Host chitinase 3-like-1 is a universal therapeutic target for SARS-CoV-2 viral variants in COVID-19
Curation statements for this article:-
Curated by eLife
Evaluation Summary:
The observation that CHI3L1 has the potential to modulate SARS-CoV-2 variants is timely and of potentially great significance. This is a novel approach to treatment and if sufficient additional data can be provided regarding in vivo efficacy this would be of great interest.
(This preprint has been reviewed by eLife. We include the public reviews from the reviewers here; the authors also receive private feedback with suggested changes to the manuscript.The reviewers remained anonymous to the authors.)
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- Immunology and Inflammation (eLife)
Abstract
Coronavirus disease 2019 (COVID-19) is the disease caused by severe acute respiratory syndrome coronavirus-2 (SARS-CoV-2; SC2), which has caused a worldwide pandemic with striking morbidity and mortality. Evaluation of SC2 strains demonstrated impressive genetic variability, and many of these viral variants are now defined as variants of concern (VOC) that cause enhanced transmissibility, decreased susceptibility to antibody neutralization or therapeutics, and/or the ability to induce severe disease. Currently, the delta (δ) and omicron (ο) variants are particularly problematic based on their impressive and unprecedented transmissibility and ability to cause breakthrough infections. The delta variant also accumulates at high concentrations in host tissues and has caused waves of lethal disease. Because studies from our laboratory have demonstrated that chitinase 3-like-1 (CHI3L1) stimulates ACE2 and Spike (S) priming proteases that mediate SC2 infection, studies were undertaken to determine if interventions that target CHI3L1 are effective inhibitors of SC2 viral variant infection. Here, we demonstrate that CHI3L1 augments epithelial cell infection by pseudoviruses that express the alpha, beta, gamma, delta, or omicron S proteins and that the CHI3L1 inhibitors anti-CHI3L1 and kasugamycin inhibit epithelial cell infection by these VOC pseudovirus moieties. Thus, CHI3L1 is a universal, VOC-independent therapeutic target in COVID-19.
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Author Response
Reviewer #1 (Public Review):
In this manuscript, Kamle and colleagues report that inhibition of host constitutively-expressed chitinase 3-like-1 (CHI3L1) increased epithelial expression of ACE2 and SPP, resulting in epithelial cell viral uptake of pseudoviruses that express the alpha, beta, gamma, delta or omicron S proteins. They further show that antagonism of CHI3L1 using anti-CHI3L1 or kasugamycin inhibits epithelial cell infection by the pseudoviruses with ancestral, alpha, beta, gamma S protein mutations. The in vitro data has relevance to SARS-CoV-2 pathogenesis and potentially has therapeutic implications in that the anti-CHI3L1 antibody and/or kasugamycin might be a treatment for this pandemic virus. These in vitro data are novel and the results are clear and convincing.
We are pleased that the reviewer …
Author Response
Reviewer #1 (Public Review):
In this manuscript, Kamle and colleagues report that inhibition of host constitutively-expressed chitinase 3-like-1 (CHI3L1) increased epithelial expression of ACE2 and SPP, resulting in epithelial cell viral uptake of pseudoviruses that express the alpha, beta, gamma, delta or omicron S proteins. They further show that antagonism of CHI3L1 using anti-CHI3L1 or kasugamycin inhibits epithelial cell infection by the pseudoviruses with ancestral, alpha, beta, gamma S protein mutations. The in vitro data has relevance to SARS-CoV-2 pathogenesis and potentially has therapeutic implications in that the anti-CHI3L1 antibody and/or kasugamycin might be a treatment for this pandemic virus. These in vitro data are novel and the results are clear and convincing.
We are pleased that the reviewer found these studies to be clear and convincing. We are also pleased that the reviewer recognizes the therapeutic potential of anti-CHI3L1 and kasugamycin in COVID 19.
The most important challenge with this manuscript is whether these in vitro findings translate into inhibiting SARS-CoV-2 variants in vivo. Are the effects of anti-CHI3L1 or kasugamycin great enough to change the course of the disease? Given the limitations of the mouse model of human ACE2 expression, determining how effective this strategy is in disease pathogenesis is difficult to discern. Without in vivo results, the importance of the data in this manuscript is unknown and this is a significant limitation that should be certainly noted in the discussion and possibly the abstract.
We agree that in vivo studies could add significantly to our understandings of the therapeutic potential of anti-CHI3L1 and kasugamycin. We can not undertake these investigations at the present time due to our lack of access to a BSL-3 lab facility. As requested, we have added a paragraph to the discussion that addresses this limitation. This new paragraph can be seen on pages 13-14. We have also modified that final paragraph in the discussion to highlight the importance of in vivo investigations and the limitations of the K18-hACE2 mouse model of SC2 infection.
Reviewer #2 (Public Review):
The paper by Khamle et al shows that CHI3L1 augments SARS-COV2 pseudovirus uptake in cells and that blocking CHI3L1 partially reduces uptake but the effect is not as efficient as some mAbs or soluble ACE2. A major limitation of the work is all of the data are based solely on experiments with pseudovirus. To be impactful, work would need to be performed with live virus assays as well as in vivo with either K18 mice or hamster models.
We agree that in vivo experimentation has the potential to strengthen the therapeutic potential of these studies. The reasons why these can not be undertaken at the present time are noted above and are addressed in the new paragraph that has been added to the discussion on pages 13-14. In addition, the limitations of the K18-hACE2 mouse model are also commented on in the new paragraph and addressed above.
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Evaluation Summary:
The observation that CHI3L1 has the potential to modulate SARS-CoV-2 variants is timely and of potentially great significance. This is a novel approach to treatment and if sufficient additional data can be provided regarding in vivo efficacy this would be of great interest.
(This preprint has been reviewed by eLife. We include the public reviews from the reviewers here; the authors also receive private feedback with suggested changes to the manuscript.The reviewers remained anonymous to the authors.)
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Reviewer #2 (Public Review):
The paper by Khamle et al shows that CHI3L1 augments SARS-COV2 pseudovirus uptake in cells and that blocking CHI3L1 partially reduces uptake but the effect is not as efficient as some mAbs or soluble ACE2. A major limitation of the work is all of the data are based solely on experiments with pseudovirus. To be impactful, work would need to be performed with live virus assays as well as in vivo with either K18 mice or hamster models.
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Reviewer #1 (Public Review):
In this manuscript, Kamle and colleagues report that inhibition of host constitutively-expressed chitinase 3-like-1 (CHI3L1) increased epithelial expression of ACE2 and SPP, resulting in epithelial cell viral uptake of pseudoviruses that express the alpha, beta, gamma, delta or omicron S proteins. They further show that antagonism of CHI3L1 using anti-CHI3L1 or kasugamycin inhibits epithelial cell infection by the pseudoviruses with ancestral, alpha, beta, gamma S protein mutations. The in vitro data has relevance to SARS-CoV-2 pathogenesis and potentially has therapeutic implications in that the anti-CHI3L1 antibody and/or kasugamycin might be a treatment for this pandemic virus. These in vitro data are novel and the results are clear and convincing.
The most important challenge with this manuscript is …
Reviewer #1 (Public Review):
In this manuscript, Kamle and colleagues report that inhibition of host constitutively-expressed chitinase 3-like-1 (CHI3L1) increased epithelial expression of ACE2 and SPP, resulting in epithelial cell viral uptake of pseudoviruses that express the alpha, beta, gamma, delta or omicron S proteins. They further show that antagonism of CHI3L1 using anti-CHI3L1 or kasugamycin inhibits epithelial cell infection by the pseudoviruses with ancestral, alpha, beta, gamma S protein mutations. The in vitro data has relevance to SARS-CoV-2 pathogenesis and potentially has therapeutic implications in that the anti-CHI3L1 antibody and/or kasugamycin might be a treatment for this pandemic virus. These in vitro data are novel and the results are clear and convincing.
The most important challenge with this manuscript is whether these in vitro findings translate into inhibiting SARS-CoV-2 variants in vivo. Are the effects of anti-CHI3L1 or kasugamycin great enough to change the course of the disease? Given the limitations of the mouse model of human ACE2 expression, determining how effective this strategy is in disease pathogenesis is difficult to discern. Without in vivo results, the importance of the data in this manuscript is unknown and this is a significant limitation that should be certainly noted in the discussion and possibly the abstract.
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SciScore for 10.1101/2022.01.21.477274: (What is this?)
Please note, not all rigor criteria are appropriate for all manuscripts.
Table 1: Rigor
Ethics not detected. 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
Antibodies Sentences Resources Generation of monoclonal antibodies against CHI3L1 (FRG): The murine monoclonal anti-CHI3L1 antibody (FRG) was generated using peptide antigen (amino acid 223-234 of human CHI3L1) as immunogen through Abmart Inc (Berkeley Heights, NJ). CHI3L1suggested: NoneThen the cells on the slides were fixed, permeabilized, and treated with blocking buffer then incubated with anti-ACE2 antibody (R&D, AF933) for overnight at 4°C. anti-ACE2suggested: NoneThe photographs of cellular immunofluorescence … SciScore for 10.1101/2022.01.21.477274: (What is this?)
Please note, not all rigor criteria are appropriate for all manuscripts.
Table 1: Rigor
Ethics not detected. 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
Antibodies Sentences Resources Generation of monoclonal antibodies against CHI3L1 (FRG): The murine monoclonal anti-CHI3L1 antibody (FRG) was generated using peptide antigen (amino acid 223-234 of human CHI3L1) as immunogen through Abmart Inc (Berkeley Heights, NJ). CHI3L1suggested: NoneThen the cells on the slides were fixed, permeabilized, and treated with blocking buffer then incubated with anti-ACE2 antibody (R&D, AF933) for overnight at 4°C. anti-ACE2suggested: NoneThe photographs of cellular immunofluorescence of GFP (+) pseudovirus and Cy-5 (+) ACE2 expression was taken with fluorescent microscopes Western blotting (Immunoblotting): 25 µg lung or cell lysates were subjected to immunoblot analysis using antibodies against phosphorylated (p) ERK (pERK), total ERK(ERK), Phosphorylated(p) AKT (pAKT), total AKT(AKT) (Cell Signaling Tech, MA, USA). GFPsuggested: Noneantibodies against phosphorylated (p) ERKsuggested: NonePhosphorylated(p) AKT (pAKT), total AKT(AKT) (Cell Signaling Tech, MAsuggested: NoneExperimental Models: Cell Lines Sentences Resources Cell lines and primary cells in culture: Calu-3 (HTB-55) lung epithelial cells were purchased from American Tissue Type Collection (ATCC) and maintained at 37°C in Dulbecco’s modified eagle medium (DMEM) supplemented with high glucose, L-Glutamine, minimal essential media (MEM) non-essential amino acids, penicillin/streptomycin and 10% fetal bovine serum (FBS) until used. Calu-3suggested: ATCC Cat# HTB-55, RRID:CVCL_0609)HEK-293T cells were transfected with the FRG construct using Lipofectamine™ 3000 (Invitrogen, # L3000015). HEK-293Tsuggested: NoneSoftware and Algorithms Sentences Resources Flow cytometry analysis of GFP (+) cells was carried out 48 h after infection on a BD LSRII flow cytometer and analyzed with the FlowJo software. FlowJosuggested: (FlowJo, RRID:SCR_008520)Quantification and Statistical analysis: Statistical evaluations were undertaken with GraphPad Prism software. GraphPad Prismsuggested: (GraphPad Prism, RRID:SCR_002798)Results 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: An explicit section about the limitations of the techniques employed in this study was not found. We encourage authors to address study limitations.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: Please consider improving the rainbow (“jet”) colormap(s) used on page 29. At least one figure is not accessible to readers with colorblindness and/or is not true to the data, i.e. not perceptually uniform.
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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