Heparin Inhibits Cellular Invasion by SARS-CoV-2: Structural Dependence of the Interaction of the Spike S1 Receptor-Binding Domain with Heparin

Courtney J. Mycroft-West; Dunhao Su; Isabel Pagani; Timothy R. Rudd; Stefano Elli; Neha S. Gandhi; Scott E. Guimond; Gavin J. Miller; Maria C. Z. Meneghetti; Helena B. Nader; Yong Li; Quentin M. Nunes; Patricia Procter; Nicasio Mancini; Massimo Clementi; Antonella Bisio; Nicholas R. Forsyth; Vito Ferro; Jeremy E. Turnbull; Marco Guerrini; David G. Fernig; Elisa Vicenzi; Edwin A. Yates; Marcelo A. Lima; Mark A. Skidmore

doi:10.1055/s-0040-1721319

Thrombosis and Haemostasis, Table of Contents

Thromb Haemost 2020; 120(12): 1700-1715
DOI: 10.1055/s-0040-1721319

Blood Cells, Inflammation and Infection

Heparin Inhibits Cellular Invasion by SARS-CoV-2: Structural Dependence of the Interaction of the Spike S1 Receptor-Binding Domain with Heparin

Courtney J. Mycroft-West^∗∗

¹Molecular and Structural Biosciences, School of Life Sciences, Keele University, Newcastle-Under-Lyme, Staffordshire, United Kingdom

,

Dunhao Su^∗∗

²Department of Biochemistry and Systems Biology, Institute of Systems, Molecular and Integrative Biology, University of Liverpool, Liverpool, United Kingdom

,

Isabel Pagani^∗∗

³Viral Pathogenesis and Biosafety Unit, Division of Immunology, Transplantation and Infectious Diseases, IRCCS San Raffaele Scientific Institute, Milan, Italy

,

Timothy R. Rudd^∗∗

⁴Analytical and Biological Sciences Division, National Institute for Biological Standards and Control, Potters Bar, Hertfordshire, United Kingdom

,

Stefano Elli^∗∗

⁵Istituto di Ricerche Chimiche e Biochimiche G. Ronzoni, Milan, Italy

,

Neha S. Gandhi^∗∗

⁶School of Chemistry and Physics, Queensland University of Technology, Brisbane, Australia

⁷Institute of Health and Biomedical Innovation, Queensland University of Technology, Brisbane, Australia

,

Scott E. Guimond^∗∗

⁸School of Medicine, Keele University, Newcastle-Under-Lyme, Staffordshire, United Kingdom

,

Gavin J. Miller

⁹School of Chemical and Physical Sciences, Keele University, Newcastle-Under-Lyme, Staffordshire, United Kingdom

,

Maria C. Z. Meneghetti

¹⁰Biochemistry Department, Federal University of São Paulo (UNIFESP), São Paulo, SP Brazil

,

Helena B. Nader

¹⁰Biochemistry Department, Federal University of São Paulo (UNIFESP), São Paulo, SP Brazil

,

Yong Li

²Department of Biochemistry and Systems Biology, Institute of Systems, Molecular and Integrative Biology, University of Liverpool, Liverpool, United Kingdom

,

Quentin M. Nunes

¹¹Department of Molecular and Clinical Cancer Medicine, Institute of Systems, Molecular and Integrative Biology, University of Liverpool, Liverpool, United Kingdom

,

Patricia Procter

¹Molecular and Structural Biosciences, School of Life Sciences, Keele University, Newcastle-Under-Lyme, Staffordshire, United Kingdom

,

Nicasio Mancini

¹²Università Vita-Salute San Raffaele, Milan, Italy

,

Massimo Clementi

¹²Università Vita-Salute San Raffaele, Milan, Italy

,

Antonella Bisio

⁵Istituto di Ricerche Chimiche e Biochimiche G. Ronzoni, Milan, Italy

,

Nicholas R. Forsyth

¹³Guy Hilton Research Centre, School of Pharmacy and Bioengineering, Keele University, Hartshill, Stoke-on-Trent, Staffordshire, United Kingdom

,

Vito Ferro

¹⁴School of Chemistry and Molecular Biosciences, The University of Queensland, Brisbane, Australia

¹⁵Australian Infectious Diseases Research Centre, The University of Queensland, Brisbane, Australia

,

Jeremy E. Turnbull^∗

²Department of Biochemistry and Systems Biology, Institute of Systems, Molecular and Integrative Biology, University of Liverpool, Liverpool, United Kingdom

,

Marco Guerrini^∗

⁵Istituto di Ricerche Chimiche e Biochimiche G. Ronzoni, Milan, Italy

,

David G. Fernig^∗

²Department of Biochemistry and Systems Biology, Institute of Systems, Molecular and Integrative Biology, University of Liverpool, Liverpool, United Kingdom

,

Elisa Vicenzi^∗

³Viral Pathogenesis and Biosafety Unit, Division of Immunology, Transplantation and Infectious Diseases, IRCCS San Raffaele Scientific Institute, Milan, Italy

,

Edwin A. Yates^∗

¹Molecular and Structural Biosciences, School of Life Sciences, Keele University, Newcastle-Under-Lyme, Staffordshire, United Kingdom

²Department of Biochemistry and Systems Biology, Institute of Systems, Molecular and Integrative Biology, University of Liverpool, Liverpool, United Kingdom

,

Marcelo A. Lima^∗

¹Molecular and Structural Biosciences, School of Life Sciences, Keele University, Newcastle-Under-Lyme, Staffordshire, United Kingdom

,

Mark A. Skidmore ^∗

¹Molecular and Structural Biosciences, School of Life Sciences, Keele University, Newcastle-Under-Lyme, Staffordshire, United Kingdom

²Department of Biochemistry and Systems Biology, Institute of Systems, Molecular and Integrative Biology, University of Liverpool, Liverpool, United Kingdom

› Author Affiliations

Abstract

The dependence of development and homeostasis in animals on the interaction of hundreds of extracellular regulatory proteins with the peri- and extracellular glycosaminoglycan heparan sulfate (HS) is exploited by many microbial pathogens as a means of adherence and invasion. Heparin, a widely used anticoagulant drug, is structurally similar to HS and is a common experimental proxy. Exogenous heparin prevents infection by a range of viruses, including S-associated coronavirus isolate HSR1. Here, we show that heparin inhibits severe acute respiratory syndrome coronavirus-2 (SARS-CoV-2) invasion of Vero cells by up to 80% at doses achievable through prophylaxis and, particularly relevant, within the range deliverable by nebulisation. Surface plasmon resonance and circular dichroism spectroscopy demonstrate that heparin and enoxaparin, a low-molecular-weight heparin which is a clinical anticoagulant, bind and induce a conformational change in the spike (S1) protein receptor-binding domain (S1 RBD) of SARS-CoV-2. A library of heparin derivatives and size-defined fragments were used to probe the structural basis of this interaction. Binding to the RBD is more strongly dependent on the presence of 2-O or 6-O sulfate groups than on N-sulfation and a hexasaccharide is the minimum size required for secondary structural changes to be induced in the RBD. It is likely that inhibition of viral infection arises from an overlap between the binding sites of heparin/HS on S1 RBD and that of the angiotensin-converting enzyme 2. The results suggest a route for the rapid development of a first-line therapeutic by repurposing heparin and its derivatives as antiviral agents against SARS-CoV-2 and other members of the Coronaviridae.

Keywords

heparin - coronavirus - SARS-CoV-2 - COVID-19 - spike - S1 - RBD - circular dichroism - surface plasmon resonance - molecular modelling

Full Text

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