The distinct roles of exosomes in innate immune responses and therapeutic applications in cancer.

The innate immune system is one of the major constituents of the host's defense against invading pathogens and extracellular vesicles (EVs) are involved in regulating its responses. Exosomes, a subclass of EVs, released from eukaryotic cells, contribute to intracellular communication and drive various biological processes by transferring nuclei acids, proteins, lipids, and carbohydrates between cells, protecting cargo from enzymatic degradation and immune recognition and consequent elimination by the immune system. A growing body of evidence has revealed that exosomes produced from host cells, infected cells, tumor cells, and immune cells regulate innate immune signaling and responses and thus play a significant role in the propagation of pathogens. Immune cells can recognize exosomes-bearing components including DNA strands, viral RNAs, and even proteins by various mechanisms such as through Toll-like receptor/NF-κB signaling, inducing cytokine production and reprogramming the innate immune responses, immunosuppression or immunesupportive. There is persuasive preclinical and clinical evidence that exosomes are therapeutic strategies for immunotherapy, cancer vaccine, drug-delivery system, and diagnostic biomarker. However, further scrutiny is essential to validate these findings. In this review, we describe the current facts on the regulation of innate immune responses by exosomes. We also describe the translational application of exosomes as cancer-therapy agents and immunotherapy.

A review on exosomes application in clinical trials: perspective, questions, and challenges.

BACKGROUND: Exosomes are progressively known as significant mediators of cell-to-cell communication. They convey active biomolecules to target cells and have vital functions in several physiological and pathological processes, and show substantial promise as novel treatment strategies for diseases. METHODS: In this review study, we studied numerous articles over the past two decades published on application of exosomes in different diseases as well as on perspective and challenges in this field. RESULTS: The main clinical application of exosomes are using them as a biomarker, cell-free therapeutic agents, drug delivery carriers, basic analysis for exosome kinetics, and cancer vaccine. Different exosomes from human or plant sources are utilized in various clinical trials. Most researchers used exosomes from the circulatory system for biomarker experiments. Mesenchymal stem cells (MSCs) and dendritic cells (DCs) are two widely held cell sources for exosome use. MSCs-derived exosomes are commonly used for inflammation treatment and drug delivery, while DCs-exosomes are used to induce inflammation response in cancer patients. However, the clinical application of exosomes faces various questions and challenges. In addition, translation of exosome-based clinical trials is required to conform to specific good manufacturing practices (GMP). In this review, we summarize exosomes in the clinical trials according to the type of application and disease. We also address the main questions and challenges regarding exosome kinetics and clinical applications. CONCLUSIONS: Exosomes are promising platforms for treatment of many diseases in clinical trials. This exciting field is developing hastily, understanding of the underlying mechanisms that direct the various observed roles of exosomes remains far from complete and needs further multidisciplinary research in working with these small vesicles. Video Abstract.

Inhibitory effects of gallic acid on the activity of exosomal secretory pathway in breast cancer cell lines: A possible anticancer impact.

Introduction: Breast cancer cells produce exosomes that promote tumorigenesis. The anticancer properties of gallic acid have been reported. However, the mechanism underlying its anticancer effect on the exosomal secretory pathway is still unclear. We investigated the effect of gallic acid on exosome biogenesis in breast cancer cell lines. Methods: The cytotoxic effect of gallic acid on MCF-10a, MCF-7, and MDA-MD-231 cells was measured by MTT assay after 48 hours treatment. Expression of miRNAs including miRNA-21, -155, and 182 as well as exosomal genes such as Rab27a, b, Rab11, Alix, and CD63; along with HSP-70 (autophagy gene), was determined using Q-PCR. The subcellular distribution of it was monitored by flow cytometry analysis. Isolated exosomes were characterized by transmission and scanning electron microscopes and flow cytometry. Acetylcholinesterase activity is used to measure the number of exosomes in supernatants. In addition, autophagy markers including LC3 and P62 were measured by ELISA. Results: Data showed that gallic acid was cytotoxic to cells (P < 0.05). Gallic acid modulated expression of miRNAs and down-regulated transcript levels of exosomal genes and up-regulated the HSP-70 gene in three cell lines (P < 0.05). The surface CD63/total CD63 ratio as well as acetylcholinesterase activity decreased in treated cells (P < 0.05). The protein level of LC3 was increased in three cell lines, while the expression of P62 increased in MCF-7 and MDA-MB-231 cancer cell lines. Conclusion: Together, gallic acid decreased the activity of the exosomal secretory pathway in breast cancer cell lines, providing evidence for its anti-cancer effects.

Bystander effects induced by electron beam-irradiated MCF-7 cells: a potential mechanism of therapy resistance.

PURPOSE: The distinct direct and non-targeting effects of electron beam radiation on MCF-7 cells remain obscure. We aimed to investigate the effect of electron beam irradiation (EBI) and conditioned media (CM) of the irradiated MCF-7 cells on MCF-7 cells. The cytotoxic effects of CM from irradiated MCF-7 cells on the mesenchymal stem cells and human umbilical vein endothelial cells (HUVECs) were also examined. METHODS: Cell viability and apoptosis were assayed via MTT and flow cytometry analysis, respectively. The production of ROS (reactive oxygen species) was evaluated by the chemical fluorometric method, while the amount of extracellular vesicles was detected via acetylcholinesterase activity assay. Expression of genes involved in apoptosis, including caspase-3, -8, -9, and stemness such as Sox-2 and Oct-4, were calculated through qPCR. The wound healing rate of cells was monitored via in vitro scratch assay. RESULTS: Compared to the control group, EBI groups showed decreased cell viability but increased apoptosis and ROS as well as acetylcholinesterase activity dose-dependently (P < 0.05). Concurrently with increasing the dose of the electron beam, the transcript levels of apoptotic genes (caspase-3, -8, -9) and stemness-related genes (Sox-2 and Oct-4) were up-regulated following EBI. The wound healing rate of irradiated MCF-7 cells increased dose-dependently (P < 0.05). Similar results were observed after treatment with CM from irradiated MCF-7 cells. Additionally, CM from irradiated MCF-7 cells decreased the viability of MCF-7 cells, mesenchymal stem cells, and HUVECs (P < 0.05). CONCLUSION: MCF-7 cells treated with an electron beam and CMs from irradiated MCF-7 cells exhibit an up-regulation in both genes involved in the apoptosis pathway and stemness. As a result, EBI can affect apoptosis and stemness in MCF-7 cells in direct and bystander manners. However, specific signaling pathways require careful evaluation to provide an understanding of the mechanisms involved in the EBI-induced alternation in tumor cell dynamics.

Tumor-derived extracellular vesicles: reliable tools for Cancer diagnosis and clinical applications.

BACKGROUND: Studies have recently revealed that almost every type of cells including tumor cells abundantly release small vesicles known as extracellular vesicles (EVs) into the extracellular milieu. EVs carry a repertoire of biological molecules including nucleic acids, proteins, lipids, and carbohydrates and transport their cargo between cells in the vicinity as well as distantly located cells and hence act as messengers of intercellular communication. In this review, we aimed to discuss the tumor-derived exosome biology and the pivotal roles of exosomes in cancer diagnosis and treatment. METHODS: In the present review study, the authors studied several articles over the past two decades published on the kinetics of EVs in tumor environment as well as on the application of these vesicles in cancer diagnosis and therapy. RESULTS: A growing body of evidence indicates that nucleic acids such as microRNAs (miRNAs) transferring by EVs participate to create a conducive tumor environment. As EV-associated miRNAs are tissue-specific and present in most biological fluids, they hold great potential for clinical application in cancer early diagnosis, prognosis, and treatment response. Furthermore, exosomes can serve as drug delivery vehicles transferring miRNAs as well as therapeutic agents to target cells. These nano-vesicles exhibit ideal properties in comparison with the synthetic carriers that attracted scientist's attention in the field of nanotechnology medicine. Scientists have employed different strategies to build exosomes-based drug delivery system. In general, two methods (direct engineering and indirect engineering) are being utilized to produce artificial exosomes. Para-clinical data have confirmed the beneficial effects of engineering exosomes in cancer therapy. CONCLUSION: Exosomal miRNAs hold great promise for clinical application in early diagnosis and treatment of cancers. In addition, in spite of enthusiastic results obtained by engineered exosomes, however, there is an increasing concern over the use of optimal methods for engineering exosomes and the safety of engineered exosomes in clinical trials is still unclear.