(T1330-02-09) Gyrolab Generic Anti-drug Antibody Assay Streamlines Preclinical Immunogenicity Assessment

Purpose: Antibody therapies can elicit an immune responses in the host organisms leading to anti-drug antibody (ADA) generation.  These ADAs can significantly impact therapeutic efficacy, pharmacokinetics, and safety in pre-clinical- and clinical studies. The prevalent assay format used in the preclinical setting to detect ADA is the bridging ELISA employed in ADA assessment in clinical studies, where the antibody therapeutic is used as both the capture and detection reagent. The advantage of this format is the ability to detect all Ig isotypes with adequate sensitivity and drug tolerance. However, this approach requires generation of assay reagents and optimization of assay for each molecule which can be time and resource intensive. A preclinical ADA assay employing a plug and play format suitable for all types of mAB therapeutics would offer a streamlined, optimization-free approach to accelerate drug development programs in preclinical research.

Methods: In our previous work, we introduced a generic ADA assay that utilizes the same capture, detection and positive control reagents for assessment of total ADA in non-clinical cynomolgus monkey studies. Although this format works well it has a limited dynamic range which does not allow for replacement of titer values with signal to noise ratio. In this study, we demonstrate the adaptability of this generic assay format for use on the Gyrolab, an automated immunoassay platform. This adaptation significantly reduces hands-on time and enhances assay reproducibility.

Results: We first evaluated the commercial Gyrolab generic cyno ADA kit reagents and observed that signal-to-noise can be dramatically improved by using optimized assay reagents. As with the ELISA platform, our assay performs similarly across therapeutic mAb with different IgG isotypes backbones, exhibiting similar sensitivity and detection range. Our observations reveal that the Gyrolab-based ADA assay possesses an expanded signal dynamic range and greater drug tolerance as compared to the total ADA assay performed using ELISA. In a study involving cynomolgus monkeys treated with a single dose of a mAb therapeutic, we assessed the ADA response using bridging ELISA, total ADA ELISA and total ADA Gyrolab assays. We noted that the Gyrolab-based assay detected host ADA responses at earlier time points, particularly in cases with unexpected drug clearance profiles, when compared to the bridging ELISA method. In addition, the correlation between signal-to-noise and titer is excellent on the Gyrolab platform (Spearman’s r > 0.9).

Conclusion: Collectively, our findings underscore the robustness of adapting the total ADA Gyrolab assay for non-clinical biotherapeutic studies enabling reliable detection of ADA responses, as well as level of ADA response without a need for titer evaluations, facilitating the interpretation of pharmacological data with minimal need for assay optimization.

References: M. Carrasco-Triguero, H. Davis, Y. Zhu, D. Coleman, D. Nazzal, P. Vu, S. Kaur, Application of a Plug-and-Play Immunogenicity Assay in Cynomolgus Monkey Serum for ADCs at Early Stages of Drug Development, J Immunol Res 2016 (2016) 2618575.

Acknowledgements: The authors are employees of Genentech, Inc., and own stock in F. Hoffman-La Roche Ltd.

Figure 1. ADA assay formats used in this study. A) Therapeutic-specific biotin-DIG ELISA, also known as the bridging ELISA. B) Total ADA assay using ELISA. C) Total ADA assay using Gyrolab. The total ADA assays detect both drug-bound and unbound ADA as excess drug is added during the sample pre-incubation step.