METHOD FOR THE SOLID PHASE-BASED PRODUCTION OF PHOSPHATE-BRIDGED NUCLEOSIDE CONJUGATES

• US 20120116067A1
• Filed: 05/06/2010
• Published: 05/10/2012
• Est. Priority Date: 05/07/2009
• Status: Active Grant

• Current Assignee: Universität Hamburg
• Original Assignee: Universität Hamburg
• Inventors: Meier, Chris, Tonn, Viktoria Caroline
• Granted Patent: US 8,742,094 B2
• Time in Patent Office: Days
• US Class Current: 536/26.14
• CPC Class Codes: C07H 19/11 containing cyclic phosphate

Abstract

The invention relates to a method for producing phosphate-bridged nucleoside conjugates. In the method, a cyclosaligenyl nucleotide is produced first, to which a linker is added, which is used to perform the immobilization on a solid phase. A subsequent reaction with corresponding nucleophiles results in the desired phosphate-bridged nucleoside conjugates, which can then again be cleaved from the solid phase-bound linker.

Claims

1. A method for the solid-phase based production of a compound of the general formula (I):

2. The method according to claim 1, wherein in step a) a linker (L) according to the general formula (IV)

3. The method according to claim 2, wherein n is 2, and R⁵ and R⁶ are both H.

4. The method according to claim 1, wherein the linker (L) is covalently bound to an OH group of the sugar component of the nucleoside or to a component analogous to a sugar component of the nucleoside analog of R¹.

5. The method according to claim 4, wherein in step a) a compound according to the following general formula (IIIb)

6. The method according to claim 5, wherein B is one of the nucleobases guanine, adenine, cytosine, thymine or uracil.

7. The method according to claim 1, wherein the linker (L) is covalently bound to a nitrogen atom of the base component of the nucleoside or a nitrogen atom of a component analogous to a base component of the nucleoside analog of R¹.

8. The method according to claim 7, wherein the linker (L) is a benzoyl linker of the formula —

OC—

C₆H₄—

, wherein the bond to R¹ occurs via the functional group —

OC—

.

9. The method according to claim 1, wherein polystyrene, preferably aminomethyl polystyrene, is used as a solid phase.

10. The method according to claim 1, wherein X, in case of multiple substituents X independently from each other, is selected from the group consisting of H, Me, OMe, MeSO₂, ketone, formyl, ester, ═

O, C═

O, COOH, NO₂ and halogen.

11. The method according to claim 1, wherein the nucleophile is selected from the group consisting of phosphate, pyrophosphate, glycosyl phosphate, nucleoside, nucleoside monophosphate, nucleoside diphosphate, nucleoside triphosphate, nucleoside analog, nucleoside monophosphate analog, nucleoside diphosphate analog, nucleoside triphosphate analog, α

-deprotonated glycosyl, amines, amino acids, or salts thereof.

12. The method according to claim 1, wherein Nucl is selected from the group consisting of adenosine, guanosine, cytidine, thymidine, uridine, deoxyadenosine, deoxyguanosine, inosine, deoxycytidine, deoxyuridine, deoxythymidine, 2-thiocytidine, N⁴-acetyl-cytidine, 2′

-o-methyl-cytidine, 3-methyl-cytidine, 5-methyl-cytidine, 2-thiouridine, pseudouridine, dihydrouridine, 5-(carboxyhydroxymethyl)-uridine, 5-carboxymethylaminomethyl-uridine, 5-methylaminomethyl-uridine, 5-methoxy-carbonylmethyl-uridine, 5-methoxy-uridine, ribothymidine, 1-methyl-adenosine, 2-methyl-adenosine, N⁶-methyl-adenosine, inosine, 1-methyl-inosine, guanosine, N²-2,2-dimethyl-guanosine, N²-2-methyl-guanosine, 7⁺-methyl-guanosine and 2′

-O-methyl-guanosine.

13. The method according to claim 11, wherein the nucleoside or nucleoside analog Nucl in compound (II) and the nucleophile nucleoside or nucleophile nucleoside analog are the same or different, preferably different.

14. The method according to claim 1, characterized by the further step of

d) releasing the linker (L) from the residue R¹.

15. The method according to claim 1, wherein the method is carried out under inert gas atmosphere, preferably under nitrogen gas.

20. The compound according to claim 16, claim 1 Nucl is selected from the group consisting of adenosine, guanosine, cytidine, thymidine, uridine, deoxyadenosine, deoxyguanosine, inosine, deoxycytidine, deoxyuridine, deoxythymidine, 2-thiocytidine, N⁴-acetyl-cytidine, 2′

-O-methyl-cytidine, 3-methyl-cytidine, 5-methyl-cytidine, 2-thiouridine, pseudouridine, dihydrouridine, 5-(carboxyhydroxymethyl)-uridine, 5-carboxymethylaminomethyl-uridine, 5-methylaminomethyl-uridine, 5-methoxy-carbonylmethyl-u of uridine, 5-methoxy-uridine, ribo-thymidine, 1-methyl-adenosine, 2-methyl-adenosine, N⁶-methyl-adenosine, inosine, 1-methyl-inosine, guanosine, N²-2,2-dimethyl-guanosine, N²-2-methyl-guanosine, 7⁺-methyl-guanosine and 2′

-O-methyl-guanosine.

16. A compound of the general formula (IIIa)

18. The compound according to claim 16, wherein X, in case of multiple substituents X independently from each other, is selected from the group consisting of H, Me, OMe, MeSO₂, ketone, formyl, ester, ═

O, C═

O, COOH, NO₂ and halogen.

17. The compound of the general formula (IIIb)

19. The compound according to claim 17, wherein B is one of the nucleobases guanine, adenine, cytosine, thymine or uracil.