Effect of insilico predicted and designed potential siRNAs on inhibition of SARS-CoV-2 in HEK-293 cells

Sohrab, Sayed Sartaj; El‐Kafrawy, Sherif A.; Azhar, Esam I.

doi:10.1016/j.jksus.2022.101965

Cited by 7 publications

(13 citation statements)

References 35 publications

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“…To reduce such risks, it would probably be necessary to track viral variant emergence and to update siRNA desgin periodically, always administering combinations of siRNAs targeting distinct viral regions (hence: unlikely to mutate in the same variant) rather than a single This study Ambike et al, 2022Becker et al, 2022Chang et al, 2022Friedrich et al, 2022Sohrab et al, 2022bTolksdorf et al, 2021Idris et al, 2021Khaitov et al, 2021Nabiabad et al, 2022Niktab et al, 2021 . "High activity" was defined as a significant (p¡0.05) repression of viral RNA accumulation by 50% in infected cultured cells Khaitov et al, 2021 ;Tolksdorf et al, 2021 ;Sohrab et al, 2022b ;Sohrab et al, 2022a ;Chang et al, 2022 ;Becker et al, 2022 (and this study), or significant repression of viral plaque count by 50% Idris et al, 2021 ;Friedrich et al, 2022, or a measured IC 50 for cell infection inhibition being 10 nM Ambike et al, 2022. siRNAs have unclear activity when they were assessed in a siRNA mixture rather than individually Niktab et al, 2021 or when they were combined with an anti-hACE2 aptamer before administration to cultured cells Nabiabad et al, 2022 (gray arrows; also denoted by a question mark for the number of highly active siRNAs in column "High activity siRNAs / total"). SARS-CoV-2 ORF coordinates are from GenBank ACC # OP278726.1; mature proteins generated from the Orf1ab polyprotein are indicated by pink rectangles.…”

Section: Discussionmentioning

confidence: 99%

“…Significance was assessed using Dunnett's test, with the "∅" siRNA as a reference; p-values are indicated above each graph, with p-values lower than 0.05 being shown in red. designed siRNAs with high activity ranges from 0 Sohrab et al, 2022b ;Sohrab et al, 2022a 4; note that siRNAs designed by Yogev et al, 2022 could not be analyzed because their sequence was not released). Our approach, yielding 6/8 siRNAs with high activity, proved efficient in designing potent siRNAs with a pure bioinformatics approach, without requiring high-throughput experimental screening.…”

Section: Comparison To Previously-published Sirnas and Recent Viral V...mentioning

confidence: 99%

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Biochemistry-informed design selects potent siRNAs against SARS-CoV-2

Houbron

Mockly

Rafasse

et al. 2022

Preprint

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RNA interference (RNAi) offers an efficient way to repress genes of interest, and it is widely used in research settings. Clinical applications emerged more recently, with 5 approved siRNAs (the RNA guides of the RNAi effector complex) against human diseases. The development of siRNAs against the SARS-CoV-2 virus could therefore provide the basis of novel Covid-19 treatments, while being easily adaptable to future variants or to other, unrelated viruses. Because the biochemistry of RNAi is very precisely described, it is now possible to design siRNAs with high predicted activity and specificity using only computational tools. While previous siRNA design algorithms tended to rely on simplistic strategies (raising fully complementary siRNAs against targets of interest), our approach uses the most up-to-date mechanistic description of RNAi to allow mismatches at tolerable positions and to force them at beneficial positions, while optimizing siRNA duplex asymmetry. Our pipeline proposes 8 siRNAs against SARS-CoV-2, and ex vivo assessment confirms the high antiviral activity of 6 out of 8 siRNAs, also achieving excellent variant coverage (with several 3-siRNA combinations recognizing each correctly-sequenced variant as of September 2022). Our approach is easily generalizable to other viruses as long as a variant genome database is available. With siRNA delivery procedures being currently improved, RNAi could therefore become an efficient and versatile antiviral therapeutic strategy.

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Section: Discussionmentioning

confidence: 99%

Section: Comparison To Previously-published Sirnas and Recent Viral V...mentioning

confidence: 99%

Biochemistry-informed design selects potent siRNAs against SARS-CoV-2

Houbron

Mockly

Rafasse

et al. 2022

Preprint

View full text Add to dashboard Cite

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“…The global coronavirus disease 19 (COVID- 19) pandemic is caused by severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) [1]. The virus has a higher rates human-to-human transmission rate, facilitating rapid spread across the world [2].…”

Section: Introductionmentioning

confidence: 99%

“…The ongoing pandemic prompted multiple research groups to evaluate siRNAbased therapies for COVID-19. While the most of the published studies so far reviewed the potential of RNAi to treat COVID-19 [14][15][16][17], describe insilico studies [18,19], initial proof-of-concept that SARS-CoV-2 can be inhibited by siRNAs was also provided [8,20,21]. The designed siRNA targeting the Orf1a/b region of the SARS-CoV-2 RNA genome, encoding for non-structural proteins (nsp), was tested.…”

Section: Introductionmentioning

confidence: 99%

“…The designed siRNA targeting the Orf1a/b region of the SARS-CoV-2 RNA genome, encoding for non-structural proteins (nsp), was tested. A siRNA with highly efficient inhibition of SARS-CoV-2 replication was identified [19,22]. The adenoassociated virus vectors co-expressing a cocktail of the short hairpin RNAs (shRNA) directed against the SARS-CoV-2 RdRp and N genes were tested.…”

Section: Introductionmentioning

confidence: 99%

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RNAi Degrades Spike Protein Gene Transcripts of the SARS-CoV-2 for Developing Drugs to Treat of COVID-19

Zhang¹,

Huang²,

Huang³

et al. 2022

Nano BioMed ENG

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COVID-19 is caused by severe acute respiratory SARS-CoV-2. Regardless of the availability of treatment strategies for COVID-19, effective therapy will remain essential. A promising approach to tackle the SARS-CoV-2 could be small interfering (si) RNAs. Here we designed the small hairpin RNA (named as shRNA688) for targeting the prepared 813 bp Est of the S protein genes (Delta). The conserved and mutated regions of the S protein genes from the genomes of the SARS-CoV-2 variants in the public database were analyzed. A 813 bp fragment encoding the most part of the RBD and partial downstream RBD of the S protein was cloned into the upstream red florescent protein gene (RFP) as a fusing gene in the pCMV-S-Protein RBD-Est-RFP plasmid for expressing a potential target for RNAi. The double stranded of the DNA encoding for shRNA688 was constructed in the downstream human H1 promoter of the plasmid in which CMV promoter drives enhanced green fluorescent protein (EGFP) marker gene expression. These two kinds of the constructed plasmids were co-transfected into HEK293T via Lipofectamine 2000. The degradation of the transcripts of the SARS-CoV-2 S protein fusing gene expressed in the transfected HEK293T treated by RNAi was analyzed by RT-qPCR with a specific probe of the targeted SARS-CoV-2 S protein gene transcripts. Our results showed that shRNA688 targeting the conserved region of the S protein genes could effectively reduce the transcripts of the S protein genes. This study provides a cell model and technical support for the research and development of the broad-spectrum small nucleic acid RNAi drugs against SARS-CoV-2 or the RNAi drugs for the other hazard viruses which cause human diseases.

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