A Novel Approach for Quantifying the Pharmacological Activity of T-Cell Engagers Utilizing In Vitro Time Course Experiments and Streamlined Data Analysis

Abstract

CD3-bispecific antibodies are a new class of immunotherapeutic drugs against cancer. The pharmacological activity of CD3-bispecifics is typically assessed through in vitro assays of cancer cell lines co-cultured with human peripheral blood mononuclear cells (PBMCs). Assay results depend on experimental conditions such as incubation time and the effector-to-target cell ratio, which can hinder robust quantification of pharmacological activity. In order to overcome these limitations, we developed a new, holistic approach for quantification of the in vitro dose–response relationship. Our experimental design integrates a time-independent analysis of the dose–response across different time points as an alternative to the static, “snap-shot” analysis based on a single time point commonly used in dose–response assays. We show that the potency values derived from static in vitro experiments depend on the incubation time, which leads to inconsistent results across multiple assays and compounds. We compared the potency values from the time-independent analysis with a model-based approach. We find comparably accurate potency estimates from the model-based and time-independent analyses and that the time-independent analysis provides a robust quantification of pharmacological activity. This approach may allow for an improved head-to-head comparison of different compounds and test systems and may prove useful for supporting first-in-human dose selection.

Strategies for clinical dose optimization of T cell-engaging therapies in oncology

“…It was then demonstrated that a more robust determination of EC 50 could be obtained when taking into account multiple time points by using the area under the effect curve to derive a time-independent measure of potency. 52 Other factors such as the inter-individual variability across peripheral blood mononuclear cell (PBMC) donors can also have an impact on the potency values determined using these assays. The above analysis for cibisatamab showed that the time-independent approach actually gave a similar EC 50 value compared to single time point approach as long as the most appropriate time point was chosen, and the authors proposed a pragmatic workflow in which the time course of biological activity could initially be used to determine potency using relatively few PBMC donors, and then the most relevant time point could be selected and EC 50 determined with a greater number of PBMC donors in order to evaluate inter-donor variability.…”