Oliviero, Claudia ; Hinz, Steffen C. ; Bogen, Jan P. ; Kornmann, Henri ; Hock, Björn ; Kolmar, Harald ; Hagens, Gerrit (2024)
Generation of a host cell line containing a MAR-rich landing pad for site-specific integration and expression of transgenes.
In: Biotechnology Progress, 2022, 38 (4)
doi: 10.26083/tuprints-00027669
Artikel, Zweitveröffentlichung, Verlagsversion
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Kurzbeschreibung (Abstract)
In recent years, targeted gene integration (TI) has been introduced as a strategy for the generation of recombinant mammalian cell lines for the production of biotherapeutics. Besides reducing the immense heterogeneity within a pool of recombinant transfectants, TI also aims at shortening the duration of the current cell line development process. Here we describe the generation of a host cell line carrying Matrix-Attachment Region (MAR)-rich landing pads (LPs), which allow for the simultaneous and site-specific integration of multiple genes of interest (GOIs). We show that several copies of each chicken lysozyme 5'MAR-based LP containing either BxB1 wild type or mutated recombination sites, integrated at one random chromosomal locus of the host cell genome. We further demonstrate that these LP-containing host cell lines can be used for the site-specific integration of several GOIs and thus, generation of transgene-expressing stable recombinant clones. Transgene expression was shown by site-specific integration of heavy and light chain genes coding for a monospecific antibody (msAb) as well as for a bi-specific antibody (bsAb). The genetic stability of the herein described LP-based recombinant clones expressing msAb or bsAb was demonstrated by cultivating the recombinant clones and measuring antibody titers over 85 generations. We conclude that the host cell containing multiple copies of MAR-rich landing pads can be successfully used for stable expression of one or several GOIs.
| Typ des Eintrags: | Artikel |
|---|---|
| Erschienen: | 2024 |
| Autor(en): | Oliviero, Claudia ; Hinz, Steffen C. ; Bogen, Jan P. ; Kornmann, Henri ; Hock, Björn ; Kolmar, Harald ; Hagens, Gerrit |
| Art des Eintrags: | Zweitveröffentlichung |
| Titel: | Generation of a host cell line containing a MAR-rich landing pad for site-specific integration and expression of transgenes |
| Sprache: | Englisch |
| Publikationsjahr: | 20 August 2024 |
| Ort: | Darmstadt |
| Publikationsdatum der Erstveröffentlichung: | 2022 |
| Ort der Erstveröffentlichung: | Malden, MA |
| Verlag: | Wiley |
| Titel der Zeitschrift, Zeitung oder Schriftenreihe: | Biotechnology Progress |
| Jahrgang/Volume einer Zeitschrift: | 38 |
| (Heft-)Nummer: | 4 |
| Kollation: | 16 Seiten |
| DOI: | 10.26083/tuprints-00027669 |
| URL / URN: | https://tuprints.ulb.tu-darmstadt.de/27669 |
| Zugehörige Links: | |
| Herkunft: | Zweitveröffentlichungsservice |
| Kurzbeschreibung (Abstract): | In recent years, targeted gene integration (TI) has been introduced as a strategy for the generation of recombinant mammalian cell lines for the production of biotherapeutics. Besides reducing the immense heterogeneity within a pool of recombinant transfectants, TI also aims at shortening the duration of the current cell line development process. Here we describe the generation of a host cell line carrying Matrix-Attachment Region (MAR)-rich landing pads (LPs), which allow for the simultaneous and site-specific integration of multiple genes of interest (GOIs). We show that several copies of each chicken lysozyme 5'MAR-based LP containing either BxB1 wild type or mutated recombination sites, integrated at one random chromosomal locus of the host cell genome. We further demonstrate that these LP-containing host cell lines can be used for the site-specific integration of several GOIs and thus, generation of transgene-expressing stable recombinant clones. Transgene expression was shown by site-specific integration of heavy and light chain genes coding for a monospecific antibody (msAb) as well as for a bi-specific antibody (bsAb). The genetic stability of the herein described LP-based recombinant clones expressing msAb or bsAb was demonstrated by cultivating the recombinant clones and measuring antibody titers over 85 generations. We conclude that the host cell containing multiple copies of MAR-rich landing pads can be successfully used for stable expression of one or several GOIs. |
| Freie Schlagworte: | antibody, CHO cells, protein expression, recombinant protein, site-specific integration |
| Status: | Verlagsversion |
| URN: | urn:nbn:de:tuda-tuprints-276693 |
| Sachgruppe der Dewey Dezimalklassifikatin (DDC): | 500 Naturwissenschaften und Mathematik > 540 Chemie 500 Naturwissenschaften und Mathematik > 570 Biowissenschaften, Biologie |
| Fachbereich(e)/-gebiet(e): | 07 Fachbereich Chemie 07 Fachbereich Chemie > Clemens-Schöpf-Institut > Fachgebiet Biochemie 07 Fachbereich Chemie > Clemens-Schöpf-Institut |
| Hinterlegungsdatum: | 20 Aug 2024 13:27 |
| Letzte Änderung: | 21 Aug 2024 05:20 |
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| Suche nach Titel in: | TUfind oder in Google |
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- Generation of a host cell line containing a MAR-rich landing pad for site-specific integration and expression of transgenes. (deposited 20 Aug 2024 13:27) [Gegenwärtig angezeigt]
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