The approved pediatric drug suramin identified as a clinical candidate for the treatment of EV71 infection-suramin inhibits EV71 infection in vitro and in vivo

Abstract

Enterovirus 71 (EV71) causes severe central nervous system infections, leading to cardiopulmonary complications and death in young children. There is an urgent unmet medical need for new pharmaceutical agents to control EV71 infections. Using a multidisciplinary approach, we found that the approved pediatric antiparasitic drug suramin blocked EV71 infectivity by a novel mechanism of action that involves binding of the naphtalentrisulonic acid group of suramin to the viral capsid. Moreover, we demonstrate that when suramin is used in vivo at doses equivalent to or lower than the highest dose already used in humans, it significantly decreased mortality in mice challenged with a lethal dose of EV71 and peak viral load in adult rhesus monkeys. Thus, suramin inhibits EV71 infection by neutralizing virus particles prior to cell attachment. Consequently, these findings identify suramin as a clinical candidate for further development as a therapeutic or prophylactic treatment for severe EV71 infection.

Keywords: anti-viral; drug discovery; enterovirus 71; hand, foot and mouth disease; suramin.

Figure 1

Suramin was identified as an EV71 inhibitor. (A) Suramin inhibits the replication of EV71 isolates Fuyang573, SH12-036, SH12-276 and SEP-4, without cytotoxicity. Viral load was measured by quantitative RT-PCR, and expressed as the EV71 genome equivalent to TCID₅₀/mL. Data represents the mean±s.e.m. of results of duplicated experiment. (B) Suramin reduces the progeny virus yield. Data represents the mean±s.e.m. of results of two independent experiments which are duplicated.

Figure 2

Suramin blocks EV71 virus-cell attachment. (A) Time of addition assay. 10 µM suramin was added at different stage in viral infection as shown in figure. The viral load in supernatant represents the means±s.e.m. of results of experiment with three replicates, and the intracellular viral load represents the result of a single test. (B) Inhibition of viral attachment by suramin (cell-virus adsorption at 4°C). Data represents the means±s.e.m. of results of experiment with three replications.

Figure 3

Sulfonated and sulfated compounds inhibit the replication of the EV71 Fuyang573 isolate. (A) Sulfated suramin analogs inhibit EV71 infection. Data represents the means±s.e.m. of results of experiment with three replications. (B) Sulfonated and sulfated compounds not analogous to suramin inhibit EV71 infection. Data represents the means±s.e.m. of results of experiment with three replications.

Figure 4

STD NMR assay of suramin-EV71 particle binding. (A) STD NMR of suramin with the EV71 particle. ¹H (proton) NMR spectrum of suramin, shown in black, and STD NMR spectrum of suramin in complex with EV71 particles, shown in red. (B) Left: structure of suramin, labeled with proton positions and relative intensity percentages. Right: ¹H NMR spectrum of suramin, with proton numbers shown.

Figure 5

Anti-EV71 efficacy in vivo. (A) Infect 10-day-old ICR mice with 1×10⁷ TCID₅₀ (lethal dose) of the mouse-adapted EV71 strain MP10. Then inject 20 or 50 mg/kg body weight suramin by i.p. injection, twice daily for 7 days. There are 10 mice in each group. (B) Suramin at a dose of 50 mg/kg body weight decreases EV71 viremia in adult rhesus monkeys. Challenge adult monkeys with 1×10^6.5 CCID₅₀ EV71 FY-23 strain, and inject 50 mg/kg suramin i.v., on the day before virus challenge and on days 1, 3 and 5 post challenge. There are five monkeys in each group. ICR, Institute of Cancer Research; i.p., intraperitoneal; i.v., intravenously.