Summary of Study ST001921

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 PR001212. The data can be accessed directly via it's Project DOI: 10.21228/M81X3N 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	ST001921
Study Title	An Airway Organoid-Based Screen Identifies a Role for the HIF1α-Glycolysis Axis in SARS-CoV-2 Infection
Study Summary	SARS-CoV-2 causes the COVID-19 pandemic. It is urgent to develop disease models to dissect mechanisms regulating SARS-CoV-2 infection. Here, we derive airway organoids from human pluripotent stem cells (hPSC-AOs). The hPSC-AOs, particularly ciliated-like cells, are permissive to SARS-CoV-2 infection. Using this platform, we perform a high content screen and identify GW6471, which blocks SARS-CoV-2 infection. GW6471 can also block infection of the B.1.351 SARS-CoV-2 variant. RNA-seq analysis suggests that GW6471 blocks SARS-CoV-2 infection at least in part by inhibiting HIF1α, which is further validated by chemical inhibitor and genetic perturbation targeting HIF1α. Metabolic profiling identifies decreased rates of glycolysis upon GW6471 treatment, consistent with transcriptome profiling. Finally, xanthohumol, 5-(Tetradecyloxy)-2-furoic acid, and ND-646, three compounds that suppress fatty acid biosynthesis, also block SARS-CoV-2 infection. Together, a high content screen coupled with transcriptome and metabolic profiling reveals a key role of the HIF1α-glycolysis axis in mediating SARS-CoV-2 infection of human airway epithelium.
Institute	Weill Cornell Medicine
Last Name	Chen
First Name	Shuibing
Address	A 827B, 1300 York Ave
Email	shc2034@med.cornell.edu
Phone	2127465431
Submit Date	2021-09-24
Raw Data Available	Yes
Raw Data File Type(s)	raw(Waters)
Analysis Type Detail	GC-MS
Release Date	2021-10-20
Release Version	1

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Project:

Project ID:	PR001212
Project DOI:	doi: 10.21228/M81X3N
Project Title:	An Airway Organoid-Based Screen Identifies a Role for the HIF1α-Glycolysis Axis in SARS-CoV-2 Infection
Project Type:	MS
Project Summary:	SARS-CoV-2 causes the COVID-19 pandemic. It is urgent to develop disease models to dissect mechanisms regulating SARS-CoV-2 infection. Here, we derive airway organoids from human pluripotent stem cells (hPSC-AOs). The hPSC-AOs, particularly ciliated-like cells, are permissive to SARS-CoV-2 infection. Using this platform, we perform a high content screen and identify GW6471, which blocks SARS-CoV-2 infection. GW6471 can also block infection of the B.1.351 SARS-CoV-2 variant. RNA-seq analysis suggests that GW6471 blocks SARS-CoV-2 infection at least in part by inhibiting HIF1α, which is further validated by chemical inhibitor and genetic perturbation targeting HIF1α. Metabolic profiling identifies decreased rates of glycolysis upon GW6471 treatment, consistent with transcriptome profiling. Finally, xanthohumol, 5-(Tetradecyloxy)-2-furoic acid, and ND-646, three compounds that suppress fatty acid biosynthesis, also block SARS-CoV-2 infection. Together, a high content screen coupled with transcriptome and metabolic profiling reveals a key role of the HIF1α-glycolysis axis in mediating SARS-CoV-2 infection of human airway epithelium.
Institute:	Weill Cornell Medical College
Department:	Surgery
Last Name:	Chen
First Name:	Shuibing
Address:	A 827B, 1300 York Ave, New York, NY, 10065, USA
Email:	shc2034@med.cornell.edu
Phone:	2127465431

Subject:

Subject ID:	SU001999
Subject Type:	Cultured cells
Subject Species:	Homo sapiens
Taxonomy ID:	9606

Factors:

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

mb_sample_id	local_sample_id	Treatment
SA177907	GW+SARS_CoV_2_3	GW+SARS_CoV_2
SA177908	GW+SARS_CoV_2_2	GW+SARS_CoV_2
SA177909	GW+SARS_CoV_2_1	GW+SARS_CoV_2
SA177914	Mock2	mock
SA177915	Mock3	mock
SA177916	Mock4	mock
SA177917	Mock1	mock
SA177910	SARS_CoV_2_4	SARS_CoV_2
SA177911	SARS_CoV_2_1	SARS_CoV_2
SA177912	SARS_CoV_2_3	SARS_CoV_2
SA177913	SARS_CoV_2_2	SARS_CoV_2
SA177918	shHIF1+SARS+CoV-2_2	scramble shRNA+SARS+CoV-2
SA177919	shHIF1+SARS+CoV-2_3	scramble shRNA+SARS+CoV-2
SA177920	scramble shRNA+SARS+CoV-2_3	scramble shRNA+SARS+CoV-2
SA177921	scramble shRNA+SARS+CoV-2_1	scramble shRNA+SARS+CoV-2
SA177922	scramble shRNA+SARS+CoV-2_2	scramble shRNA+SARS+CoV-2
SA177923	shHIF1+SARS+CoV-2_1	scramble shRNA+SARS+CoV-2

Showing results 1 to 17 of 17

Showing results 1 to 17 of 17

Collection:

Collection ID:	CO001992
Collection Summary:	hPSC-AOs were collected into 150 mM ammonium acetate. After centrifugation, cell pellets were collected for metabolite profiling. Metabolite extraction was performed with 200 μl of 80% methanol (20% water) with internal standards (U13C succinate, and U13C citrate, and heptadecanoate).
Sample Type:	Lung organoids

Treatment:

Treatment ID:	TR002011
Treatment Summary:	SARS-CoV-2 variants, including USA-WA1/2020 and B.1.351, were obtained from the World Reference Center for Emerging Viruses and Arboviruses located at the University of Texas, Medical Branch via the CDC. SARS-CoV-2 was propagated in Vero E6 cells (ATCC) in EMEM supplemented with 10% FCS, 1 mM Sodium Pyruvate and 10 mM HEPES. hPSC-AOs were fragmented into small cell clusters and plated on 10% matrigel-coated plates. The infection was performed in the culture media at the indicated MOIs at 37°C. For pre-infection treatment experiments, hPSC-AOs were pretreated with DMSO (control), 10 µM GW6471 for 4 hours prior to infection. HIF1a knockdown in hPSC-AOs Two shRNAs against HIF1α and one scrambled negative control non-effective shRNA in lentiviral GFP vectors were purchased from OriGene company. The shRNA sequences are shown in Table S3. To generate lentivirus particles, shRNA vectors and lentivirus packaging vectors were co-transfected into 293T cells. The day 15 lung progenitor cells were infected with the collected lentivirus (MOI=0.5) with 8 μg/ml polybrene. To increase the infection efficiency, the cells were centrifuged at 2300 rpm for 1 hour at 30℃. At 24 hpi, the cells were selected with 1 μg/ml puromycin for an additional 72 hours. After selection, the cells were dissociated into single cells and seeded into 24 well plates at 300-400 cells/μl for 3D organoid differentiation.

Treatment ID:

TR002011

Treatment Summary:

SARS-CoV-2 variants, including USA-WA1/2020 and B.1.351, were obtained from the World Reference Center for Emerging Viruses and Arboviruses located at the University of Texas, Medical Branch via the CDC. SARS-CoV-2 was propagated in Vero E6 cells (ATCC) in EMEM supplemented with 10% FCS, 1 mM Sodium Pyruvate and 10 mM HEPES. hPSC-AOs were fragmented into small cell clusters and plated on 10% matrigel-coated plates. The infection was performed in the culture media at the indicated MOIs at 37°C. For pre-infection treatment experiments, hPSC-AOs were pretreated with DMSO (control), 10 µM GW6471 for 4 hours prior to infection. HIF1a knockdown in hPSC-AOs Two shRNAs against HIF1α and one scrambled negative control non-effective shRNA in lentiviral GFP vectors were purchased from OriGene company. The shRNA sequences are shown in Table S3. To generate lentivirus particles, shRNA vectors and lentivirus packaging vectors were co-transfected into 293T cells. The day 15 lung progenitor cells were infected with the collected lentivirus (MOI=0.5) with 8 μg/ml polybrene. To increase the infection efficiency, the cells were centrifuged at 2300 rpm for 1 hour at 30℃. At 24 hpi, the cells were selected with 1 μg/ml puromycin for an additional 72 hours. After selection, the cells were dissociated into single cells and seeded into 24 well plates at 300-400 cells/μl for 3D organoid differentiation.

Sample Preparation:

Sampleprep ID:	SP002005
Sampleprep Summary:	In brief, the samples were methoximized with 50 µl of methoxyamine hydrochloride (MOA, 15 mg/mL in pyridine) at 30°C for 90 minutes. The silylation step was done with 50 µl of N,O-Bis(trimethylsilyl)trifluoroacetamide (BSTFA, containing 1% TMCS, Supelco, PA, USA) at 70 °C for 60 minutes. After the samples cooled to room temperature, they were analyzed by GC-TOF/MS. Separation was performed with a 30-meter DB-5MS column coupled with 10-meter guard column. Helium was used as carrier gas at a flow of 1 ml/min. The oven program is as the following: started at 60 °C for 1 minute, 10°C /min to 320 minutes and kept for 3 minutes.

Sampleprep ID:

SP002005

Sampleprep Summary:

In brief, the samples were methoximized with 50 µl of methoxyamine hydrochloride (MOA, 15 mg/mL in pyridine) at 30°C for 90 minutes. The silylation step was done with 50 µl of N,O-Bis(trimethylsilyl)trifluoroacetamide (BSTFA, containing 1% TMCS, Supelco, PA, USA) at 70 °C for 60 minutes. After the samples cooled to room temperature, they were analyzed by GC-TOF/MS. Separation was performed with a 30-meter DB-5MS column coupled with 10-meter guard column. Helium was used as carrier gas at a flow of 1 ml/min. The oven program is as the following: started at 60 °C for 1 minute, 10°C /min to 320 minutes and kept for 3 minutes.

Combined analysis:

Analysis ID	AN003122
Analysis type	MS
Chromatography type	GC
Chromatography system	Agilent 7890A
Column	Agilent DB5-MS (30m)
MS Type	EI
MS instrument type	GC-TOF
MS instrument name	Unspecified
Ion Mode	POSITIVE
Units	Area

Chromatography:

Chromatography ID:	CH002307
Chromatography Summary:	Separation was performed with a 30-meter DB-5MS column coupled with 10-meter guard column. Helium was used as carrier gas at a flow of 1 ml/min. The oven program is as the following: started at 60 °C for 1 minute, 10°C /min to 320 minutes and kept for 3 minutes.
Instrument Name:	Agilent 7890A
Column Name:	Agilent DB5-MS (30m)
Chromatography Type:	GC

MS:

MS ID:	MS002903
Analysis ID:	AN003122
Instrument Name:	Unspecified
Instrument Type:	GC-TOF
MS Type:	EI
MS Comments:	The transfer line temperature was set to 250 °C. The data was acquired with MassLynx software (Waters). The data was collected with the scan time of 0.3 s and interscan delay of 0.1 s. The mass range was set to 35 to 650 for EI. Raw data files (.raw) generated from GC-TOF/MS were analyzed in Refiner MS (Genedata Expressionist, Basel, Switzerland) software. The output from Refiner MS contains retention time, quantification mass, and peak volume for each metabolite.
Ion Mode:	POSITIVE