Recommendations for The Development and Validation of Confirmatory Anti-Drug Antibody Assays

Abstract

Identification and characterization of anti-drug antibodies is a critical component of biopharmaceutical drug development. The tiered approach for immunogenicity testing consists of screening, confirmatory, and characterization assays. Herein, we provide recommendations for confirmatory assays by expanding upon published guidance and present common practices across the industry. The authors recommend scientific approaches for development and validation of confirmatory assays using competition methods in ligand-binding assays, along with statistical formulae for routine use and validation. The paper will assist in understanding the confirmatory assay, and carefully implementing validation criteria a priori, as well as during sample analysis. These approaches represent the authors' current knowledge and practices, with the aim that more uniform practices will be applied across the industry.

Assessment of clinical immunogenicity of inotuzumab ozogamicin in patients with non-Hodgkin lymphoma and acute lymphoblastic leukemia

“…Samples for immunogenicity assessments were collected based on time points specified in each individual clinical study. During early NHL studies, immunogenicity response was measured using an ELISA platform, which was considered a standard of practice at the time, for the detection of Hoofring et al, 2013;Jani et al, 2015;Shankar et al, 2008). Similar bridging assay platforms have successfully been used for other approved ADCs (CarrascoTriguero et al, 2013).…”

“…For the confirmatory assay, confirmatory cut point was calculated by testing serum samples from 50 treatment-naive patients with ALL (BioreclamationIVT). The serum samples were incubated with an excess amount of InO before analysis in a competitive binding format (Jani et al, 2015). For each sample, the percentage inhibition values in the confirmatory assay were calculated.…”

“…The screening and confirmatory assay cut points were statistically determined based on the current industry white papers (Jani et al, 2015;Shankar et al, 2008). A screening cut point was derived from 50 treatment-naive ALL patients (BioreclamationIVT) by targeting a 5% false-positive rate to minimize potential false-negatives.…”

“…The screen cut point factor (SCPF) and confirmatory/ characterization cut point (CCP) were determined using 50 commercially procured ALL serum samples according to proposed methods (Jani et al, 2015;Shankar et al, 2008) (95% CI for screen cut point [SCP] and 99.9% CI for CCPs for InO, G544, and calicheamicin-BSA). To obtain the SCP for the assay, the NC signal was multiplied by the SCPF of 1.47.…”

“…Independent anti-InO and anti-rituximab ELISA methods were developed and validated as per the scientific experience and best practices of laboratories where assays were developed. In the recent ALL studies, a Meso Scale Discovery (MSD) based bridging electrochemiluminescence (ECL) assay was developed and validated as per the regulatory expectations and current industry white papers (Center for Drug Evaluation and Research, US Food and Drug Administration, 2009; Committee for Medicinal Products for Human Use, 2007;Hoofring et al, 2013;Jani et al, 2015;Shankar et al, 2008). Similar bridging assay platforms have successfully been used for other approved ADCs (Carrasco-Triguero et al, 2013).…”