermo Fisher Scientific, San Jose, CA, USA) and was performed with polarity switching. Scans used were in the positive and negative ionization mode in a range of m/z = 70–1000, with the resolution set at 70,000, the AGC target at 1 × 10^6, and the maximum injection time (Max IT) at 40 msec. |
| Ion Mode: | UNSPECIFIED |
| MS ID: | MS007080 |
| Analysis ID: | AN007387 |
| Instrument Name: | Thermo Q Exactive Orbitrap |
| Instrument Type: | Orbitrap |
| MS Type: | ESI |
| MS Comments: | MS data acquisition was performed using a QExactive benchtop orbitrap mass spectrometer equipped with an Ion Max source and a HESI II probe (Thermo Fisher Scientific, San Jose, CA, USA) and was performed with polarity switching. Scans used were in the positive and negative ionization mode in a range of m/z = 70–1000, with the resolution set at 70,000, the AGC target at 1 × 10^6, and the maximum injection time (Max IT) at 40 msec. |
| Ion Mode: | UNSPECIFIED |
