Extracellular Vesicles as Biological Shuttles for Targeted Therapies.

The development of effective nanosystems for drug delivery represents a key challenge for the improvement of most current anticancer therapies. Recent progress in the understanding of structure and function of extracellular vesicles (EVs)-specialized membrane-bound nanocarriers for intercellular communication-suggests that they might also serve as optimal delivery systems of therapeutics. In addition to carrying proteins, lipids, DNA and different forms of RNAs, EVs can be engineered to deliver specific bioactive molecules to target cells. Exploitation of their molecular composition and physical properties, together with improvement in bio-techniques to modify their content are critical issues to target them to specific cells/tissues/organs. Here, we will discuss the current developments in the field of animal and plant-derived EVs toward their potential use for delivery of therapeutic agents in different pathological conditions, with a special focus on cancer

Cancer relevance of signal recognition particle RNA and other non-coding RNAs in extracellular vesicles

[No abstract available

Extracellular Vesicles from Thyroid Carcinoma: The New Frontier of Liquid Biopsy

The diagnostic approach to thyroid cancer is one of the most challenging issues in oncology of the endocrine system because of its high incidence (3.8% of all new cancer cases in the US) and the difficulty to distinguish benign from malignant non-functional thyroid nodules and establish the cervical lymph node involvement during staging. Routine diagnosis of thyroid nodules usually relies on a fine-needle aspirate biopsy, which is invasive and often inaccurate. Therefore, there is an urgent need to identify novel, accurate, and non-invasive diagnostic procedures. Liquid biopsy, as a non-invasive approach for the detection of diagnostic biomarkers for early tumor diagnosis, prognosis, and disease monitoring, may be of particular benefit in this context. Extracellular vesicles (EVs) are a consistent source of tumor-derived RNA due to their prevalence in circulating bodily fluids, the well-established isolation protocols, and the fact that RNA in phospholipid bilayer-enclosed vesicles is protected from blood-borne RNases. Recent results in other types of cancer, including our recent study on plasma EVs from glioblastoma patients suggest that information derived from analysis of EVs from peripheral blood plasma can be integrated in the routine diagnostic tumor approach. In this review, we will examine the diagnostic and prognostic potential of liquid biopsy to detect tumor-derived nucleic acids in circulating EVs from patients with thyroid carcinoma

Anti-Human CD9 Antibody Fab Fragment Impairs the Internalization of Extracellular Vesicles and the Nuclear Transfer of their Cargo Proteins.

The intercellular communication mediated by extracellular vesicles (EVs) has gained international interest during the last decade. Interfering with the mechanisms regulating this cellular process might find application particularly in oncology where cancer cell-derived EVs play a role in tumour microenvironment transformation. Although several mechanisms were ascribed to explain the internalization of EVs, little is our knowledge about the fate of their cargos, which are crucial to mediate their function. We recently demonstrated a new intracellular pathway in which a fraction of endocytosed EV-associated proteins is transported into the nucleoplasm of the host cell via a subpopulation of late endosomes penetrating into the nucleoplasmic reticulum. Silencing tetraspanin CD9 both in EVs and recipient cells strongly decreased the endocytosis of EVs and abolished the nuclear transfer of their cargos. Here, we investigated whether monovalent Fab fragments derived from 5H9 anti-CD9 monoclonal antibody (referred hereafter as CD9 Fab) interfered with these cellular processes. To monitor the intracellular transport of proteins, we used fluorescent EVs containing CD9-green fluorescent protein fusion protein and various melanoma cell lines and bone marrow-derived mesenchymal stromal cells as recipient cells. Interestingly, CD9 Fab considerably reduced EV uptake and the nuclear transfer of their proteins in all examined cells. In contrast, the divalent CD9 antibody stimulated both events. By impeding intercellular communication in the tumour microenvironment, CD9 Fab-mediated inhibition of EV uptake, combined with direct targeting of cancerous cells could lead to the development of novel anti-melanoma therapeutic strategies

Breast Cancer-Derived Extracellular Vesicles: Characterization and Contribution to the Metastatic Phenotype

The study of extracellular vesicles (EVs) in cancer progression is a complex and rapidly evolving field. Whole categories of cellular interactions in cancer which were originally presumed to be due solely to soluble secreted molecules have now evolved to include membrane-enclosed extracellular vesicles (EVs), which include both exosomes and shed microvesicles (MVs), and can contain many of the same molecules as those secreted in soluble form but many different molecules as well. EVs released by cancer cells can transfer mRNA, miRNA, and proteins to different recipient cells within the tumor microenvironment, in both an autocrine and paracrine manner, causing a significant impact on signaling pathways, mRNA transcription, and protein expression. The transfer of EVs to target cells, in turn, supports cancer growth, immunosuppression, and metastasis formation. This review focuses exclusively on breast cancer EVs with an emphasis on breast cancer-derived exosomes, keeping in mind that breast cancer-derived EVs share some common physical properties with EVs of other cancers

Breast Cancer-Derived Extracellular Vesicles: Characterization and Contribution to the Metastatic Phenotype

The study of extracellular vesicles (EVs) in cancer progression is a complex and rapidly evolving field. Whole categories of cellular interactions in cancer which were originally presumed to be due solely to soluble secreted molecules have now evolved to include membrane-enclosed extracellular vesicles (EVs), which include both exosomes and shed microvesicles (MVs), and can contain many of the same molecules as those secreted in soluble form but many different molecules as well. EVs released by cancer cells can transfer mRNA, miRNA, and proteins to different recipient cells within the tumor microenvironment, in both an autocrine and paracrine manner, causing a significant impact on signaling pathways, mRNA transcription, and protein expression. The transfer of EVs to target cells, in turn, supports cancer growth, immunosuppression, and metastasis formation. This review focuses exclusively on breast cancer EVs with an emphasis on breast cancer-derived exosomes, keeping in mind that breast cancer-derived EVs share some common physical properties with EVs of other cancers

HIV-1-induced nuclear invaginations mediated by VAP-A, ORP3, and Rab7 complex explain infection of activated T cells

The mechanism of human immunodeficiency virus 1 (HIV-1) nuclear entry, required for productive infection, is not fully understood. Here, we report that in HeLa cells and activated CD4+ T cells infected with HIV-1 pseudotyped with VSV-G and native Env protein, respectively, Rab7+ late endosomes containing endocytosed HIV-1 promote the formation of nuclear envelope invaginations (NEIs) by a molecular mechanism involving the VOR complex, composed of the outer nuclear membrane protein VAP-A, hyperphosphorylated ORP3 and Rab7. Silencing VAP-A or ORP3 and drug-mediated impairment of Rab7 binding to ORP3-VAP-A inhibited the nuclear transfer of the HIV-1 components and productive infection. In HIV-1-resistant quiescent CD4+ T cells, ORP3 was not hyperphosphorylated and neither VOR complex nor NEIs were formed. This new cellular pathway and its molecular players are potential therapeutic targets, perhaps shared by other viruses that require nuclear entry to complete their life cycle

Extracellular vesicle-mediated transfer of CLIC1 protein is a novel mechanism for the regulation of glioblastoma growth

Little progresses have been made in the treatment of glioblastoma (GBM), the most aggressive and lethal among brain tumors. Recently we have demonstrated that Chloride Intracellular Channel-1 (CLIC1) is overexpressed in GBM compared to normal tissues, with highest expression in patients with poor prognosis. Moreover, CLIC1-silencing in cancer stem cells (CSCs) isolated from human GBM patients negatively influences proliferative capacity and self-renewal properties in vitro and impairs the in vivo tumorigenic potential. Here we show that CLIC1 exists also as a circulating protein, secreted via extracellular vesicles (EVs) released by either cell lines or GBM-derived CSCs. Extracellular vesicles (EVs), comprising exosomes and microvesicles based on their composition and biophysical properties, have been shown to sustain tumor growth in a variety of model systems, including GBM. Interestingly, treatment of GBM cells with CLIC1-containing EVs stimulates cell growth both in vitro and in vivo in a CLIC1-dose dependent manner. EVs derived from CLIC1-overexpressing GBM cells are strong inducers of proliferation in vitro and tumor engraftment in vivo. These stimulations are significantly attenuated by treatment of GBM cells with EVs derived from CLIC1-silenced cells. However, CLIC1 modulation appears to have no direct role in EV structure, biogenesis and secretion. These findings reveal that, apart from the function of CLIC1 cellular reservoir, CLIC1 contained in EVs is a novel regulator of GBM growth

Itraconazole inhibits nuclear delivery of extracellular vesicle cargo by disrupting the entry of late endosomes into the nucleoplasmic reticulum

Extracellular vesicles (EVs) are mediators of intercellular communication under bothhealthy and pathological conditions, including the induction of pro-metastatic traits,but it is not yet known how and where functional cargoes of EVs are delivered to theirtargets in host cell compartments. We have described that after endocytosis, EVsreach Rab+late endosomes and a fraction of these enter the nucleoplasmic reticu-lum and transport EV biomaterials to the host cell nucleoplasm. Their entry thereinand docking to outer nuclear membrane occur through a tripartite complex formedby the proteins VAP-A, ORP and Rab (VOR complex). Here, we report that theantifungal compound itraconazole (ICZ), but not its main metabolite hydroxy-ICZor ketoconazole, disrupts the binding of Rab to ORP–VAP-A complexes, leadingto inhibition of EV-mediated pro-metastatic morphological changes including cellmigration behaviour of colon cancer cells. With novel, smaller chemical drugs, inhi-bition of the VOR complex was maintained, although the ICZ moieties responsiblefor antifungal activity and interference with intracellular cholesterol distributionwere removed. Knowing that cancer cells hijack their microenvironment and thatEVs derived from them determine the pre-metastatic niche, small-sized inhibitors ofnuclear transfer of EV cargo into host cells could nd cancer therapeutic applications,particularly in combination with direct targeting of cancer cell