A Morpholino Oligomer Targeting Highly Conserved Internal Ribosome Entry Site Sequence Is Able To Inhibit Multiple Species of Picornavirus

Stone, Jeffrey K.; Rijnbrand, René; Stein, David A.; Ma, Yanxia; Yang, Yan; Iversen, Patrick L.; Andino, Raul

doi:10.1128/aac.00011-08

Cited by 40 publications

(42 citation statements)

References 58 publications

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“…These results suggest that CSDE1 is a universal factor for translation initiation in human enteroviruses. Stone et al (2008) reported that morpholino oligomers targeting IRES inhibited multiple species of picornaviruses. Using a similar approach targeting the IRES region, which interacts with CSDE1, potential broad therapeutic inhibitors against diverse human enteroviruses could be developed.…”

Section: Discussionmentioning

confidence: 99%

Arrayed CRISPR screen with image-based assay reliably uncovers host genes required for coxsackievirus infection

Lee

Kim

et al. 2018

Genome Res.

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Pooled CRISPR screens based on lentiviral systems have been widely applied to identify the effect of gene knockout on cellular phenotype. Although many screens were successful, they also have the limitation that genes conferring mild phenotypes or those essential for growth can be overlooked, as every genetic perturbation is incorporated in the same population. Arrayed screens, on the other hand, incorporate a single genetic perturbation in each well and could overcome these limitations. However, arrayed screens based on siRNA-mediated knockdown were recently criticized for low reproducibility caused by incomplete inhibition of gene expression. To overcome these limitations, we developed a novel arrayed CRISPR screen based on a plasmid library expressing a single guide RNA (sgRNA) and disrupted 1514 genes, encoding kinases, proteins related to endocytosis, and Golgi-localized proteins, individually using 4542 sgRNAs (three sgRNAs per gene). This screen revealed host factors required for infection by coxsackievirus B3 (CVB3) from Picornaviridae, which includes human pathogens causing diverse diseases. Many host factors that had been overlooked in a conventional pooled screen were identified for CVB3 infection, including entry-related factors, translational initiation factors, and several replication factors with different functions, demonstrating the advantage of the arrayed screen. This screen was quite reliable and reproducible, as most genes identified in the primary screen were confirmed in secondary screens. Moreover, ACBD3, whose phenotype was not affected by siRNA-mediated knockdown, was reliably identified. We propose that arrayed CRISPR screens based on sgRNA plasmid libraries are powerful tools for arrayed genetic screening and applicable to larger-scale screens.

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Section: Discussionmentioning

confidence: 99%

Arrayed CRISPR screen with image-based assay reliably uncovers host genes required for coxsackievirus infection

Lee

Kim

et al. 2018

Genome Res.

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“…This same double-target approach was used against several other viruses that included clinically important RSV, HSV1 (87,88), the picornavirus family (89) and influenza viruses (90–92). Oligomers were targeted to the AUG sites or the highly conserved terminal sequence of appropriate genes.…”

Section: Rna-blocking Oligonucleotides For Other Indicationsmentioning

confidence: 99%

RNA therapeutics: beyond RNA interference and antisense oligonucleotides

Kole

Krainer

Altman³

2012

Nat Rev Drug Discov

1,041

828

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Here we discuss three RNA therapeutic technologies exploiting various oligonucleotides that bind RNA by base-pairing in a sequence-specific manner yet have different mechanisms of action and effects. RNA interference and antisense oligonucleotides downregulate gene expression by enzyme-dependent degradation of targeted mRNA. Steric blocking oligonucleotides block access of cellular machinery to pre-mRNA and mRNA without degrading the RNA. Through this mechanism, blocking oligonucleotides can redirect alternative splicing, repair defective RNA, restore protein production or also downregulate gene expression. Moreover, they can be extensively chemically modified, resulting in more drug-like properties. The ability of RNA blocking oligonucleotides to restore gene function makes them suited for treatment of genetic disorders. Positive results from clinical trials for the treatment of Duchenne muscular dystrophy show that this technology is close to realizing its clinical potential.

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“…These arginine-based peptides with the 6-aminohexanoicspaced oligoarginine (R-Ahx-R)4 have successfully delivered Morpholinos to the nucleus and cytosol in vitro (11) and in vivo (12), including sustained induction of dystrophin expression in mdx mice (13). They have also been shown to actively or prophy-lactically knock down viral titers in various tissues of mice infected with corona virus (14), picornavirus (15), respiratory syncytial virus (16), and influenza A (17). However, it should be noted that arginine-based peptides are not generally available to the research community and that their greatest successes have been in delivering Morpholinos to the cytosol of what would be considered easily deliverable tissues like liver (18) or leaky muscle (19).…”

Section: Introductionmentioning

confidence: 99%

Vivo-Morpholinos: A Non-Peptide Transporter Delivers Morpholinos into a Wide Array of Mouse Tissues

2008

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We have developed a new transporter structure that provides effective delivery of Morpholino antisense oligomers into a wide variety of tissues in living mice. This transporter comprises a dendritic structure assembled around a triazine core which serves to position eight guanidinium head groups in a conformation effective to penetrate cell membranes. This transporter structure is conjugated to a Morpholino oligomer to form a delivery-enabled product referred to as a Vivo-Morpholino. Vivo-Morpholinos are shown to effectively enter and function within cultured cells in the presence of 100% serum using a rigorous positive test system based on correction of a defined splicing error in a pre-messenger RNA. In addition, Vivo-Morpholinos are demonstrated to enter into a wide variety of tissues in a similar positive test system in transgenic mice, as evidenced by correction of the targeted splicing error in all tissues assessed, including near-complete splice correction in the small intestine, colon, stomach, liver kidney, and a number of muscles. Finally, Vivo-Morpholinos, which target the exon-skipping of exon 23 harboring a premature termination codon in the mdx mouse model, effectively restore the reading frame of dystrophin and restore expression of a functional dystrophin protein.

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A Morpholino Oligomer Targeting Highly Conserved Internal Ribosome Entry Site Sequence Is Able To Inhibit Multiple Species of Picornavirus

Abstract: Members of the genera Enterovirus and Rhinovirus (family

Cited by 40 publications

References 58 publications

Arrayed CRISPR screen with image-based assay reliably uncovers host genes required for coxsackievirus infection

Arrayed CRISPR screen with image-based assay reliably uncovers host genes required for coxsackievirus infection

RNA therapeutics: beyond RNA interference and antisense oligonucleotides

Vivo-Morpholinos: A Non-Peptide Transporter Delivers Morpholinos into a Wide Array of Mouse Tissues

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