Inhibition of influenza A virus by mixed siRNAs, targeting the PA, NP, and NS genes, delivered by hybrid microcarriers

Antiviral Res. 2018 Oct;158:147-160. doi: 10.1016/j.antiviral.2018.08.003. Epub 2018 Aug 6.

Abstract

In the present study, a highly effective carrier system has been developed for the delivery of antiviral siRNA mixtures. The developed hybrid microcarriers, made of biodegradable polymers and SiO2 nanostructures, more efficiently mediate cellular uptake of siRNA than commercially available liposome-based reagents and polyethyleneimine (PEI); they also demonstrate low in vitro toxicity and protection of siRNA from RNase degradation. A series of siRNA designs (targeting the most conserved regions of three influenza A virus (IAV) genes: NP, NS, and PA) were screened in vitro using RT-qPCR, ELISA analysis, and hemagglutination assay. Based on the results of screening, the three most effective siRNAs (PA-1630, NP-717, and NS-777) were selected for in situ encapsulation into hybrid microcarriers. It was revealed that pre-treatment of cells with a mixture of PA-1630, NP-717, and NS-777 siRNAs, delivered by hybrid microcarriers, provided stronger inhibition of viral M1 mRNA expression and control of NP protein level, after viral infection, than single pre-treatment by any of three encapsulated siRNAs used in the study. Moreover, the effective inhibition of replication in several IAV subtypes (H1N1, H1N1pdm, H5N2, and H7N9) using a cocktail of the three selected siRNAs, delivered by our hybrid capsules to the cells, was achieved. In conclusion, we have developed a proof-of-principle which shows that our hybrid microcarrier technology (utilizing a therapeutic siRNA cocktail) may represent a promising approach in anti-influenza therapy.

Keywords: Hybrid microcontainers; Influenza A virus; RNAi therapy; siRNA delivery.

Publication types

  • Research Support, Non-U.S. Gov't

MeSH terms

  • A549 Cells
  • Animals
  • Antiviral Agents / pharmacology*
  • Cell Line
  • Cell Survival / drug effects
  • Dogs
  • Drug Delivery Systems / methods*
  • Epithelial Cells
  • Gene Expression Regulation, Viral / drug effects
  • Humans
  • Influenza A Virus, H1N1 Subtype / genetics
  • Influenza A Virus, H5N2 Subtype / genetics
  • Influenza A Virus, H7N9 Subtype / genetics
  • Influenza A virus / drug effects*
  • Influenza A virus / genetics*
  • Influenza A virus / pathogenicity
  • Liposomes
  • Madin Darby Canine Kidney Cells
  • Mice
  • Mice, Inbred BALB C
  • Nucleocapsid Proteins
  • Polyethyleneimine
  • RNA, Messenger / metabolism
  • RNA, Small Interfering / metabolism
  • RNA, Small Interfering / pharmacology*
  • RNA-Binding Proteins / genetics*
  • RNA-Binding Proteins / metabolism
  • RNA-Dependent RNA Polymerase / genetics*
  • RNA-Dependent RNA Polymerase / metabolism
  • Silicon Dioxide
  • Viral Core Proteins / genetics*
  • Viral Core Proteins / metabolism
  • Viral Matrix Proteins
  • Viral Nonstructural Proteins / genetics*
  • Viral Nonstructural Proteins / metabolism
  • Viral Proteins / genetics*
  • Viral Proteins / metabolism
  • Virus Replication / drug effects

Substances

  • Antiviral Agents
  • Liposomes
  • M1 protein, Influenza A virus
  • NP protein, Influenza A virus
  • NS protein, influenza virus
  • Nucleocapsid Proteins
  • PA protein, influenza viruses
  • RNA, Messenger
  • RNA, Small Interfering
  • RNA-Binding Proteins
  • Viral Core Proteins
  • Viral Matrix Proteins
  • Viral Nonstructural Proteins
  • Viral Proteins
  • Silicon Dioxide
  • Polyethyleneimine
  • RNA-Dependent RNA Polymerase