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

Abstract

The ongoing COVID-19/Severe Acute Respiratory Syndrome CoV-2 (SARS-CoV-2) pandemic has become a significant threat to public health and has hugely impacted societies globally. Targeting conserved SARS-CoV-2 RNA structures and sequences essential for viral genome translation is a novel approach to inhibit viral infection and progression. This new pharmacological modality compasses two classes of RNA-targeting molecules: 1) synthetic small molecules that recognize secondary or tertiary RNA structures and 2) antisense oligonucleotides (ASOs) that recognize the RNA primary sequence. These molecules can also serve as a "bait" fragment in RNA degrading chimeras to eliminate the viral RNA genome. This new type of chimeric RNA degrader is recently named ribonuclease targeting chimera or RIBOTAC. This review paper summarizes the sequence conservation in SARS-CoV-2 and the current development of RNA-targeting molecules to combat this virus. These RNA-binding molecules will also serve as an emerging class of antiviral drug candidates that might pivot to address future viral outbreaks.

Keywords: RIBOTAC; RNA-targeting; SARS-CoV-2; antisense oligonucleotide; antiviral; programmed frameshift; small molecule; untranslated region.

FIGURE 1

SARS-CoV-2 Life cycle and viral targets for antiviral development. ① The virus enters the host cell through endocytosis using spike protein-angiotensin-converting enzyme 2 (ACE2) interaction. ② The host ribosome then translates the positive-sense RNA genome. ③ The long polypeptide precursor is subsequently cleaved by the viral proteases into non-structural proteins (nsp), which will assemble the replication-transcription complex (RTC) for ④ viral RNA genome replication in the 3’→5’ direction and ⑤ transcription in the 5’→3’ direction for the whole genome and sub-genomic sequences. ⑥ The host ribosome further translates the sub-genomic sequences that encode the nucleocapsid proteins. ⑦–⑧ Newly synthesized nucleocapsid components are assembled in the endoplasmic reticulum-Golgi intermediate compartment (ERGIC) to form the infectious virions, which are ⑨ released from the cell by maturation in the budding process. Some anti-SARS-CoV-2 agents illustrated in this figure include spike protein neutralizing antibody bamlanivimab, main protease inhibitors PF-07321332, MPI8, and bepridil, and RdRP inhibitors remdesivir and molnupiravir.

FIGURE 2

Nucleotide mutations in five VOCs compared to the original SARS-CoV-2 sequence discovered in Wuhan, China (RefSeq NC_045512). GenBank accession numbers: B.1.1.7 (Alpha): MZ344997, B.1.351 (Beta): MW598419, P.1 (Gamma): MZ169911, and B.1.617.2 (Delta): MZ359841. GISAID accession number: B.1.1.529 (Omicron): EPI_ISL_6795188.

FIGURE 3

The RNA structure and nucleotide conservation of the (A) PFS element in SARS-CoV-2, (B) 5’ UTR, and (C) 3’ UTR. The ribosome acts on the slippery sequence to produce a –1 PFS is illustrated in (A).

FIGURE 4

The small molecules with antiviral activities targeting the SARS-CoV-2 RNA genome.

FIGURE 5

(A) Antiviral ASO binding sites in the SARS-CoV-2 genome. (B) Chemical composition of the anti-SARS-CoV-2 ASOs.

FIGURE 6

Structures of (A) small molecule-based and (B) ASO-based RIBOTACs targeting the SARS-CoV-2 RNA genome.