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New Antisense Strategies: Chemical Synthesis of RNA Oligomers

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Nucleic Acid Drugs

Part of the book series: Advances in Polymer Science ((POLYMER,volume 249))

Abstract

In this chapter, we introduce our recent work on the development of new antisense strategies in RNA synthetic technology and RNA drug discovery. In the first part of the chapter, we briefly introduce selected chemical modifications of antisense oligomers and recent developments in antisense RNA. In the second part, we describe our research on RNA, including synthetic approaches to stereodefined backbone-modified oligoribonucleotides with chiral linkages, and the development of a method for the synthesis of RNA based on the use of 2-cyanoethoxymethyl (CEM) as the 2′-hydroxyl protecting group. The CEM method has been applied to the synthesis of single and double short hairpin RNA, pre-microRNA, and designed mRNA molecules up to 170 nucleotides in length, which were physicochemically identified and shown to be biologically active. We hope that the work presented here will contribute to the pushing forward of the frontiers of knowledge and understanding in RNA biology and RNA drug discovery.

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Abbreviations

ACE:

Bis(2-acetoxyethoxy)methyl

ApbA:

Diadenosine 3′,5′-boranophosphate

Bcl-2:

B-cell lymphoma 2

BH3-ODN:

Boranophosphate oligodeoxynucleotide

BH3-ORN:

Boranophosphate oligoribonucleotide

BNA:

Bridged nucleic acid

CEE:

2-Cyanoethoxyethyl

CEM:

2-Cyanoethoxymethyl

CEM-Cl:

Chloromethyl ether

DMTr:

4,4′-Dimethoxytrityl

dshRNA:

Double short hairpin RNA

dsRNA:

Double-stranded RNA

ELISA:

Enzyme-linked immunosorbent assay

FDA:

Food and Drug Administration

GLP-1:

Glucagon-like peptide-1

HOX:

Homeobox

HPLC:

High-performance liquid chromatography

IFN:

Interferon

LNA:

Locked nucleic acid

miRNA:

micro RNA

mRNA:

Messenger RNA

NMR:

Nuclear magnetic resonance

NOE:

Nuclear Overhauser effect

nP1:

Nuclease P1

nt:

Nucleotide(s)

ODN:

Oligodeoxynucleotide

poly(A):

Polyadenylic acid

poly(U):

Polyuridylic acid

pre-miRNA:

Precursor micro RNA

PS-ODN:

Phosphorothioate oligodeoxynucleotide

PS-ORN:

Phosphorothioate oligoribonucleotide

RISC:

RNA-induced silencing complex

RNAi:

RNA interference

RNase H:

Ribonuclease H

RT-PCR:

Real-time polymerase chain reaction

shRNA:

Short hairpin RNA

siRNA:

Small interfering RNA

svPDE:

Snake venom phosphodiesterase

TBDMS:

2′-O-t-butyldimethylsilyl

TEM:

2-(4-Tolylsulfonyl)ethoxymethyl

TOM:

Triisopropylsilyloxymethyl

UTR:

Untranslated region

References

  1. Crooke ST (ed) (2001) Antisense drug technology: principles, strategies, and applications. Marcel Dekker, New York

    Google Scholar 

  2. Crooke ST (ed) (2008) Antisense drug technology: principles, strategies, and applications, 2nd edn. CRC, Boca Raton

    Google Scholar 

  3. Crooke ST, Lebleu B (eds) (1993) Antisense research and applications. CRC, Boca Raton

    Google Scholar 

  4. Zon G (1993) History of antisense drug discovery. In: Crooke ST, Lebleu B (eds) Antisense research and applications. CRC, Boca Raton, pp 1–5

    Google Scholar 

  5. Lima W, Wu H, Crooke ST (2008) The RNase H mechanism. In: Crooke ST (ed) Antisense drug technology: principles, strategies, and applications, 2nd edn. CRC, Boca Raton, pp 47–74

    Google Scholar 

  6. Zamecnik PC, Stephenson ML (1978) Inhibition of Rous sarcoma virus replication and cell transformation by a specific oligodeoxynucleotide. Proc Natl Acad Sci USA 75:280–284

    CAS  Google Scholar 

  7. Stephenson ML, Zamecnik PC (1978) Inhibition of Rous sarcoma viral RNA translation by a specific oligodeoxyribonucleotide. Proc Natl Acad Sci USA 75:285–288

    CAS  Google Scholar 

  8. Agrawal S (1999) Importance of nucleotide sequence and chemical modifications of antisense oligonucleotides. Biochim Biophys Acta 1489:53–68

    CAS  Google Scholar 

  9. Swayze EE, Bhat B (2008) The medicinal chemistry of oligonucleotides. In: Crooke ST (ed) Antisense drug technology: principles, strategies, and applications, 2nd edn. CRC, Boca Raton, pp 143–182

    Google Scholar 

  10. Monia BP, Yu RZ, Lima W et al (2008) Optimization of second-generation antisense drugs: going beyond generation 2.0. In: Crooke ST (ed) Antisense drug technology: principles, strategies, and applications, 2nd edn. CRC, Boca Raton, pp 487–506

    Google Scholar 

  11. Cech TR, Bass BL (1986) Biological catalysis by RNA. Annu Rev Biochem 55:599–629

    CAS  Google Scholar 

  12. Altman S (1989) Ribonuclease P: an enzyme with a catalytic RNA subunit. Adv Enzymol Relat Areas Mol Biol 62:1–36

    CAS  Google Scholar 

  13. Rossi JJ, Sarver N (1990) RNA enzymes (ribozymes) as antiviral therapeutic agents. Trends Biotechnol 8:179–183

    CAS  Google Scholar 

  14. Rossi JJ (1999) The application of ribozymes to HIV infection. Curr Opin Mol Ther 1:316–322

    CAS  Google Scholar 

  15. Goodchild J (2002) Hammerhead ribozymes for target validation. Expert Opin Ther Targets 6:235–247

    CAS  Google Scholar 

  16. Mulhbacher J, St-Pierre P, Lafontaine DA (2010) Therapeutic applications of ribozymes and riboswitches. Curr Opin Pharmacol 10:551–556

    CAS  Google Scholar 

  17. Fire A, Xu S, Montgomery MK et al (1998) Potent and specific genetic interference by double-stranded RNA in Caenorhabditis elegans. Nature 391:806–811

    CAS  Google Scholar 

  18. Hannon GJ (ed) (2003) RNAi: a guide to gene silencing, 2nd edn. Cold Spring Harbor Laboratory Press, Cold Spring Harbor

    Google Scholar 

  19. de Fougerolles AR, Maraganore JM (2008) Discovery and development of RNAi therapeutics. In: Crooke ST (ed) Antisense drug technology: principles, strategies, and applications, 2nd edn. CRC, Boca Raton, pp 465–484

    Google Scholar 

  20. Harborth J, Elbashir SM, Bechert K et al (2001) Identification of essential genes in cultured mammalian cells using small interfering RNAs. J Cell Sci 114:4557–4565

    CAS  Google Scholar 

  21. de Fougerolles A, Vornlocher HP, Maraganore J et al (2007) Interfering with disease: a progress report on siRNA-based therapeutics. Nat Rev Drug Discov 6:443–453

    Google Scholar 

  22. Castanotto D, Rossi JJ (2009) The promises and pitfalls of RNA-interference-based therapeutics. Nature 457:426–433

    CAS  Google Scholar 

  23. Li J, Liang Z (2010) The consideration of synthetic short interfering RNA for therapeutic use. Basic Clin Pharmacol Toxicol 106:22–29

    CAS  Google Scholar 

  24. Mattick JS (2005) The functional genomics of noncoding RNA. Science 309:1527–1528

    CAS  Google Scholar 

  25. Ghildiyal M, Zamore PD (2009) Small silencing RNAs: an expanding universe. Nat Rev Genet 10:94–108

    CAS  Google Scholar 

  26. Bartel DP (2004) MicroRNAs: genomics, biogenesis, mechanism, and function. Cell 116:281–297

    CAS  Google Scholar 

  27. Kim VN, Nam JW (2006) Genomics of microRNA. Trends Genet 22:165–173

    CAS  Google Scholar 

  28. Watanabe Y, Tomita M, Kanai A (2007) Computational methods for microRNA target prediction. Methods Enzymol 427:65–86

    CAS  Google Scholar 

  29. Weiler J, Hunziker J, Hall J (2006) Anti-miRNA oligonucleotides (AMOs): ammunition to target miRNAs implicated in human disease. Gene Ther 13:496–502

    CAS  Google Scholar 

  30. Hammond SM (2006) MicroRNA therapeutics: a new niche for antisense nucleic acids. Trends Mol Med 12:99–101

    CAS  Google Scholar 

  31. Miller PS, Yano J, Yano E et al (1979) Nonionic nucleic acid analogues. Synthesis and characterization of dideoxyribonucleoside methylphosphonates. Biochemistry 18:5134–5143

    CAS  Google Scholar 

  32. Miller PS, Ts’o POP, Hogrefe RI et al (1993) Anticode oligonucleoside methylphosphonates and their psoralen derivatives. In: Crooke ST, Lebleu B (eds) Antisense research and applications. CRC, Boca Raton, pp 189–203

    Google Scholar 

  33. Reynolds MA, Hogrefe RI, Jaeger JA et al (1996) Synthesis and thermodynamics of oligonucleotides containing chirally pure RP methylphosphonate linkages. Nucleic Acids Res 24:4584–4591

    CAS  Google Scholar 

  34. Cohen JS (1993) Phosphorothioate oligodeoxynucleotides. In: Crooke ST, Lebleu B (eds) Antisense research and applications. CRC, Boca Raton, pp 205–221

    Google Scholar 

  35. De Clercq E, Eckstein F, Sternbach H et al (1970) The antiviral activity of thiophosphate-substituted polyribonucleotides in vitro and in vivo. Virology 42:421–428

    Google Scholar 

  36. Summers JS, Shaw BR (2001) Boranophosphates as mimics of natural phosphodiesters in DNA. Curr Med Chem 8:1147–1155

    CAS  Google Scholar 

  37. Micklefield J (2001) Backbone modification of nucleic acids: synthesis, structure and therapeutic applications. Curr Med Chem 8:1157–1179

    CAS  Google Scholar 

  38. Rait VK, Shaw BR (1999) Boranophosphates support the RNase H cleavage of polyribonucleotides. Antisense Nucleic Acid Drug Dev 9:53–60

    CAS  Google Scholar 

  39. Hall AH, Wan J, Shaughnessy EE et al (2004) RNA interference using boranophosphate siRNAs: structure–activity relationships. Nucleic Acids Res 32:5991–6000

    CAS  Google Scholar 

  40. Sproat BS, Lamond AI (1993) 2′-O-Alkyloligoribonucleotides. In: Crooke ST, Lebleu B (eds) Antisense research and applications. CRC, Boca Raton, pp 351–362

    Google Scholar 

  41. Dean NM, Butler M, Monia BP et al (2001) Pharmacology of 2′-O-(2-methoxy)ethyl-modified antisense oligonucleotides. In: Crooke ST (ed) Antisense drug technology: principles, strategies, and applications. Marcel Dekker, New York, pp 319–338

    Google Scholar 

  42. Bennett CF (2008) Pharmacological properties of 2′-O-methoxyethyl-modified oligonucleotides. In: Crooke ST (ed) Antisense drug technology: principles, strategies, and applications, 2nd edn. CRC, Boca Raton, pp 273–303

    Google Scholar 

  43. Iribarren AM, Sproat BS, Neuner P et al (1990) 2′-O-Alkyl oligoribonucleotides as antisense probes. Proc Natl Acad Sci USA 87:7747–7751

    CAS  Google Scholar 

  44. Sproat BS, Lamond AI, Garcia RG et al (1991) 2′-O-Alkyloligoribonucleotides, synthesis and applications in molecular biology. Nucleic Acids Symp Ser 1991:59–62

    Google Scholar 

  45. Cotten M, Oberhauser B, Brunar H et al (1991) 2′-O-Methyl, 2′-O-ethyl oligoribonucleotides and phosphorothioate oligodeoxyribonucleotides as inhibitors of the in vitro U7 snRNP-dependent mRNA processing event. Nucleic Acids Res 19:2629–2635

    CAS  Google Scholar 

  46. Koch T, Ørum H (2008) Locked nucleic acid. In: Crooke ST (ed) Antisense drug technology: principles, strategies, and applications, 2nd edn. CRC, Boca Raton, pp 519–564

    Google Scholar 

  47. Wengel J (2001) LNA (locked nucleic acid). In: Crooke ST (ed) Antisense drug technology: principles, strategies, and applications. Marcel Dekker, New York, pp 339–357

    Google Scholar 

  48. Singh SK, Koshkin AA, Wengel J et al (1998) LNA (locked nucleic acids): synthesis and high-affinity nucleic acid recognition. Chem Commun 1998:455–456

    Google Scholar 

  49. Veedu RN, Wengel J (2009) Locked nucleic acid as a novel class of therapeutic agents. RNA Biol 6:321–323

    CAS  Google Scholar 

  50. Frieden M, Ørum H (2008) Locked nucleic acid holds promise in the treatment of cancer. Curr Pharm Des 14:1138–1142

    CAS  Google Scholar 

  51. Lanford RE, Hildebrandt-Eriksen ES, Petri A et al (2010) Therapeutic silencing of microRNA-122 in primates with chronic hepatitis C virus infection. Science 327:198–201

    CAS  Google Scholar 

  52. Abdur Rahman SM, Sato H, Tsuda N et al (2010) RNA interference with 2′,4′-bridged nucleic acid analogues. Bioorg Med Chem 18:3474–3480

    CAS  Google Scholar 

  53. Ketting RF, Tijsterman M, Plasterk RHA (2003) RNAi in Caenorhabditis elegans. In: Hannon GJ (ed) RNAi: a guide to gene silencing, 2nd edn. Cold Spring Harbor Laboratory Press, Cold Spring Harbor, pp 65–85

    Google Scholar 

  54. Sigova A, Zamore PD (2008) Small RNA silencing pathways. In: Crooke ST (ed) Antisense drug technology: principles, strategies, and applications, 2nd edn. CRC, Boca Raton, pp 75–88

    Google Scholar 

  55. Bartel DP (2009) microRNAs: target recognition and regulatory functions. Cell 136:215–233

    CAS  Google Scholar 

  56. Lewis BP, Burge CB, Bartel DP (2005) Conserved seed pairing, often flanked by adenosines, indicates that thousands of human genes are microRNA targets. Cell 120:15–20

    CAS  Google Scholar 

  57. Griffiths-Jones S (2004) The microRNA registry. Nucleic Acids Res 32(Database issue):D109–111

    Google Scholar 

  58. Griffiths-Jones S, Saini HK, van Dongen S et al (2008) miRBase: tools for microRNA genomics. Nucleic Acids Res 36(Database issue):D154–158

    Google Scholar 

  59. Griffiths-Jones S (2006) miRBase: the microRNA sequence database. Methods Mol Biol 342:129–138

    CAS  Google Scholar 

  60. Griffiths-Jones S, Grocock RJ, van Dongen S et al (2006) miRBase: microRNA sequences, targets and gene nomenclature. Nucleic Acids Res 34(Database issue):D140–144

    Google Scholar 

  61. Backes C, Meese E, Lenhof HP et al (2010) A dictionary on microRNAs and their putative target pathways. Nucleic Acids Res 38:4476–4486

    CAS  Google Scholar 

  62. Thum T, Gross C, Fiedler J et al (2008) MicroRNA-21 contributes to myocardial disease by stimulating MAP kinase signalling in fibroblasts. Nature 456:980–984

    CAS  Google Scholar 

  63. Melo SA, Esteller M (2011) Dysregulation of microRNAs in cancer: playing with fire. FEBS Lett 585:2087–2099

    Google Scholar 

  64. Medina PP, Slack FJ (2008) microRNAs and cancer: an overview. Cell Cycle 7:2485–2492

    CAS  Google Scholar 

  65. Krützfeldt J, Rajewsky N, Braich R et al (2005) Silencing of microRNAs in vivo with ‘antagomirs’. Nature 438:685–689

    Google Scholar 

  66. Esau C, Davis S, Murray SF et al (2006) miR-122 regulation of lipid metabolism revealed by in vivo antisense targeting. Cell Metab 3:87–98

    CAS  Google Scholar 

  67. De Guire V, Caron M, Scott N et al (2010) Designing small multiple-target artificial RNAs. Nucleic Acids Res. doi:10.1093/nar/gkq354

  68. Kosaka N, Iguchi H, Yoshioka Y et al (2010) Secretory mechanisms and intercellular transfer of microRNAs in living cells. J Biol Chem 285:17442–17452

    CAS  Google Scholar 

  69. Zhou SL, Wang LD (2010) Circulating microRNAs: novel biomarkers for esophageal cancer. World J Gastroenterol 16:2348–2354

    CAS  Google Scholar 

  70. Uno T, Hirabayashi K, Murai M (2005) The role of IFN regulatory factor-3 in the cytotoxic activity of NS-9, a polyinosinic-polycytidylic acid/cationic liposome complex, against tumor cells. Mol Cancer Ther 4:799–805

    CAS  Google Scholar 

  71. Yano J, Hirabayashi K, Nakagawa S et al (2004) Antitumor activity of small interfering RNA/cationic liposome complex in mouse models of cancer. Clin Cancer Res 10:7721–7726

    CAS  Google Scholar 

  72. Guga P, Stec WJ (2003) Synthesis of phosphorothioate oligonucleotides with stereodefined phosphorothioate linkages. Curr Protoc Nucleic Acid Chem 4.17:1–18 doi:10.1002/0471142700.nc0417s14

    Google Scholar 

  73. Lu Y (2006) Recent advances in the stereocontrolled synthesis of antisense phosphorothioates. Mini Rev Med Chem 6:319–330

    CAS  Google Scholar 

  74. Lesnikowski ZJ (1992) The first stereocontrolled synthesis of thiooligoribonucleotide: (RpRp)- and (SpSp)-UpsUpsU. Nucleosides Nucleotides 11:1621–1638

    CAS  Google Scholar 

  75. Sierzchała A, Okruszek A, Stec WJ (1996) Oxathiaphospholane method of stereocontrolled synthesis of diribonucleoside 3′,5′-phosphorothioates. J Org Chem 61:6713–6716

    Google Scholar 

  76. Almer H, Stawinski J, Strömberg R (1994) Synthesis of stereochemically homogeneous oligoribonucleoside all-R P-phosphorothioates by combining H-phosphonate chemistry and enzymatic digestion. J Chem Soc Chem Commun 1994:1459–1460

    Google Scholar 

  77. Almer H, Stawinski J, Strömberg R (1996) Solid support synthesis of all-Rp-oligo(ribonucleoside phosphorothioate)s. Nucleic Acids Res 24:3811–3820

    CAS  Google Scholar 

  78. Slim G, Gait MJ (1991) Configurationally defined phosphorothioate-containing oligoribonucleotides in the study of the mechanism of cleavage of hammerhead ribozymes. Nucleic Acids Res 19:1183–1188

    CAS  Google Scholar 

  79. Yano J, Ohgi T, Ueda T (2006) Optically active oligonucleic acid compound containing phosphorothioate bond. International application published under the patent cooperation treaty (PCT), WO 2006/043521 A1 (in Japanese with English abstract)

    Google Scholar 

  80. Oka N, Kondo T, Fujiwara S et al (2009) Stereocontrolled synthesis of oligoribonucleoside phosphorothioates by an oxazaphospholidine approach. Org Lett 11:967–970

    CAS  Google Scholar 

  81. Oka N, Yamamoto M, Sato T et al (2008) Solid-phase synthesis of stereoregular oligodeoxyribonucleoside phosphorothioates using bicyclic oxazaphospholidine derivatives as monomer units. J Am Chem Soc 130:16031–16037

    CAS  Google Scholar 

  82. Hyodo M, Ando H, Nishitani H et al (2005) Eur J Org Chem 2005:5216–5223

    Google Scholar 

  83. Oka N, Kondo T, Fujiwara S et al (2009) Stereocontrolled synthesis of oligoribonucleoside phosphorothioates by an oxazaphospholidine approach. Org Lett 11:967–970

    Google Scholar 

  84. Burgers PMJ, Eckstein F (1978) Absolute configuration of the diastereomers of adenosine 5′-O-(1-thiotriphosphate): consequences for the stereochemistry of polymerization by DNA-dependent RNA polymerase from Escherichia coli. Proc Natl Acad Sci USA 75:4798–4801

    CAS  Google Scholar 

  85. Bryant FR, Benkovic SJ (1979) Stereochemical course of the reaction catalyzed by 5′-nucleotide phosphodiesterase from snake venom. Biochemistry 18:2825–2829

    CAS  Google Scholar 

  86. Potter BVL, Connolly BA, Eckstein F (1983) Synthesis and configurational analysis of a dinucleoside phosphate isotopically chiral at phosphorus. Stereochemical course of Penicillium citrum nuclease P1 reaction. Biochemistry 22:1369–1377

    CAS  Google Scholar 

  87. Burgers PMJ, Eckstein F (1979) Diastereomers of 5′-O-adenosyl 3′-O-uridyl phosphorothioate: chemical synthesis and enzymatic properties. Biochemistry 18:592–596

    CAS  Google Scholar 

  88. Griffiths AD, Potter BVL, Eperon IC (1987) Stereospecificity of nucleases towards phosphorothioate-substituted RNA: stereochemistry of transcription by T7 RNA polymerase. Nucleic Acids Res 15:4145–4162

    CAS  Google Scholar 

  89. Sood A, Shaw BR, Spielvogel BF (1990) Boron-containing nucleic acids. 2. Synthesis of oligodeoxynucleoside boranophosphates. J Am Chem Soc 112:9000–9001

    CAS  Google Scholar 

  90. Li P, Sergueeva ZA, Dobrikov M et al (2007) Nucleoside and oligonucleoside boranophosphates: chemistry and properties. Chem Rev 107:4746–4796

    CAS  Google Scholar 

  91. Wada T, Shimizu M, Oka N et al (2002) A new boranophosphorylation reaction for the synthesis of deoxyribonucleoside boranophosphates. Tetrahedron Lett 43:4137–4140

    CAS  Google Scholar 

  92. Enya Y, Nagata S, Masutomi Y et al (2008) Chemical synthesis of diastereomeric diadenosine boranophosphates (ApbA) from 2′-O-(2-cyanoethoxymethyl)adenosine by the boranophosphotriester method. Bioorg Med Chem 16:9154–9160

    CAS  Google Scholar 

  93. Chen Y-Q, Qu F-C, Zhang Y-B (1995) Diuridine 3′,5′-boranophosphate: preparation and properties. Tetrahedron Lett 36:745–748

    CAS  Google Scholar 

  94. Li H, Huang F, Shaw BR (1997) Conformational studies of dithymidine boranomonophosphate diastereoisomers. Bioorg Med Chem 5:787–795

    CAS  Google Scholar 

  95. Tazawa I, Tazawa S, Stempel LM et al (1970) Conformation and interaction of dinucleoside mono- and diphosphates. 3. L-Adenylyl-(3′-5′)-L-adenosine and L-adenylyl-(2′-5′)-L-adenosine. Biochemistry 9:3499–3514

    CAS  Google Scholar 

  96. Ishiyama K, Smyth GE, Ueda T et al (2004) Homo-N-oligonucleotides (N1/N9−C1′ methylene bridge oligonucleotides): nucleic acids with left-handed helicity. J Am Chem Soc 126:7476–7485

    CAS  Google Scholar 

  97. Helm M, Brulé H, Giegé R, Florentz C (1999) More mistakes by T7 RNA polymerase at the 5′ ends of in vitro-transcribed RNAs. RNA 5:618–621

    CAS  Google Scholar 

  98. Ohgi T, Yano J (2010) Development of a novel synthetic method for RNA oligomers. In: Shioiri T, Izawa K, Konoike T (eds) Pharmaceutical process chemistry. Wiley-VCH, Weinheim, pp 345–362

    Google Scholar 

  99. Usman N, Ogilvie KK, Jiang MY et al (1987) Automated chemical synthesis of long oligoribonucleotides using 2′-O-silylated ribonucleoside 3′-O-phosphoramidites on a controlled-pore glass support: synthesis of a 43-nucleotide sequence similar to the 3′-half molecule of an Escherichia coli formylmethionine tRNA. J Am Chem Soc 109:7845–7854

    CAS  Google Scholar 

  100. Schwartz ME, Breaker RR, Asteriadis GT et al (1992) Rapid synthesis of oligoribonucleotides using 2′-O-(o-nitrobenzyloxymethyl)-protected monomers. Bioorg Med Chem Lett 2:1019–1024

    CAS  Google Scholar 

  101. Yamakage S, Sakatsume O, Furuyama E et al (1989) 1-(2-Chloroethoxy)ethyl group for the protection of 2′-hydroxyl group in the synthesis of oligoribonucleotides. Tetrahedron Lett 30:6361–6364

    CAS  Google Scholar 

  102. Matysiak S, Fitznar HP, Schnell R et al (1998) Nucleosides. Part XIII. Acetals as new 2′-O-protecting functions for the synthesis of oligoribonucleotides: synthesis of uridine building blocks and evaluation of their relative acid stability. Helv Chim Acta 81:1545–1566

    CAS  Google Scholar 

  103. Gasparutto D, Molko D, Téoule R (1990) Studies on the formation of the internucleotidic bond in RNA synthesis using dialkylamino phosphoroamidites. Nucleosides Nucleotides Nucleic Acids 9:1087–1098

    CAS  Google Scholar 

  104. Reese CB (2000) Protection of 2′-hydroxy functions of ribonucleosides. Curr Protoc Nucleic Acid Chem 2.2:1–24 doi:10.1002/0471142700.nc0202s00

    Google Scholar 

  105. Gough GR, Miller TJ, Mantick NA (1996) p-Nitrobenzyloxymethyl: a new fluoride-removable protecting group for ribonucleoside 2′-hydroxyls. Tetrahedron Lett 37:981–982

    CAS  Google Scholar 

  106. Welz R, Müller S (2002) 5-(Benzylmercapto)-1H-tetrazole as activator for 2′-O-TBDMS phosphoramidite building blocks in RNA synthesis. Tetrahedron Lett 43:795–797

    CAS  Google Scholar 

  107. Scaringe SA, Wincott FE, Caruthers MH (1998) Novel RNA synthesis method using 5′-O-silyl-2′-O-orthoester protecting groups. J Am Chem Soc 120:11820–11821

    CAS  Google Scholar 

  108. Pitsch S, Weiss PA, Jenny L et al (2001) Reliable chemical synthesis of oligoribonucleotides (RNA) with 2′-O-[(triisopropylsilyl)oxy]methyl(2′-O-tom)-protected phosphoramidites. Helv Chim Acta 84:3773–3795

    CAS  Google Scholar 

  109. Umemoto T, Wada T (2004) Oligoribonucleotide synthesis by the use of 1-(2-cyanoethoxy)ethyl (CEE) as a 2′-hydroxy protecting group. Tetrahedron Lett 45:9529–9531

    CAS  Google Scholar 

  110. Ohgi T, Masutomi Y, Ishiyama K et al (2005) A new RNA synthetic method with a 2′-O-(2-cyanoethoxymethyl) protecting group. Org Lett 7:3477–3480

    CAS  Google Scholar 

  111. Shiba Y, Masuda H, Watanabe N et al (2007) Chemical synthesis of a very long oligoribonucleotide with 2-cyanoethoxymethyl (CEM) as the 2′-O-protecting group: structural identification and biological activity of a synthetic 110mer precursor-microRNA candidate. Nucleic Acids Res 35:3287–3296

    CAS  Google Scholar 

  112. Ohgi T, Kitagawa H, Yano J (2008) Chemical synthesis of oligoribonucleotides with 2′-O-(2-cyanoethoxymethyl)-protected phosphoramidites. Curr Protoc Nucleic Acid Chem 2.15:1–19 doi:10.1002/0471142700.nc0215s34

    Google Scholar 

  113. Zhou C, Honcharenko D, Chattopadhyaya J (2007) 2-(4-Tolylsulfonyl)ethoxymethyl (TEM)—a new 2′-OH protecting group for solid-supported RNA synthesis. Org Biomol Chem 5:333–343

    CAS  Google Scholar 

  114. Murchison EP, Hannon GJ (2004) miRNAs on the move: miRNA biogenesis and the RNAi machinery. Curr Opin Cell Biol 16:223–229

    CAS  Google Scholar 

  115. Zhang Y, Zhang J, Hara H et al (2005) Insights into the mRNA cleavage mechanism by MazF, an mRNA interferase. J Biol Chem 280:3143–3150

    CAS  Google Scholar 

  116. Yekta S, Shih I-H, Bartel DP (2004) MicroRNA-directed cleavage of HOXB8 mRNA. Science 304:594–596

    CAS  Google Scholar 

  117. Watanabe N, Masuda H, Takagaki K et al (2007) Biological activity of a synthetic 110mer precursor-microRNA candidate. Poster presented at the 17th antisense symposium, Kanazawa, Japan. Antisense DNA/RNA Research Association (in Japanese with English abstract)

    Google Scholar 

  118. Matranga C, Tomari Y, Shin C et al (2005) Passenger-strand cleavage facilitates assembly of siRNA into Ago2-containing RNAi enzyme complexes. Cell 123:607–620

    CAS  Google Scholar 

  119. Rand TA, Petersen S, Du F et al (2005) Argonaute2 cleaves the anti-guide strand of siRNA during RISC activation. Cell 123:621–629

    CAS  Google Scholar 

  120. Gregory RI, Chendrimada TP, Cooch N et al (2005) Human RISC couples microRNA biogenesis and posttranscriptional gene silencing. Cell 123:631–640

    CAS  Google Scholar 

  121. Schwarz DS, Hutvágner G, Du T et al (2003) Asymmetry in the assembly of the RNAi enzyme complex. Cell 115:199–208

    CAS  Google Scholar 

  122. Khvorova A, Reynolds A, Jayasena SD (2003) Functional siRNAs and miRNAs exhibit strand bias. Cell 115:209–216

    CAS  Google Scholar 

  123. McManus MT, Conklin DS (2003) shRNA-mediated silencing of mammalian gene expression. In: Hannon GJ (ed) RNAi: a guide to gene silencing, 2nd edn. Cold Spring Harbor Laboratory Press, Cold Spring Harbor, pp 109–127

    Google Scholar 

  124. Siolas D, Lerner C, Burchard J et al (2005) Synthetic shRNAs as potent RNAi triggers. Nat Biotechnol 23:227–231

    CAS  Google Scholar 

  125. Ge Q, Dallas A, Ilves H et al (2010) Effects of chemical modification on the potency, serum stability, and immunostimulatory properties of short shRNAs. RNA 16:118–130

    CAS  Google Scholar 

  126. Bramsen JB, Laursen MB, Nielsen AF et al (2009) A large-scale chemical modification screen identifies design rules to generate siRNAs with high activity, high stability and low toxicity. Nucleic Acids Res 37:2867–2881

    CAS  Google Scholar 

  127. Masuda H, Watanabe N, Naruoka H et al (2010) Synthesis, gene-silencing activity and nuclease resistance of 3′-3′-linked double short hairpin RNA. Bioorg Med Chem 18:8277–8283

    CAS  Google Scholar 

  128. Judge AD, Bola G, Lee AC et al (2006) Design of noninflammatory synthetic siRNA mediating potent gene silencing in vivo. Mol Ther 13:494–505

    CAS  Google Scholar 

  129. Nagata S, Hamasaki T, Uetake K et al (2010) Synthesis and biological activity of artificial mRNA prepared with novel phosphorylating reagents. Nucleic Acids Res 38:7845–7857

    CAS  Google Scholar 

  130. Holst JJ (2007) The physiology of glucagon-like peptide 1. Physiol Rev 87:1409–1439

    CAS  Google Scholar 

  131. Kozak M (1987) An analysis of 5′-noncoding sequences from 699 vertebrate messenger RNAs. Nucleic Acids Res 15:8125–8148

    CAS  Google Scholar 

  132. Kuge H, Brownlee GG, Gershon PD et al (1998) Cap ribose methylation of c-mos mRNA stimulates translation and oocyte maturation in Xenopus laevis. Nucleic Acids Res 26:3208–3214

    CAS  Google Scholar 

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Acknowledgments

Although most of the research described in this chapter was carried out in our laboratories, we are pleased to introduce in Sect. 5.2 some of the excellent work carried out by Professor Takeshi Wada and his coworkers at the University of Tokyo. We thank Professor Wada for his kind permission to include this material. We are grateful to all of the individual researchers who carried out the work and to those who helped in the preparation of the manuscript, including Dr. Kazuchika Takagaki, Mr. Toshihiro Ueda, Dr. Hidetoshi Kitagawa, Messrs. Hirofumi Masuda, Seigo Nagata, and Kimihiko Higashiyama, and Ms. Yukiko Enya. We are especially grateful to Dr. Makoto Miyagishi of the National Institute of Advanced Industrial Science and Technology (AIST), Tsukuba, Japan, for his careful reading of the manuscript.

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  1. Discovery Research Laboratories, Nippon Shinyaku Co., Ltd, 3-14-1 Sakura, Tsukuba, Ibaraki, 305-0003, Japan

    Junichi Yano & Gerald E. Smyth

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  1. Junichi Yano
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  2. Gerald E. Smyth
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Correspondence to Junichi Yano .

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  1. Dept. Biomolecular Engineering, Kyoto Institute of Technology, Matsugasaki, Kyoto, 606-8585, Japan

    Akira Murakami

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Yano, J., Smyth, G.E. (2011). New Antisense Strategies: Chemical Synthesis of RNA Oligomers. In: Murakami, A. (eds) Nucleic Acid Drugs. Advances in Polymer Science, vol 249. Springer, Berlin, Heidelberg. https://doi.org/10.1007/12_2011_136

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