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Alias id Notation Equivalent Sequence
341401 ... AAGTAAGGACCAGAGACAA (Sense)
341391 ... TTGTCTCTGGTCCTTACTT (Antisense)
375761 ... TTGTCTCTGGTCCTTACTT (Antisense)
375762 ... TTGTCTCTGGTCCTTACTT (Antisense)
Key: o = phosphodiester linkage r = ribose sugar; s = 4'-thioribose sugar m = 2'-Omethylribose sugar x = 2'-0-methyl-4'-thioribose sugar
341391 375761 375762 UTC UTC UTC
341401 341401 341401
Treatment (ass)
1
OLIGOMERIC COMPOUNDS COMPRISING
4'-THIONUCLEOSIDES FOR USE IN GENE
MODULATION
CROSS-REFERENCE TO RELATED
APPLICATIONS
This application claims priority to U.S. provisional application Ser. No. 60/503,997 filed Sep. 18, 2003, which is incorporated herein by reference in its entirety.
FIELD OF THE INVENTION
The present invention provides monomeric and oligomeric compounds comprising 4'-thionucleosides. More particularly, the present invention provides oligomeric compounds and compositions comprising at least one 4'-thionucleoside of the invention. In some embodiments, the oligomeric compounds and compositions of the present invention hybridize to a portion of a target RNA resulting in loss of normal function of the target RNA.
BACKGROUND OF THE INVENTION
Targeting disease-causing gene sequences was first suggested more than thirty years ago (Belikova et al., Tet. Lett., 1967, 37, 3557-3562), and antisense activity was demonstrated in cell culture more than a decade later (Zamecnik et al, Proc. Natl. Acad. Sci. U.S.A., 1978, 75, 280-284). One advantage of antisense technology in the treatment of a disease or condition that stems from a disease-causing gene is that it is a direct genetic approach that has the ability to modulate (increase or decrease) the expression of specific disease-causing genes. Another advantage is that validation of a therapeutic target using antisense compounds results in direct and immediate discovery of the drug candidate; the antisense compound is the potential therapeutic agent.
Generally, the principle behind antisense technology is that an antisense compound hybridizes to a target nucleic acid and modulates gene expression activities or function, such as transcription or translation. The modulation of gene expression can be achieved by, for example, target degradation or occupancy-based inhibition. An example of modulation of RNA target function by degradation is RNase H-based degradation of the target RNA upon hybridization with a DNAlike antisense compound. Another example of modulation of gene expression by target degradation is RNA interference (RNAi). RNAi generally refers to antisense-mediated gene silencing involving the introduction of dsRNA leading to the sequence-specific reduction of targeted endogenous mRNA levels. Regardless of the specific mechanism, this sequencespecificity makes antisense compounds extremely attractive as tools for target validation and gene functionalization, as well as therapeutics to selectively modulate the expression of genes involved in the pathogenesis of malignancies and other diseases.
Antisense compounds have been employed as therapeutic agents in the treatment of disease states in animals, including humans. Antisense oligonucleotide drugs are being safely and effectively administered to humans in numerous clinical trials. In 1998, the antisense compound, Vitravene® (fomivirsen; developed by Isis Pharmaceuticals Inc., Carlsbad, Calif.) was the first antisense drug to achieve marketing clearance from the U.S. Food and Drug Administration (FDA), and is currently used in the treatment of cytomegalovirus (CMV)induced retinitis in AIDS patients. A New Drug Application (NDA) for GenasenseTM (oblimersen sodium; developed by
2
Genta, Inc., Berkeley Heights, N.J.), an antisense compound which targets the Bcl-2 mRNA overexpressed in many cancers, was accepted by the FDA. Many other antisense compounds are in clinical trials, including those targeting c-myc
5 (NeuGene® AVI-4126, AVI BioPharma, Ridgefield Park, N.J.), TNF-alpha (ISIS 104838, developed by Isis Pharmaceuticals, Inc.), VLA4 (ATL1102, Antisense Therapeutics Ltd, Toorak, Victoria, Australia) and DNA methyltransferase (MG98, developed by MGI Pharma, Bloomington, Minn.), to
10 name a few.
New chemical modifications have improved the potency and efficacy of antisense compounds, uncovering the potential for oral delivery as well as enhancing subcutaneous administration, decreasing potential for side effects, and lead
15 ing to improvements in patient convenience. Chemical modifications increasing potency of antisense compounds allow administration of lower doses, which reduces the potential for toxicity, as well as decreasing overall cost of therapy. Modifications increasing the resistance to degradation result in
20 slower clearance from the body, allowing for less frequent dosing. Different types of chemical modifications can be combined in one compound to further optimize the compound's efficacy.
Antisense technology is an effective means for reducing
25 the expression of one or more specific gene products and can therefore prove to be uniquely useful in a number of therapeutic, diagnostic, and research applications.
Consequently, there remains a long-felt need for agents that specifically regulate gene expression via antisense
30 mechanisms. Disclosed herein are antisense compounds useful for modulating gene expression pathways, including those relying on mechanisms of action such as RNaseH, RNAi and dsRNA enzymes, as well as other antisense mechanisms based on target degradation or target occupancy. One having
35 skill in the art, once armed with this disclosure will be able, without undue experimentation, to identify, prepare and exploit antisense compounds for these uses.
In many species, introduction of double-stranded RNA (dsRNA) induces potent and specific gene silencing. This
40 phenomenon occurs in both plants and animals and has roles in viral defense and transposon silencing mechanisms. This phenomenon was originally described more than a decade ago by researchers working with the petunia flower. While trying to deepen the purple color of these flowers, Jorgensen
45 et al. introduced a pigment-producing gene under the control of a powerful promoter. Instead of the expected deep purple color, many of the flowers appeared variegated or even white. Jorgensen named the observed phenomenon "cosuppression", since the expression of both the introduced gene and
50 the homologous endogenous gene was suppressed (Napoli et al. Plant Cell, 1990, 2, 279-289; Jorgensen et al. Plant Mol. Biol, 1996, 31, 957-973).
Cosuppression has since been found to occur in many species of plants, fungi, and has been particularly well char
55 acterized in Neurospora crassa, where it is known as "quelling" (Cogoni et al. Genes Dev. 2000, 10, 638-643; and Guru, Nature, 2000, 404, 804-808).
The first evidence that dsRNA could lead to gene silencing in animals came from work in the nematode, Caenorhabditis
60 elegans. In 1995, researchers Guo and Kemphues were attempting to use antisense RNA to shut down expression of the par-1 gene in order to assess its function. As expected, injection of the antisense RNA disrupted expression of par-1, but quizzically, injection of the sense-strand control also dis
65 rupted expression (Guo etal. Cell, 1995, 81, 611-620). This result was a puzzle until Fire et al. injected dsRNA (a mixture of both sense and antisense strands) into C. elegans. This
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