A Therapeutic Non-self-reactive SARS-CoV-2 Antibody Protects from Lung Pathology in a COVID-19 Hamster Model

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1. Version published to 10.1016/j.cell.2020.09.049

   Nov 1, 2020
2. 

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   Aug 17, 2020

   SciScore for 10.1101/2020.08.15.252320: (What is this?)

   Please note, not all rigor criteria are appropriate for all manuscripts.

   Table 1: Rigor

   Institutional Review Board Statement

   Consent: SARS-CoV-2-infected individuals and sample collection: The patients have given written informed consent and analyses were approved by the Institutional Review Board of Charité - Universitätsmedizin Berlin, corporate member of Freie Universität Berlin, Humboldt-Universität Berlin, and Berlin Institute of Health, Berlin. IRB: SARS-CoV-2-infected individuals and sample collection: The patients have given written informed consent and analyses were approved by the Institutional Review Board of Charité - Universitätsmedizin Berlin, corporate member of Freie Universität Berlin, Humboldt-Universität Berlin, and Berlin Institute of Health, Berlin.

   Randomization

   For …

   SciScore for 10.1101/2020.08.15.252320: (What is this?)

   Please note, not all rigor criteria are appropriate for all manuscripts.

   Table 1: Rigor

   Institutional Review Board Statement	Consent: SARS-CoV-2-infected individuals and sample collection: The patients have given written informed consent and analyses were approved by the Institutional Review Board of Charité - Universitätsmedizin Berlin, corporate member of Freie Universität Berlin, Humboldt-Universität Berlin, and Berlin Institute of Health, Berlin. IRB: SARS-CoV-2-infected individuals and sample collection: The patients have given written informed consent and analyses were approved by the Institutional Review Board of Charité - Universitätsmedizin Berlin, corporate member of Freie Universität Berlin, Humboldt-Universität Berlin, and Berlin Institute of Health, Berlin.
   Randomization	For the SARS-CoV-2 challenge experiments, hamsters were randomly distributed into three groups: In the first group (prophylaxis group), animals received an intraperitoneal (i.p.) injection of 18 mg per kg bodyweight of SARS-CoV-2 neutralizing mAb CV07-209 24 hours prior to infection.
   Blinding	For histopathology, slides were stained with hematoxylin and eosin (HE) followed by blinded microscopic evaluation by board-certified veterinary pathologists as previously described (Dietert et al., 2017; Osterrieder et al., 2020).
   Power Analysis	not detected.
   Sex as a biological variable	Twenty-seven six-week old female and male golden Syrian hamsters (Mesocricetus auratus; outbred hamster strain RjHan:AURA, Janvier Labs) were kept in groups of 3 animals in enriched, individually ventilated cages.
   Cell Line Authentication	not detected.

   Table 2: Resources

   Antibodies
   Sentences	Resources
   The first patients (CV01 (first time point), CV03, and CV05) were stained with a divergent set of antibodies: CD19-PE 1:50 (clone HIB19, BioLegend, 302207), CD38-PEcy7 1:50	CV01 suggested: None CV03 suggested: None CD19-PE suggested: None CD38-PEcy7 suggested: None
   To this end, a fusion protein (RBD-Fc) of the signal peptide of the NMDA receptor subunit GluN1, the RBD-SD1 part of SARS-CoV2-S1 (amino acids 319-591) and the constant region of rabbit IgG1 heavy chain (Fc) was expressed in HEK293T cells and immobilized onto 96-well plates from cell culture supernatant via anti-rabbit IgG (Dianova, 111-035-045) antibodies.	anti-rabbit IgG suggested: (Jackson ImmunoResearch Labs Cat# 111-035-045, RRID:AB_2337938)
   ACE2-HA binding was detected using anti-HA antibody HA.11 (clone 16B12, BioLegend, San Diego, CA, MMS-101P)	anti-HA suggested: (Covance Cat# MMS-101P, RRID:AB_2314672)
   For experiments regarding the competition between mAbs for RBD binding, purified monoclonal antibodies were biotinylated using EZ-Link Sulfo-NHS-Biotin (Thermo Fisher) according to the manufacturer’s instructions.	Sulfo-NHS-Biotin suggested: None
   For co-staining, the following antibodies were used: mouse Smooth Muscle Actin (clone 1A4, Agilent, 172	mouse Smooth Muscle Actin suggested: None
   Experimental Models: Cell Lines
   Sentences	Resources
   PBMC collection and FACS staining: Recombinant SARS-CoV-2-S1 protein produced in HEK cells (Creative Diagnostics, DAGC091) was covalently labeled using CruzFluor647 (Santa Cruz Biotechnology, sc-362620) according to the manufacturer’s instructions.	HEK suggested: None
   For mAb expression, human embryonic kidney cells (HEK293T) were transiently transfected with matching Ig heavy and light chains.	HEK293T suggested: NCBI_Iran Cat# C498, RRID:CVCL_0063)
   HCoV-NL63, HCoV-HKU1, and HCoV-229E and applied mAbs at 1 µg/ml.	HCoV-NL63 suggested: RRID:CVCL_RW88)
   After 24 hours, transfected as well as untransfected VeroB4 cells were harvested and resuspended in DMEM/10% FCS to achieve a cell density of 2.5×105 cells/ml each.	VeroB4 suggested: CCLV Cat# CCLV-RIE 1146, RRID:CVCL_1912)
   HEp2 cell assay: HEp-2 cell reactivity was investigated using the NOVA Lite HEp-2 ANA Kit (Inova Diagnostics) according to the manufacturer’s instructions using mAb containing culture supernatant (screening of all S1+ mAbs) or purified mAbs at 50 µg/ml (polyreactivity testing of CV07-200, CV07-209, CV07-222, CV07-255, CV07-270 and CV38-148) and examined under an inverted fluorescence microscope.	HEp2 suggested: None HEp-2 suggested: None
   Viruses were propagated on Vero E6 cells (ATCC CRL-1586) in minimal essential medium (MEM; PAN Biotech) supplemented with 10% fetal bovine serum (PAN Biotech) 100 IU/ml Penicillin G and 100 µg/ml Streptomycin (Carl Roth).	Vero E6 suggested: None
   Experimental Models: Organisms/Strains
   Sentences	Resources
   Murine tissue reactivity screening: Preparations of brain, lung, heart, liver, kidney and gut from 8-12 weeks old C57BL/6J mice were frozen in -50°C 2-methylbutane, cut on a cryostat in 20 µm sections and mounted on glass slides.	C57BL/6J suggested: None
   Software and Algorithms
   Sentences	Resources
   Cloning was considered successful when sequence identify >99.5% was given, verified by the cBASE module of BASE software.	BASE suggested: (BASE, RRID:SCR_010937)
   For in vivo experiments, mAbs were concentrated using Pierce™ 3K Protein Concentrator PES (Thermo Scientific).	Pierce™ suggested: None
   After identification of public clonotypes, they were plotted in a Circos plot using the R package circlize (Gu et al., 2014).	Circos suggested: (Circos, RRID:SCR_011798)
   Iterative model building and refinement were carried out in COOT (Emsley and Cowtan, 2004) and PHENIX (Adams et al., 2010), respectively.	COOT suggested: (Coot, RRID:SCR_014222) PHENIX suggested: (Phenix, RRID:SCR_014224)
   Statistical Analysis: All statistical tests were performed using GraphPad Prism, version 8.4.	GraphPad Prism suggested: (GraphPad Prism, RRID:SCR_002798)

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   Results from OddPub: Thank you for sharing your code and data.

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   Results from LimitationRecognizer: An explicit section about the limitations of the techniques employed in this study was not found. We encourage authors to address study limitations.

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   Results from TrialIdentifier: No clinical trial numbers were referenced.

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   Results from Barzooka: We did not find any issues relating to the usage of bar graphs.

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   Results from JetFighter: We did not find any issues relating to colormaps.

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   Results from rtransparent:

   • Thank you for including a conflict of interest statement. Authors are encouraged to include this statement when submitting to a journal.
   • Thank you for including a funding statement. Authors are encouraged to include this statement when submitting to a journal.
   • No protocol registration statement was detected.

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3. 

   Version published to 10.1101/2020.08.15.252320 on bioRxiv

   Aug 16, 2020