Isolation and characterization of exosomes for cancer research

Abstract

Exosomes are a subset of extracellular vesicles that carry specific combinations of proteins, nucleic acids, metabolites, and lipids. Mounting evidence suggests that exosomes participate in intercellular communication and act as important molecular vehicles in the regulation of numerous physiological and pathological processes, including cancer development. Exosomes are released by various cell types under both normal and pathological conditions, and they can be found in multiple bodily fluids. Moreover, exosomes carrying a wide variety of important macromolecules provide a window into altered cellular or tissue states. Their presence in biological fluids renders them an attractive, minimally invasive approach for liquid biopsies with potential biomarkers for cancer diagnosis, prediction, and surveillance. Due to their biocompatibility and low immunogenicity and cytotoxicity, exosomes have potential clinical applications in the development of innovative therapeutic approaches. Here, we summarize recent advances in various technologies for exosome isolation for cancer research. We outline the functions of exosomes in regulating tumor metastasis, drug resistance, and immune modulation in the context of cancer development. Finally, we discuss prospects and challenges for the clinical development of exosome-based liquid biopsies and therapeutics.

Keywords: Biomarker; Cancer; Characterization; Exosomes; Isolation; Therapy.

Fig. 1

Exosomes from tumor or immune cells stimulate or suppress anti-cancer immunity. a Tumor-derived exosomes bearing antigens stimulate CD8⁺ T cells via dendritic cells (DCs). HSPs-containing exosomes promote the activation and enhance the cytotoxicity of NK cells. b RNA-containing exosomes activate regulatory T cells. Tumor-derived exosomes can stimulate tumor-associated macrophages to adopt a tumor-promoting M2-like phenotype. Similarly, tumor-derived exosomes promote N2-like polarization in tumor-associated neutrophils. PD-L1-bearing exosomes inhibit the cytotoxicity of CD8⁺ T cells. c B cell-derived exosomes carrying MHC II stimulate CD4⁺ T cells by specifically targeting receptors on CD4⁺ T cells. DCs present antigens and MHC I to CD8⁺ T cells, resulting in their activation. Macrophages present antigens to CD4⁺ T cells via DCs, ultimately triggering CD4⁺ T cells activation. d Antigen-bearing exosomes suppress the function of CD8⁺ T cells and CD4⁺ T cells in bone marrow-derived cells, tumor-associated macrophages, and regulatory T cells

Fig. 2

Clinical applications of exosomes in cancer. Exosomes can be extracted from bodily fluids, including cerebrospinal fluid, saliva, milk, lymph, bile, blood, and urine (among others). Analysis of the molecular contents of exosomes, including proteins, nucleic acids, metabolites, and lipids, could provide unique opportunities in the context of liquid biopsies for gaining information about the presence, molecular profile, and behavior of cancer. Exosomes can be used as biomarkers in cancer diagnosis, prediction, and surveillance. Clinical treatment mainly involves three strategies: First, cargo, including drugs, DNAs, RNAs, and proteins, can be encapsulated in exosomes and targeted to cancer sites. Second, immunotherapy can be used in cancer therapy. DC-derived exosomes inhibit tumor progression. CAR-containing exosomes, unlike CAR-T cells, suppress tumor progression via receptor binding. The interaction between SIRPα on macrophages and CD47 on tumor cells can be blocked by engineered exosomes. Therapeutics inhibit the release of PD-L1-bearing exosomes. Finally, inhibition of exosome biogenesis, secretion and uptake are relevant to cancer therapy. Exosome secretion and biogenesis can be prevented via a p300/CBP inhibitor or genetic knockout of Rab27a and nSMase2. The uptake process can be prevented by inhibitors such as heparin, cytochalasin D, methyl-β-cyclodextrin, and dynasore