Oligonucleotide analogs having modified dimers

• US 6,420,549 B1
• Filed: 08/07/1998
• Issued: 07/16/2002
• Est. Priority Date: 06/06/1995
• Status: Expired due to Fees

• Current Assignee: ISIS Pharmaceuticals Incorporated (Ionis Pharmaceuticals, Inc.)
• Original Assignee: ISIS Pharmaceuticals Incorporated (Ionis Pharmaceuticals, Inc.)
• Inventors: Cook, Phillip Dan, Bhat, Balkrishen, Manoharan, Muthiah
• Primary Examiner(s): Riley, Jezia
• Application Number:
  US09/131,102
• Time in Patent Office: 1,439 Days
• Field of Search: 435/6, 435/7.1, 435/91.1, 435/91.2, 514/1, 514/44, 536/22.1, 536/23.1, 536/25.3, 536/24.2
• US Class Current: 536/24.2
• 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: C12N 15/1135 against oncogenes or tumor suppressor genes
• CPC Class Codes: C12N 2310/315 Phosphorothioates
• CPC Class Codes: C12N 2310/318 where the PO2 is completely replaced, e.g. MMI or formacetal
• CPC Class Codes: C12N 2310/321 2'-O-R Modification
• CPC Class Codes: C12N 2310/322 2'-R Modification
• CPC Class Codes: C12N 2310/341 Gapmers, i.e. of the type ===---===
• CPC Class Codes: C12N 2310/345 having at least two different backbone modifications
• CPC Class Codes: C12N 2310/346 having a combination of backbone and sugar modifications
• CPC Class Codes: C12N 2310/3525 MOE, methoxyethoxy
• CPC Class Codes: C12Q 1/6876 Nucleic acid products used in the analysis of nucleic acids, e.g. primers or probes
• CPC Class Codes: Y02P 20/582 Recycling of unreacted starting or intermediate materials

Abstract

Modified dimers having a ribose sugar moiety in the 5′ nucleoside and a 2′ modified sugar in the 3′ nucleoside are provided. The modified dimers are useful in the preparation of oligonucleotide analogs having enhanced properties compared to native oligonucleotides, including increased nuclease resistance, enhanced binding affinity and improved protein binding.

Claims

1. A compound having Formula:

wherein;

Z is a covalent intersugar linkage selected from the group consisting of 3′

-CH₂—

NH—

O—

, 3′

-CH₂—

N(CH₃)—

O—

, 3′

-amino phosphoramidate, and 3′

-amino phosphorothioamidate;

each T₁ and T₂ is, independently, —

OH, —

OR₁, —

CH₂R₁, —

NH(R₁), —

SH, —

SR₁, or a blocked hydroxyl;

R₁ is C₁-C₁₂ alkyl;

B_X is a heterocyclic base;

X is F, —

O—

R, —

S—

R or —

NR(R₂);

R is alkyl, or a ring system having from about 4 to about 7 carbon atoms or having from about 3 to about 6 carbon atoms and 1 or 2 hetero atoms wherein said hetero atoms are selected from oxygen, nitrogen and sulfur and wherein said ring system is aliphatic, unsaturated aliphatic, aromatic or heterocyclic;

and wherein any available hydrogen atom of said ring system is each replaceable with an alkoxy, alkylamino, urea or alkylurea group;

or R has one of the formulas;

 

wherein

Q is O, S or NR₂;

m is from 1 to 10;

y is from 0 to 10;

E is N(R₂)(R₃), N═

C(R₂)(R₃), C₁-C₁₀ alkyl, or C₁-C₁₀ substituted alkyl wherein said substituent is N(R₂)(R₃); and

each R₂ and R₃ is, independently, H, C₁-C₁₀ alkyl, alkylthioalkyl, a nitrogen protecting group, or R₂ and R₃, together, are a nitrogen protecting group or wherein R₂ and R₃ are joined in a ring structure that can include at least one heteroatom selected from N and O.

2. The compound of claim 1 wherein X is —

O—

R.

3. The compound of claim 2 wherein R is —

CH₃.

4. The compound of claim 2 wherein R is —

CH₂—

CH₂—

O—

CH₃.

5. The compound of claim 1 wherein Z is —

CH₂—

N(CH₃)—

O—

.

6. The compound of claim 2 wherein R is —

CH₂—

CH₂—

O—

N(CH₃)₂.

7. An oligonucleotide analog comprising at least one moiety having Formula:

wherein

each Z is a covalent intersugar linkage selected from the group consisting of 3′

-CH₂—

N—

H—

O—

, 3′

-CH₂—

N(CH₃)—

O—

, 3′

-amino phosphoramidate, and 3′

-amino phosphorothioamidate;

T₃ is a nucleotide other than a ribonucleotide, a nucleoside other than a ribonucleoside, a hydroxyl, a blocked hydroxyl, or an oligonucleotide wherein the 3′

-terminal nucleotide of said oligonucleotide is not a ribonucleotide;

T₄ is a nucleotide, a nucleoside, an oligonucleotide, a hydroxyl or a blocked hydroxyl;

with the proviso that at least one of said T₃ and T₄ is not a hydroxyl, or blocked hydroxyl;

B_X is a heterocyclic base;

each X is F, —

O—

R, —

S—

R or —

NR(R₂);

R is alkyl, or a ring system having from about 4 to about 7 carbon atoms or having from about 3 to about 6 carbon atoms and 1 or 2 hetero atoms wherein said hetero atoms are selected from oxygen, nitrogen and sulfur and wherein said ring system is aliphatic, unsaturated aliphatic, aromatic or heterocyclic;

and wherein any available hydrogen atom of said ring system is each replaceable with an alkoxy, alkylamino, urea or alkylurea group;

or R has one of the formulas;

 

wherein

Q is O, S or NR₂;

m is from 1 to 10;

y is from 1 to 10;

E is N(R₂)(R₃), N═

C(R₂)(R₃), C₁-C₁₀ alkyl, or C₁-C₁₀ substituted alkyl wherein said substituent is N(R₂)(R₃);

each R₂ and R₃ is, independently, H, C₁-C₁₀ alkyl, alkylthioalkyl, a nitrogen protecting group, or R₂ and R₃, together, are a nitrogen protecting group or wherein R₂ and R₃ are joined in a ring structure that can include at least one heteroatom selected from N and O; and

R₁ is H or C₁-C₁₂ alkyl.

8. The oligonucleotide analog of claim 7 wherein at least one X is —

O—

R.

9. The oligonucleotide analog of claim 8 wherein R is —

CH₃.

10. The oligonucleotide analog of claim 8 wherein R is —

CH₂—

CH₂—

O—

CH₃.

11. The oligonucleotide analog of claim 7 having a length of from 1 to 200 subunits.

12. The oligonucleotide analog of claim 7 having a length of from 10 to 25 subunits.

13. The oligonucleotide analog of claim 7 having a length of from 12 to 20 subunits.

14. The compound of claim 7 wherein Z is —

CH₂—

N(CH₃)—

O—

.

15. The oligonucleotide analog of claim 7 wherein R is —

CH₂—

CH₂—

O—

N(CH₃)₂.

16. A compound comprising a moiety of Formula:

wherein;

Z is a covalent intersugar linkage selected from the group consisting of 3′

-CH₂—

NH—

O—

, 3′

-CH₂—

N(CH₃)—

O—

, 3′

-amino phosphoramidate, and 3′

-amino phosphorothioamidate;

B_X is a heterocyclic base;

X is F, —

O—

R, —

S—

R or —

NR(R₂);

R is alkyl, or a ring system having from about 4 to about 7 carbon atoms or having from about 3 to about 6 carbon atoms and 1 or 2 hetero atoms wherein said hetero atoms are selected from oxygen, nitrogen and sulfur and wherein said ring system is aliphatic, unsaturated aliphatic, aromatic or heterocyclic;

and wherein any available hydrogen atom of said ring system is each replaceable with an alkoxy, alkylamino, urea or alkylurea group;

or R has one of the formulas;

 

wherein

Q is O, S or NR₂;

m is from 1 to 10;

y is from 0 to 10;

E is N(R₂)(R₃), N═

C(R₂)(R₃), C₁-C₁₀ alkyl, or C₁-C₁₀ substituted alkyl wherein said substituent is N(R₂)(R₃);

each R₂ and R₃ is, independently, H, C₁-C₁₀ alkyl, alkylthioalkyl, a nitrogen protecting group, or R₂ and R₃, together, are a nitrogen protecting group or wherein R₂ and R₃ are joined in a ring structure that can include at least one heteroatom selected from N and O;

T₃ is a nucleotide, a nucleoside, an oligonucleotide, a hydroxyl or a blocked hydroxyl, wherein said nucleotide is not a ribonucleotide, and said nucleoside is not a ribonucleoside, and wherein the 3′

-terminal nucleotide of said oligonucleotide is not a ribonucleotide; and

R₁ is H or C₁-C₁₂ alkyl.

17. The compound of claim 16 wherein X is —

O—

R.

18. The compound of claim 17 wherein R is —

CH₃.

19. The compound of claim 17 wherein R is —

CH₂—

CH₂—

O—

CH₃.

20. The compound of claim 16 wherein Z is —

CH₂—

N(CH₃)—

O—

.

21. The compound of claim 17 wherein R is —

CH₂—

CH₂—

O—

N(CH₃)₂.