Ex vivo pembrolizumab pharmacology for personalized PD-1 inhibitor therapy reveals a critical gap between receptor occupancy and T cell functionality
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Updated: May 17, 2025
Identifying PD-1/PD-L1 Inhibitors with Surface Plasmon Resonance Technology
Published on: May 2, 2025
Judith D J Verdonk1, Berber Piet2, Rob Ter Heine3
1Laboratory of Medical Immunology, Department of Laboratory Medicine, Radboudumc, Nijmegen, The Netherlands.
Purpose:
Targeting the programmed death (ligand)-1 (PD-1/PD-L1) axis with immune checkpoint inhibitors (ICIs), like pembrolizumab, has improved cancer survival. Unfortunately, the optimal dose remains unknown and less than 50 % of patients respond. Understanding PD-1 receptor pharmacology and developing early-response biomarkers are crucial to personalize therapy. We hypothesize that characterizing individual pre-treatment variability in immune responses to pembrolizumab will enhance PD-1 receptor pharmacology insights and improve treatment response prediction. Hence, this study evaluates the performance of a newly developed ex vivo immunopharmacological bioassay under healthy and pathological states.
Methods:
Peripheral blood mononuclear cells from healthy individuals and non-small cell lung cancer (NSCLC) patients were stimulated in vitro in the presence of pembrolizumab. PD-1 expression, interleukin-2 (IL-2) secretion, T cell differentiation, expression of activation markers and phosphorylation of T cell signalling molecules were analysed.
Results:
In healthy individuals, receptor saturation occurred at >0.025 μg/mL pembrolizumab. Yet IL-2 production still increased significantly beyond this concentration. NSCLC patients showed significantly higher PD-1 expression and IL-2 production than healthy individuals. Nevertheless, pembrolizumab induced IL-2 production similarly in both cohorts. In NSCLC patients, pembrolizumab inhibited differentiation towards CD4 central memory T cells. Remarkably, phosphorylation of proximal phospho-markers in response to pembrolizumab varied between NSCLC patients, potentially discriminating responders from non-responders.
Conclusions:
We highlight the importance of evaluating T cell functionality alongside PD-1 receptor occupancy. We identified PD-1-induced modulation of phosphorylation of proximal signalling molecules as potential predictors for ICI treatment response in NSCLC. We recommend further development of this immunopharmacological bioassay for personalization of ICI treatment.
Insights
This study reveals that analyzing T cell signaling molecule phosphorylation can predict responses to pembrolizumab in non-small cell lung cancer (NSCLC) patients, aiding personalized immunotherapy.
Area of Science:
- Immunology
- Pharmacology
- Oncology
Background:
- Immune checkpoint inhibitors (ICIs) like pembrolizumab targeting the PD-1/PD-L1 axis improve cancer survival.
- Optimal dosing and predicting response to ICIs remain challenges, with less than 50% of patients responding.
- Understanding PD-1 receptor pharmacology and identifying early biomarkers are crucial for personalized cancer therapy.
Purpose of the Study:
- To evaluate an ex vivo immunopharmacological bioassay for characterizing individual immune responses to pembrolizumab.
- To investigate pre-treatment variability in immune responses for enhanced PD-1 receptor pharmacology insights.
- To improve prediction of treatment response in non-small cell lung cancer (NSCLC) patients.
Main Methods:
- Peripheral blood mononuclear cells (PBMCs) from healthy donors and NSCLC patients were stimulated with pembrolizumab in vitro.
- Analyzed PD-1 expression, interleukin-2 (IL-2) secretion, T cell differentiation, and activation markers.
- Assessed phosphorylation of T cell signaling molecules in response to pembrolizumab.
Main Results:
- In healthy individuals, PD-1 receptor saturation occurred at >0.025 μg/mL pembrolizumab, but IL-2 production increased beyond this concentration.
- NSCLC patients exhibited higher PD-1 expression and IL-2 production than healthy individuals, with similar IL-2 induction by pembrolizumab.
- Pembrolizumab inhibited CD4 central memory T cell differentiation in NSCLC patients, and phosphorylation patterns varied, potentially distinguishing responders from non-responders.
Conclusions:
- Evaluating T cell functionality alongside PD-1 receptor occupancy is important for immunotherapy.
- PD-1-induced modulation of proximal signaling molecule phosphorylation may predict ICI treatment response in NSCLC.
- The developed immunopharmacological bioassay shows potential for personalizing ICI treatment.
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