Cellular Engineering with Membrane Fusogenic Liposomes to Produce Functionalized Extracellular Vesicles

Abstract

Engineering of extracellular vesicles (EVs) without affecting biological functions remains a challenge, limiting the broad applications of EVs in biomedicine. Here, we report a method to equip EVs with various functional agents, including fluorophores, drugs, lipids, and bio-orthogonal chemicals, in an efficient and controlled manner by engineering parental cells with membrane fusogenic liposomes, while keeping the EVs intact. As a demonstration of how this method can be applied, we prepared EVs containing azide-lipids, and conjugated them with targeting peptides using copper-free click chemistry to enhance targeting efficacy to cancer cells. We believe that this liposome-based cellular engineering method will find utility in studying the biological roles of EVs and delivering therapeutic agents through their innate pathway.

Therapeutic biomaterials based on extracellular vesicles: classification of bio‐engineering and mimetic preparation routes

“…Based on the study of exosome biogenesis-related mechanisms, several methods have been developed [50,57–62]. The following criteria have been identified for their classification (see Table 1):…”

“…A similar method was used to modify the composition of EVs, with a special focus on the modification of the properties of the EV membrane [57]. Dyes, fluorescent lipids with different lengths and saturation grade of acyl chains, and chemotherapeutics were loaded into cells by means of EVs.…”

Gesicle-Mediated Delivery of CRISPR/Cas9 Ribonucleoprotein Complex for Inactivating the HIV Provirus

“…EVs have become a topic of focus because of their implications as disease biomarkers in cancer and immunological disorders, 20,21 functions in normal physiology such as cell-to-cell communication, 22,23 and bioengineering-restorative capabilities. [24][25][26][27] EVs are endogenously produced membranous vessels that are heterogeneous in size, shape, and cargo. In general, EVs can be characterized into three subpopulations based on size and process of formation: microvesicles, exosomes, and apoptotic bodies (reviewed in Kalra et al, 28 Momen-Heravi et al, 29 and Akyurekli et al 30 ).…”

Microfluidic On-demand Engineering of Exosomes towards Cancer Immunotherapy

“…Bioengineered exosomes as emerging immunotherapeutics have gained substantial attention in developing a new generation of cancer vaccines, including recent phase-II trial using IFN-DC-derived exosomes loaded with MHC I/II restricted cancer antigens to promote T cell and natural killer (NK) cell-based immune responses in non-small cell lung cancer patients 21,[37][38][39][40][41] . Unfortunately, current exosome engineering approaches, such as the transfection or extrusion of parent cells and membrane permeabilization of secreted exosomes, suffer from poor yield, low purity, and time-consuming operations 38,[42][43][44][45] . Therefore, in this paper, we introduce a facile, 3D-printing molded PDMS microfluidic culture chip for solving this bottleneck problem.…”