# US 2025/0084418 A1 · Compositions and methods for silencing carbonic anhydrase 2 expression

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- Number: US 2025/0084418 A1
- Document ID: US20250084418A1
- Status: Published application
- Kind: A1
- Application number: 18/820,997
- Filed: 2024-08-30
- Published: 2025-03-13
- Earliest claimed: 2021-05-27
- Assignees: Alnylam Pharmaceuticals, Inc.
- Applicants: Alnylam Pharmaceuticals, Inc.
- Inventors: JEFFREY ZUBER; JAMES D. MCININCH; MARK K. SCHLEGEL; ADAM CASTORENO
- Web page: https://patentgrep.com/patent/US20250084418A1

## CPC

- C12N 15/1137: {against enzymes (viral enzymes C12N15/1131; receptors C12N15/1138)}
- A61K 31/713: Double-stranded nucleic acids or oligonucleotides
- A61K 47/542: {Carboxylic acids, e.g. a fatty acid or an amino acid}
- A61K 47/549: {Sugars, nucleosides, nucleotides or nucleic acids}
- C12Y 402/01001: Carbonate dehydratase (4.2.1.1), i.e. carbonic anhydrase
- C12N 2310/14: interfering nucleic acids [NA]
- C12N 2310/3125: Methylphosphonates
- C12N 2310/315: Phosphorothioates
- C12N 2310/323: modified ring structure
- C12N 2310/3341: 5-Methylcytosine
- C12N 2310/343: having patterns, e.g. ==--==--==--
- C12N 2310/346: having a combination of backbone and sugar modifications
- C12N 2310/351: Conjugate
- C12N 2310/3515: Lipophilic moiety, e.g. cholesterol
- C12Y 402/01: Hydro-lyases (4.2.1)
- 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)
- C12Y: ENZYMES

## Abstract

The disclosure relates to double-stranded ribonucleic acid (dsRNA) compositions targeting carbonic anhydrase 2 (CA2), and methods of using such dsRNA compositions to alter (e.g., inhibit) expression of carbonic anhydrase 2.

## Claims

### Claim 1

A double stranded ribonucleic acid (dsRNA) agent for inhibiting expression of Carbonic anhydrase 2 (CA2), wherein the dsRNA agent comprises a sense strand and an antisense strand forming a double stranded region, wherein the antisense strand comprises a nucleotide sequence comprising at least 15 contiguous nucleotides, with 0, 1, 2, or 3 mismatches, from one of the antisense sequences listed in any one of Tables 3-10, and wherein the sense strand comprises a nucleotide sequence comprising at least 15 contiguous nucleotides, with 0, 1, 2, or 3 mismatches, from a sense sequence listed in any one of Tables 3-10 that corresponds to the antisense sequence and wherein the dsRNA agent comprises at least one modified nucleotide.

### Claim 2 (depends on 1)

The dsRNA agent of claim 1, wherein at least one of the sense strand and the antisense strand is conjugated to one or more lipophilic moieties.

### Claim 3 (depends on 2)

The dsRNA agent of claim 2, wherein the lipophilic moiety is conjugated via a linker or carrier.

### Claim 4 (depends on 2, 3)

The dsRNA agent of claim 2 or 3, wherein one or more lipophilic moieties are conjugated to one or more internal positions on at least one strand.

### Claim 5 (depends on 4)

The dsRNA agent of claim 4, wherein the one or more lipophilic moieties are conjugated to one or more internal positions on at least one strand via a linker or carrier.

### Claim 6 (depends on 2, 3, 4, 5)

The dsRNA agent of any one of claims 2-5, wherein the lipophilic moiety is an aliphatic, alicyclic, or polyalicyclic compound.

### Claim 7 (depends on 6)

The dsRNA agent of claim 6, wherein the lipophilic moiety contains a saturated or unsaturated C16 hydrocarbon chain.

### Claim 8 (depends on 2, 3, 4, 5, 6, 7)

The dsRNA agent of any one of claims 2-7, wherein the lipophilic moiety is conjugated via a carrier that replaces one or more nucleotide(s) in the internal position(s) or the double stranded region.

### Claim 9 (depends on 2, 3, 4, 5, 6, 7)

The dsRNA agent of any one of claims 2-7, wherein the lipophilic moiety is conjugated to the double-stranded iRNA agent via a linker containing an ether, thioether, urea, carbonate, amine, amide, maleimide-thioether, disulfide, phosphodiester, sulfonamide linkage, a product of a click reaction, or carbamate.

### Claim 10 (depends on 2, 3, 4, 5, 6, 7, 8)

The double-stranded iRNA agent of any one of claims 2-8, wherein the lipophilic moiety is conjugated to a nucleobase, sugar moiety, or internucleosidic linkage.

### Claim 11

The dsRNA agent of any of the preceding claims, wherein no more than five of the sense strand nucleotides and not more than five of the nucleotides of the antisense strand are unmodified nucleotides.

### Claim 12

The dsRNA agent of any of the preceding claims, wherein all of the nucleotides of the sense strand and all of the nucleotides of the antisense strand comprise a modification.

### Claim 13

The dsRNA agent of any of the preceding claims, wherein at least one of the modified nucleotides is selected from the group consisting of a deoxy-nucleotide, a 3′-terminal deoxythimidine (dT) nucleotide, a 2′-O-methyl modified nucleotide, a 2′-fluoro modified nucleotide, a 2′-deoxy-modified nucleotide, a locked nucleotide, an unlocked nucleotide, a conformationally restricted nucleotide, a constrained ethyl nucleotide, an abasic nucleotide, a 2′-amino-modified nucleotide, a 2′-O-allyl-modified nucleotide, 2′-C-alkyl-modified nucleotide, a 2′-methoxyethyl modified nucleotide, a 2′-O-alkyl-modified nucleotide, a morpholino nucleotide, a phosphoramidate, a non-natural base comprising nucleotide, a tetrahydropyran modified nucleotide, a 1,5-anhydrohexitol modified nucleotide, a cyclohexenyl modified nucleotide, a nucleotide comprising a phosphorothioate group, a nucleotide comprising a methylphosphonate group, a nucleotide comprising a 5′-phosphate, a nucleotide comprising a 5′-phosphate mimic, a glycol modified nucleotide, and a 2-O—(N-methylacetamide) modified nucleotide; and combinations thereof.

### Claim 14

The dsRNA agent of any of the preceding claims, wherein at least one strand comprises a 3′ overhang of at least 2 nucleotides.

### Claim 15

The dsRNA agent of any of the preceding claims, wherein the double stranded region is 15-30 nucleotide pairs in length.

### Claim 16 (depends on 15)

The dsRNA agent of claim 15, wherein the double stranded region is 17-23 nucleotide pairs in length.

### Claim 17

The dsRNA agent of any of the preceding claims, wherein each strand has 19-30 nucleotides.

### Claim 18

The dsRNA agent of any of the preceding claims, wherein the agent comprises at least one phosphorothioate or methylphosphonate internucleotide linkage.

### Claim 19 (depends on 2, 3, 4, 5, 6, 7, 8, 9, 10, 11, 12, 13, 14, 15, 16, 17, 18)

The dsRNA agent of any one of claims 2-18, further comprising a targeting ligand, e.g., a ligand that targets an ocular tissue.

### Claim 20 (depends on 19)

The dsRNA agent of claim 19, wherein the ocular tissue is ciliary epithelium, an optic nerve, a trabecular meshwork, a juxtacanalicular tissue, a ganglion (e.g., including a retinal ganglion), episcleral veins or a Schlemm's canal (e.g., including an endothelial cell).

### Claim 21

The dsRNA agent of any one of the preceding claims, further comprising a phosphate or phosphate mimic at the 5′-end of the antisense strand.

### Claim 22 (depends on 21)

The dsRNA agent of claim 21, wherein the phosphate mimic is a 5′-vinyl phosphonate (VP).

### Claim 23 (depends on 1, 2, 3, 4, 5, 6, 7, 8, 9, 10, 11, 12, 13, 14, 15, 16, 17, 18, 19, 20, 21, 22)

The dsRNA of any one of claims 1-22 wherein the dsRNA agent targets a hotspot region of an mRNA encoding CA2.

### Claim 24

A dsRNA agent that targets a hotspot region of a Carbonic anhydrase 2 (CA2) mRNA.

### Claim 25 (depends on 1, 2, 3, 4, 5, 6, 7, 8, 9, 10, 11, 12, 13, 14, 15, 16, 17, 18, 19, 20, 21, 22, 23, 24)

A cell containing the dsRNA agent of any one of claims 1-24.

### Claim 26 (depends on 1, 2, 3, 4, 5, 6, 7, 8, 9, 10, 11, 12, 13, 14, 15, 16, 17, 18, 19, 20, 21, 22, 23, 24)

A pharmaceutical composition for inhibiting expression of a CA2, comprising the dsRNA agent of any one of claims 1-24 and a pharmaceutically acceptable buffer.

### Claim 27 (depends on 1, 2, 3, 4, 5, 6, 7, 8, 9, 10, 11, 12, 13, 14, 15, 16, 17, 18, 19, 20, 21, 22, 23, 24, 26)

A method of inhibiting expression of CA2 in a cell, the method comprising:
a. contacting the cell with the dsRNA agent of any one of claims 1-24, or a pharmaceutical composition of claim 26; and
b. maintaining the cell produced in step (a) for a time sufficient to reduce levels of CA2 mRNA, CA2 protein, or both of CA2 mRNA and protein, thereby inhibiting expression of CA2 in the cell.

### Claim 28 (depends on 27)

The method of claim 27, wherein the cell is within a subject.

### Claim 29 (depends on 28)

The method of claim 28, wherein the subject is a human.

### Claim 30 (depends on 29)

The method of claim 29, wherein the subject has been diagnosed with a CA2-associated disorder.

### Claim 31 (depends on 1, 2, 3, 4, 5, 6, 7, 8, 9, 10, 11, 12, 13, 14, 15, 16, 17, 18, 19, 20, 21, 22, 23, 24, 26)

A method of treating a subject diagnosed with a CA2-associated disorder comprising administering to the subject a therapeutically effective amount of the dsRNA agent of any one of claims 1-24 or a pharmaceutical composition of claim 26, thereby treating the disorder.

### Claim 32 (depends on 31)

The method of claim 31, wherein the CA2-associated disorder is glaucoma.

### Claim 33 (depends on 31, 32)

The method of claim 31 or 32, wherein treating comprises amelioration of at least one sign or symptom of the disorder.

### Claim 34 (depends on 31, 32, 33)

The method of any one of claims 31-33, wherein the treating comprises one or more of (a) inhibiting or reducing intraocular pressure; (b) inhibiting or reducing the expression or activity of CA2; (c) decreasing the amount of aqueous humor; (d) inhibiting or reducing optic nerve damage; (e) inhibiting or reducing retinal ganglion cell death; (f) medication to reduce intraocular pressure; (g) laser treatment; (h) surgery; (i) or trabeculectomy.

### Claim 35 (depends on 24, 25, 26, 27, 28, 29, 30, 31, 32, 33, 34)

The method of any one of claims 24-34, wherein the dsRNA agent is administered to the subject intraocularly, intravenously, or topically.

### Claim 36 (depends on 35)

The method of claim 35, wherein the intraocular administration comprises intravitreal administration (e.g., intravitreal injection), transscleral administration (e.g., transscleral injection), subconjunctival administration (e.g., subconjunctival injection), retrobulbar administration (e.g., retrobulbar injection), intracameral administration (e.g., intracameral injection), or subretinal administration (e.g., subretinal injection).

### Claim 37 (depends on 24, 25, 26, 27, 28, 29, 30, 31, 32, 33, 34, 35, 36)

The method of any one of claims 24-36, further comprising administering to the subject an additional agent or therapy comprising one or more of a prostaglandin analog, a beta blocker, an alpha-adrenergic agonist, a carbonic anhydrase inhibitor, or an anti-CA2 agent suitable for treatment or prevention of a CA2-associated disorder.