Tu161 - Flow Cytometry–based Functional Assay to Study Rituximab, Obinutuzumab and Blinatumomab Mechanisms of Action and Capture Donor Heterogeneity

Donata DEMARCO – Hoffmann-La Roche, Basel, Switzerland; Corinne PLOIX – Hoffmann-La Roche, Basel, Switzerland; Gethin THOMAS – Hoffmann-La Roche, Basel, Switzerland; Anneliese SCHNEIDER – Hoffmann-La Roche, Basel, Switzerland; Jean-Marc BUSNEL – Beckman Coulter Life Sciences, R&D department, Marseille, France

Abstract Text: Many B-cell targeting biologics already available today have proven efficacious treatment modalities, and more are in clinical development. The result is a wide range of therapeutics that differ in structure, specificity, and mechanism of action (MoA). As patients further constitute a very heterogeneous population, patient stratification and treatment personalization are a major challenge to achieve the highest treatment efficacy and the lowest magnitude of adverse events in an individual patient.

In this study, we used a whole blood–based functional assay strategy to capture subject heterogeneity in terms of engagement of effector cells and activation of bystander cells, and to characterize drug MoA. Using three flow cytometry panels, we examined the activation phenotype of whole blood immune cells after overnight stimulation with Rituximab, Obinutuzumab or Blinatumomab.

As expected, Rituximab and Obinutuzumab were associated with target cell depletion and NK activation, with a strong antibody-dependent cellular cytotoxicity (ADCC) in Obinutuzumab. Blinatumomab was associated with a strong T cell response and a high target cell activation. In bystander cells, membrane markers and cytokine production were strongly induced by Obinutuzumab and Blinatumomab. Interestingly, in patient samples, the effector response was higher than in healthy donors, whereas bystander activation was weaker. In both healthy donors and patients, the range of responses was variable, demonstrating the ability to capture subject heterogeneity.

The proposed approach allowed to capture donor heterogeneity and drug MoA. We believe it might represent a valuable option to guide treatment personalization and needs to be further evaluated in translational research.