WO2004074499A3 - Combinatorial dna library for producing modified n-glycans in lower eukaryotes - Google Patents

Combinatorial dna library for producing modified n-glycans in lower eukaryotes Download PDF

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Publication number
WO2004074499A3
WO2004074499A3 PCT/US2004/005244 US2004005244W WO2004074499A3 WO 2004074499 A3 WO2004074499 A3 WO 2004074499A3 US 2004005244 W US2004005244 W US 2004005244W WO 2004074499 A3 WO2004074499 A3 WO 2004074499A3
Authority
WO
WIPO (PCT)
Prior art keywords
modified
glycosylation
host cells
glycans
mannosidases
Prior art date
Application number
PCT/US2004/005244
Other languages
French (fr)
Other versions
WO2004074499A2 (en
Inventor
Tillman U Gerngross
Stefan Wildt
Byung-Kwon Choi
Juergen Hermann Nett
Piotr Bobrowicz
Stephen R Hamilton
Robert C Davidson
Original Assignee
Tillman U Gerngross
Stefan Wildt
Byung-Kwon Choi
Juergen Hermann Nett
Piotr Bobrowicz
Stephen R Hamilton
Robert C Davidson
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 Tillman U Gerngross, Stefan Wildt, Byung-Kwon Choi, Juergen Hermann Nett, Piotr Bobrowicz, Stephen R Hamilton, Robert C Davidson filed Critical Tillman U Gerngross
Priority to CA002516440A priority Critical patent/CA2516440A1/en
Priority to EP04713437A priority patent/EP1599596B1/en
Priority to AT04713437T priority patent/ATE469236T1/en
Priority to JP2006503788A priority patent/JP4875486B2/en
Priority to DE602004027351T priority patent/DE602004027351D1/en
Priority to AU2004213869A priority patent/AU2004213869B2/en
Publication of WO2004074499A2 publication Critical patent/WO2004074499A2/en
Publication of WO2004074499A3 publication Critical patent/WO2004074499A3/en
Priority to IL170347A priority patent/IL170347A/en
Priority to IL198906A priority patent/IL198906A0/en

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    • C—CHEMISTRY; METALLURGY
    • C12—BIOCHEMISTRY; BEER; SPIRITS; WINE; VINEGAR; MICROBIOLOGY; ENZYMOLOGY; MUTATION OR GENETIC ENGINEERING
    • C12P—FERMENTATION OR ENZYME-USING PROCESSES TO SYNTHESISE A DESIRED CHEMICAL COMPOUND OR COMPOSITION OR TO SEPARATE OPTICAL ISOMERS FROM A RACEMIC MIXTURE
    • C12P21/00—Preparation of peptides or proteins
    • C12P21/005—Glycopeptides, glycoproteins
    • C—CHEMISTRY; METALLURGY
    • C07—ORGANIC CHEMISTRY
    • C07K—PEPTIDES
    • C07K14/00—Peptides having more than 20 amino acids; Gastrins; Somatostatins; Melanotropins; Derivatives thereof
    • C07K14/435—Peptides having more than 20 amino acids; Gastrins; Somatostatins; Melanotropins; Derivatives thereof from animals; from humans
    • C07K14/46—Peptides having more than 20 amino acids; Gastrins; Somatostatins; Melanotropins; Derivatives thereof from animals; from humans from vertebrates
    • C07K14/47—Peptides having more than 20 amino acids; Gastrins; Somatostatins; Melanotropins; Derivatives thereof from animals; from humans from vertebrates from mammals
    • 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
    • C12N15/1082—Preparation or screening gene libraries by chromosomal integration of polynucleotide sequences, HR-, site-specific-recombination, transposons, viral vectors
    • 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
    • C12N15/63—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
    • 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
    • C12N9/10—Transferases (2.)
    • C12N9/1048—Glycosyltransferases (2.4)
    • C12N9/1051—Hexosyltransferases (2.4.1)

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  • Health & Medical Sciences (AREA)
  • Life Sciences & Earth Sciences (AREA)
  • Chemical & Material Sciences (AREA)
  • Genetics & Genomics (AREA)
  • Engineering & Computer Science (AREA)
  • Organic Chemistry (AREA)
  • Zoology (AREA)
  • Wood Science & Technology (AREA)
  • Bioinformatics & Cheminformatics (AREA)
  • Biotechnology (AREA)
  • General Engineering & Computer Science (AREA)
  • Biomedical Technology (AREA)
  • Molecular Biology (AREA)
  • Biochemistry (AREA)
  • General Health & Medical Sciences (AREA)
  • Microbiology (AREA)
  • Biophysics (AREA)
  • Plant Pathology (AREA)
  • Physics & Mathematics (AREA)
  • Medicinal Chemistry (AREA)
  • Bioinformatics & Computational Biology (AREA)
  • Crystallography & Structural Chemistry (AREA)
  • Virology (AREA)
  • Proteomics, Peptides & Aminoacids (AREA)
  • Gastroenterology & Hepatology (AREA)
  • Toxicology (AREA)
  • Chemical Kinetics & Catalysis (AREA)
  • General Chemical & Material Sciences (AREA)
  • Preparation Of Compounds By Using Micro-Organisms (AREA)
  • Micro-Organisms Or Cultivation Processes Thereof (AREA)
  • Peptides Or Proteins (AREA)
  • Enzymes And Modification Thereof (AREA)
  • Polysaccharides And Polysaccharide Derivatives (AREA)
  • Breeding Of Plants And Reproduction By Means Of Culturing (AREA)
  • Saccharide Compounds (AREA)

Abstract

The present invention relates to eukaryotic host cells having modified oligosaccharides which may be modified further by heterologous expression of a set of glycosyltransferases, sugar transporters and mannosidases to become host-strains for the production of mammalian, e.g., human therapeutic glycoproteins. The invention provides nucleic acid molecules and combinatorial libraries which can be used to successfully target and express mammalian enzymatic activities such as those involved in glycosylation to intracellular compartments in a eukaryotic host cell. The process provides an engineered host cell which can be used to express and target any desirable gene(s) involved in glycosylation. Host cells with modified oligosaccharides are created or selected. N-glycans made in the engineered host cells have a Man5GlcNAc2 core structure which may then be modified further by heterologous expression of one or more enzymes, e.g., glycosyltransferases, sugar transporters and mannosidases, to yield human-like glycoproteins. For the production of therapeutic proteins, this method may be adapted to engineer cell lines in which any desired glycosylation structure may be obtained.
PCT/US2004/005244 2003-02-20 2004-02-20 Combinatorial dna library for producing modified n-glycans in lower eukaryotes WO2004074499A2 (en)

Priority Applications (8)

Application Number Priority Date Filing Date Title
CA002516440A CA2516440A1 (en) 2003-02-20 2004-02-20 Combinatorial dna library for producing modified n-glycans in lower eukaryotes
EP04713437A EP1599596B1 (en) 2003-02-20 2004-02-20 Combinatorial dna library for producing modified n -glycans in lower eukaryotes
AT04713437T ATE469236T1 (en) 2003-02-20 2004-02-20 COMBINATORY DNA LIBRARY FOR PRODUCING MODIFIED N-GLUCANS IN LOWER EUKARYOTES
JP2006503788A JP4875486B2 (en) 2003-02-20 2004-02-20 Combinatorial DNA library for producing modified N-glycans in lower eukaryotes
DE602004027351T DE602004027351D1 (en) 2003-02-20 2004-02-20 COMBINATIVE DNA LIBRARY FOR THE PREPARATION OF MODIFIED N-GLUCANES IN LOWER EUKARYOTES
AU2004213869A AU2004213869B2 (en) 2003-02-20 2004-02-20 Combinatorial DNA library for producing modified N-glycans in lower eukaryotes
IL170347A IL170347A (en) 2003-02-20 2005-08-18 Methods for producing modified n-glycans in eukaryotes
IL198906A IL198906A0 (en) 2003-02-20 2009-05-24 Dna encoding a cellular targeting signal peptide and a glycosylation enzyme and uses thereof

Applications Claiming Priority (2)

Application Number Priority Date Filing Date Title
US10/371,877 US7449308B2 (en) 2000-06-28 2003-02-20 Combinatorial DNA library for producing modified N-glycans in lower eukaryotes
US10/371,877 2003-02-20

Publications (2)

Publication Number Publication Date
WO2004074499A2 WO2004074499A2 (en) 2004-09-02
WO2004074499A3 true WO2004074499A3 (en) 2005-01-27

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Family Applications (1)

Application Number Title Priority Date Filing Date
PCT/US2004/005244 WO2004074499A2 (en) 2003-02-20 2004-02-20 Combinatorial dna library for producing modified n-glycans in lower eukaryotes

Country Status (10)

Country Link
US (6) US7449308B2 (en)
EP (3) EP2196540B1 (en)
JP (2) JP4875486B2 (en)
AT (1) ATE469236T1 (en)
AU (2) AU2004213869B2 (en)
CA (2) CA2516440A1 (en)
DE (1) DE602004027351D1 (en)
ES (2) ES2415955T3 (en)
IL (2) IL170347A (en)
WO (1) WO2004074499A2 (en)

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US8877462B2 (en) 2000-06-28 2014-11-04 Glycofi, Inc. Combinatorial DNA library for producing modified N-glycans in lower eukaryotes
US8986949B2 (en) 2003-02-20 2015-03-24 Glycofi, Inc. Endomannosidases in the modification of glycoproteins in eukaryotes
US8999671B2 (en) 2003-02-20 2015-04-07 Glycofi, Inc. Production of sialylated N-glycans in lower eukaryotes
US9598682B2 (en) 2009-09-29 2017-03-21 Vib Vzw Hydrolysis of mannose-1-phospho-6-mannose linkage to phospho-6-mannose
CN107810271A (en) * 2014-11-20 2018-03-16 耶路撒冷希伯来大学伊萨姆研发公司 Composition and method for polypeptide of the production with the glycosylation pattern changed in plant cell

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