The present disclosure provides a DNA-targeting RNA that comprises a targeting sequence and, together with a modifying polypeptide, provides for site-specific modification of a target DNA and/or a polypeptide associated with the target DNA. The present disclosure further provides site-specific modifying polypeptides. The present disclosure further provides methods of site-specific modification of a target DNA and/or a polypeptide associated with the target DNA The present disclosure provides methods of modulating transcription of a target nucleic acid in a target cell, generally involving contacting the target nucleic acid with an enzymatically inactive Cas9 polypeptide and a DNA-targeting RNA. Kits and compositions for carrying out the methods are also provided. The present disclosure provides genetically modified cells that produce Cas9; and Cas9 transgenic non-human multicellular organisms.

Among other things, the present disclosure provides technologies for altering splicing, particularly for increasing inclusion of exons in splicing products. In some embodiments, the present disclosure provides SMN2 oligonucleotides, compositions, and methods thereof. In some embodiments, the present disclosure provides chirally controlled SMN2 oligonucleotide compositions. In some embodiments, provided oligonucleotides and compositions can increase level of an exon 7-containing SMN2 splicing product and/or a gene product thereof. In some embodiments, the present disclosure provides methods for treatment of splicing-related conditions, disorders and diseases. In some embodiments, the present disclosure provides methods for treating SMN2-related conditions, disorders and diseases such as SMA (spinal muscular atrophy) and ALS (amyotrophic lateral sclerosis).

The present invention relates to antisense oligonucleotides that are complementary to mammalian CD73 (NT5E) pre-mRNA, wherein the antisense oligonucleotides are capable of modulating the splicing of mammalian CD73 pre-mRNA exon 7. Splice modulation of mammalian CD73 exon 7 is beneficial for a range of medical disorders, including disorders in the field of immune-oncology.

Disclosed herein are oligonucleotides, such as nucleic acid inhibitor molecules, having a 4′-phosphate analog and methods of using the same, for example, to modulate the expression of a target gene in a cell. The phosphate analogs are bound to the 4′-carbon of the sugar moiety (e.g., a ribose or deoxyribose or analog thereof) of the 5′-terminal nucleotide of an oligonucleotide. Typically, the phosphate analog is an oxymethylphosphonate, where the oxygen atom of the oxymethyl group is bound to the 4′-carbon of the sugar moiety or analog thereof.

The present disclosure provides compounds comprising modified oligonucleotides for use in CRISPR. In certain embodiments, such modified oligonucleotides provide improved properties of crRNA. In certain embodiments, such modified oligonucleotides provide improved properties of scrRNA.

The present invention provides RIG-I agonists. In certain embodiments, the agonists of the invention can be used to induce a type I interferon response in a cell.

Antisense molecules capable of binding to a selected target site in the human dystrophin gene to induce exon 53 skipping are described.

Provided is a polymerizable compound represented by a Formula E-1 or Formula E-2 shown in the description. In Formula E-1 or Formula E-2, R.sup.1 represents an alkoxy group, —NR.sup.N.sub.2, a hydroxy group, an aryl group, or an alkyl group, wherein R.sup.N each independently represents a hydrogen atom or an alkyl group having 1 to 10 carbon atoms; n represents an integer from 1 to 5; R.sup.3 represents a hydrogen atom, an acetyl group, a phenoxyacetyl group, a pivaloyl group, a benzyl group, a 4-methoxybenzyl group, a benzoyl group, a triphenylmethyl group, a 4,4′-dimethoxytrityl (DMTr) group, a 4-methoxytrityl (MMTr) group, a 9-phenylxanthenyl group, a trimethylsilyl group, a cyanomethoxymethyl group, a 2-(cyanoethoxy)ethyl group, or a cyanoethoxymethyl group; and X represents a structure represented by any one of Formula B-1 to Formula B-5 shown in the description.

Disclosed herein are double-stranded polyribonucleotides comprising a sense strand with 24 to 30 nucleotides in length and an antisense strand with 24 to 30 nucleotides in length, wherein the sense strand and the antisense strand form a fully complementary region of at least 24 base pairs with a blunt end at the 5′-end of the sense strand and the 3′-end of the antisense strand; and wherein the first 24 ribonucleotides at 5′-end of the sense strand further have at least one 2′-flourine modification at a ribonucleotide at a position selected from the group consisting of position number 2, 4, 6, 9, 10, 16, 21, 22, and 24, and no 2′-flourine modification at a ribonucleotide at a position selected from the group consisting of position number 1, 3, 8, and 14, and/or wherein the last 24 ribonucleotides at 3′-end of the antisense strand further have at least one 2′-flourine modification at a ribonucleotide at a position selected from the group consisting of position number 5, and 13, and no 2′-flourine modification at a ribonucleotide at a position selected from the group consisting of position 18 and 23; wherein all positions are counted from 5′ to 3′.

Antisense molecules capable of binding to a selected target site in the human dystrophin gene to induce exon 53 skipping are described.