Mechanism through Which Retrocyclin Targets Flavivirus Multiplication.

Xiaoying Jia,Alfredo Garzino-Demo,Lingling Sun,Minmin Zhou,Jiao Guo,Wuyuan Lu,Yang Liu,Weirong Yuan,Wei Wang,Gengfu Xiao

doi:10.1128/jvi.00560-21

Xiaoying Jia, Alfredo Garzino-Demo + Show 8 more

Open Access

https://doi.org/10.1128/jvi.00560-21

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Abstract

ABSTRACTCurrently, there are no approved drugs for the treatment of flavivirus infection. Accordingly, we tested the inhibitory effects of the novel θ-defensin retrocyclin-101 (RC-101) against flavivirus infection and investigated the mechanism underlying the potential inhibitory effects. First, RC-101 robustly inhibited both Japanese encephalitis virus (JEV) and Zika virus (ZIKV) infections. RC-101 exerted inhibitory effects on the entry and replication stages. Results also indicated that the nonstructural protein NS2B-NS3 serine protease might serve as a potential viral target. Furthermore, RC-101 inhibited protease activity at the micromolar level. We also demonstrated that with respect to the glycoprotein E protein of flavivirus, the DE loop of domain III (DIII), which is the receptor-binding domain of the E protein, might serve as another viral target of RC-101. Moreover, a JEV DE mutant exhibited resistance to RC-101, which was associated with deceased binding affinity of RC-101 to DIII. These findings provide a basis for the development of RC-101 as a potential candidate for the treatment of flavivirus infection.IMPORTANCE Retrocyclin is an artificially humanized circular θ-defensin peptide, containing 18 residues, previously reported to possess broad antimicrobial activity. In this study, we found that retrocyclin-101 inhibited flavivirus (ZIKV and JEV) infections. Retrocyclin-101 inhibited NS2B-NS3 serine protease activity, suggesting that the catalytic triad of the protease is the target. Moreover, retrocyclin-101 bound to the DE loop of the E protein of flavivirus, which prevented its entry.

Highlights

There are no approved drugs for the treatment of flavivirus infection
An immunofluorescence antibody (IFA) staining plaque assay for the antiviral effect of RC101 against Zika virus (ZIKV) PRVABC 59 showed a dose-dependent inhibition, with an IC50 of 7.033 mM (Fig. 1D to F)
We evaluated the antiviral effects of RC-101 against flaviviruses and elucidated the mechanism of action

Summary

Introduction

There are no approved drugs for the treatment of flavivirus infection. we tested the inhibitory effects of the novel u -defensin retrocyclin-101 (RC-101) against flavivirus infection and investigated the mechanism underlying the potential inhibitory effects. Q258, located in DII, and T410, located in the stem, are indispensable for low-pH-triggered conformational changes, in which the stem region undergoes zippering along with DII, leading to the postfusion conformation and membrane fusion [9,10,11] As it envelops the surface of the virion, the E protein is the natural target for antibodies and the design of entry inhibitors to prevent receptor binding and membrane fusion [4, 9, 12, 13]. Likewise, viral proteases such as NS2B-NS3 protease-helicase and the NS5 RNA-dependent RNA polymerase represent attractive drug targets in an attempt to identify replication inhibitors [14, 15]. Some viral proteases with negatively charged surfaces might serve as targets for RC-1 [20]

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