PD-L1, also known as Programmed Death Ligand-1, is a protein that plays a crucial role in immune cell tolerance. It acts as an immune checkpoint ligand for the co-inhibitory receptor Programmed cell death protein 1 (PD-1), which is found on T-cells. However, when PD-L1 binds to PD-1, it inhibits the growth and proliferation of T-cells, ultimately promoting the growth of tumor cells. As a therapeutic approach, multiple anti-PD-L1 antibodies have been used to target the PD-L1 protein, thereby reducing immune cell tolerance. Notably, these tumor cells produce small extracellular vesicles (sEVs) that express PD-L1 on their surface, which further masks the anti-PD-L1 antibodies, leading to a reduction in their effectiveness. Therefore, a quantitative detection system is necessary to identify and capture PD-L1-positive sEVs. The primary objective of this project is to create an aptamer that specifically targets PD-L1-positive sEVs.
In this study, candidate aptamers obtained from SELEX were screened against recombinant PD-L1 protein. The most promising aptamers that bound to the recombinant PD-L1 protein were selected and characterized using ELASA and flow cytometry.
Through SELEX, twenty-nine candidate aptamers were obtained. Aptamers PD-L1-24-apt and PD-L1-29-apt showed promising binding to the recombinant PD-L1 protein. Using ELASA, the affinities of PD-L1-24-apt and PD-L1-29-apt were determined to be 310 nM and 286 nM against 90 nM of recombinant PD-L1 protein, respectively. Moreover, these aptamers have shown substantial selective binding to the native structure of PD-L1 protein anchored on the cell surface.
These aptamers will undergo further engineering and development to enhance their binding affinity and specificity. The final selected aptamers should exhibit a rapid on-rate and a slow off-rate, enabling the establishment of an aptamer-based capture system for PD-L1 positive sEVs through liquid biopsy. However, additional verification is needed to confirm the efficacy of these aptamers.