Engineered virus-like particles for efficient in vivo delivery of therapeutic proteins

Cell, Год журнала: 2022, Номер 185(2), С. 250 - 265.e16

Опубликована: Янв. 1, 2022

Methods to deliver gene editing agents in vivo as ribonucleoproteins could offer safety advantages over nucleic acid delivery approaches. We report the development and application of engineered DNA-free virus-like particles (eVLPs) that efficiently package base editor or Cas9 ribonucleoproteins. By engineering VLPs overcome cargo packaging, release, localization bottlenecks, we developed fourth-generation eVLPs mediate efficient several primary mouse human cell types. Using different glycoproteins alters their cellular tropism. Single injections into mice support therapeutic levels multiple tissues, reducing serum Pcsk9 78% following 63% liver editing, partially restoring visual function a model genetic blindness. In vitro off-target from was virtually undetected, an improvement AAV plasmid delivery. These results establish promising vehicles for macromolecule combine key both viral nonviral

Язык: Английский

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CRISPR/Cas9 therapeutics: progress and prospects

Signal Transduction and Targeted Therapy, Год журнала: 2023, Номер 8(1)

Опубликована: Янв. 16, 2023

Abstract Clustered regularly interspaced short palindromic repeats (CRISPR)/CRISPR-associated protein 9 (Cas9) gene-editing technology is the ideal tool of future for treating diseases by permanently correcting deleterious base mutations or disrupting disease-causing genes with great precision and efficiency. A variety efficient Cas9 variants derivatives have been developed to cope complex genomic changes that occur during diseases. However, strategies effectively deliver CRISPR system diseased cells in vivo are currently lacking, nonviral vectors target recognition functions may be focus research. Pathological physiological resulting from disease onset expected serve as identifying factors targeted delivery targets gene editing. Diseases both varied complex, choice appropriate methods different important. Meanwhile, there still many potential challenges identified when targeting CRISPR/Cas9 treatment. This paper reviews current developments three aspects, namely, type, vector, characteristics. Additionally, this summarizes successful examples clinical trials finally describes possible problems associated applications.

Язык: Английский

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Milk exosomes-mediated miR-31-5p delivery accelerates diabetic wound healing through promoting angiogenesis

Drug Delivery, Год журнала: 2022, Номер 29(1), С. 214 - 228

Опубликована: Янв. 5, 2022

The refractory diabetic wound has remained a worldwide challenge as one of the major health problems. impaired angiogenesis phase during healing partly contributes to pathological process. MicroRNA (miRNA) is an essential regulator gene expression in crucial biological processes and promising nucleic acid drug therapeutic fields wound. However, miRNA therapies have limitations due lacking effective delivery system. In present study, we found significant reduction miR-31-5p full-thickness wounds mice compared normal mice. Further, been proven promote proliferation, migration, endothelial cells. Thus, conceived idea exogenously supplementing mimics treat We used milk-derived exosomes novel system for successfully encapsulated into milk through electroporation. Then, proved that loaded achieved higher cell uptake was able resist degradation. Moreover, our miRNA-exosomal formulation demonstrated dramatically improved functions vitro, together with promotion enhanced vivo. Collectively, data showed feasibility scalable, biocompatible, cost-effective enhance bioavailability efficacy miRNAs.

Язык: Английский

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Current Strategies for Exosome Cargo Loading and Targeting Delivery

Cells, Год журнала: 2023, Номер 12(10), С. 1416 - 1416

Опубликована: Май 17, 2023

Extracellular vesicles (EVs) such as ectosomes and exosomes have gained attention promising natural carriers for drug delivery. Exosomes, which range from 30 to 100 nm in diameter, possess a lipid bilayer are secreted by various cells. Due their high biocompatibility, stability, low immunogenicity, favored cargo carriers. The membrane of also offers protection against degradation, making them desirable candidate However, loading into remains be challenge. Despite strategies incubation, electroporation, sonication, extrusion, freeze–thaw cycling, transfection that been developed facilitate loading, inadequate efficiency still persists. This review an overview current delivery using summarizes recent approaches small-molecule, nucleic acid, protein drugs exosomes. With insights these studies, we provide ideas more efficient effective molecules

Язык: Английский

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Engineered extracellular vesicles as versatile ribonucleoprotein delivery vehicles for efficient and safe CRISPR genome editing

Journal of Extracellular Vesicles, Год журнала: 2021, Номер 10(5)

Опубликована: Март 1, 2021

Abstract Transient delivery of CRISPR‐based genome editing effectors is important to reduce off‐target effects and immune responses. Recently extracellular vesicles (EVs) have been explored for Cas9 ribonucleoprotein (RNP) delivery. However, lack mechanisms enrich RNPs into EVs limited the efficiency as a RNP vehicle. Here we describe mechanism actively EVs. We used specific interaction between RNA aptamer aptamer‐binding protein (ABP) inserted com single guide (sgRNA), fused com‐binding ABP Com both termini tetraspan CD63 that abundant in exosomes. found Com/com enriched adenine base editor (ABE) EVs, via forming three‐component complex including CD63‐Com fusion protein, com‐modified sgRNA or ABE. The are efficient transiently expressed. system capable delivering targeting multiple loci multiplex editing. In addition, from different species can be together. EV‐delivered active vivo. data show interactions utilized improved safety.

Язык: Английский

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Development of Extracellular Vesicle Therapeutics: Challenges, Considerations, and Opportunities

Frontiers in Cell and Developmental Biology, Год журнала: 2021, Номер 9

Опубликована: Сен. 20, 2021

Extracellular vesicles (EVs) hold great promise as therapeutic modalities due to their endogenous characteristics, however, further bioengineering refinement is required address clinical and commercial limitations. Clinical applications of EV-based therapeutics are being trialed in immunomodulation, tissue regeneration recovery, delivery vectors for combination therapies. Native/biological EVs possess diverse properties that offer stability facilitate crossing biological barriers molecular cargo cells, acting a form intercellular communication regulate function phenotype. Moreover, important components paracrine signaling stem/progenitor cell-based therapies, employed standalone can be used drug system. Despite remarkable utility native/biological EVs, they improved using bio/engineering approaches potential. engineered harbor specific pharmaceutical content, enhance stability, modify surface epitopes tropism targeting cells tissues vivo. Limitations currently challenging the full realization include scalability standardization generation, characterization design regulation, potency assessment, targeted delivery. The fields' utilization advanced technologies (imaging, quantitative analyses, multi-omics, labeling/live-cell reporters), biocompatible natural sources producing (plants, bacteria, milk) will play an role overcoming these Advancements EV engineering methodologies development therapeutics, revolutionizing current landscape.

Язык: Английский

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Chondrocyte-specific genomic editing enabled by hybrid exosomes for osteoarthritis treatment

Theranostics, Год журнала: 2022, Номер 12(11), С. 4866 - 4878

Опубликована: Янв. 1, 2022

Rationale: A cell-specific delivery vehicle is required to achieve gene editing of the disease-associated cells, so hereditable genome reactions are confined within these cells without affecting healthy cells. hybrid exosome-based nano-sized derived by fusion engineered exosomes and liposomes will be able encapsulate deliver CRISPR/Cas9 plasmids selectively chondrocytes embedded in articular cartilage attenuate condition damage. Methods: Chondrocyte-targeting (CAP-Exo) were constructed genetically fusing a chondrocyte affinity peptide (CAP) at N-terminus exosomal surface protein Lamp2b. Membrane CAP-Exo with formed CAP-exosomes (hybrid CAP-Exo) which used plasmids. By intra-articular (IA) administration, CAP-Exo/Cas9 sgMMP-13 entered rats damages that mimicked osteoarthritis. Results: The deep region matrix arthritic on IA delivered plasmid Cas9 chondrocytes, knocked down metalloproteinase 13 (MMP-13), efficiently ablated expression MMP-13 attenuated hydrolytic degradation extracellular proteins cartilage. Conclusion: Chondrocyte-specific knockdown mitigates or prevents rats, showing may alleviate

Язык: Английский

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Biomembrane-based nanostructures for cancer targeting and therapy: From synthetic liposomes to natural biomembranes and membrane-vesicles

Advanced Drug Delivery Reviews, Год журнала: 2021, Номер 178, С. 113974 - 113974

Опубликована: Сен. 13, 2021

Язык: Английский

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Pharmacokinetics and biodistribution of extracellular vesicles administered intravenously and intranasally to Macaca nemestrina

Journal of Extracellular Biology, Год журнала: 2022, Номер 1(10)

Опубликована: Окт. 1, 2022

Abstract Extracellular vesicles (EVs) have potential in disease treatment since they can be loaded with therapeutic molecules and engineered for retention by specific tissues. However, questions remain on optimal dosing, administration pharmacokinetics. Previous studies addressed biodistribution pharmacokinetics rodents, but little evidence is available larger animals. Here, we investigated the of Expi293F‐derived EVs labelled a highly sensitive nanoluciferase reporter (palmGRET) non‐human primate model ( Macaca nemestrina ), comparing intravenous (IV) intranasal (IN) over 125‐fold dose range. We report that administered IV had longer circulation times plasma than previously reported mice were detectable cerebrospinal fluid after 30–60 min. EV association peripheral blood mononuclear cells, especially B‐cells, was observed as early 1‐min post‐administration. detected liver spleen within 1 h administration. IN delivery minimal, suggesting pretreatment approaches may needed large Furthermore, strongly decreased repeated administration, possibly due to immune responses clear implications xenogeneic EV‐based therapeutics. hope our findings from this baseline study macaques will help inform future research development EVs.

Язык: Английский

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Engineered virus-like particles for transient delivery of prime editor ribonucleoprotein complexes in vivo

Nature Biotechnology, Год журнала: 2024, Номер 42(10), С. 1526 - 1537

Опубликована: Янв. 8, 2024

Prime editing enables precise installation of genomic substitutions, insertions and deletions in living systems. Efficient vitro vivo delivery prime components, however, remains a challenge. Here we report editor engineered virus-like particles (PE-eVLPs) that deliver proteins, guide RNAs nicking single as transient ribonucleoprotein complexes. We systematically v3 v3b PE-eVLPs with 65- to 170-fold higher efficiency human cells compared PE-eVLP construct based on our previously reported base eVLP architecture. In two mouse models genetic blindness, injections resulted therapeutically relevant levels the retina, protein expression restoration partial visual function rescue. Optimized support ribonucleoproteins, enhancing potential safety by reducing off-target obviating possibility oncogenic transgene integration.

Язык: Английский

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