COMPREHENSIVE EVALUATION OF PD-L1 EXPRESSION IN CIRCULATING TUMOR CELLS (CTCs) AND PLASMA-DERIVED EXOSOMES IN TRIPLE NEGATIVE BREAST CANCER PATIENTS

Introduction: Triple-negative (TN) is the most aggressive subtype of breast cancer (BC) accounting for approximately 15% of all BC cases. Available target therapies are limited and often insufficient for these patients. Liquid biopsy is a non-invasive method for identifying cancer biomarkers and potential therapeutic targets in body fluids, examining different components such as circulating tumor cells (CTCs), nucleic acids, and extracellular vesicles/exosomes. Aim: Our study aimed to evaluate the presence of programmed cell death ligand 1(PD-L1) in CTCs and plasma-derived exosomes, from TNBC patients, and address their clinical relevance. Methods: Our research enrolled 35 TNBC patients and 12 healthy donors. CTCs were isolated by ficoll density gradient centrifugation, followed by triple immunofluorescence experiments and VyCAP analysis. Additionally, exosomes were isolated from the plasma of the same cohort of patients and characterized by transmission electron microscopy and Western blot analysis. Results: CTCs were detected in 20% of cases. Notably, PD-L1 was identified in 6 out of 7 CK-positive patients (85.7%), while PD-L1 negative CTCs were detected only in 1 case (14.3%). The CK+PD-L1+CD45- phenotype was associated with shorter progression-free survival (PFS) (log-rank p=0.006, HR=7.8). On the other hand, PD-L1 exosomal expression in patients was lower compared to healthy donors. However, in 22.9% of patients, PD-L1 exosomal expression exceeded the median observed in healthy donors and was associated with significantly poorer PFS (log-rank p=0.046, HR=6.4). Finally, the combination of (CK+PD-L1+CD45-) phenotype in CTCs and PD-L1 exosomal overexpression showcased a statistically significant worse PFS (log-rank p=0.018, HR=6.4). Conclusions: Overexpression of PD-L1 in either CTCs or exosomes is associated with worse clinical outcome. Furthermore, the combined evaluation of exosomes and CTCs provides an interesting approach to classify patients at higher risk of relapse. Furthermore, PD-L1 overexpression could potentially serve as a therapeutic target for these patients.