Functional biopolymer modification reagents and uses thereof

• US 7,102,024 B1
• Filed: 08/01/2000
• Issued: 09/05/2006
• Est. Priority Date: 08/01/2000
• Status: Active Grant

• Current Assignee: Vector Laboratories
• Original Assignee: Solulink, Incorporated (Maravai Life Sciences Holdings LLC)
• Inventors: Schwartz, David A., Hogrefe, Richard I.
• Primary Examiner(s): Crane, L. E.
• Application Number:
  US09/630,060
• Time in Patent Office: 2,226 Days
• Field of Search: 558/70, 536/25.3, 536/23.1, 536/25.32, 536/26.8, 546/305
• US Class Current: 558/70
• CPC Class Codes: A61K 47/61 the organic macromolecular compound being a polysaccharide or a derivative thereof
• CPC Class Codes: A61K 47/62 the modifying agent being a protein, peptide or polyamino acid
• CPC Class Codes: A61K 47/643 Albumins, e.g. HSA, BSA, ovalbumin or a Keyhole Limpet Hemocyanin [KHL]
• CPC Class Codes: A61K 47/645 Polycationic or polyanionic oligopeptides, polypeptides or polyamino acids, e.g. polylysine, polyarginine, polyglutamic acid or peptide TAT
• CPC Class Codes: A61K 47/6801 Drug-antibody or immunoglobulin conjugates defined by the pharmacologically or therapeutically active agent
• CPC Class Codes: C04B 35/632 Organic additives
• CPC Class Codes: C07D 207/452 with hydrocarbon radicals, substituted by hetero atoms, directly attached to the ring nitrogen atom
• CPC Class Codes: C07D 207/46 with hetero atoms directly attached to the ring nitrogen atom
• CPC Class Codes: C07D 213/82 in position 3
• CPC Class Codes: C07D 401/12 linked by a chain containing hetero atoms as chain links
• CPC Class Codes: C07F 7/1804 Compounds having Si-O-C linkages Si-O-acyl linkages C07F7/1896
• CPC Class Codes: C07F 9/2408 of hydroxyalkyl compounds
• CPC Class Codes: C07F 9/58 Pyridine rings
• CPC Class Codes: C07H 21/00 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 Class Codes: C08G 65/329 with organic compounds

Abstract

Hydrazino, oxyamino and carbonyl-based reagents and methods for incorporation into oligonucleotides during their solid phase synthesis are provided. Modified oligonucleotides are provided that incorporate the reagents provided here in. Immobilized oligonucleotides and oligonucleotide conjugates that contain covalent hydrazone or oxime linkages are provided. Methods for preparation of surface bound oligonucleotides are provided. Methods for the preparation of oligonucleotide conjugates are also provided.

Claims

1. A compound that has formula (I):

P¹—

M—

X 

 

(1)

wherein;

P¹ is a phosphorous based coupling group selected from the group consisting of phosphoramidite, H-phosphonate and phosphotriester,

M is a divalent group of the formula;

—

(CH₂)_n—

NH—

(C═

R₁)—

R₂—

wherein R₁ is S or O;

R₂ is a direct bond, —

C₆H₄—

or —

C₅H₃N—

; and

n is 2, 3, 4, 5, 6, 7, 8, 9 or 10

X is selected from the group consisting of;

—

NHNH—

R₄, —

NHN═

C(CH₃)₂, —

(C═

O)CH₃, —

C═

O)H and —

ONHR₄

wherein R₄ is a protecting group.

2. The compound of claim 1, wherein R₄ is a protecting groups selected from the group consisting of selected from monomethoxytrityl (MMT), dimethoxytrityl (DMT), 9-fluorenylmethoxycarbonyl (FMOC) and trifluororacetyl (TFA).

3. The compound of claim 1, wherein P¹ is a phosphoramidite.

4. The compound of claim 1, wherein P¹ is H-phosphonate.

5. The compound of claim 1, wherein P¹ is phosphotriester.

6. The compound of claim 1, R₁ is O.

7. The compound of claim 1, R₁ is S.

8. The compound of claim 1, wherein R₂ is a direct bond.

9. The compound of claim 1, wherein R₂ is —

C₆H₄—

.

10. The compound of claim 1, wherein R₂ is —

C₅H₃N—

.

11. The compound of claim 1, wherein n is 2.

12. The compound of claim 1, wherein n is 6.

13. The compound of claim 1, wherein X is —

NHNH—

R₄—

.

14. The compound of claim 1, wherein X is —

NHN═

C(CH₃)₂—

.

15. The compound of claim 1, wherein X is —

C(═

O)CH₃.

16. The compound of claim 1, wherein X is —

C(═

O)H.

17. The compound of claim 1, wherein X is —

ONR₄—

.

18. The compound of claim 1, wherein R₄ is MMT.

19. The compound of claim 1, wherein R₄ is DMT.

20. The compound of claim 1, wherein R₄ is FMOC.

21. The compound of claim 1, wherein R₄ is TFA.

22. The compound of claim 1, wherein M has the formula:

24. The compound of claim 1, wherein M has the formula:

26. The compound of claim 1 that is N-monomethoxytrityl-O-{5-[8-2-cyanoethyl-N,N-diisopropylphosphoramidyloxy)-2-aza-1-oxooct-1-yl]pyrid-2-yl}oxyamine, having the structure:

23. The compounds N-dimethoxytrityl-N′

-{5-[8-(2-cyanoethyl-N, N-diisopropylphosphoramidyloxy)-2-aza-1-oxooct-1-yl]pyrid-2-yl}hydrazine and N-(9-fluorenylmethoxycarbonyl)-N′

-[8-(2-cyanoethyl-N,N-diisopropylphosphoramidyloxy)-2-aza-1-thiooct-1-yl]hydrazine, having the structures;

25. The compounds 4-[8-(2-cyanoethyl-N,N-diisopropylphosphoramnidyloxy)-2-aza-1-oxooct-1-yl]benzaldehyde, 4-[8-(2-cyanoethyl-N,N-diisopropylphosphoramidyl-oxy)-2-aza-1-oxooct-1-yl]acetophenone and N-[6-(2-cyanoethyl-N,N-diisopropylphosphoramnidyloxy)hexyl]4-ketopentanamide, having the structures: