title: Validation of serum IGF-I as a biomarker to monitor the bioactivity of exogenous growth hormone agonists and antagonists in rabbits.
abstract: The development of new growth hormone (GH) agonists and growth hormone antagonists (GHAs) requires animal models for pre-clinical testing. Ideally, the effects of treatment are monitored using the same pharmacodynamic marker that is later used in clinical practice. However, intact rodents are of limited value for this purpose because serum IGF-I, the most sensitive pharmacodynamic marker for the action of GH in humans, shows no response to treatment with recombinant human GH and there is little evidence for the effects of GHAs, except when administered at very high doses or when overexpressed. As an alternative, more suitable model, we explored pharmacodynamic markers of GH action in intact rabbits. We performed the first validation of an IGF-I assay for the analysis of rabbit serum and tested precision, sensitivity, linearity and recovery using an automated human IGF-I assay (IDS-iSYS). Furthermore, IGF-I was measured in rabbits of different strains, age groups and sexes, and we monitored IGF-I response to treatment with recombinant human GH or the GHA Pegvisomant. For a subset of samples, we used LC-MS/MS to measure IGF-I, and quantitative western ligand blot to analyze IGF-binding proteins (IGFBPs). Although recovery of recombinant rabbit IGF-I was only 50% in the human IGF-I assay, our results show that the sensitivity, precision (1.7-3.3% coefficient of variation) and linearity (90.4-105.6%) were excellent in rabbit samples. As expected, sex, age and genetic background were major determinants of IGF-I concentration in rabbits. IGF-I and IGFBP-2 levels increased after single and multiple injections of recombinant human GH (IGF-I: 286±22 versus 434±26 ng/ml; P<0.01) and were highly correlated (P<0.0001). Treatment with the GHA lowered IGF-I levels from the fourth injection onwards (P<0.01). In summary, we demonstrated that the IDS-iSYS IGF-I immunoassay can be used in rabbits. Similar to rodents, rabbits display variations in IGF-I depending on sex, age and genetic background. Unlike in rodents, the IGF-I response to treatment with recombinant human GH or a GHA closely mimics the pharmacodynamics seen in humans, suggesting that rabbits are a suitable new model to test human GH agonists and antagonists.
authors.affiliation: Endocrine Research Unit, Medizinische Klinik und Poliklinik IV, Ludwig-Maximilians University, 80336 Munich, Germany.
authors.forename: Maximilian
authors.initials: M
authors.lastname: Bielohuby
authors.affiliation: Department of Biology, Faculty of Sciences, University of Isfahan, Isfahan 81746-73695, Iran. Department of Immunology, Medical School, Isfahan University of Medical Sciences, Isfahan 81746-73461, Iran. The Department of Human Metabolism, The University of Sheffield, Sheffield S10 2JF, UK.
authors.forename: Sayyed Hamid
authors.initials: SH
authors.lastname: Zarkesh-Esfahani
authors.affiliation: Immunodiagnostic Systems, Boldon, Tyne and Wear NE35 9PD, UK.
authors.forename: Jenny
authors.initials: J
authors.lastname: Manolopoulou
authors.affiliation: Ligandis GbR, Wilhelm-Stahl-Allee 2, 18196 Dummerstorf, Germany.
authors.forename: Elisa
authors.initials: E
authors.lastname: Wirthgen
authors.affiliation: German Sport University Cologne, Institute of Biochemisty/Center for Preventive Doping Research, 50933 Cologne, Germany.
authors.forename: Katja
authors.initials: K
authors.lastname: Walpurgis
authors.affiliation: Department of Immunology, Medical School, Isfahan University of Medical Sciences, Isfahan 81746-73461, Iran.
authors.forename: Mohaddeseh
authors.initials: M
authors.lastname: Toghiany Khorasgani
authors.affiliation: Department of Biology, Faculty of Sciences, University of Isfahan, Isfahan 81746-73695, Iran.
authors.forename: Zahra Sadat
authors.initials: ZS
authors.lastname: Aghili
authors.affiliation: The Department of Human Metabolism, The University of Sheffield, Sheffield S10 2JF, UK.
authors.forename: Ian Robert
authors.initials: IR
authors.lastname: Wilkinson
authors.affiliation: Leibniz Institute for Farm Animal Biology (FBN), Institute of Genome Biology, 18196 Dummerstorf, Germany.
authors.forename: Andreas
authors.initials: A
authors.lastname: Hoeflich
authors.affiliation: German Sport University Cologne, Institute of Biochemisty/Center for Preventive Doping Research, 50933 Cologne, Germany.
authors.forename: Mario
authors.initials: M
authors.lastname: Thevis
authors.affiliation: The Department of Human Metabolism, The University of Sheffield, Sheffield S10 2JF, UK.
authors.forename: Richard J
authors.initials: RJ
authors.lastname: Ross
authors.affiliation: Endocrine Research Unit, Medizinische Klinik und Poliklinik IV, Ludwig-Maximilians University, 80336 Munich, Germany. Martin.Bidlingmaier@med.uni-muenchen.de.
authors.forename: Martin
authors.initials: M
authors.lastname: Bidlingmaier
mesh_terms: D000818:Animals; D015415:Biomarkers / Q000097:blood*; D002853:Chromatography, Liquid; D013006:Growth Hormone / Q000819:agonists* / Q000037:antagonists & inhibitors*; D007334:Insulin-Like Growth Factor I / Q000378:metabolism*; D057230:Limit of Detection; D011817:Rabbits; D015203:Reproducibility of Results; D053719:Tandem Mass Spectrometry
chemical_list: D015415:Biomarkers; D007334:Insulin-Like Growth Factor I; D013006:Growth Hormone
keywords: Acromegaly; Animal model; Growth hormone deficiency; Pharmacodynamic marker