Summary of study ST001135

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 PR000760. The data can be accessed directly via it's Project DOI: 10.21228/M8FH6C This work is supported by NIH grant, U2C- DK119886.

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Study IDST001135
Study TitleDifferent dose exposure of OPC-163493 on HepG2 cells (part-I)
Study TypeCompound dosage test
Study SummaryMetabolomics analysis were on 8 samples of HepG2 cells that were treated with compound OPC-163493 (DMSO control, 1, 3, or 10µM; each n=2) exposure for 30 min.
Institute
Otsuka Pharmaceutical Co., Ltd.
Last NameKanemoto
First NameNaohide
Address463-10 Kagasuno Kawauchi-cho Tokushima 771-0192, Japan
EmailKanemoto.Naohide@otsuka.jp
Phone81-03-6717-1400
Submit Date2019-02-07
Analysis Type DetailLC-MS
Release Date2019-03-06
Release Version1
Naohide Kanemoto Naohide Kanemoto
https://dx.doi.org/10.21228/M8FH6C
ftp://www.metabolomicsworkbench.org/Studies/ application/zip

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Project:

Project ID:PR000760
Project DOI:doi: 10.21228/M8FH6C
Project Title:Antidiabetic and cardiovascular beneficial effects of a liver-localized mitochondrial uncoupler
Project Summary:Inducing mitochondrial uncoupling (mUncoupling) is an attractive therapeutic strategy for treating metabolic diseases because it leads to calorie-wasting by reducing the efficiency of oxidative phosphorylation (OXPHOS) in mitochondria. Here we report a safe mUncoupler, OPC-163493, which has unique pharmacokinetic characteristics. OPC-163493 shows a good bioavailability upon oral administration and primarily distributed to specific organs: the liver and kidneys, avoiding systemic toxicities. It exhibitsinsulin-independent antidiabetic effects in multiple animal models of type I and type II diabetes and antisteatotic effects in fatty liver models. These beneficial effects can be explained by the improvement of glucose metabolism and enhancement of energy expenditure by OPC-163493 in the liver. Moreover, OPC-163493 treatment lowered blood pressure, extended survival, and improved renal function in the rat model of stroke/hypertension, possibly by enhancing NO bioavailability in blood vessels and reducing mitochondrial ROS production. OPC-163493 is a liver-localized/targeted mUncoupler that ameliorates various complications of diabetes.
Institute:Otsuka Pharmaceutical Co., Ltd.
Last Name:Kanemoto
First Name:Naohide
Address:463-10 Kagasuno Kawauchi-cho, Tokushima, Tokusima, 770-0865, Japan
Email:Kanemoto.Naohide@otsuka.jp
Phone:+81-88-665-2126

Subject:

Subject ID:SU001196
Subject Type:Cultured cells
Subject Species:Homo sapiens
Taxonomy ID:9606
Gender:Not applicable
Cell Strain Details:HepG2 cells

Factors:

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

mb_sample_id local_sample_id Treatment
SA078229OPC-0uM-5-
SA078230OPC-0uM-1-
SA078231OPC-1uM-21
SA078232OPC-1uM-61
SA078235OPC-10uM-810
SA078236OPC-10uM-410
SA078233OPC-3uM-33
SA078234OPC-3uM-73
Showing results 1 to 8 of 8

Collection:

Collection ID:CO001190
Collection Summary:Culture medium of HepG2 cells in 100-mm dish was aspirated and cells were washed twice by 5% mannitol solution, and then processed with sampleprep steps.
Sample Type:HepG2 cells
Storage Conditions:Room temperature

Treatment:

Treatment ID:TR001211
Treatment Summary:HepG2 cells were seeded into a 100mm dish the day before OPC-163493 treatment. OPC-163493 treatments (DMSO control, 1, 3, or 10uM, each n=2) were performed for 30min in FBS-free DMEM with high glucose (25mM).
Treatment Dose:0uM, 1uM, 3uM, 10uM
Treatment Doseduration:30 min
Cell Media:MEM with Earle`s salts, L-Glutamine and Non-Essiontial Amino Acids, 10% fetal bovine serum, and 1mM sodium pyruvate solution

Sample Preparation:

Sampleprep ID:SP001204
Sampleprep Summary:Culture medium of HepG2 cells in 100-mm dish was aspirated and cells were washed twice by 5% mannitol solution, and then the cells were treated with methanol. The cell extract was treated with Mili-Q water containing internal standard and filtered with 5-kDa cutoff filter. The filtrate was centrifugally concentrated and re-suspended in 50 µL of Milli-Q water.
Processing Storage Conditions:Room temperature
Extract Storage:-80℃
Sample Resuspension:50 uL Mili-Q

Combined analysis:

Analysis ID AN001860 AN001861
Analysis type MS MS
Chromatography type CE CE
Chromatography system Agilent 7100 CE Agilent 7100 CE
Column Fused silica capillary, i.d. 50 μm × 80 cm Fused silica capillary, i.d. 50 μm × 80 cm
MS Type ESI ESI
MS instrument type Other Triple quadrupole
MS instrument name Agilent 6210 TOF Agilent 6460 QQQ
Ion Mode UNSPECIFIED UNSPECIFIED
Units Concentration (pmol/1000000 cells) Concentration (pmol/1000000 cells)

Chromatography:

Chromatography ID:CH001347
Chromatography Summary:capillary electrophoresis was connected with time-of-flight mass spectrometry (CE-TOFMS) for cation analysis and tandem mass spectrometry (CE-MS/MS) for anion.
Instrument Name:Agilent 7100 CE
Column Name:Fused silica capillary, i.d. 50 μm × 80 cm
Chromatography Type:CE

MS:

MS ID:MS001720
Analysis ID:AN001860
Instrument Name:Agilent 6210 TOF
Instrument Type:Other
MS Type:ESI
MS Comments:The spectrometer was scanned from m/z 50 to 1,000. Peaks were extracted using automatic integration software MasterHands (Keio University, Tsuruoka, Japan) in order to obtain peak information including m/z, migration time for CE-TOFMS measurement (MT) and peak area. Signal peaks corresponding to isotopomers, adduct ions, and other product ions of known metabolites were excluded, and remaining peaks were annotated with putative metabolites from the HMT metabolite database based on their MTs and m/z values determined by TOFMS. The tolerance range for the peak annotation was configured at ±0.5 min for MT and ±10 ppm for m/z. In addition, peak areas were normalized against those of the internal standards and then the resultant relative area values were further normalized by sample amount.
Ion Mode:UNSPECIFIED
  
MS ID:MS001721
Analysis ID:AN001861
Instrument Name:Agilent 6460 QQQ
Instrument Type:Triple quadrupole
MS Type:ESI
MS Comments:The spectrometer was scanned from m/z 50 to 1,000. Peaks were extracted using MasterHands, automatic integration software (Keio University, Tsuruoka, Yamagata, Japan) and MassHunter Quantitative Analysis B.04.00 (Agilent Technologies) in order to obtain peak information including m/z, peak area, and migration time (MT). Signal peaks were annotated according to the HMT metabolite database based on their m/z values with the MTs. The peak area of each metabolite was normalized with respect to the area of the internal standard and metabolite concentration was evaluated by standard curves with three-point calibrations using each standard compound.
Ion Mode:UNSPECIFIED
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