Method for the synthesis of phosphorus atom modified nucleic acids

Patent metadata

Field	Value
Publication number	US-10329318-B2
Application number	US-201715608123-A
Country	US
Kind code	B2
Filing date	May 30, 2017
Priority date	Dec 2, 2008
Publication date	Jun 25, 2019
Grant date	Jun 25, 2019

Abstract

Described herein are methods of syntheses of phosphorous atom-modified nucleic acids comprising chiral X-phosphonate moieties. The methods described herein provide backbone-modified nucleic acids in high diasteteomeric purity via an asymmetric reaction of an achiral molecule comprising a chemically stable H-phophonate moiety with a nucleoside/nucleotide.

First claim

The invention claimed is:

1. A method for the synthesis of a nucleic acid comprising a chiral X-phosphonate moiety comprising:
reacting a molecule comprising an achiral H-phosphonate moiety, a chiral reagent, and a nucleoside comprising a 5′-OH moiety to form a condensed intermediate; and
converting the condensed intermediate to the nucleic acid comprising a chiral X-phosphonate moiety, wherein:
the nucleic acid comprises a modified sugar, wherein the modified sugar contains one or more substituents at the 2′-position selected from the group consisting of F, CF 3 , CN, N 3 , NO, NO 2 , O-, S-, or N-alkyl; O-, S-, or N-alkenyl; O-, S-, or N-alkynyl; or O-alkyl-O-alkyl, O-alkyl-N-alkyl or N-alkyl-O-alkyl wherein the alkyl, alkenyl and alkynyl may be substituted or unsubstituted C 1 -C 10 alkyl or C 2 -C 10 alkenyl and alkynyl;
each instance of X is independently alkyl, alkoxy, aryl, alkylthio, acyl, —NR f R f , alkenyloxy, alkynyloxy, alkenylthio, alkynylthio, —S − Z + , —Se − Z + , or —BH 3 − Z + ;
each instance of R f is independently hydrogen, alkyl, alkenyl, alkynyl, or aryl;
Z + is ammonium ion, alkylammonium ion, heteroaromatic iminium ion, or heterocyclic iminium ion, any of which is primary, secondary, tertiary or quaternary, or Z + is a monovalent metal ion.

2. The method of claim 1 , wherein a substituent at the 2′-position of the modified sugar is O-alkyl-O-alkyl, wherein alkyl is substituted or unsubstituted C 1 -C 10 alkyl.

3. The method of claim 2 , wherein a substituent at the 2′-position of the modified sugar is —O(CH 2 ) n OCH 3 , wherein n is from 1 to about 10.

4. The method of claim 3 , wherein n is 1.

5. The method of claim 2 , wherein a substituent at the 2′-position of the modified sugar is O-methoxyethyl.

6. The method of claim 1 , wherein a substituent at the 2′-position of the modified sugar is F.

7. The method of claim 1 , wherein a substituent at the 2′-position of the modified sugar is O-alkyl, wherein alkyl is substituted or unsubstituted C 1 -C 10 alkyl.

8. The method of claim 1 , wherein the nucleic acid comprises one or more natural sugars of DNA or RNA.

9. The method of claim 1 , wherein the step of reacting the molecule comprising an achiral H-phosphonate moiety and the nucleoside comprising a 5′-OH moiety to form a condensed intermediate is a one-pot reaction.

10. The method of claim 1 , wherein the nucleic acid comprising a chiral X— phosphonate moiety is a compound of Formula 1:

wherein R 1 is —OH, —SH, —NR d R d , —N 3 , halogen, hydrogen, alkyl, alkenyl, alkynyl, alkyl-Y 1 —, alkenyl-Y 1 —, alkynyl-Y 1 —, aryl-Y 1 —, heteroaryl-Y 1 —, —P(O)(R e ) 2 , —HP(O)(R e ), —OR a or —SR c ;
Y 1 is O, NR d , S, or Se;
R a is a blocking moiety;
R e is a blocking group;
each instance of R d is independently hydrogen, alkyl, alkenyl, alkynyl, aryl, acyl, substituted silyl, carbamate, —P(O)(R e ) 2 , or —HP(O)(R e );
each instance of R e is independently hydrogen, alkyl, aryl, alkenyl, alkynyl, alkyl-Y 2 —, alkenyl-Y 2 —, alkynyl-Y 2 —, aryl-Y 2 —, or heteroaryl-Y 2 —, or a cation which is Na +1 , Li +1 , or K +1 ;
Y 2 is O, NH, N-alkyl, N-alkenyl, N-alkynyl, N-aryl, N-acyl, N-silyl, N-carbamate, or S;
each instance of R 2 is independently hydrogen, —OH, —SH, —NR d R d , —N 3 , halogen, alkyl, alkenyl, alkynyl, alkyl-Y 1 —, alkenyl-Y 1 —, alkynyl-Y 1 —, aryl-Y 1 —, heteroaryl-Y 1 —, —OR b , or —SR c , wherein R b is a blocking moiety;
R 3 is hydrogen, a blocking group, a linking moiety connected to a solid support or a linking moiety connected to a nucleic acid; and
n is an integer of 1 to about 200.

11. The method of claim 10 , wherein:
each instance of R d is independently hydrogen, alkyl, alkenyl, alkynyl, aryl, acyl, or substituted silyl;
each instance of R e is independently hydrogen, alkyl, aryl, alkenyl, alkynyl, alkyl-Y 2 —, alkenyl-Y 2 —, alkynyl-Y 2 —, aryl-Y 2 —, or heteroaryl-Y 2 ; and
Y 2 is O, NH, N-alkyl, N-alkenyl, N-alkynyl, N-aryl, N-acyl, N-silyl, N-carbamate, or S.

12. The method of claim 10 , wherein n is about 10 to about 200.

13. The method of claim 10 , wherein each X-phosphonate moiety of the compound of Formula 1 is more than 98% diastereomerically pure as determined by 31 P NMR spectroscopy or reverse-phase HPLC.

14. The method of claim 1 , wherein the chiral reagent is a compound of Formula 3:

wherein:
W 1 and W 2 are independently —NG 5 -, —O—, or —S—;
G 1 , G 2 , G 3 , G 4 , and G 5 are independently hydrogen, alkyl, aralkyl, cycloalkyl, cycloalkylalkyl, heterocyclyl, hetaryl, or aryl, or two of G 1 , G 2 , G 3 , G 4 , and G 5 are taken together to form G 6 a saturated, partially unsaturated or unsaturated carbocyclic or heteroatom-containing ring of up to about 20 ring atoms which is monocyclic or polycyclic, fused or unfused, and wherein no more than four of G 1 , G 2 , G 3 , G 4 , and G 5 are taken together to form are G 6 .

15. The method of claim 14 , wherein G 5 and one of G 3 and G 4 are taken together to form G 6 .

16. The method of claim 14 , wherein the chiral reagent is the compound of Formula 3 wherein W 1 is NG 5 and W 2 is O.

17. The method of claim 1 , wherein the chiral reagent is Formula O, Formula P, Formula Q or Formula R:

Assignees

• Wave Life Sciences Ltd

Inventors

• Wada Takeshi
• Shimizu Mamoru

Classifications

• C07H1/00

  Processes for the preparation of sugar derivatives · CPC title

• C07H21/00 Primary

  Compounds containing two or more mononucleotide units having separate phosphate or polyphosphate groups linked by saccharide radicals of nucleoside groups, e.g. nucleic acids · CPC title

• C12N15/11

  DNA or RNA fragments; Modified forms thereof (DNA or RNA not used in recombinant technology, C07H21/00); {Non-coding nucleic acids having a biological activity} · CPC title

• C07H21/02

  with ribosyl as saccharide radical · CPC title

• C07H19/20

  with the saccharide radical esterified by phosphoric or polyphosphoric acids · CPC title