Systematic Evolution of Ligands by Exosome Enrichment: A Proof‐of‐Concept Study for Exosome‐Based Targeting Peptide Screening

Selection of a peptide that binds preferentially to targeted cells or tissues is a prerequisite for targeted therapy. Although in vivo phage display is a high-throughput method, it is restricted in identifying target ligands specific for different vascular beds. In this study, the exosomes are repurposed for targeting peptide screening. Briefly, the signal peptide region of Lamp2b (a membrane protein on the exosomes) in the N-terminus is engineered to fuse with 10 aa long random peptides, while the C-terminus of Lamp2b is fused with the MS2 coating protein (MCP). Then, the whole Lamp2b-MCP open reading frame (ORF) is further engineered to harbor a 3'UTR sequence consisting of MS2. The resultant exosomes from engineered Lamp2b-MCP expressing cells display the 10 aa peptides on the outside while containing the genetic information inside. By proof-of-principle experiments, the exosomes with different peptides could preferentially distribute to different tissues besides the spleen and liver. Furthermore, detailed target sequences for different tissues are enriched by rounds of selection. In summary, the established novel targeted peptide screening strategy, namely, "exosome display," has broad applicability, especially for displaying and screening targeted peptides for the cells outside the capillary with condense barriers, like the neurons in the brain.