{"status":"ok","message-type":"work","message-version":"1.0.0","message":{"indexed":{"date-parts":[[2026,9,16]],"date-time":"2026-09-16T14:07:20Z","timestamp":1789567640093,"version":"build-2803163510"},"reference-count":41,"publisher":"American Chemical Society (ACS)","issue":"36","license":[{"start":{"date-parts":[[2026,9,2]],"date-time":"2026-09-02T00:00:00Z","timestamp":1788307200000},"content-version":"vor","delay-in-days":0,"URL":"https:\/\/creativecommons.org\/licenses\/by-nc-nd\/4.0\/"}],"content-domain":{"domain":[],"crossmark-restriction":false},"short-container-title":[],"published-print":{"date-parts":[[2026,9,16]]},"abstract":"<jats:title>Abstract<\/jats:title>\n                  <jats:p>\u03b1-(l)-(3\u2032\u20132\u2032)-Threofuranosyl nucleic acid (TNA) cross-pairs with DNA and RNA and is not degraded by nucleases and thus has potential for use in diagnostics and therapeutics based on nucleic acids. Here, we describe the synthesis of 15 building blocks of chiral 4\u2032-(R)- and 15 building blocks of chiral (S)-methyl-\u03b1-(l)-threofuranosyl nucleic acid (4\u2032-Me-TNA) monomers and their incorporation into oligonucleotides. Crystallographic studies of RNAs containing 4\u2032-Me-TNA modifications revealed the impact of the methyl substituent on the sugar pucker and duplex conformation. Biostability studies showed that a terminal 4\u2032-Me-TNA modification on a deoxyoligonucleotide imparted resistance to nuclease digestion equivalent to unmethylated TNA, which is markedly superior to that of more conventional 2\u2032-ribo and 2\u2032-deoxy modifications. A single nucleotide incorporation of the 4\u2032-(R)-Me-TNA destabilized an otherwise all-RNA 12-mer duplex by 6.5 to 12 \u00b0C, depending on the base, relative to the unmodified RNA control duplex; the (S) isomer was less destabilizing than the (R) isomer. Computational models were used to rationalize the high levels of nuclease resistance and the thermal destabilization. The 4\u2032-Me-TNA triphosphates are poorly recognized by an engineered TNA polymerase making it unlikely that the 4\u2032-Me-TNA triphosphates will be recognized by natural polymerases. Together, these properties warrant the evaluation of 4\u2032-Me-TNA in nucleic acid-based therapeutics.<\/jats:p>","DOI":"10.1021\/jacs.6c07051","type":"journal-article","created":{"date-parts":[[2026,9,3]],"date-time":"2026-09-03T16:48:46Z","timestamp":1788454126000},"page":"38692-38706","source":"Crossref","is-referenced-by-count":0,"title":["Chiral-4\u2032-Methyl-\u03b1-(\n                    <scp>l<\/scp>\n                    )-Threofuranosyl Nucleic Acids: Synthesis, Structure,\nBinding\nAffinity, Nuclease Resistance, and Polymerase Recognition"],"prefix":"10.1021","volume":"148","author":[{"given":"Atsushi","family":"Mikami","sequence":"first","affiliation":[{"name":"Alnylam Pharmaceuticals , 675 West Kendall Street , , ,","place":["Cambridge, Massachusetts, United States, 02142"]}],"role":[{"vocabulary":"crossref","role":"author"}]},{"ORCID":"https:\/\/orcid.org\/0000-0003-2077-1294","authenticated-orcid":true,"given":"Dhrubajyoti","family":"Datta","sequence":"additional","affiliation":[{"name":"Alnylam Pharmaceuticals , 675 West Kendall Street , , ,","place":["Cambridge, Massachusetts, United States, 02142"]}],"role":[{"vocabulary":"crossref","role":"author"}]},{"ORCID":"https:\/\/orcid.org\/0000-0003-2669-8112","authenticated-orcid":true,"given":"Jayanta","family":"Kundu","sequence":"additional","affiliation":[{"name":"Alnylam Pharmaceuticals , 675 West Kendall Street , , ,","place":["Cambridge, Massachusetts, United States, 02142"]}],"role":[{"vocabulary":"crossref","role":"author"}]},{"ORCID":"https:\/\/orcid.org\/0000-0002-2837-419X","authenticated-orcid":true,"given":"Vathan","family":"Kumar","sequence":"additional","affiliation":[{"name":"Alnylam Pharmaceuticals , 675 West Kendall Street , , ,","place":["Cambridge, Massachusetts, United States, 02142"]}],"role":[{"vocabulary":"crossref","role":"author"}]},{"given":"Namrata","family":"Erande","sequence":"additional","affiliation":[{"name":"Alnylam Pharmaceuticals , 675 West Kendall Street , , ,","place":["Cambridge, Massachusetts, United States, 02142"]}],"role":[{"vocabulary":"crossref","role":"author"}]},{"given":"Esau","family":"Medina","sequence":"additional","affiliation":[{"name":"University of California , , , ,","place":["Irvine, California, United States, 92697-3958"]}],"role":[{"vocabulary":"crossref","role":"author"}]},{"given":"Victoria A.","family":"Maola","sequence":"additional","affiliation":[{"name":"University of California , , , ,","place":["Irvine, California, United States, 92697-3958"]}],"role":[{"vocabulary":"crossref","role":"author"}]},{"given":"Erica M.","family":"Lee","sequence":"additional","affiliation":[{"name":"University of California , , , ,","place":["Irvine, California, United States, 92697-3958"]}],"role":[{"vocabulary":"crossref","role":"author"}]},{"given":"Shigeo","family":"Matsuda","sequence":"additional","affiliation":[{"name":"Alnylam Pharmaceuticals , 675 West Kendall Street , , ,","place":["Cambridge, Massachusetts, United States, 02142"]}],"role":[{"vocabulary":"crossref","role":"author"}]},{"given":"Joel M.","family":"Harp","sequence":"additional","affiliation":[{"name":"Vanderbilt University, School of Medicine , , , ,","place":["Nashville, Tennessee, United States, 37232"]}],"role":[{"vocabulary":"crossref","role":"author"}]},{"ORCID":"https:\/\/orcid.org\/0000-0003-4145-356X","authenticated-orcid":true,"given":"Martin","family":"Egli","sequence":"additional","affiliation":[{"name":"Vanderbilt University, School of Medicine , , , ,","place":["Nashville, Tennessee, United States, 37232"]}],"role":[{"vocabulary":"crossref","role":"author"}]},{"ORCID":"https:\/\/orcid.org\/0000-0003-1393-135X","authenticated-orcid":true,"given":"John C.","family":"Chaput","sequence":"additional","affiliation":[{"name":"University of California , , , ,","place":["Irvine, California, United States, 92697-3958"]}],"role":[{"vocabulary":"crossref","role":"author"}]},{"ORCID":"https:\/\/orcid.org\/0000-0002-7931-1172","authenticated-orcid":true,"given":"Muthiah","family":"Manoharan","sequence":"additional","affiliation":[{"name":"Alnylam Pharmaceuticals , 675 West Kendall Street , , ,","place":["Cambridge, Massachusetts, United States, 02142"]}],"role":[{"vocabulary":"crossref","role":"author"}]}],"member":"316","published-online":{"date-parts":[[2026,9,2]]},"reference":[{"key":"2026091609115515900_cit1","doi-asserted-by":"publisher","first-page":"1347","DOI":"10.1126\/science.290.5495.1347","article-title":"Chemical etiology of nucleic acid structure: the \u03b1-threofuranosyl-(3\u2032\u22122\u2032)\noligonucleotide system","volume":"290","author":"Scho\u0308ning","year":"2000","journal-title":"Science"},{"key":"2026091609115515900_cit2","doi-asserted-by":"publisher","first-page":"2118","DOI":"10.1126\/science.284.5423.2118","article-title":"Chemical etiology\nof nucleic acid structure","volume":"284","author":"Eschenmoser","year":"1999","journal-title":"Science"},{"key":"2026091609115515900_cit3","doi-asserted-by":"publisher","first-page":"289","DOI":"10.1007\/s00239-007-9017-9","article-title":"Experimental evidence\nthat GNA and TNA were not sequential polymers in the prebiotic evolution\nof RNA","volume":"65","author":"Yang","year":"2007","journal-title":"J. Mol. Evol."},{"key":"2026091609115515900_cit4","doi-asserted-by":"publisher","first-page":"1144","DOI":"10.1021\/acssynbio.9b00028","article-title":"Thermostability\nTrends of TNA:DNA Duplexes Reveal Strong Purine Dependence","volume":"8","author":"Lackey","year":"2019","journal-title":"ACS Synth. Biol."},{"key":"2026091609115515900_cit5","doi-asserted-by":"publisher","first-page":"15105","DOI":"10.1021\/ja8041959","article-title":"The structure of a TNA-TNA complex in solution: NMR study of the\noctamer duplex derived from \u03b1-(l)-threofuranosyl-(3\u2032\u22122\u2032)-CGAATTCG","volume":"130","author":"Ebert","year":"2008","journal-title":"J. Am. Chem. Soc."},{"key":"2026091609115515900_cit6","doi-asserted-by":"publisher","first-page":"13716","DOI":"10.1021\/ja0207807","article-title":"Crystal structure of a B-form DNA\nduplex containing (l)-\u03b1-threofuranosyl\n(3\u2032\u22122\u2032) nucleosides: a four-carbon sugar is easily\naccommodated into the backbone of DNA","volume":"124","author":"Wilds","year":"2002","journal-title":"J. Am.\nChem. Soc."},{"key":"2026091609115515900_cit7","doi-asserted-by":"publisher","first-page":"5893","DOI":"10.1002\/anie.200352553","article-title":"Why does TNA cross-pair\nmore strongly\nwith RNA than with DNA? An answer from X-ray analysis","volume":"42","author":"Pallan","year":"2003","journal-title":"Angew. Chem., Int. Ed."},{"key":"2026091609115515900_cit8","doi-asserted-by":"publisher","first-page":"2418","DOI":"10.1016\/j.bmcl.2016.03.118","article-title":"Evaluating TNA stability under simulated physiological\nconditions","volume":"26","author":"Culbertson","year":"2016","journal-title":"Bioorg. Med. Chem. Lett."},{"key":"2026091609115515900_cit9","doi-asserted-by":"publisher","first-page":"9542","DOI":"10.1093\/nar\/gkad716","article-title":"Stability and mechanism\nof threose nucleic acid toward\nacid-mediated degradation","volume":"51","author":"Lee","year":"2023","journal-title":"Nucleic Acids Res."},{"key":"2026091609115515900_cit10","doi-asserted-by":"publisher","first-page":"1056","DOI":"10.1021\/acs.accounts.0c00886","article-title":"Redesigning\nthe Genetic Polymers of Life","volume":"54","author":"Chaput","year":"2021","journal-title":"Acc. Chem. Res."},{"key":"2026091609115515900_cit11","doi-asserted-by":"publisher","first-page":"1007","DOI":"10.1093\/nar\/gkv1472","article-title":"The structural diversity\nof artificial genetic polymers","volume":"44","author":"Anosova","year":"2016","journal-title":"Nucleic Acids\nRes."},{"key":"2026091609115515900_cit12","doi-asserted-by":"publisher","first-page":"19691","DOI":"10.1021\/jacs.3c04744","article-title":"Shorter Is Better: The  \u03b1-(l)-Threofuranosyl Nucleic\nAcid Modification Improves Stability, Potency,\nSafety, and Ago2 Binding and Mitigates Off-Target Effects of Small\nInterfering RNAs","volume":"145","author":"Matsuda","year":"2023","journal-title":"J. Am. Chem. Soc."},{"key":"2026091609115515900_cit13","doi-asserted-by":"publisher","first-page":"100299","DOI":"10.1016\/j.mtbio.2022.100299","article-title":"Cellular\nuptake,\ntissue penetration, biodistribution, and biosafety of threose nucleic\nacids: Assessing in vitro and in vivo delivery","volume":"15","author":"Wang","year":"2022","journal-title":"Mater. Today Bio"},{"key":"2026091609115515900_cit14","doi-asserted-by":"publisher","first-page":"25789","DOI":"10.1021\/jacs.3c09497","article-title":"Functionally\nEnhanced XNA Aptamers Discovered by Parallelized\nLibrary Screening","volume":"145","author":"Lozoya-Colinas","year":"2023","journal-title":"J. Am. Chem. Soc."},{"key":"2026091609115515900_cit15","doi-asserted-by":"publisher","first-page":"183","DOI":"10.1038\/nchem.1241","article-title":"Darwinian evolution of an alternative\ngenetic system provides support for TNA as an RNA progenitor","volume":"4","author":"Yu","year":"2012","journal-title":"Nat. Chem."},{"key":"2026091609115515900_cit16","doi-asserted-by":"publisher","first-page":"8154","DOI":"10.1021\/jacs.1c02895","article-title":"A Threose\nNucleic Acid Enzyme with\nRNA Ligase Activity","volume":"143","author":"Wang","year":"2021","journal-title":"J. Am. Chem. Soc."},{"key":"2026091609115515900_cit17","doi-asserted-by":"publisher","DOI":"10.1017\/s0033583520000050","article-title":"Engineering\npolymerases\nfor applications in synthetic biology","volume":"53","author":"Nikoomanzar","year":"2020","journal-title":"Q. Rev.\nBiophys."},{"key":"2026091609115515900_cit18","doi-asserted-by":"publisher","first-page":"11235","DOI":"10.1038\/ncomms11235","article-title":"A general strategy\nfor expanding polymerase function\nby droplet microfluidics","volume":"7","author":"Larsen","year":"2016","journal-title":"Nat. Commun."},{"key":"2026091609115515900_cit19","doi-asserted-by":"publisher","first-page":"9894","DOI":"10.1039\/D0CC04356G","article-title":"Efficient\nconstruction of a stable linear gene based on a TNA loop modified\nprimer pair for gene delivery","volume":"56","author":"Lu","year":"2020","journal-title":"Chem. Commun."},{"key":"2026091609115515900_cit20","doi-asserted-by":"publisher","first-page":"10101","DOI":"10.1093\/nar\/gkaa750","article-title":"Synthesis, chirality-dependent\nconformational and biological properties of siRNAs containing 5\u2032-(R)-\nand 5\u2032-(S)-C-methyl-guanosine","volume":"48","author":"Mikami","year":"2020","journal-title":"Nucleic\nAcids Res."},{"key":"2026091609115515900_cit21","doi-asserted-by":"publisher","first-page":"14517","DOI":"10.1021\/jacs.2c01679","article-title":"Role of\na \u201cMagic\u201d Methyl: 2\u2032-Deoxy-2\u2032-\u03b1-F-2\u2032-\u03b2-C-methyl\nPyrimidine Nucleotides Modulate RNA Interference Activity through\nSynergy with 5\u2032-Phosphate Mimics and Mitigation of Off-Target\nEffects","volume":"144","author":"Guenther","year":"2022","journal-title":"J. Am. Chem. Soc."},{"key":"2026091609115515900_cit22","doi-asserted-by":"publisher","first-page":"15","DOI":"10.3891\/acta.chem.scand.40b-0015","article-title":"Synthesis of three 3-C-hydroxymethylpentoses\nwith the D-ribo-D-xylo- and L-lyxo-configurations. Identification\nof the latter with a monosaccharide isolated from phase I Coxiella\nburnetii lipopolysaccharide","volume":"40","author":"Dahlman","year":"1986","journal-title":"Acta Chem. Scand.\nB"},{"key":"2026091609115515900_cit23","doi-asserted-by":"publisher","first-page":"498","DOI":"10.1021\/acsomega.9b03042","article-title":"Methoxymethyl\nThreofuranosyl Thymidine (4\u2019-MOM-TNA-T) at the T7 Position\nof the Thrombin-Binding Aptamer Boosts Anticoagulation Activity, Thermal\nStability, and Nuclease Resistance","volume":"5","author":"Varada","year":"2020","journal-title":"ACS Omega"},{"key":"2026091609115515900_cit24","doi-asserted-by":"publisher","first-page":"144","DOI":"10.1038\/s42004-025-01545-8","article-title":"An epimer\nof threose nucleic acid enhances oligonucleotide exonuclease resistance\nthrough end capping","volume":"8","author":"Wen","year":"2025","journal-title":"Commun. Chem."},{"key":"2026091609115515900_cit25","doi-asserted-by":"publisher","first-page":"881","DOI":"10.3390\/md10040881","article-title":"First Total\nSynthesis of a Naturally Occurring Iodinated 5\u2032-Deoxyxylofuranosyl\nMarine Nucleoside","volume":"10","author":"Sun","year":"2012","journal-title":"Mar. Drugs"},{"key":"2026091609115515900_cit26","doi-asserted-by":"publisher","first-page":"1.28.1","DOI":"10.1002\/0471142700.nc0128s50","article-title":"Solid-Phase Chemical\nSynthesis of 5\u2032-Triphosphate DNA, RNA, and Chemically Modified\nOligonucleotides","volume":"50","author":"Zlatev","year":"2012","journal-title":"Curr. Protoc. Nucleic Acid\nChem."},{"key":"2026091609115515900_cit27","doi-asserted-by":"publisher","first-page":"4.54.1","DOI":"10.1002\/0471142700.nc0454s52","article-title":"Synthesis of Threose\nNucleic Acid (TNA) Triphosphates\nand Oligonucleotides by Polymerase-Mediated Primer Extension","volume":"52","author":"Zhang","year":"2013","journal-title":"Curr. Protoc. Nucleic Acid Chem."},{"key":"2026091609115515900_cit28","doi-asserted-by":"publisher","first-page":"363","DOI":"10.1006\/jmbi.1997.1586","article-title":"Structural principles for the inhibition of the 3\u2032-5\u2032\nexonuclease activity of Escherichia coli DNA polymerase I by phosphorothioates","volume":"277","author":"Brautigam","year":"1998","journal-title":"J. Mol. Biol."},{"key":"2026091609115515900_cit29","doi-asserted-by":"publisher","first-page":"600","DOI":"10.1016\/j.molcel.2011.02.004","article-title":"Coupled\n5\u2032 nucleotide recognition\nand processivity in Xrn1-mediated mRNA decay","volume":"41","author":"Jinek","year":"2011","journal-title":"Mol. Cell"},{"key":"2026091609115515900_cit30","doi-asserted-by":"publisher","first-page":"1605","DOI":"10.1002\/jcc.20084","article-title":"UCSF Chimera\u2500a\nvisualization\nsystem for exploratory research and analysis","volume":"25","author":"Pettersen","year":"2004","journal-title":"J. Comput. Chem."},{"key":"2026091609115515900_cit31","doi-asserted-by":"publisher","first-page":"1668","DOI":"10.1002\/jcc.20290","article-title":"The Amber biomolecular simulation\nprograms","volume":"26","author":"Case","year":"2005","journal-title":"J. Comput. Chem."},{"key":"2026091609115515900_cit32","doi-asserted-by":"publisher","first-page":"137413","DOI":"10.1016\/j.cplett.2020.137413","article-title":"DNA\/TNA mesoscopic\nmodeling of melting temperatures suggests weaker hydrogen bonding\nof CG than in DNA\/RNA","volume":"749","author":"Muniz","year":"2020","journal-title":"Chem. Phys. Lett."},{"key":"2026091609115515900_cit33","doi-asserted-by":"publisher","first-page":"2798","DOI":"10.1021\/acs.jctc.4c01316","article-title":"Molecular\nModeling on Duplexes with\nThreose-Based TNA and TPhoNA Reveals Structural Basis for Different\nHybridization Affinity toward Complementary Natural Nucleic Acids","volume":"21","author":"Reynders","year":"2025","journal-title":"J. Chem. Theory Comput."},{"key":"2026091609115515900_cit34","doi-asserted-by":"publisher","first-page":"2425","DOI":"10.1021\/acs.biochem.4c00387","article-title":"DNA Replication across \u03b1-l-(3\u2032\u22122\u2032)-Threofuranosyl\nNucleotides Mediated by Human DNA Polymerase \u03b7","volume":"63","author":"Tomar","year":"2024","journal-title":"Biochemistry"},{"issue":"11","key":"2026091609115515900_cit35","doi-asserted-by":"publisher","first-page":"1173","DOI":"10.1038\/s41929-024-01233-1","article-title":"Directed\nevolution of a highly efficient TNA polymerase\nachieved by homologous recombination","volume":"7","author":"Maola","year":"2024","journal-title":"Nat. Catal."},{"key":"2026091609115515900_cit36","doi-asserted-by":"publisher","first-page":"5215","DOI":"10.1021\/cr200060g","article-title":"The Methylation Effect in Medicinal\nChemistry","volume":"111","author":"Barreiro","year":"2011","journal-title":"Chem. Rev."},{"key":"2026091609115515900_cit37","doi-asserted-by":"publisher","first-page":"1157","DOI":"10.3390\/ph16081157","article-title":"The Magic Methyl and Its Tricks in\nDrug Discovery and Development","volume":"16","author":"Pinheiro","year":"2023","journal-title":"Pharmaceuticals"},{"key":"2026091609115515900_cit38","doi-asserted-by":"publisher","first-page":"723","DOI":"10.1038\/s41467-018-02989-4","article-title":"Selection of GalNAc-conjugated\nsiRNAs with limited off-target-driven rat hepatotoxicity","volume":"9","author":"Janas","year":"2018","journal-title":"Nat. Commun."},{"key":"2026091609115515900_cit39","doi-asserted-by":"publisher","first-page":"10851","DOI":"10.1093\/nar\/gkab916","article-title":"Overcoming GNA\/RNA base-pairing\nlimitations using isonucleotides improves the pharmacodynamic activity\nof ESC+ GalNAc-siRNAs","volume":"49","author":"Schlegel","year":"2021","journal-title":"Nucleic Acids Res."},{"key":"2026091609115515900_cit40","doi-asserted-by":"publisher","first-page":"6656","DOI":"10.1093\/nar\/gkac539","article-title":"From bench to bedside:\nImproving the clinical safety\nof GalNAc-siRNA conjugates using seed-pairing destabilization","volume":"50","author":"Schlegel","year":"2022","journal-title":"Nucleic Acids Res."},{"key":"2026091609115515900_cit41","doi-asserted-by":"publisher","first-page":"402","DOI":"10.1261\/rna.079526.122","article-title":"Acyclic (S)-glycol nucleic acid (S-GNA)\nmodification of siRNAs improves the safety of RNAi therapeutics while\nmaintaining potency","volume":"29","author":"Egli","year":"2023","journal-title":"RNA"}],"container-title":["Journal of the American\nChemical Society"],"original-title":[],"language":"en","link":[{"URL":"https:\/\/pubs.acs.org\/doi\/pdf\/10.1021\/jacs.6c07051","content-type":"application\/pdf","content-version":"vor","intended-application":"unspecified"},{"URL":"https:\/\/pubs.acs.org\/jacsat\/article-pdf\/148\/36\/38692\/68223188\/jacs.6c07051.pdf","content-type":"application\/pdf","content-version":"vor","intended-application":"syndication"},{"URL":"https:\/\/pubs.acs.org\/jacsat\/article-pdf\/148\/36\/38692\/68223188\/jacs.6c07051.pdf","content-type":"unspecified","content-version":"vor","intended-application":"similarity-checking"}],"deposited":{"date-parts":[[2026,9,16]],"date-time":"2026-09-16T13:29:16Z","timestamp":1789565356000},"score":1,"resource":{"primary":{"URL":"https:\/\/pubs.acs.org\/jacsat\/article\/148\/36\/38692\/5406725\/Chiral-4-Methyl-l-Threofuranosyl-Nucleic-Acids"}},"subtitle":[],"short-title":[],"issued":{"date-parts":[[2026,9,2]]},"references-count":41,"journal-issue":{"issue":"36","published-online":{"date-parts":[[2026,9,2]]},"published-print":{"date-parts":[[2026,9,16]]}},"URL":"https:\/\/doi.org\/10.1021\/jacs.6c07051","relation":{},"ISSN":["0002-7863","1520-5126"],"issn-type":[{"value":"0002-7863","type":"print"},{"value":"1520-5126","type":"electronic"}],"subject":[],"published":{"date-parts":[[2026,9,2]]}}}