Purpose: Pharmacokinetic (PK) analyses can substantially aid in the selection of an optimal human/humanized IgG-based therapeutic among other similar drug candidates. We developed and validated (fit-for-purpose) an immunoassay capable of accurately quantitating human IgG in plasma of mice, rats, and monkeys. The method allows for PK analyses of human IgG-based therapeutic candidates in these species without needing de novo method development and characterization for go/no-go decision on applicable drug candidates.
Methods: Meso Scale Discovery (MSD) electrochemiluminescence (ECL) was the assay platform. Standard MSD assay plates were coated (in PBS) with goat anti-human IgG that had been extensively adsorbed with monkey IgG. Assay diluent was PBS containing 1% BSA and 0.05% Tween-20. Matrices included K2EDTA plasma from mice, rats, and monkeys, diluted to the minimum required dilution of 1:100 after spiking neat (≥95%). Binding of human IgG to the assay wells was detected with monkey IgG-adsorbed, biotinylated goat anti-human IgG, followed by streptavidin-SULFO-TAG. All incubations subsequent to plate coating were for 1 hour at room temperature.
Results: Standard curves of human IgG in mouse, rat, and monkey plasma were essentially superimposable at the MRD of 1:100. Separate Accuracy and Precision runs were conducted for each species with validation samples in the corresponding matrix for LLOQ, LQC, IQC, HQC, and ULOQ. An assay range of 7.5 ng/mL LLOQ to 5.5 g/mL ULOQ was established for all species with MRD factored in the calculation. Regardless of species, intra- and inter-assay % deviation values were all within 0.9%-18.5% of nominal, with total error values ranging from 7% to 28% in all cases. Other standard validation parameters tested in the 3 matrices including linearity of dilution, hook effect, and selectivity, all of which met acceptance criteria.
Conclusion: This method allows for rapid PK assessment of a broad spectrum of human/humanized IgG-based therapeutic candidates without the need for individualized method development and validation for each drug candidate. Go/no-go decisions on individual drug candidates based on this method would then help drive selection for full validation of the optimal molecular entity.
Trang Le– MicroConstants, Inc.
Alvin Yee– MicroConstants, Inc.
Dustin Lu– MicroConstants, Inc.
Veronica Ortega– MicroConstants, Inc.
John Marcelletti– MicroConstants, Inc.
John Marcelletti– Senior Laboratory Director, MicroConstants, Inc., San Diego