Circulating extracellular vesicles carrying PD-1, PD-L1 and CTLA-4 inform resistance to anti-PD-L1-based therapy in
João Gorgulho1,2, Ramsha Masood3,4, Gustav Buescher3,4
1Department of Oncology, Hematology and Bone Marrow Transplantation with Section of Pneumology, University Medical Center Hamburg-Eppendorf, Hamburg, Germany.
Background:
First-line immune checkpoint inhibitor (ICI) therapy has contributed to improved outcomes in hepatocellular carcinoma (HCC). However, only around 30% of patients respond, highlighting the need for biomarker-driven patient selection. Considering the lack of tissue acquirement in HCC in clinical routine, liquid biopsy holds great promise.
Objective:
We aimed at profiling immune checkpoints (IC) on circulating extracellular vesicles (EVs) as potential liquid biopsy-based biomarkers for prediction of treatment response.
Design:
Three distinct cohorts were analysed in this study: (1) HCC explorer cohort (n=40), testing the presence of membrane-bound ICs on EVs; (2) early-stage HCC cohort (n=37 with paired blood and tissue), assessing the interplay between EV and tissue ICs alongside clinicopathological parameters and (3) treatment cohort (n=202 with 600 sequential blood samples), comprising an ICI treated cohort with a training (n=79, n=402 sequential samples) and a validation group (n=82, n=146 sequential samples), alongside a tyrosine-kinase inhibitor (TKI) treated cohort (n=41, n=52 sequential samples), to identify predictors of immunotherapy response through IC profiling of circulating EVs via multiplex immunoassay.
Results:
Programmed cell death protein 1 (PD-1), programmed death-ligand 1 (PD-L1) and cytotoxic T-lymphocyte associated protein 4 (CTLA-4) were enriched in EV fractions compared to EV-depleted serum. Baseline and early dynamics of EV-IC levels significantly discriminated between responders and non-responders in the ICI-treated training and validation cohorts, while simultaneously predicting progression-free survival (PFS) and overall survival (OS). These findings were not reproducible in the TKI-treated cohort. In addition, dynamic changes of EV-IC levels during therapy in patients with initial response predicted acquired therapeutic resistance, approximately 36-42 weeks before progression was traceable on imaging.
Conclusion:
EVs carrying PD-1, PD-L1 and CTLA-4 represent readily quantifiable, non-invasive biomarkers to predict response and survival in patients with advanced HCC undergoing ICI therapy and serve to identify biological ICI resistance much earlier than current standards with imaging.
Insights
Extracellular vesicles (EVs) carrying immune checkpoints like PD-1, PD-L1, and CTLA-4 can predict treatment response and survival in advanced hepatocellular carcinoma (HCC) patients receiving immune checkpoint inhibitor (ICI) therapy. This liquid biopsy approach identifies resistance earlier than imaging.
Area of Science:
- Oncology
- Immunology
- Biomarker Discovery
Background:
- Immune checkpoint inhibitor (ICI) therapy improves outcomes in hepatocellular carcinoma (HCC), but response rates are low (~30%).
- Biomarker-driven patient selection is crucial for optimizing ICI therapy in HCC.
- Liquid biopsy offers a promising alternative to tissue biopsy for biomarker analysis in HCC due to difficulties in tissue acquisition.
Purpose of the Study:
- To profile immune checkpoints (ICs) on circulating extracellular vesicles (EVs).
- To evaluate EVs as potential liquid biopsy-based biomarkers for predicting treatment response to ICI therapy in HCC.
- To assess the predictive value of EV-IC profiling for survival and therapeutic resistance.
Main Methods:
- Analysis of three distinct HCC cohorts including an explorer cohort (n=40), an early-stage cohort with paired blood and tissue (n=37), and a large treatment cohort (n=202) with sequential samples.
- Profiling of membrane-bound ICs, including programmed cell death protein 1 (PD-1), programmed death-ligand 1 (PD-L1), and cytotoxic T-lymphocyte associated protein 4 (CTLA-4), on circulating EVs using multiplex immunoassay.
- Comparison of EV-IC levels with treatment response, progression-free survival (PFS), overall survival (OS), and imaging-based progression in both ICI- and tyrosine-kinase inhibitor (TKI)-treated patients.
Main Results:
- PD-1, PD-L1, and CTLA-4 were significantly enriched on EVs compared to EV-depleted serum.
- Baseline and early dynamic changes in EV-IC levels effectively discriminated between responders and non-responders in ICI-treated cohorts, predicting PFS and OS.
- Dynamic changes in EV-IC levels during therapy predicted acquired resistance to ICI therapy 36-42 weeks before imaging detection.
Conclusions:
- Circulating EVs carrying PD-1, PD-L1, and CTLA-4 are quantifiable, non-invasive biomarkers for predicting response and survival in advanced HCC patients undergoing ICI therapy.
- EV-IC profiling enables earlier identification of biological ICI resistance compared to current imaging standards.
- This liquid biopsy approach facilitates personalized treatment strategies for HCC patients receiving immunotherapy.
