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首页|Functional diversification of hybridoma produced antibodies by CRISPR/HDR genomic engineering
10.1101/551382
来源：bioRxiv_logobioRxiv

Functional diversification of hybridoma produced antibodies by CRISPR/HDR genomic engineering

Functional diversification of hybridoma produced antibodies by CRISPR/HDR genomic engineering

Becker Anouk M.D. 1Dalen Duco van 1van Bruggen J. Armando C. 2Bentlage Arthur E.H. 3Heck Albert J.R. 4Scheeren Ferenc A. 5Nederend Maaike 6Hagemans Iris M. 1Fransen Marieke F. 7Vidarsson Gestur 3der Schoot Johan M.S. van 1Dolen Yusuf 1Valente Michael 1Cevirgel Alper 1Le Gall Camille M. 1Figdor Carl G. 1Leusen Jeanette H.W. 6Caval Tomislav 4Verdoes Martijn 1Engelfriet M 2Fennemann Felix L.1

1. Department of Tumor Immunology, Radboud Institute for Molecular Life Sciences, Radboud University Medical Center 2. Division of Immunology, The Netherlands Cancer Institute 3. Sanquin Research, Department of Experimental Immunohematology, Amsterdam, The Netherlands, and Landsteiner Laboratory, Amsterdam UMC, University of Amsterdam 4. Biomolecular Mass Spectrometry and Proteomics, Bijvoet Center for Biomolecular Research and Utrecht Institute for Pharmaceutical Sciences, Utrecht University 5. Department of Medical Oncology, Department of Surgery and Department of Immunohematology and Blood Transfusion, Leiden University Medical Center (LUMC) 6. Laboratory for Translational Immunology, UMC Utrecht 7. Department of Immunohematology and Blood Transfusion, Leiden University Medical Center (LUMC)

医学研究方法生物科学研究方法、生物科学研究技术生物工程学

Becker Anouk M.D.,Dalen Duco van,van Bruggen J. Armando C.,Bentlage Arthur E.H.,Heck Albert J.R.,Scheeren Ferenc A.,Nederend Maaike,Hagemans Iris M.,Fransen Marieke F.,Vidarsson Gestur,der Schoot Johan M.S. van,Dolen Yusuf,Valente Michael,Cevirgel Alper,Le Gall Camille M.,Figdor Carl G.,Leusen Jeanette H.W.,Caval Tomislav,Verdoes Martijn,Engelfriet M,Fennemann Felix L..Functional diversification of hybridoma produced antibodies by CRISPR/HDR genomic engineering[EB/OL].(2025-03-28)[2025-09-24].https://www.biorxiv.org/content/10.1101/551382.点此复制

Abstract Hybridoma technology is instrumental for the development of novel antibody therapeutics and diagnostics. Recent preclinical and clinical studies highlight the importance of antibody isotype for therapeutic efficacy. However, since the sequence encoding the constant domains is fixed, tuning antibody function in hybridomas has been restricted. Here, we demonstrate a versatile CRISPR/HDR platform to rapidly engineer the constant immunoglobulin domains to obtain recombinant hybridomas which secrete antibodies in the preferred format, species and isotype. Using this platform, we obtained recombinant hybridomas secreting Fab’ fragments, isotype switched chimeric antibodies, and Fc-silent mutants. These antibody products are stable, retain their antigen specificity, and display their intrinsic Fc-effector functions in vitro and in vivo. Furthermore, we can site-specifically attach cargo to these antibody products via chemo-enzymatic modification. We believe this versatile platform facilitates antibody engineering for the entire scientific community, empowering preclinical antibody research. One Sentence SummaryWe demonstrate a universal CRISPR/HDR based platform for rapid genetic engineering of hybridomas to obtain functionally diverse antibody isotype panels in the species and format of choice.
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