Heparin inhibits cellular invasion by SARS-CoV-2: structural dependence of the interaction of the surface protein (spike) S1 receptor binding domain with heparin
cell biology drug discovery/repurposing structural biology
Authors: Mycroft-West et al.
Link to paper: https://www.biorxiv.org/content/10.1101/2020.04.28.066761v1
Journal/ Pre-Print: MedRxiv
Tags: Drug discovery/Drug repurpose
1. Addition of 100 ug/ml of heparin to vero cells showed a 70% inhibition of infection by SARS-CoV-2.
2. Heparin shown to be more approximately 30 times more potent (on a weight basis) at inhibiting SARS-CoV-2 S1 receptor-binding domain (RBD) than the low-molecular weight heparin enoxaparin.
3. Predicts possible sites on the RBD that heparin binds to.
In line with a report that heparin treatment improves prognosis in severe COVID-19 patients with coagulopathy the authors suggest heparin interferes with SARS-CoV-2 viral infection using an in vitro assay (vero cell plaque assay), as reported previously for SARS-CoV. They demonstrate an interaction between the Spike protein S1 RBD and immobilised heparin by SPR, and show competition for binding with heparin-derived oligo- and polysaccharides. Circular dichroism (CD) spectroscopy shows heparin and certain derivatives induce secondary structural changes in the SARS-CoV-2 S1 RBD, and conduct an analysis of the RBD sequence and modelled structure for potential heparin binding sites, which they compare with sequences from known heparin-binding proteins.
Impact for SARS-CoV2/COVID19 research efforts
· Understand the virology and/or cell biology of SARS-CoV2/COVID19
· Treatment of SARS-CoV2/COVID19 positive individuals
· In vitro study
Strengths and limitations of the paper
Novelty: Shows interactions between SARS-CoV-2 S1 RBD and heparin
Standing in the field: Low-molecular weight heparin has been found to have beneficial outcomes in severe Covid-19 patients with sepsis-induced coagulopathy or elevated D-dimer, but no significant effect for other severe Covid-19 patients (https://doi.org/10.1111/jth.14817) - however the former of these results are controversial.
Appropriate statistics: Needs error bars and/or error calculations for SPR and CD measurements. Especially for the percentage secondary structure graph to show that changes in CD spectra caused by heparin are statistically significant. Figure 1 lacks a multiple comparison post-test correction.
Viral model used: SARS-CoV-2 (Italy/UniSR1/2020 strain) used predominantly, but SARS-CoV used in addition for some experiments
Translatability: Heparin derivatives might be used to treat severe Covid-19 patients with sepsis-induced coagulopathy, by inhibiting the virus and treating the coagulopathy.
· The viral plaque assay should have a control without virus. Would also be worth including some modified heparin derivatives in the assay, and varying concentrations.
· No indication of the quality of the SARS-CoV-2 S1 RBD (possibly aggregated) in SPR binding measurements since it wasn’t purified by size-exclusion chromatography, and no SDS-PAGE to show protein quality. No RBD concentration range for SPR binding measurement.
· SPR data set is weak, very small binding response compared to FGF2 suggesting their analyte is inactive. No attempt to calculate an equilibrium dissociation constant (KD). No range of concentrations for RBD and no binding curve.
· Better to have measured affinity of heparin to RBD directly rather than through competition with the sensor surface.
· It would have been valuable to use mutagenesis to probe the heparin binding sites on RBD.
· Varying and increasing concentrations of heparin for CD measurements with RBD would have been helpful to show conformational changes of RBD.