Your browser doesn't support javascript.
Show: 20 | 50 | 100
Results 1 - 9 de 9
Filter
1.
Nat Commun ; 13(1): 4503, 2022 Aug 03.
Article in English | MEDLINE | ID: covidwho-1972603

ABSTRACT

The COVID-19 pandemic is exacting an increasing toll worldwide, with new SARS-CoV-2 variants emerging that exhibit higher infectivity rates and that may partially evade vaccine and antibody immunity. Rapid deployment of non-invasive therapeutic avenues capable of preventing infection by all SARS-CoV-2 variants could complement current vaccination efforts and help turn the tide on the COVID-19 pandemic. Here, we describe a novel therapeutic strategy targeting the SARS-CoV-2 RNA using locked nucleic acid antisense oligonucleotides (LNA ASOs). We identify an LNA ASO binding to the 5' leader sequence of SARS-CoV-2 that disrupts a highly conserved stem-loop structure with nanomolar efficacy in preventing viral replication in human cells. Daily intranasal administration of this LNA ASO in the COVID-19 mouse model potently suppresses viral replication (>80-fold) in the lungs of infected mice. We find that the LNA ASO is efficacious in countering all SARS-CoV-2 "variants of concern" tested both in vitro and in vivo. Hence, inhaled LNA ASOs targeting SARS-CoV-2 represents a promising therapeutic approach to reduce or prevent transmission and decrease severity of COVID-19 in infected individuals. LNA ASOs are chemically stable and can be flexibly modified to target different viral RNA sequences and could be stockpiled for future coronavirus pandemics.


Subject(s)
COVID-19 , SARS-CoV-2 , Administration, Intranasal , Animals , Humans , Mice , Oligonucleotides, Antisense/pharmacology , Oligonucleotides, Antisense/therapeutic use , Pandemics/prevention & control , RNA, Viral/genetics
2.
Proc Natl Acad Sci U S A ; 119(9)2022 03 01.
Article in English | MEDLINE | ID: covidwho-1684241

ABSTRACT

SARS-CoV-2 is a highly pathogenic virus that evades antiviral immunity by interfering with host protein synthesis, mRNA stability, and protein trafficking. The SARS-CoV-2 nonstructural protein 1 (Nsp1) uses its C-terminal domain to block the messenger RNA (mRNA) entry channel of the 40S ribosome to inhibit host protein synthesis. However, how SARS-CoV-2 circumvents Nsp1-mediated suppression for viral protein synthesis and if the mechanism can be targeted therapeutically remain unclear. Here, we show that N- and C-terminal domains of Nsp1 coordinate to drive a tuned ratio of viral to host translation, likely to maintain a certain level of host fitness while maximizing replication. We reveal that the stem-loop 1 (SL1) region of the SARS-CoV-2 5' untranslated region (5' UTR) is necessary and sufficient to evade Nsp1-mediated translational suppression. Targeting SL1 with locked nucleic acid antisense oligonucleotides inhibits viral translation and makes SARS-CoV-2 5' UTR vulnerable to Nsp1 suppression, hindering viral replication in vitro at a nanomolar concentration, as well as providing protection against SARS-CoV-2-induced lethality in transgenic mice expressing human ACE2. Thus, SL1 allows Nsp1 to switch infected cells from host to SARS-CoV-2 translation, presenting a therapeutic target against COVID-19 that is conserved among immune-evasive variants. This unique strategy of unleashing a virus' own virulence mechanism against itself could force a critical trade-off between drug resistance and pathogenicity.


Subject(s)
5' Untranslated Regions/genetics , Immune Evasion/genetics , Protein Biosynthesis , SARS-CoV-2/genetics , Viral Nonstructural Proteins/genetics , Animals , Base Sequence , Chlorocebus aethiops , HEK293 Cells , Host-Pathogen Interactions/drug effects , Host-Pathogen Interactions/genetics , Humans , Immune Evasion/drug effects , Mice, Transgenic , Models, Biological , Oligonucleotides, Antisense/pharmacology , Protein Biosynthesis/drug effects , SARS-CoV-2/drug effects , Vero Cells , Virus Replication/drug effects
3.
Molecules ; 27(2)2022 Jan 15.
Article in English | MEDLINE | ID: covidwho-1625662

ABSTRACT

Antisense oligonucleotides (ASOs) are an increasingly represented class of drugs. These small sequences of nucleotides are designed to precisely target other oligonucleotides, usually RNA species, and are modified to protect them from degradation by nucleases. Their specificity is due to their sequence, so it is possible to target any RNA sequence that is already known. These molecules are very versatile and adaptable given that their sequence and chemistry can be custom manufactured. Based on the chemistry being used, their activity may significantly change and their effects on cell function and phenotypes can differ dramatically. While some will cause the target RNA to decay, others will only bind to the target and act as a steric blocker. Their incredible versatility is the key to manipulating several aspects of nucleic acid function as well as their process, and alter the transcriptome profile of a specific cell type or tissue. For example, they can be used to modify splicing or mask specific sites on a target. The entire design rather than just the sequence is essential to ensuring the specificity of the ASO to its target. Thus, it is vitally important to ensure that the complete process of drug design and testing is taken into account. ASOs' adaptability is a considerable advantage, and over the past decades has allowed multiple new drugs to be approved. This, in turn, has had a significant and positive impact on patient lives. Given current challenges presented by the COVID-19 pandemic, it is necessary to find new therapeutic strategies that would complement the vaccination efforts being used across the globe. ASOs may be a very powerful tool that can be used to target the virus RNA and provide a therapeutic paradigm. The proof of the efficacy of ASOs as an anti-viral agent is long-standing, yet no molecule currently has FDA approval. The emergence and widespread use of RNA vaccines during this health crisis might provide an ideal opportunity to develop the first anti-viral ASOs on the market. In this review, we describe the story of ASOs, the different characteristics of their chemistry, and how their characteristics translate into research and as a clinical tool.


Subject(s)
Drug Development/methods , Oligonucleotides, Antisense/chemistry , Oligonucleotides, Antisense/pharmacology , Animals , COVID-19/therapy , Drug Approval , Drug Design , Humans , Oligonucleotides, Antisense/therapeutic use , SARS-CoV-2/drug effects , United States , United States Food and Drug Administration
4.
Wiley Interdiscip Rev RNA ; 13(4): e1703, 2022 07.
Article in English | MEDLINE | ID: covidwho-1540186

ABSTRACT

The COVID-19 crisis and the development of the first approved mRNA vaccine have highlighted the power of RNA-based therapeutic strategies for the development of new medicines. Aside from RNA-vaccines, antisense oligonucleotides (ASOs) represent a new and very promising class of RNA-targeted therapy. Few drugs have already received approval from the Food and Drug Administration. Here, we underscored why and how ASOs hold the potential to change the therapeutic landscape to beat SARS-CoV-2 viral infections. This article is categorized under: RNA Interactions with Proteins and Other Molecules > Small Molecule-RNA Interactions.


Subject(s)
COVID-19 , Oligonucleotides, Antisense , COVID-19/drug therapy , Humans , Oligonucleotides , Oligonucleotides, Antisense/genetics , Oligonucleotides, Antisense/pharmacology , RNA , SARS-CoV-2 , United States , Vaccines, Synthetic , mRNA Vaccines
5.
Nat Commun ; 12(1): 4396, 2021 07 20.
Article in English | MEDLINE | ID: covidwho-1387353

ABSTRACT

Rapid development of antisense therapies can enable on-demand responses to new viral pathogens and make personalized medicine for genetic diseases practical. Antisense phosphorodiamidate morpholino oligomers (PMOs) are promising candidates to fill such a role, but their challenging synthesis limits their widespread application. To rapidly prototype potential PMO drug candidates, we report a fully automated flow-based oligonucleotide synthesizer. Our optimized synthesis platform reduces coupling times by up to 22-fold compared to previously reported methods. We demonstrate the power of our automated technology with the synthesis of milligram quantities of three candidate therapeutic PMO sequences for an unserved class of Duchenne muscular dystrophy (DMD). To further test our platform, we synthesize a PMO that targets the genomic mRNA of SARS-CoV-2 and demonstrate its antiviral effects. This platform could find broad application not only in designing new SARS-CoV-2 and DMD antisense therapeutics, but also for rapid development of PMO candidates to treat new and emerging diseases.


Subject(s)
Chemistry Techniques, Synthetic/instrumentation , Chemistry, Pharmaceutical/instrumentation , High-Throughput Screening Assays/instrumentation , Morpholinos/chemical synthesis , Oligonucleotides, Antisense/chemical synthesis , Animals , COVID-19/drug therapy , COVID-19/virology , Chlorocebus aethiops , Communicable Diseases, Emerging/drug therapy , Communicable Diseases, Emerging/microbiology , Disease Models, Animal , High-Throughput Screening Assays/methods , Humans , Morpholinos/pharmacology , Morpholinos/therapeutic use , Muscular Dystrophy, Duchenne/drug therapy , Muscular Dystrophy, Duchenne/genetics , Oligonucleotides, Antisense/pharmacology , Oligonucleotides, Antisense/therapeutic use , Precision Medicine/methods , RNA, Messenger/antagonists & inhibitors , RNA, Viral/antagonists & inhibitors , SARS-CoV-2/genetics , Time Factors , Vero Cells
6.
Nat Struct Mol Biol ; 28(9): 747-754, 2021 09.
Article in English | MEDLINE | ID: covidwho-1370728

ABSTRACT

Drug discovery campaigns against COVID-19 are beginning to target the SARS-CoV-2 RNA genome. The highly conserved frameshift stimulation element (FSE), required for balanced expression of viral proteins, is a particularly attractive SARS-CoV-2 RNA target. Here we present a 6.9 Å resolution cryo-EM structure of the FSE (88 nucleotides, ~28 kDa), validated through an RNA nanostructure tagging method. The tertiary structure presents a topologically complex fold in which the 5' end is threaded through a ring formed inside a three-stem pseudoknot. Guided by this structure, we develop antisense oligonucleotides that impair FSE function in frameshifting assays and knock down SARS-CoV-2 virus replication in A549-ACE2 cells at 100 nM concentration.


Subject(s)
COVID-19/prevention & control , Cryoelectron Microscopy/methods , Frameshift Mutation/genetics , Oligonucleotides, Antisense/genetics , RNA, Viral/genetics , Response Elements/genetics , SARS-CoV-2/genetics , A549 Cells , Animals , Base Sequence , COVID-19/virology , Cell Line, Tumor , Chlorocebus aethiops , Genome, Viral/genetics , Humans , Models, Molecular , Nucleic Acid Conformation , Oligonucleotides, Antisense/pharmacology , RNA, Viral/chemistry , RNA, Viral/ultrastructure , SARS-CoV-2/physiology , SARS-CoV-2/ultrastructure , Vero Cells , Virus Replication/drug effects , Virus Replication/genetics
7.
Angew Chem Int Ed Engl ; 60(40): 21662-21667, 2021 09 27.
Article in English | MEDLINE | ID: covidwho-1363645

ABSTRACT

There is an urgent need to develop antiviral drugs and alleviate the current COVID-19 pandemic. Herein we report the design and construction of chimeric oligonucleotides comprising a 2'-OMe-modified antisense oligonucleotide and a 5'-phosphorylated 2'-5' poly(A)4 (4A2-5 ) to degrade envelope and spike RNAs of SARS-CoV-2. The oligonucleotide was used for searching and recognizing target viral RNA sequence, and the conjugated 4A2-5 was used for guided RNase L activation to sequence-specifically degrade viral RNAs. Since RNase L can potently cleave single-stranded RNA during innate antiviral response, degradation efficiencies with these chimeras were twice as much as those with only antisense oligonucleotides for both SARS-CoV-2 RNA targets. In pseudovirus infection models, chimera-S4 achieved potent and broad-spectrum inhibition of SARS-CoV-2 and its N501Y and/or ΔH69/ΔV70 mutants, indicating a promising antiviral agent based on the nucleic acid-hydrolysis targeting chimera (NATAC) strategy.


Subject(s)
Antiviral Agents/pharmacology , Endoribonucleases/metabolism , Enzyme Activation/drug effects , Oligonucleotides, Antisense/pharmacology , SARS-CoV-2/drug effects , Animals , Chlorocebus aethiops , Coronavirus Envelope Proteins/genetics , Drug Design , HEK293 Cells , Humans , Hydrolysis/drug effects , Microbial Sensitivity Tests , Mutation , RNA, Viral/metabolism , Spike Glycoprotein, Coronavirus/genetics , Vero Cells
8.
Pharmacol Rev ; 72(4): 862-898, 2020 10.
Article in English | MEDLINE | ID: covidwho-767668

ABSTRACT

RNA-based therapies, including RNA molecules as drugs and RNA-targeted small molecules, offer unique opportunities to expand the range of therapeutic targets. Various forms of RNAs may be used to selectively act on proteins, transcripts, and genes that cannot be targeted by conventional small molecules or proteins. Although development of RNA drugs faces unparalleled challenges, many strategies have been developed to improve RNA metabolic stability and intracellular delivery. A number of RNA drugs have been approved for medical use, including aptamers (e.g., pegaptanib) that mechanistically act on protein target and small interfering RNAs (e.g., patisiran and givosiran) and antisense oligonucleotides (e.g., inotersen and golodirsen) that directly interfere with RNA targets. Furthermore, guide RNAs are essential components of novel gene editing modalities, and mRNA therapeutics are under development for protein replacement therapy or vaccination, including those against unprecedented severe acute respiratory syndrome coronavirus pandemic. Moreover, functional RNAs or RNA motifs are highly structured to form binding pockets or clefts that are accessible by small molecules. Many natural, semisynthetic, or synthetic antibiotics (e.g., aminoglycosides, tetracyclines, macrolides, oxazolidinones, and phenicols) can directly bind to ribosomal RNAs to achieve the inhibition of bacterial infections. Therefore, there is growing interest in developing RNA-targeted small-molecule drugs amenable to oral administration, and some (e.g., risdiplam and branaplam) have entered clinical trials. Here, we review the pharmacology of novel RNA drugs and RNA-targeted small-molecule medications, with a focus on recent progresses and strategies. Challenges in the development of novel druggable RNA entities and identification of viable RNA targets and selective small-molecule binders are discussed. SIGNIFICANCE STATEMENT: With the understanding of RNA functions and critical roles in diseases, as well as the development of RNA-related technologies, there is growing interest in developing novel RNA-based therapeutics. This comprehensive review presents pharmacology of both RNA drugs and RNA-targeted small-molecule medications, focusing on novel mechanisms of action, the most recent progress, and existing challenges.


Subject(s)
RNA/drug effects , RNA/pharmacology , Aptamers, Nucleotide/pharmacology , Aptamers, Nucleotide/therapeutic use , Betacoronavirus , COVID-19 , Chemistry Techniques, Analytical/methods , Chemistry Techniques, Analytical/standards , Clustered Regularly Interspaced Short Palindromic Repeats , Coronavirus Infections/drug therapy , Drug Delivery Systems/methods , Drug Development/organization & administration , Drug Discovery , Humans , MicroRNAs/pharmacology , MicroRNAs/therapeutic use , Oligonucleotides, Antisense/pharmacology , Oligonucleotides, Antisense/therapeutic use , Pandemics , Pneumonia, Viral/drug therapy , RNA/adverse effects , RNA, Antisense/pharmacology , RNA, Antisense/therapeutic use , RNA, Guide/pharmacology , RNA, Guide/therapeutic use , RNA, Messenger/drug effects , RNA, Messenger/pharmacology , RNA, Ribosomal/drug effects , RNA, Ribosomal/pharmacology , RNA, Small Interfering/pharmacology , RNA, Small Interfering/therapeutic use , RNA, Viral/drug effects , Ribonucleases/metabolism , Riboswitch/drug effects , SARS-CoV-2
9.
Clin Sci (Lond) ; 134(10): 1143-1150, 2020 05 29.
Article in English | MEDLINE | ID: covidwho-342905

ABSTRACT

Angiotensin-converting enzyme 2 (ACE2) plays an essential role in maintaining the balance of the renin-angiotensin system and also serves as a receptor for the SARS-CoV-2, SARS-CoV, and HCoV-NL63. Following the recent outbreak of SARS-CoV-2 infection, there has been an urgent need to develop therapeutic interventions. ACE2 is a potential target for many treatment approaches for the SARS-CoV-2. With the help of bioinformatics, we have predicted several novel exons of the human ACE2 gene. The inclusion of novel exons located in the 5'UTR/intronic region in the mature transcript may remove the critical ACE2 residues responsible for the interaction with the receptor-binding domain (RBD) of SARS-CoV-2, thus preventing their binding and entry into the cell. Additionally, inclusion of a novel predicted exons located in the 3'UTR by alternative splicing may remove the C-terminal transmembrane domain of ACE2 and generate soluble ACE2 isoforms. Splice-switching antisense oligonucleotides (SSOs) have been employed effectively as a therapeutic strategy in several disease conditions. Alternative splicing of the ACE2 gene could similarly be modulated using SSOs to exclude critical domains required for the entry of SARS-CoV-2. Strategies can also be designed to deliver these SSOs directly to the lungs in order to minimize the damage caused by SARS-CoV-2 pathogenesis.


Subject(s)
Alternative Splicing , Coronavirus Infections/genetics , Oligonucleotides, Antisense/pharmacology , Peptidyl-Dipeptidase A/genetics , Pneumonia, Viral/genetics , Virus Internalization , Angiotensin-Converting Enzyme 2 , Betacoronavirus/physiology , COVID-19 , Computational Biology , Coronavirus Infections/therapy , Exons , Humans , Models, Molecular , Pandemics , Pneumonia, Viral/therapy , Protein Domains , Receptors, Virus/genetics , SARS-CoV-2
SELECTION OF CITATIONS
SEARCH DETAIL