Synthesis of metformin-derived fluorescent quantum dots: uptake, cytotoxicity, and inhibition in human breast cancer cells through autophagy pathway.

BACKGROUND: Breast cancer remains a challenge for physicians. Metformin, an antidiabetic drug, show promising anticancer properties against cancers. An emerging quantum dot (QD) material improves therapeutic agents' anticancer and imaging properties. QD are nano-sized particles with extreme application in nanotechnology captured by cells and accumulated inside cells, suggesting bioimaging and effective anticancer outcomes. In this study, a simple one-pot hydrothermal method was used to synthesize fluorescent metformin-derived carbon dots (M-CDs) and then investigated the cytotoxic effects and imaging features on two human breast cancer cell lines including, MCF-7 and MDA-MB-231 cells. RESULTS: Results showed that M-CDs profoundly decreased the viability of both cancer cells. IC50 values showed that M-CDs were more cytotoxic than metformin either 24-48 h post-treatment. Cancer cells uptake M-CDs successfully, which causes morphological changes in cells and increased levels of intracellular ROS. The number of Oil Red O-positive cells and the expression of caspase-3 protein were increased in M-CDs treated cells. Authophagic factors including, AMPK, mTOR, and P62 were down-regulated, while p-AMPK, Becline-1, LC3 I, and LC3 II were up-regulated in M-CDs treated cells. Finally, M-CDs caused a decrease in the wound healing rate of cells. CONCLUSIONS: For the first, M-CDs were synthesized by simple one-pot hydrothermal treatment without further purification. M-CDs inhibited both breast cancer cells through modulating autophagy signalling.

Exosomes biogenesis was increased in metformin-treated human ovary cancer cells; possibly to mediate resistance.

BACKGROUND: Exosomes derived from tumor cells contribute to the pathogenesis of cancers. Metformin, the most usually used drug for type 2 diabetes, has been frequently investigated for anticancer effects. Here, we examined whether metformin affects exosomes signaling in human ovary cancer cells in vitro. METHODS: Human ovary cancer cells, including A2780 and Skov3 cells, were treated with metformin for either 24-48 h. Cell viability and caspase-3 activity were determined by MTT (3-[4,5-dimethylthiazol-2-yl]-2,5 diphenyl tetrazolium bromide) and colorimetric assays respectively. Oil-Red-O staining and in vitro, scratch assays were used to examine cellular toxicity and wound healing rate. After treatment with metformin, exosomes were isolated from cells and quantified by acetylcholinesterase (AChE) assay, Dynamic Light Scattering (DLS), and their markers. Genes related to exosomes signaling were analyzed by real-time PCR or western blotting. RESULTS: Our results showed that metformin decreased the viability of both cells dose/time-dependently (P < 0.05). Metformin increased the activity of caspase-3 (P < 0.05) as well as the number of Oil-Red-O positive cells in both cell lines. In vitro scratch assay showed that the cell migration rate of metformin-treated cells was decreased (P < 0.05), whereas AChE activity of exosomes from metformin-treated cells was increased (P < 0.05). Concurrent with an increase in CD63 protein levels, expression of Alix, CD63, CD81, Lamp-2, and Rab27b up-regulated in treated cells (P < 0.05). CONCLUSION: Results indicated that metformin had a cytotoxic effect on ovary cancer cells and enhanced exosome biogenesis and secretion.

Melatonin reduces lung injury in type 1 diabetic mice by the modulation of autophagy.

BACKGROUND: In recent years, the role of autophagy has been highlighted in the pathogenesis of diabetes and inflammatory lung diseases. In this study, using a diabetic model of mice, we investigated the expression of autophagy-related genes in the lung tissues following melatonin administration. RESULTS: Data showed histopathological remodeling in lung tissues of the D group coincided with an elevated level of IL-6, Becline-1, LC3, and P62 compared to the control group (p < 0.05). After melatonin treatment, histopathological remodeling was improved D + Mel group. In addition, expression levels of IL-6, Becline-1, LC3, and P62 were decreased in D + Mel compared to D group (P < 0.05). Statistically significant differences were not obtained between Mel group and C group (p > 0.05). CONCLUSION: Our results showed that melatonin injection can be effective in the amelioration of lung injury in diabetic mice presumably by modulating autophagy-related genes.

Cell-mediated barriers in cancer immunosurveillance.

The immune cells within the tumor microenvironment (TME) exert multifaceted functions ranging from tumor-antagonizing or tumor-promoting activities. During the initial phases of tumor development, the tumor-antagonizing immune cells in the TME combat cancer cells in an immune surveillance process. However, with time, cancer cells can evade detection and impede the immune cells' effectiveness through diverse mechanisms, such as decreasing immunogenic antigen presentation on their surfaces and/or secreting anti-immune factors that cause tolerance in TME. Moreover, some immune cells cause immunosuppressive situations and inhibit antitumoral immune responses. Physical and cellular-mediated barriers in the TME, such as cancer-associated fibroblasts, tumor endothelium, the altered lipid composition of tumor cells, and exosomes secreted from cancer cells, also mediate immunosuppression and prevent extravasation of immune cells. Due to successful clinical outcomes of cancer treatment strategies the potential barriers must be identified and addressed. We need to figure out how to optimize cancer immunotherapy strategies, and how to combine therapeutic approaches for maximum clinical benefit. This review provides a detailed overview of various cells and molecules in the TME, their association with escaping from immune surveillance, therapeutic targets, and future perspectives for improving cancer immunotherapy.

Tumor immune escape: extracellular vesicles roles and therapeutics application.

BACKGROUND: Immune escape, a process by which tumor cells evade immune surveillance, remains a challenge for cancer therapy. Tumor cells produce extracellular vesicles (EVs) that participate in immune escape by transferring bioactive molecules between cells. EVs refer to heterogeneous vesicles that participate in intercellular communication. EVs from tumor cells usually carry tumor antigens and have been considered a source of tumor antigens to induce anti-tumor immunity. However, evidence also suggests that these EVs can accelerate immune escape by carrying heat shock proteins (HSPs), programmed death-ligand 1 (PD-L1), etc. to immune cells, suppressing function and exhausting the immune cells pool. EVs are progressively being evaluated for therapeutic implementation in cancer therapies. EVs-based immunotherapies involve inhibiting EVs generation, using natural EVs, and harnessing engineering EVs. All approaches are associated with advantages and disadvantages. The EVs heterogeneity and diverse physicochemical properties are the main challenges to their clinical applications. SHORT CONCLUSION: Although EVs are criminal; they can be useful for overcoming immune escape. This review discusses the latest knowledge on EVs population and sheds light on the function of tumor-derived EVs in immune escape. It also describes EVs-based immunotherapies with a focus on engineered EVs, followed by challenges that hinder the clinical translation of EVs that are essential to be addressed in future investigations. Video Abstract.

Mesenchymal stem cells-derived exosomes: novel carriers for nanoparticle to combat cancer.

BACKGROUND: The advancement in novel cancer therapeutics brought a platform combining the properties of exosomes with nanoparticles to precision medicine. The novel therapeutic approach aim is cancer-targeted therapy. Exosomes from mesenchymal stem cells (MSCs-Exo) exhibit unique properties in cancer therapies, which makes them an ideal tool for delivering therapeutic agents into tumor cells. The key role of natural MSCs-Exo is controversial in cancer therapy; however, they can be engineered at their surface or cargo to serve as a smart drug delivery system for cancer-targeted therapy. In the last few years, researchers harnessed nanotechnology to enforce MSCs-Exo for cancer management including, tumor cell tracking, imaging, and tumor cell killing. Different nanoparticles such as gold nanoparticles have particularly been incorporated into MSCs-Exo, which showed an efficient accumulation at the site of tumor with improved anticancer impact. These findings indicate that a hybrid of exosomes-nanoparticles may serve as combination therapy for the effective removal of cancers. SHORT CONCLUSION: Although exhibiting impressive potential, the use of nanoparticle-loaded MSCs-Exo as a drug-delivery tool has been troubled by some challenges, therefore, translation to clinic prerequisites further scrutiny. In this review, we focus on nanoparticle-loaded MSCs-Exo as a new cancer therapy and discuss engineered MSC-Exo for target therapy.

Tumor cells-derived exosomal noncoding RNAs in cancer angiogenesis: Molecular mechanisms and prospective.

Exosomes, heterogeneous, membrane-bound nanoparticles that originated from eukaryotic cells, contribute to intracellular communication by transferring various biomolecules both on their surface and as internal cargo. One of the most significant current discussions on cancer progression is noncoding RNAs cargo of exosomes, which can regulate angiogenesis in tumor. A growing body of evidence shows that exosomes from tumor cells contain various microRNAs, long noncoding RNAs, and circular RNAs that can promote tumor progression by inducing angiogenesis. However, some noncoding RNAs may inhibit cancer angiogenesis. Targeting angiogenic noncoding RNA of exosomes may serve as a hopeful implement for cancer therapy. In this review, we discuss the latest knowledge of the roles of exosomal noncoding RNAs in tumor angiogenesis Understanding the biology of exosomal noncoding RNAs can help scientists plan exosomes-based innovations for the treatment of cancer angiogenesis and cancer biomarkers.

Association between Socioeconomic Factors and the Risk of Gastric Cancer Incidence: Results from an Ecological Study.

Background: Gastric cancer (GC), one of the most common cancer worldwide, remains the third leading cause of cancer-related mortality. The etiology of GC may arise from genetic and environmental factors. This study aimed to determine the association between GC incidence and socioeconomic status in Iran. Methods: An ecological study was designed to investigate the relationship between socioeconomic factors and the risk of GC incidence. The data of socioeconomic variables such as income changes, unemployment rate, urbanization ratio, inflation rate, and air pollution changes in 31 provinces were collected from the Statistical Center of Iran, and the data of GC of 31 provinces were provided from the Iranian National Population-based Cancer Registry (INPCR). Data from 2014 to 2017 was analyzed using panel data analysis, the fixed effects model by EViews software. Results: Panel data model was suitable for the present study. Results showed that there was a positive and significant relationship between GC incidence and socioeconomic factors including income changes (P≤ 0.001), unemployment rate (P≤0.01), inflation rate (P≤ 0.05), and air pollution changes (P≤ 0.001). The urbanization ratio showed a negative relationship and was not statistically associated with GC incidence (P> 0.05). Conclusion: Our findings suggest a positive and significant association between socioeconomic status and GC incidence, proposing a GC risk factor. The key public health policies and welfare policies' priority should therefore be to schedule for the GC management.

High-fat diet-induced biogenesis of pulmonary exosomes in an experimental rat model.

BACKGROUND: High-fat diets (HFD) have recently become a public health concern. We hypothesize that HFD induces exosomes biogenesis in the lung tissue of rat model. METHODS AND RESULTS: Sixteen adult male Wistar rats were fed with HFD or a regular chow diet for 3 months. The histopathological changes in lung tissues were measured by hematoxylin and eosin (H&E) staining. Bronchoalveolar lavage (BAL) was performed to assay exosomes by acetylcholinesterase enzyme (AhCE) activity. Real-time PCR (qPCR) was used to evaluate Rab27-b, Alix, and IL-1ß expression, while the immunohistochemical examination was performed for CD81 expression in lung tissues. In addition, expression of IL-1ß was detected by ELISA. We found pathological alterations in the lung tissue of HFD animals. AhCE activity along with the expression level of Rab27-b, Alix, and IL-1ß was increased in HFD animals (p < 0.05). Immunohistochemical staining showed that expression of CD81 was increased in lung tissues of HFD animals compared with the control group (p < 0.05). CONCLUSION: Hence, HFD induced exosomes biogenesis and histopathological changes with IL-1ß expression in rats' lung tissues.

Swimming training reduced inflammation and apoptotic changes in pulmonary tissue in type 1 diabetic mice.

Background: Despite the vulnerability of pulmonary tissue to diabetic conditions, there are few reports related to the detrimental effects of hyperglycemia and therapeutic modalities on lung parenchyma. Here, the apoptotic changes were monitored in the diabetic pulmonary tissue of mice (DM1) subjected to a four‒week swimming plan. Methods: The mice were randomly allocated into Control; Control + Swimming (S); Diabetic group (D); and Diabetic + Swimming (D + S) groups (each in 8 mice). In the D and D + S groups, mice received intraperitoneally 50 mg/kg of streptozotocin (STZ). After 14 days, swimming exercise was done for four weeks. The expression of il-1ß, bcl-2, bax, and caspase-3 was investigated using real-time PCR analysis. A histological examination was performed using H&E staining. Results: DM1 significantly upregulated il-1ß, bax, and caspase-3, and down-regulated bcl-2 compared to the non-diabetic mice (p < 0.05). We noted that swimming exercises reversed the expression pattern of all genes in the diabetic mice and closed to basal levels (p < 0.05). Data indicated that swimming exercise could diminish emphysematous changes, and interstitial pneumonitis induced by STZ. Along with these changes, swimming exercise had protective effects to reduce the thickness of the inter-alveolar septum and mean alveolar area in diabetic mice. Conclusion: These data demonstrated that swimming exercises could decrease DM1-related pathologies in mouse lungs by regulating apoptosis and inflammatory response.

Therapeutic application of mesenchymal stem cells derived exosomes in neurodegenerative diseases: A focus on non-coding RNAs cargo, drug delivery potential, perspective.

Despite the massive efforts advanced over recent years in emerging therapies for neurodegenerative diseases, effective treatment for these diseases is still an urgent need. The application of mesenchymal stem cells (MSCs) derived exosomes (MSCs-Exo) as a novel therapy for neurodegenerative diseases holds great promise. A growing body of data now suggests that an innovative cell-free therapy, MSCs-Exo, may establish a fascinating alternative therapy due to their unique advantages over MSCs. Notable, MSCs-Exo can infiltrate the blood-brain barrier and then well distribute non-coding RNAs into injured tissues. Research shows that non-coding RNAs of MSCs-Exo are vital effectors that participate in the treatment of neurodegenerative diseases through neurogeneration and neurite outgrowth, modulation of the immune system, reducing neuroinflammation, repairmen of damaged tissue, and promotion of neuroangiogenesis. In addition, MSCs-Exo can serve as a drug delivery system for delivering non-coding RNAs to neurons in neurodegenerative conditions. In this review, we summarize the recent progress in the therapeutic role of non-coding RNAs of MSCs-Exo for various neurodegenerative diseases. This study also discusses the potential drug delivery role of MSCs-Exo and challenges and opportunities in the clinical translation of MSCs-Exo-based therapies for neurodegenerative diseases in the future.

Nitrogen and copper-doped saffron-based carbon dots: Synthesis, characterization, and cytotoxic effects on human colorectal cancer cells.

AIM: Doped carbon dots (CDs) have attracted tremendous attention in cancer therapy. We aimed to synthesize copper, nitrogen-doped carbon dots (Cu, N-CDs) from saffron and investigated their effects on HCT-116 and HT-29 colorectal cancer (CRC) cells. MAIN METHODS: CDs were synthesized by hydrothermal method and characterized by transmission electron microscopy (TEM), energy-dispersive X-ray (EDX), Fourier transform infrared (FT-IR) spectroscopy, ultraviolet-visible (UV-Vis) absorption spectroscopy, and fluorescence spectroscopy. HCT-116 and HT-29 cells were incubated with saffron, N-CDs, and Cu, N-CDs for 24 and 48 h for cell viability. Cellular uptake and intracellular reactive oxygen species (ROS) were evaluated by immunofluorescence microscopy. Oil Red O staining was used to monitor lipid accumulation. Apoptosis was evaluated using acridine orange/propidium iodide (AO/PI) staining and quantitative real-time polymerase chain reaction (Q-PCR) assay. The expression of miRNA-182 and miRNA-21 was measured by Q-PCR, while the generation of nitric oxide (NO) and lysyl oxidase (LOX) activity was calculated by colorimetric methods. KEY FINDINGS: CDs were successfully prepared and characterized. Cell viability decreased in the treated cells dose- and time-dependently. HCT-116 and HT-29 cells uptook Cu, N-CDs with a high level of ROS generation. The Oil Red O staining showed lipid accumulation. Concomitant with an up-regulation of apoptotic genes (p < 0.05), AO/PI staining showed increased apoptosis in the treated cells. In comparison to control cells, NO generation, and miRNA-182 and miRNA-21 expression significantly changed in the Cu, N-CDs treated cells (p < 0.05). SIGNIFICANCE: The results indicated that Cu, N-CDs could inhibit CRC cells through the induction of ROS generation and apoptosis.

Type 2 diabetes mellitus induced autophagic response within pulmonary tissue in the rat model.

Introduction: The current experiment aimed to address the impact of type 2 diabetes mellitus on autophagy status in the rat pulmonary tissue. Methods: In this study, 20 male Wistar rats were randomly allocated into two groups as follows: control and diabetic groups. To induce type 2 diabetes mellitus, rats received a combination of streptozotocin (STZ) and a high-fat diet. After confirmation of diabetic condition, rats were maintained for 8 weeks and euthanized for further analyses. The pathological changes were assessed using H&E staining. We also measured transforming growth factor-ß (TGF-ß), bronchoalveolar lavage fluid (BALF), and tumor necrosis factor-α (TNF-α) in the lungs using ELISA and real-time PCR analyses, respectively. Malondialdehyde (MDA) and superoxide dismutase (SOD) levels were monitored in diabetic lungs to assess oxidative status. We also measured the expression of becline-1, LC3, and P62 to show autophagic response under diabetic conditions. Using immunofluorescence staining, protein levels of LC3 was also monitored. Results: H&E staining showed pathological changes in diabetic rats coincided with the increase of TNF-α (~1.4-fold) and TGF-ß (~1.3-fold) compared to those in the normal rats (P<0.05). The levels of MDA (5.6 ± 0.4 versus 6.4 ± 0.27 nM/mg protein) were increased while SOD (4.2 ± 0.28 versus 3.8 ± 0.13 U/mL) activity decreased in the diabetic rats (P<0.05). Real-time polymerase chain reaction (PCR) analysis showed the up-regulation of Becline-1 (~1.35-fold) and LC3 (~2-fold) and down-regulation of P62 (~0.8-fold) (P<0.05), showing incomplete autophagic flux. We noted the increase of LC3+ cells in diabetic condition compared to that in the control samples. Conclusion: The prolonged diabetic condition could inhibit the normal activity of autophagy flux, thereby increasing pathological outcomes.

Type 2 diabetes mellitus stimulated pulmonary vascular inflammation and exosome biogenesis in rats.

It has been shown that type 2 Diabetes Mellitus (T2DM) changes the paracrine activity of several cell types. Whether the biogenesis of exosomes is changed during diabetic conditions is the subject of debate. Here, we investigated the effect of T2M on exosome biogenesis in rat pulmonary tissue. Rats received a high-fat diet regime and a single low dose of Streptozocin to mimic the T2DM-like condition. A total of 8 weeks after induction of T2DM, rats were subjected to several analyses. Besides histological examination, vascular cell adhesion molecule 1 (VCAM-1) levels were detected using immunohistochemistry (IHC) staining. Transcription of several genes such as IL-1ß, Alix, and Rab27b was calculated by real-time polymerase chain reaction assay. Using western blot analysis, intracellular CD63 levels were measured. The morphology and exosome secretion activity were assessed using acetylcholinesterase (AChE) assay and scanning electron microscopy, respectively. Histological results exhibited a moderate-to-high rate of interstitial pneumonia with emphysematous changes. IHC staining showed an increased VCAM-1 expression in the diabetic lungs compared with the normal conditions (p < .05). Likewise, we found the induction of IL-1ß, and exosome-related genes Alix and Rab27b under diabetic conditions compared with the control group (p < .05). Along with these changes, protein levels of CD63 and AChE activity were induced upon the initiation of T2DM, indicating accelerated exosome biogenesis. Taken together, current data indicated the induction of exosome biogenesis in rat pulmonary tissue affected by T2DM. It seems that the induction of inflammatory niche is touted as a stimulatory factor to accelerate exosome secretion.

Differential Expression of Serum Exosomal miRNAs in Breast Cancer Patients and Healthy Controls.

Purpose: Breast cancer has become as a serious public health concern worldwide. Breast cancer cells release exosomes into the circulatory system, which are easily accessible for further analysis like cancer diagnosis. In this study, we aimed to investigate expression of circulating exosomal miRNAs (miRs) in the serum of individuals with breast cancer and healthy controls. Methods: Exosomes were collected from serum samples using a commercial kit and characterized by scanning electron microscopy (SEM) and flow cytometry analysis. Expression of miRs such as miR-21, miR-155, miR-182, miR-373, and miR-126 were evaluated by real-time PCR. Results: The result showed that the expression level of exosomal miR-21, miR-155, miR-182, and miR-373 in the serum of breast cancer patients was higher than of those controls (P<0.05). However, expression of miR-126 did not change between breast cancer and control individuals (P > 0.05). Conclusion: Our results showed a different miRs expression pattern between breast cancer and healthy samples, supposing potential biomarkers for breast cancer. Further studies focusing on these miRs are required to confirm our findings.

Harnessing Normal and Engineered Mesenchymal Stem Cells Derived Exosomes for Cancer Therapy: Opportunity and Challenges.

There remains a vital necessity for new therapeutic approaches to combat metastatic cancers, which cause globally over 8 million deaths per year. Mesenchymal stem cells (MSCs) display aptitude as new therapeutic choices for cancer treatment. Exosomes, the most important mediator of MSCs, regulate tumor progression. The potential of harnessing exosomes from MSCs (MSCs-Exo) in cancer therapy is now being documented. MSCs-Exo can promote tumor progression by affecting tumor growth, metastasis, immunity, angiogenesis, and drug resistance. However, contradictory evidence has suggested that MSCs-Exo suppress tumors through several mechanisms. Therefore, the exact association between MSCs-Exo and tumors remains controversial. Accordingly, the applications of MSCs-Exo as novel drug delivery systems and standalone therapeutics are being extensively explored. In addition, engineering MSCs-Exo for targeting tumor cells has opened a new avenue for improving the efficiency of antitumor therapy. However, effective implementation in the clinical trials will need the establishment of standards for MSCs-Exo isolation and characterization as well as loading and engineering methods. The studies outlined in this review highlight the pivotal roles of MSCs-Exo in tumor progression and the promising potential of MSCs-Exo as therapeutic drug delivery vehicles for cancer treatment.

Effects of crocin on T-bet/GATA-3 ratio, and miR-146a and miR-106a expression levels in lung tissue of ovalbumin-sensitized mice.

Objectives: Although various studies have revealed the beneficial effects of crocin (derived from saffron), such as anti-inflammatory, anti-cancer, antioxidant, and immune modulator, however, its exact mechanism is unknown. The present study aimed to investigate the effect of crocin on the expression ratio of T-bet/GATA-3 as an indicator of altered immune responses in the lung tissue of ovalbumin (OVA)-sensitized mice. In addition, the effect of crocin on the expression level of miR-146a and miR-106a in the lung tissue OVA-sensitized mice was investigated. Materials and Methods: Mice were randomly divided into five groups (n=6): Control; OVA, OVA + Crocin 25, OVA + Cro 50, and OVA + Cro100 groups. Crocin was administrated intraperitoneally at doses of 25, 50, and 100 mg/kg for five consecutive days. One day after asthma induction, animals were euthanized, and lungs were sampled for pathological and gene expression analysis. Results: OVA-sensitization led to increased inflammation and histopathological changes in the lung tissue of mice. In addition, GATA-3 expression increased (P<0.001) and T-bet expression decreased (P<0.001) in OVA-sensitized groups. The T-bet/GATA3 ratio was also reduced markedly in asthma groups (P<0.001). Furthermore, increased expression of miR-146a and miR-106a levels was evident in the lung tissue of OVA-sensitized mice (P<0.001 for both). Intervention with high concentrations of crocin (50 and 100 mg/kg) significantly reduced airway inflammation, GATA-3 expression, miR-146a expression, and miR-106a expression and corrected the T-bet/GATA-3 ratio (P<0.05 to P<0.001). Conclusion: Treatment with crocin led to a decrease in the severity of lung inflammation in OVA-sensitized mice, which is probably through the reduction of the T-bet/GATA-3 ratio, and mir-146a and mir-106a expression level.

Engineered extracellular vesicles: A novel platform for cancer combination therapy and cancer immunotherapy.

Extracellular vesicles (EVs), phospholipid membrane-bound vesicles, produced by most cells, contribute to cell-cell communication. They transfer several proteins, lipids, and nucleic acids between cells both locally and systemically. Owing to the biocompatibility and immune activity of EVs, therapeutic approaches using these vesicles as drug delivery systems are being developed. Different methods are used to design more effective engineered EVs, which can serve as smart tools in cancer therapy and immunotherapy. Recent progress in the field of targeted-cancer therapy has led to the gradual use of engineered EVs in combinational therapy to combat heterogeneous tumor cells and multifaceted tumor microenvironments. The high plasticity, loading ability, and genetic manipulation capability of engineered EVs have made them the ideal platforms to realize numerous combinations of cancer therapy approaches. From the combination therapy view, engineered EVs can co-deliver chemotherapy with various therapeutic agents to target tumor cells effectively, further taking part in immunotherapy-related cancer combination therapy. However, a greater number of studies were done in pre-clinical platforms and the clinical translation of these studies needs further scrutiny because some challenges are associated with the application of engineered EVs. Given the many therapeutic potentials of engineered EVs, this review discusses their function in various cancer combination therapy and immunotherapy-related cancer combination therapy. In addition, this review describes the opportunities and challenges associated with the clinical application of engineered EVs.

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.