Metalloligands for cleaving nucleic acids

Details Metalloligands for cleaving nucleic acids Metalloligands for cleaving nucleic acids

1999-07-06

2002-06-11

Wilson, James O. (Department: 1623)

Organic compounds -- part of the class 532-570 series

Organic compounds

Carbohydrates or derivatives

C536S004100, C536S013100, C536S013200, C536S013600, C536S014000, C536S016600, C536S025400, C536S025410, C536S025420, C536S055100, C536S055300, C536S101000, C536S121000, C536S124000, C435S074000, C435S081000, C435S082000, C435S083000, C549S230000, C549S273000, C549S414000, C549S415000

Reexamination Certificate

active

06403777

ABSTRACT:

BACKGROUND
At present in vitro degradation of nucleic acids is accomplished using enzymes known as nucleases. Nucleases which catalyze random scission of DNA and RNA are referred to, respectively, as DNases and RNases. DNases and RNases are used to purify and isolate proteins from cellular extracts Unfortunately, nucleases are not stable at ambient temperature, much less at physiological temperatures. Moreover, nucleases are sensitive to phosphate, and, thus, are not useful for cleaving nucleic acids in solutions comprising phosphate ions.
DNAses are also used to footprint DNA, i.e., to map those regions of isolated DNA that act as binding sites for proteins or peptides that function as transcription factors. However, DNases are relatively large macromolecules and, therefore, cannot identify the exact point of contact between the protein and the substrate DNA. Attempts have been made to overcome this problem of poor resolution by developing smaller molecules such as for example iron EDTA and copper dimethylphenanthroline, both of which are capable of catalyzing the cleavage of nucleic acids, particularly DNA. However, these small molecules produce reactive species which are diffusible and which may chemically react non-specifically, thereby leading to removal of the bases from the DNA molecule.
Furthermore, RNases are not selective, i.e., they do not preferentially catalyze cleavage of any particular RNA structure or sequence. Accordingly, RNases are not useful to characterize the secondary and tertiary structures RNA molecules assume in solution and, thus, they cannot be used to identity those RNA regions with which transcription factors and translation factors are believed to interact.
Thus, it is desirable to have new compounds and methods for catalyzing the cleavage of nucleic acids.
SUMMARY OF THE INVENTION
The present invention provides soluble transition metal complexes, referred to hereinafter as “metalloligands”, that catalyze the degradation of DNA under hydrolytic conditions or mild oxidative conditions. Advantageously, the metalloligands also selectively catalyze the cleavage of RNA in regions of secondary and tertiary structure. In one embodiment, the metalloligand has the following structure:
wherein R
1
is an amino group, i.e. an NH, or an alkylamino group comprising 1 or 2 carbon atoms; wherein R
2
is selected from the group consisting of an amino group, a hydroxyl group, i.e., O(H), an alkylamino group comprising 1 or 2 carbon atoms; and an alkylhydroxyl group comprising 1 or 2 carbon atoms; wherein J is a ligand which comprises at least one carbon-containing five-membered or six-membered ring structure; and wherein M is a transition metal ion which is bound via coordinate bonds to R
1
and R
2
.
In another embodiment the metalloligand has the following structure:
wherein R
1
′ and R
1
″ are the same or different and wherein R1 ′ and R1″ are an amino group or an alkylamino group comprising 1 or 2 carbon atoms; wherein R2′ and R2″ are the same or different and wherein R2′ and R2″ are selected from the group consisting of an amino group, a hydroxyl group, an alkylamino group comprising 1 or 2 carbon atoms, and an alkylhydroxyl group comprising one or two carbon atoms; wherein J′ and J″ are the same or different and wherein J′ and J″ are ligands which comprise at least one carbon-containing five-membered or six-membered ring structure; and wherein M is a transition metal ion which is bound via coordinate bonds to R1′, R1″, R2′ and R2″.
The present invention also relates to methods of making the metalloligands. The present invention also relates to methods of using the metalloligands to catalyze the cleavage or degradation of DNA and to catalyze the cleavage of RNA at select sites, i.e. at sites which assume a certain secondary and tertiary structural motif when the RNA is in solution.


REFERENCES:
patent: 5665890 (1997-09-01), Jacobsen et al.
patent: 5929232 (1999-07-01), Jacobsen et al.
patent: WO 00/01797 (2000-01-01), None
“Efficient catalytic cleavage of DNA mediated by metalloaminoglycosides” by Sreedhara, et al.,Chem. Cummun.1998, pp. 1737-1738.
“Novel reagents for targeted cleavage of RNA sequences: towards a new family of inorganic pharmaceuticals” by Sreedhara, et al.,Chem. Commun.,1998, pp. 1147-1148.
“Specific binding of aminoglycoside antibiotics to RNA” by Wang, et al.,Chemistry & Biology,May 1995, 2:281-290.
“Small Molecules That Selectively Block RNA Binding of HIV-1 Rev Protein Inhibit Rev. Function and Viral Production” by Zapp, et al.,Cell,vol. 74, Sep. 24, 1993, pp. 939-978.

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