siRNA Therapeutics against Respiratory Viral Infections-What Have We Learned for Potential COVID-19 Therapies?

Abstract

Acute viral respiratory tract infections (AVRIs) are a major burden on human health and global economy and amongst the top five causes of death worldwide resulting in an estimated 3.9 million lives lost every year. In addition, new emerging respiratory viruses regularly cause outbreaks such as SARS-CoV-1 in 2003, the "Swine flu" in 2009, or most importantly the ongoing SARS-CoV-2 pandemic, which intensely impact global health, social life, and economy. Despite the prevalence of AVRIs and an urgent need, no vaccines-except for influenza-or effective treatments were available at the beginning of the COVID-19 pandemic. However, the innate RNAi pathway offers the ability to develop nucleic acid-based antiviral drugs. siRNA sequences against conserved, essential regions of the viral genome can prevent viral replication. In addition, viral infection can be averted prophylactically by silencing host genes essential for host-viral interactions. Unfortunately, delivering siRNAs to their target cells and intracellular site of action remains the principle hurdle toward their therapeutic use. Currently, siRNA formulations and chemical modifications are evaluated for their delivery. This progress report discusses the selection of antiviral siRNA sequences, delivery techniques to the infection sites, and provides an overview of antiviral siRNAs against respiratory viruses.

Keywords: SARS-CoV-2; inhalation; nanomedicine; pulmonary delivery; respiratory virus; siRNA.

Figure 1

A) The mechanism of the RNA interference pathway. Double‐stranded RNA is cleaved by Dicer into short interfering RNAs. After incorporation into the RISC, the siRNA recognizes base‐complementary mRNA and guides its cleavage. When artificially applied via nanoparticles, siRNAs are taken up by the cells where they enter the early endosomes and following endosomal escape, siRNAs are released into the cytoplasm where they are taken up into the RISC. Figure created with Biorender.com. B) Endosomal escape of polyamine nanocarriers via the hypothesized proton sponge effect. Protons are pumped into the endosomes during endosomal ripening. Due to the buffering capacity of the polyamine, the proton flux increases and chloride counterions enter the endosomal compartment. This increased osmotic pressure leads to additional influx of water and the eventual burst of the endosome releasing the nanocarrier.

Figure 2

The SARS‐CoV‐2 infection lifecycle. After binding to the cellular ACE2 membrane receptor, SARS‐CoV‐2 is endocytosed and the viral genome released into the cytoplasm. Here, the polyprotein 1ab is translated directly from the (+) sense genome and cleaved into individual non‐structural proteins which form the replication transcription complex around the RNA genome. Now, full‐length genomic as well as subgenomic RNAs (sgRNA) are transcribed via negative sense RNA intermediates, and sgRNAs are subsequently translated into proteins, including the structural N, S, M and E proteins as well as ORF 3a, 6, 7a/b, and 8. Now the structural proteins assemble around full‐length RNA genomes to form progeny virus which is exocytosed. siRNA can be designed to inhibit the host–virus interactions by targeting host factors such as the ACE2 receptor, or viral genomic or subgenomic RNAs. Figure created with BioRender.com

Figure 3

Schematic presentation of ALI cell culture and air‐lift (top) as well as an example of ALI‐cultured Calu‐3 cells (bottom, mucus stained with FITC‐wheat‐germ‐agglutinin (green), cytoskeleton (red), nuclei (blue)).

Figure 4

Schematic presentation of the spray drying process or siRNA nanocarriers into dry powder nano‐in‐micro formulations for inhalation. Reproduced with permission.^{[ 123 ]} Copyright 2019, Elsevier.