The Suramin Derivative NF449 Interacts with the 5-fold Vertex of the Enterovirus A71 Capsid to Prevent Virus Attachment to PSGL-1 and Heparan Sulfate

Abstract

NF449, a sulfated compound derived from the antiparasitic drug suramin, was previously reported to inhibit infection by enterovirus A71 (EV-A71). In the current work, we found that NF449 inhibits virus attachment to target cells, and specifically blocks virus interaction with two identified receptors—the P-selectin ligand, PSGL-1, and heparan sulfate glycosaminoglycan—with no effect on virus binding to a third receptor, the scavenger receptor SCARB2. We also examined a number of commercially available suramin analogues, and newly synthesized derivatives of NF449; among these, NF110 and NM16, like NF449, inhibited virus attachment at submicromolar concentrations. PSGL-1 and heparan sulfate, but not SCARB2, are both sulfated molecules, and their interaction with EV-A71 is thought to involve positively charged capsid residues, including a conserved lysine at VP1-244, near the icosahedral 5-fold vertex. We found that mutation of VP1-244 resulted in resistance to NF449, suggesting that this residue is involved in NF449 interaction with the virus capsid. Consistent with this idea, NF449 and NF110 prevented virus interaction with monoclonal antibody MA28-7, which specifically recognizes an epitope overlapping VP1-244 at the 5-fold vertex. Based on these observations we propose that NF449 and related compounds compete with sulfated receptor molecules for a binding site at the 5-fold vertex of the EV-A71 capsid.

In Vitro and In Vivo Inhibition of the Infectivity of Human Enterovirus 71 by a Sulfonated Food Azo Dye, Brilliant Black BN

“…suramin has been reported to interact with EV71 virions in a saturation transfer difference nuclear magnetic resonance (STD-NMR) study (22), and the binding sites of NF449 on EV71 virions included amino acids VP1-98E and VP1-244K at the vertex of the 5-fold axis of viral capsid (23). Similarly, dye E151 interacted with virions to prevent viral FIG 9 Protection of EV71-challenged AG129 mice by E151.…”

“…In contrast, sulfated P-selectin glycoprotein ligand-1 (PSGL-1) and glycosaminoglycans (GAGs) on the cell surface interact with the positively charged vertex of the 5-fold axis of viral capsid to facilitate EV71 attachment and infection (15)(16)(17)(18)(19)(20). Similarly, sulfated/sulfonated heparin, suramin, and suramin derivatives have been shown to bind to the vertex of the 5-fold axis to block EV71 attachment to host cells (21)(22)(23). The cellular protein cyclophilin A has been determined to be an EV71-uncoating regulating factor through interacting with and modifying the conformation of the H-I loop of VP1 at the vertex of the 5-fold axis (24).…”

A Single Mutation in the VP1 of Enterovirus 71 Is Responsible for Increased Virulence and Neurotropism in Adult Interferon-Deficient Mice

“…H37R restored these interactions, allowing viral replication to occur. Position 244 of VP1 is a critical residue for EV71 receptor attachment and is conserved among almost all EV71 isolates (5,40). It has been proposed that a positively charged lysine at position 244 is needed for the EV71 capsid to interact with PSGL-1 and heparan sulfate (both EV71 receptors) (5,11).…”

In Vitro Assessment of Combinations of Enterovirus Inhibitors against Enterovirus 71

“…These inhibitors have distinct mechanisms of action and different resistance profiles. Suramin and its analog NF449 blocked EV-A71 infection at the step of virus binding (18)(19)(20)(21), and NF449-resistant viruses contain two mutations (E98Q and K244R) in the VP1 protein (21,22). ITZ exhibited broad-spectrum antienterovirus activity by target-ing host oxysterol-binding protein (OSBP) (23), and ITZ-resistant EV-A71 contains a single mutation in the 3A protein (V51L or V75A) (24).…”