title: Development of Immunoassays for the Quantitative Assessment of Amyloid-β in the Presence of Therapeutic Antibody: Application to Pre-Clinical Studies.
abstract: Utilizing decision making biomarkers in drug development requires thorough assay validation. Special considerations need to be taken into account when monitoring biomarkers using immunoassays in the presence of therapeutic antibodies. We have developed robust and sensitive assays to assess target engagement and proof of mechanism to support the clinical progression of a human monoclonal antibody against the neurotoxic amyloid-β (Aβ)42 peptide. Here we present the introduction of novel pre-treatment steps to ensure drug-tolerant immunoassays and describe the validation of the complete experimental procedures to measure total Aβ42 concentration (bound and unbound) in cerebrospinal fluid (CSF) and plasma, free Aβ42 concentration (unbound) in CSF, and Aβ40 concentration in CSF. The difference in composition of the matrices (CSF and plasma) and antigen levels therein, in combination with the hydrophobic properties of Aβ protein, adds to the complexity of validation. Monitoring pharmacodynamics of an Aβ42 specific monoclonal antibody in a non-human primate toxicology study using these assays, we demonstrated a 1500-fold and a 3000-fold increase in total Aβ42 in plasma, a 4-fold and 8-fold increase in total Aβ42 in CSF together with a 95% and 96% reduction of free Aβ42 in CSF following weekly intravenous injections of 10 mg/kg and 100 mg/kg, respectively. Levels of Aβ40 were unchanged. The accuracy of these data is supported by previous pre-clinical studies as well as predictive pharmacokinetic/pharmacodynamics modeling. In contrast, when analyzing the same non-human primate samples excluding the pre-treatment steps, we were not able to distinguish between free and total Aβ42. Our data clearly demonstrate the importance of thorough evaluation of antibody interference and appropriate validation to monitor different types of biomarkers in the presence of a therapeutic antibody.
authors.affiliation: AstraZeneca Translational Science Center, Personalized Healthcare & Biomarkers AstraZeneca R&D Innovative Medicines and Department of Clinical Neuroscience, Solna, Sweden; Science for Life Laboratory, Karolinska Institutet, Sweden.
authors.forename: Anna
authors.initials: A
authors.lastname: Bogstedt
authors.affiliation: Antibody Discovery and Protein Engineering, MedImmune Limited, Granta Park, Cambridge, UK.
authors.forename: Maria
authors.initials: M
authors.lastname: Groves
authors.affiliation: Translational Science, MedImmune Limited, Granta Park Cambridge, UK.
authors.forename: Keith
authors.initials: K
authors.lastname: Tan
authors.affiliation: Clinical Pharmacology & DMPK, MedImmune Limited, Gaithersburg, MD, USA.
authors.forename: Rajesh
authors.initials: R
authors.lastname: Narwal
authors.affiliation: Biologics Safety Assessment, Translational Science, MedImmune Limited, Cambridge, UK.
authors.forename: Mary
authors.initials: M
authors.lastname: McFarlane
authors.affiliation: AstraZeneca Translational Science Center, Personalized Healthcare & Biomarkers AstraZeneca R&D Innovative Medicines and Department of Clinical Neuroscience, Solna, Sweden; Science for Life Laboratory, Karolinska Institutet, Sweden.
authors.forename: Kina
authors.initials: K
authors.lastname: Höglund
mesh_terms: D000544:Alzheimer Disease / Q000097:blood / Q000134:cerebrospinal fluid / Q000139:chemically induced / Q000628:therapy*; D016229:Amyloid beta-Peptides / Q000097:blood / Q000134:cerebrospinal fluid* / Q000276:immunology* / Q000633:toxicity*; D000818:Animals; D000911:Antibodies, Monoclonal / Q000627:therapeutic use*; D004195:Disease Models, Animal; D018450:Disease Progression; D004353:Drug Evaluation, Preclinical; D005260:Female; D006801:Humans; D007118:Immunoassay / Q000379:methods*; D008252:Macaca fascicularis; D008297:Male; D010446:Peptide Fragments / Q000097:blood / Q000134:cerebrospinal fluid* / Q000276:immunology* / Q000633:toxicity*; D015203:Reproducibility of Results; D013997:Time Factors
chemical_list: D016229:Amyloid beta-Peptides; D000911:Antibodies, Monoclonal; D010446:Peptide Fragments; C078592:amyloid beta-protein (1-40); C075222:amyloid beta-protein (1-42)
keywords: Alzheimer’s disease; amyloid-β; biomarkers; cerebrospinal fluid; immunotherapy; plasma; pre-clinical