idr0094-study.txt

# FILL IN AS MUCH INFORMATION AS YOU CAN.  HINTS HAVE BEEN PUT IN SOME FIELDS AFTER THE HASH # SYMBOL. REPLACE THE HINT WITH TEXT WHERE APPROPRIATE.																
# STUDY DESCRIPTION SECTION							
# Section with generic information about the study including title, description, publication details (if applicable) and contact details																														

Comment[IDR Study Accession]	idr0094																			

Study Title	SARS-COV-2 drug repurposing - Caco2 cell line																			

Study Type	high content screen of cells treated with a compound library	infection						

Study Type Term Source REF	EFO	EFO																			

Study Type Term Accession	EFO_0007553	EFO_0000544																	

Study Description	To identify possible candidates for progression towards clinical studies against SARS-CoV-2, we screened a well-defined collection of 5632 compounds including 3488 compounds which have undergone clinical investigations (marketed drugs, phases 1 -3, and withdrawn) across 600 indications. Compounds were screened for their inhibition of viral induced cytotoxicity using the human epithelial colorectal adenocarcinoma cell line Caco-2 and a SARS-CoV-2 isolate. The primary screen of 5632 compounds gave 271 hits. A total of 64 compounds with IC50 <20 μM were identified, including 19 compounds with IC50 < 1 μM. Of this confirmed hit population, 90% have not yet been previously reported as active against SARS-CoV-2 in-vitro cell assays. Some 37 of the actives are launched drugs, 19 are in phases 1-3 and 10 pre-clinical. Several inhibitors were associated with modulation of host pathways including kinase signaling P53 activation, ubiquitin pathways and PDE activity modulation, with long chain acyl transferases were effective viral inhibitors.																			

Study Key Words	SARS-CoV-2	compound screening	inhibition of viral induced cytotoxicity	human epithelial colorectal adenocarcinoma cell line Caco-2																			

Study Organism	Homo sapiens	Severe acute respiratory syndrome coronavirus 2						

Study Organism Term Source REF	NCBITaxon	NCBITaxon																			

Study Organism Term Accession	9606	2697049																				

Study Screens Number	2																			

Study External URL	


Study BioImage Archive Accession	S-BIAD29									

Study Public Release Date	2020-10-06							
																																	
# Study Publication																				

Study PubMed ID	33637768																			

Study Publication Title	A SARS-CoV-2 cytopathicity dataset generated by high-content screening of a large drug repurposing collection																			

Study Author List	Ellinger B, Bojkova D, Zaliani A, Cinatl J, Claussen C, Westhaus S, Keminer O, Reinshagen J, Kuzikov M, Wolf M, Geisslinger G, Gribbon P, Ciesek S																			

Study PMC ID	PMC7910569 																									

Study DOI	https://doi.org/10.1038/s41597-021-00848-4							
																																	
# Study Contacts																				

Study Person Last Name	Ellinger																			

Study Person First Name	Bernhard																			

Study Person Email	bernhard.ellinger@ime.fraunhofer.de																			

Study Person Address	Schnackenburgallee 114, 22525 Hamburg, Germany						

Study Person ORCID	0000-0001-5703-3901																			

Study Person Roles	submitter						
																																	
# Study License and Data DOI																				

Study License	CC0 1.0																			

Study License URL	https://creativecommons.org/publicdomain/zero/1.0/						

Study Copyright	Ellinger et al.																			

Study Data Publisher	University of Dundee																			

Study Data DOI	https://doi.org/10.17867/10000148	
																													

Term Source Name	NCBITaxon	EFO	CMPO	Fbbi																													

Term Source File	http://purl.obolibrary.org/obo/	http://www.ebi.ac.uk/efo/	http://www.ebi.ac.uk/cmpo/	http://purl.obolibrary.org/obo/																
																																	

# SCREEN SECTION							

# Screen Section containing all information relative to each screen in the study including materials used, protocols names and description, phenotype names and description. 	
# For multiple screens this section should be repeated.  Copy and paste the whole section below and fill out for the next screen.																		

Screen Number	1																			

Comment[IDR Screen Name]	idr0094-ellinger-sarscov2/screenA		


Screen Data DOI	https://doi.org/10.17867/10000148a


Screen Sample Type	cell																		

Screen Description	Primary Screen																			

Screen Size	Plates: 	96 5D Images: 	96x9 fields	Planes: 	1 Average Image Dimension (XYZCT): 	1080x1080x1x1x1	Total	Total Tb: 	0.3 TB											

Screen Example Images																			

Screen Imaging Method	spinning disk confocal microscopy	phase contrast microscopy						

Screen Imaging Method Term Source REF	Fbbi	Fbbi																			

Screen Imaging Method Term Accession	FBbi_00000253	FBbi_00000247						

Screen Technology Type	compound screen																			

Screen Technology Type Term Source REF	EFO																			

Screen Technology Type Term Accession	EFO_0007553																			

Screen Type	primary assay																			

Screen Type Term Source REF	BAO																			

Screen Type Term Accession	BAO_0000031					


Screen Comments							
																																	

# Library section. The library file should be supplied separately and it should contain  the reagents description including, at the absolute minimum: reagent ID, sequences and position in the layout (= plate + position in the plate)																				

Library File Name	idr0000-PrimaryScreen-DPC-library.txt																			

Library File Format	tab-delimited text																			

Library Type	compound library																			

Library Type Term Source REF	EFO																			

Library Type Term Accession	EFO_0007569																		

Library Manufacturer	Specs, specs.net																			

Library Version	1.0																			

Library Experimental Conditions	Cytotoxicity Assay BAO http://www.bioassayontology.org/bao#BAO_0000090 or http://www.bioassayontology.org/bao#BAO_0002993																			

Library Experimental Conditions Term Source REF	EFO																			

Library Experimental Conditions Term Accession																				

Quality Control Description	row 23 virus only + DMSO, row 24 DMSO, quality control was based on z' 																																		

# Protocols																																	

Protocol Name	growth protocol	treatment protocol	HCS image acquisition and feature extraction protocol	HCS data analysis protocol																

Protocol Type	growth protocol	treatment protocol	HCS image acquisition and feature extraction protocol	HCS data analysis protocol																

Protocol Type Term Source REF	EFO	EFO	EFO	EFO																

Protocol Type Term Accession	EFO_0003789	EFO_0007571	EFO_0007572	EFO_0007573			

Protocol Description	Cell culture: Caco-2 cells (Human epithelial cell line derived from colon carcinoma) were obtained from the Deutsche Sammlung von Mikroorganismen und Zellkulturen (DSMZ). Cells were grown in Minimal Essential Medium (MEM) supplemented with 10% fetal bovine serum (FBS) and containing 100 IU/ml penicillin and 100 μg/ml streptomycin at 37 °C. All culture reagents were purchased from Sigma. Virus culture: SARS-CoV-2 was isolated from samples of travelers returning from Wuhan (China) to Frankfurt (Germany) using Caco-2 cells. SARS-CoV-2 stocks used in the experiments had undergone one passage on Caco-2 cells and were stored at -80° C. Virus titers were determined as TCID50/ml in confluent cells in 96-well microtiter plates.	Compounds were added to confluent layers of Caco–2 cells in MEM supplemented with 1% FBS in 96-well plates. For the primary screen final compound concentration was 10 μM (0.1 % DMSO final) in singlicate. Following the addition of compounds, cells were immediately infected with SARS-CoV-2 at MOI 0.01. Control wells (+ virus and - virus) also contained DMSO at 0.1 % DMSO final. After 48 hours, cells were fixed using 3 % PFA in PBS, and the plates sealed and disinfected to inactivate SARS-CoV-2	The collection of 5641 compounds was assembled by an external partner (SPECS) in a manner aligned to the recommendations from the Broad Institute (Cambridge Mass., USA). In assembling this “mirror” collection, compounds were purchased from the same set of >70 high quality suppliers identified by the Broad, (personal communication Dr Joshua Bitker), and quality controlled by LC/MS for purity and identity (minimum purity > 90%). Compounds were stored at 10mM in 100% DMSO at -20 °C. A curated database is available containing the compound, indication, primary target (where known) and mechanism of action, as well as analysis tools which can assist in mechanism of action determination and target elucidation.	Quantification of viral inhibition (based upon Caco-2 cell viability relative to controls) was performed using high content imaging of unlabeled cells. For high content imaging an Operetta CLS microscope from PerkinElmer was used. Cells were analysed in label free mode using digital phase contrast, with maximum contrast as a read out. Images were acquired using 10x objective with nine imaged fields per well and analysed using the manufacturer's software (PerkinElmer, Columbus v.2.9.0.1546).	The analysis sequence started with cell detection (method: c, common threshold: 0.05, area >100 μm, splitting coefficient: 6.5, individual threshold: 0.05, contrast >0.05) and was followed by calculating morphology, intensity and position properties as well as cell confluence. Well level data were analysed using ActivityBase (IDBS) and R (v.3.6.1). Test well results were normalized relative to the corresponding intra-plate control (no virus assigned as 100% inhibition of cellular toxicity, with virus assigned as 0% inhibition of cellular toxicity). Outliers were eliminated according to 3-sigma method. Plate level statistical performance was assessed using the standard Z’ calculation5. Initial hit criteria was an inhibition of virus induced cellular toxicity of 75%.																																												

# Phenotypes																				

Phenotype Name	number of cells																			

Phenotype Description	count number of cells and compare relative to control						

Phenotype Score Type	automatic																			

Phenotype Term Source REF	CMPO																			

Phenotype Term Name	increased cell viability in population  						

Phenotype Term Accession	http://www.ebi.ac.uk/cmpo/CMPO_0000161	
																																	

# Raw Data Files																																	

Raw Image Data Format	https://docs.openmicroscopy.org/bio-formats/6.4.0/formats/perkinelmer-operetta.html										

Raw Image Organization	64 plates, 96 wells, 9 fields						
													
																				

# Feature Level Data Files																				

Feature Level Data File Name																				

Feature Level Data File Description																				

Feature Level Data File Format																				

Feature Level Data Column Name																				

Feature Level Data Column Description							

																																	

#  Processed Data Files 																				

Processed Data File Name	idr0000-PrimaryScreen-DPC-processed.txt						

Processed Data File Format	tab-delimited text																			

Processed Data File Description	absolute number of cells per well, relative number of cells per well based on plate control																			

Processed Data Column Name	Plate	Well	CompoundName	SMILES	IUPAC_NAME	pubchem_cid	unichem_link	broad_ids% Inhibition (DPC)	Compound a hit ? (over 75% activity)	Compound a hit ? (over 75% activity)	Phenoptye Annotation Level	NumberOfCells (DPC)								

Processed Data Column Type	plate	well	reagent identifier	reagent identifier	reagent identifier	reagent identifier	reagent identifier	reagent identifier	data	phenotype	phenotype	phenotype	phenotype																				

Processed Data Column Annotation Level									well	well	well	well	well																				

Processed Data Column Description									cell viability compared control				number of cells													

Processed Data Column Link To Library File	Link to Library file	Link to Library file	Link to Library file	Link to Library file	Link to Library file	Link to Library file	Link to Library file	Link to Library file																
																																	

Screen Number	2																			

Comment[IDR Screen Name]	idr0094-ellinger-sarscov2/screenB	


Screen Data DOI	https://doi.org/10.17867/10000148b


Screen Sample Type	cell																		

Screen Description	Dose response screen																			

Screen Size	Plates: 	5D Images: 	Planes: 	Average Image Dimension (XYZCT): 	Total Tb: 	0.1														

Screen Example Images																				

Screen Imaging Method	phase contrast microscopy																			

Screen Imaging Method Term Source REF	Fbbi																			

Screen Imaging Method Term Accession	FBbi_00000247									

Screen Technology Type	compound screen																			

Screen Technology Type Term Source REF	EFO																			

Screen Technology Type Term Accession	EFO_0007553		


Screen Type	multiple concentration																			

Screen Type Term Source REF	BAO									


Screen Type Term Accession	BAO_0000535				


Screen Comments																																	
																																	

# Library section. The library file should be supplied separately and it should contain  the reagents description including, at the absolute minimum: reagent ID, sequences and position in the layout (= plate + position in the plate)																				

Library File Name	idr0000-HitProfiling-DPC-library.txt																			

Library File Format	tab-delimited text																			

Library Type	compound library																			

Library Type Term Source REF	EFO																			

Library Type Term Accession	# leave blank																			

Library Manufacturer	Specs, specs.net																			

Library Version	1.0																			

Library Experimental Conditions	Cytotoxicity Assay BAO http://www.bioassayontology.org/bao#BAO_0000090 or http://www.bioassayontology.org/bao#BAO_0002993																			

Library Experimental Conditions Term Source REF	EFO																			

Library Experimental Conditions Term Accession	# leave blank																			

Quality Control Description	row 23 virus only + DMSO, row 24 DMSO, quality control was based on z' 																																
																																	
# Protocols																																	
Protocol Name	growth protocol	treatment protocol	HCS library protocol	HCS image acquistion and feature extraction protocol	HCS data analysis protocol																												
Protocol Type	growth protocol	treatment protocol	HCS library protocol	HCS image acquistion and feature extraction protocol	HCS data analysis protocol																												
Protocol Type Term Source REF	EFO	EFO	EFO	EFO	EFO																												
Protocol Type Term Accession	EFO_0003789	EFO_0003969	EFO_0007571	EFO_0007572	EFO_0007573																												
Protocol Description	Cell culture: Caco-2 cells (Human epithelial cell line derived from colon carcinoma) were obtained from the Deutsche Sammlung von Mikroorganismen und Zellkulturen (DSMZ). Cells were grown in Minimal Essential Medium (MEM) supplemented with 10% fetal bovine serum (FBS) and containing 100 IU/ml penicillin and 100 μg/ml streptomycin at 37 °C. All culture reagents were purchased from Sigma. Virus culture: SARS-CoV-2 was isolated from samples of travelers returning from Wuhan (China) to Frankfurt (Germany) using Caco-2 cells. SARS-CoV-2 stocks used in the experiments had undergone one passage on Caco-2 cells and were stored at -80° C. Virus titers were determined as TCID50/ml in confluent cells in 96-well microtiter plates.	Compounds were added to confluent layers of Caco–2 cells in MEM supplemented with 1% FBS in 96-well plates. Dose response profiling of selected priority compounds was performed with a range of eight different concentrations in three independent replicates (maximum 20 μM, minimum 20 nM, half log dilution factor, 0.1 % DMSO final). Following the addition of compounds, cells were immediately infected with SARS-CoV-2 at MOI 0.01. Control wells (+ virus and - virus) also contained DMSO at 0.1 % DMSO final. After 48 hours, cells were fixed using 3 % PFA in PBS, and the plates sealed and disinfected to inactivate SARS-CoV-2	The collection of 5641 compounds was assembled by an external partner (SPECS) in a manner aligned to the recommendations from the Broad Institute (Cambridge Mass., USA). In assembling this “mirror” collection, compounds were purchased from the same set of >70 high quality suppliers identified by the Broad, (personal communication Dr Joshua Bitker), and quality controlled by LC/MS for purity and identity (minimum purity > 90%). Compounds were stored at 10mM in 100% DMSO at -20 °C. A curated database is available containing the compound, indication, primary target (where known) and mechanism of action, as well as analysis tools which can assist in mechanism of action determination and target elucidation.	Quantification of viral inhibition (based upon Caco-2 cell viability relative to controls) was performed using high content imaging of unlabeled cells. For high content imaging an Operetta CLS microscope from PerkinElmer was used. Cells were analysed in label free mode using digital phase contrast, with maximum contrast as a read out. Images were acquired using 10x objective with nine imaged fields per well and analysed using the manufacturer's software (PerkinElmer, Columbus v.2.9.0.1546).	The analysis sequence started with cell detection (method: c, common threshold: 0.05, area >100 μm, splitting coefficient: 6.5, individual threshold: 0.05, contrast >0.05) and was followed by calculating morphology, intensity and position properties as well as cell confluence. Well level data were analysed using ActivityBase (IDBS) and R (v.3.6.1). Test well results were normalized relative to the corresponding intra-plate control (no virus assigned as 100% inhibition of cellular toxicity, with virus assigned as 0% inhibition of cellular toxicity). Outliers were eliminated according to 3-sigma method. Plate level statistical performance was assessed using the standard Z’ calculation5. Dose response curves fitted in GraphPad Prism v.8.0.0 (GraphPad Software) using a 4-parameter logarithmic (least squares fit). Hits were further selected for dose response studies based on the progression of the compounds in clinical development. Therefore drugs in clinical use were priorised over phase 2 compounds, which were priorized over phase 1 or phase 1/phase 2 compounds.																												
																																	
																																	
# Phenotypes																																	
Phenotype Name	number of cells																																
Phenotype Description	count number of cells and compare relative to control																																
Phenotype Score Type	automatic																																
Phenotype Term Source REF	CMPO																																
Phenotype Term Name	increased cell viability in population  																																
Phenotype Term Accession	http://www.ebi.ac.uk/cmpo/CMPO_0000161																																
																																	
# Raw Data Files																																	
Raw Image Data Format	https://docs.openmicroscopy.org/bio-formats/6.4.0/formats/perkinelmer-operetta.html																																
Raw Image Organization	102 plates, 96 wells, 9 fields																																
																																	
# Feature Level Data Files																																	
Feature Level Data File Name																																	
Feature Level Data File Description																																	
Feature Level Data File Format																																	
Feature Level Data Column Name																																	
Feature Level Data Column Description																																	
																																	
#  Processed Data Files 																																	
Processed Data File Name	idr0000-HitProfiling-DPC-processed.txt																																
Processed Data File Format	tab-delimited text																																
Processed Data File Description	Concentration of the compound, % Inhibition and number of cells																																
Processed Data Column Name	Plate Id	Well Reference	Compound Id	Concentration [µM]	% Inhibition	Cells - Number of Objects	NAME	Compound ID (2)	SMILES	IUPAC_NAME	pubchem_cid	unichem_link	broad_ids	Compound a hit ? (over 75% activity)	Phenoptye Annotation Level																		
Processed Data Column Type	plate	well	reagent identifier	experimental condition	data	reagent identifier	reagent identifier	reagent identifier	reagent identifier	reagent identifier	reagent identifier	reagent identifier	reagent identifier	data	phenotype																		
Processed Data Column Annotation Level					multiple replicates of reagent and experimental condition	multiple replicates of reagent and experimental condition								multiple replicates of reagent and experimental condition	multiple replicates of reagent and experimental condition																		
Processed Data Column Description					% Inhibition	Number of Cells																											
Processed Data Column Link To Library File	Link to Library file	Link to Library file	Link to Library file				Link to Library file	Link to Library file	Link to Library file	Link to Library file	Link to Library file	Link to Library file	Link to Library file