# US 2022/0275366 A1 · Rna interference mediated inhibition of gene expression using chemically modified short interfering nucleic acid (sina)

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- Number: US 2022/0275366 A1
- Document ID: US20220275366A1
- Status: Published application
- Kind: A1
- Application number: 17/497,747
- Filed: 2021-10-08
- Published: 2022-09-01
- Earliest claimed: 2001-05-18
- Assignees: Sirna Therapeutics, Inc.
- Applicants: Sirna Therapeutics, Inc.
- Inventors: James McSwiggen; Leonid Beigelman
- Web page: https://patentgrep.com/patent/US20220275366A1

## CPC

- C12N 15/113: Non-coding nucleic acids modulating the expression of genes, e.g. antisense oligonucleotides; {Antisense DNA or RNA; Triplex- forming oligonucleotides; Catalytic nucleic acids, e.g. ribozymes; Nucleic acids used in co-suppression or gene silencing (when used in plants C12N15/8218)}
- A61K 47/54: the modifying agent being an organic compound
- A61K 47/544: {Phospholipids}
- A61K 47/549: {Sugars, nucleosides, nucleotides or nucleic acids}
- A61K 47/551: {one of the codrug's components being a vitamin, e.g. niacinamide, vitamin B3, cobalamin, vitamin B12, folate, vitamin A or retinoic acid}
- A61K 47/554: {the modifying agent being a steroid plant sterol, glycyrrhetic acid, enoxolone or bile acid}
- A61K 47/60: the organic macromolecular compound being a polyoxyalkylene oligomer, polymer or dendrimer, e.g. PEG, PPG, PEO or polyglycerol
- A61K 47/64: Drug-peptide, drug-protein or drug-polyamino acid conjugates, i.e. the modifying agent being a peptide, protein or polyamino acid which is covalently bonded or complexed to a therapeutically active agent (peptidic linkers A61K47/65)
- C07H 21/02: with ribosyl as saccharide radical
- C12N 15/111: {General methods applicable to biologically active non-coding nucleic acids}
- C12N 15/1131: {against viruses}
- C12N 15/1138: {against receptors or cell surface proteins}
- C12N 15/8218: {Antisense, co-suppression, viral induced gene silencing [VIGS], post-transcriptional induced gene silencing [PTGS]}
- A61K 38/00: Medicinal preparations containing peptides (peptides containing beta-lactam rings A61K31/00; cyclic dipeptides not having in their molecule any other peptide link than those which form their ring, e.g. piperazine-2,5-diones, A61K31/00; ergot alkaloids of the cyclic peptide type A61K31/48; containing macromolecular compounds having statistically distributed amino acid units A61K31/74; medicinal preparations containing antigens or antibodies A61K39/00; medicinal preparations characterised by the non-active ingredients, e.g. peptides as drug carriers, A61K47/00)
- C12N 2310/111: spanning the whole gene, or a large part of it
- C12N 2310/14: interfering nucleic acids [NA]
- C12N 2310/315: Phosphorothioates
- C12N 2310/317: with an inverted bond, e.g. a cap structure
- C12N 2310/318: where the PO2 is completely replaced, e.g. MMI or formacetal
- C12N 2310/321: 2'-O-R Modification
- C12N 2310/322: 2'-R Modification
- C12N 2310/331: Universal or degenerate base
- C12N 2310/332: Abasic residue
- C12N 2310/346: having a combination of backbone and sugar modifications
- C12N 2310/351: Conjugate
- C12N 2310/3515: Lipophilic moiety, e.g. cholesterol
- C12N 2310/53: partially self-complementary or closed
- C12N 2320/11: for the determination of target sites, i.e. of active nucleic acids
- C12N 2320/51: modulating the chemical stability, e.g. nuclease-resistance
- C12N 2330/30: chemically synthesised
- C12N: MICROORGANISMS OR ENZYMES; COMPOSITIONS THEREOF; PROPAGATING, PRESERVING, OR MAINTAINING MICROORGANISMS; MUTATION OR GENETIC ENGINEERING; CULTURE MEDIA (microbiological testing media C12Q1/00)
- A61K: PREPARATIONS FOR MEDICAL, DENTAL OR TOILETRY PURPOSES (devices or methods specially adapted for bringing pharmaceutical products into particular physical or administering forms A61J3/00; chemical aspects of, or use of materials for deodorisation of air, for disinfection or sterilisation, or for bandages, dressings, absorbent pads or surgical articles A61L; soap compositions C11D)
- C07H: SUGARS; DERIVATIVES THEREOF; NUCLEOSIDES; NUCLEOTIDES; NUCLEIC ACIDS (derivatives of aldonic or saccharic acids C07C, C07D; aldonic acids, saccharic acids C07C59/105, C07C59/285; cyanohydrins C07C255/16; glycals C07D; compounds of unknown constitution C07G; polysaccharides, derivatives thereof C08B; DNA or RNA concerning genetic engineering, vectors, e.g. plasmids, or their isolation, preparation or purification C12N15/00; sugar industry C13)

## Abstract

The present invention concerns methods and reagents useful in modulating gene expression in a variety of applications, including use in therapeutic, diagnostic, target validation, and genomic discovery applications. Specifically, the invention relates to synthetic chemically modified small nucleic acid molecules, such as short interfering nucleic acid (siNA), short interering RNA (siRNA), double-stranded RNA (dsRNA), micro-RNA (miRNA), and short hairpin NA (shRNA) molecules capable of mediating RNA interference (RNAi) against target nucleic acid sequences. The small nucleic acid molecules are useful in the treatment of any disease or condition that responds to modulation of gene expression or activity in a cell, tissue, or organism.

## Claims

### Claim 1

A short interfering nucleic acid (siNA) molecule comprising a sense region and an antisense region that down-regulates expression of a target nucleic acid, wherein said siNA molecule does not require a 2′-hydroxyl group containing ribonucleotide, each strand of said double-stranded structure of the siNA molecule comprises about 21 nucleotides and the antisense region comprises a nucleotide sequence that is complementary to a nucleotide sequence of said target nucleic acid or a portion thereof, wherein said sense region is connected to the antisense region via a linker molecule.

### Claim 2 (depends on 1)

The siNA molecule of claim 1, wherein said siNA molecule is assembled from two separate oligonucleotide fragments wherein one fragment comprises the sense region and the second fragment comprises the antisense region of said siNA molecule.

### Claim 3 (depends on 2)

The siNA molecule of claim 2, wherein the 5′-end of the fragment comprising said antisense region optionally includes a phosphate group.

### Claim 4 (depends on 1)

The siNA molecule of claim 1 wherein said linker molecule is a polynucleotide linker.

### Claim 5 (depends on 1)

The siNA molecule of claim 1 wherein said linker molecule is a non-nucleotide linker.

### Claim 6 (depends on 4)

The siNA molecule of claim 4, wherein the polynucleotide linker is:
(i) greater or equal to 2 nucleotides in length; or
(ii) 3, 4, 5, 6, 7, 8, 9, or 10 nucleotides in length.

### Claim 7 (depends on 4)

The siNA molecule of claim 4, wherein the polynucleotide linker is an aptamer.

### Claim 8 (depends on 5)

The siNA molecule of claim 5, wherein said non-nucleotide linker comprises an abasic nucleotide, polyether, polyamine, polyamide, peptide, carbohydrate, lipid, polyhydrocarbon, or a polyethyelene glycol.

### Claim 9 (depends on 1)

The siNA of claim 1, wherein said target nucleic acid is an endogenous gene.

### Claim 10 (depends on 1)

The siNA molecule of claim 1, wherein said target nucleic acid is an exogenous gene.

### Claim 11 (depends on 1)

The siNA molecule of claim 1, wherein said target nucleic acid is a mammalian gene, plant gene, human gene, or a viral nucleic acid.

### Claim 12 (depends on 11)

The siNA molecule of claim 11, wherein said viral nucleic acid is:
(i) a mammalian viral nucleic acid chosen from the group consisting of: hepatitis C virus, human immunodeficiency virus, hepatitis B virus, herpes simplex virus, cytomegalovirus, human papilloma virus, respiratory syncytial virus, influenza virus, severe acute respiratory syndrome virus; or
(ii) a plant viral nucleic acid.

### Claim 13 (depends on 1)

The siNA of claim 1, wherein said target nucleic acid is an RNA.

### Claim 14 (depends on 1)

The siNA molecule of claim 1, wherein said siNA molecule comprises nucleotide sequence having complementarity to nucleotide sequence of RNA or a portion thereof encoded by said target nucleic acid or a portion thereof.

### Claim 15 (depends on 1)

The siNA molecule of claim 1, wherein each of sense and the antisense regions of said siNA molecule comprise 21 nucleotides.

### Claim 16 (depends on 2)

The siNA molecule of claim 2, wherein each of the two fragments of said siNA molecule comprise 21 nucleotides.

### Claim 17 (depends on 16)

The siNA molecule of claim 16, wherein about 19 nucleotides of each fragment of the siNA molecule are base-paired to the complementary nucleotides of the other fragment of the siNA molecule and wherein at least two 3′ terminal nucleotides of each fragment of the siNA molecule are not base-paired to the nucleotides of the other fragment of the siNA molecule.

### Claim 18 (depends on 16)

The siNA molecule of claim 16, wherein all 21 nucleotides of each fragment of the siNA molecule are base-paired to the complementary nucleotides of the other fragment of the siNA molecule.

### Claim 19 (depends on 16)

The siNA molecule of claim 16, wherein about 19 or 21 nucleotides of the antisense region of the siNA molecule are base-paired to the nucleotide sequence or a portion thereof of the target nucleic acid.

### Claim 20 (depends on 1)

A pharmaceutical composition comprising the siNA molecule of claim 1, in an acceptable carrier or diluent.