Dendritic Cell-derived Exosomes Research Articles

Hybrid nanovesicles comprising induced pluripotent stem cells and dendritic cell-derived exosomes enhance antitumor immunity

Induced pluripotent stem cells (iPSCs) tailored to express tumor-specific antigens have emerged as a promising strategy for cancer immunoprevention. Despite the potential of this approach, a number of challenges remain to be addressed—such as ensuring that iPSCs are safely transported to immune-processing organs, mitigating the risk of oncogenesis, and increasing the degree of antitumor efficacy. To address these, we developed innovative vesicles that merge the antigenic repertoire of iPSCs with the lymph node homing and dendritic cell (DC)-targeting capabilities of DC-derived exosomes (DEXO), further boosted by the use of CpG oligodeoxynucleotides (CpG ODNs) as adjuvants. Our engineered vesicles are crafted to engage with immune system gatekeepers, streamlining the delivery of tumor antigens and priming T-cell responses. Laboratory analyses clarified their proficiency in terms of navigating to lymph nodes and initiating potent activation of both DCs and T-cells, thereby intensifying antitumor effects. Further tests in animal models corroborated their therapeutic potential, offering significant defense against tumor initiation, growth, spread, and recurrence, alongside significant enhancements in survival rates. Our findings indicate a promising pathway for innovative iPSC-based cancer vaccine therapies, laying a foundation for future clinical applications.

Tumor Antigen-Primed Dendritic Cell-Derived Exosome Synergizes with Colony Stimulating Factor-1 Receptor Inhibitor by Modulating the Tumor Microenvironment and Systemic Immunity.

Dendritic cell-derived exosomes (Dex) have overcome the disadvantages associated with dendritic cell (DC) vaccines, such as cost effectiveness, stability, and sensitivity to the systemic microenvironment. However, in clinical trials, Dex failed to provide satisfactory results because of many reasons, including inadequate maturation of DC as well as the immunosuppressive tumor microenvironment (TME). Hence, culturing DCs in the presence of a maturation cocktail showed an induced expression of MHCs and co-stimulatory molecules. Additionally, targeting the colony stimulating factor-1 (CSF-1)/CSF-1 receptor (CSF-1R) signaling pathway by a CSF-1R inhibitor could deplete tumor-associated macrophages (TAMs) and myeloid-derived suppressor cells (MDSCs) which are responsible for immunosuppressive TME. Hence, in this study, mDexTA were isolated from bone marrow-derived DC cultured in the presence of a novel maturation cocktail and tumor antigen. mDexTA showed elevated expression of major histocompatibility complexes (MHCs) and co-stimulatory molecules and was found capable of activating naïve DC and T cells in vitro more efficiently when compared to imDexTA isolated from immature DCs. In addition, PLX-3397, a small molecule inhibitor of CSF-1/CSF-1R, was used in combination to enhance the antitumor efficacy of mDexTA. PLX-3397 showed dose-dependent toxicity against bone marrow-derived macrophages (BMDMs). In the B16-F10 murine melanoma model, we found that the combination treatment delayed tumor growth and improved survival compared to the mice treated with mDexTA alone by enhancing the CD8 T cells infiltration in TME. mDexTA when combined with PLX-3397 modulated the TME by shifting the Th1/Th2 toward a dominant Th1 population and depleting the TAMs and MDSCs. Interestingly, PLX-3397-induced FoxP3 expression was diminished when it was used in combination with mDexTA. Combination treatment also induced favorable systemic antitumor immunity in the spleen and lymph node. In conclusion, our findings provide insights into the synergy between mDexTA-based immunotherapy and PLX-3397 as the combination overcame the disadvantages associated with monotherapy and offer a therapeutic strategy for the treatment of solid tumors including melanoma.

Dendritic Cell-Derived Exosomes Stimulated by Treponema pallidum Induce Endothelial Cell Inflammatory Response through the TLR4/MyD88/NF-κB Signaling Pathway.

Exosomes have been implicated in vascular damage in recent research. The influence of dendritic cell-derived exosomes generated by Treponema pallidum (T. pallidum) on the inflammatory process of vascular cells was examined in this study. Human umbilical vein endothelial cells (HUVECs) were cocultured with exosomes isolated from dendritic cells induced by T. pallidum. Western blot and reverse transcription-quantitative real-time polymerase chain reaction were used to assess toll-like receptor 4 (TLR4) expression and the quantity of proinflammatory cytokines. The findings showed that the expression of TLR4 was considerably upregulated, and TLR4 knockdown dramatically reduced interleukin-1β (IL-1β), interleukin-6 (IL-6), and tumor necrosis factor-α (TNF-α) production in exosome-treated HUVECs. Furthermore, TLR4 silencing reduced myeloid differentiation primary response protein 88 (MyD88) and nuclear factor kappa light chain enhancer of activated B cells (NF-κB) levels in exosome-treated HUVECs. Additionally, suppression of the activity of NF-κB with BAY11-7082, an NF-κB inhibitor, also reduced the exosome-treated inflammatory response. Our results suggested that dendritic cell-derived exosomes stimulated by T. pallidum induced endothelial cell inflammation, and the TLR4/MyD88/NF-κB signal axis was activated, significantly increasing IL-1β, IL-6, and TNF-α expression. This may have a significant role in the vascular inflammatory response in syphilis, which would contribute to the understanding of the pathogenesis of syphilis and the host immunological response to T. pallidum.

Dendritic Cell-Derived Exosomes in Cancer Immunotherapy.

Exosomes are nanoscale vesicles released by diverse types of cells for complex intercellular communication. Numerous studies have shown that exosomes can regulate the body's immune response to tumor cells and interfere with the tumor microenvironment (TME). In clinical trials on dendritic cell (DC)-based antitumor vaccines, no satisfactory results have been achieved. However, recent studies suggested that DC-derived exosomes (DEXs) may be superior to DC-based antitumor vaccines in avoiding tumor cell-mediated immunosuppression. DEXs contain multiple DC-derived surface markers that capture tumor-associated antigens (TAAs) and promote immune cell-dependent tumor rejection. These findings indicate the necessity of the further development and improvement of DEX-based cell-free vaccines to complement chemotherapy, radiotherapy, and other immunotherapies. In this review, we highlighted the recent progress of DEXs in cancer immunotherapy, particularly by concentrating on landmark studies and the biological characterization of DEXs, and we summarized their important role in the tumor immune microenvironment (TIME) and clinical application in targeted cancer immunotherapy. This review could enhance comprehension of advances in cancer immunotherapy and contribute to the elucidation of how DEXs regulate the TIME, thereby providing a reference for utilizing DEX-based vaccines in clinical practice.

Mesenchymal stem cells, as glioma exosomal immunosuppressive signal multipliers, enhance MDSCs immunosuppressive activity through the miR-21/SP1/DNMT1 positive feedback loop

BackgroundThe immunosuppressive microenvironment in glioma induces immunotherapy resistance and is associated with poor prognosis. Glioma-associated mesenchymal stem cells (GA-MSCs) play an important role in the formation of the immunosuppressive microenvironment, but the mechanism is still not clear.ResultsWe found that GA-MSCs promoted the expression of CD73, an ectonucleotidase that drives immunosuppressive microenvironment maintenance by generating adenosine, on myeloid-derived suppressor cells (MDSCs) through immunosuppressive exosomal miR-21 signaling. This process was similar to the immunosuppressive signaling mediated by glioma exosomal miR-21 but more intense. Further study showed that the miR-21/SP1/DNMT1 positive feedback loop in MSCs triggered by glioma exosomal CD44 upregulated MSC exosomal miR-21 expression, amplifying the glioma exosomal immunosuppressive signal. Modified dendritic cell-derived exosomes (Dex) carrying miR-21 inhibitors could target GA-MSCs and reduce CD73 expression on MDSCs, synergizing with anti-PD-1 monoclonal antibody (mAb).ConclusionsOverall, this work reveals the critical role of MSCs in the glioma microenvironment as signal multipliers to enhance immunosuppressive signaling of glioma exosomes, and disrupting the positive feedback loop in MSCs with modified Dex could improve PD-1 blockade therapy.Graphical abstract

Dendritic Cell-Derived Exosomes Driven Drug Co-Delivery Biomimetic Nanosystem for Effective Combination of Malignant Melanoma Immunotherapy and Gene Therapy.

Malignant melanoma (MM), the most lethal skin cancer, is highly invasive and metastatic. These qualities are related to not only genetic mutations in MM itself but also the interaction of MM cells with the immune system and microenvironment. This study aimed to construct a combined immunotherapy and gene therapy drug delivery system for the effective treatment of MM. Mature dendritic cell (mDC) exosomes (mDexos) with immune induction functions were used as carriers. BRAF siRNA (siBRAF) with the ability to silence mutated BRAF in MM was encapsulated in mDexos by electroporation to construct a biomimetic nanosystem for the codelivery of immunotherapy and gene therapy drugs (siBRAF-mDexos) to the MM microenvironment. Then, we investigated the nanosystem's serum stability and biocompatibility, uptake efficiency in mouse melanoma cells (B16-F10 cells), cytotoxicity against B16-F10 cells and inhibitory effect on BRAF expression. Furthermore, we evaluated its antimelanoma activity and safety in vivo. SiBRAF-mDexos were nanosized. Compared to siBRAF, siBRAF-mDexos displayed significantly increased serum stability, biocompatibility, uptake efficiency in B16-F10 cells, and cytotoxicity to B16-F10 melanoma cells; they also had a significantly greater inhibitory effect on BRAF expression and induced T-lymphocyte proliferation. Moreover, compared with siBRAF, siBRAF-mDexos showed significantly enhanced anti-MM activity and a high level of safety in vivo. The study suggests that the siBRAF-mDexo biomimetic drug codelivery system can be used to effectively treat MM, which provides a new strategy for combined gene therapy and immunotherapy for MM.

Abstract 5077: Engineering dendritic cell-derived exosomes forbrain metastases immunotherapy

Abstract Brain metastases (BrMs) represent an unmet challenge for the therapy of most aggressive cancers owing to the extremely poor survival and the limited efficacy of available therapies. Currently, NO curative therapy for breast cancer BrM (BCBrM) is available and we urgently need to develop new and effective therapies. Immune checkpoint therapies (ICT) have revolutionized cancer treatment, but are generally unsuccessful in treating BrMs, including BCBrM. A crucial reason for this is the unique brain immune microenvironment with low antigen exposure/presentation in BrMs and fewer antigen-specific T cells in the brain parenchyma. The priming and activation of antigen-specific T cells require professional antigen-presenting dendritic cells (DCs) for effective antigen presentation through multifaceted communication between DCs and T cells by direct intercellular contact and indirect distant communication via exosomes and other means. DC-derived exosomes (DCEXs) carry functional MHC-I/peptide complexes (pMHC-I) and costimulatory molecules, hence facilitating systemic T cell immune responses. DCEXs have been developed as immunotherapies which exhibit advantages over DC-based immunotherapies. However, DCEXs in ongoing clinical trials showed limited efficacy. We found that increased non-receptor protein tyrosine phosphatase SHP1, a DC-intrinsic inhibitory checkpoint, in monocyte-derived DCs (MoDCs) ex vivo led to DC exhaustion that limits DCEXs function resulting in poor systemic T cells stimulation in vivo. To improve DCEXs functions, we developed next-generation DCEXs by genetic knockout of SHP1 in DCs from which DCEXs (iSHP1-DCEXs) were isolated. Remarkably, the modified iSHP1-DCEXs exhibited significantly increased pMHC-I expression and more effectively improved T cell proliferation compared to control DCEXs, indicating a greater capacity of antigen presentation of engineered iSHP1-DCEXs. In the EO771-OVA mammary tumor cells-bearing mouse model, iSHP1-DCEXs treatment greatly increased tumor infiltration of OVA antigen-specific CD8+ T (OT-1) cells resulting in tumor inhibition and prolonged mouse survival, indicating that iSHP1-DCEXs elicited an effective and specific antitumor immunity in vivo. Moreover, combining iSHP1-DCEXs with anti-PD1 Ab yielded a strong syngenetic therapeutic response in mice bearing the anti-PD1-resistant EO771 tumors, as indicated by delayed tumor growth and prolonged survival. importantly, iSHP1-DCEXs also significantly increased tumor-infiltrating antigen-specific T cells and synergized with PD-1 Ab leading to tumors clearance in mice bearing the B16-GMCSF melanoma. To further test the iSHP1-DCEXs effect in BCBrM, we co-injected EO771-OVA and iSHP1-DCEXs into mice and found that iSHP1-DCEXs inhibited BrM. Together, the engineered iSHP1-DCEXs offer a promising option to generate a potent antigen-specific T cell immune response for BCBrM inhibition. Citation Format: Hao-Nien Chen, Xiangliang Yuan, Yimin Duan, Yi Xiao, Shao-Ping Yang, DIhua Yu. Engineering dendritic cell-derived exosomes forbrain metastases immunotherapy. [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2023; Part 1 (Regular and Invited Abstracts); 2023 Apr 14-19; Orlando, FL. Philadelphia (PA): AACR; Cancer Res 2023;83(7_Suppl):Abstract nr 5077.

Release of exosomes in polytraumatized patients: The injury pattern is reflected by the surface epitopes.

Trauma is still a leading cause of morbidity and mortality, especially in the younger population. Trauma patients need a precise, early diagnostic to avoid complications like multiorgan failure and sepsis. Exosomes were described as markers and mediators in trauma. The aim of the present study was to analyze, whether the surface epitopes of plasma-exosomes can reflect the injury pattern in polytrauma. Polytraumatized patients (Injury Severity Score = ISS ≥16, n = 38) were subdivided according to the predominant injury in either abdominal trauma, chest trauma or traumatic brain injury (TBI). Plasma exosomes were isolated via size exclusion chromatography. The concentration and size distribution of the plasma exosomes from emergency room samples were measured by nanoparticle tracking analysis. The exosomal surface antigens were investigated by bead-based multiplex flow cytometry and compared with healthy controls (n=10). In contrast to other studies, we did not observe an increase in the total amount of plasma exosomes in polytrauma patients (1,15x109 vs. 1,13x109 particles/ml), but found changes in the exosomal surface epitopes. We found a significant reduction of CD42a+ (platelet-derived) exosomes in polytrauma patients, CD209+ (dendritic cell-derived) exosomes in the patients with predominant abdominal trauma, and CD11+ (monocyte-derived) exosomes in the patients with chest trauma. The group of patients with TBI was characterized in contrast by an increase of CD62p+ (endothelial/platelet-derived) exosomes (*p<0.05). Our data showed that the polytrauma injury pattern might be reflected by the cellular origin/surface epitopes of plasma-released exosomes immediately after trauma. The observed reduction of CD42+ exosomes in polytrauma patients was not associated with a reduction of total platelets in polytrauma patients.

A novel costimulatory molecule gene-modified leukemia cell-derived exosome-targeted CD4+ T cell vaccine efficiently enhances anti-leukemia immunity.

Previous studies demonstrated that CD4+ T cells can uptake tumor antigen-pulsed dendritic cell-derived exosomes (DEXO), which harbor tumor antigen peptide/pMHC I complex and costimulatory molecules and show potent effects on inducing antitumor immunity. However, in preliminary study, CD4+ T cells targeted by leukemia cell-derived exosomes (LEXs) did not show the expected effects in inducing effective anti-leukemia immunity, indicating that LEX is poorly immunogenetic largely due to an inadequate costimulatory capacity. Therefore, LEX-based anti-leukemia vaccines need to be optimized. In this study, we constructed a novel LEX-based vaccine by combining CD4+ T cells with costimulatory molecules gene-modified LEXs, which harbor upregulated CD80 and CD86, and the anti-leukemia immunity of CD80 and CD86 gene-modified LEX-targeted CD4+ T cells was investigated. We used lentiviral vectors encoding CD80 and CD86 to successfully transduced the L1210 leukemia cells, and the expression of CD80 and CD86 was remarkably upregulated in leukemia cells. The LEXs highly expressing CD80 and CD86 were obtained from the supernatants of gene-transduced leukemia cells. Our data have shown that LEX-CD8086 could promote CD4+ T cell proliferation and Th1 cytokine secretion more efficiently than control LEXs. Moreover, CD4+ TLEX-CD8086 expressed the acquired exosomal costimulatory molecules. With acquired costimulatory molecules, CD4+ TLEX-CD8086 can act as APCs and are capable of directly stimulating the leukemia cell antigen-specific CD8+ CTL response. This response was higher in potency compared to that noted by the other formulations. Furthermore, the animal study revealed that the CD4+ TLEX-CD8086 significantly inhibited tumor growth and prolonged survival of tumor-bearing mice than other formulations did in both protective and therapeutic models. In conclusion, this study revealed that CD4+ TLEX-CD8086 could effectively induce more potential anti-leukemia immunity than LEX-CD8086 alone, suggesting that the utilization of a costimulatory molecule gene-modified leukemia cell-derived exosome-targeted CD4+ T cell vaccine may have promising potential for leukemia immunotherapy.

Recent progress of dendritic cell-derived exosomes (Dex) as an anti-cancer nanovaccine.

Although cancer vaccines such as dendritic cell (DC) vaccines and peptide vaccines have become appealing and attractive anticancer immunotherapy options in recent decades, some obstacles have hindered their successful application in the clinical setting. The difficulties associated with the high cost of DC preparation, storage of DC vaccines, tumor-mediated immunosuppressive environment, identification of specific tumor antigens, and high degradation of antigen peptides in vivo limit the clinical application and affect the outcomes of these cancer vaccines. Recently, nanocarriers have been considered as a new approach for vaccine delivery. As biogenic nanocarriers, exosomes are small membrane vesicles secreted by cells that carry various proteins, RNAs, and lipids. More importantly, DC-derived exosomes (Dex) express tumor antigens, MHC molecules, and co-stimulatory molecules on their surface, which trigger the release of antigen-specific CD4+ and CD8+ T cells. With their membrane structure, Dex can avoid high degradation while ensuring favorable biocompatibility and biosafety in vivo. In addition, Dex can be stored in vitro for a longer period, which facilitates a significant reduction in production costs. Furthermore, they have shown better antitumor efficacy in preclinical studies compared with DC vaccines owing to their higher immunogenicity and stronger resistance to immunosuppressive effects. However, the clinical efficacy of Dex vaccines remains limited. In this review, we aimed to evaluate the efficacy of Dex as an anticancer nanovaccine.

Modified dendritic cell-derived exosomes activate both NK cells and T cells through the NKG2D/NKG2D-L pathway to kill CML cells with or without T315I mutation

BackgroundTyrosine kinase inhibitors have achieved quite spectacular advances in the treatment of chronic myeloid leukemia (CML), but disease progression and drug resistance that related to the T315I mutation, remain major obstacles. Dendritic cell-derived exosomes (Dex) induce NK cell immunity, but have yet to achieve satisfactory clinical efficacy. An approach to potentiate antitumor immunity by inducing both NK- and T-cell activation is urgently needed. Retinoic acid early inducible-1γ (RAE-1γ), a major ligand of natural killer group 2 member D (NKG2D), plays an important role in NK-cell and T-lymphocyte responses. We generated RAE-1γ enriched CML-specific Dex (CML-RAE-1γ-Dex) from dendritic cells (DCs) pulsed with lysates of RAE-1γ-expressing CML cells or T315I-mutant CML cells, aiming to simultaneously activate NK cells and T lymphocytes.MethodsWe generated novel CML-RAE-1γ-Dex vaccines, which expressed RAE-1γ, and were loaded with CML tumor cell lysates. NK cells or T lymphocytes were coincubated with CML-RAE-1γ-Dex vaccines. Flow cytometry was performed to evaluate the activation and proliferation of these immune cells. Cytokine production and cytotoxicity toward CML cells with or without the T315I mutation were detected by ELISPOT, ELISA and LDH assays. CML models induced by BCR-ABL or BCR-ABLT315I were used to determine the immunological function of Dex in vivo.ResultsHerein, CML-RAE-1γ-Dex were prepared. CML-RAE-1γ-Dex effectively enhanced the proliferation and effector functions of NK cells, CD4+ T cells and CD8+ T cells, which in turn produced strong anti-CML efficacy in vitro. Moreover, CML-RAE-1γ-Dex-based immunotherapy inhibited leukemogenesis and generated durable immunological memory in CML mouse models. Similar immune responses were also observed with imatinib-resistant CML cells carrying the T315I mutation.ConclusionsThis approach based on CML-RAE-1γ-Dex vaccines may be a promising strategy for CML treatment, especially for cases with the T315I mutation.

Dendritic cells a critical link to alveolar bone loss and systemic disease risk in periodontitis: Immunotherapeutic implications.

Extensive research in humans and animal models has begun to unravel the complex mechanisms that drive the immunopathogenesis of periodontitis. Neutrophils mount an early and rapid response to the subgingival oral microbiome, producing destructive enzymes to kill microbes. Chemokines and cytokines are released that attract macrophages, dendritic cells, and T cells to the site. Dendritic cells, the focus of this review, are professional antigen-presenting cells on the front line of immune surveillance. Dendritic cells consist of multiple subsets that reside in the epithelium, connective tissues, and major organs. Our work in humans and mice established that myeloid dendritic cells are mobilized in periodontitis. This occurs in lymphoid and nonlymphoid oral tissues, in the bloodstream, and in response to Porphyromonas gingivalis. Moreover, the dendritic cells mature in situ in gingival lamina propria, forming immune conjugates with cluster of differentiation(CD) 4+ T cells, called oral lymphoid foci. At such foci, the decisions are made as to whether to promote bone destructive Thelper17 or bone-sparing regulatory T cell responses. Interestingly, dendritic cells lack potent enzymes and reactive oxygen species needed to kill and degrade endocytosed microbes. The keystone pathogen P.gingivalis exploits this vulnerability by invading dendritic cells in the tissues and peripheral blood using its distinct fimbrial adhesins. This promotes pathogen dissemination and inflammatory disease at distant sites, such as atherosclerotic plaques. Interestingly, our recent studies indicate that such P.gingivalis-infected dendritic cells release nanosized extracellular vesicles called exosomes, in higher numbers than uninfected dendritic cells do. Secreted exosomes and inflammasome-related cytokines are a key feature of the senescence-associated secretory phenotype. Exosomes communicate in paracrine with neighboring stromal cells and immune cells to promote and amplify cellular senescence. We have shown that dendritic cell-derived exosomes can be custom tailored to target and reprogram specific immune cells responsible for inflammatory bone loss in mice. The long-term goal of these immunotherapeutic approaches, ongoing in our laboratory and others, is to promote human health and longevity.

A CAR T-inspiring platform based on antibody-engineered exosomes from antigen-feeding dendritic cells for precise solid tumor therapy

Chimeric antigen receptor T (CAR T) cell therapy has achieved remarkable results treating patients with hematological malignancies in clinical studies. Although promising, extensive research has also revealed that CAR T therapy is unsatisfactory for the treatment of solid tumors. In addition, the production of CAR T cells is time-consuming and it's hard for storage and transportation. In this work, inspired by the construction of CAR T cell, we developed an antibody-engineered exosomes from antigen-feeding dendritic cells for beyond CAR T therapy of solid tumor by in situ T cells activation and cancer cell targeting. We have confirmed that tumor antigen-stimulated dendritic cell-derived exosomes (tDC-Exo) provided major histocompatibility (MHC)-antigen complexes and CD86 co-stimulating molecules, which were the same as CAR of CAR T cell acting as the necessary signals for T cell activation. Furthermore, anti-CD3 and anti-EGFR were then engineered on tDC-Exo to promote the binding of T cell to cancer cells for precise therapy. Our CAR T cell therapy-mimicking system have shown an efficient endogenous T cells activation and their crosslinking with cancer cells for enhanced solid tumor therapy. More interestingly, we found that immune activation significantly up-regulated PD-L1 expression, and thus we confirmed the combination with anti-PD-L1 antibodies further enhanced the efficacy of our CAR T cell therapy-mimicking platform.

Exogenous and Endogenous Dendritic Cell-Derived Exosomes: Lessons Learned for Immunotherapy and Disease Pathogenesis.

Immune therapeutic exosomes, derived exogenously from dendritic cells (DCs), the ‘directors’ of the immune response, are receiving favorable safety and tolerance profiles in phase I and II clinical trials for a growing number of inflammatory and neoplastic diseases. DC-derived exosomes (EXO), the focus of this review, can be custom tailored with immunoregulatory or immunostimulatory molecules for specific immune cell targeting. Moreover, the relative stability, small size and rapid uptake of EXO by recipient immune cells offer intriguing options for therapeutic purposes. This necessitates an in-depth understanding of mechanisms of EXO biogenesis, uptake and routing by recipient immune cells, as well as their in vivo biodistribution. Against this backdrop is recognition of endogenous exosomes, secreted by all cells, the molecular content of which is reflective of the metabolic state of these cells. In this regard, exosome biogenesis and secretion is regulated by cell stressors of chronic inflammation and tumorigenesis, including dysbiotic microbes, reactive oxygen species and DNA damage. Such cell stressors can promote premature senescence in young cells through the senescence associated secretory phenotype (SASP). Pathological exosomes of the SASP amplify inflammatory signaling in stressed cells in an autocrine fashion or promote inflammatory signaling to normal neighboring cells in paracrine, without the requirement of cell-to-cell contact. In summary, we review relevant lessons learned from the use of exogenous DC exosomes for immune therapy, as well as the pathogenic potential of endogenous DC exosomes.

An efficient and safe MUC1-dendritic cell-derived exosome conjugate vaccine elicits potent cellular and humoral immunity and tumor inhibition in vivo

Antitumor vaccines are a promising strategy for preventing or treating cancers by eliciting antitumor immune responses and inducing protective immunity against specific antigens expressed on tumor cells. Vaccine formulations that enhance the humoral and cellular immune responses of vaccine candidates would be highly beneficial but are still limited. Here we developed an antitumor vaccine candidate by conjugating a MUC1 glycopeptide antigen to dendritic cell-derived exosomes (Dex). In vivo, the MUC1-Dex construct induced high MUC1-specific IgG antibody titers with strong binding affinities for MUC1-positive tumor cells and promoted cytokine secretion. Moreover, CD8+ T cells from immunized mice exhibited strong cytotoxicity against MUC1-positive tumor cells. Importantly, in both preventative and therapeutic tumor-bearing mouse models, the construct inhibited tumor growth and prolonged survival. Collectively, these results demonstrate that Dex is a promising vaccine carrier that can be used as adjuvant to enhance the immunological efficacy of tumor vaccines. Statement of significance•We prepared an efficient and biosafe antitumor vaccine candidate by conjugating Dex with MUC1 glycopeptide using a two-step biorthogonal crosslinking strategy.•MUC1-Dex enhanced the expression of costimulatory molecules CD80 and CD86 on DCs to promote their maturation whilst simultaneously upregulating MHC I and MHC II expression to increase antigen presentation by DCs.•MUC1-Dex conjugates stimulated the production of MUC1-specific IgG antibodies in vivo, and the antibodies bound strongly to MUC1-expressing tumor cells.•The constructed vaccines induced a strong antibody response and antigen specific CTLs to kill the tumor and delayed tumor development in a therapeutic and prophylactic tumor bearing mouse model.

Hydrogel-load exosomes derived from dendritic cells improve cardiac function via Treg cells and the polarization of macrophages following myocardial infarction

BackgroudMyocardial infarction (MI) is one of the leading causes of global death. Dendritic cell-derived exosomes (DEXs) provide us with the possibility of improving cardiac function after MI but are limited by low retention times and short-lived therapeutic effects. In this study, we developed a novel drug delivery system incorporating alginate hydrogel that continuously releases DEXs and investigated the mechanisms underlying the action of DEXs in the improvement of cardiac function after MI.ResultsWe incorporated DEXs with alginate hydrogel (DEXs-Gel) and investigated controlled released ability and rheology, and found that DEXs-Gel release DEXs in a sustainable mammer and prolonged the retention time of DEXs but had no detrimental effects on the migration in vivo. Then DEXs-Gel was applicated in the MI model mice, we found that DEXs-Gel siginificantly enhanced the therapeutic effects of DEXs with regards to improving cardiac function after MI. Flow cytometry and immunofluorescence staining revealed that DEXs significantly upregulated the infiltration of Treg cells and M2 macrophages into the border zoom after MI, and DEXs activated regulatory T (Treg) cells and shifted macrophages to reparative M2 macrophages, both in vitro and in vivo.ConclusionOur novel delivery method provides an innovative tool for enhancing the therapeutic effects of DEXs after MI. Further analysis revealed that DEXs exert effect by activating Treg cells and by modifying the polarization of macrophages.Graphic

Transfer of exosomal microRNA-203-3p from dendritic cells to bone marrow-derived macrophages reduces development of atherosclerosis by downregulating Ctss in mice.

Dendritic cell-derived exosomes have been proven to be efficient adjuvant options for anti-tumor vaccines in cancer immunotherapy. However, their potency in atherosclerosis remains unclear. Here we summarize the association of microRNA-203-3p (miR-203-3p) with dendritic cell-derived exosomes and atherosclerosis. Firstly, dendritic cell-derived exosomes and bone marrow-derived macrophages were isolated, after which expression of miR-203-3p and cathepsin S was determined. After the establishment of atherosclerosis mouse models, gain- and loss-of-function experiments were conducted for the analysis of effects of miR-203-3p and cathepsin S on foam-cell formation, lipid accumulation, collagen deposition and serum total cholesterol. The results found high expression of cathepsin S in atherosclerosis mice and downregulation of miR-203-3p in the serum of atherosclerosis patients and ox-LDL-simulated bone marrow-derived macrophages. Cathepsin S was the target gene of miR-203-3p. miR-203-3p transporting from exosomes to bone marrow-derived macrophages resulted in inhibition of cathepsin S expression and atherosclerosis-related phenotypes in bone marrow-derived macrophages, thus alleviating atherosclerosis in mice, and this process was found to involve the p38/MAPK signaling pathway. These findings provided evidence that the transfer of miR-203-3p by dendritic cell-derived exosomes targeted cathepsin S in bone marrow-derived macrophages to attenuate atherosclerosis progression in mice, serving as a promising clinical target for atherosclerosis.

Development of surface engineered antigenic exosomes as vaccines for respiratory syncytial virus

Abstract Human respiratory syncytial virus (HRSV) is one of the main drivers of lower respiratory tract illness in infants and young children worldwide. There are no licensed vaccines for HRSV available. Exosomes are cell-derived extracellular nanovesicles containing various biomolecules for intracellular communication. Exosomes secreted by antigen presenting cells such as dendritic cells can elicit immune responses by carrying MHC-I with antigenic peptide complex and other co-stimulating factors. Therefore, exosomes have emerged as potential vaccines to prevent viral infections or to treat cancers. Our prior work has demonstrated that a PDMS microfluidic culture chip device can be used to generate dendritic cell-derived tumor antigenic exosomes, with the capacity to activate tumor-specific CD8+ T cells. Herein, we further extended the work to surface engineer dendritic cell-derived exosomes with antigenic peptides from RSV (M187–195 and NS161–75). A mouse model of RSV infection was used to define the immunogenicity of surface engineered exosomes for activating virus-specific immune responses in vitro and in vivo. In vitro assays demonstrated that surface engineered exosomes successfully carried the engineered peptides of interest and had the capacity to elicit IFNγ production by activated, virus-specific CD8+ T cells. However, vaccination with engineered exosomes did not prime an in vivo activation of antigen-specific CD8+ T cell response, although surface engineered exosomes exhibit the safe administration. Additional experiments are necessary to optimize the potency of surface engineered exosomes as a vaccine platform for the prevention of viral infections such as RSV.

Reactive oxygen species-responsive dendritic cell-derived exosomes for rheumatoid arthritis

Although tolerogenic dendritic cell-derived exosomes (TolDex) have emerged as promising therapeutics for rheumatoid arthritis (RA), their clinical applications have been hampered by their poor in vivo disposition after systemic administration. Herein, we report the development of stimuli-responsive TolDex that induces lesion-specific immunoregulation in RA. Responsiveness to reactive oxygen species (ROS), a physiological stimulus in the RA microenvironment, was conferred on TolDex by introducing a thioketal (TK) linker-embedded poly(ethylene glycol) (PEG) on TolDex surface via hydrophobic insertion. The detachment of PEG following overproduction of ROS facilitates the cellular uptake of ROS-responsive TolDex (TKDex) into activated immune cells. Notably, TolDex and TKDex downregulated CD40 in mature dendritic cells (mDCs) and regulated secretion of pro-inflammatory cytokines, including tumor necrosis factor (TNF)-α and interleukin-6 (IL-6) at the cellular level. In the collagen-induced arthritis (CIA) mouse model, PEG prolonged the blood circulation of TKDex following intravenous administration and enhanced their accumulation in the joints. In addition, TKDex decreased IL-6, increased transforming growth factor-β, and induced the CD4+CD25+Foxp3+ regulatory T cells in CIA mice. Overall, ROS-responsive TolDex might have potential as therapeutic agents for RA. Statement of significanceTolerogenic dendritic cell-derived exosomes (TolDex) are emerging immunoregulators of autoimmune diseases, including rheumatoid arthritis (RA). However, their lack of long-term stability and low targetability are still challenging. To overcome these issues, we developed reactive oxygen species (ROS)-responsive TolDex (TKDex) by incorporating the ROS-sensitive functional group-embedded poly(ethylene glycol) linker into the exosomal membrane of TolDex. Surface-engineered TKDex were internalized in mature DCs because of high ROS-sensitivity and enhanced accumulation in the inflamed joint in vivo. Further, for the first time, we investigated the potential mechanism of action of TolDex relevant to CD40 downregulation and attenuation of tumor necrosis factor (TNF)-α secretion. Our strategy highlighted the promising nanotherapeutic effects of stimuli-sensitive TolDex, which induces immunoregulation.

Analysis of differential expression of long non‑coding RNAs inexosomes derived from mature and immature dendritic cells.

Dendritic cells release bioactive exosomes involved in immune regulation. Long non-coding RNAs (lncRNAs) are implicated in a number of immunoregulatory mechanisms. However, the roles of lncRNAs in dendritic cell-derived exosomes remain to be elucidated. The present study aimed to investigate the roles of lncRNAs in exosomes derived from mature and immature dendritic cells and to find specific lncRNAs with immunoregulatory function. The expression profiles of lncRNAs in exosomes derived from bone marrow dendritic cells of C57 mice were illustrated. Gene Ontology (GO), Kyoto Encyclopedia of Genes and Genomes (KEGG) pathway analyses and Gene Set Enrichment Analysis were performed to identify potential targets correlated with immune regulation. In addition, lncRNA-miRNA-mRNA networks were predicted using bioinformatics methods. Representative lncRNAs were further validated via reverse transcription-quantitative PCR. A total of 437 lncRNAs were analyzed using RNA-seq. Among these, the expression of ~87 lncRNAs was upregulated and 21 lncRNAs was downregulated in mature dendritic cell-derived exosomes (Dex) compared with immature Dex. GO analyses indicated the involvement of upregulated lncRNAs in multiple biological functions, such as the immune system process, while downregulated lncRNAs were involved in poly(A) RNA binding. Analysis of the KEGG pathway identified the relationship of TNF signaling and ribosome pathway with upregulated lncRNAs and downregulated lncRNAs, respectively. The results of gene set enrichment analysis identified that three lncRNA-associated transcripts (Procr-203, Clec4e-202 and Traf1-203) were highly associated with immunoregulatory functions including T helper cell differentiation and Janus kinase-STAT signaling pathway. The results indicated the involvement of candidate lncRNAs in immunoregulation and suggested a new perspective on the modulation of lncRNAs in Dex.