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Review 1: "Antisense oligonucleotides target a nearly invariant structural element from the SARS-CoV-2 genome and drive RNA degradation"

This manuscript offers a gapmer-based therapeutic strategy targeting the highly conserved s2m element present in the SARS-CoV-2 genome. While potentially informative, the study's claims require testing with wildtype SARS-CoV-2 virus and animal models to be substantiated.

Published onNov 05, 2020
Review 1: "Antisense oligonucleotides target a nearly invariant structural element from the SARS-CoV-2 genome and drive RNA degradation"
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Antisense oligonucleotides target a nearly invariant structural element from the SARS-CoV-2 genome and drive RNA degradation
Antisense oligonucleotides target a nearly invariant structural element from the SARS-CoV-2 genome and drive RNA degradation
Description

Summary The SARS-CoV-2 virus contains an unusually large, single-stranded RNA genome that is punctuated with structured elements of unknown function, such as the s2m element located in the 3’ untranslated region. The evolutionary conservation of the s2m element and its occurrence in all viral subgenomic transcripts implicates a key role in the viral infection cycle. In order to exploit this element as a potential therapeutic target, we have designed antisense “gapmer” oligonucleotides that efficiently base-pair to the s2m region. These oligonucleotides, composed of locked nucleic acids (LNA) flanking a central DNA core, successfully remodel the s2m structure and induce sequence-specific RNA cleavage by RNase H in vitro. Gapmers are also effective in human cells as they reduce the fluorescence signal in GFP reporter assays and cause a dose-dependent reduction in replication in a model replicon system based on a human astrovirus. Overall, these oligonucleotides show promise as anti-viral agents and may serve as a helpful starting point to develop treatments for COVID-19.

RR:C19 Evidence Scale rating by reviewer:

  • Reliable. The main study claims are generally justified by its methods and data. The results and conclusions are likely to be similar to the hypothetical ideal study. There are some minor caveats or limitations, but they would/do not change the major claims of the study. The study provides sufficient strength of evidence on its own that its main claims should be considered actionable, with some room for future revision.

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Review:

Claims are generally supported by the data and methods used. Decision-makers should consider the claims in this study actionable with limitations based on the methods and data. Also, this research is well-structured and well-written by using clear and accurate language. The manuscript has a high degree of novelty which contributes to broader research understandings. I can recommend it for publishing.

Reviewer recommendations:

1. Figure 1D-E requires more clear explanation like “what was the main purpose to do these experiments”, and “how does it contribute to support the idea in case of structures formed.”

2. Figure 2C is not clearly explained. There are some questions coming tp mind there; what are the meanings of multiple bands in +all, +some, and +RNaseH samples, how is it possible to be a degradation in +all and +some samples without RNaseH enzymes. These must be answered.

3. There is no in vivo study in this manuscript, so don’t use in vivo terms, as studies on cell lines are mostly accepted as in vitro studies as they are happening inside a tube.

4. Figure 5D should be discussed more. Possible questions: How do you explain the saturated replicon activity between 50 and 500ul doses in gapmer 6 sample for HEK293T cell lines? Why does the replicon activity significantly decrease in gapmer scr samples for HEK293T cell lines from 0 to 500ul dose?

5. The discussion section seems incomplete and more like a conclusion. That’s why there should be more efforts in that section to build a compact study.

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