Cell-derived Nanovesicles Research Articles

In vivo liquid biopsy for glioblastoma malignancy by the AFM and LSPR based sensing of exosomal CD44 and CD133 in a mouse model

Glioblastoma (GBM) is the fatal brain tumor in which secreted lactate enhances the expression of cluster of differentiation 44 (CD44) and the release of exosomes, cell-derived nanovesicles (30–200 nm), and therefore promotes tumor malignant progression. This study found that lactate-driven upregulated CD44 in malignant Glioblastoma cells (GMs) enhanced the release of CD44-enriched exosomes which increased GMs' migration and endothelial cells’ tube formation, and CD44 in the secreted exosomes was sensitively detected by “capture and sensing” Titanium Nitride (TiN) - Nanoholes (NH) - discs immunocapture (TIC) - atomic force microscopy (AFM) and ultrasensitive TiN–NH-localized surface plasmon resonance (LSPR) biosensors. The limit of detection for exosomal CD44 with TIC-AFM- and TiN–NH-LSPR-biosensors was 5.29 × 10-1 μg/ml and 3.46 × 10-3 μg/ml in exosome concentration, respectively. Importantly, this work first found that label-free sensitive TiN–NH-LSPR biosensor could detect and quantify enhanced CD44 and CD133 levels in immunocaptured GMs-derived exosomes in the blood and the cerebrospinal fluid of a mouse model of GBM, supporting its potential application in a minimally invasive molecular diagnostic for GBM progression as liquid biopsy.

Logic-Gated Cell-Derived Nanovesicles via DNA-Based Smart Recognition Module.

Engineering cell-derived nanovesicles with active-targeting ligands is an important strategy to enhance the targeting efficiency. However, the enhanced binding capability to targeting cells also leads to the binding with nontarget cells that share the same biomarkers. DNA-based logic gate is a kind of molecular system that responds to chemical inputs by generating output signals, and the relationship between the input and the output is based on a certain logic. Thus, the DNA-based logic gate could provide a new approach to improve the delivery efficiency of the nanovesicle. In this work, we developed a DNA logic-gated module that coupled two tumor cell-targeting factors (e.g., low pH and a tumor cell biomarker) in a Boolean manner. Immobilization of this module on the surface of the nanovesicle enables the nanovesicle to sense tumor cell-targeting factors and regard these cues as inputs AND logic gate. With the guide of DNA-based logic gate, gold carbon dots (GCDs) encapsulated within nanovesicles were delivered into target cells, and then the intracellular redox status variation was reflected by fluorescence change of GCDs. Overall, we developed DNA logic-gated nanovesicles that contract different targeting factors into a unique tag for target cells. This facile functionalization strategy can pave the way for constructing smart nanovesicles and would broaden their application in the field of precision medicine and personalized treatment.

Emerging Role of Exosomes in Tuberculosis: From Immunity Regulations to Vaccine and Immunotherapy.

Exosomes are cell-derived nanovesicles carrying protein, lipid, and nucleic acid for secreting cells, and act as significant signal transport vectors for cell-cell communication and immune modulation. Immune-cell-derived exosomes have been found to contain molecules involved in immunological pathways, such as MHCII, cytokines, and pathogenic antigens. Tuberculosis (TB), caused by Mycobacterium tuberculosis (MTB), remains one of the most fatal infectious diseases. The pathogen for tuberculosis escapes the immune defense and continues to replicate despite rigorous and complicate host cell mechanisms. The infected-cell-derived exosomes under this circumstance are found to trigger different immune responses, such as inflammation, antigen presentation, and activate subsequent pathways, highlighting the critical role of exosomes in anti-MTB immune response. Additionally, as a novel kind of delivery system, exosomes show potential in developing new vaccination and treatment of tuberculosis. We here summarize recent research progress regarding exosomes in the immune environment during MTB infection, and further discuss the potential of exosomes as delivery system for novel anti-MTB vaccines and therapies.

Does KRAS Play a Role in the Regulation of Colon Cancer Cells-Derived Exosomes?

Exosomes are cell-derived nanovesicles, and lately, cancer-derived exosomes have been reported to carry KRAS protein, which contributes to the malignancy of many cancers. In this study, farnesylthiosalicylic acid (FTS) was used to inhibit the activities of mutated KRAS in colon cancer SW480 cells to discover the potential link between KRAS activities and cancer-derived exosomes. We observed that FTS inhibits KRAS activity in SW480 cells, but promotes their exosome production. When the exosomal proteins of SW480 cells were profiled, a total of 435 proteins were identified with 16 of them showing significant changes (greater than or equal to two-fold) in response to FTS treatment. Protein network analysis suggests KRAS inhibition may trigger stress in the cells. In addition, a high level of acetyl-coA synthetase family member 4 protein which plays an important role in colon cancer survival was identified in the exosomes secreted by FTS-treated SW480 cells. The uptake of these exosomes suppresses the growth of some cell types, but in general exosomes from FTS-treated cells enhance the recipient cell survival when compared to that of untreated cells. Together our findings suggest that FTS may trigger stress in SW480 cells, and induce more exosomes secretion as the survival messenger to mitigate the impact of KRAS inhibition in colon cancer cells.

Neural progenitor cell-derived nanovesicles promote hair follicle growth via miR-100

BackgroundAccumulating evidence shows that mesenchymal stem cell-derived extracellular vesicles (EVs) hold great promise to promote hair growth. However, large-scale production of EVs is still a challenge. Recently, exosome-mimetic nanovesicles (NV) prepared by extruding cells have emerged as an alternative strategy for clinical-scale production. Here, ReNcell VM (ReN) cells, a neural progenitor cell line was serially extruded to produce NV.ResultsReN-NV were found to promote dermal papilla cell (DPC) proliferation. In addition, in a mouse model of depilation-induced hair regeneration, ReN-NV were injected subcutaneously, resulting in an acceleration of hair follicle (HF) cycling transition at the site. The underlying mechanism was indicated to be the activation of Wnt/β-catenin signaling pathway. Furthermore, miR-100 was revealed to be abundant in ReN-NV and significantly up-regulated in DPCs receiving ReN-NV treatment. miR-100 inhibition verified its important role in ReN-NV-induced β-catenin signaling activation.ConclusionThese results provide an alternative agent to EVs and suggest a strategy for hair growth therapy.

Engineering exosomes for targeted drug delivery.

Exosomes are cell-derived nanovesicles that are involved in the intercellular transportation of materials. Therapeutics, such as small molecules or nucleic acid drugs, can be incorporated into exosomes and then delivered to specific types of cells or tissues to realize targeted drug delivery. Targeted delivery increases the local concentration of therapeutics and minimizes side effects. Here, we present a detailed review of exosomes engineering through genetic and chemical methods for targeted drug delivery. Although still in its infancy, exosome-mediated drug delivery boasts low toxicity, low immunogenicity, and high engineerability, and holds promise for cell-free therapies for a wide range of diseases.

Cell-Derived Nanovesicles as Exosome-Mimetics for Drug Delivery Purposes: Uses and Recommendations.

Cell-derived Drug Delivery Systems (DDSs), particularly exosomes, have grown in popularity and have been increasingly explored as novel DDSs, due to their intrinsic targeting capabilities. However, clinical translation of exosomes is impeded by the tedious isolation procedures and poor yield. Cell-derived nanovesicles (CDNs) have recently been produced and proposed as exosome-mimetics. Various methods for producing exosome-mimetics have been developed. In this chapter, we present a simple, efficient, and cost-effective CDNs production method that uses common laboratory equipment (microcentrifuge) and spin cups. Through a series of extrusion and size exclusion steps, CDNs are produced from in vitro cell culture and are found to highly resemble the endogenous exosomes. Thus, we envision that this strategy holds great potential as a viable alternative to exosomes in the development of ideal DDS.

Bioinspired cell-derived nanovesicles protect the heart from ischemia reperfusion injury

Abstract Introduction Exosomes have been proven to alleviate myocardial ischemia reperfusion (I/R) injury in preclinical studies. However, the laborious and low-yield production process of naturally secreted exosomes has been impeding their translation into clinical trials. Purpose We aim to develop a simple and cost-effective protocol to produce exosome mimetics, bioinspired Cell-Derived Nanovesicles (CDNs), and examine their intrinsic bioactivity in a mouse model of I/R injury. Methods CDNs were produced from human U937 monocytes using cell shearing approach and characterized by dynamic light scattering (DLS), nanoparticle tracking analysis (NTA) and transmission electron microscopy (TEM). Lipid composition of cells and CDNs was analysed by lipidomics. I/R injury was induced by transient occlusion of left coronary artery. Data was analysed with Mann-Whitney U test. P<0.05 was considered statically significant. Results We obtained 538 mg (protein content) of CDNs, or 3x109 CDNs, via cell shearing approach from 2×107 cells, approximately 15 folds of exosomes via natural secretion from the same number of cells. CDNs were 125±8 nm in diameter with negative surface charge (zeta potential −7.0±0.8) and presented as double-membranous vesicles under TEM. In vitro, CDNs showed strong antioxidant activity and could be taken up by bone marrow-derived macrophages. Following intravenous administration, as demonstrated by the IVIS Spectrum imaging system, CDNs accumulated specifically in the infarct area of the heart within 3 hours. Compared with saline treatment, CDNs reduced myocardial infarct size by 31.6% (p<0.01) after 24 hours of I/R injury. Intriguingly, CDNs generated from human mesenchymal stem cells showed similar therapeutic efficacy. Mechanistically, CDNs inhibited infiltration of inflammatory cells (macrophages) and promoted upregulation of the anti-inflammatory cytokine interleukin 10 (IL10) in the I/R injured hearts. In the blood stream, CDNs increased IL10 protein level and exerted antioxidant activity. Furthermore, CDNs reduced I/R injury-induced cell apoptosis in the myocardium. Conclusion We have established a cost-effective approach to produce exosome mimetics, bioinspired CDNs, which protect the heart from I/R injury via inhibition of inflammation, oxidative stress and cardiac cell apoptosis. CDNs have intrinsic cardioprotective capability in heart injury, comparable to exosomes. Funding Acknowledgement Type of funding source: Public Institution(s). Main funding source(s): National University Health System.

Nanomedicine for Ischemic Stroke.

Stroke is a severe brain disease leading to disability and death. Ischemic stroke dominates in stroke cases, and there are no effective therapies in clinic, partly due to the challenges in delivering therapeutics to ischemic sites in the brain. This review is focused on the current knowledge of pathogenesis in ischemic stroke, and its potential therapies and diagnosis. Furthermore, we present recent advances in developments of nanoparticle-based therapeutics for improved treatment of ischemic stroke using polymeric NPs, liposomes and cell-derived nanovesicles. We also address several critical questions in ischemic stroke, such as understanding how nanoparticles cross the blood brain barrier and developing in vivo imaging technologies to address this critical question. Finally, we discuss new opportunities in developing novel therapeutics by targeting activated brain endothelium and inflammatory neutrophils to improve the current therapies for ischemic stroke.

Umbilical Cord Mesenchymal Stem Cell-Derived Nanovesicles Potentiate the Bone-Formation Efficacy of Bone Morphogenetic Protein 2

Recombinant human bone morphogenetic protein 2 (rhBMP-2) is one of the most potent osteogenic factors used to treat bone loss. However, at higher doses, rhBMP-2 does not necessarily increase bone formation but rather increases the incidence of adverse side effects. Here, we investigated whether umbilical cord mesenchymal stem cell (UCMSC)-derived nanovesicles (NVs) further increase the in vivo bone formation at high doses of rhBMP-2. In the presence of UCMSC-derived NVs, proliferation, migration, and tube formation of human umbilical vein endothelial cells were stimulated in vitro. Furthermore, migration and osteogenesis of human bone marrow-derived mesenchymal stem cells were stimulated. To examine the efficacy of UCMSC-derived NVs on in vivo bone formation, collagen sponges soaked with rhBMP-2 and UCMSC-derived NVs were used in athymic nude mice with calvarial defects. At a high rhBMP-2 dosage (500 ng/mL), UCMSC-derived NVs significantly promoted bone formation in calvarial defects; however, the UCMSC-derived NVs alone did not induce in vivo bone formation. Our results indicate that UCMSC-derived NVs can potentiate the bone formation efficacy of rhBMP-2 at a high dosage.

Exosome mediated delivery of functional nucleic acid nanoparticles (NANPs)

RNAi-based technologies have shown biomedical potential; however, safe and efficient delivery of RNA remains a barrier for their broader clinical applications. Nucleic acid nanoparticles (NANPs) programmed to self-assemble and organize multiple therapeutic nucleic acids (TNAs) also became attractive candidates for diverse therapeutic options. Various synthetic nanocarriers are used to deliver TNAs and NANPs, but their clinical translation is limited due to immunotoxicity. Exosomes are cell-derived nanovesicles involved in cellular communication. Due to their ability to deliver biomolecules, exosomes are a novel delivery choice. In this study, we explored the exosome-mediated delivery of NANPs designed to target GFP. We assessed the intracellular uptake, gene silencing efficiency, and immunostimulation of exosomes loaded with NANPs. We also confirmed that interdependent RNA/DNA fibers upon recognition of each other after delivery, can conditionally activate NF-kB decoys and prevent pro-inflammatory cytokines. Our study overcomes challenges in TNA delivery and demonstrates future studies in drug delivery systems.

Targeted Delivery of Mesenchymal Stem Cell-Derived Nanovesicles for Spinal Cord Injury Treatment.

Due to the safety issues and poor engraftment of mesenchymal stem cell (MSC) implantation, MSC-derived exosomes have been spotlighted as an alternative therapy for spinal cord injury (SCI). However, insufficient productivity of exosomes limits their therapeutic potential for clinical application. Moreover, low targeting ability of unmodified exosomes is a critical obstacle for their further applications as a therapeutic agent. In the present study, we fabricated macrophage membrane-fused exosome-mimetic nanovesicles (MF-NVs) from macrophage membrane-fused umbilical cord blood-derived MSCs (MF-MSCs) and confirmed their therapeutic potential in a clinically relevant mouse SCI model (controlled mechanical compression injury model). MF-NVs contained larger quantity of ischemic region-targeting molecules compared to normal MSC-derived nanovesicles (N-NVs). The targeting molecules in MF-NVs, which were derived from macrophage membranes, increased the accumulation of MF-NVs in the injured spinal cord after the in vivo systemic injection. Increased accumulation of MF-NVs attenuated apoptosis and inflammation, prevented axonal loss, enhanced blood vessel formation, decreased fibrosis, and consequently, improved spinal cord function. Synthetically, we developed targeting efficiency-potentiated exosome-mimetic nanovesicles and present their possibility of clinical application for SCI.

Matrix Effect in the Isolation of Breast Cancer-Derived Nanovesicles by Immunomagnetic Separation and Electrochemical Immunosensing-A Comparative Study.

Exosomes are cell-derived nanovesicles released into biological fluids, which are involved in cell-to-cell communication. The analysis of the content and the surface of the exosomes allow conclusions about the cells they are originating from and the underlying condition, pathology or disease. Therefore, the exosomes are currently considered good candidates as biomarkers to improve the current methods for clinical diagnosis, including cancer. However, due to their low concentration, conventional procedures for exosome detection including biosensing usually require relatively large sample volumes and involve preliminary purification and preconcentration steps by ultracentrifugation. In this paper, the immunomagnetic separation is presented as an alternative method for the specific isolation of exosomes in serum. To achieve that, a rational study of the surface proteins in exosomes, which can be recognized by magnetic particles, is presented. The characterization was performed in exosomes obtained from cell culture supernatants of MCF7, MDA-MB-231 and SKBR3 breast cancer cell lines, including TEM and nanoparticle tracking analysis (NTA). For the specific characterization by flow cytometry and confocal microscopy, different commercial antibodies against selected receptors were used, including the general tetraspanins CD9, CD63 and CD81, and cancer-related receptors (CD24, CD44, CD54, CD326 and CD340). The effect of the serum matrix on the immunomagnetic separation was then carefully evaluated by spiking the exosomes in depleted human serum. Based on this study, the exosomes were preconcentrated by immunomagnetic separation on antiCD81-modified magnetic particles in order to achieve further magnetic actuation on the surface of the electrode for the electrochemical readout. The performance of this approach is discussed and compared with classical characterization methods.

Biofabrication of Cell-Derived Nanovesicles: A Potential Alternative to Extracellular Vesicles for Regenerative Medicine.

Extracellular vesicles (EVs) are mediators of intercellular communication by transferring functional biomolecules from their originating cells to recipient cells. This intrinsic ability has gained EVs increased scientific interest in their use as a direct therapeutic in the field of regenerative medicine or as vehicles for drug delivery. EVs derived from stem cells or progenitor cells can act as paracrine mediators to promote repair and regeneration of damaged tissues. Despite substantial research efforts into EVs for various applications, their use remains limited by the lack of highly efficient and scalable production methods. Here, we present the biofabrication of cell-derived nanovesicles (NVs) as a scalable, efficient, and cost-effective production alternative to EVs. We demonstrate that NVs have a comparable size and morphology as EVs, but lack standard EV (surface) markers. Additionally, in vitro uptake experiments show that human fetal cardiac fibroblast, endothelial cells, and cardiomyocyte progenitor cells internalize NVs. We observed that cardiac progenitor cell-derived NVs and EVs are capable of activating mitogen-activated protein kinase 1/2 (MAPK1/2)-extracellular signal-regulated kinase, and that both NVs and EVs derived from A431 and HEK293 cells can functionally deliver Cre-recombinase mRNA or protein to other cells. These observations indicate that NVs may have similar functional properties as EVs. Therefore, NVs have the potential to be applied for therapeutic delivery and regenerative medicine purposes.

Mesenchymal stromal cell-derived exosomes ameliorate peripheral neuropathy in a mouse model of diabetes.

Diabetic peripheral neuropathy (DPN) is one of the major complications of diabetes, which contributes greatly to morbidity and mortality. There is currently no effective treatment for this disease. Exosomes are cell-derived nanovesicles and play an important role in intercellular communications. The present study investigated whether mesenchymal stromal cell (MSC)-derived exosomes improve neurological outcomes of DPN. Exosomes were isolated from the medium of cultured mouse MSCs by ultracentrifugation. Diabetic mice (BKS.Cg-m+/+Leprdb/J, db/db) at the age of 20weeks were used as DPN models. Heterozygous mice (db/m) of the same age were used as the control. MSC-exosomes were administered weekly via the tail vein for 8weeks. Neurological function was evaluated by testing motor and sensory nerve conduction velocities, and thermal and mechanical sensitivity. Morphometric analysis was performed by myelin sheath staining and immunohistochemistry. Macrophage markers and circulating cytokines were measured by western blot and ELISA. MicroRNA (miRNA) array and bioinformatics analyses were performed to examine the exosomal miRNA profile and miRNA putative target genes involved in DPN. Treatment of DPN with MSC-exosomes markedly decreased the threshold for thermal and mechanical stimuli and increased nerve conduction velocity in diabetic mice. Histopathological analysis showed that MSC-exosomes markedly augmented the density of FITC-dextran perfused blood vessels and increased the number of intraepidermal nerve fibres (IENFs), myelin thickness and axonal diameters of sciatic nerves. Western blot analysis revealed that MSC-exosome treatment decreased and increased M1 and M2 macrophage phenotype markers, respectively. Moreover, MSC-exosomes substantially suppressed proinflammatory cytokines. Bioinformatics analysis revealed that MSC-exosomes contained abundant miRNAs that target the Toll-like receptor (TLR)4/NF-κB signalling pathway. MSC-derived exosomes alleviate neurovascular dysfunction and improve functional recovery in mice with DPN by suppression of proinflammatory genes.

Decoy exosomes as a novel biologic reagent to antagonize inflammation.

Background: Exosomes are ubiquitous naturally secreted stable nanovesicles that can be engineered to target and deliver novel therapeutics to treat a host of human diseases.Methods: We engineered the surfaces of cell-derived nanovesicles to act as decoys in the treatment of inflammation by antagonizing the major proinflammatory cytokine, tumor necrosis factor alpha (TNFα).Results: Decoy exosomes were generated by displaying the TNFα binding domain of human TNF receptor-1 (hTNFR1) on the outer surface of exosomes using stably transfected HEK293 cells. We developed an efficient method to purify the engineered exosomes from conditioned medium based on sequential centrifugation, ultrafiltration, and precipitation. We characterized decoy exosomes using immune-quantification, nanoparticle tracking analysis, and confocal microscopy to confirm that they retain the correct orientation, size, and shape of naturally produced exosomes. We demonstrated the engineered decoy exosomes specifically antagonize activities of TNFα using an inflammatory reporter cell line.Conclusions: Decoy exosomes produced in human cells serve as a novel biologic reagent for antagonizing inflammatory signaling mediated by TNFα.

Exploiting Exosomes in Cancer Liquid Biopsies and Drug Delivery.

Exosomes are cell-derived nanovesicles that transfer molecular cargo from donor to recipient cells and mediate intercellular communication. Advancement in elucidating the biological capabilities and functionalities of exosomes has revealed the striking roles of exosomes as conveyors of bioactive molecules across the biological barriers. Tumor-derived exosomes hold great promise to serve as a liquid biopsy tool for cancer diagnosis and prognosis, as large quantities of exosomes are excreted by tumor cells continuously into the circulation, carrying the molecular cargo (DNA, RNA, proteins) reflective of the genetic and signaling alterations in tumor cells. Two inherent characteristics of exosomes offer important opportunities for drug delivery: their superb transcellular permeability and biocompatibility. Exosomes are uniquely capable of encapsulating a variety of payloads and deliver them to the target tissues. This review discusses the potential of tumor-derived exosomes in cancer liquid biopsies as well as the underlying mechanisms. Furthermore, the recent progress of developing exosomes as highly versatile and efficient drug carriers is also summarized.

Red blood cell-derived nanovesicles for safe and efficient macrophage-targeted drug delivery in vivo.

Macrophage-targeted drug delivery has great therapeutic potential for the treatment of cancers and inflammatory diseases. There is also an unmet need for efficient and nontoxic means of in vivo macrophage depletion to determine the role of macrophages under normal and disease settings. Herein, we explored the potential of red blood cell (RBC)-derived nanovesicles (RDNVs) as drug nanocarriers to specifically deplete macrophages. We show that RDNVs are effective hydrophilic drug carriers and can effectively deliver drugs into macrophages both in vitro and in vivo. Nanovesicles derived from both wild-type mouse RBCs (WT-RDNVs) and CD47 KO mouse RBCs (KO-RDNVs) can encapsulate clodronate with good stability in PBS for long-term storage. However, KO-RDNVs were more efficiently engulfed by macrophages in vitro and more rapidly cleared in vivo than WT-RDNVs, indicating that CD47 also serves as a "don't eat me" molecule for RDNVs as it does for RBCs. Accordingly, clodronate-encapsulated KO-RDNVs (KO-RDNV/CLD) were significantly more toxic to mouse macrophages in vitro than drug-loaded WT-RDNVs (WT-RDNV/CLD). Furthermore, WT-RDNV/CLD showed prolonged accumulation in tissues (e.g., liver and lung) and macrophage depletion versus KO-RDNV/CLD. Importantly, RBC-derived nanovesicles are more biocompatible and less toxic in vivo than clodronate-encapsulated liposomes-the current gold-standard macrophage-depleting reagent. This study offers a useful strategy for macrophage-targeted drug delivery.

Exosomes in tuberculosis: Still terra incognita?

Today, diagnosis, vaccination, and treatment of tuberculosis (TB) remain major clinical challenges. Therefore, an introduction of new diagnostic measures and biomarkers is necessary to improve infection control. The ideal biomarker for TB infection can be defined as a host or pathogen-derived biomolecule, which is potent for identifying infection and determining its clinical stage. Exosomes, defined as cell-derived nanovesicles released into biological fluids, are involved in cell-cell communication and immune modulation. These vesicles have emerged as a new platform for improving the clinical diagnosis and prognosis of different infectious diseases and cancers. The role of these nanovehicles, as alternative biomarkers for the improvement of TB diagnosis and treatment, has been demonstrated in a significant body of literature. In this review, we summarized recent progress in the clinical application of exosome-based biomarkers in TB infection.

Cancer Cell-Derived, Drug-Loaded Nanovesicles Induced by Sulfhydryl-Blocking for Effective and Safe Cancer Therapy.

Extracellular vesicles (EVs) pose great promise as therapeutic carriers due to their ideal size range and intrinsic biocompatibility. Limited scalability, poor quality control during production, and cumbersome isolation and purification processes have caused major setbacks in the progression of EV therapeutics to the clinic. Here, we overcome these setbacks by preparing cell-derived nanovesicles induced by sulfhydryl-blocking (NIbS), in the desirable size range for therapeutic delivery, that can be further loaded with the chemotherapeutic drug, doxorubicin (DOX), resulting in NIbS/DOX. Applicable to most cell types, this chemical blebbing approach enables efficient, quick, and simple harvest and purification as well as easily scalable production. Cellular uptake and intracellular release of DOX was improved using NIbS/DOX compared to a liposomal formulation. We also confirmed that in tumor-challenged C57BL/6 mice NIbS/DOX significantly slowed tumor growth and led to improved survival compared to treatment with free drug or liposomal drug. NIbS are a promising therapeutic carrier for improving cancer treatment outcomes since they are easy to prepare at a large scale, good candidates for drug loading, and capable of efficient administration of therapeutic agents with avoided nonspecific major distribution in vital organs. In addition, the utility of NIbS can be easily expanded to immunotherapy, gene therapy, and cell therapy when they are derived from applicable cell types.