Summary of Study ST001024

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 PR000684. The data can be accessed directly via it's Project DOI: 10.21228/M8868G This work is supported by NIH grant, U2C- DK119886.

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Study IDST001024
Study TitleTCA cycle metabolomics of H3K27M cells grown in regular glutamine media, glutamine free media, and glutamine free media with alpha-ketoglutarate (Part-II)
Study SummaryTesting TCA concentrations of Diffuse Intrinsic Pontine Gliomas (DIPG) cellines with H3K27M mutations. Preliminary studies show H3K27M tumor cells are addicted to Gln for survival. Removal of Gln from media resulted in tumor cell death which was rescued by the addition of α-KG. These data show that Gln is taken up and metabolized by H3K27M tumor cells and that Gln derived α-KG is critical for the survival of these tumors. Interestingly, tumor cell death with Gln deprivation was similar to the effect of the JMJD3 inhibitor GSKJ4. Therefore, Gln derived α-KG may be required for both anaplerosis and to drive JMJD3 demethylation. We hypothesize that H3K27M tumors are reliant on α-KG that is derived from Gln to drive the TCA cycle and further decrease H3K27 methylation levels. Furthermore, inhibition of Gln metabolism may represent a novel therapeutic approach for tumors with this mutation. In this study, TCA cycle metabolomics are analyzed of H3K27M cells grown in regular glutamine media, glutamine free media, and glutamine free media with alpha-ketoglutarate.
Institute
Mayo Clinic
Last NameDaniels
First NameDavid
Address200 First Street SW Rochester, MN 55905
Emaildaniels.david@mayo.edu
Phone507-284-2511
Submit Date2018-07-18
Analysis Type DetailGC-MS
Release Date2020-07-15
Release Version1
David Daniels David Daniels
https://dx.doi.org/10.21228/M8868G
ftp://www.metabolomicsworkbench.org/Studies/ application/zip

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Project:

Project ID:PR000684
Project DOI:doi: 10.21228/M8868G
Project Title:Mayo Pilot and Feasibility: H3K27M cells and glutamine metabolomics quatitation studies
Project Summary:In children, tumors affecting the brain and nervous system result in more cancer-related deaths than any other type of tumor. It is thus critical to identify new approaches for therapy. Among pediatric patients, one of the most devastating brain tumor types is Diffuse Intrinsic Pontine Gliomas (DIPG). Our understanding of this deadly disease has recently been advanced by important discoveries, including the discovery that the majority of DIPG tumors harbor the histone H3K27M mutation. This mutation results in global hypomethylation of H3K27 residues and is the pathological hallmark for this disease. Glutamine (Gln) addiction has been reported in many cancers including malignant adult gliomas. Glutamine likely promotes cancer cell proliferation and survival likely through generation of the TCA cycle intermediate alpha-ketoglutarate (α-KG). Importantly, α-KG is a critical co-factor for histone lysine demethylases including JMJD3, the enzyme responsible for removing the methyl groups from H3K27me3. Our preliminary data shows H3K27M tumor cells require Gln for survival, and if Gln is removed from the culture media, cells can be rescued by the addition of α-KG. Furthermore, Gln deprivation leads to an increase in H3K27 trimethylation similar to direct inhibition of JMJD3. It is for these reasons we hypothesize that H3K27M tumors are dependent on Gln derived α-KG both for feeding the TCA cycle and for further decreasing H3K27 trimethylation. Inhibition of Gln metabolism will likely uncover novel therapeutic targets for this deadly disease. In Aim 1 we will study Gln and glucose metabolism in H3K27M tumor cells and compare this to Wild Type (WT) tumors and Embryonic Stem Cells (ESCs). In Aim 2 we will validate the therapeutic validity of inhibiting Gln metabolism in H3K27M tumors.
Institute:Mayo Clinic
Last Name:Daniels
First Name:David
Address:200 First Street SW Rochester, MN 55905
Email:daniels.david@mayo.edu
Phone:507-284-2511

Subject:

Subject ID:SU001063
Subject Type:Human
Subject Species:Homo sapiens
Taxonomy ID:9606

Factors:

Subject type: Human; Subject species: Homo sapiens (Factor headings shown in green)

mb_sample_id local_sample_id Sample type Grouping
SA064320Sample # 1Cells IV Control
SA064321Sample # 2Cells IV Control
SA064322Sample # 3Cells IV Control
SA064323Sample # 7Cells IV Treatment 1
SA064324Sample # 8Cells IV Treatment 1
SA064325Sample # 9Cells IV Treatment 1
SA064326Sample # 14Cells IV Treatment 2
SA064327Sample # 15Cells IV Treatment 2
SA064328Sample # 13Cells IV Treatment 2
SA064329Sample # 20Cells XVII Control
SA064330Sample # 21Cells XVII Control
SA064331Sample # 19Cells XVII Control
SA064332Sample # 27Cells XVII Treatment 1
SA064333Sample # 26Cells XVII Treatment 1
SA064334Sample # 25Cells XVII Treatment 1
SA064335Sample # 32Cells XVII Treatment 2
SA064336Sample # 33Cells XVII Treatment 2
SA064337Sample # 31Cells XVII Treatment 2
SA064338Sample # 5Media IV Control
SA064339Sample # 6Media IV Control
SA064340Sample # 4Media IV Control
SA064341Sample # 12Media IV Treatment 1
SA064342Sample # 11Media IV Treatment 1
SA064343Sample # 10Media IV Treatment 1
SA064344Sample # 18Media IV Treatment 2
SA064345Sample # 17Media IV Treatment 2
SA064346Sample # 16Media IV Treatment 2
SA064347Sample # 22Media XVII Control
SA064348Sample # 24Media XVII Control
SA064349Sample # 23Media XVII Control
SA064350Sample # 28Media XVII Treatment 1
SA064351Sample # 30Media XVII Treatment 1
SA064352Sample # 29Media XVII Treatment 1
SA064353Sample # 36Media XVII Treatment 2
SA064354Sample # 34Media XVII Treatment 2
SA064355Sample # 35Media XVII Treatment 2
Showing results 1 to 36 of 36

Collection:

Collection ID:CO001057
Collection Summary:DIPG IV and DIPG XVII cell lines are collected in this experiment. Susupension cells are harvested using centrifugation at 1200 rpm for 5 min. 1 mL of the supernatant media werr collected in an eppendorf tube and snapped frozen. The cell pellets were broken up into single cell suspension and counted. 1 million cells were taken from the stock and washed 1 x with PBS using table top centrifuge with 10 sec quick spin. The resulting cell pellet was snap frozen. Both the frozen media and cell pellet are stored in -80 C prior transfer.
Sample Type:Glioma cells

Treatment:

Treatment ID:TR001077
Treatment Summary:Glutamine is required for survival in H3K27M cell lines. DIPG cell lines with the H3K27M mutation were grown in regular media (45mM glucose + 4.5 mM glutamine), glutamine (Gln) free media, and Gln free media with alpha-ketoglutarate (α-KG). Study Design Factors IV Control / Treatment 1 / Treatment2 = DIPG IV celline grown in regular media / glutamine free media / glutamine free meida with alpha-ketoglutarate XVII Control / Treatment 1 / Treatment2 = DIPG XVII celline grown in regular media / glutamine free media / glutamine free meida with alpha-ketoglutarate

Sample Preparation:

Sampleprep ID:SP001070
Sampleprep Summary:TCA Concentrations in glioma cell lines

Combined analysis:

Analysis ID AN001681
Analysis type MS
Chromatography type GC
Chromatography system Agilent 7890B
Column Agilent HP5-MS (30m × 0.25mm, 0.25 um)
MS Type EI
MS instrument type Single quadrupole
MS instrument name Agilent 5977A
Ion Mode POSITIVE
Units nmol/vial

Chromatography:

Chromatography ID:CH001182
Instrument Name:Agilent 7890B
Column Name:Agilent HP5-MS (30m × 0.25mm, 0.25 um)
Chromatography Type:GC

MS:

MS ID:MS001556
Analysis ID:AN001681
Instrument Name:Agilent 5977A
Instrument Type:Single quadrupole
MS Type:EI
Ion Mode:POSITIVE
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