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WO2024095245A3 - Evolved adenine deaminases and rna-guided nuclease fusion proteins with internal insertion sites and methods of use - Google Patents

Evolved adenine deaminases and rna-guided nuclease fusion proteins with internal insertion sites and methods of use Download PDF

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Publication number
WO2024095245A3
WO2024095245A3 PCT/IB2023/061192 IB2023061192W WO2024095245A3 WO 2024095245 A3 WO2024095245 A3 WO 2024095245A3 IB 2023061192 W IB2023061192 W IB 2023061192W WO 2024095245 A3 WO2024095245 A3 WO 2024095245A3
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WIPO (PCT)
Prior art keywords
fusion proteins
methods
deaminases
rna
guided nuclease
Prior art date
Legal status (The legal status is an assumption and is not a legal conclusion. Google has not performed a legal analysis and makes no representation as to the accuracy of the status listed.)
Ceased
Application number
PCT/IB2023/061192
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French (fr)
Other versions
WO2024095245A9 (en
WO2024095245A2 (en
Inventor
Michael Coyle
Alexandra Briner CRAWLEY
Tedd D. Elich
Salem FAHAM
Drew KELSO
Adrian Pickar OLIVER
Lucas Ferreira RIBEIRO
Ryan RICKELS
Sarah COMPTON
Current Assignee (The listed assignees may be inaccurate. Google has not performed a legal analysis and makes no representation or warranty as to the accuracy of the list.)
Lifeedit Therapeutics Inc
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Lifeedit Therapeutics Inc
Priority date (The priority date is an assumption and is not a legal conclusion. Google has not performed a legal analysis and makes no representation as to the accuracy of the date listed.)
Filing date
Publication date
Application filed by Lifeedit Therapeutics Inc filed Critical Lifeedit Therapeutics Inc
Priority to IL320573A priority Critical patent/IL320573A/en
Priority to AU2023374241A priority patent/AU2023374241A1/en
Priority to JP2025525634A priority patent/JP2025537710A/en
Priority to KR1020257018452A priority patent/KR20250126155A/en
Priority to CN202380090553.8A priority patent/CN121127583A/en
Priority to EP23805171.8A priority patent/EP4612284A2/en
Publication of WO2024095245A2 publication Critical patent/WO2024095245A2/en
Publication of WO2024095245A9 publication Critical patent/WO2024095245A9/en
Publication of WO2024095245A3 publication Critical patent/WO2024095245A3/en
Anticipated expiration legal-status Critical
Ceased legal-status Critical Current

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    • C—CHEMISTRY; METALLURGY
    • C12—BIOCHEMISTRY; BEER; SPIRITS; WINE; VINEGAR; MICROBIOLOGY; ENZYMOLOGY; MUTATION OR GENETIC ENGINEERING
    • C12N—MICROORGANISMS OR ENZYMES; COMPOSITIONS THEREOF; PROPAGATING, PRESERVING, OR MAINTAINING MICROORGANISMS; MUTATION OR GENETIC ENGINEERING; CULTURE MEDIA
    • C12N9/00—Enzymes; Proenzymes; Compositions thereof; Processes for preparing, activating, inhibiting, separating or purifying enzymes
    • C12N9/14—Hydrolases (3)
    • C12N9/16—Hydrolases (3) acting on ester bonds (3.1)
    • C12N9/22—Ribonucleases [RNase]; Deoxyribonucleases [DNase]
    • C—CHEMISTRY; METALLURGY
    • C12—BIOCHEMISTRY; BEER; SPIRITS; WINE; VINEGAR; MICROBIOLOGY; ENZYMOLOGY; MUTATION OR GENETIC ENGINEERING
    • C12N—MICROORGANISMS OR ENZYMES; COMPOSITIONS THEREOF; PROPAGATING, PRESERVING, OR MAINTAINING MICROORGANISMS; MUTATION OR GENETIC ENGINEERING; CULTURE MEDIA
    • C12N9/00—Enzymes; Proenzymes; Compositions thereof; Processes for preparing, activating, inhibiting, separating or purifying enzymes
    • C12N9/14—Hydrolases (3)
    • C12N9/16—Hydrolases (3) acting on ester bonds (3.1)
    • C12N9/22—Ribonucleases [RNase]; Deoxyribonucleases [DNase]
    • C12N9/222—Clustered regularly interspaced short palindromic repeats [CRISPR]-associated [CAS] enzymes
    • C—CHEMISTRY; METALLURGY
    • C12—BIOCHEMISTRY; BEER; SPIRITS; WINE; VINEGAR; MICROBIOLOGY; ENZYMOLOGY; MUTATION OR GENETIC ENGINEERING
    • C12N—MICROORGANISMS OR ENZYMES; COMPOSITIONS THEREOF; PROPAGATING, PRESERVING, OR MAINTAINING MICROORGANISMS; MUTATION OR GENETIC ENGINEERING; CULTURE MEDIA
    • C12N15/00—Mutation or genetic engineering; DNA or RNA concerning genetic engineering, vectors, e.g. plasmids, or their isolation, preparation or purification; Use of hosts therefor
    • C12N15/09—Recombinant DNA-technology
    • C12N15/10—Processes for the isolation, preparation or purification of DNA or RNA
    • C12N15/102—Mutagenizing nucleic acids
    • C—CHEMISTRY; METALLURGY
    • C12—BIOCHEMISTRY; BEER; SPIRITS; WINE; VINEGAR; MICROBIOLOGY; ENZYMOLOGY; MUTATION OR GENETIC ENGINEERING
    • C12N—MICROORGANISMS OR ENZYMES; COMPOSITIONS THEREOF; PROPAGATING, PRESERVING, OR MAINTAINING MICROORGANISMS; MUTATION OR GENETIC ENGINEERING; CULTURE MEDIA
    • C12N15/00—Mutation or genetic engineering; DNA or RNA concerning genetic engineering, vectors, e.g. plasmids, or their isolation, preparation or purification; Use of hosts therefor
    • C12N15/09—Recombinant DNA-technology
    • C12N15/10—Processes for the isolation, preparation or purification of DNA or RNA
    • C12N15/1034—Isolating an individual clone by screening libraries
    • C12N15/1058—Directional evolution of libraries, e.g. evolution of libraries is achieved by mutagenesis and screening or selection of mixed population of organisms
    • C—CHEMISTRY; METALLURGY
    • C12—BIOCHEMISTRY; BEER; SPIRITS; WINE; VINEGAR; MICROBIOLOGY; ENZYMOLOGY; MUTATION OR GENETIC ENGINEERING
    • C12N—MICROORGANISMS OR ENZYMES; COMPOSITIONS THEREOF; PROPAGATING, PRESERVING, OR MAINTAINING MICROORGANISMS; MUTATION OR GENETIC ENGINEERING; CULTURE MEDIA
    • C12N15/00—Mutation or genetic engineering; DNA or RNA concerning genetic engineering, vectors, e.g. plasmids, or their isolation, preparation or purification; Use of hosts therefor
    • C12N15/09—Recombinant DNA-technology
    • C12N15/11—DNA or RNA fragments; Modified forms thereof; Non-coding nucleic acids having a biological activity
    • 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
    • C—CHEMISTRY; METALLURGY
    • C12—BIOCHEMISTRY; BEER; SPIRITS; WINE; VINEGAR; MICROBIOLOGY; ENZYMOLOGY; MUTATION OR GENETIC ENGINEERING
    • C12N—MICROORGANISMS OR ENZYMES; COMPOSITIONS THEREOF; PROPAGATING, PRESERVING, OR MAINTAINING MICROORGANISMS; MUTATION OR GENETIC ENGINEERING; CULTURE MEDIA
    • C12N9/00—Enzymes; Proenzymes; Compositions thereof; Processes for preparing, activating, inhibiting, separating or purifying enzymes
    • C12N9/14—Hydrolases (3)
    • C12N9/78—Hydrolases (3) acting on carbon to nitrogen bonds other than peptide bonds (3.5)
    • C—CHEMISTRY; METALLURGY
    • C12—BIOCHEMISTRY; BEER; SPIRITS; WINE; VINEGAR; MICROBIOLOGY; ENZYMOLOGY; MUTATION OR GENETIC ENGINEERING
    • C12Y—ENZYMES
    • C12Y305/00—Hydrolases acting on carbon-nitrogen bonds, other than peptide bonds (3.5)
    • C12Y305/04—Hydrolases acting on carbon-nitrogen bonds, other than peptide bonds (3.5) in cyclic amidines (3.5.4)
    • C12Y305/04002—Adenine deaminase (3.5.4.2)
    • C—CHEMISTRY; METALLURGY
    • C07—ORGANIC CHEMISTRY
    • C07K—PEPTIDES
    • C07K2319/00—Fusion polypeptide
    • C—CHEMISTRY; METALLURGY
    • C07—ORGANIC CHEMISTRY
    • C07K—PEPTIDES
    • C07K2319/00—Fusion polypeptide
    • C07K2319/85—Fusion polypeptide containing an RNA binding domain
    • C—CHEMISTRY; METALLURGY
    • C12—BIOCHEMISTRY; BEER; SPIRITS; WINE; VINEGAR; MICROBIOLOGY; ENZYMOLOGY; MUTATION OR GENETIC ENGINEERING
    • C12N—MICROORGANISMS OR ENZYMES; COMPOSITIONS THEREOF; PROPAGATING, PRESERVING, OR MAINTAINING MICROORGANISMS; MUTATION OR GENETIC ENGINEERING; CULTURE MEDIA
    • C12N2310/00—Structure or type of the nucleic acid
    • C12N2310/10—Type of nucleic acid
    • C12N2310/20—Type of nucleic acid involving clustered regularly interspaced short palindromic repeats [CRISPR]

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  • Life Sciences & Earth Sciences (AREA)
  • Health & Medical Sciences (AREA)
  • Chemical & Material Sciences (AREA)
  • Engineering & Computer Science (AREA)
  • Genetics & Genomics (AREA)
  • Organic Chemistry (AREA)
  • Bioinformatics & Cheminformatics (AREA)
  • Wood Science & Technology (AREA)
  • Zoology (AREA)
  • General Engineering & Computer Science (AREA)
  • Biomedical Technology (AREA)
  • Biotechnology (AREA)
  • Molecular Biology (AREA)
  • General Health & Medical Sciences (AREA)
  • Biochemistry (AREA)
  • Microbiology (AREA)
  • Physics & Mathematics (AREA)
  • Biophysics (AREA)
  • Plant Pathology (AREA)
  • Medicinal Chemistry (AREA)
  • Crystallography & Structural Chemistry (AREA)
  • Ecology (AREA)
  • Bioinformatics & Computational Biology (AREA)
  • Micro-Organisms Or Cultivation Processes Thereof (AREA)
  • Peptides Or Proteins (AREA)
  • Enzymes And Modification Thereof (AREA)
  • Medicines That Contain Protein Lipid Enzymes And Other Medicines (AREA)
  • Pharmaceuticals Containing Other Organic And Inorganic Compounds (AREA)
  • Medicines Containing Material From Animals Or Micro-Organisms (AREA)
  • Preparation Of Compounds By Using Micro-Organisms (AREA)

Abstract

Compositions and methods comprising deaminases for targeted editing of nucleic acids are provided. Also provided are compositions and methods for localizing a heterologous polypeptide to a target DNA molecule and for targeted editing of nucleic acids are provided. Fusion proteins comprising an RNA-guided nuclease (RGN) and at least one heterologous polypeptide inserted therein and fusion proteins comprising a DNA-binding polypeptide and a deaminase are provided. The heterologous polypeptide can be a prime editing polypeptide or a base-editing polypeptide. Compositions also include nucleic acid molecules encoding the deaminases or the fusion proteins. Vectors and host cells comprising the nucleic acid molecules encoding the deaminases or fusion proteins are also provided.
PCT/IB2023/061192 2022-11-04 2023-11-06 Evolved adenine deaminases and rna-guided nuclease fusion proteins with internal insertion sites and methods of use Ceased WO2024095245A2 (en)

Priority Applications (6)

Application Number Priority Date Filing Date Title
IL320573A IL320573A (en) 2022-11-04 2023-11-06 Evolved adenine deaminases and rna-guided nuclease fusion proteins with internal insertion sites and methods of use
AU2023374241A AU2023374241A1 (en) 2022-11-04 2023-11-06 Evolved adenine deaminases and rna-guided nuclease fusion proteins with internal insertion sites and methods of use
JP2025525634A JP2025537710A (en) 2022-11-04 2023-11-06 Evolved adenine deaminase and RNA-guided nuclease fusion proteins with internal insertion sites and methods of use
KR1020257018452A KR20250126155A (en) 2022-11-04 2023-11-06 Evolved adenine deaminase and RNA-guided nuclease fusion proteins having internal insertion sites and methods of use
CN202380090553.8A CN121127583A (en) 2022-11-04 2023-11-06 Evolved adenine deaminase and RNA-guided nuclease fusion proteins with internal insertion sites and methods of use
EP23805171.8A EP4612284A2 (en) 2022-11-04 2023-11-06 Evolved adenine deaminases and rna-guided nuclease fusion proteins with internal insertion sites and methods of use

Applications Claiming Priority (4)

Application Number Priority Date Filing Date Title
US202263382344P 2022-11-04 2022-11-04
US63/382,344 2022-11-04
US202363485642P 2023-02-17 2023-02-17
US63/485,642 2023-02-17

Publications (3)

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WO2024095245A2 WO2024095245A2 (en) 2024-05-10
WO2024095245A9 WO2024095245A9 (en) 2024-07-18
WO2024095245A3 true WO2024095245A3 (en) 2024-11-14

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EP (1) EP4612284A2 (en)
JP (1) JP2025537710A (en)
KR (1) KR20250126155A (en)
CN (1) CN121127583A (en)
AU (1) AU2023374241A1 (en)
IL (1) IL320573A (en)
TW (1) TW202434726A (en)
WO (1) WO2024095245A2 (en)

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* Cited by examiner, † Cited by third party
Publication number Priority date Publication date Assignee Title
WO2025022367A2 (en) 2023-07-27 2025-01-30 Life Edit Therapeutics, Inc. Rna-guided nucleases and active fragments and variants thereof and methods of use
WO2025083619A1 (en) 2023-10-18 2025-04-24 Life Edit Therapeutics, Inc. Rna-guided nucleases and acive fragments and variants thereof and methods of use

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AU2023374241A1 (en) 2025-06-12
CN121127583A (en) 2025-12-12
WO2024095245A9 (en) 2024-07-18
JP2025537710A (en) 2025-11-20
WO2024095245A2 (en) 2024-05-10
TW202434726A (en) 2024-09-01
KR20250126155A (en) 2025-08-22
EP4612284A2 (en) 2025-09-10
IL320573A (en) 2025-07-01

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