Compounds and methods for modulating target nuclear and sub-nuclear nucleic acid molecules in cells and animals

#The present application is being filed along with a Sequence Listing in electronic format. The Sequence Listing is provided as a file entitled CORE0093USC1SEQ_ST25.txt, created on Mar. 3, 2016 and is 32 Kb in size. The information in the electronic format of the sequence listing is incorporated herein by reference in its entirety.

#Antisense compounds have been shown to be effective for modulating the amount, activity, and/or function of cellular target nucleic acids. Certain chemical modifications have been incorporated into antisense oligonucleotides to enhance one or more properties of such antisense compounds. In certain instances, such chemically modified oligonucleotides have been shown to possess desirable characteristics compared to unmodified oligonucleotides, such as improved affinity for target and/or resistance to degradation. Chemically modified antisense oligonucleotides have been shown to have value as research tools, diagnostic reagents, and therapeutic agents, depending on the target nucleic acid and chemical modifications.

#Certain nucleic acid molecules have been shown to localize to cellular sub-organelles. Certain such nucleic acid molecules are RNA molecules, including non-coding RNA molecules. For example, small nucleolar RNA molecules (snoRNA) localize to the nucleolus inside the nucleus of eukaryotic cells. In certain instances, such snoRNA have been shown to be associated with precursors of ribosomal RNA (rRNA). Accordingly, certain snoRNAs have been reported to be involved in nucleotide modification and processing of pre-rRNA. Nucleic acids have also been found in Cajal bodies within the nucleus. RNA found in Cajal bodies have been referred to as small Cajal body-specific RNA (scaRNA). Certain scaRNA have been reported to be involved in nucleotide modification of spliceosomal small nuclear RNAs (snRNAs).

#FIG. 1 shows composition of the 5-10-5 RNA-DNA chimeric ASOs described in Example 1. All nucleotides are linked by phosphorothioate backbone (PS). “R” and “d” mean ribonucleoside and deoxyribonucleoside, respectively. “m” means 2′-O-methyloxylethyl nucleoside (MOE).

#FIG. 2 shows active ASOs described in Example 1. UTC=untreated cells. Lower panel shows U16 snoRNA regions targeted by the ASOs. The gray box indicates coding region of the snoRNA, whereas the white boxes represent the junctions between the coding region and flanking sequences. The thin line indicates flanking sequences.

#FIG. 3 shows structures for probing U16 snoRNA described in Example 1. The nucleotides accessible to DMS are identified.

#Unless specific definitions are provided, the nomenclature used in connection with, and the procedures and techniques of, analytical chemistry, synthetic organic chemistry, and medicinal and pharmaceutical chemistry described herein are those well known and commonly used in the art. Standard techniques may be used for chemical synthesis, and chemical analysis. Certain such techniques and procedures may be found for example in “Carbohydrate Modifications in Antisense Research” Edited by Sangvi and Cook, American Chemical Society, Washington D.C., 1994; “Remington's Pharmaceutical Sciences,” Mack Publishing Co., Easton, Pa., 18th edition, 1990; and “Antisense Drug Technology, Principles, Strategies, and Applications” Edited by Stanley T. Crooke, CRC Press, Boca Raton, Fla.; and Sambrook et al., “Molecular Cloning, A laboratory Manual,” 2nd Edition, Cold Spring Harbor Laboratory Press, 1989, which are hereby incorporated by reference for any purpose. Where permitted, all patents, applications, published applications and other publications and other data referred to throughout in the disclosure herein are incorporated by reference in their entirety.

#Unless otherwise indicated, the following terms have the following meanings:

#As used herein, “nucleoside” means a compound comprising a nucleobase moiety and a sugar moiety. Nucleosides include, but are not limited to, naturally occurring nucleosides (as found in DNA and RNA) and modified nucleosides. Nucleosides may be linked to a phosphate moiety.