DK3218490T3 - Dna-bindende domæne af crispr-system til produktion af ikke-fucosylerede og delvist fucosylerede proteiner - Google Patents

Dna-bindende domæne af crispr-system til produktion af ikke-fucosylerede og delvist fucosylerede proteiner Download PDF

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DK3218490T3
DK3218490T3 DK15816521.7T DK15816521T DK3218490T3 DK 3218490 T3 DK3218490 T3 DK 3218490T3 DK 15816521 T DK15816521 T DK 15816521T DK 3218490 T3 DK3218490 T3 DK 3218490T3
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crispr
dna
cell
lys
gmd
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DK15816521.7T
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Bhargav Prasad
Divya Unnikrishnan
Jahnabi Hazarika
Kavitha Iyer Rodrigues
Maloy Ghosh
Pavithra M
Pravin Kumar D
Sanghamitra Bhattacharjee
Sathyabalan M
Sankaranarayanan Srinivasan
Sohang Chatterjee
Sunit Maity
Veeresha K
Vivek Halan
M Yogendra Manjunath B
Anuradha Hora
Bairavabalakumar N
Karthika Nair
Aswini Thanigaivel
Amol Maliwalave
Bharath R Shenoy
Anisha Kurup
Sahana Bhima Rao
Rajeshwari Pendse
Prabhat Kumar Pathak
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Zumutor Biologics Inc
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    • C—CHEMISTRY; METALLURGY
    • C07—ORGANIC CHEMISTRY
    • C07K—PEPTIDES
    • C07K16/00—Immunoglobulins [IGs], e.g. monoclonal or polyclonal antibodies
    • C07K16/06—Immunoglobulins [IGs], e.g. monoclonal or polyclonal antibodies from serum
    • C07K16/065—Purification, fragmentation
    • C—CHEMISTRY; METALLURGY
    • C12—BIOCHEMISTRY; BEER; SPIRITS; WINE; VINEGAR; MICROBIOLOGY; ENZYMOLOGY; MUTATION OR GENETIC ENGINEERING
    • C12Y—ENZYMES
    • C12Y402/00—Carbon-oxygen lyases (4.2)
    • C12Y402/01—Hydro-lyases (4.2.1)
    • C12Y402/01047—GDP-mannose 4,6-dehydratase (4.2.1.47), i.e. GMD
    • A—HUMAN NECESSITIES
    • A61—MEDICAL OR VETERINARY SCIENCE; HYGIENE
    • A61K—PREPARATIONS FOR MEDICAL, DENTAL OR TOILETRY PURPOSES
    • A61K48/00—Medicinal preparations containing genetic material which is inserted into cells of the living body to treat genetic diseases; Gene therapy
    • A61K48/0008—Medicinal preparations containing genetic material which is inserted into cells of the living body to treat genetic diseases; Gene therapy characterised by an aspect of the 'non-active' part of the composition delivered, e.g. wherein such 'non-active' part is not delivered simultaneously with the 'active' part of the composition
    • A61K48/0016—Medicinal preparations containing genetic material which is inserted into cells of the living body to treat genetic diseases; Gene therapy characterised by an aspect of the 'non-active' part of the composition delivered, e.g. wherein such 'non-active' part is not delivered simultaneously with the 'active' part of the composition wherein the nucleic acid is delivered as a 'naked' nucleic acid, i.e. not combined with an entity such as a cationic lipid
    • C—CHEMISTRY; METALLURGY
    • C07—ORGANIC CHEMISTRY
    • C07K—PEPTIDES
    • C07K16/00—Immunoglobulins [IGs], e.g. monoclonal or polyclonal antibodies
    • 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
    • C12N15/11—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
    • C12N15/1137—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 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
    • C12N5/00—Undifferentiated human, animal or plant cells, e.g. cell lines; Tissues; Cultivation or maintenance thereof; Culture media therefor
    • C12N5/06—Animal cells or tissues; Human cells or tissues
    • C12N5/0601—Invertebrate cells or tissues, e.g. insect cells; Culture media therefor
    • 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
    • C12N5/00—Undifferentiated human, animal or plant cells, e.g. cell lines; Tissues; Cultivation or maintenance thereof; Culture media therefor
    • C12N5/10—Cells modified by introduction of foreign genetic material
    • C12N5/12—Fused cells, e.g. hybridomas
    • C12N5/16—Animal cells
    • C12N5/163—Animal cells one of the fusion partners being a B or a T lymphocyte
    • C—CHEMISTRY; METALLURGY
    • C12—BIOCHEMISTRY; BEER; SPIRITS; WINE; VINEGAR; MICROBIOLOGY; ENZYMOLOGY; MUTATION OR GENETIC ENGINEERING
    • C12Y—ENZYMES
    • C12Y204/00—Glycosyltransferases (2.4)
    • C12Y204/01—Hexosyltransferases (2.4.1)
    • C12Y204/01068—Glycoprotein 6-alpha-L-fucosyltransferase (2.4.1.68), i.e. FUT8
    • C—CHEMISTRY; METALLURGY
    • C07—ORGANIC CHEMISTRY
    • C07K—PEPTIDES
    • C07K2317/00—Immunoglobulins specific features
    • C07K2317/40—Immunoglobulins specific features characterized by post-translational modification
    • C07K2317/41—Glycosylation, sialylation, or fucosylation
    • 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 [CRISPRs]

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  • Genetics & Genomics (AREA)
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  • Plant Pathology (AREA)
  • Proteomics, Peptides & Aminoacids (AREA)
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  • Virology (AREA)
  • Crystallography & Structural Chemistry (AREA)
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  • Veterinary Medicine (AREA)
  • Pharmacology & Pharmacy (AREA)
  • Public Health (AREA)
  • Animal Behavior & Ethology (AREA)
  • Micro-Organisms Or Cultivation Processes Thereof (AREA)
  • Enzymes And Modification Thereof (AREA)
  • Peptides Or Proteins (AREA)
  • Preparation Of Compounds By Using Micro-Organisms (AREA)
  • Measuring Or Testing Involving Enzymes Or Micro-Organisms (AREA)
  • Medicines That Contain Protein Lipid Enzymes And Other Medicines (AREA)

Claims (13)

1. DNA-bindende domæne af CRISPR-system, hvor det DNA-bindende domæne omfatter RNA-sekvens transskriberet fra nukleotidsekvens valgt fra gruppen bestående af SEQ ID NO: 41, SEQ ID NO: 43 og SEQ ID NO: 45.
2. DNA-bindende domæne ifølge krav 1, hvor RNA-sekvensen, der transskriberes fra SEQ ID NO: 41, SEQ ID NO: 43 og SEQ ID NO: 45, binder til GMD-gensekvens; og hvor SEQ ID NO: 41 transskriberes til SEQ ID NO: 42; SEQ ID NO: 43 transskriberes til SEQ ID NO: 44, og SEQ ID NO: 45 transskriberes til SEQ ID NO: 46.
3. CRISPR-nukleasekompleks, som omfatter det DNA-bindende domæne ifølge krav 1 og nuklease.
4. Kompleks ifølge krav 3, hvor nukleasen er Cas9-endonuklease eller Cas9n-endonuklease.
5. Vektor, som omfatter nukleotidsekvensen, der koder for DNA-bindende domæne ifølge krav 1.
6. Vektor ifølge krav 5, som endvidere omfatter nukleotidsekvens, der koder for nuklease.
7. Celle, der omfatter en vektor ifølge krav 5.
8. Celle ifølge krav 7, hvor cellen er valgt fra gruppen bestående af COS, CHO-S, CHO-K1, CHO-K1 GS (-/-), CHO-DG44, CHO -DUXB11, CHO-DUKX, CHOK1SV, VERO, MDCK, W138, V79, B14AF28-G3, BHK, HaK, NSO, SP2/0-Agl4, HeLa, HEK293-F, HEK293-H, HEK293 -T, YB23HL.P2.Gl1.16Ag.20, perC6, antistofproducerende hybridomacelle, embryonial stamcelle, Namalwa-celle, insektcellelinje fra Spodoptera fugiperda (Sf), Pichia, Saccharomyces og Schizosaccharomyces.
9. In vi tro-fremgangsmåde til frembringelse af en fucose-knockout-celle, hvilken fremgangsmåde omfatter trin, der går ud på: a) at frembringe en CRISPR-nukleasekonstruktion, som omfatter nukleotidsekvens, der koder for det DNA-bindende domæne ifølge krav 1; og b) at transfektere en celle med konstruktionen fra trin (a) for at frembringe en fucose-knockout-celle, hvor CRISPR-nukleasekonstruktionen tilvejebringer CRISPR-nukleasekompleks, der omfatter det DNA-bindende domæne og nuklease; og hvor komplekset spalter GMD-gensekvens i cellen.
10. In vi tro-fremgangsmåde til frembringelse af protein med fucosylering i området fra 0 % til 100 %, hvilken fremgangsmåde omfatter trin, der går ud på: a) at dyrke cellen, der er frembragt ved fremgangsmåden ifølge krav 9, hvor cellen har en fucosyleringsaktivitet i området fra 0 % til 100 %; og b) at frembringe proteinet, der udtrykkes af cellen fra trin (a).
11. Fremgangsmåde ifølge krav 9, hvor GMD-gensekvensens, der koder for a-GDP-D-mannose-4,6-dehydratase-enzym, spaltes i exon valgt fra gruppen bestående af Exon 3, Exon 4 og kombination deraf; og hvor cellen er valgt fra gruppen bestående af COS, CHO-S, CHO-K1, CHO-K1 GS (-/-), CHO-DG44, CHO -DUXB11, CHO-DUKX, CHOK1SV, VERO, MDCK, W138, V79, B14AF28-G3, BHK, HaK, NSO, SP2/0-Agl4, HeLa, HEK293-F, HEK293- H, HEK293 -T, YB23HL.P2.GI1.16Ag.20, perC6, antistofproducerende hybridomacelle, embryonial stamcelle, Namalwa-celle, insektcellelinje fra Spodoptera fugiperda (Sf) , Pichia, Saccharomyces og Schizosaccharomyces.
12. Fremgangsmåde ifølge krav 10, hvor fremgangsmåden endvidere omfatter tilsætning af L-fucose til vækstmediet.
13. Fremgangsmåde ifølge krav 10, hvor proteinet er et antistof; hvor antistoffet er et monoklonalt antistof; eller hvor cellen producerer et endogent protein, der kodes for at et gen, som er indsat i cellen.
DK15816521.7T 2014-11-15 2015-11-13 Dna-bindende domæne af crispr-system til produktion af ikke-fucosylerede og delvist fucosylerede proteiner DK3218490T3 (da)

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IN5767CH2014 2014-11-15
PCT/IB2015/058777 WO2016075662A2 (en) 2014-11-15 2015-11-13 Dna-binding domain, non-fucosylated and partially fucosylated proteins, and methods thereof

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US (2) US10752674B2 (da)
EP (2) EP3467110A1 (da)
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DK (1) DK3218490T3 (da)
ES (1) ES2712303T3 (da)
WO (1) WO2016075662A2 (da)

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JP7090421B2 (ja) 2022-06-24
EP3218490A2 (en) 2017-09-20
EP3218490B1 (en) 2018-10-31
ES2712303T3 (es) 2019-05-10
WO2016075662A2 (en) 2016-05-19
US20200339666A1 (en) 2020-10-29
WO2016075662A3 (en) 2016-07-07
US20190112358A1 (en) 2019-04-18
US11548937B2 (en) 2023-01-10
JP2018502595A (ja) 2018-02-01
JP7134207B2 (ja) 2022-09-09
EP3467110A1 (en) 2019-04-10
JP2021007397A (ja) 2021-01-28

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