Engineering methods for loading therapeutic agents into exosomes. The techniques for loading cargo into exosomes can be divided into four approaches. (A) Using naïve exosomes (e.g., MSC-derived exosomes) requires relatively simple techniques to generate therapeutic exosomes, but the drawback is the difficulty in controlling the bioactive molecules contained in the exosomes. (B) Exogenous cargo loading is based on the use of sonication, repeated freeze/thaw cycles, or electroporation to destabilize the integrity of exosomal membranes and thus allow drugs to be introduced into the exosomes. (C, D) Endogenous cargo loading spontaneously loads molecules of interest by hijacking the natural exosome biogenesis pathway. These techniques are divided into two approaches based on whether the cargo is anchored onto the exosomal membrane (C) or resides as a free form inside the lumen of the exosome (D).
