- unfractionated Heparin Inhibits SARS-CoV-2 S Protein Binding in RT4 Cells
- low Molecular Weight Heparins are Partial Inhibitors of SARS-CoV-2 S Protein

Heparin Studies Using RT4 Cells Pre-Treated with Anti-ACE2 Antibodies

RT4 cells were treated with 15µg/ml anti-ACE-2 antibody (alpha-ACE2).  A significant, 54.25% increase in S protein (S1 and S2 subunits) interaction with the cells was observed (p=0.0025).  The antibodies bound to ACE2 may cause the receptors to cluster together, and this may make them an easier target for SARS-CoV-2 S protein.  

Michigan State University

Partridge et al. determined which human cell line model to use for studying SARS-CoV-2 S protein attachment based off of the mRNA expression of different membrane proteins important to the pathogenesis of COVID-19.  They investigated the following cell types:

- RT4 urinary bladder epithelial cell line
- A549  lung adenocarcinoma alveolar basal epithelial cell line
- Caco2 colorectal adenocarcinoma cell line
- HaCaT skin keratinocyte cell line

RT4 was determined to be the best cell line to use because it expresses both ACE2 and TMPRSS2 which directly interact with SARS-CoV-2 S protein.  It also expresses the following accessory proteins relevant to S protein attachment:

- ADAM17: a metalloprotease that processes ACE2
- rhomboid-like 2 (RHBDL2): a protease that is involved with the function of ADAM17
- CD9: an adaptor protein that controls ADAM17

Cell Lines Developed by Partridge et al. to Study COVID-19

Partridge, L. J., Urwin, L., Nicklin, M. J., James, D. C., Green, L. R., & Monk, P. N. (2020). ACE2-independent interaction of SARS-CoV-2 spike protein to human epithelial cells can be inhibited by unfractionated heparin. bioRxiv.
https://www.biorxiv.org/content/10.1101/2020.05.21.107870v2
doi: 10.1101/2020.05.21.107870

ACE2-independent interaction of SARS-CoV-2 spike protein to human epithelial cells can be inhibited by unfractionated heparin

A549 are a cell line that do not express ACE2 receptors.  Partridge et al. observed that SARS-CoV-2 S protein interact with these cells even though they do not express ACE2.  Further studies were conducted using two types of 293T cells: cells that express low amounts of ACE2 and cells that over-express ACE2.  S protein interacted with both cell types, but the binding affinity for those expressing low amounts of ACE2 was nearly undetectable.  This suggests that the S protein, specifically the S1 and S2 subunits, may initially interact with host cells via  a different receptor.  However, binding affinity studies suggest that the viral receptor binding domain (RBD) requires higher expression levels of ACE2 to interact with cells.

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