Nucleic acid therapeutics最新文献

{"title":"Enhancing SIRT1 Gene Expression Using Small Activating RNAs: A Novel Approach for Reversing Metabolic Syndrome.","authors":"Pinelopi Andrikakou,&nbsp;Vikash Reebye,&nbsp;Daniel Vasconcelos,&nbsp;Sorah Yoon,&nbsp;Jon Voutila,&nbsp;Andrew J T George,&nbsp;Piotr Swiderski,&nbsp;Robert Habib,&nbsp;Matthew Catley,&nbsp;David Blakey,&nbsp;Nagy A Habib,&nbsp;John J Rossi,&nbsp;Kai-Wen Huang","doi":"10.1089/nat.2021.0115","DOIUrl":"https://doi.org/10.1089/nat.2021.0115","url":null,"abstract":"<p><p>Metabolic syndrome (MetS) is a pathological condition characterized by abdominal obesity, insulin resistance, hypertension, and hyperlipidemia. Sirtuin 1 (SIRT1), a highly conserved histone deacetylase, is characterized as a key metabolic regulator and protector against aging-associated pathologies, including MetS. In this study, we investigate the therapeutic potential of activating SIRT1 using small activating RNAs (saRNA), thereby reducing inflammatory-like responses and re-establishing normal lipid metabolism. SIRT1 saRNA significantly increased SIRT1 messenger RNA (mRNA) and protein levels in both lipopolysaccharide-stimulated and nonstimulated macrophages. SIRT1 saRNA significantly decreased inflammatory-like responses, by reducing mRNA levels of key inflammatory cytokines, such as Tumor Necrosis Factor alpha, Interleukin 1 beta (IL-1β), Interleukin 6 (IL-6), and chemokines Monocyte Chemoattractant Protein-1 and keratinocyte chemoattractant. SIRT1 overexpression also significantly reduced phosphorylation of nuclear factor-κB and c-Jun N-terminal kinase, both key signaling molecules for the inflammatory pathway. To investigate the therapeutic effect of SIRT1 upregulation, we treated a high-fat diet model with SIRT1 saRNA conjugated to a transferrin receptor aptamer for delivery to the liver and cellular internalization. Animals in the SIRT1 saRNA treatment arm demonstrated significantly decreased weight gain with a significant reduction in white adipose tissue, triglycerides, fasting glucose levels, and intracellular lipid accumulation. These suggest treatment-induced changes to lipid and glucose metabolism in the animals. The results of this study demonstrate that targeted activation of SIRT1 by saRNAs is a potential strategy to reverse MetS.</p>","PeriodicalId":19412,"journal":{"name":"Nucleic acid therapeutics","volume":"32 6","pages":"486-496"},"PeriodicalIF":4.0,"publicationDate":"2022-12-01","publicationTypes":"Journal Article","fieldsOfStudy":null,"isOpenAccess":false,"openAccessPdf":"","citationCount":null,"resultStr":null,"platform":"Semanticscholar","paperid":"10486735","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":"Inflammatory Non-CpG Antisense Oligonucleotides Are Signaling Through TLR9 in Human Burkitt Lymphoma B Bjab Cells.","authors":"Adam J Pollak,&nbsp;Patrick Cauntay,&nbsp;Todd Machemer,&nbsp;Suzanne Paz,&nbsp;Sagar Damle,&nbsp;Scott P Henry,&nbsp;Sebastien A Burel","doi":"10.1089/nat.2022.0034","DOIUrl":"https://doi.org/10.1089/nat.2022.0034","url":null,"abstract":"<p><p>Nucleic acid-based phosphorothioate containing antisense oligonucleotides (PS-ASOs) have the potential to activate cellular innate immune responses, and the level of activation can vary quite dramatically with sequence. Minimizing the degree of proinflammatory effect is one of the main selection criteria for compounds intended to move into clinical trials. While a recently developed human peripheral blood mononuclear cell (hPBMC)-based assay showed excellent ability to detect innate immune active PS-ASOs, which can then be discarded from the developmental process, this assay is highly resource intensive and easily affected by subject variability. This compelled us to develop a more convenient high-throughput assay. In this study, we describe a new <i>in vitro</i> assay, utilizing a cultured human Bjab cell line, which was developed and validated to identify PS-ASOs that may cause innate immune activation. The assay was calibrated to replicate results from the hPBMC assay. The Bjab assay was designed to be high throughput and more convenient by using RT-qPCR readout of mRNA of the chemokine Ccl22. The Bjab assay was also shown to be highly reproducible and to provide a large dynamic range in determining the immune potential of PS-ASOs through comparison to known benchmark PS-ASO controls that were previously shown to be safe or inflammatory in clinical trials. In addition, we demonstrate that Bjab cells can be used to provide mechanistic information on PS-ASO TLR9-dependent innate immune activation.</p>","PeriodicalId":19412,"journal":{"name":"Nucleic acid therapeutics","volume":"32 6","pages":"473-485"},"PeriodicalIF":4.0,"publicationDate":"2022-12-01","publicationTypes":"Journal Article","fieldsOfStudy":null,"isOpenAccess":false,"openAccessPdf":"","citationCount":null,"resultStr":null,"platform":"Semanticscholar","paperid":"10793628","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":"Nucleic Acid Aptamers Increase the Anticancer Efficiency and Reduce the Toxicity of Cisplatin-Arabinogalactan Conjugates <i>In Vivo</i>.","authors":"Tatiana N Zamay,&nbsp;Alexander K Starkov,&nbsp;Olga S Kolovskaya,&nbsp;Galina S Zamay,&nbsp;Dmitry V Veprintsev,&nbsp;Natalia Luzan,&nbsp;Elena D Nikolaeva,&nbsp;Kirill A Lukyanenko,&nbsp;Polina V Artyushenko,&nbsp;Irina A Shchugoreva,&nbsp;Yury E Glazyrin,&nbsp;Anastasia A Koshmanova,&nbsp;Alexey V Krat,&nbsp;Dariya S Tereshina,&nbsp;Sergey S Zamay,&nbsp;Yuriy S Pats,&nbsp;Ruslan A Zukov,&nbsp;Felix N Tomilin,&nbsp;Maxim V Berezovski,&nbsp;Anna S Kichkailo","doi":"10.1089/nat.2022.0024","DOIUrl":"https://doi.org/10.1089/nat.2022.0024","url":null,"abstract":"<p><p>Cisplatin is an effective drug for treating various cancer types. However, it is highly toxic for both healthy and tumor cells. Therefore, there is a need to reduce its therapeutic dose and increase targeted bioavailability. One of the ways to achieve this could be the coating of cisplatin with polysaccharides and specific carriers for targeted delivery. Nucleic acid aptamers could be used as carriers for the specific delivery of medicine to cancer cells. Cisplatin-arabinogalactan-aptamer (Cis-AG-Ap) conjugate was synthesized based on Cis-dichlorodiammineplatinum, Siberian larch arabinogalactan, and aptamer AS-42 specific to heat-shock proteins (HSP) 71 kDa (Hspa8) and HSP 90-beta (Hsp90ab1). The antitumor effect was estimated using ascites and metastatic Ehrlich tumor models. Cis-AG-Ap toxicity was assessed by blood biochemistry on healthy mice. Here, we demonstrated enhanced anticancer activity of Cis-AG-Ap and its specific accumulation in tumor foci. It was shown that targeted delivery allowed a 15-fold reduction in the therapeutic dose of cisplatin and its toxicity. Cis-AG-Ap sufficiently suppressed the growth of Ehrlich's ascites carcinoma, the mass and extent of tumor metastasis <i>in vivo</i>. Arabinogalactan and the aptamers promoted cisplatin efficiency by enhancing its bioavailability. The described strategy could be very promising for targeted anticancer therapy.</p>","PeriodicalId":19412,"journal":{"name":"Nucleic acid therapeutics","volume":"32 6","pages":"497-506"},"PeriodicalIF":4.0,"publicationDate":"2022-12-01","publicationTypes":"Journal Article","fieldsOfStudy":null,"isOpenAccess":false,"openAccessPdf":"","citationCount":null,"resultStr":null,"platform":"Semanticscholar","paperid":"10486741","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":"Assessment of the Immunogenicity Potential for Oligonucleotide-Based Drugs.","authors":"Scott P Henry,&nbsp;Cecilia Arfvidsson,&nbsp;Josh Arrington,&nbsp;Jasna Canadi,&nbsp;Dave Crowe,&nbsp;Shalini Gupta,&nbsp;Sabine Lohmann,&nbsp;Benoit Massonnet,&nbsp;Daniel Mytych,&nbsp;Tina Rogers,&nbsp;Hobart Rogers,&nbsp;Chris Stebbins,&nbsp;Craig Stovold,&nbsp;Daniela Verthelyi,&nbsp;Adam Vigil,&nbsp;Chi Xuan,&nbsp;Yuanxin Xu,&nbsp;Rosie Yu,&nbsp;Thomas Klem","doi":"10.1089/nat.2021.0112","DOIUrl":"https://doi.org/10.1089/nat.2021.0112","url":null,"abstract":"<p><p>Therapeutic oligonucleotides (ONs) have characteristics of both small molecules and biologics. Although safety assessment of ONs largely follows guidelines established for small molecules, the unique characteristics of ONs often require incorporation of concepts from the safety assessment of biologics. The assessment of immunogenicity for ON therapeutics is one area where the approach is distinct from either established small molecule or biologic platforms. Information regarding immunogenicity of ONs is limited, but indicates that administration of ONs can result in antidrug antibody formation. In this study, we summarize the collective experience of the Oligonucleotide Safety Working Group in designing the immunogenicity assessment appropriate for this class of therapeutic, including advice on assay development, clinical monitoring, and evaluation of the impact of immunogenicity on exposure, efficacy, and safety of therapeutic ONs.</p>","PeriodicalId":19412,"journal":{"name":"Nucleic acid therapeutics","volume":" ","pages":"369-377"},"PeriodicalIF":4.0,"publicationDate":"2022-10-01","publicationTypes":"Journal Article","fieldsOfStudy":null,"isOpenAccess":false,"openAccessPdf":"","citationCount":null,"resultStr":null,"platform":"Semanticscholar","paperid":"40384926","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":"The Combination of Mesyl-Phosphoramidate Inter-Nucleotide Linkages and 2'-<i>O</i>-Methyl in Selected Positions in the Antisense Oligonucleotide Enhances the Performance of RNaseH1 Active PS-ASOs.","authors":"Lingdi Zhang,&nbsp;Xue-Hai Liang,&nbsp;Cheryl Li De Hoyos,&nbsp;Michael Migawa,&nbsp;Joshua G Nichols,&nbsp;Graeme Freestone,&nbsp;Jun Tian,&nbsp;Punit P Seth,&nbsp;Stanley T Crooke","doi":"10.1089/nat.2022.0005","DOIUrl":"https://doi.org/10.1089/nat.2022.0005","url":null,"abstract":"<p><p>Antisense oligonucleotides (ASOs) that mediate RNA target degradation by RNase H1 are used as drugs to treat various diseases. Previously we found that introduction of a single 2'-<i>O</i>-methyl (2'-OMe) modification in position 2 of the central deoxynucleotide region of a gapmer phosphorothioate (PS) ASO, in which several residues at the termini are 2'-methoxyethyl, 2' constrained ethyl, or locked nucleic acid, dramatically reduced cytotoxicity with only modest effects on potency. More recently, we demonstrated that replacement of the PS linkage at position 2 or 3 in the gap with a mesyl-phosphoramidate (MsPA) linkage also significantly reduced toxicity without meaningful loss of potency and increased the elimination half-life of the ASOs. In this study, we evaluated the effects of the combination of MsPA linkages and 2'-OMe nucleotides on PS ASO performance. We found that two MsPA modifications at the 5' end of the gap or in the 3'-wing of a Gap 2'-OMe PS ASO substantially increased the activity of ASOs with OMe at position 2 of the gap without altering the safety profile. Such effects were observed with multiple sequences in cells and animals. Thus, the MsPA modification improves the RNase H1 cleavage rate of PS ASOs with a 2'-OMe in the gap, significantly reduces binding of proteins involved in cytotoxicity, and prolongs elimination half-lives.</p>","PeriodicalId":19412,"journal":{"name":"Nucleic acid therapeutics","volume":" ","pages":"401-411"},"PeriodicalIF":4.0,"publicationDate":"2022-10-01","publicationTypes":"Journal Article","fieldsOfStudy":null,"isOpenAccess":false,"openAccessPdf":"https://ftp.ncbi.nlm.nih.gov/pub/pmc/oa_pdf/29/a2/nat.2022.0005.PMC9595634.pdf","citationCount":null,"resultStr":null,"platform":"Semanticscholar","paperid":"40525095","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":"Delivery of RNA Therapeutics: The Great Endosomal Escape!","authors":"Steven F Dowdy,&nbsp;Ryan L Setten,&nbsp;Xian-Shu Cui,&nbsp;Satish G Jadhav","doi":"10.1089/nat.2022.0004","DOIUrl":"10.1089/nat.2022.0004","url":null,"abstract":"<p><p>RNA therapeutics, including siRNAs, antisense oligonucleotides, and other oligonucleotides, have great potential to selectively treat a multitude of human diseases, from cancer to COVID to Parkinson's disease. RNA therapeutic activity is mechanistically driven by Watson-Crick base pairing to the target gene RNA without the requirement of prior knowledge of the protein structure, function, or cellular location. However, before widespread use of RNA therapeutics becomes a reality, we must overcome a billion years of evolutionary defenses designed to keep invading RNAs from entering cells. Unlike small-molecule therapeutics that are designed to passively diffuse across the cell membrane, macromolecular RNA therapeutics are too large, too charged, and/or too hydrophilic to passively diffuse across the cellular membrane and are instead taken up into cells by endocytosis. However, similar to the cell membrane, endosomes comprise a lipid bilayer that entraps 99% or more of RNA therapeutics, even in semipermissive tissues such as the liver, central nervous system, and muscle. Consequently, before RNA therapeutics can achieve their ultimate clinical potential to treat widespread human disease, the rate-limiting delivery problem of endosomal escape must be solved in a clinically acceptable manner.</p>","PeriodicalId":19412,"journal":{"name":"Nucleic acid therapeutics","volume":"32 5","pages":"361-368"},"PeriodicalIF":4.0,"publicationDate":"2022-10-01","publicationTypes":"Journal Article","fieldsOfStudy":null,"isOpenAccess":false,"openAccessPdf":"https://www.ncbi.nlm.nih.gov/pmc/articles/PMC9595607/pdf/nat.2022.0004.pdf","citationCount":null,"resultStr":null,"platform":"Semanticscholar","paperid":"10351888","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":"Inhibition of Epidermal Growth Factor Receptor Signaling by Antisense Oligonucleotides as a Novel Approach to Epidermal Growth Factor Receptor Inhibition.","authors":"Thushara W Madanayake,&nbsp;Eric A Welsh,&nbsp;Lancia N F Darville,&nbsp;John M Koomen,&nbsp;Charles E Chalfant,&nbsp;Eric B Haura,&nbsp;Timothy J Robinson","doi":"10.1089/nat.2021.0101","DOIUrl":"10.1089/nat.2021.0101","url":null,"abstract":"<p><p>We report a novel method to inhibit epidermal growth factor receptor (EGFR) signaling using custom morpholino antisense oligonucleotides (ASOs) to drive expression of dominant negative mRNA isoforms of EGFR by ASO-induced exon skipping within the transmembrane (16) or tyrosine kinase domains (18 and 21). <i>In vivo</i> ASO formulations induced >95% exon skipping in several models of nonsmall cell lung cancer (NSCLC) and were comparable in efficacy to erlotinib in reducing colony formation, cell viability, and migration in EGFR mutant NSCLC (PC9). However, unlike erlotinib, ASOs maintained their efficacy in both erlotinib-resistant subclones (PC9-GR) and wild-type overexpressing EGFR models (H292), in which erlotinib had no significant effect. The most dramatic ASO-induced phenotype resulted from targeting the EGFR kinase domain directly, which resulted in maximal inhibition of phosphorylation of EGFR, Akt, and Erk in both PC9 and PC9GR cells. Phosphoproteomic mass spectrometry confirmed highly congruent impacts of exon 16-, 18-, and 21-directed ASOs compared with erlotinib on PC9 genome-wide cell signaling. Furthermore, <i>EGFR</i>-directed ASOs had no impact in EGFR-independent NSCLC models, confirming an EGFR-specific therapeutic mechanism. Further exploration of synergy of ASOs with existing tyrosine kinase inhibitors may offer novel clinical models to improve EGFR-targeted therapies for both mutant and wild-type NSCLC patients.</p>","PeriodicalId":19412,"journal":{"name":"Nucleic acid therapeutics","volume":" ","pages":"391-400"},"PeriodicalIF":4.0,"publicationDate":"2022-10-01","publicationTypes":"Journal Article","fieldsOfStudy":null,"isOpenAccess":false,"openAccessPdf":"https://www.ncbi.nlm.nih.gov/pmc/articles/PMC9595651/pdf/nat.2021.0101.pdf","citationCount":null,"resultStr":null,"platform":"Semanticscholar","paperid":"40526775","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":"Host-Directed Targeting of LincRNA-MIR99AHG Suppresses Intracellular Growth of <i>Mycobacterium tuberculosis</i>.","authors":"Lorna Gcanga,&nbsp;Ousman Tamgue,&nbsp;Mumin Ozturk,&nbsp;Shandre Pillay,&nbsp;Raygaana Jacobs,&nbsp;Julius Ebua Chia,&nbsp;Stanley Kimbung Mbandi,&nbsp;Malika Davids,&nbsp;Keertan Dheda,&nbsp;Sebastian Schmeier,&nbsp;Tanvir Alam,&nbsp;Sugata Roy,&nbsp;Harukazu Suzuki,&nbsp;Frank Brombacher,&nbsp;Reto Guler","doi":"10.1089/nat.2022.0009","DOIUrl":"https://doi.org/10.1089/nat.2022.0009","url":null,"abstract":"<p><p>Tuberculosis (TB) caused by <i>Mycobacterium tuberculosis</i> (Mtb) kills 1.6 million people worldwide every year, and there is an urgent need for targeting host-pathogen interactions as a strategy to reduce mycobacterial resistance to current antimicrobials. Noncoding RNAs are emerging as important regulators of numerous biological processes and avenues for exploitation in host-directed therapeutics. Although long noncoding RNAs (lncRNAs) are abundantly expressed in immune cells, their functional role in gene regulation and bacterial infections remains understudied. In this study, we identify an immunoregulatory long intergenic noncoding RNA, lincRNA-MIR99AHG, which is upregulated in mouse and human macrophages upon IL-4/IL-13 stimulation and downregulated after clinical Mtb HN878 strain infection and in peripheral blood mononuclear cells from active TB patients. To evaluate the functional role of lincRNA-MIR99AHG, we used antisense locked nucleic acid (LNA) GapmeR-mediated antisense oligonucleotide (ASO) lncRNA knockdown experiments. Knockdown of lincRNA-MIR99AHG with ASOs significantly reduced intracellular Mtb growth in mouse and human macrophages and reduced pro-inflammatory cytokine production. In addition, <i>in vivo</i> treatment of mice with MIR99AHG ASOs reduced the mycobacterial burden in the lung and spleen. Furthermore, in macrophages, lincRNA-MIR99AHG is translocated to the nucleus and interacts with high affinity to hnRNPA2/B1 following IL-4/IL-13 stimulation and Mtb HN878 infection. Together, these findings identify lincRNA-MIR99AHG as a positive regulator of inflammation and macrophage polarization to promote Mtb growth and a possible target for adjunctive host-directed therapy against TB.</p>","PeriodicalId":19412,"journal":{"name":"Nucleic acid therapeutics","volume":"32 5","pages":"421-437"},"PeriodicalIF":4.0,"publicationDate":"2022-10-01","publicationTypes":"Journal Article","fieldsOfStudy":null,"isOpenAccess":false,"openAccessPdf":"https://www.ncbi.nlm.nih.gov/pmc/articles/PMC7613730/pdf/nat.2022.0009.pdf","citationCount":null,"resultStr":null,"platform":"Semanticscholar","paperid":"10550983","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":"Binary Antisense Oligonucleotide Agent for Cancer Marker-Dependent Degradation of Targeted RNA.","authors":"Valeriia S Drozd,&nbsp;Ahmed A Eldeeb,&nbsp;Dmitry M Kolpashchikov,&nbsp;Daria D Nedorezova","doi":"10.1089/nat.2021.0108","DOIUrl":"https://doi.org/10.1089/nat.2021.0108","url":null,"abstract":"<p><p>Antisense oligonucleotide technology is one of the most successful gene therapy (GT) approaches. However, low selectivity of antisense agents limits their application as anticancer drugs. To achieve activation of antisense agent selectively in cancer cells, herein, we propose the concept of binary antisense oligonucleotide (biASO) agent. biASO recognizes an RNA sequence of a gene associated with cancer development (marker) and then activates RNase H-dependent cleavage of a targeted messenger RNA. biASO was optimized to produce only the background cleavage of the targeted RNA in the absence of the activator. The approach lays the foundation for the development of highly selective and efficient GT agents.</p>","PeriodicalId":19412,"journal":{"name":"Nucleic acid therapeutics","volume":" ","pages":"412-420"},"PeriodicalIF":4.0,"publicationDate":"2022-10-01","publicationTypes":"Journal Article","fieldsOfStudy":null,"isOpenAccess":false,"openAccessPdf":"","citationCount":null,"resultStr":null,"platform":"Semanticscholar","paperid":"40628325","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":"Antisense Oligonucleotide Rescue of Deep-Intronic Variants Activating Pseudoexons in the 6-Pyruvoyl-Tetrahydropterin Synthase Gene.","authors":"Ainhoa Martínez-Pizarro,&nbsp;Fátima Leal,&nbsp;Lise Lolle Holm,&nbsp;Thomas K Doktor,&nbsp;Ulrika S S Petersen,&nbsp;María Bueno,&nbsp;Beat Thöny,&nbsp;Belén Pérez,&nbsp;Brage S Andresen,&nbsp;Lourdes R Desviat","doi":"10.1089/nat.2021.0066","DOIUrl":"https://doi.org/10.1089/nat.2021.0066","url":null,"abstract":"<p><p>We report two new 6-pyruvoyl-tetrahydropterin synthase splicing variants identified through genomic sequencing and transcript analysis in a patient with tetrahydrobiopterin deficiency, presenting with hyperphenylalaninemia and monoamine neurotransmitter deficiency. Variant c.243 + 3A>G causes exon 4 skipping. The deep-intronic c.164-672C>T variant creates a potential 5' splice site that leads to the inclusion of four overlapping pseudoexons, corresponding to exonizations of an antisense short interspersed nuclear element <i>AluSq</i> repeat sequence. Two of the identified pseudoexons have been reported previously, activated by different deep-intronic variants, and were also detected at residual levels in control cells. Interestingly, the predominant pseudoexon is nearly identical to a disease causing activated pseudoexon in the <i>F8</i> gene, with the same 3' and 5' splice sites. Splice switching antisense oligonucleotides (SSOs) were designed to hybridize with splice sites and/or predicted binding sites for regulatory splice factors. Different SSOs corrected the aberrant pseudoexon inclusion, both in minigenes and in fibroblasts from patients carrying the new variant c.164-672C>T or the previously described c.164-716A>T. With SSO treatment PTPS protein was recovered, illustrating the therapeutic potential of the approach, for patients with different pseudoexon activating variants in the region. In addition, the natural presence of pseudoexons in the wild type context suggests the possibility of applying the antisense strategy in patients with hypomorphic <i>PTS</i> variants with the purpose of upregulating their expression to increase overall protein and activity.</p>","PeriodicalId":19412,"journal":{"name":"Nucleic acid therapeutics","volume":" ","pages":"378-390"},"PeriodicalIF":4.0,"publicationDate":"2022-10-01","publicationTypes":"Journal Article","fieldsOfStudy":null,"isOpenAccess":false,"openAccessPdf":"https://www.ncbi.nlm.nih.gov/pmc/articles/PMC9595628/pdf/","citationCount":null,"resultStr":null,"platform":"Semanticscholar","paperid":"40521384","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}