title: Identification and elimination of target-related matrix interference in a neutralizing anti-drug antibody assay.
abstract: Biopharmaceuticals administered to the human body have the potential to trigger the production of anti-drug (also called anti-therapeutic) antibodies (ADA) that can neutralize the therapeutic activity. For antibody therapeutics, cell-based neutralizing ADA assays are frequently used to evaluate ADA in clinical studies. We developed a method to detect neutralizing antibodies against MEDI-575, a fully human IgG2κ antagonistic antibody against PDGFR-α. We evaluated three assay formats, two of which measured late responses, cell proliferation and apoptosis, whereas the third assay detected an early signaling event, phosphorylation of PDGFR-α. Measuring phosphorylation provided a superior assay window and therefore was developed as a neutralizing ADA (NAb) assay. Matrix interference, however, was significant, and could be identified to be caused by PDGF-AA and PDGF-AB, apparently the two most abundant ligands of PDGFR-α present in human serum samples. A simple pre-treatment step, addition of an inhibitory antibody to PDGF-A, a subunit present in PDGF-AA and PDGF-AB, was found to eliminate matrix interference, increasing assay reliability and sensitivity. We integrated the pre-treatment step into assay development and qualified a robust NAb assay.
authors.affiliation: MedImmune, Clinical Pharmacology and DMPK, 24500 Clawiter Rd, Hayward, CA, United States. Electronic address: schwickartm@medimmune.com.
authors.forename: Martin
authors.initials: M
authors.lastname: Schwickart
authors.affiliation: MedImmune, Clinical Pharmacology and DMPK, 24500 Clawiter Rd, Hayward, CA, United States.
authors.forename: Freshta
authors.initials: F
authors.lastname: Mehrzai
authors.affiliation: MedImmune, Clinical Pharmacology and DMPK, 24500 Clawiter Rd, Hayward, CA, United States.
authors.forename: Jennifer
authors.initials: J
authors.lastname: Pearson
authors.affiliation: MedImmune, Clinical Pharmacology and DMPK, 24500 Clawiter Rd, Hayward, CA, United States.
authors.forename: Nabila
authors.initials: N
authors.lastname: Shaghasi
authors.affiliation: MedImmune, Clinical Pharmacology and DMPK, 24500 Clawiter Rd, Hayward, CA, United States.
authors.forename: Carlos
authors.initials: C
authors.lastname: Chavez
authors.affiliation: MedImmune, Clinical Pharmacology and DMPK, 24500 Clawiter Rd, Hayward, CA, United States.
authors.forename: Amy
authors.initials: A
authors.lastname: Schneider
authors.affiliation: MedImmune, Clinical Pharmacology and DMPK, 24500 Clawiter Rd, Hayward, CA, United States.
authors.forename: Spencer
authors.initials: S
authors.lastname: Wu
authors.affiliation: MedImmune, Clinical Pharmacology and DMPK, 24500 Clawiter Rd, Hayward, CA, United States.
authors.forename: Lorin
authors.initials: L
authors.lastname: Roskos
authors.affiliation: MedImmune, Clinical Pharmacology and DMPK, 24500 Clawiter Rd, Hayward, CA, United States.
authors.forename: Meina
authors.initials: M
authors.lastname: Liang
mesh_terms: D000911:Antibodies, Monoclonal / Q000009:adverse effects / Q000276:immunology*; D057134:Antibodies, Neutralizing / Q000097:blood*; D017209:Apoptosis / Q000187:drug effects; D045744:Cell Line, Tumor; D049109:Cell Proliferation / Q000187:drug effects; D002470:Cell Survival / Q000187:drug effects; D004305:Dose-Response Relationship, Drug; D006801:Humans; D007118:Immunoassay / Q000379:methods*; D008024:Ligands; D010766:Phosphorylation; D010982:Platelet-Derived Growth Factor / Q000276:immunology / Q000378:metabolism; D011237:Predictive Value of Tests; D020796:Receptor, Platelet-Derived Growth Factor alpha / Q000037:antagonists & inhibitors* / Q000276:immunology / Q000378:metabolism; D015203:Reproducibility of Results; D015398:Signal Transduction / Q000187:drug effects
chemical_list: D000911:Antibodies, Monoclonal; D057134:Antibodies, Neutralizing; D008024:Ligands; D010982:Platelet-Derived Growth Factor; C075278:platelet-derived growth factor A; C095477:platelet-derived growth factor AB; D020796:Receptor, Platelet-Derived Growth Factor alpha
keywords: Cell-based; Immunogenicity; MEDI-575; Matrix interference; Neutralizing antibody; PDGF