Anti-sense oligonucleotides targeted against exon 9 of IL-23R-alpha gene and method of using same to induce exon skipping and to treat inflammatory bowel diseases
The present invention relates to anti-sense oligonucleotides (AONs) used to induce exon 9 skipping in IL-23Rα gene. Exon 9 skipping of the IL23Rα gene ultimately causes specific induction of a novel soluble truncated IL-23Rα (Δ9) protein, characterized by a lack in a transmembrane domain and has a unique eight (8) amino acids (GLKEGSYC) at its C-terminus end as a result of frame-shift. The present invention provides a utility application of the use of AONs to induce production of a Δ9 protein which inhibits IL-23R-mediated cell signaling. More particularly, Δ9 protein blocks STAT3 formation as well as Th17 maturation. There is provided a therapeutic application of AONs in treating a mammal such as a human patient inflicted with Crohn's disease.
1. A method of producing a soluble truncated IL-23Rα protein, comprising the step of exposing an anti-sense RNA oligonucleotide to a mammalian cell,
wherein said anti-sense RNA oligonucleotide is targeted against exon 9 of IL-23Rα gene to induce skipping of exon 9 in said mammalian cell, and is selected from the group consisting of SEQ ID NO: 10, SEQ ID NO: 12, SEQ ID NO: 13, SEQ ID NO: 14, and SEQ ID NO: 15, and
wherein said anti-sense RNA oligonucleotide comprises a modified inter-nucleoside linkage.
2. The method of claim 1 , wherein said modified inter-nucleoside linkage is a phosphorothioate linkage.
3. The method of claim 1 , wherein said anti-sense oligonucleotide comprises a modified sugar moiety.
4. The method of claim 3 , wherein said modified sugar moiety is 2′-O-methyl sugar moiety.
5. The method of claim 1 , further comprising the step of quantifying said soluble truncated IL-23Rα protein after said exposing step.
6. The method of claim 5 , wherein said soluble truncated IL-23Rα protein is quantified using an enzyme-linked immunosorbent assay (ELISA).
7. The method of claim 1 , further comprising the step of quantifying mRNA of said soluble truncated IL-23Rα protein after said exposing step.
8. The method of claim 7 , wherein said mRNA of said soluble truncated IL-23Rα protein is quantified using qRT-PCR.
9. The method of claim 1 , where said mammalian cell is a peripheral blood mononuclear cell (PBMC), a CD4+ T cell, or a Th17 cell.