TY  - JOUR
AU  - Walter, Justin D
AU  - Beffinger, Michal
AU  - Egloff, Pascal
AU  - Zimmermann, Iwan
AU  - Hürlimann, Lea M
AU  - Ackle, Fabian
AU  - Seifert, Matthias
AU  - Kobold, Sebastian
AU  - Vom Berg, Johannes
AU  - Seeger, Markus A
TI  - Flycodes enable simultaneous preclinical analysis for dozens of antibodies in single cassette-dosed mice.
JO  - Proceedings of the National Academy of Sciences of the United States of America
VL  - 122
IS  - 12
SN  - 0027-8424
CY  - Washington, DC
PB  - National Acad. of Sciences
M1  - DKFZ-2025-00671
SP  - e2426481122
PY  - 2025
AB  - Protein therapeutics such as antibodies require in-depth in vivo characterization during development and consequently account for a large proportion of laboratory animal consumption in the pharmaceutical industry. Currently, antibody candidates are exhaustively tested one-by-one in animal models to determine pharmacokinetic and pharmacodynamic (PK/PD) profiles. The simultaneous analysis of antibody mixtures in single animals, called cassette-dosing, could in principle overcome this bottleneck, but is currently limited to small cassette sizes. Here, we demonstrate how the use of genetically encoded peptide tags (flycodes), designed for maximal detectability in liquid chromatography-mass spectrometry, can allow for the simultaneous characterization of large pools of drug candidates, from single cassette-dosed mice. We demonstrate the simultaneous assessment of PK parameters for a group of >20 marketed/development-stage antibodies. Biodistribution experiments in mice bearing EGFR-expressing tumors correctly identified the two pool members recognizing EGFR, while organ analysis registered liver accumulation of an antibody targeting glucagon receptor, a protein profoundly expressed in that organ. In analogy to an early-phase drug development campaign, we performed biophysical and PK analysis for a cassette of 80 unique bispecific DARPin-sybody molecules. The data shown in this study originate from only 18 cassette-dosed mice, thereby demonstrating how flycode technology efficiently advances preclinical discovery pipelines allowing a direct comparison of drug candidates under identical experimental conditions.
KW  - Animals
KW  - Mice
KW  - ErbB Receptors: immunology
KW  - Humans
KW  - Tissue Distribution
KW  - Antibodies: immunology
KW  - Chromatography, Liquid: methods
KW  - Antibodies, Monoclonal: pharmacokinetics
KW  - Antibodies, Monoclonal: immunology
KW  - Drug Evaluation, Preclinical: methods
KW  - Female
KW  - antibodies (Other)
KW  - cassette dosing (Other)
KW  - flycodes (Other)
KW  - mass spectrometry (Other)
KW  - sybodies (Other)
KW  - ErbB Receptors (NLM Chemicals)
KW  - Antibodies (NLM Chemicals)
KW  - Antibodies, Monoclonal (NLM Chemicals)
LB  - PUB:(DE-HGF)16
C6  - pmid:40096612
DO  - DOI:10.1073/pnas.2426481122
UR  - https://inrepo02.dkfz.de/record/300194
ER  -