In vitrotoxicology studies of extracellular vesicles

Abstract

Extracellular vesicles (EVs) are membrane-bound vesicles released from cells into the extracellular environment. There is emerging interest in the use of EVs as potential therapeutic interventions. We sought to evaluate the safety of EVs that may be therapeutically used by performing in vitro toxicological assessments. EVs were obtained from mesenchymal stem cells (MSC-EV) or from bovine milk (BM-EV) by differential ultracentrifugation, and quantitated using nanoparticle tracking analysis. Genotoxic effects, hematological effects, immunological effects and endotoxin production were evaluated at two dose levels. Neither MSC-EVs nor BM-EVs elicited detectable genotoxic effects using either the alkaline comet assay or micronucleus assay. Hemolysis was observed with BM-EVs but not with MSC-EVs. MSC-EVs did not have any significant effect on either spontaneous or collagen-induced platelet aggregation. In contrast, BM-EVs were noted to increase collagen-induced platelet aggregation, even though no spontaneous increase in platelet aggregation was noted. Both types of EVs induced leukocyte proliferation, which was greater with BM-EV. Neither MSC-EVs nor BM-EVs induced HL-60 phagocytosis, although BM-EVs decreased zymosan-induced phagocytosis. Furthermore, neither MSC-EVs nor BM-EVs induced nitric oxide production. Unlike MSC-EVs, BM-EVs tested positive for endotoxin and induced complement activation. There are significant differences in toxicological profiles between MSC-EVs and BM-EVs that may reflect variations in techniques for EV isolation, EV content or cross-species differences. The safety of MSC-EV supports their use for disease therapeutics, whereas detailed safety and toxicological assessment will be necessary before the use of BM-EVs. Copyright © 2016 John Wiley & Sons, Ltd.

Extracellular Vesicle‐Based Therapeutic Targeting of β‐Catenin to Modulate Anticancer Immune Responses in Hepatocellular Cancer

“…For these studies, we developed a therapeutic approach involving hepatic delivery of siRNA using EVs as a biological nanoparticle delivery vehicle. We have established scalable procedures for production of EVs and have demonstrated the safety profile of these MNVs in vitro . Proof‐of‐principle studies have demonstrated the ability to generate therapeutic EVs (tMNVs) by loading these nanovesicles with siRNA and for these tMNVs to deliver their cargo into liver cells in vitro and in vivo .…”

“…The benefits of targeting the immune system for cancer therapy are being increasingly recognized. Immunotherapy with checkpoint inhibitors targeting anti‐programmed death 1 (anti‐PD‐1), anti‐programmed death ligand 1 (anti‐PD‐L1), and cytotoxic T lymphocyte antigen 4 (CTLA4) has resulted in durable responses and is currently approved for use in several types of aggressive cancers . The PD‐1/PD‐L1 interaction enables tumor cells to escape from the attack of cytotoxic T cells .…”

Safety of bovine milk derived extracellular vesicles used for delivery of RNA therapeutics in zebrafish and mice

“…In recent preclinical studies we have demonstrated the feasibility of using EV isolated from milk, or milk‐derived nanovesicles (MNV), for therapeutic applications as drug delivery vehicles (George, Yan, & Patel, ; Matsuda, Ishiguro, Yan, & Patel, ). Systematic toxicological studies of MNV in vitro have revealed a favorable safety profile (Maji et al, ). However, prior to their clinical adoption, there is an essential need to demonstrate their safety and toxicology in vivo.…”

Mesenchymal stem cell derived extracellular vesicles: a promising new therapeutic approach for hepatic injury

“…Recent studies have highlighted the involvement of EV, endogenous membrane-bound cellular nanoparticles released from cells, as a mediator of intercellular communication, and in maintenance of tissue homeostasis. EV released from cells can be taken up by other cells, with transfer of their cargo which can include proteins, lipids, RNA or DNA (2). These EV can include several different types of particles such as exosomes, microvesicles, apoptotic bodies and others.…”

“…EV have been postulated to contribute to many physiological and pathological processes such as cancer progression, immune modulation, neurodegenerative diseases, or tissue regeneration (1,2,7,8). EV released from stem cells have been shown to modulate angiogenesis, prevent apoptosis or promote cell proliferation.…”