Nanoparticle-mediated delivery of functional macromolecules is a promising method for treating a variety of human diseases. Among nanoparticles, cell-derived exosomes have recently been highlighted as a new therapeutic strategy for the in vivo delivery of nucleotides and chemical drugs.
Here, KAIST researchers describe a new tool for intracellular delivery of target proteins, named ‘exosomes for protein loading via optically reversible protein-protein interactions’ (EXPLORs). By integrating a reversible protein-protein interaction module controlled by blue light with the endogenous process of exosome biogenesis, they are able to successfully load cargo proteins into newly generated exosomes. Treatment with protein-loaded EXPLORs is shown to significantly increase intracellular levels of cargo proteins and their function in recipient cells in vitro and in vivo. These results clearly indicate the potential of EXPLORs as a mechanism for the efficient intracellular transfer of protein-based therapeutics into recipient cells and tissues.
Schematic diagram of EXPLOR technology
In EXPLOR-producing donor cells, CRY2 protein was fused to a cargo protein, and CIBN was conjugated with a representative marker of exosomes, CD9 protein. Blue light illumination induces the reversible PPI between CIBN and CRY2 fusion proteins. With continuous blue light irradiation, the cargo proteins are guided to the inner surface of the cell membrane or the surface of early endosomes. Mature multi-vesicular bodies (MVBs) then readily secrete cargo protein-carrying exosomes (EXPLORs) from the cells by membrane fusion with the plasma membrane. After exocytosis, EXPLORs can be easily isolated and purified in vitro. Purified EXPLORs can be used for delivery of the cargo proteins into target cells via membrane fusion or endocytosis processes. Bottom grey boxes highlight the essential steps from EXPLORs biogenesis to target cell delivery.