# US 2026/0308602 A1 · Nucleic acids encoding crispr-associated proteins and uses thereof

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- Number: US 2026/0308602 A1
- Document ID: US20260308602A1
- Status: Published application
- Kind: A1
- Application number: 19/712,910
- Filed: 2026-06-18
- Published: 2026-10-08
- Earliest claimed: 2023-07-03
- Assignees: CureVac AG
- Applicants: CureVac SE
- Inventors: Frédéric CHEVESSIER-TÜNNESEN; Marion POENISCH; Thomas SCHLAKE
- Web page: https://patentgrep.com/patent/US20260308602A1

## CPC

- C12N 15/67: General methods for enhancing the expression
- A61K 31/7088: Compounds having three or more nucleosides or nucleotides
- A61K 48/0066: {Manipulation of the nucleic acid to modify its expression pattern, e.g. enhance its duration of expression, achieved by the presence of particular introns in the delivered nucleic acid}
- C12N 9/22: Ribonucleases {[RNase]; Deoxyribonucleases [DNase]}
- 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)}
- C12N 15/62: DNA sequences coding for fusion proteins
- C12N 2310/20: involving clustered regularly interspaced short palindromic repeats [CRISPR]
- C12N 2310/531: Stem-loop; Hairpin
- 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)

## Abstract

The present invention relates to the field of biomedicine, and in particular to the field of therapeutic nucleic acids. The present invention provides artificial nucleic acids, in particular RNAs, encoding CRISPR-associated proteins. A (pharmaceutical) composition and kit-of-parts comprising the same are also provided. Furthermore, the present invention relates to the artificial nucleic acid, (pharmaceutical) composition, or kit-of-parts for use in medicine, and in particular in the treatment and/or prophylaxis of diseases amenable to treatment with CRISPR-associated proteins.

## Claims

### Claim 1

An RNA molecule comprising:
(a) a 5′ Cap 1 structure;
(b) a 5′ untranslated region (5′ UTR) that is heterologous to a coding sequence;
(c) the coding sequence encoding a Cas9 protein that comprises at least one nuclear localization sequence (NLS) at the N-terminus and the C-terminus, wherein:
(i) the Cas9 protein comprises an amino acid sequence having at least 95% sequence identity to the amino acid sequence of SEQ ID NO: 1362; and
(ii) the coding sequence comprises a nucleic acid sequence having at least 80% sequence identity to the nucleic acid sequence of SEQ ID NO: 14518;
(d) a 3′ untranslated region (3′ UTR) that is heterologous to the coding sequence; and
(e) a poly(A) sequence located at the 3′ end of the mRNA and comprising about 60 to 250 adenosine nucleotides.

### Claim 2 (depends on 1)

The RNA molecule of claim 1, wherein the coding sequence comprises a nucleic acid sequence having at least 85% sequence identity to the nucleic acid sequence of SEQ ID NO: 14518.

### Claim 3 (depends on 1)

The RNA molecule of claim 1, wherein the Cas9 protein comprises an amino acid sequence having at least 99% sequence identity to the amino acid sequence of SEQ ID NO: 1362.

### Claim 4 (depends on 1)

The RNA molecule of claim 1, wherein the RNA molecule comprises an nucleic acid sequence having at least 85% sequence identity to the nucleic acid sequence of SEQ ID NOs 14524 or 14527.

### Claim 5 (depends on 1)

The RNA molecule of claim 1, wherein the G/C content of the coding sequence is increased compared to the G/C content of a corresponding wild-type or reference coding sequence.

### Claim 6 (depends on 1)

The RNA molecule of claim 1, wherein the 5′ UTR region comprises a nucleic acid sequence derived from a 5′UTR of a gene selected from the group consisting of SLC7A3, ATP5A1, RPL32, HSD17B4, NOSIP, ASAH1, RPL31, TUBB4B, UBQLN2, MP68 and NDUFA4.

### Claim 7 (depends on 1)

The RNA molecule of claim 1, wherein the 3′ UTR region comprises a nucleic acid sequence derived from a 3′ UTR of a gene selected from the group consisting of GNAS, CASP1, PSMB3, ALB, COX6B1, NDUFA1 and RPS9.

### Claim 8 (depends on 1)

The RNA molecule of claim 1, wherein the 5′ UTR region comprises a nucleic acid sequence derived from a 5′UTR of a HSD17B4 gene and wherein the 3′ UTR region comprises a nucleic acid sequence derived from a 3′ UTR of a GNAS gene.

### Claim 9 (depends on 1)

The RNA molecule of claim 1, wherein the poly(A) sequence comprises about 60 to 100 adenosine nucleotides.

### Claim 10 (depends on 1)

The RNA molecule of claim 1, wherein the Cas9 protein has the structure NLS2-Cas9-NLS4.

### Claim 11 (depends on 10)

The RNA molecule of claim 10, wherein the NLS2 has at least 95% sequence identity to the amino acid sequence of SEQ ID NO: 426, and NLS4 has at least 95% sequence identity to the amino acid sequence of SEQ ID NO: 427.

### Claim 12 (depends on 1)

A composition comprising the RNA molecule of claim 1, wherein the RNA molecule is formulated in a lipid nanoparticle.

### Claim 13 (depends on 12)

The composition of claim 12, wherein the lipid nanoparticles (LNPs) comprise (a) a cationic lipid, (b) an aggregation reducing lipid, (c) a neutral lipid, and (d) a sterol.

### Claim 14 (depends on 13)

The composition of claim 13, wherein the LNPs comprise the lipids in a molar ratio of about 20-60% cationic lipid; 5-25% neutral lipid; 25-55% sterol; and 0.5-15% aggregation reducing lipid.

### Claim 15 (depends on 14)

The composition of claim 14, wherein the LNPs have a median diameter size of from about 50 nm to about 200 nm.

### Claim 16 (depends on 12)

The composition of claim 12, further comprising at least one guide RNA (gRNA) or a nucleic acid encoding at least one gRNA.

### Claim 17

An mRNA having at least 85% sequence identity to the nucleic acid sequence of SEQ ID NO: 14524 or SEQ ID NO: 14527, wherein the mRNA molecule encodes a Cas9 that comprises at least one nuclear localization sequence (NLS) at the N-terminus and the C-terminus, wherein the Cas9 protein comprises an amino acid sequence having at least 95% sequence identity to the amino acid sequence of SEQ ID NO: 1362.

### Claim 18 (depends on 17)

A composition comprising the mRNA of claim 17, wherein the mRNA is formulated in lipid nanoparticles (LNPs).

### Claim 19 (depends on 18)

The composition of claim 18, wherein the LNPs comprise (a) a cationic lipid, (b) an aggregation reducing lipid, (c) a neutral lipid, and (d) a sterol.

### Claim 20 (depends on 19)

The composition of claim 19, wherein the LNPs comprise the lipids in a molar ratio of about 20-60% cationic lipid; 5-25% neutral lipid; 25-55% sterol; and 0.5-15% aggregation reducing lipid.

### Claim 21 (depends on 20)

The composition of claim 20, wherein the LNPs have a median diameter size of from about 50 nm to about 200 nm.

### Claim 22 (depends on 18)

The composition of claim 18, further comprising at least one guide RNA (gRNA) or a nucleic acid encoding at least one gRNA.

### Claim 23 (depends on 1)

A method of treating or preventing a disorder in a subject in need thereof, the method comprising administering to the subject an effective amount of:
(a) the RNA molecule of claim 1;
wherein the disorder is amenable to modification of a gene of interest by knock-in, knock-out, mutation, or modulation of gene expression.