WO2023173062A3 - Nucleic acid editing systems, methods, and uses thereof - Google Patents

Nucleic acid editing systems, methods, and uses thereof Download PDF

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Publication number
WO2023173062A3
WO2023173062A3 PCT/US2023/064097 US2023064097W WO2023173062A3 WO 2023173062 A3 WO2023173062 A3 WO 2023173062A3 US 2023064097 W US2023064097 W US 2023064097W WO 2023173062 A3 WO2023173062 A3 WO 2023173062A3
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WIPO (PCT)
Prior art keywords
methods
nucleic acid
editing systems
acid editing
nuclease
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PCT/US2023/064097
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French (fr)
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WO2023173062A2 (en
Inventor
Tilmann BUERCKSTUEMMER
Henrik Schmidt
Anna SIEBER
Christoph ILSINGER
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Intima Bioscience, Inc.
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Publication of WO2023173062A2 publication Critical patent/WO2023173062A2/en
Publication of WO2023173062A3 publication Critical patent/WO2023173062A3/en

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    • C—CHEMISTRY; METALLURGY
    • C12—BIOCHEMISTRY; BEER; SPIRITS; WINE; VINEGAR; MICROBIOLOGY; ENZYMOLOGY; MUTATION OR GENETIC ENGINEERING
    • C12Q—MEASURING OR TESTING PROCESSES INVOLVING ENZYMES, NUCLEIC ACIDS OR MICROORGANISMS; COMPOSITIONS OR TEST PAPERS THEREFOR; PROCESSES OF PREPARING SUCH COMPOSITIONS; CONDITION-RESPONSIVE CONTROL IN MICROBIOLOGICAL OR ENZYMOLOGICAL PROCESSES
    • C12Q1/00—Measuring or testing processes involving enzymes, nucleic acids or microorganisms; Compositions therefor; Processes of preparing such compositions
    • C12Q1/68—Measuring or testing processes involving enzymes, nucleic acids or microorganisms; Compositions therefor; Processes of preparing such compositions involving nucleic acids
    • C12Q1/6806—Preparing nucleic acids for analysis, e.g. for polymerase chain reaction [PCR] assay
    • C—CHEMISTRY; METALLURGY
    • C12—BIOCHEMISTRY; BEER; SPIRITS; WINE; VINEGAR; MICROBIOLOGY; ENZYMOLOGY; MUTATION OR GENETIC ENGINEERING
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    • 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
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    • 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
    • C12N15/1138—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 against receptors or cell surface proteins
    • C—CHEMISTRY; METALLURGY
    • C12—BIOCHEMISTRY; BEER; SPIRITS; WINE; VINEGAR; MICROBIOLOGY; ENZYMOLOGY; MUTATION OR GENETIC ENGINEERING
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    • 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
    • C12N15/62—DNA sequences coding for fusion proteins
    • C—CHEMISTRY; METALLURGY
    • C12—BIOCHEMISTRY; BEER; SPIRITS; WINE; VINEGAR; MICROBIOLOGY; ENZYMOLOGY; MUTATION OR GENETIC ENGINEERING
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    • 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
    • C12N15/63—Introduction of foreign genetic material using vectors; Vectors; Use of hosts therefor; Regulation of expression
    • C12N15/79—Vectors or expression systems specially adapted for eukaryotic hosts
    • C12N15/85—Vectors or expression systems specially adapted for eukaryotic hosts for animal cells
    • C12N15/86—Viral vectors
    • C—CHEMISTRY; METALLURGY
    • C12—BIOCHEMISTRY; BEER; SPIRITS; WINE; VINEGAR; MICROBIOLOGY; ENZYMOLOGY; MUTATION OR GENETIC ENGINEERING
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    • 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
    • C12N15/87—Introduction of foreign genetic material using processes not otherwise provided for, e.g. co-transformation
    • C12N15/90—Stable introduction of foreign DNA into chromosome
    • C12N15/902—Stable introduction of foreign DNA into chromosome using homologous recombination
    • C12N15/907—Stable introduction of foreign DNA into chromosome using homologous recombination in mammalian cells
    • C—CHEMISTRY; METALLURGY
    • C12—BIOCHEMISTRY; BEER; SPIRITS; WINE; VINEGAR; MICROBIOLOGY; ENZYMOLOGY; MUTATION OR GENETIC ENGINEERING
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    • 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 RNAses, DNAses
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    • C12—BIOCHEMISTRY; BEER; SPIRITS; WINE; VINEGAR; MICROBIOLOGY; ENZYMOLOGY; MUTATION OR GENETIC ENGINEERING
    • C12Q—MEASURING OR TESTING PROCESSES INVOLVING ENZYMES, NUCLEIC ACIDS OR MICROORGANISMS; COMPOSITIONS OR TEST PAPERS THEREFOR; PROCESSES OF PREPARING SUCH COMPOSITIONS; CONDITION-RESPONSIVE CONTROL IN MICROBIOLOGICAL OR ENZYMOLOGICAL PROCESSES
    • C12Q1/00—Measuring or testing processes involving enzymes, nucleic acids or microorganisms; Compositions therefor; Processes of preparing such compositions
    • C12Q1/68—Measuring or testing processes involving enzymes, nucleic acids or microorganisms; Compositions therefor; Processes of preparing such compositions involving nucleic acids
    • C12Q1/6813—Hybridisation assays
    • C12Q1/6827—Hybridisation assays for detection of mutation or polymorphism
    • C12Q1/683—Hybridisation assays for detection of mutation or polymorphism involving restriction enzymes, e.g. restriction fragment length polymorphism [RFLP]
    • C—CHEMISTRY; METALLURGY
    • C07—ORGANIC CHEMISTRY
    • C07K—PEPTIDES
    • C07K2319/00—Fusion polypeptide
    • C07K2319/01—Fusion polypeptide containing a localisation/targetting motif
    • C07K2319/09—Fusion polypeptide containing a localisation/targetting motif containing a nuclear localisation signal
    • C—CHEMISTRY; METALLURGY
    • C12—BIOCHEMISTRY; BEER; SPIRITS; WINE; VINEGAR; MICROBIOLOGY; ENZYMOLOGY; MUTATION OR GENETIC ENGINEERING
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    • 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 [CRISPRs]
    • C—CHEMISTRY; METALLURGY
    • C12—BIOCHEMISTRY; BEER; SPIRITS; WINE; VINEGAR; MICROBIOLOGY; ENZYMOLOGY; MUTATION OR GENETIC ENGINEERING
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    • C12N2740/00—Reverse transcribing RNA viruses
    • C12N2740/00011—Details
    • C12N2740/10011—Retroviridae
    • C12N2740/16011—Human Immunodeficiency Virus, HIV
    • C12N2740/16041—Use of virus, viral particle or viral elements as a vector
    • C12N2740/16043—Use of virus, viral particle or viral elements as a vector viral genome or elements thereof as genetic vector

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  • Life Sciences & Earth Sciences (AREA)
  • Health & Medical Sciences (AREA)
  • Genetics & Genomics (AREA)
  • Chemical & Material Sciences (AREA)
  • Engineering & Computer Science (AREA)
  • Organic Chemistry (AREA)
  • Wood Science & Technology (AREA)
  • Zoology (AREA)
  • Bioinformatics & Cheminformatics (AREA)
  • Biomedical Technology (AREA)
  • Biotechnology (AREA)
  • General Engineering & Computer Science (AREA)
  • Molecular Biology (AREA)
  • Microbiology (AREA)
  • Biochemistry (AREA)
  • General Health & Medical Sciences (AREA)
  • Physics & Mathematics (AREA)
  • Biophysics (AREA)
  • Plant Pathology (AREA)
  • Proteomics, Peptides & Aminoacids (AREA)
  • Analytical Chemistry (AREA)
  • Immunology (AREA)
  • Virology (AREA)
  • Chemical Kinetics & Catalysis (AREA)
  • Crystallography & Structural Chemistry (AREA)
  • Medicinal Chemistry (AREA)
  • Cell Biology (AREA)
  • Mycology (AREA)
  • Enzymes And Modification Thereof (AREA)
  • Measuring Or Testing Involving Enzymes Or Micro-Organisms (AREA)

Abstract

Nucleic acid editing systems, methods, and uses thereof are provided. Various nuclease amino acid sequences and guide RNA sequences are described that promote cleavage of a target nucleic acid sequence. Methods of treating a disease and methods of detecting an alternation in a gene are described. Scaffolds and fusion proteins comprising nuclease mutants and fragments thereof are also provided.
PCT/US2023/064097 2022-03-11 2023-03-10 Nucleic acid editing systems, methods, and uses thereof WO2023173062A2 (en)

Applications Claiming Priority (4)

Application Number Priority Date Filing Date Title
US202263318906P 2022-03-11 2022-03-11
US63/318,906 2022-03-11
US202363479274P 2023-01-10 2023-01-10
US63/479,274 2023-01-10

Publications (2)

Publication Number Publication Date
WO2023173062A2 WO2023173062A2 (en) 2023-09-14
WO2023173062A3 true WO2023173062A3 (en) 2023-11-23

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PCT/US2023/064097 WO2023173062A2 (en) 2022-03-11 2023-03-10 Nucleic acid editing systems, methods, and uses thereof

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Citations (5)

* Cited by examiner, † Cited by third party
Publication number Priority date Publication date Assignee Title
US20180237770A1 (en) * 2013-03-14 2018-08-23 Caribou Biosciences, Inc. Compositions and Methods of Nucleic Acid-Targeting Nucleic Acids
US20180245099A1 (en) * 2015-08-07 2018-08-30 Caribou Biosciences, Inc. Compositions and methods of engineered crispr-cas9 systems using split-nexus cas9-associated polynucleotides
US20200263190A1 (en) * 2016-04-19 2020-08-20 The Broad Institute, Inc. Novel crispr enzymes and systems
US11098325B2 (en) * 2017-06-30 2021-08-24 Intima Bioscience, Inc. Adeno-associated viral vectors for gene therapy
US20210284979A1 (en) * 2017-07-18 2021-09-16 Howard Hughes Medical Institute Methods and compositions for genetically manipulating genes and cells

Patent Citations (5)

* Cited by examiner, † Cited by third party
Publication number Priority date Publication date Assignee Title
US20180237770A1 (en) * 2013-03-14 2018-08-23 Caribou Biosciences, Inc. Compositions and Methods of Nucleic Acid-Targeting Nucleic Acids
US20180245099A1 (en) * 2015-08-07 2018-08-30 Caribou Biosciences, Inc. Compositions and methods of engineered crispr-cas9 systems using split-nexus cas9-associated polynucleotides
US20200263190A1 (en) * 2016-04-19 2020-08-20 The Broad Institute, Inc. Novel crispr enzymes and systems
US11098325B2 (en) * 2017-06-30 2021-08-24 Intima Bioscience, Inc. Adeno-associated viral vectors for gene therapy
US20210284979A1 (en) * 2017-07-18 2021-09-16 Howard Hughes Medical Institute Methods and compositions for genetically manipulating genes and cells

Non-Patent Citations (5)

* Cited by examiner, † Cited by third party
Title
BĄBAŁA NIKOLINA, BOVENS ASTRID, DE VRIES EVERT, IGLESIAS-GUIMARAIS VICTORIA, AHRENDS TOMASZ, KRUMMEL MATTHEW F., BORST JANNIE, BIN: "Subcellular Localization of Antigen in Keratinocytes Dictates Delivery of CD4+ T-cell Help for the CTL Response upon Therapeutic DNA Vaccination into the Skin", CANCER IMMUNOLOGY RESEARCH, vol. 6, no. 7, 1 July 2018 (2018-07-01), US , pages 835 - 847, XP093113555, ISSN: 2326-6066, DOI: 10.1158/2326-6066.CIR-17-0408 *
DATABASE Nucleotide 31 May 2016 (2016-05-31), ANONYMOUS: "PREDICTED: Anolis carolinensis EF-hand calcium-binding domain-containing protein 2-like(LOC100563576), mRNA", XP093113556, retrieved from Genbank Database accession no. XM_003230113 *
DATABASE Protein 13 October 2019 (2019-10-13), ANONYMOUS: "type V CRISPR-associated protein Cas12a/Cpf1 [Xylanibacter ruminicola]", XP093113549, retrieved from Genbank Database accession no. WP_073043853 *
DATABASE Protein 22 April 2021 (2021-04-22), ANONYMOUS: "MAG: type V CRISPR-associated protein Cas12a/Cpf1 [Prevotella sp.", XP093113564, retrieved from Genbank Database accession no. MBR3078291 *
S.-T. CHOU, Q. LENG, J. MIXSON: "Zinc finger nucleases: Tailor-made for gene therapy", DRUGS OF THE FUTURE, vol. 37, no. 3, 1 January 2012 (2012-01-01), ES , pages 183 - 196, XP055512500, ISSN: 0377-8282, DOI: 10.1358/dof.2012.037.03.1779022 *

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