Nucleic acid therapeutics最新文献

{"title":"Positive Allosteric Modulation of Antithrombin's Inhibitory Activity by RNA Aptamers.","authors":"Khalequz Zaman,&nbsp;Adi Breitman,&nbsp;Isa Malik,&nbsp;Yolanda M Fortenberry","doi":"10.1089/nat.2022.0047","DOIUrl":"https://doi.org/10.1089/nat.2022.0047","url":null,"abstract":"<p><p>The leading cause of death in adults in the United States is cardiovascular disease, with mortality and morbidity mainly attributed to thromboembolism. Heparin is the most common therapy used for treating venous and arterial thrombosis. Heparin effectively accelerates the inhibition of coagulation proteases thrombin and factor Xa through the serine protease inhibitor (serpin) antithrombin (AT). Heparin is an essential therapeutic anticoagulant because of its effectiveness and the availability of protamine sulfate as an antidote. However, heparin therapy has several limitations. Thus, new anticoagulants, including direct thrombin inhibitors (ie, argatroban) and low-molecular-weight heparins (ie, fondaparinux), are used to treat some thromboembolic disorders. We developed and characterized a family of novel RNA-based aptamers that bind AT using two novel selection schemes. One of the aptamers, AT-16, accelerates factor Xa inhibition by AT in the absence of heparin. AT-16's effect on thrombin inhibition by AT is less effective compared to factor Xa. AT-16 induces a conformational change in AT that is different from that induced by heparin. This study demonstrates that an AT-specific RNA aptamer, AT-16, exhibits a positive allosteric modulator effect on AT's inhibition of factor Xa.</p>","PeriodicalId":19412,"journal":{"name":"Nucleic acid therapeutics","volume":"33 4","pages":"277-286"},"PeriodicalIF":4.0,"publicationDate":"2023-08-01","publicationTypes":"Journal Article","fieldsOfStudy":null,"isOpenAccess":false,"openAccessPdf":"","citationCount":null,"resultStr":null,"platform":"Semanticscholar","paperid":"10028836","PeriodicalName":null,"FirstCategoryId":null,"ListUrlMain":null,"RegionNum":2,"RegionCategory":"医学","ArticlePicture":[],"TitleCN":null,"AbstractTextCN":null,"PMCID":"","EPubDate":null,"PubModel":null,"JCR":null,"JCRName":null,"Score":null,"Total":0}

{"title":"mTOR Inhibition Enhances Delivery and Activity of Antisense Oligonucleotides in Uveal Melanoma Cells.","authors":"Shanna Dewaele,&nbsp;Louis Delhaye,&nbsp;Boel De Paepe,&nbsp;Bram Bogaert,&nbsp;Ramiro Martinez,&nbsp;Jasper Anckaert,&nbsp;Nurten Yigit,&nbsp;Justine Nuytens,&nbsp;Rudy Van Coster,&nbsp;Sven Eyckerman,&nbsp;Koen Raemdonck,&nbsp;Pieter Mestdagh","doi":"10.1089/nat.2023.0008","DOIUrl":"https://doi.org/10.1089/nat.2023.0008","url":null,"abstract":"<p><p>Uveal melanoma (UM) is the most common primary intraocular malignancy in adults. Owing to a lack of effective treatments, patients with metastatic disease have a median survival time of 6-12 months. We recently demonstrated that the Survival Associated Mitochondrial Melanoma Specific Oncogenic Non-coding RNA <i>(SAMMSON)</i> is essential for UM cell survival and that antisense oligonucleotide (ASO)-mediated silencing of <i>SAMMSON</i> impaired cell viability and tumor growth <i>in vitro</i> and <i>in vivo</i>. By screening a library of 2911 clinical stage compounds, we identified the mammalian target of rapamycin (mTOR) inhibitor GDC-0349 to synergize with <i>SAMMSON</i> inhibition in UM. Mechanistic studies revealed that mTOR inhibition enhanced uptake and reduced lysosomal accumulation of lipid complexed <i>SAMMSON</i> ASOs, improving <i>SAMMSON</i> knockdown and further decreasing UM cell viability. We found mTOR inhibition to also enhance target knockdown in other cancer cell lines as well as normal cells when combined with lipid nanoparticle complexed or encapsulated ASOs or small interfering RNAs (siRNAs). Our results are relevant to nucleic acid treatment in general and highlight the potential of mTOR inhibition to enhance ASO and siRNA-mediated target knockdown.</p>","PeriodicalId":19412,"journal":{"name":"Nucleic acid therapeutics","volume":"33 4","pages":"248-264"},"PeriodicalIF":4.0,"publicationDate":"2023-08-01","publicationTypes":"Journal Article","fieldsOfStudy":null,"isOpenAccess":false,"openAccessPdf":"","citationCount":null,"resultStr":null,"platform":"Semanticscholar","paperid":"10020238","PeriodicalName":null,"FirstCategoryId":null,"ListUrlMain":null,"RegionNum":2,"RegionCategory":"医学","ArticlePicture":[],"TitleCN":null,"AbstractTextCN":null,"PMCID":"","EPubDate":null,"PubModel":null,"JCR":null,"JCRName":null,"Score":null,"Total":0}

{"title":"Experimental Model Systems Used in the Preclinical Development of Nucleic Acid Therapeutics.","authors":"Haiyan Zhou, Virginia Arechavala-Gomeza, Alejandro Garanto","doi":"10.1089/nat.2023.0001","DOIUrl":"10.1089/nat.2023.0001","url":null,"abstract":"<p><p>Preclinical evaluation of nucleic acid therapeutics (NATs) in relevant experimental model systems is essential for NAT drug development. As part of COST Action \"DARTER\" (Delivery of Antisense RNA ThERapeutics), a network of researchers in the field of RNA therapeutics, we have conducted a survey on the experimental model systems routinely used by our members in preclinical NAT development. The questionnaire focused on both cellular and animal models. Our survey results suggest that skin fibroblast cultures derived from patients is the most commonly used cellular model, while induced pluripotent stem cell-derived models are also highly reported, highlighting the increasing potential of this technology. Splice-switching antisense oligonucleotide is the most frequently investigated RNA molecule, followed by small interfering RNA. Animal models are less prevalent but also widely used among groups in the network, with transgenic mouse models ranking the top. Concerning the research fields represented in our survey, the mostly studied disease area is neuromuscular disorders, followed by neurometabolic diseases and cancers. Brain, skeletal muscle, heart, and liver are the top four tissues of interest reported. We expect that this snapshot of the current preclinical models will facilitate decision making and the share of resources between academics and industry worldwide to facilitate the development of NATs.</p>","PeriodicalId":19412,"journal":{"name":"Nucleic acid therapeutics","volume":"33 4","pages":"238-247"},"PeriodicalIF":4.0,"publicationDate":"2023-08-01","publicationTypes":"Journal Article","fieldsOfStudy":null,"isOpenAccess":false,"openAccessPdf":"https://www.ncbi.nlm.nih.gov/pmc/articles/PMC10457615/pdf/","citationCount":null,"resultStr":null,"platform":"Semanticscholar","paperid":"10472910","PeriodicalName":null,"FirstCategoryId":null,"ListUrlMain":null,"RegionNum":2,"RegionCategory":"医学","ArticlePicture":[],"TitleCN":null,"AbstractTextCN":null,"PMCID":"OA","EPubDate":null,"PubModel":null,"JCR":null,"JCRName":null,"Score":null,"Total":0}

{"title":"Considerations for the Terminal Sterilization of Oligonucleotide Drug Products.","authors":"Daniel Paul DeCollibus,&nbsp;Justin Searcy,&nbsp;Anna Tivesten,&nbsp;Nadim Akhtar,&nbsp;Christian Lindenberg,&nbsp;Nounja Abarrou,&nbsp;Sujana Pradhan,&nbsp;Maggie Fiandaca,&nbsp;Jenny Franklin,&nbsp;Geetha Govindan,&nbsp;Hung-Yi Liu,&nbsp;David Royle,&nbsp;Patrick Lim Soo,&nbsp;Kirsten Storch","doi":"10.1089/nat.2022.0073","DOIUrl":"https://doi.org/10.1089/nat.2022.0073","url":null,"abstract":"<p><p>A primary function of the parenteral drug product manufacturing process is to ensure sterility of the final product. The two most common methods for sterilizing parenteral drug products are terminal sterilization (TS), whereby the drug product is sterilized in the final container following filling and finish, and membrane sterilization, whereby the product stream is sterilized by membrane filtration and filled into presterilized containers in an aseptic processing environment. Although TS provides greater sterility assurance than membrane sterilization and aseptic processing, not all drug products are amenable to TS processes, which typically involve heat treatment or exposure to ionizing radiation. Oligonucleotides represent an emerging class of therapeutics with great potential for treating a broad range of indications, including previously undruggable targets. Owing to their size, structural complexity, and relative lack of governing regulations, several challenges in drug development are unique to oligonucleotides. This exceptionality justifies a focused assessment of traditional chemistry, manufacturing, and control strategies before their adoption. In this article, we review the current state of sterile oligonucleotide drug product processing, highlight the key aspects to consider when assessing options for product sterilization, and provide recommendations to aid in the successful evaluation and development of TS processes. We also explore current regulatory expectations and provide our interpretation as it pertains to oligonucleotide drug products.</p>","PeriodicalId":19412,"journal":{"name":"Nucleic acid therapeutics","volume":"33 3","pages":"159-177"},"PeriodicalIF":4.0,"publicationDate":"2023-06-01","publicationTypes":"Journal Article","fieldsOfStudy":null,"isOpenAccess":false,"openAccessPdf":"https://www.ncbi.nlm.nih.gov/pmc/articles/PMC10277985/pdf/","citationCount":null,"resultStr":null,"platform":"Semanticscholar","paperid":"10021709","PeriodicalName":null,"FirstCategoryId":null,"ListUrlMain":null,"RegionNum":2,"RegionCategory":"医学","ArticlePicture":[],"TitleCN":null,"AbstractTextCN":null,"PMCID":"OA","EPubDate":null,"PubModel":null,"JCR":null,"JCRName":null,"Score":null,"Total":0}

{"title":"DNAzymes: Expanding the Potential of Nucleic Acid Therapeutics.","authors":"Leon M Larcher,&nbsp;Ianthe L Pitout,&nbsp;Niall P Keegan,&nbsp;Rakesh N Veedu,&nbsp;Sue Fletcher","doi":"10.1089/nat.2022.0066","DOIUrl":"https://doi.org/10.1089/nat.2022.0066","url":null,"abstract":"<p><p>Nucleic acids drugs have been proven in the clinic as a powerful modality to treat inherited and acquired diseases. However, key challenges including drug stability, renal clearance, cellular uptake, and movement across biological barriers (foremost the blood-brain barrier) limit the translation and clinical efficacy of nucleic acid-based therapies, both systemically and in the central nervous system. In this study we provide an overview of an emerging class of nucleic acid therapeutic, called DNAzymes. In particular, we review the use of chemical modifications and carrier molecules for the stabilization and/or delivery of DNAzymes in cell and animal models. Although this review focuses on DNAzymes, the strategies described are broadly applicable to most nucleic acid technologies. This review should serve as a general guide for selecting chemical modifications to improve the therapeutic performance of DNAzymes.</p>","PeriodicalId":19412,"journal":{"name":"Nucleic acid therapeutics","volume":"33 3","pages":"178-192"},"PeriodicalIF":4.0,"publicationDate":"2023-06-01","publicationTypes":"Journal Article","fieldsOfStudy":null,"isOpenAccess":false,"openAccessPdf":"https://www.ncbi.nlm.nih.gov/pmc/articles/PMC10278027/pdf/","citationCount":null,"resultStr":null,"platform":"Semanticscholar","paperid":"9670408","PeriodicalName":null,"FirstCategoryId":null,"ListUrlMain":null,"RegionNum":2,"RegionCategory":"医学","ArticlePicture":[],"TitleCN":null,"AbstractTextCN":null,"PMCID":"OA","EPubDate":null,"PubModel":null,"JCR":null,"JCRName":null,"Score":null,"Total":0}

{"title":"Rapid and Reliable Quantification of Prime Editing Targeting Within the Porcine <i>ABCA4</i> Gene Using a BRET-Based Sensor.","authors":"Tobias Wimmer,&nbsp;Hannah Sawinski,&nbsp;Anne M Urban,&nbsp;Jan Motlik,&nbsp;Knut Stieger","doi":"10.1089/nat.2022.0037","DOIUrl":"https://doi.org/10.1089/nat.2022.0037","url":null,"abstract":"Stargardt disease (STGD) leads to blindness in children and young adults. So far, no curative therapy is available and gene augmentation therapies have not yet advanced to the clinics, in part, due to the limited packaging capacity of adeno-associated viruses used to transfer genes into photoreceptor cells. Prime editing offers a new perspective to treat mutations on the genomic level. A nicking variant of Cas9 fused to a reverse transcriptase complex with an elongated guideRNA force intracellular mismatch repair to correct the targeted mutation even in postmitotic cells such as photoreceptors in the eye. Using a custom-made bioluminescence resonance energy transfer (BRET)-based editing sensor in HEK293 cells, we tested 27 different prime editing guide RNAs (pegRNAs) and additional 4 nicking guide RNAs (ngRNAs) with regard to their efficiency to induce sequences changes in exon 43 of the porcine ATP binding cassette subfamily A member 4 (ABCA4) gene that eliminate a mutagenic adenine frameshift insertion, which has been associated with STGD in humans. We identified nine working pegRNAs, and in combination with ngRNAs, we achieved a correction rate of up to ≈92% measured with the BRET-based reporter system. Our data prove the high efficiency of prime editors to correct mutations and highlight the importance of optimal ngRNA design, thus offering a promising editing tool to correct ABCA4 mutations in the disease context.","PeriodicalId":19412,"journal":{"name":"Nucleic acid therapeutics","volume":"33 3","pages":"226-232"},"PeriodicalIF":4.0,"publicationDate":"2023-06-01","publicationTypes":"Journal Article","fieldsOfStudy":null,"isOpenAccess":false,"openAccessPdf":"https://www.ncbi.nlm.nih.gov/pmc/articles/PMC10278032/pdf/","citationCount":null,"resultStr":null,"platform":"Semanticscholar","paperid":"9666363","PeriodicalName":null,"FirstCategoryId":null,"ListUrlMain":null,"RegionNum":2,"RegionCategory":"医学","ArticlePicture":[],"TitleCN":null,"AbstractTextCN":null,"PMCID":"OA","EPubDate":null,"PubModel":null,"JCR":null,"JCRName":null,"Score":null,"Total":0}

{"title":"Next Generation Exon 51 Skipping Antisense Oligonucleotides for Duchenne Muscular Dystrophy.","authors":"Judith van Deutekom,&nbsp;Chantal Beekman,&nbsp;Suzanne Bijl,&nbsp;Sieto Bosgra,&nbsp;Rani van den Eijnde,&nbsp;Dennis Franken,&nbsp;Bas Groenendaal,&nbsp;Bouchra Harquouli,&nbsp;Anneke Janson,&nbsp;Paul Koevoets,&nbsp;Melissa Mulder,&nbsp;Daan Muilwijk,&nbsp;Galyna Peterburgska,&nbsp;Bianca Querido,&nbsp;Janwillem Testerink,&nbsp;Ruurd Verheul,&nbsp;Peter de Visser,&nbsp;Rudie Weij,&nbsp;Annemieke Aartsma-Rus,&nbsp;Jukka Puoliväli,&nbsp;Timo Bragge,&nbsp;Charles O'Neill,&nbsp;Nicole A Datson","doi":"10.1089/nat.2022.0063","DOIUrl":"https://doi.org/10.1089/nat.2022.0063","url":null,"abstract":"<p><p>In the last two decades, antisense oligonucleotides (AONs) that induce corrective exon skipping have matured as promising therapies aimed at tackling the dystrophin deficiency that underlies the severe and progressive muscle fiber degeneration in Duchenne muscular dystrophy (DMD) patients. Pioneering first generation exon 51 skipping AONs like drisapersen and eteplirsen have more recently been followed up by AONs for exons 53 and 45, with, to date, a total of four exon skipping AON drugs having reached (conditional) regulatory US Food and Drug Administration (FDA) approval for DMD. Nonetheless, considering the limited efficacy of these drugs, there is room for improvement. The aim of this study was to develop more efficient [2'-<i>O</i>-methyl-modified phosphorothioate (2'OMePS) RNA] AONs for <i>DMD</i> exon 51 skipping by implementing precision chemistry as well as identifying a more potent target binding site. More than a hundred AONs were screened in muscle cell cultures, followed by a selective comparison in the hDMD and hDMDdel52/<i>mdx</i> mouse models. Incorporation of 5-methylcytosine and position-specific locked nucleic acids in AONs targeting the drisapersen/eteplirsen binding site resulted in 15-fold higher exon 51 skipping levels compared to drisapersen in hDMDdel52/<i>mdx</i> mice. However, with similarly modified AONs targeting an alternative site in exon 51, 65-fold higher skipping levels were obtained, restoring dystrophin up to 30% of healthy control. Targeting both sites in exon 51 with a single AON further increased exon skipping (100-fold over drisapersen) and dystrophin (up to 40%) levels. These dystrophin levels allowed for normalization of creatine kinase (CK) and lactate dehydrogenase (LDH) levels, and improved motor function in hDMDdel52/<i>mdx</i> mice. As no major safety observation was obtained, the improved therapeutic index of these next generation AONs is encouraging for further (pre)clinical development.</p>","PeriodicalId":19412,"journal":{"name":"Nucleic acid therapeutics","volume":"33 3","pages":"193-208"},"PeriodicalIF":4.0,"publicationDate":"2023-06-01","publicationTypes":"Journal Article","fieldsOfStudy":null,"isOpenAccess":false,"openAccessPdf":"https://www.ncbi.nlm.nih.gov/pmc/articles/PMC10277991/pdf/","citationCount":null,"resultStr":null,"platform":"Semanticscholar","paperid":"10022270","PeriodicalName":null,"FirstCategoryId":null,"ListUrlMain":null,"RegionNum":2,"RegionCategory":"医学","ArticlePicture":[],"TitleCN":null,"AbstractTextCN":null,"PMCID":"OA","EPubDate":null,"PubModel":null,"JCR":null,"JCRName":null,"Score":null,"Total":0}

{"title":"Complement C3d/C4d Deposition on Platelets Correlates with 2'-O-Methoxyethyl Antisense Oligonucleotide-Induced Thrombocytopenia in Monkeys.","authors":"Lijiang Shen,&nbsp;Andrea Wong,&nbsp;Satoru Oneda,&nbsp;Brian R Curtis,&nbsp;Joe Schroeder,&nbsp;Tom Zanardi,&nbsp;Jeffery A Engelhardt,&nbsp;Scott P Henry,&nbsp;Padmakumar Narayanan","doi":"10.1089/nat.2022.0042","DOIUrl":"https://doi.org/10.1089/nat.2022.0042","url":null,"abstract":"<p><p>2'-O-Methoxyethyl antisense oligonucleotide (2'-MOE ASO)-induced severe thrombocytopenia (TCP) [platelet (PLT) count <50 K/μL] was observed in the Asian-sourced cynomolgus monkeys with low incidence (2%-4% at doses >5 mg/kg/week). The potential mechanisms for TCP were studied using the Mauritian-sourced cynomolgus monkeys, which were shown to be more susceptible to ASO-induced TCP, along with the Asian-sourced animals. ISIS 405879, a 2'-MOE ASO, induced severe TCP (PLT <50 K/μL) in seven of nine Mauritian-sourced monkeys but not in the Asian-sourced monkeys after 16 weeks of treatment at 40 mg/kg/week. Marked increases in PLT-bound C3d/C4d were detected in all thrombocytopenic Mauritian-sourced monkeys but not in the unaffected Mauritian- or Asian-sourced monkeys, suggesting increased PLT clearance due to complement deposition on the PLTs. However, this effect was independent of the ASO-mediated fluid-phase alternative complement activation. A correlation was also observed between serum antiglycoprotein (GP) IIb/IIIa immunoglobulin G (IgG) and PLT reduction. In addition, increases in total serum IgM, anti-PLT IgM, and anti-PLT factor 4 IgM levels were observed in monkeys from both sources but were more evident in the Mauritian-sourced monkeys. These data suggest an enhanced innate immune cell activation to ISIS 405879, leading to increased PLT destruction through complement fixation on the PLTs or PLT crossreacting polyclonal antibody production.</p>","PeriodicalId":19412,"journal":{"name":"Nucleic acid therapeutics","volume":"33 3","pages":"209-225"},"PeriodicalIF":4.0,"publicationDate":"2023-06-01","publicationTypes":"Journal Article","fieldsOfStudy":null,"isOpenAccess":false,"openAccessPdf":"","citationCount":null,"resultStr":null,"platform":"Semanticscholar","paperid":"9953445","PeriodicalName":null,"FirstCategoryId":null,"ListUrlMain":null,"RegionNum":2,"RegionCategory":"医学","ArticlePicture":[],"TitleCN":null,"AbstractTextCN":null,"PMCID":"","EPubDate":null,"PubModel":null,"JCR":null,"JCRName":null,"Score":null,"Total":0}

{"title":"Time Is a Critical Factor When Evaluating Oligonucleotide Therapeutics in hERG Assays.","authors":"Yusheng Qu,&nbsp;Robert Kirby,&nbsp;Richard Davies,&nbsp;Ayesha Jinat,&nbsp;Stefano Stabilini,&nbsp;Bin Wu,&nbsp;Longchuan Yu,&nbsp;BaoXi Gao,&nbsp;Hugo M Vargas","doi":"10.1089/nat.2022.0043","DOIUrl":"https://doi.org/10.1089/nat.2022.0043","url":null,"abstract":"<p><p>In accord with International Conference on Harmonization S7B guidelines, an <i>in vitro</i> human ether-a-go-go-related gene (hERG) assay is one component of an integrated risk assessment for delayed ventricular repolarization. Function of hERG could be affected by direct (acute) mechanisms, or by indirect (chronic) mechanisms. Some approved oligonucleotide therapeutics had submitted hERG data to regulatory agents, which were all collected with the same protocol used for small-molecule testing (incubation time <20 min; acute), however, oligonucleotides have unique mechanisms and time courses of action (indirect). To reframe the hERG testing strategy for silencing RNA (siRNA), an investigation was performed to assess the time course for siRNA-mediated inhibition of hERG function and gene expression. Commercially available siRNAs of hERG were evaluated in a stable hERG-expressed cell line by whole-cell voltage clamp using automated electrophysiology and polymerase chain reaction. In the acute hERG study, no effects were observed after treatment with 100 nM siRNA for 20 min. The chronic effects of 100 nM siRNAs on hERG function were evaluated and recorded over 8-48 h following transfection. At 8 h there was no significant effect, whereas 77% reduction was observed at 48 h. Measurement of hERG mRNA levels demonstrated a 79% and 93% decrease of hERG mRNA at 8 and 48 h, respectively, consistent with inhibition of hERG transcription. The results indicate that an anti-hERG siRNA requires a long exposure time (48 h) in the hERG assay to produce a maximal reduction in hERG current; short exposures (20 min-8 h) had no effect. These findings imply that off-target profiling of novel oligonucleotides could benefit from using hERG protocol with long incubation times to de-risk potential off-target (indirect) effects on the hERG channel. This hERG assay modification may be important to consider if the findings are used to support an integrated nonclinical-clinical risk assessment for QTc (the duration of the QT interval adjusted for heart rate) prolongation.</p>","PeriodicalId":19412,"journal":{"name":"Nucleic acid therapeutics","volume":"33 2","pages":"132-140"},"PeriodicalIF":4.0,"publicationDate":"2023-04-01","publicationTypes":"Journal Article","fieldsOfStudy":null,"isOpenAccess":false,"openAccessPdf":"https://www.ncbi.nlm.nih.gov/pmc/articles/PMC10066779/pdf/","citationCount":null,"resultStr":null,"platform":"Semanticscholar","paperid":"9332968","PeriodicalName":null,"FirstCategoryId":null,"ListUrlMain":null,"RegionNum":2,"RegionCategory":"医学","ArticlePicture":[],"TitleCN":null,"AbstractTextCN":null,"PMCID":"OA","EPubDate":null,"PubModel":null,"JCR":null,"JCRName":null,"Score":null,"Total":0}

{"title":"Chemical Modifications and Design Influence the Potency of <i>Huntingtin</i> Anti-Gene Oligonucleotides.","authors":"Osama Saher,&nbsp;Eman M Zaghloul,&nbsp;Tea Umek,&nbsp;Daniel W Hagey,&nbsp;Negin Mozafari,&nbsp;Mathias B Danielsen,&nbsp;Alaa S Gouda,&nbsp;Karin E Lundin,&nbsp;Per T Jørgensen,&nbsp;Jesper Wengel,&nbsp;C I Edvard Smith,&nbsp;Rula Zain","doi":"10.1089/nat.2022.0046","DOIUrl":"https://doi.org/10.1089/nat.2022.0046","url":null,"abstract":"<p><p>Huntington's disease is a neurodegenerative, trinucleotide repeat (TNR) disorder affecting both males and females. It is caused by an abnormal increase in the length of CAG•CTG TNR in exon 1 of the <i>Huntingtin</i> gene (<i>HTT</i>). The resultant, mutant HTT mRNA and protein cause neuronal toxicity, suggesting that reduction of their levels would constitute a promising therapeutic approach. We previously reported a novel strategy in which chemically modified oligonucleotides (ONs) directly target chromosomal DNA. These anti-gene ONs were able to downregulate both <i>HTT</i> mRNA and protein. In this study, various locked nucleic acid (LNA)/DNA mixmer anti-gene ONs were tested to investigate the effects of varying ON length, LNA content, and fatty acid modification on <i>HTT</i> expression. Altering the length did not significantly influence the ON potency, while LNA content was critical for activity. Utilization of palmitoyl-modified LNA monomers enhanced the ON activity relatively to the corresponding nonmodified LNA under serum starvation conditions. Furthermore, the number of palmitoylated LNA monomers and their positioning greatly affected ON potency. In addition, we performed RNA sequencing analysis, which showed that the anti-gene ONs affect the \"immune system process, mRNA processing, and neurogenesis.\" Furthermore, we observed that for repeat containing genes, there is a higher tendency for antisense off-targeting. Taken together, our findings provide an optimized design of anti-gene ONs that could potentially be developed as DNA-targeting therapeutics for this class of TNR-related diseases.</p>","PeriodicalId":19412,"journal":{"name":"Nucleic acid therapeutics","volume":"33 2","pages":"117-131"},"PeriodicalIF":4.0,"publicationDate":"2023-04-01","publicationTypes":"Journal Article","fieldsOfStudy":null,"isOpenAccess":false,"openAccessPdf":"https://www.ncbi.nlm.nih.gov/pmc/articles/PMC10066784/pdf/","citationCount":null,"resultStr":null,"platform":"Semanticscholar","paperid":"9333519","PeriodicalName":null,"FirstCategoryId":null,"ListUrlMain":null,"RegionNum":2,"RegionCategory":"医学","ArticlePicture":[],"TitleCN":null,"AbstractTextCN":null,"PMCID":"OA","EPubDate":null,"PubModel":null,"JCR":null,"JCRName":null,"Score":null,"Total":0}