Common cardiac medications potently inhibit ACE2 binding to the SARS-CoV-2 Spike, and block virus penetration and infectivity in human lung cells

Hung Caohuy,Ofer Eidelman,Alakesh Bera,Shufeng Liu,Tony T Wang,Qingfeng Yang,Harvey B Pollard,Tinghua Chen,Nathan I Walton

doi:10.1038/s41598-021-01690-9

Hung Caohuy, Ofer Eidelman + Show 7 more

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https://doi.org/10.1038/s41598-021-01690-9

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To initiate SARS-CoV-2 infection, the Receptor Binding Domain (RBD) on the viral spike protein must first bind to the host receptor ACE2 protein on pulmonary and other ACE2-expressing cells. We hypothesized that cardiac glycoside drugs might block the binding reaction between ACE2 and the Spike (S) protein, and thus block viral penetration into target cells. To test this hypothesis we developed a biochemical assay for ACE2:Spike binding, and tested cardiac glycosides as inhibitors of binding. Here we report that ouabain, digitoxin, and digoxin, as well as sugar-free derivatives digitoxigenin and digoxigenin, are high-affinity competitive inhibitors of ACE2 binding to the Original [D614] S1 and the α/β/γ [D614G] S1 proteins. These drugs also inhibit ACE2 binding to the Original RBD, as well as to RBD proteins containing the β [E484K], Mink [Y453F] and α/β/γ [N501Y] mutations. As hypothesized, we also found that ouabain, digitoxin and digoxin blocked penetration by SARS-CoV-2 Spike-pseudotyped virus into human lung cells, and infectivity by native SARS-CoV-2. These data indicate that cardiac glycosides may block viral penetration into the target cell by first inhibiting ACE2:RBD binding. Clinical concentrations of ouabain and digitoxin are relatively safe for short term use for subjects with normal hearts. It has therefore not escaped our attention that these common cardiac medications could be deployed worldwide as inexpensive repurposed drugs for anti-COVID-19 therapy.

Highlights

To initiate SARS-CoV-2 infection, the Receptor Binding Domain (RBD) on the viral spike protein must first bind to the host receptor ACE2 protein on pulmonary and other ACE2-expressing cells
We found similar results for inhibition of ACE2 binding to the α,β,γ mutant [D614G] S1 (Supplemental Fig. S4c and S4d), and to the Original RBD protein (Supplementary Fig. S4e and S4f)
An additional limitation is that while we discovered that the α/β/γ, [D614G] mutation in the S1 protein, which is at some distance from the RBD, raises the KD for ACE2 binding to the RBD, the present data do not reveal the mechanism

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Introduction

To initiate SARS-CoV-2 infection, the Receptor Binding Domain (RBD) on the viral spike protein must first bind to the host receptor ACE2 protein on pulmonary and other ACE2-expressing cells. Based on the foregoing, we hypothesized that cardiac glycoside drugs such as digitoxin, digoxin ouabain, might block the binding reaction between ACE2 and the viral Spike (S) protein, and block viral penetration into target cells (Supplemental Fig. S1) To test this hypothesis we first developed a biochemical method to measure the kinetics of ACE2 binding to immobilized variants of the Spike S1 and RBD domains. The most potent was ouabain, followed by digitoxin, with digoxin being the least active This relationship of inhibitory potencies was observed when we tested, in vivo, whether these cardiac glycoside drugs could block viral entry into human lung cells with SARS-CoV-2-Spike-pseudotyped virus, and whether they could block infectivity by native SARS-CoV-2. It is possible that these drugs could be repurposed for COVID-19 prevention and therapy

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