Modified small interfering RNA molecules and methods of use

Abstract

The present invention provides double-stranded RNA molecules that mediate RNA interference in target cells, preferably hepatic cells. The invention also provides double-stranded RNA molecules that are modified to be resistant to nuclease degradation, which inactivates a virus, and more specifically, hepatitis C virus (HCV). The invention also provides a method of using these modified RNA molecules to inactivate virus in mammalian cells and a method of making modified small interfering RNAs (siRNAs) using human Dicer.

Description

BACKGROUND OF THE INVENTION [0001] The present invention relates to the field of nucleic acid detection and to the phenomenon of RNA silencing, or RNA interference (RNAi). RNA silencing constitutes a phenomenon wherein non-coding RNA molecules mediate specific gene suppression in an organism. In nature, the phenomenon protects an organism's genome from foreign, invading nucleic acids such as transposons, trangenes and viral genes. [0002] The introduction of double-stranded RNA (dsRNA) into a cell triggers RNA silencing, which then degrades endogenous MRNA corresponding to the dsRNA. RNA silencing pathways involve a conversion of dsRNA into short interfering RNAs (siRNAs) that direct ribonucleases to homologous mRNA targets (Baulcombe et al., 2001). An enzyme called Dicer processes the dsRNA into siRNAs, which are 20-25 nucleotides long. The siRNAs then assemble into endoribonuclease-containing complexes known as RNA-induced silencing complexes (RISCs). Subsequently, the siRNAs guide...

AI Technical Summary

Benefits of technology

[0007] The present invention provides double-stranded RNA (dsRNA) molecules that mediate RNA interference in target cells. In particular, it provides small interfering RNAs (siRNAs) that inhibit viral replication in infected cells. Preferred dsRNA molecules of the invention correspond to hepatitis C virus (HCV) nucleic acids, and inhibit replication of HCV in hepatic cells.

[0008] In another aspect, the invention provides modified dsRNA, including siRNA, molecules that are protected against nuclease degradation, but are able to inhibit viral replication in mammalian cells.

[0009] The invention also provides methods of inhibiting viral replication in infected cells by administering dsRNA or siRNA molecules. Modified dsRNA and siRNA molecules are particularly useful in these methods, as they are nuclease resistant, yet retain the biological activity of being able to inhibit viral rep

Problems solved by technology

However, it is not clear that all RNA or DNA sequences of a mammalian cell's genome are susceptible to siRNA.

It is also uncertain that every mammalian cell type possesses the necessary machinery for effectuating gene-specific suppression using siRNA.

Further, siRNA is of limited use for at least two reasons: (a) the transient nature of the suppression effect seen in cells where the siRNA has been administered, and (b) the necessity for chemical synthesis of siRNAs before their use (Tuschl, T., Nature Biotech., 20: 446-448 (2002)).

Also, since siRNAs are unstable in vivo, their long-term effectiveness is limited.

Current therapies for such viral infections are very limited, and tend to have poor response rates.

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