Oligonucleotide n3'-->p5' thiophosphoramidates: their synthesis and use
AssigneeGeron Corporation
InventorsSergei Gryaznov, Krisztina Pongracz, Tracy Matray
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Abstract
Oligonucleotides with a novel sugar-phosphate backbone containing at least one internucleoside 3′-NHP(O)(S−)O-5′ linkage, and methods of synthesizing and using the inventive oligonucleotides are provided. The inventive thiophosphoramidate oligonucleotides were found to retain the high RNA binding affinity of the parent oligonucleotide N3′→P5′ phosphoramidates and to exhibit a much higher acid stability.
Claims27
A polynucleotide comprising a non-homopolymeric sequence of nucleoside subunits joined by at least one inter-subunit linkage that is a N3′→P5′ thiophosphoramidate.
A polynucleotide comprising a sequence of nucleoside subunits containing at least one subunit defined by the formula:
- wherein B is a purine or pyrimidine or an analog thereof;
- Z is OR, SR, or methyl, wherein R is selected from the group consisting of hydrogen, alkyl, aryl and salts thereof;
- and
- R1 is selected from the group consisting of hydrogen, O—R2, S—R2, and halogen, wherein R2 is H, alkyl, or (CH2)nW(CH2)mH, where n is between 1-10, m is between 0-10 and W is O, S, or NH, with the proviso that when Z is methy or OMe, R1 is not H.
A method of synthesizing an oligonucleotide N3′→P5′ thiophosphoramidate, the method comprising:
- (a) providing a first 5′-succinyl-3′-aminotrityl-2′,3′-dideoxy nucleoside attached to a solid phase support, the first nucleoside having a protected 3′ amino group;
- (b) deprotecting the protected 3′ amino group to form a free 3′ amino group;
- (c) reacting the free 3′ amino group with a 3′-protected aminonucleoside-5′-O-cyanoethyl-N,N-diisopropylaminophosphoramidite monomer to form an internucleoside N3′→P5′ phosphoramidite linkage;
- and
- (d) sulfurizing the internucleaside phosphoramidite group to form a N3′→P5′ thiophosphoramidate.
Dependencies are inferred from the claim text.