Synthesis of 2'-O-[2-[(N,N-dimethylamino)oxy]ethyl] modified nucleosides and oligonucleotides.
A versatile synthetic route has been developed for the synthesis of 2'-O-[2-[(N,N-dimethylamino)oxy]ethyl] (abbreviated as 2'-O-DMAOE) modified purine and pyrimidine nucleosides and their corresponding nucleoside phosphoramidites and solid supports. To synthesize 2'-O-DMAOE purine nucleosides, the key intermediate B (Scheme 1) was obtained from the 2'-O-allyl purine nucleosides (13a and 15) via oxidative cleavage of the carbon-carbon bond to the corresponding aldehydes followed by reduction. To synthesize pyrimidine nucleosides, opening the 2,2'-anhydro-5-methyluridine 5 with the borate ester of ethylene glycol gave the key intermediate B. The 2'-O-(2-hydroxyethyl) nucleosides were converted, in excellent yield, by a regioselective Mitsunobu reaction, to the corresponding 2'-O-[2-[(1,3-dihydro-1,3-dioxo-2H-isoindol-2-yl)oxy]ethyl] nucleosides (18, 19, and 20). These compounds were subsequently deprotected and converted into the 2'-O-[2-[(methyleneamino)oxy]ethyl] derivatives (22, 23, and 24). Reduction and a second reductive amination with formaldehyde yielded the corresponding 2'-O-[2-[(N,N-dimethylamino)oxy]ethyl] nucleosides (25, 26, and 27). These nucleosides were converted to their 3'-O-phosphoramidites and controlled-pore glass solid supports in excellent overall yield. Using these monomers, modified oligonucleotides containing pyrimidine and purine bases were synthesized with phosphodiester, phosphorothioate, and both linkages (phosphorothioate and phosphodiester) present in the same oligonucleotide as a chimera in high yields. The oligonucleotides were characterized by HPLC, capillary gel electrophoresis, and ESMS. The effect of this modification on the affinity of the oligonucleotides for complementary RNA and on nuclease stability was evaluated. The 2'-O-DMAOE modification enhanced the binding affinity of the oligonucleotides for the complementary RNA (and not for DNA). The modified oligonucleotides that possessed the phosphodiester backbone demonstrated excellent resistance to nuclease with t(1/2) > 24 h.
Prakash
Thazha P
TP
Department of Medicinal Chemistry, Isis Pharmaceuticals Inc., 2292 Faraday Avenue, Carlsbad, California 92008, USA.
Kawasaki
Andrew M
AM
Fraser
Allister S
AS
Vasquez
Guillermo
G
Manoharan
Muthiah
M
D000818:Animals; D001483:Base Sequence; D002384:Catalysis; D002625:Chemistry, Organic / Q000379:methods; D002851:Chromatography, High Pressure Liquid; D018076:DNA, Complementary / Q000737:chemistry; D013058:Mass Spectrometry; D008968:Molecular Conformation; D008969:Molecular Sequence Data; D015394:Molecular Structure; D009693:Nucleic Acid Hybridization; D016376:Oligonucleotides, Antisense / Q000138:chemical synthesis* / Q000737:chemistry / Q000378:metabolism; D010084:Oxidation-Reduction; D043264:Phosphodiesterase I; D010727:Phosphoric Diester Hydrolases / Q000378:metabolism*; D011684:Purine Nucleosides / Q000138:chemical synthesis* / Q000737:chemistry / Q000378:metabolism; D011741:Pyrimidine Nucleosides / Q000138:chemical synthesis* / Q000737:chemistry / Q000378:metabolism; D018075:RNA, Complementary / Q000737:chemistry; D012910:Snake Venoms / Q000201:enzymology; D013379:Substrate Specificity
D018076:DNA, Complementary; D016376:Oligonucleotides, Antisense; D011684:Purine Nucleosides; D011741:Pyrimidine Nucleosides; D018075:RNA, Complementary; D012910:Snake Venoms; D010727:Phosphoric Diester Hydrolases; D043264:Phosphodiesterase I; C470790:phosphodiesterase I, snake venom