Predictive Analysis in Oral Cancer Immunotherapy: Profiling Dual PD-L1-Positive Extracellular Vesicle Subtypes with Step-Wedge Microfluidic Chips

• Published in: Analytical chemistry (Washington) 2024-09, Vol.96 (37), p.14980-14988
• Main Authors: Yu, Zi-Li, Wu, Zhou-Yang, Liu, Xing-Chi, Ji, Chang-Xin, Wang, Xuan, Fu, Qiu-Yun, Chen, Gang, Wu, Min, Hong, Shao-Li, Jia, Jun
• Format: Article
• Language: English
• Subjects: B7-H1 Antigen - analysis; B7-H1 Antigen - metabolism; Biomarkers; Biomarkers, Tumor - analysis; Biomarkers, Tumor - metabolism; Biopsy; Cancer; Cancer immunotherapy; CD45 antigen; Enzyme-linked immunosorbent assay; Epithelial Cell Adhesion Molecule - metabolism; Extracellular vesicles; Extracellular Vesicles - chemistry; Extracellular Vesicles - metabolism; Fluorescence; Humans; Immune system; Immunotherapy; Lab-On-A-Chip Devices; Microfluidics; Microspheres; Mouth Neoplasms - metabolism; Mouth Neoplasms - pathology; Mouth Neoplasms - therapy; Nanoparticles; Oral cancer; Oral carcinoma; Oral squamous cell carcinoma; PD-L1 protein; Saliva; Saliva - chemistry; Saliva - metabolism; Subpopulations; Tumor cells; Tumors; Vesicles
• ISSN: 0003-2700; 1520-6882; 1520-6882
• DOI: 10.1021/acs.analchem.4c03101

PD-L1-positive extracellular vesicles (PD-L1+ EVs) play a pivotal role as predictive biomarkers in cancer immunotherapy. These vesicles, originating from immune cells (I-PD-L1+ EVs) and tumor cells (T-PD-L1+ EVs), hold distinct clinical predictive values, emphasizing the importance of deeply differentiating the PD-L1+ EV subtypes for effective liquid biopsy analyses. However, current methods such as ELISA lack the ability to differentiate their cellular sources. In this study, a novel step-wedge microfluidic chip that combines magnetic microsphere separation with single-layer fluorescence counting is developed. This chip integrates magnetic microspheres modified with anti-PD-L1 antibodies and fluorescent nanoparticles targeting EpCAM (tumor cell marker) or CD45 (immunocyte marker), enabling simultaneous quantification and sensitive analysis of PD-L1+ EV subpopulations in oral squamous cell carcinoma (OSCC) patients’ saliva without background interference. Analysis results indicate reduced levels of I-PD-L1+ EVs in OSCC patients compared to those in healthy individuals, with varying levels of heterogeneous PD-L1+ EVs observed among different patient groups. During immunotherapy, responders exhibit decreased levels of total PD-L1+ EVs and T-PD-L1+ EVs, accompanied by reduced levels of I-PD-L1+ EVs. Conversely, nonresponders show increased levels of I-PD-L1+ EVs. Utilizing the step-wedge microfluidic chip allows for simultaneous detection of PD-L1+ EV subtypes, facilitating the precise prediction of oral cancer immunotherapy outcomes.