Development of an electrochemiluminescence-based bridging assay to detect antibodies against a PTH inverse agonist in human plasma.

BACKGROUND

Therapeutic peptide-induced anti-drug antibodies (ADA) can have various effects, including adverse events and drug-neutralizing activity which can lead to loss of efficacy. This has led regulatory agencies to require nonclinical and clinical testing of biotherapeutics to assess immunogenicity risk.

RESEARCH DESIGN AND

METHODS

An electrochemiluminescence bridging assay was developed to detect antibodies against parathyroid hormone receptor inverse agonist (PTH-IA), a novel synthetic peptide that is being developed to treat Jansen's Metaphyseal Chondrodysplasia. Biotinylated PTH-IA and ruthenylated PTH-IA reagents were synthesized via amine-coupling chemistry, and rat polyclonal anti-PTH-IA positive control antibody was generated by immunizing rats with PTH-IA conjugated to keyhole limpet hemocyanin. The reagents were utilized to determine cut points for the screening and confirmatory assays and assay sensitivity in drug-naïve human plasma.

RESULTS

Sensitivity of the screening assay was determined to be 3.61 ng/mL at cut point. The cut point for the confirmatory assay was determined to be 9.34% inhibition, and the cut point factor used to establish the floating plate cut point from the negative control signal was 1.055. The assay was found to be selective and reproducible.

CONCLUSION

The assay's performance characteristics demonstrated its suitability for detecting anti-PTH-IA antibodies in human plasma.