Extracellular vesicles (EVs) are membrane-delimitated sub-cellular entities released by cells in a constitutive manner or in response to stress. EVs contain membrane proteins and lipids as well as cytoplasm components in a pattern which depends on the type of stimulation and physiopathology of parental cells. EVs constitute a far-reaching intercellular communication pathway controlling cell signaling. Such unique feature stems from their capacity to shelter their internal cargo (proteins and genomic material) from the harsh extracellular environment and from their extraordinary ability to transfer material between neighbor and distal cells.
EVs have the plasticity to be engineered to transport drugs and therapeutic nanoparticles. We were the first to demonstrate that exogenous material such as iron oxide nanoparticles (Small, 2008; Biomaterials, 2010) and carbon nanotubes (Nano Letters, 2012) may exit the cell via the extracellular vesicle pathway, being able to transfer the nanoparticle cargo in a homotypic as hetorotypic manner (Pharmaceutical Research, 2012). Notably, we took advantage of this cell detoxification strategy to design vesicles loaded with drugs and nanoparticles. Crossing the boundaries of materials science and cell biology, our aim was to translate such cell communication effectors into an intrinsically biocompatible bio-inspired vector. We showed the feasibility of encapsulating different drug molecules into vesicles regardless their molecular weight, hydrophobic, hydrophilic and amphiphilic character (Nanomedicine, 2015). We equally demonstrated that such unique biogenic nanoplatform was able to encapsulate a set of nanoparticles, regardless their chemistry or shape (Nanoscale, 2013) (Figure 1). Therefore, we could design of different hybrid EVs: magnetic-fluorescent and magnetic-metallic vesicles, either single component or multicomponent, combining the advantageous properties of each constituent. These vesicles delivered thermal therapy as they heated when submitted to an alternating magnetic field and could be monitored using fluorescent imaging (Nanoscale, 2013) (Figure 2).
Figure 1 : Engineering EVs to encapsulate nanoparticles. (A) Transmission electron microscopy images of iron oxide spherical nanoparticles (IONP) and nanocubes (IONC), quantum dots (QD), gold-based nanoparticles (AuNP) and gold-iron oxide nanodimers (Au/IONP). (B) Schematic representation of cell-derived microvesicles enclosing such nanoparticles. (C) Transmission electron micrographs of a series of vesicles released from precursor cells loaded with IONP; IONP + QD; IONP + AuNP; IONC or Au/IONP (Silva et al, Nanoscale. 2013).
Figure 2 : Magnetic nanoparticle encapsulation into EVs confer them heating properties and magnetophoresis. (A) Micromagnetophoresis experiment under fluorescence microscopy of macrophage vesicles conjugated with a green fluorophore and encapsulating iron oxide nanoparticles moving toward a micromagnet tip. (B) Temperature rise as a function of the time for Huvec vesicles loaded with iron oxide nanoparticles at 320, 600 and 900 kHz (Silva et al, Nanoscale. 2013 and Silva et al Nanomedicine 2012).
Engineering of endothelial-cell derived microvesicles with magnetic nanoparticles enabled in vivo MRI imaging of vesicle biodistribution (Radiology, 2012). We also provided evidence of the therapeutic and imaging functions of these hybrid vesicles both in vitro and in vivo in tumor-bearing mice (ACS Nano, 2013) (Figures 2-3).
Figure 3 . Schematic representation of theranosome production from drug-loaded magnetic precursor cells and their application for photodynamic therapy and dual-mode MRI and fluorescence imaging (ACS Nano, 2013).
Figure 4 : Theranosome dual-mode imaging in vitro and in vivo and theranosome-mediated therapeutic effect in vivo. (a) False-color images of fluorescence emission from nude mice bearing TC-1 tumor before and after theranosome or m-THPC intratumoral injection, as a function of time. (b) MRI scans of TC-1 tumors 48 h after PBS intra-tumoral injection (control group) or 48 h after theranosome intra-tumoral injection. (c) Tumor growth curves (tumor volume is normalized to day 0) for mice intra-tumorally injected with: theranosomes (25 µM m-THPC concentration) or m-THPC (25 µM) both followed by light exposure (λ=630nm - 30J/cm²: 77mW/cm² for 390 s) 20 h later, compared with untreated control (ACS Nano, 2013).
We are currently interested in new high-throughput scalable methods to produce and load EVs and the validation of the produced vesicles as drug/nanoparticle biocamouflaged carriers in orthotopic animal models.