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      Progress, opportunity, and perspective on exosome isolation - efforts for efficient exosome-based theranostics

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          Abstract

          Exosomes are small extracellular vesicles with diameters of 30-150 nm. In both physiological and pathological conditions, nearly all types of cells can release exosomes, which play important roles in cell communication and epigenetic regulation by transporting crucial protein and genetic materials such as miRNA, mRNA, and DNA. Consequently, exosome-based disease diagnosis and therapeutic methods have been intensively investigated. However, as in any natural science field, the in-depth investigation of exosomes relies heavily on technological advances. Historically, the two main technical hindrances that have restricted the basic and applied researches of exosomes include, first, how to simplify the extraction and improve the yield of exosomes and, second, how to effectively distinguish exosomes from other extracellular vesicles, especially functional microvesicles. Over the past few decades, although a standardized exosome isolation method has still not become available, a number of techniques have been established through exploration of the biochemical and physicochemical features of exosomes. In this work, by comprehensively analyzing the progresses in exosome separation strategies, we provide a panoramic view of current exosome isolation techniques, providing perspectives toward the development of novel approaches for high-efficient exosome isolation from various types of biological matrices. In addition, from the perspective of exosome-based diagnosis and therapeutics, we emphasize the issue of quantitative exosome and microvesicle separation.

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          Extracellular Vesicle Heterogeneity: Subpopulations, Isolation Techniques, and Diverse Functions in Cancer Progression

          Cells release membrane enclosed nano-sized vesicles termed extracellular vesicles (EVs) that function as mediators of intercellular communication by transferring biological information between cells. Tumor-derived EVs have emerged as important mediators in cancer development and progression, mainly through transfer of their bioactive content which can include oncoproteins, oncogenes, chemokine receptors, as well as soluble factors, transcripts of proteins and miRNAs involved in angiogenesis or inflammation. This transfer has been shown to influence the metastatic behavior of primary tumors. Moreover, tumor-derived EVs have been shown to influence distant cellular niches, establishing favorable microenvironments that support growth of disseminated cancer cells upon their arrival at these pre-metastatic niches. It is generally accepted that cells release a number of major EV populations with distinct biophysical properties and biological functions. Exosomes, microvesicles, and apoptotic bodies are EV populations most widely studied and characterized. They are discriminated based primarily on their intracellular origin. However, increasing evidence suggests that even within these EV populations various subpopulations may exist. This heterogeneity introduces an extra level of complexity in the study of EV biology and function. For example, EV subpopulations could have unique roles in the intricate biological processes underlying cancer biology. Here, we discuss current knowledge regarding the role of subpopulations of EVs in cancer development and progression and highlight the relevance of EV heterogeneity. The position of tetraspanins and integrins therein will be highlighted. Since addressing EV heterogeneity has become essential for the EV field, current and novel techniques for isolating EV subpopulations will also be discussed. Further dissection of EV heterogeneity will advance our understanding of the critical roles of EVs in health and disease.
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            A phase I study of dexosome immunotherapy in patients with advanced non-small cell lung cancer

            Background There is a continued need to develop more effective cancer immunotherapy strategies. Exosomes, cell-derived lipid vesicles that express high levels of a narrow spectrum of cell proteins represent a novel platform for delivering high levels of antigen in conjunction with costimulatory molecules. We performed this study to test the safety, feasibility and efficacy of autologous dendritic cell (DC)-derived exosomes (DEX) loaded with the MAGE tumor antigens in patients with non-small cell lung cancer (NSCLC). Methods This Phase I study enrolled HLA A2+ patients with pre-treated Stage IIIb (N = 4) and IV (N = 9) NSCLC with tumor expression of MAGE-A3 or A4. Patients underwent leukapheresis to generate DC from which DEX were produced and loaded with MAGE-A3, -A4, -A10, and MAGE-3DPO4 peptides. Patients received 4 doses of DEX at weekly intervals. Results Thirteen patients were enrolled and 9 completed therapy. Three formulations of DEX were evaluated; all were well tolerated with only grade 1–2 adverse events related to the use of DEX (injection site reactions (N = 8), flu like illness (N = 1), and peripheral arm pain (N = 1)). The time from the first dose of DEX until disease progression was 30 to 429+ days. Three patients had disease progression before the first DEX dose. Survival of patients after the first DEX dose was 52–665+ days. DTH reactivity against MAGE peptides was detected in 3/9 patients. Immune responses were detected in patients as follows: MAGE-specific T cell responses in 1/3, increased NK lytic activity in 2/4. Conclusion Production of the DEX vaccine was feasible and DEX therapy was well tolerated in patients with advanced NSCLC. Some patients experienced long term stability of disease and activation of immune effectors
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              Characterization and comprehensive proteome profiling of exosomes secreted by hepatocytes.

              Exosomes represent a discrete population of vesicles that are secreted from various cell types to the extracellular media. Their protein and lipid composition are a consequence of sorting events at the level of the multivesicular body, a central organelle which integrates endocytic and secretory pathways. Characterization of exosomes from different biological samples has shown the presence of common as well as cell-type specific proteins. Remarkably, the protein content of the exosomes is modified upon pathological or stress conditions. Hepatocytes play a central role in the body response to stress metabolizing potentially harmful endogenous substances as well as xenobiotics. In the present study, we described and characterized for the first time exosome secretion in nontumoral hepatocytes, and with the use of a systematic proteomic approach, we establish the first extensive proteome of a hepatocyte-derived exosome population which should be useful in furthering our understanding of the hepatic function and in the identification of components that may serve as biomarkers for hepatic alterations. Our analysis identifies a significant number of proteins previously described among exosomes derived from others cell types as well as proteins involved in metabolizing lipoproteins, endogenous compounds and xenobiotics, not previously described in exosomes. Furthermore, we demonstrated that exosomal membrane proteins can constitute an interesting tool to express nonexosomal proteins into exosomes with therapeutic purposes.
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                Author and article information

                Journal
                Theranostics
                Theranostics
                thno
                Theranostics
                Ivyspring International Publisher (Sydney )
                1838-7640
                2020
                19 February 2020
                : 10
                : 8
                : 3684-3707
                Affiliations
                [1 ]Department of Neurosurgery of Hebi People's Hospital; Hebi Neuroanatomical Laboratory, Hebi, 458030, China.
                [2 ]School of Nursing, Zhengzhou University, Zhengzhou, 450001, China.
                [3 ]Henan Key Laboratory of Ion-beam Bioengineering, Zhengzhou University, Zhengzhou, China, 450000.
                [4 ]Guangdong Key Laboratory for Research and Development of Nature Drugs, School of Pharmacy, Guangdong Medical University, Zhanjiang 524023, China.
                [5 ]School of Statistics, Henan University of Economics and Law, Zhengzhou 450046, China.
                [6 ]Centre for Molecular Medicine and Innovative Therapeutics, Murdoch University, Perth 6150, Australia.
                [7 ]General Practice Centre, Nanhai Hospital, Southern Medical University, 528244, Foshan, China.
                [8 ]School of Medicine, Wake Forest University, Winston Salem, NC 27101, USA.
                [9 ]School of Medicine, and Centre for Molecular and Medical Research, Deakin University, 3216, Australia.
                [10 ]The First Affiliated Hospital of Zheng Zhou University, Zhengzhou 450001, China.
                [11 ]Perron Institute for Neurological and Translational Science, Perth 6009, Australia
                Author notes
                ✉ Corresponding author: Tao Wang, wangtaomary@ 123456zzu.edu.cn ; Rakesh N Veedu, R.Veedu@ 123456murdoch.edu.au ; Changying Chen, changying@ 123456zzu.edu.cn .

                #These authors contribute equally to this work.

                Competing Interests: The authors have declared that no competing interest exists.

                Article
                thnov10p3684
                10.7150/thno.41580
                7069071
                32206116
                3016f58d-b8cf-428b-842d-ed43b5b2dd9d
                © The author(s)

                This is an open access article distributed under the terms of the Creative Commons Attribution License ( https://creativecommons.org/licenses/by/4.0/). See http://ivyspring.com/terms for full terms and conditions.

                History
                : 29 October 2019
                : 8 February 2020
                Categories
                Review

                Molecular medicine
                exosome,microvesicle,extracellular vesicle,microfluidic,diagnosis,separation
                Molecular medicine
                exosome, microvesicle, extracellular vesicle, microfluidic, diagnosis, separation

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