Integrin/TGF-β1 Inhibitor GLPG-0187 Blocks SARS-CoV-2 Delta and Omicron Pseudovirus Infection of Airway Epithelial Cells In Vitro, Which Could Attenuate Disease Severity
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Abstract
As COVID-19 continues to pose major risk for vulnerable populations, including the elderly, immunocompromised, patients with cancer, and those with contraindications to vaccination, novel treatment strategies are urgently needed. SARS-CoV-2 infects target cells via RGD-binding integrins, either independently or as a co-receptor with surface receptor angiotensin-converting enzyme 2 (ACE2). We used pan-integrin inhibitor GLPG-0187 to demonstrate the blockade of SARS-CoV-2 pseudovirus infection of target cells. Omicron pseudovirus infected normal human small airway epithelial (HSAE) cells significantly less than D614G or Delta variant pseudovirus, and GLPG-0187 effectively blocked SARS-CoV-2 pseudovirus infection in a dose-dependent manner across multiple viral variants. GLPG-0187 inhibited Omicron and Delta pseudovirus infection of HSAE cells more significantly than other variants. Pre-treatment of HSAE cells with MEK inhibitor (MEKi) VS-6766 enhanced the inhibition of pseudovirus infection by GLPG-0187. Because integrins activate transforming growth factor beta (TGF-β) signaling, we compared the plasma levels of active and total TGF-β in COVID-19+ patients. The plasma TGF-β1 levels correlated with age, race, and number of medications upon presentation with COVID-19, but not with sex. Total plasma TGF-β1 levels correlated with activated TGF-β1 levels. Moreover, the inhibition of integrin signaling prevents SARS-CoV-2 Delta and Omicron pseudovirus infectivity, and it may mitigate COVID-19 severity through decreased TGF-β1 activation. This therapeutic strategy may be further explored through clinical testing in vulnerable and unvaccinated populations.
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SciScore for 10.1101/2022.01.02.22268641: (What is this?)
Please note, not all rigor criteria are appropriate for all manuscripts.
Table 1: Rigor
Ethics Field Sample Permit: This is based on the fact that the project used deidentified specimens from a biobank with a determination that this project did not meet the definition of human subjects research based on specific criteria as described below.
Consent: The original samples were collected at Rhode Island hospital by the Lifespan Brown COVID-19 Biobank through an IRB-approved protocol that involved informed consent that was used by the biobank.Sex as a biological variable not detected. Randomization not detected. Blinding not detected. Power Analysis not detected. Cell Line Authentication not detected. Table 2: Resources
Experimental Models: Cell Lines Sentences Resources Briefly, 293FT cells … SciScore for 10.1101/2022.01.02.22268641: (What is this?)
Please note, not all rigor criteria are appropriate for all manuscripts.
Table 1: Rigor
Ethics Field Sample Permit: This is based on the fact that the project used deidentified specimens from a biobank with a determination that this project did not meet the definition of human subjects research based on specific criteria as described below.
Consent: The original samples were collected at Rhode Island hospital by the Lifespan Brown COVID-19 Biobank through an IRB-approved protocol that involved informed consent that was used by the biobank.Sex as a biological variable not detected. Randomization not detected. Blinding not detected. Power Analysis not detected. Cell Line Authentication not detected. Table 2: Resources
Experimental Models: Cell Lines Sentences Resources Briefly, 293FT cells (Invitrogen) at 75% confluency were co-transfected with the backbone vector pHAGE-fullEF1α-Luciferase-IRES-ZsGreen, plasmids expressing lentiviral proteins Tat, Rev and Gag/Pol, and plasmids expressing D614 or D614G S protein (a gift from Dr. Hyeryun Choe, The Scripps Research Institute, Jupiter, FL), or S protein with N501Y, E484K, N501Y+E484K or N501Y+E484K+K417N mutations. 293FTsuggested: NoneFollowing drug treatment, HSAE cells were spin-infected with SARS-CoV-2 pseudoviruses or a pantropic VsVg positive control lentivirus in a 12-well plate (931 g, 2 hours, 30°C with 8 μg/ml polybrene). HSAEsuggested: NoneRecombinant DNA Sentences Resources Briefly, 293FT cells (Invitrogen) at 75% confluency were co-transfected with the backbone vector pHAGE-fullEF1α-Luciferase-IRES-ZsGreen, plasmids expressing lentiviral proteins Tat, Rev and Gag/Pol, and plasmids expressing D614 or D614G S protein (a gift from Dr. Hyeryun Choe, The Scripps Research Institute, Jupiter, FL), or S protein with N501Y, E484K, N501Y+E484K or N501Y+E484K+K417N mutations. pHAGE-fullEF1α-Luciferase-IRES-ZsGreensuggested: NoneAn S protein expression plasmid construct containing all Omicron variant (B.1.1.529) mutations (28) was custom-made by GenScript (Piscataway, NJ): pcDNA3.1(+)-SARS-CoV-2-Omicron-(6xHis)-Spike (human codon). pcDNA3.1suggested: RRID:Addgene_79663)Software and Algorithms Sentences Resources Analysis of ZsGreen+ cells was conducted by flow cytometry 20-24 hours after infection using a BD LSRII flow cytometer and FlowJo software. FlowJosuggested: (FlowJo, RRID:SCR_008520)We completed a human subjects determination form for the Human Subjects Protection Program at Brown University. Human Subjects Protection Programsuggested: NoneSince we answered “no” to all these questions, our proposed project did NOT involve “Human Subjects.” Based on the information included in the Human Subjects Determination Form, The Human Research Protection Program at Brown University agreed with the investigator’s self-determination that the project does not meet the definition of human subjects research. Human Research Protection Programsuggested: NoneResults from OddPub: We did not detect open data. We also did not detect open code. Researchers are encouraged to share open data when possible (see Nature blog).
Results from LimitationRecognizer: We detected the following sentences addressing limitations in the study:Two limitations of this study include the small sample size of plasma samples from patients with COVID-19, as well as the lack of serial samples over time from the same patient. Because we only analyzed plasma from patients upon admission to the emergency department, we may have missed fluctuations in TGF-β concentrations, which are thought to peak during the first two weeks post-infection in severe COVID-19 cases (37). Future work should monitor levels of TGF-β in serial patient samples. Since its first identification in South Africa in November, 2021, the SARS-CoV-2 Omicron variant raised serious concerns of a significant reduction in efficacy of vaccines and monoclonal antibody treatments and an increased risk of reinfection due to numerous mutations in its spike protein, which is the antigenic target of infection- and vaccine-elicited antibodies against SARS-CoV-2. Currently, the Omicron variant is on track to outcompete the Delta variant as cases have soared to record highs in parts of Europe and now the U.S. according to the data released by Johns Hopkins University (https://coronavirus.jhu.edu/data). A number of recent studies suggest that much of the Omicron variant’s dominance comes down to its ability to evade the body’s immune defenses (28, 30, 38-40). However, earlier analyses of patients in South Africa suggest Omicron-infected individuals had a reduced risk of severe disease when compared to Delta-infected individuals (39). In the first findings on how the Omicr...
Results from TrialIdentifier: No clinical trial numbers were referenced.
Results from Barzooka: We did not find any issues relating to the usage of bar graphs.
Results from JetFighter: We did not find any issues relating to colormaps.
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.
Results from scite Reference Check: We found no unreliable references.
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