Nucleic acids encoding CRISPR-associated proteins and uses thereof

Patent metadata

Field	Value
Publication number	US-12371699-B2
Application number	US-202318346686-A
Country	US
Kind code	B2
Filing date	Jul 3, 2023
Priority date	Mar 24, 2017
Publication date	Jul 29, 2025
Grant date	Jul 29, 2025

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.

First claim

The invention claimed is:

1. An artificial nucleic acid molecule comprising
a. at least one coding region encoding at least one CRISPR-associated protein, wherein the at least one coding region of said artificial nucleic acid molecule comprises or consists of a nucleic acid sequence at least 90% identical to the RNA sequence of SEQ ID NO: 14518, said sequence encoding a polypeptide that is at least 90% identical to SEQ ID NO: 1362;
b. at least one 5′ untranslated region (5′ UTR) element, which is heterologous relative to the at least one coding region; and
c. at least one 3′ untranslated region (3′ UTR) element, which is heterologous relative to the at least one coding region, wherein said artificial nucleic acid molecule is an RNA, which comprises a 5′ Cap and a Poly(A) sequence.

2. The artificial nucleic acid molecule of claim 1 , wherein the at least one 5′ UTR element is derived from a 5′UTR of a HSD17B4 gene; or derived from a 5′UTR of a NDUFA4 gene.

3. The artificial nucleic acid molecule of claim 1 , wherein said CRISPR-associated protein comprises at least one further effector domain, selected from KRAB, CSD, WRPW, VP64, p65AD and Mxi.

4. The artificial nucleic acid molecule of claim 1 , wherein said artificial nucleic acid further comprises at least one nucleic acid sequence encoding a nuclear localization signal (NLS).

5. The artificial nucleic acid molecule of claim 1 , wherein the RNA is an mRNA.

6. The artificial nucleic acid molecule of claim 1 , which comprises at least one histone stem-loop.

7. The artificial nucleic acid molecule of claim 1 , wherein the poly(A) sequence comprises 10 to 200 adenosine nucleotides.

8. The artificial nucleic acid molecule of claim 1 , which comprises, in 5′ to 3′ direction, the following elements:
a) a 5′-CAP structure,
b) the 5′-UTR element,
c) the at least one coding sequence,
d) the 3′-UTR element,
e) a poly(A) tail,
f) optionally a poly(C) tail, and
g) optionally a histone stem-loop (HSL).

9. A composition comprising the artificial nucleic acid molecule of claim 1 and a pharmaceutically acceptable carrier and/or excipient.

10. The composition according to claim 9 , wherein the artificial nucleic acid molecule is complexed with one or more cationic or polycationic lipids.

11. A kit comprising the artificial nucleic acid molecule of claim 1 , and optionally a liquid vehicle and/or optionally technical instructions with information on the administration and dosage of the artificial nucleic acid molecule or the composition.

12. The artificial nucleic acid molecule of claim 1 , wherein the at least one 5′ UTR element is derived from a 5′UTR of a HSD17B4 gene.

13. The artificial nucleic acid molecule of claim 1 , wherein the at least one 5′ UTR element comprises a nucleic acid sequence at least 90% identical to the RNA sequence of SEQ ID NO: 2.

14. The artificial nucleic acid molecule of claim 1 , wherein the Poly(A) sequence comprises 10 to 100 adenosine nucleotides and is positioned at the 3′ end of the RNA.

15. The artificial nucleic acid molecule of claim 13 , wherein the at least one 3′ UTR element is derived from a 3′ UTR of a GNAS, CASP1, PSMB3, ALB, COX6B1, NDUFA1, or RPS9 gene.

16. The artificial nucleic acid molecule of claim 15 , wherein the at least one 3′ UTR element is derived from a Y UTR of a GNAS gene.

Assignees

• CureVac SE

Inventors

• Chevessier-Tünnesen Frédéric
• Poenisch Marion
• Schlake Thomas

Classifications

• C12N2310/531

  Stem-loop; Hairpin · CPC title

• C12N15/113

  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)} · CPC title

• C12N9/22

  Ribonucleases {[RNase]; Deoxyribonucleases [DNase]} · CPC title

• A61K48/0066

  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 · CPC title

• A61K31/7088

  Compounds having three or more nucleosides or nucleotides · CPC title