Harnessing genetically engineered cell membrane-derived vesicles as biotherapeutics
0
Abstract
Cell membrane-derived vesicles (CMVs) are particles generated from living cells, including extracellular vesicles (EVs) and artificial extracellular vesicles (aEVs) prepared from cell membranes. CMVs possess considerable potential in drug delivery, regenerative medicine, immunomodulation, disease diagnosis, etc. owing to their stable lipid bilayer structure, favorable biocompatibility, and low toxicity. Although the majority of CMVs inherit certain attributes from the original cells, it is still difficult to execute distinct therapeutic functions, such as organ targeting, signal regulation, and exogenous biotherapeutic supplementation. Hence, engineering CMVs by genetic engineering, chemical modification, and hybridization is a promising way to endow CMVs with specific functions and open up novel vistas for applications. In particular, there is a growing interest in genetically engineered CMVs harnessed to exhibit biotherapeutics. Herein, we outline the preparation strategies and their characteristics for purifying CMVs. Additionally, we review the advances of genetically engineered CMVs utilized to target organs, regulate signal transduction, and deliver biomacromolecules and chemical drugs. Furthermore, we also summarize the emerging therapeutic applications of genetically engineered CMVs in addressing tumors, diabetes, systemic lupus erythematosus, and cardiovascular diseases.
Keywords
INTRODUCTION
CMVs are membrane particles secreted or prepared from almost all living cells, including prokaryotes and eukaryotes, which are delimited by the phospholipid bilayer membrane-bound structure[1]. Classified by source, CMVs could be divided into naturally occurring EVs and aEVs prepared from cell membranes. Given differences in derivation and biomolecular components, the EVs are classified into three main subtypes (exosomes, ectosomes, and apoptotic EVs). Exosomes are the most-studied type of EVs, and they are secreted from multivesicular bodies (MVBs) via the endosomal pathway, while ectosomes are outward shedding from the surface of cell membranes directly[2,3]. Apoptotic EVs are released as fragments of cells undergoing apoptosis[4]. EVs spontaneously sort biomacromolecule cargos such as protein and nucleic acids into nanovesicles and deliver them to recipient cells without degradation and biological barriers, interpreted as information transmitters in normal physiological function, immune response, and disease development[1,3,4]. Although the mechanisms of cargo sorting into EVs are still not fully understood, there are several instances in which EVs are utilized to load endogenous biomacromolecules including protein, DNA, and RNA[5]. The aEVs are generally obtained through manual methods such as ultrasonic extrusion in vitro, which also maintain the profile of EVs. In contrast to polymer-based nanoparticles, CMVs possess properties including stable lipid bilayer structure, favorable biocompatibility, and lower toxicity, which endow CMVs with great potential as drug carriers utilized for therapeutic purposes[5,6].
Nevertheless, natural CMVs have inherent limitations that have restrained scientists’ research and restricted the application of CMVs as biotherapeutics. For instance, the most significant constraint is yield and heterogeneity[7]. The heterogeneity of CMVs caused by different original cells, discrepant biogenetic routes, and distinction in purification strategies, posed challenges to deeply understanding the components and functional characteristics of the distinct secreted components, confounded our analyses and limited their efficiency as biotherapeutics[7,8]. Moreover, CMVs innately have biological components derived from original cells and could deliver information to recipient cells through the progress of intercellular communication[9]. Although most of CMVs inherit certain attributes of the original cells, it is still difficult for them to execute distinct therapeutic functions, such as inadequate organ targeting, insufficient signal regulation, and a lack of exogenous biological therapeutic supplementation.
Further studies have demonstrated that engineered CMVs are promising nanomedicine with a broad range of specific applications[10,11]. Through strategies including genetic engineering, click chemistry, and other engineering methods such as hybridization, sonication, and electroporation[5], CMVs’ internal and membrane surface components are optimized, thus enabling enhanced or brand-new functions. Hence, genetically engineered CMVs harnessed to exhibit biotherapeutics elicit increasing interest. Herein, we outline the preparation strategies and their characteristics for purifying CMVs. Additionally, we review the advances of genetically engineered CMVs utilized to target organs, regulate signal transduction, and deliver biomacromolecules and chemical drugs. Furthermore, we also summarize the emerging therapeutics involving genetically engineered CMVs in the treatment of tumors, diabetes, systemic lupus erythematosus, and cardiovascular diseases, which will be beneficial to a better understanding of the development and challenges of this field.
PREPARATION OF CELL MEMBRANE VESICLES
Purification methods of EVs
EVs could be generated via normal biogenesis and released to the cellular space. We have developed a better understanding of mechanisms during this process. Exosomes originate from the formation of endosomes, assumed to be a heterogeneous subgroup that are secreted via inward membrane budding of early endosomes to generate intraluminal vesicles (ILVs), the precursor of exosomes, within MVB. Original cells released ILVs to the peripheral cytoplasm space as exosomes with a scale of 30 ~ 150 nm through power-driven membrane fusion between MVB and the plasma membrane[4,7,12]. Ectosomes are directly derived through outward budding from the plasma membrane. There is more than one type of ectosome with size of 150~1000 nm[4]. Apoptotic EVs are identified as fragments released by original cells that are undergoing apoptosis, arising from the outward blebbing of the membrane, with a size range of 100 nm ~ 5 μm[13,14]. Given the existence of different subpopulations of EVs with similar morphology and overlapped size ranges, the proper isolation and purification strategies are crucial for understanding EVs’ action mechanisms and are conducive to their extended application in biomedicine. Purification techniques with unique features have been adopted to facilitate EV isolation. Given that the natural EVs’ biogenesis process occurs on the genetically engineered donor cells, the purification procedures are applicable to the genetically engineered donor cells as well [Figure 1].
Figure 1. Schematic diagram of genetically engineered modification mechanism of CMVs and series of strategies for separation and purification. (A) Strategies for modifying CMVs by genetic engineering; (B) Transfection and delivery of gene or microRNAs sequence via vectors including LNPs and vectors are core steps in the process of cell genetic engineering. Hereafter, the donor cell’s biosynthetic machinery is employed to produce specific biomacromolecules which will execute specific functions inside and outside the cell. Thus, CMVs generated from original cells were modified to load exogenous biotherapeutics and used for disease treatment through genetic engineering methods; (C) The aEVs are acquired by series of sonication and extrusion; (D-E) Ultracentrifugation depends on size or molecular weight employing a series of centrifugal forces and duration, including two types of preparative ultracentrifugation--differential ultracentrifugation and density gradient ultracentrifugation; (F) Size-Exclusion Chromatography; (G) Microfluidics-based Technologies. LNP: lipid nano particle; ILVs: intraluminal vesicles; MVBs: multivesicular body; ER: endoplasmic reticulum; CMVs: cell membrane-derived vesicles.
For method I: Ultracentrifugation. Ultracentrifugation remains the most widely employed purification strategy[15]. It briefly relies on the different size and weight characteristics of EVs for isolation, utilizing centrifugal forces and specific duration. Ultracentrifugation encompasses techniques such as differential ultracentrifugation and density gradient ultracentrifugation[16]. For differential ultracentrifugation: after removing cells and debris during a series of low-speed centrifugation processes, the samples undergo ultracentrifugation at 10,000× g to collect ectosomes and at 100,000× g to obtain exosomes. For density gradient ultracentrifugation: EVs are separated via continuous or discontinuous gradients. This method has been extensively used to purify EVs. However, it has certain limitations when isolating EVs from vesicles with comparable sedimentation velocities. Compared ultracentrifugation with density gradient ultracentrifugation, Tauro et al. revealed that all separated products contained nanovesicles with close size and similar expression profiles of protein markers on the membrane surface, but they had different relative abundances of particular proteins[17]. In addition, high-speed centrifugation typically revealed heterogeneous EV aggregates[18]. Compared to differential ultracentrifugation, density gradient centrifugation gives the purer exosome population[19].
For method II: Size-Exclusion Chromatography. EVs with similar sizes are isolated together through sequential elution from the specific column[16]. The porous gel filtration polymer as the stationary phase allows differential elution in order: bigger particles tracing fewer pores are eluted firstly, and the smaller particles follow closely behind the bigger ones[14]. In contrast with EVs isolated by ultracentrifugation, the golden standard for EV isolation, the yielded EVs using size-exclusion chromatography possess more intact biophysical properties and bear higher functionality[20]. In a study conducted by Sidhom et al., size-exclusion chromatography was reviewed for its arrestive purity, functionality, replicability, and scalability, without the specialized equipment or expertise[14]. Recently, the practice of applying size-exclusion chromatography to EV purification has remarkably increased.
For method III: Microfluidics-based Technologies. Microfluidics-based isolation technologies depend on the immunoaffinity, size, and density of EVs. These liquid biopsy systems employ a series of methods to isolate EVs, such as immunofluorescence, nanoporous membrane sieving, nanowires on microcolumns for exosome capture, acoustic-based microfluidic devices, and viscoelastic microfluidic sorting[21]. One of the advantages of using microfluidic devices is high throughput and efficiency. Microfluidics-based isolation technologies have great potential in clinical applications, although it is not a standard isolation method.
Preparation methods of aEVs
In addition to the methods of obtaining EVs, the cell membrane could also be extracted by mechanical methods to produce aEVs. A previous review noted that the first step of extracting the original cell membrane is processing the cells into a cell lysis mixture through mechanical homogenization[22].
For method I: extrusion. The original cells are successively lysed and extruded several times through polycarbonate membrane filters of different diameters using a hand-held extruder to harvest aEVs with a core-shell structure[23]. The drugs and other target biotherapeutics could be mixed with original cells in the extrusion procedure so that the cargo-loaded aEVs are collected in parallel. In an earlier study, the cell suspension was sequentially extruded three times through polycarbonate membrane filters and the drug-free or drug-loaded aEVs were acquired for carrying chemotherapeutics targeting the tumor[23]. Similarly, other studies also reported cases utilizing this approach to enable aEVs with the ability for drug delivery[24,25]. Jhan et al. reported the sequential extrusion of mixtures including the cell suspension and lipid through orifices of different diameters: 400 nm, 200 nm, and 100 nm[26]. At each step, the mixtures were squeezed more than 25 times manually. After that, the engineered extracellular vesicles were harvested to establish an efficient gene delivery system[26].
For method II: sonication. The cell homogenized mixture was generally sonicated with the following settings: 20% power, 4 sec of pulsing followed by 2 sec of pause, repeated 6 times, and then cooled on ice for 2 min. After that, gradient centrifugation was carried out to harvest high-purity cell membranes[27]. Haney et al. used the following settings: 20% power, 6 cycles, 30 sec on/off each time, lasting for 3 min, with 2 min of cooling between each cycle to load chemotherapeutics into aEVs simultaneously[25].
It is worth noting that the study by Wen et al. supports the previous studies with the notion that aEVs have comparable performance with EVs in physicochemical properties (size, morphology, and classical markers, etc.)[27]. Moreover, Chul’s research revealed that the yield of aEVs is 50-100 times more than EVs, while the cost is less than 10% of EVs[23,27]. However, extrusion and sonication have definite defects of compromising the membrane integrity. Therefore, some researchers choose to incubate the mixture at 37 °C to promote recovery of the impaired exosomal membrane structure[24].
GENETICALLY ENGINEERED STRATEGIES IN CELL MEMBRANE VESICLES
In the past decades, genetic engineering has been one of the most widely practiced engineering strategies, documented and tested to induce cells to express specific products. The emerging gene vectors for delivering nucleic acid fragments include viral and nonviral vectors[22]. Transfection and delivery of genes or RNA sequences are core steps in the process of cell genetic engineering. Hereafter, the original cell’s biosynthetic machinery is employed to produce specific biomacromolecules that will execute specific functions inside or outside the cell. Hence, genetic engineering is increasingly supposed to be highly efficient for engineering CMVs, including membrane modification and exogenous cargo loading according to requirements[28]. With genetic engineering, researchers have achieved their aim of feasibly loading certain biotherapeutics (protein, nucleic acid, chemical molecules, etc.) into original cells and then transferring them to secret EVs through the natural biogenesis process or aEVs via an artificial manufacturing process. In the subsequent sections, we focus on strategies for how CMVs may be modified to load exogenous biotherapeutics and used for disease treatment through genetic engineering methods [Table 1].
Modification characteristics and advantages of genetic engineering strategies for CMVs
| Engineered strategies | Types | Modification technique | Modified form | Advantages |
| Protein | Receptor or ligand, antibody and targeting peptide | Genetic engineering vectors | Surface labeling and loaded delivery | Natural pathways， Protective delivery， Specific Targeting |
| Chemical drug | Chemotherapeutic drugs and photosensitive materials | Incubation and targeted molecular modification on the surface | Targeting, Enhanced therapeutic efficient | |
| Nucleic acids | DNA, siRNA, miRNA and mRNA | Electroporation, electrotransfection and genetic engineering vectors | Genome editing, efficient delivery, prolong circulation |
Protein cargo loading into CMVs
In general, the biotherapeutic proteins including ligands, antibodies, and targeted peptides are primarily fused with proteins ( such as LAMP-2B, CD63, etc.) expressed on CMVs membrane surface[9,10]. Since CMVs are generated from cell membranes, the overexpressed proteins located on the cell membranes will be transferred onto them. Certainly, the aEVs prepared from the cell membrane will also display the membrane-located proteins including the overexpressed proteins. These bio-functional proteins are displayed on the CMV membrane so that these strategies could enhance the targeting and therapeutic effect for various diseases[29].
Genetic engineering protein receptor or Ligand on the surface of CMVs
Wu et al. established a stable cell line with overexpression of the central nervous system (CNS) lesion -targeting ligand PDGFRα and collected the EVs[30]. Combined with the utilization of Bryostatin-1 (Bryo-1), the microglia pro-inflammatory phenotype in the CNS is dramatically altered and the clinical disease development of EAE mice is significantly ameliorated[30]. Tyrosine phosphatase-2 (SHP2) highly expressed EVs derived from MSCs were utilized to penetrate the blood-brain barrier, induce mitophagy in neuronal cells, and diminish their apoptosis, contributing to the treatment of Alzheimer's disease (AD)[31]. The oncolytic adenoviruses camouflaged by CMVs harboring the targeting ligands achieve the purpose of suppressing the intracorporal immune responses against oncolytic adenoviruses (OA) and enhance OA’s targeting capabilities to a greater extent[32,33]. The genetically modified genes of targeted protein ligands, including human epidermal growth factor (hEGF) and anti-HER2 Affibody, were introduced into cells to collect CMVs which presented tumor-targeting ligands on the membrane. The CMVs exhibit enhanced targeting capability via excellent ligand-mediated affinity to epidermal growth factor receptor (EGFR)or HER2 on the tumor surface[33].
Single protein modification for original cells or CMVs is often of limited benefit. Thus, some reports have carried out combined modifications of multiple functional protein types for disease treatment. CMVs based on the genetically engineered dendritic cell were primarily created as a cancer nanovaccine. Dendritic cells, which were infected by recombinant adenovirus displayed the specific peptide-major histocompatibility complex class I (pMHC-I), anti-programmed death 1 (PD-1) antibody, and B7 costimulatory molecules. In this way, the CMVs distinctly improved antigen delivery and triggered a wider range of T-cell immune responses for established tumors[34].
Overexpressing Antibody and Nanobody on CMVs
Antibody-drug conjugates (ADCs) act to target the delivery of drugs to specific tissues through high-affinity binding between antigens and antibodies, which significantly contributes to the precise delivery of biotherapeutics in many diseases. However, limited drug release efficiency and drug inactivation during drug delivery are prompting increasing concerns[35]. Engineered EVs may circumvent these problems because they can encapsulate the chemical drugs and protein drugs within or on the membranes of EVs. A study explores the synthetic multivalent antibodies retargeted exosomes (SMART-exosome), expressing antibodies specific for CD3 and the EGFR. The cross-linking between T lymphocytes and EGFR-positive cancer cells was promoted. Additionally, the SMART-exosome has a good performance in provoking intensive anticancer immunity via recruiting and activating cytotoxic T cells[36]. Similarly, another recent study by the same research team suggested that genetically engineered exosomes armed with not only CD3, EGFR but also the programmed death 1 and OX40 ligand (OX40L) obviously killed the EGFR-positive cancer cells and inhibited progression of established tumors[37]. Xue et al. established aEVs displaying anti-PD-1 single-chain variable fragment antibody (aPD-1-scFv) to counteract immune resistance mediated by the PD-1/PD-L1 axis in cancer[38]. The CPI-444 was loaded with aPD-1-scFv aEVs to simultaneously antagonize adenosine[38]. The CPI-444-aPD-1-scFv aEVs not only activated T cells to exhibit the capability of antitumor but also intensively increased the infiltrating T cells density, which inhibited the tumor progression and metastasis[38].
Displaying targeting peptides on the surface of CMVs
Functional peptides have been synthesized through various peptide design technologies, which are utilized widely in the production of therapeutic agents for targeted therapy[39]. Surface presentation of targeting peptides on CMVs can be realized by the sequence expression of targeting peptides fused to transmembrane proteins of CMVs[35]. In particular, cell-specific targeting peptides can be genetically engineered at the N-terminus of LAMP-2B. The αγ integrin-specific iRGD peptide (CRGDKGPDC) was displayed on exosomes via fusion expression with LAMP-2B, and it enhanced the targeting efficiency of doxorubicin (DOX) to cancer cells[40]. Similarly, KRAS small interfering RNA (siRNA) was delivered specifically to tumors via fusion expression between iRGD and LAMP-2B. The results indicated that these engineered EVs led to KRAS gene expression down-regulation and tumor growth halt[41]. In addition, other types of fusion proteins have been reported and applied. For example, Zhu et al. prepared the c(RGDyK)-modified and paclitaxel (PTX)-loaded ESC-exosomes, and confirmed they have much better performance in improving the therapeutic activity of PTX via enhanced targeting capability vs. the free drug alone in glioblastoma (GBM)[42]. In addition, another study established a “production center” based on synovial mesenchymal stromal cells (SMSCs) that produced CRY2-ZEB1 and CIBN-CD9, two recombinant fusion proteins, which were used to generate ZEB1-loaded EVs with c(RGDfC) surface modification simultaneously for improving the bone defect regeneration[43].
CMVs loading and target delivery of chemical drug
CMVs-mediated drug delivery has the advantages of favorable biocompatibility and lower toxicity in vivo, which endow CMVs with great potential as carrier systems to improve drug stability in circulation and ameliorate drug accumulation in recipient sites. A study developed exosomes that target sigma receptors and are loaded with PTX, combined with an aminoethyl ethanolamine-polyethylene glycol (AA-PEG) carrier. The AA-PEG-exoPTX significantly accumulated where the tumor cells have been established and improved the drug’s therapeutic outcomes[44]. Ultrasonication was utilized to collect genetically engineered CMVs, sorting the photosensitizer (ICG) or DOX. The drug-loaded CMVs have a better performance on antitumor progression[33]. The ExoSTING is derived from EVs that were engineered to exogenously carry cyclic dinucleotide (CDN), and it was proved that it can enhance the potency of CDN and significantly activate antigen-presenting cells[45].
Engineering CMVs sequester and delivery of nucleic acid cargo
Transfection of exogenous DNA into CMVs
Transfection of recipient cells by plasmid vectors containing specific DNA sequences is a traditional genetic engineering method to achieve direct gene delivery, further fundamentally changing certain biological properties of recipient cells. CMVs could also be genetically modified via this approach. Huang et al. genetically modified human bone marrow-derived MSCs (HMSCs) for expressing bone morphogenetic protein 2 (BMP2), and they proved that transfected cells produced EVs that potentiated the BMP2 signaling cascade and thus enhanced osteoinductive properties[46]. Research indicated that the myotube formation of sarcoblast was promoted while downregulating the expression of fibrotic genes with the delivery of miR29-EVs. In tibialis anterior (TA) muscle injury model, the miR29-EVs conduced to the consolidation of primary myoblasts and host muscle[47].
CMVs were also designed to enhance the CRISPR/Cas9 system for gene delivery. McAndrews et al. demonstrated that CRISPR/Cas9-CMVs can target the KrasG12D and successfully suppress the proliferation and growth of tumor[48]. Gee et al. have developed an all-in-one CRISPR/Cas9 ribonucleoprotein delivery platform (NanoMEDIC) using EVs, which were efficiently involved in genome editing in various hard-to-transfect cells[49]. The com-com interaction occurs by forming a ternary complex consisting of CD63-Com fusion protein, com-modified sgRNA, and Cas9 or ABE, enriching Cas9 and Adenine Base Editor (ABE) RNPs into EVs. These RNP-enriched EVs exhibit high-performance genome editing capabilities and can be instantly expressed for high-efficiency and safe CRISPR genome editing[50].
Introduction of RNA into CMVs
Recently, CMVs have emerged as a promising carrier system for transporting therapeutic nucleic acids in multiple disease models. RNA (including mRNA, miRNA, siRNA, and other non-coding RNA) is a kind of favorable cargo loaded into CMVs in order to realize certain curative effects. With the utilization of electroporation, various targeting RNA are transfected into donor cells or secreted EVs. Moreover, the aim of RNA precision delivery in vivo is achieved by designing a targeting system, such as that reviewed above. For example, synthesized siRNA as therapeutic is camouflaged into targeting tLyp-1 exosomes that are transfected by constructed tLyp-1-LAMP-2B plasmids in advance. After that, lung cancer or cancer stem cells uptook the gathered exosomes and synthesized siRNA working to knock down the SOX2 gene[51]. In another study, Bellavia et al. engineered HEK293T cells to display the LAMP-2B fused with Interleukin 3 (IL3) segment, and collected the IL3-LAMP-2B exosomes to load with Imatinib or with BCR-ABL siRNA, targeting CML cells and inhibiting cancer progression in vitro and in vivo[52]. Lu et al. designed a small interfering RNA against PAK4 (siPAK4) and drove them to form the nanocomplex core assembling with a photoactivatable ROS-sensitive polymer, which is further encapsulated by EVs from M1 macrophages through reaped extrusion[53]. The delivery of siPAK4 breached the defense of tumor-cell-intrinsic “guard” PAK4, boosted intratumoral infiltration, and elicited robust anticancer immunity[53]. A multifunctional biomimetic nanoplatform that combines small interfering RNA against PD-L1 (PD-L1 siRNA), Ru-TePt nanorods, and CMVs was designed. The anti-PD-L1-siRNA caused PD-L1 gene silencing and considerably actived cytotoxic T cells, which work synergistically with an enhanced reactive oxygen species provoked to trigger by Ru-TePt to trigger antitumor immune response. Hence, the Ru-TePt@siRNA-MVs nanosystems evoked the impressive immune response triggered by oxidative stress and suppressed immune resistance mediated by the immune checkpoint[54]. CMVs derived from MSCs encapsulated siRNAs to target the Myc and were localized to orthotopic GBM[55].
GENETICALLY ENGINEERED CELL MEMBRANE VESICLES AS DISEASE TREATMENT STRATEGY
Genetically engineered CMVs maintain the inherent advantages of native CMVs, including a stable lipid bilayer structure, favorable biocompatibility, and lower toxicity[5]. Critically, CMVs modified through genetic engineering will endow them with overexpressed protein that function as a targeting molecule, biotherapeutics, or immune modulators. Still not only such, in consideration of modification strategies’ diversity, genetically engineered cell membrane vesicles exhibit distinctive advantages in the realm of chemical drug carriers. Genetically engineered cell membrane vesicles have been inarguably meaningful for disease treatment. Herein, we focus on the advanced applications of CMVs within common disease treatment strategies [Figure 2 and Table 2].
Figure 2. Applications in treatment of common diseases model with genetically engineered CMVs. Genetically engineered CMVs work through an overexpressed proteins or nucleic acid which function as a targeting molecules, biotherapeutics, or immune modulators in therapy for tumors, systemic lupus erythematosus, diabetes, and cardiovascular diseases. CMVs: cell membrane-derived vesicles.
Advanced applications of genetically engineered CMVs in cancer treatment
| Disease model | CMVs source | Types | Engineered strategy | Key features | Mechanisms | Refs |
| Immunotherapy | CTLL-2 | EVs | PD-1 | Restart the activity of CD8+ T cells and enhance the tumor elimination | Interrupt the PD-1/PD-L1 immune inhibitory axis | [61] |
| HEK 293T | aEVs | PD-1 | Reactivate CD8+ T cells and increase the extent of CD8+ in tumors | Interrupt the PD-1/PD-L1 immune inhibitory axis | [62] | |
| BMDC | EVs | aCD19 scFv, PD-1 | Accumulate in huCD19-expressing solid tumors and reverse the immune landscape | Interrupt the PD-1/PD-L1 immune inhibitory axis | [63] | |
| DC2.4 | aEVs | MHC I, a PD-1, B7 | Eliminate established tumors | Improve antigen delivery to lymphoid organs and generate broad-spectrum T-cell responses | [34] | |
| Raw264.7 | aEVs | aPD-1-scFv, CPI-444 | Activate T cells and inhibit the tumor progression and metastasis | Interrupt the PD-1/PD-L1 immune inhibitory axis | [38] | |
| HEK 293T | aEVs | CD64, aPD-L1, cyclophosphamide | Inhibit the tumor growth and prolong survival time | Interrupt the immune escape mediated by PD-L1 and control the inhibition of regulatory T cells | [64] | |
| NIH 3T3 | aEVs | IL-15/IL-15Rα, PD-1/PD-L1 inhibitor 1 | Enhance the activation and proliferation of tumor-infiltrated T cells | Interrupt the PD-1/PD-L1 immune inhibitory axis | [65] | |
| ReN cells | EVs | siPD-L1, RGDyK | Increase CD8+ T cell activity, halt tumor growth and prolong survival | Enhance the targeting efficiency of RGD-EV to murine GBM, while the loaded siRNA reverse radiation-stimulated PD-L1 expression on tumor cells | [66] | |
| Chemotherapy | imDCs | EVs | LAMP-2B, iRGD, DOX | Decelerate the tumor growth | Show excellent targeting and DOX convey capacity to αv integrin-positive cancer cells | [40] |
| HEK 293T | EVs | CL4, DARS-AS1 siRNA, DOX | Demonstrate strong anti-proliferative, anti-migratory, and pro-apoptotic effects on TNBC cells. | Suppress the TGF-β/Smad3 signaling pathway-induced autophagy and increase the sensitivity of tumor cells to DOX | [73] | |
| Raw264.7, BMM | EVs | AA-PEG vector, PTX | Superior antineoplastic effect and increase survival times | Excellent ability to target σ receptors | [33] | |
| Embryonic stem cells | EVs | c(RGDyK), PTX | Inhibit the GBM growth | Improve the curative effects of PTX in GBM via enhanced targeting | [42] | |
| Immunotherapy+ Chemotherapy | BM-MSC | EVs | galectin-9 siRNA, OXA | Reverse immunosuppressive TME, increase infiltration of antitumoral cytotoxic T lymphocytes, and increase drug accumulation in tumor site | Disrupt the galectin-9/dectin 1 axis | [74] |
| Photodynamic therapy + Chemotherapy | HEK293T | aEVs | hEGF or anti-HER2 Affibody, ICG, DOX | Improve antitumor therapeutic outcomes and reduce side effects | Enhance targeting capacity | [33] |
| Radiotherapy | MSCs | EVs | miR-34c | Inhibit tumor progression and increase the efficiency of radiotherapy | [76] | |
| NPC cells | EVs | miR-197-3p | Reduce the proliferation and migration of NPC cells, alleviate tumor growth, and enhance their radiosensitivity | Regulate AKT/mTOR phosphorylation activation and HSPA5-mediated autophagy | [77] | |
| Geng therapy | HEK 293T | EVs | SOX2 siRNA | High transfection efficiency into tumor cells | Targeting tLyp-1 exosomes are successfully engineered | [51] |
| HEK293T | EVs | LAMP-2B, IL-3, Imatinib or BCR-ABL siRNA | Target CML cells and inhibit in vitro and in vivo cancer cell growth | Enhance targeting capacity | [52] | |
| M1 macrophages | EVs | PAK4 siRNA, ROS-sensitive polymer | Robust anticancer immunity | Prime the TME through immunogenic phototherapy, boost intratumoral infiltration and immune activation | [53] |
Harnessing genetically engineered CMVs in cancer treatment
CMVs in Immunotherapy
Immune checkpoint blockade therapy reinvigorates the host’s anti-tumor immune response for cancer treatment[56]. With application of genetically engineered CMVs, we have another perspective to modulate immune checkpoint therapy for cancer treatment. Cancer cells overexpress PD-L1 on the membrane, enhancing the binding ability to effector T cells with high expression of PD-1, inducing T cell exhaustion, and contributing to tumor immune escape[57-60]. The excellent biotherapeutics delivery capacity of genetically engineered CMVs enables therapeutic biomacromolecules to be transported to target cells with reduced immunogenicity and enhanced efficiency for immune checkpoint-based immunotherapy. A recent study reported that the constructed T cell-derived EVs displaying PD-1 on the membrane surface to interrupt the PD-1/PD-L1 immune inhibitory axis and enhance tumor elimination. The EVs effectively restarted the activity of CD8+ T cells. Moreover, PD-1 EVs could even kill tumor cells and directly drive tumor regression[61]. Similarly, the genetically engineered HEK 293T cell expressing PD-1 receptors were utilized to harvest PD-1 aEVs from an earlier research. PD-1 on the surface of aEVs not only reactivated suppressed CD8+ T cells but also significantly increased the extent of CD8+ lymphocyte infiltration in the margin of tumor by specifically binding to PD-L1 on cancer cells[62]. In another independent investigation, researchers genetically modified aCD19 scFv and PD-1, incorporating them onto the surface of DCs for the extraction of bi-specific EVs. The engineered EVs accumulated within huCD19-expressing tumors, and reversed the immune landscape[63]. Moreover, CMVs could also display relevant antibodies against the PD-1/PD-L1 immune inhibitory axis to block checkpoints and avoid immune escape in cancer[34,38]. In addition, the combination application of CMVs, possessing the capability of targeted therapy with immune checkpoint inhibitors, also demonstrated significant tumor suppression effects. The combination reagent CD64-NVs-aPD-L1-CP prepared via incubating CD64-CMVs with PD-L1 antibody and cyclophosphamide, simultaneously interrupted the immune escape mediated by PD-L1 and regulated the inhibition of regulatory T cells (Tregs), suppressing the tumor growth and prolonging survival time[64]. Similarly, IL-15/IL-15R NVs packaging the PD-1/PD-L1 inhibitor 1 simultaneously enhanced the activation and proliferation of tumor-infiltrated T cells while preventing PD-L1-mediated CD8+ T cell exhaustion[65]. In addition to targeting immune cells or normal cells to overexpress PD-1 on CMVs membrane for maneuvering the antitumor effects, tumor cells can also be targeted for interfering with the expression of genes associated with tumor progression. For instance, EVs loaded with siRNA against PD-L1 show exceptional capability of recruiting tumor-associated myeloid cells, increasing CD8+ cytotoxic T cells activity, and suppressing tumor growth with the existence of RGD, which significantly enhanced the targeting efficiency to murine GBM[66].
CMVs in Chemotherapy
Chemotherapy, being one of the most pivotal strategies for cancer clinical treatment, has shown substantial efficacy in impeding tumor progression. However, decades of clinical data have proved that although chemotherapeutic agents effectively eliminate tumor cells and promote tumor regression, they inflict damage to normal cells within the body and lead to adverse effects including cytokine release syndrome, organ damage, and other cancer tumor lysis syndromes in patients, making the prognosis of patients after chemotherapy worse[67].The therapy even caused tumor spontaneous metastasis in certain types of cancer models[68-71]. In recent years, synthetic drug delivery carriers such as liposomes have enhanced the efficacy of chemotherapy drugs and alleviated their inherent toxicity[72]. Nevertheless, the potential immunogenicity, restricted release efficiency, and undesired side toxicity of liposomes cannot be ignored. Genetically engineered CMVs are expected to provide novel avenues for chemotherapy therapeutic with significantly reduced toxicity and attenuated chemotherapeutic drug resistance. Furthermore, the combination of spatially controllable targeting ability with biocompatibility ensures that genetically engineered CMVs-based drug reagents serve as a robust platform for the delivery of chemotherapy drugs. Genetically engineered EVs derived from imDCs, displaying the LAMP-2B fused with αv integrin-specific iRGD peptide (CRGDKGPDC), were concurrently loaded with DOX. The iRGD-exosomes showed the excellent targeting and DOX convey capacity to αv integrin-positive cancer cells, resulting in the deceleration of tumor growth without distinct toxicity[40]. Liu et al. developed CL4-modified exosomes in order to target DARS-AS1 siRNA and DOX to triple-negative breast cancer (TNBC) cells[73]. The downregulation of DARS-AS1 improved the susceptivity of TNBC cells to DOX, furthering the synergetic antitumor response. The study proved that the transport of DARS-AS1 via siRNA EXOs-CL4 holds significant potential as a novel DOX-resistant treatment strategy[73]. A study developed exosomes loaded with PTX, which are partially combined with incorporated aminoethylanisamide-polyethylene glycol (AA-PEG) vector to specifically bind to the sigma receptor. The AA-PEG-exoPTX significantly accumulated where the tumor cells have been established and improves the drug’s therapeutic outcomes[44]. As mentioned earlier, the cRGD-Exo-PTX was prepared and confirmed to exhibit much better performance in improving the PTX’s pharmaceutical effect via enhanced targeting capability in GBM[42].
The synergistic application of chemotherapy with other cancer therapies has sparked fresh insights into the realm of cancer treatment. For example, the bone marrow mesenchymal stem cell (BM-MSC) exosomes charged with galectin-9 siRNA and oxaliplatin (OXA) prodrug, integrated immunotherapy with chemotherapy and attained notable therapeutic efficacy in tumors. The combined carrier provoked potent antitumor responses by increasing the infiltration of antitumoral cytotoxic T lymphocytes and promoting drug accumulation at the tumor site, reversing the immunosuppressive TME[74]. In addition to the aforementioned, ultrasonication was utilized to co-encapsulate photosensitizer (ICG) and DOX into genetically engineered CMVs. The drug-loaded CMVs exhibit significantly improved antitumor therapeutic outcomes[33].
CMVs for Radiotherapy
Technological advancements over the past decades have elevated radiotherapy to become one of the most technologically intense disciplines in medicine[75]. Benefiting from this, the toxicity and adverse effects of radiation therapy have been minimized. Nevertheless, the issue of resistance following radiotherapy remains unresolved. In response to the challenge of radiation therapy resistance, scientists are exploring innovative strategies, including the application of combination therapies, the development of drugs targeting resistance mechanisms, and the use of novel adjuvant approaches including engineered exosomes. For instance, exosomes loaded with miR-34c demonstrated a strong inhibition in invasion, migration, and proliferation of nasopharyngeal carcinoma (NPC) cells. Furthermore, these engineered exosomes significantly heightened radiation-induced apoptosis in NPC cells, suppressing tumor progression and enhancing the efficacy of radiotherapy[76].Similarly, the overexpression of miR-197-3p by exosomes resulted in a discernible reduction in proliferation and migration capabilities of nasopharyngeal carcinoma (NPC) cells in vitro, along with decreased tumor growth and radioresistance in vivo. The authors substantiated that EXO-miR-197-3p effectively impeded the progression and radioresistance activation of AKT/mTOR phosphorylation and orchestrating autophagy via HSPA5 mediation[77].
CMVs for Gene therapy
Recently, considerably novel strategies for the clinical application of engineered-CMVs-based gene therapy have increasingly intrigued researchers. Genetically engineered CMVs facilitate gene therapy targeting the diseased genome with high specificity and admirable flexibility. The gene-modulating strategies primarily include improved viral vectors and therapeutic RNAs. Numerous studies have been dedicated to investigating the functions of RNAs delivered by genetically engineered CMVs. As claimed by previous results, RNAs loaded by genetically engineered CMVs indeed possessed the normal biofunction, effectively modulating expression of target genes associated with tumor development and establishment[51-53]. The specific siRNA was delivered by EVs to oncogenic KRASG12D , and the CD47 on exosomes prevented siRNA from phagocytosis by monocytes and extended the half-life of exosomes in the circulation. Thus, their combined utilization significantly inhibited the progression of pancreatic cancer[78]. Furthermore, genetically engineered CMVs-based gene therapy combined with other therapeutic strategies could trigger a more intensive antitumor immune response and improve the prognosis[46].
Genetically engineered CMVs for Autoimmune Diseases
The attacks launched by the immune system on normal components of the body cause autoimmune diseases[79]. In the past decades, autoimmune diseases have emerged as the third-largest category of chronic disease, profoundly impacting the quality of life of patients[80]. Therapies derived from CMVs have also brought new light to advancing the treatment of autoimmune diseases.
Type 1 diabetes mellitus (T1DM) treatment
Type 1 diabetes is a chronic disease caused by the autoimmune destruction of β cells responsible for insulin production in pancreas, which is mediated by autoreactive CD4+ and CD8+ T cells infiltrating the islets[81,82]. Inhibiting the autoreactive T cells to protect β-cells from damage is a promising strategy for T1D therapy. The PD-1/PD-L1 axis, widely acknowledged as the immune checkpoint signal, inhibits the activity, induces the exhaustion of T cells, and is promising to autoimmune attack in T1D. For example, New-Onset Type 1 Diabetes could be reversed by engineered immunosuppressive platelets expressing PD-L1. The engineered platelets not only may inhibit the effects of pancreatic aggressive T cells, but also increase the proportion of Tregs maintaining immune tolerance[83]. Becker et al. designed EVs to regulate T cell effects in the model of T1D. The K562, a lymphoblast cell line, was genetically engineered to express HLA-A*02 (HLA-A2) along with costimulatory CD80 and/or coinhibitory PD-L1[84]. EVs packaging PD-L1 caused an immunosuppressive response, which restrained activation and cytotoxicity of CD8+ T cells[84].
The favorable therapeutic outcome of EVs on autoimmune diseases has attracted widespread attention and may provide a novel prospective approach for T1D treatment. The exosomes secreted by adipose tissue macrophages carrying miRNAs lead to glucose intolerance and insulin resistance[85]. The adipose tissue discharged exosomes and exosomes were absorbed by mononuclear cells, leading to their differentiation into active macrophages with increased release of pro-inflammatory cytokines such as TNF-α. The activation intermediated by exosomes involves the TLR4/TRIF pathway and contributes to insulin resistance[86]. Additionally, exosomes secreted by M2 polarized macrophages from bone marrow contained miRNA that contributes to improved glucose tolerance and insulin sensitivity. Ying et al. found that miR-690 is highly expressed in M2 BMDM and has an insulin-sensitizing function. It suggested that miR-690 could be a novel insulin sensitizer in metabolic disease[87]. Conversely, exosomes produced by adipose-derived stem cells promoted insulin sensitivity and reduced inflammation by inducing an anti-inflammatory M2 phenotype. This confirmed that exosomes are pivotal in regulating immunity and maintaining metabolic equilibrium. The miRNAs carried by these exosomes modulated TGF-β and Wnt/β-catenin signaling, which provide the potential treatment sites and are vital in the progression of chronic inflammation[88]. The studies have confirmed that CMVs containing miRNAs improved insulin sensitivity and reduced the inflammatory response. These findings revealed the importance of CMVs in mediating the crosstalk between adipose tissue, inflammation, and insulin resistance.
Systemic lupus erythematosus treatment
Systemic lupus erythematosus (SLE), characterized by persistent and excessive inflammation and autoantibody production, is an autoimmune disease involving multiple systems and organs[89]. Generally, massive autoantibodies against self-antigens produced by overactive B cells will organize immune complexes with nucleic acids and complements, which accumulate in the skin, glomeruli, and other tissues, ultimately leading to the organ damage or dysfunction[90,91]. Although live MSCs have shown impressive efficacy in clinical practice as therapy, they pose challenges such as senescence, low cell survival rates, varying degrees of immune rejection, and potential carcinogenicity[92]. Due to their cell-free nature, exosomes present more advantages over MSCs while delivering comparable therapeutic effects, making them a new treatment option[93]. Hence, genetically engineered exosomes have progressively garnered the attention of researchers. For example, CD40 was presented on the surface of genetically engineered NIH 3T3 cells to harvest CD40-NVs. CD40-NVs disrupted the CD40/CD40L costimulatory signal axis on B cells, thereby inhibiting their capability to produce antibodies. Concurrently, it limited the normal formation of germinal center structure. Furthermore, this work encapsulated the immunosuppressive drug mycophenolate mofetil (MMF) into aEVs, and it indicated that the combination regent could be employed to deplete immunocytes[94]. Similarly, it is noteworthy that Xu et al. developed the PD-L1+ MSC-derived EVs (MSC-EVs-PD-L1) with immunosuppressive properties, reprogramming the local immune microenvironment proximity to an infected organism[95]. It has been confirmed that the above engineered EVs have the ability to reconstitute the inflammatory microenvironment and show promising therapeutic effects in the disease models of UC and psoriasis in mice[95]. It is believed that the application of this similar idea for the treatment of SLE will be a promising strategy.
Genetically engineered CMVs for complications of type 2 diabetes mellitus therapy
Chronic diabetic wounds due to diabetes
Type 2 diabetes mellitus (T2DM) is distinguished by hyperglycemia resulting from impaired insulin secretion and insulin resistance[96]. Chronic diabetic wounds are one of the most recognized challenging chronic diabetic complications in clinical diagnosis and therapy worldwide. The wound of diabetic patients is particularly difficult to heal, resulting in disability and high mortality, which significantly impacts the physical and mental health and treatment prognosis of patients[97-99]. The pathogenesis of this type of wound is complex, and the main feature is that continuous hyperglycemia leads to excessive oxidative stress in tissues, inhibits angiogenesis, and prolongs inflammatory response around the wound[100]. However, there is still a lack of forceful treatment within the clinical domain. Recently, more and more evidences has been reported that the delivery of engineered exosomes containing biotherapeutics is a prospective strategy for diabetic wound healing. In particular, the positive therapeutic effects of EVs secreted by MSCs (MSC-EVs) on autoimmune diseases have attracted considerable attention. This may provide a promising new strategy for T1D treatment, and the healing of chronic diabetic wounds in T2DM is no exception. MSC EVs loaded with the miR-155 inhibitor promoted keratinocyte migration and showed synergistic effects on anti-inflammatory meanwhile. Moreover, negative regulation of miR-155 enhanced collagen deposition, angiogenesis, and re-epithelialization, leading to accelerated wound healing[101]. MSCs, as original cells, are engineered via genetic modification to create MSC-EVs overexpressing the long non-coding RNA HOX transcript antisense RNA (HOTAIR). The results demonstrated that exosomes secreted with increased HOTAIR contributed to angiogenesis and wound healing in chronic wounds[102].
A recent study published by Chu et. al built miR-17-5p-overexpressing MSCs and identified the therapeutic effect of miR-17-5p on chronic diabetic wound healing. The overexpressed miR-17-5p caused Hypo-sEVs to possess the functions of targeting, inhibiting ROS/MAPK activity, and blocking neutrophil extracellular traps (NET) formation which has been increasingly recognized as an extremely unfavorable factor in diabetic wound healing. MiR-17-5p overexpression is conducive to recovering the fibroblasts’ function and alleviating ER stress mediated by NET formation, and is recognized as a promising NET-targeting treatment based on MSC-EVs in the treatment of chronic diabetic wounds[103].
Fracture risk due to diabetes
Diabetic bone marrow-derived macrophages (dBMDM), transfected with miR-144-5p inhibitor, significantly downregulated the expressive abundance of miR-144-5p in exosomes so that it rescued the adverse impact of dBMDM-exos on bone repair and regeneration. The results presented that miR-144-5p loaded into dBMDM-derived exosomes restrained osteogenesis differentiation and negatively impacted fracture healing[104]. Researchers upregulated the abundance of miR-140-3p in BMSC-Exos through genetic engineering and showed that compared with diabetes mellitus-Exos, miR-140-3p-Exos promoted the osteoblast genesis function of BMSCs by inhibiting the expression of plexin B1, thereby accelerating the diabetic bone wound healing and promoting bone regeneration[105]. Moreover, given the significance of improving the immunosuppressive microenvironment for fracture repair, strategies employing CMVs for the suppression of overactive immune cells can also be exploited[106]. For example, exosomes with enriched concentrations of PD-L1 generated by genetically modified umbilical vein endothelial cells were delivered and released to the surrounding microenvironment via injectable hydrogel. It was demonstrated that the PD-L1-enriched exosomes specifically inhibited the activity of T cells in peripheral lymphatic tissues without affecting the activation of T cells in distant immune organs. Meanwhile, it promoted MSCs’ differentiation towards osteogenesis in the presence of T cells, and promoted adequate fracture healing[107].
Genetically engineered CMVs for Cardiovascular diseases
Cardiovascular diseases remain the most prevalent cause of death and chronic disability worldwide[108]. Cell therapy has been recognized as the most potential treatment for cardiomyocyte injury. However, the mechanism underlying stem cell therapy for damaged hearts is the acute inflammatory wound healing response rejuvenating necrotic areas of the heart, which is not associated with the production of new cardiomyocytes, presumed to work via paracrine mechanisms[109,110]. It is worth emphasizing that exosomes executing unique biological functions from original cells hold potential applications im cardiovascular treatment[111]. Currently, there are more extensive reviews available regarding the functions and application of CMVs in the context of cardiovascular diseases[112-114].Genetic engineering can be employed to cargo mRNA and other types of nucleic acids into cells, where nucleic acids including miRNA or small interfering RNA, can be subsequently packaged into exosomes. Overexpressing miRNA-133 promotes the therapeutic efficacy of mesenchymal stem cells for acute myocardial infarction[115]. The miRNA-181a carried by MSC-exo exercised an influence over immune-suppressing regulation, and when combined with the targeting capability of MSC-exo, it exerted a more valid treatment effectiveness on myocardium ischemia/reperfusion (I/R) injury. Similarly, EVs enriched with miRNA-181a promoted Treg polarization of peripheral blood mononuclear cells and increased the EF of infarcted mouse hearts by 12%[116]. CD47-EVs collected from MSCs were generated by loading purified CD47-EV with miR-21a. Following intravenous administration of miR-21a-loaded CD47-EVs, their accumulation in the heart was observed. These vesicles effectively attenuated myocardial I/R apoptosis and alleviate cardiac inflammation, ultimately contributing to the restoration of cardiac function post-I/R injury[117].
CONCLUSION
In summary, we retrospect the preparation approaches, the modified strategies for genetically engineered CMVs, and their potential applications as a novel drug delivery system in treating cancer, diabetes, and other diseases. In comparison, unmodified CMVs retain certain attributes of original cells, with research primarily focused on exploring their basic biological characteristics[4,7]. Additionally, due to the presence of specific biomarkers, unmodified CMVs play a crucial role in clinical diagnosis[118]. However, unmodified CMVs lack specificity and exhibit poor targeting capacity. Engineered exosomes show promising clinical potential across diverse fields, encompassing tumors, diabetes, and cardiovascular diseases. They have demonstrated enhanced therapeutic effects and improved targeting capabilities compared to their natural counterparts[119,120]. In contrast to other engineered methods of original cell modification, genetic engineering has the advantage of serving as carriers for protein and nucleic acid therapeutics simultaneously[121,36]. Moreover, the engineered CMVs with receptors or ligands overexpressed on their surface can execute cell signal transduction functions, enabling them to regulate the cell function, activity, and even proliferation of the target receiver cells. Thus, some engineered CMVs with immune checkpoint ligands or receptors could modulate the immunocytes such as dendritic cells and effector T cells, making these CMVs efficacious biotherapeutics for treating cancer and autoimmune diseases. However, genetically modified CMVs have some limitations, such as inefficient packing and damage to the activity of donor cells. Additionally, the types of original cells that can be modified are limited. Moreover, it is essential to acknowledge that the research field of genetically engineered CMVs’ clinical application is still in the early stages without too many strategies reaching clinical transfection. The exoASO-STAT6 (CDK-004), exolL-12TM(CDK-003), and exoSTING (CDK-002) from Codiak BioSciences has entered the phase 1 clinical trial, and the clinical trial data is of great concern[122,123].
All candidate preparation methods introduce heterogeneity and result in component loss in CMVs. Thus, it must be noted that direct comparative studies about the methodology are necessary to support current conclusions. In addition, safety in clinical applications should also be a key concern in future studies. It is imperative to conduct comprehensive testing to assess potential issues, including the potential loss or alteration of the original exosome contents and the inadvertent introduction of unwanted substances. Moreover, the quantity of genetically engineered CMVs that can be obtained is limited, making large-scale production cost-prohibitive. It recommends improving cell culture conditions during the large-scale production of genetically engineered CMVs to enhance yield. Additionally, genetically engineered CMVs exhibit immunogenicity, and the current limitations associated with cell line-derived membrane vesicles constrain their application in humans. Genetically engineered CMVs, expressing specific proteins, are susceptible to clearance by the liver in vivo, also restricting their targeting capabilities for therapeutic purposes. Hence, choosing low-immunogenicity cells such as MSCs as donor cells or utilizing genetic engineering methods to knock out immunogenic factors aims to unlock the therapeutic potential for the application of genetically engineered CMVs in human disease therapy. While facing challenges such as technical hurdles in achieving optimal preparation efficiency, the necessity for clinical validation, ongoing research, and innovation offers substantial promise for the advancement of exosome-based drug delivery systems. These engineered systems have the potential to serve as efficacious strategies for enhancing disease treatment.
DECLARATIONS
Authors’ contributions
Contributed to literature retrieval, data analysis, drafting and revising the article, gave final approval of the version to be published, and agreed to be accountable for all aspects of the work: Li X, Wei Y, Zhang Z, Zhang X
Availability of data and materials
Not applicable.
Financial support and sponsorship
This work was supported by grants from Shenzhen Science and Technology Program (Grant No. RCYX20200714114643121); The National Natural Science Foundation of China (31971268, 32201084, 32371425); Science, Technology & Innovation Commission of Shenzhen Municipality (JCYJ20200109142610136, JCYJ20180507181654186, ZDSYS20220606100803007); Guangdong Basic and Applied Basic Research Foundation (2019A1515010855, 2020A1515110166); the Natural Science Foundation of Guangdong Province (No.2020A1515010802, No.2022A1515012289); University of Chinese Academy of Sciences-Shenzhen Hospital Research Funding (HRF2020004); the Health system scientific research project of Shenzhen Guangming District Science and innovation Bureau (2020R01073, 2020R01061); Fundamental Research Funds for the Central Universities (19lgzd453) Special fund for economic development of ShenZhen Guangming District (2021R01128); Doctoral personnel scientific research start-up Fund project of Guangdong Medical University (GDMUB2022037).
Conflicts of interest
All authors declared that there are no conflicts of interest.
Ethical approval and consent to participate
Not applicable.
Consent for publication
Not applicable.
Copyright
© The Author(s) 2024.
REFERENCES
1. Andaloussi S, Mäger I, Breakefield XO, Wood MJ. Extracellular vesicles: biology and emerging therapeutic opportunities. Nat Rev Drug Discov 2013;12:347-57.
2. Mathieu M, Martin-Jaular L, Lavieu G, Théry C. Specificities of secretion and uptake of exosomes and other extracellular vesicles for cell-to-cell communication. Nat Cell Biol 2019;21:9-17.
3. Rädler J, Gupta D, Zickler A, Andaloussi SE. Exploiting the biogenesis of extracellular vesicles for bioengineering and therapeutic cargo loading. Mol Ther 2023;31:1231-50.
4. Jeppesen DK, Zhang Q, Franklin JL, Coffey RJ. Extracellular vesicles and nanoparticles: emerging complexities. Trends Cell Biol 2023;33:667-81.
5. Zhang X, Zhang H, Gu J, et al. Engineered extracellular vesicles for cancer therapy. Adv Mater 2021;33:e2005709.
6. Munagala R, Aqil F, Jeyabalan J, Gupta RC. Bovine milk-derived exosomes for drug delivery. Cancer Lett 2016;371:48-61.
7. Jeppesen DK, Fenix AM, Franklin JL, et al. Reassessment of exosome composition. Cell 2019;177:428-445.e18.
8. Buzas EI. The roles of extracellular vesicles in the immune system. Nat Rev Immunol 2023;23:236-50.
9. Raposo G, Stahl PD. Extracellular vesicles - on the cusp of a new language in the biological sciences. Extracell Vesicles Circ Nucleic Acids 2023;4:240-54.
10. Dooley K, McConnell RE, Xu K, et al. A versatile platform for generating engineered extracellular vesicles with defined therapeutic properties. Mol Ther 2021;29:1729-43.
11. Liang Y, Duan L, Lu J, Xia J. Engineering exosomes for targeted drug delivery. Theranostics 2021;11:3183-95.
12. Harding C, Heuser J, Stahl P. Endocytosis and intracellular processing of transferrin and colloidal gold-transferrin in rat reticulocytes: demonstration of a pathway for receptor shedding. Eur J Cell Biol 1984;35:256-63.
13. Buzas EI, György B, Nagy G, Falus A, Gay S. Emerging role of extracellular vesicles in inflammatory diseases. Nat Rev Rheumatol 2014;10:356-64.
14. Sidhom K, Obi PO, Saleem A. A review of exosomal isolation methods: is size exclusion chromatography the best option? Int J Mol Sci 2020;21:6466.
15. Yang B, Chen Y, Shi J. Exosome biochemistry and advanced nanotechnology for next-generation theranostic platforms. Adv Mater 2019;31:e1802896.
16. Gurunathan S, Kang MH, Jeyaraj M, Qasim M, Kim JH. Review of the isolation, characterization, biological function, and multifarious therapeutic approaches of exosomes. Cells 2019;8:307.
17. Tauro BJ, Greening DW, Mathias RA, et al. Comparison of ultracentrifugation, density gradient separation, and immunoaffinity capture methods for isolating human colon cancer cell line LIM1863-derived exosomes. Methods 2012;56:293-304.
18. Linares R, Tan S, Gounou C, Arraud N, Brisson AR. High-speed centrifugation induces aggregation of extracellular vesicles. J Extracell Vesicles 2015;4:29509.
19. Van Deun J, Mestdagh P, Sormunen R, et al. The impact of disparate isolation methods for extracellular vesicles on downstream RNA profiling. J Extracell Vesicles 2014:3.
20. Mol EA, Goumans MJ, Doevendans PA, Sluijter JPG, Vader P. Higher functionality of extracellular vesicles isolated using size-exclusion chromatography compared to ultracentrifugation. Nanomedicine 2017;13:2061-5.
21. Contreras-Naranjo JC, Wu HJ, Ugaz VM. Microfluidics for exosome isolation and analysis: enabling liquid biopsy for personalized medicine. Lab Chip 2017;17:3558-77.
22. Cheng Q, Kang Y, Yao B, et al. Genetically engineered-cell-membrane nanovesicles for cancer immunotherapy. Adv Sci 2023;10:e2302131.
23. Jang SC, Kim OY, Yoon CM, et al. Bioinspired exosome-mimetic nanovesicles for targeted delivery of chemotherapeutics to malignant tumors. ACS Nano 2013;7:7698-710.
24. Fuhrmann G, Serio A, Mazo M, Nair R, Stevens MM. Active loading into extracellular vesicles significantly improves the cellular uptake and photodynamic effect of porphyrins. J Control Release 2015;205:35-44.
25. Haney MJ, Klyachko NL, Zhao Y, et al. Exosomes as drug delivery vehicles for Parkinson's disease therapy. J Control Release 2015;207:18-30.
26. Jhan YY, Prasca-Chamorro D, Palou Zuniga G, et al. Engineered extracellular vesicles with synthetic lipids via membrane fusion to establish efficient gene delivery. Int J Pharm 2020;573:118802.
27. Wen Y, Fu Q, Soliwoda A, et al. Cell-derived nanovesicles prepared by membrane extrusion are good substitutes for natural extracellular vesicles. Extracell Vesicle 2022;1:100004.
28. Li J, Sharkey CC, Wun B, Liesveld JL, King MR. Genetic engineering of platelets to neutralize circulating tumor cells. J Control Release 2016;228:38-47.
29. Lewis ND, Sia CL, Kirwin K, et al. Exosome surface display of IL12 results in tumor-retained pharmacology with superior potency and limited systemic exposure compared with recombinant IL12. Mol Cancer Ther 2021;20:523-34.
30. Wu WC, Tian J, Xiao D, et al. Engineered extracellular vesicles encapsulated Bryostatin-1 as therapy for neuroinflammation. Nanoscale 2022;14:2393-410.
31. Xu F, Wu Y, Yang Q, et al. Engineered extracellular vesicles with SHP2 high expression promote mitophagy for alzheimer's disease treatment. Adv Mater 2022;34:e2207107.
32. Lv P, Liu X, Chen X, et al. Genetically engineered cell membrane nanovesicles for oncolytic adenovirus delivery: a versatile platform for cancer virotherapy. Nano Lett 2019;19:2993-3001.
33. Zhang P, Zhang L, Qin Z, et al. Genetically engineered liposome-like nanovesicles as active targeted transport platform. Adv Mater 2018;30:1705350.
34. Liu C, Liu X, Xiang X, et al. A nanovaccine for antigen self-presentation and immunosuppression reversal as a personalized cancer immunotherapy strategy. Nat Nanotechnol 2022;17:531-40.
35. Komuro H, Aminova S, Lauro K, Harada M. Advances of engineered extracellular vesicles-based therapeutics strategy. Sci Technol Adv Mater 2022;23:655-81.
36. Cheng Q, Shi X, Han M, Smbatyan G, Lenz HJ, Zhang Y. Reprogramming exosomes as nanoscale controllers of cellular immunity. J Am Chem Soc 2018;140:16413-7.
37. Cheng Q, Dai Z, Smbatyan G, Epstein AL, Lenz HJ, Zhang Y. Eliciting anti-cancer immunity by genetically engineered multifunctional exosomes. Mol Ther 2022;30:3066-77.
38. Xue T, Zhang Z, Fang T, et al. Cellular vesicles expressing PD-1-blocking scFv reinvigorate T cell immunity against cancer. Nano Res 2022;15:5295-5304.
40. Tian Y, Li S, Song J, et al. A doxorubicin delivery platform using engineered natural membrane vesicle exosomes for targeted tumor therapy. Biomaterials 2014;35:2383-90.
41. Zhou Y, Yuan Y, Liu M, Hu X, Quan Y, Chen X. Tumor-specific delivery of KRAS siRNA with iRGD-exosomes efficiently inhibits tumor growth. ExRNA 2019;1:28.
42. Zhu Q, Ling X, Yang Y, et al. Embryonic stem cells-derived exosomes endowed with targeting properties as chemotherapeutics delivery vehicles for glioblastoma therapy. Adv Sci 2019;6:1801899.
43. Tao SC, Li XR, Wei WJ, et al. Polymeric coating on β-TCP scaffolds provides immobilization of small extracellular vesicles with surface-functionalization and ZEB1-Loading for bone defect repair in diabetes mellitus. Biomaterials 2022;283:121465.
44. Kim MS, Haney MJ, Zhao Y, et al. Engineering macrophage-derived exosomes for targeted paclitaxel delivery to pulmonary metastases: in vitro and in vivo evaluations. Nanomedicine 2018;14:195-204.
45. Jang SC, Economides KD, Moniz RJ, et al. ExoSTING, an extracellular vesicle loaded with STING agonists, promotes tumor immune surveillance. Commun Biol 2021;4:497.
46. Huang CC, Kang M, Lu Y, et al. Functionally engineered extracellular vesicles improve bone regeneration. Acta Biomater 2020;109:182-94.
47. Song Y, Li M, Lei S, et al. Silk sericin patches delivering miRNA-29-enriched extracellular vesicles-decorated myoblasts (SPEED) enhances regeneration and functional repair after severe skeletal muscle injury. Biomaterials 2022;287:121630.
48. McAndrews KM, Xiao F, Chronopoulos A, LeBleu VS, Kugeratski FG, Kalluri R. Exosome-mediated delivery of CRISPR/Cas9 for targeting of oncogenic Kras(G12D) in pancreatic cancer. Life Sci Alliance 2021;4:e202000875.
49. Gee P, Lung MSY, Okuzaki Y, et al. Extracellular nanovesicles for packaging of CRISPR-Cas9 protein and sgRNA to induce therapeutic exon skipping. Nat Commun 2020;11:1334.
50. Yao X, Lyu P, Yoo K, et al. Engineered extracellular vesicles as versatile ribonucleoprotein delivery vehicles for efficient and safe CRISPR genome editing. J Extracell Vesicles 2021;10:e12076.
51. Bai J, Duan J, Liu R, et al. Engineered targeting tLyp-1 exosomes as gene therapy vectors for efficient delivery of siRNA into lung cancer cells. Asian J Pharm Sci 2020;15:461-71.
52. Bellavia D, Raimondo S, Calabrese G, et al. Interleukin 3- receptor targeted exosomes inhibit in vitro and in vivo chronic myelogenous leukemia cell growth. Theranostics 2017;7:1333-45.
53. Lu M, Xing H, Shao W, et al. Photoactivatable silencing extracellular vesicle (pasev) sensitizes cancer immunotherapy. Adv Mater 2022;34:e2204765.
54. Wu S, Zhang J, Pan J, et al. Integrated nanorod-mediated PD-L1 downregulation in combination with oxidative-stress immunogene therapy against cancer. Adv Healthc Mater 2023;12:e2300110.
55. Haltom AR, Hassen WE, Hensel J, et al. Engineered exosomes targeting MYC reverse the proneural-mesenchymal transition and extend survival of glioblastoma. Extracell Vesicle 2022;1:100014.
56. Sun Q, Hong Z, Zhang C, Wang L, Han Z, Ma D. Immune checkpoint therapy for solid tumours: clinical dilemmas and future trends. Signal Transduct Target Ther 2023;8:320.
57. Ishida Y, Agata Y, Shibahara K, Honjo T. Induced expression of PD-1, a novel member of the immunoglobulin gene superfamily, upon programmed cell death. EMBO J 1992;11:3887-95.
58. Freeman GJ, Long AJ, Iwai Y, et al. Engagement of the PD-1 immunoinhibitory receptor by a novel B7 family member leads to negative regulation of lymphocyte activation. J Exp Med 2000;192:1027-34.
59. Latchman Y, Wood CR, Chernova T, et al. PD-L2 is a second ligand for PD-1 and inhibits T cell activation. Nat Immunol 2001;2:261-8.
60. Kornepati AVR, Vadlamudi RK, Curiel TJ. Programmed death ligand 1 signals in cancer cells. Nat Rev Cancer 2022;22:174-89.
61. Li B, Fang T, Li Y, et al. Engineered T cell extracellular vesicles displaying PD-1 boost anti-tumor immunity. Nano Today 2022;46:101606.
62. Zhang X, Wang C, Wang J, et al. PD-1 blockade cellular vesicles for cancer immunotherapy. Adv Mater 2018;30:e1707112.
63. Xu F, Jiang D, Xu J, et al. Engineering of dendritic cell bispecific extracellular vesicles for tumor-targeting immunotherapy. Cell Rep 2023;42:113138.
64. Li L, Miao Q, Meng F, et al. Genetic engineering cellular vesicles expressing CD64 as checkpoint antibody carrier for cancer immunotherapy. Theranostics 2021;11:6033-43.
65. Fang W, Li L, Lin Z, et al. Engineered IL-15/IL-15R α -expressing cellular vesicles promote T cell anti-tumor immunity. Extracell Vesicles 2023;2:100021.
66. Tian T, Liang R, Erel-Akbaba G, et al. Immune checkpoint inhibition in GBM primed with radiation by engineered extracellular vesicles. ACS Nano 2022;16:1940-53.
67. Kuderer NM, Desai A, Lustberg MB, Lyman GH. Mitigating acute chemotherapy-associated adverse events in patients with cancer. Nat Rev Clin Oncol 2022;19:681-97.
68. Keklikoglou I, Cianciaruso C, Güç E, et al. Chemotherapy elicits pro-metastatic extracellular vesicles in breast cancer models. Nat Cell Biol 2019;21:190-202.
69. Wills CA, Liu X, Chen L, et al. Chemotherapy-induced upregulation of small extracellular vesicle-associated PTX3 accelerates breast cancer metastasis. Cancer Res 2021;81:452-63.
70. Voloshin T, Alishekevitz D, Kaneti L, et al. Blocking IL1β pathway following paclitaxel chemotherapy slightly inhibits primary tumor growth but promotes spontaneous metastasis. Mol Cancer Ther 2015;14:1385-94.
71. Liu G, Chen Y, Qi F, et al. Specific chemotherapeutic agents induce metastatic behaviour through stromal- and tumour-derived cytokine and angiogenic factor signalling. J Pathol 2015;237:190-202.
72. Zhang Z, Yao S, Hu Y, Zhao X, Lee RJ. Application of lipid-based nanoparticles in cancer immunotherapy. Front Immunol 2022;13:967505.
73. Liu X, Zhang G, Yu T, et al. CL4-modified exosomes deliver lncRNA DARS-AS1 siRNA to suppress triple-negative breast cancer progression and attenuate doxorubicin resistance by inhibiting autophagy. Int J Biol Macromol 2023;250:126147.
74. Zhou W, Zhou Y, Chen X, et al. Pancreatic cancer-targeting exosomes for enhancing immunotherapy and reprogramming tumor microenvironment. Biomaterials 2021;268:120546.
75. Jaffray DA, Knaul F, Baumann M, Gospodarowicz M. Harnessing progress in radiotherapy for global cancer control. Nat Cancer 2023;4:1228-38.
76. Wan FZ, Chen KH, Sun YC, et al. Exosomes overexpressing miR-34c inhibit malignant behavior and reverse the radioresistance of nasopharyngeal carcinoma. J Transl Med 2020;18:12.
77. Jiang J, Tang Q, Gong J, et al. Radiosensitizer EXO-miR-197-3p inhibits nasopharyngeal carcinoma progression and radioresistance by regulating the AKT/mTOR axis and HSPA5-mediated autophagy. Int J Biol Sci 2022;18:1878-95.
78. Kamerkar S, LeBleu VS, Sugimoto H, et al. Exosomes facilitate therapeutic targeting of oncogenic KRAS in pancreatic cancer. Nature 2017;546:498-503.
80. Fugger L, Jensen LT, Rossjohn J. Challenges, progress, and prospects of developing therapies to treat autoimmune diseases. Cell 2020;181:63-80.
81. Foulis AK, McGill M, Farquharson MA. Insulitis in type 1 (insulin-dependent) diabetes mellitus in man--macrophages, lymphocytes, and interferon-gamma containing cells. J Pathol 1991;165:97-103.
82. Katsarou A, Gudbjörnsdottir S, Rawshani A, et al. Type 1 diabetes mellitus. Nat Rev Dis Primers 2017;3:17016.
83. Zhang X, Kang Y, Wang J, et al. Engineered PD-L1-expressing platelets reverse new-onset type 1 diabetes. Adv Mater 2020;32:1907692.
84. Becker MW, Peters LD, Myint T, et al. Immune engineered extracellular vesicles to modulate T cell activation in the context of type 1 diabetes. Sci Adv 2023;9:eadg1082.
85. Ying W, Riopel M, Bandyopadhyay G, et al. Adipose tissue macrophage-derived exosomal miRNAs can modulate in vivo and in vitro insulin sensitivity. Cell 2017;171:372-84.e12.
86. Deng ZB, Poliakov A, Hardy RW, et al. Adipose tissue exosome-like vesicles mediate activation of macrophage-induced insulin resistance. Diabetes 2009;58:2498-505.
87. Ying W, Gao H, Dos Reis FCG, et al. MiR-690, an exosomal-derived miRNA from M2-polarized macrophages, improves insulin sensitivity in obese mice. Cell Metab 2021;33:781-790.e5.
88. Ferrante SC, Nadler EP, Pillai DK, et al. Adipocyte-derived exosomal miRNAs: a novel mechanism for obesity-related disease. Pediatr Res 2015;77:447-54.
89. Li C, Ni YQ, Xu H, et al. Roles and mechanisms of exosomal non-coding RNAs in human health and diseases. Signal Transduct Target Ther 2021;6:383.
90. Kil LP, de Bruijn MJ, van Nimwegen M, et al. Btk levels set the threshold for B-cell activation and negative selection of autoreactive B cells in mice. Blood 2012;119:3744-56.
91. Jin X, Xu Q, Pu C, et al. Therapeutic efficacy of anti-CD19 CAR-T cells in a mouse model of systemic lupus erythematosus. Cell Mol Immunol 2021;18:1896-903.
92. Lee OJ, Luk F, Korevaar SS, et al. The importance of dosing, timing, and (in)activation of adipose tissue-derived mesenchymal stromal cells on their immunomodulatory effects. Stem Cells Dev 2020;29:38-48.
93. Liu YJ, Miao HB, Lin S, Chen Z. Current progress in treating systemic lupus erythematosus using exosomes/MicroRNAs. Cell Transplant 2023;32:9636897221148775.
94. Fang T, Li B, Li M, et al. Engineered cell membrane vesicles expressing CD40 alleviate system lupus nephritis by intervening B cell activation. Small Methods 2023;7:e2200925.
95. Xu F, Fei Z, Dai H, et al. Mesenchymal stem cell-derived extracellular vesicles with high PD-L1 expression for autoimmune diseases treatment. Adv Mater 2022;34:2106265.
96. DeFronzo RA, Ferrannini E, Groop L, et al. Type 2 diabetes mellitus. Nat Rev Dis Primers 2015;1:15039.
97. Dong J, Wu B, Tian W. How to maximize the therapeutic effect of exosomes on skin wounds in diabetes mellitus: Review and discussion. Front Endocrinol 2023;14:1146991.
98. Zhang P, Lu J, Jing Y, Tang S, Zhu D, Bi Y. Global epidemiology of diabetic foot ulceration: a systematic review and meta-analysis†. Ann Med 2017;49:106-16.
99. Chen Q, Chen J, Liu Y, Qi S, Huang L. Exosome-based drug delivery systems for the treatment of diabetes and its complications: current opinion. Extracell Vesicles Circ Nucleic Acids 2023;4:502-17.
101. Gondaliya P, Sayyed AA, Bhat P, et al. Mesenchymal stem cell-derived exosomes loaded with miR-155 inhibitor ameliorate diabetic wound healing. Mol Pharm 2022;19:1294-308.
102. Born LJ, Chang KH, Shoureshi P, et al. HOTAIR-loaded mesenchymal stem/stromal cell extracellular vesicles enhance angiogenesis and wound healing. Adv Healthc Mater 2022;11:e2002070.
103. Chu Z, Huang Q, Ma K, et al. Novel neutrophil extracellular trap-related mechanisms in diabetic wounds inspire a promising treatment strategy with hypoxia-challenged small extracellular vesicles. Bioact Mater 2023;27:257-70.
104. Zhang D, Wu Y, Li Z, et al. MiR-144-5p, an exosomal miRNA from bone marrow-derived macrophage in type 2 diabetes, impairs bone fracture healing via targeting Smad1. J Nanobiotechnology 2021;19:226.
105. Wang N, Liu X, Tang Z, et al. Increased BMSC exosomal miR-140-3p alleviates bone degradation and promotes bone restoration by targeting Plxnb1 in diabetic rats. J Nanobiotechnology 2022;20:97.
106. Qiu P, Li M, Chen K, et al. Periosteal matrix-derived hydrogel promotes bone repair through an early immune regulation coupled with enhanced angio- and osteogenesis. Biomaterials 2020;227:119552.
107. Lin Z, Xiong Y, Meng W, et al. Exosomal PD-L1 induces osteogenic differentiation and promotes fracture healing by acting as an immunosuppressant. Bioact Mater 2022;13:300-11.
108. Roth GA, Johnson C, Abajobir A, et al. Global, regional, and national burden of cardiovascular diseases for 10 causes, 1990 to 2015. J Am Coll Cardiol 2017;70:1-25.
109. Vagnozzi RJ, Maillet M, Sargent MA, et al. An acute immune response underlies the benefit of cardiac stem cell therapy. Nature 2020;577:405-9.
110. Bolli R, Ghafghazi S. Stem cells: cell therapy for cardiac repair: what is needed to move forward? Nat Rev Cardiol 2017;14:257-8.
111. Sahoo S, Adamiak M, Mathiyalagan P, Kenneweg F, Kafert-Kasting S, Thum T. Therapeutic and diagnostic translation of extracellular vesicles in cardiovascular diseases: roadmap to the clinic. Circulation 2021;143:1426-49.
112. Du Y, Wu L, Wang L, Reiter RJ, Lip GYH, Ren J. Extracellular vesicles in cardiovascular diseases: From pathophysiology to diagnosis and therapy. Cytokine Growth Factor Rev 2023;74:40-55.
113. Ma J, Lei P, Chen H, et al. Advances in lncRNAs from stem cell-derived exosome for the treatment of cardiovascular diseases. Front Pharmacol 2022;13:986683.
114. Lai J, Huang C, Guo Y, Rao L. Engineered extracellular vesicles and their mimics in cardiovascular diseases. J Control Release 2022;347:27-43.
115. Chen Y, Zhao Y, Chen W, et al. MicroRNA-133 overexpression promotes the therapeutic efficacy of mesenchymal stem cells on acute myocardial infarction. Stem Cell Res Ther 2017;8:268.
116. Wei Z, Qiao S, Zhao J, et al. miRNA-181a over-expression in mesenchymal stem cell-derived exosomes influenced inflammatory response after myocardial ischemia-reperfusion injury. Life Sci 2019;232:116632.
117. Wei Z, Chen Z, Zhao Y, et al. Mononuclear phagocyte system blockade using extracellular vesicles modified with CD47 on membrane surface for myocardial infarction reperfusion injury treatment. Biomaterials 2021;275:121000.
118. Puhka M, Takatalo M, Nordberg ME, et al. Metabolomic Profiling of Extracellular Vesicles and Alternative Normalization Methods Reveal Enriched Metabolites and Strategies to Study Prostate Cancer-Related Changes. Theranostics 2017;7:3824-41.
119. Zhang M, Hu S, Liu L, et al. Engineered exosomes from different sources for cancer-targeted therapy. Signal Transduct Target Ther 2023;8:124.
120. Liu G, Wu J, Chen G, Shang A. The potential therapeutic value and application prospect of engineered exosomes in human diseases. Front Cell Dev Biol 2022;10:1051380.
121. Ovchinnikova LA, Terekhov SS, Ziganshin RH, et al. Reprogramming extracellular vesicles for protein therapeutics delivery. Pharmaceutics 2021;13:768.
122. A Study of exoASO-STAT6 (CDK-004) in patients with advanced hepatocellular carcinoma (HCC) and patients with liver metastases from either primary gastric cancer or colorectal cancer (CRC). Available from: https://clinicaltrials.gov/study/NCT05375604#study-overview [Last accessed on 24 Jan 2024].
123. A phase 1/2a study of CDK-003 in patients with cutaneous T-cell lymphoma (CTCL). (Part B). Available from: https://clinicaltrials.gov/study/NCT05156229?term=NCT05156229&rank=1 [Last accessed on 24 Jan 2024].
Cite This Article
Export citation file: BibTeX | RIS
OAE Style
Li X, Wei Y, Zhang Z, Zhang X. Harnessing genetically engineered cell membrane-derived vesicles as biotherapeutics. Extracell Vesicles Circ Nucleic Acids 2024;5:44-63. http://dx.doi.org/10.20517/evcna.2023.58
AMA Style
Li X, Wei Y, Zhang Z, Zhang X. Harnessing genetically engineered cell membrane-derived vesicles as biotherapeutics. Extracellular Vesicles and Circulating Nucleic Acids. 2024; 5(1): 44-63. http://dx.doi.org/10.20517/evcna.2023.58
Chicago/Turabian Style
Li, Xiaohong, Yuting Wei, Zhirang Zhang, Xudong Zhang. 2024. "Harnessing genetically engineered cell membrane-derived vesicles as biotherapeutics" Extracellular Vesicles and Circulating Nucleic Acids. 5, no.1: 44-63. http://dx.doi.org/10.20517/evcna.2023.58
ACS Style
Li, X.; Wei Y.; Zhang Z.; Zhang X. Harnessing genetically engineered cell membrane-derived vesicles as biotherapeutics. Extracell. Vesicles. Circ. Nucleic. Acids. 2024, 5, 44-63. http://dx.doi.org/10.20517/evcna.2023.58
About This Article
Copyright
Data & Comments
Data
0
Cite This Article 6 clicks
Like This Article 6
likes
Comments
Comments must be written in English. Spam, offensive content, impersonation, and private information will not be permitted. If any comment is reported and identified as inappropriate content by OAE staff, the comment will be removed without notice. If you have any queries or need any help, please contact us at support@oaepublish.com.