Amido and peptido modified catalytic oligonucleotides

Details Amido and peptido modified catalytic oligonucleotides Amido and peptido modified catalytic oligonucleotides

2000-01-31

2002-04-30

McGarry, Sean (Department: 1635)

Chemistry: molecular biology and microbiology

Animal cell, per se ; composition thereof; process of...

C435S091310, C435S366000, C536S024300, C536S024500

Reexamination Certificate

active

06379954

ABSTRACT:

BACKGROUND OF THE INVENTION
This invention relates to modifications of oligonucleotides.
Usman et al., “Nucleozymes”, International Application No. PCT/US 93/00833, describes modification of the 2′-hydroxyl group of RNA to produce modified nucleotides. Such nucleotides are termed nucleic acid analogs, and may have a “good coordinating ligand” with divalent metal ions, e g, a halogen, or amine group. Acyclic analogs are also described.
Eckstein, International Application No. PCT/EP91/01811 (WO 92/07065), describes 2′-hydroxyl modifications of RNA having the following substitutions in place of the hydroxyl group: halo, sulfhydryl, azido, amino, mono-substituted amino and di-substituted amino.
Sproat et al., 1994 “Synthetic catalytic oligonucleotide structures”, U.S. Pat. No. 5,334,711, describes hammerhead ribozymes which contain nucleotides having 2′-OR modifications where, O represents oxygen and R represents H, or alkyl, alkenyl, or alkinyl.
Buhr and Matteucci, International Application No. WO 91/06556, describes 2′-hydroxyl modifications of antisense oligonucleotides with NHAC modifications.
SUMMARY OF THE INVENTION
This invention relates to replacement of the 2′-hydroxyl group of a ribonucleotide moiety with a 2′-amido or 2′-peptido moiety. In other embodiments, the 3′ and 5′ portions of the sugar of a nucleotide may be substituted, or the phosphate group may be substituted with amido or peptido moieties. Generally, such a nucleotide has the general structure shown in Formula I below:
The base (B) is any one of the standard bases or is a modified nucleotide base known to those in the art, or can be a hydrogen group. In addition, either R
1
or R
2
is H or an alkyl, alkene or alkyne group containing between 2 and 10 carbon atoms, or hydrogen, an amine (primary, secondary or tertiary, e.g., R
3
NR
4
where each R
3
and R
4
independently is hydrogen or an alkyl, alkene or alkyne having between 2 and 10 carbon atoms, or is a residue of an amino acid, i.e., an amide), an alkyl group, or an amino acid (D or L forms) or peptide containing between 2 and 5 amino acids. The zigzag lines represent hydrogen, or a bond to another base or other chemical moiety known in the art. Preferably, one of R
1
, R
2
and R
3
is an H, and the other is an amino acid or peptide.
Applicant has recognized that RNA can assume a much more complex structural form than DNA because of the presence of the 2′-hydroxyl group in RNA. This group is able to provide additional hydrogen bonding with other hydrogen donors, acceptors and metal ions within the RNA molecule. Applicant now provides molecules which have a modified amine group at the 2′ position, such that significantly more complex structures can be formed by the modified oligonucleotide. Such modification with a 2′-amido or peptido group leads to expansion and enrichment of the side-chain hydrogen bonding network. The amide and peptide moieties are responsible for complex structural formation of the oligonucleotide and can form strong complexes with other bases, and interfere with standard base pairing interactions. Such interference will allow the formation of a complex nucleic acid and protein conglomerate.
Oligonucleotides of this invention are significantly more stable than existing oligonucleotides and can potentially form biologically active bioconjugates not previously possible for oligonucleotides. They may also be used for in vitro selection of unique aptamers, that is, randomly generated oligonucleotides which can be folded into an effective ligand for a target protein, nucleic acid or polysaccharide.
Thus, in a first aspect, the invention features an oligonucleotide containing the modified base shown in Formula I, above.
In other aspects, the oligonucleotide may include a 3′ or 5′ nucleotide having a 3′ or 5′ located amino acid or aminoacyl group. In all these aspects, as well as the 2′-modified nucleotide, it will be evident that various standard modifications can be made. For example, an “O” may be replaced with an S, the sugar may lack a base (i.e., abasic) and the phosphate moiety may be modified to include other substitutions (see Sproat, supra).
Other features and advantages of the invention will be apparent from the following description of the preferred embodiments thereof, and from the claims.


REFERENCES:
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patent: 6093555 (2000-07-01), Dudycz et al.
patent: 91/06556 (1991-05-01), None
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