Structure-based design of antisense oligonucleotides that inhibit SARS-CoV-2 replication

Abstract

Antisense oligonucleotides (ASOs) are an emerging class of drugs that target RNAs. Current ASO designs strictly follow the rule of Watson-Crick base pairing along target sequences. However, RNAs often fold into structures that interfere with ASO hybridization. Here we developed a structure-based ASO design method and applied it to target severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2). Our method makes sure that ASO binding is compatible with target structures in three-dimensional (3D) space by employing structural design templates. These 3D-ASOs recognize the shapes and hydrogen bonding patterns of targets via tertiary interactions, achieving enhanced affinity and specificity. We designed 3D-ASOs that bind to the frameshift stimulation element and transcription regulatory sequence of SARS-CoV-2 and identified lead ASOs that strongly inhibit viral replication in human cells. We further optimized the lead sequences and characterized structure-activity relationship. The 3D-ASO technology helps fight coronavirus disease-2019 and is broadly applicable to ASO drug development.

Emerging roles for CNS fibroblasts in health, injury and disease

“…Despite these recent advances, we still know very little about the roles of CNS fibroblasts in the development and maintenance of the healthy, adult CNS and how they contribute to disease aside from fibrotic scarring. While we focus mostly on rodent studies in this Review, perivascular fibroblasts have been detected in human tumour samples 5 and associated with the vasculature of the human brain 6 , 7 , confirming that these cells are indeed present in the human CNS. Furthermore, perivascular fibroblasts have been shown to be dysfunctional in patients with amyotrophic lateral sclerosis (ALS) 8 .…”

Inhibition of SARS-CoV-2 by Targeting Conserved Viral RNA Structures and Sequences

“…By using 3D antisense modeling, a PMO named PRF3p was optimized to target the Stem 3 region in the PFS element ( Li et al, 2021b ) ( Figure 5 ). The PRF3p binding disrupted the pseudoknotted structure in the PFS element and inhibited the frameshift, eventually leading to a knockdown of the genes encoded by the ORF1b in the virus-infected 293T cells ( Li et al, 2021b ). Gapmers S2D, S3D-1, S2D-2, and Slp-2 targeting PFS elements were reported to have efficacy in Huh-7 inoculated with SARS-CoV-2 with a luciferase reporter ( Zhang et al, 2021 ).…”

“…A PMO named SBD1 was designed to target the conserved TRS-L region in the SARS-CoV 5’ UTR ( Figure 5 ), and thereby inhibited the “discontinuous” transcription ( Li et al, 2021b ). The suppression of sub-genomic RNA transcription ultimately led to the reduction of viral structural protein levels and virus titer ( Li et al, 2021 ). Two PMOs, 5’END-1 and 5’END-2, targeted the viral 5’ UTR and were shown to inhibit the translation pre-initiation complex ( Rosenke et al, 2021 ).…”

“…Spike protein-targeting antibodies (e.g., bamlanivimab) can effectively neutralize the virus and prevent viral entry ( Gottlieb et al, 2021 ). RNA-targeting antisense oligonucleotides (ASO) or small molecules will degrade the viral RNA genome or hinder RNA translation ( Li et al, 2021b , Li et al, 2021a. ; Lulla et al, 2021 ; Rosenke et al, 2021 ; Sun et al, 2021a ; Zhang et al, 2021 ; Zhu et al, 2021 ).…”

Translational Control of COVID-19 and Its Therapeutic Implication

“…Targeting conserved RNA structures and sequences of SARS-CoV-2 is an alternative approach to inhibiting viral infection and progression ( 209 , 210 ). The most well-known examples are antisense oligonucleotides (ASOs), which contain modifications at their positions, such as 2-O-methyl (2-OME), 2-O-methoxy (2-MOE), locked nucleic acid (LNA), morpholino, or other nucleotide modifications, which may increase RNA base pairing, metabolic stability, and/or delivery ( 209 , 211 , 212 ).…”