Preparation of engineered extracellular vesicles with full-length functional PD-1 membrane proteins by baculovirus expression system

Ishikawa, Raga; Yoshida, Shosuke; Sawada, Shin–ichi; Sasaki, Yoshihiro; Akiyoshi, Kazunari

doi:10.1016/j.bbrc.2020.03.187

Cited by 7 publications

(19 citation statements)

References 26 publications

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“…Recent studies indicate that these nanoparticles not only impact mammalian cell cultures, but are also important to consider in insect cells, 24 particularly in continuous operation processes aiming to achieve reproducible and constant culture conditions. EVs facilitate the intercellular communication by transporting functional molecules 42 ; however, the influence of EVs in insect cell bioprocesses is still not well understood. In this sense, it is important to consider that these nanoparticles are also present in the insect cell platform, especially when the product of interest is a nanoparticle, since not taking them into consideration might cause product overestimation.…”

Section: Discussionmentioning

confidence: 99%

Stable Sf9 cell pools as a system for rapid HIV‐1 virus‐like particle production

Puente‐Massaguer

Grau-Garcia

Strobl

et al. 2021

J of Chemical Tech & Biotech

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BACKGROUND: The emergence of infectious diseases is accelerating the intensification of bioprocess strategies to support the increasing demand for the manufacture of higher quantities of vaccines in short timeframes. Here, the development of stable Sf9 cell pools producing human immunodeficiency virus serotype 1 (HIV-1) Gag-eGFP virus-like particles (VLPs) is assessed.RESULTS: Fluorescence-activated cell sorting (FACS) was employed to select high producing cells, achieving an 8.1-fold increase in fluorescence intensity compared to unsorted cell pools after three rounds of cell sorting. The transferability of this system to bioreactor scale was also successfully achieved, attaining a 1.4-fold increase in VLP production and maintaining a higher cell viability than shake flask controls. Analysis of the metabolism of stable cell pools and parental Sf9 cells did not show significant differences regarding metabolite consumption and production, even though a better performance and more efficient metabolism were observed in bioreactor compared with shake flask cultures, highlighting the flexibility of these cells to adapt to different culture conditions and heterologous recombinant protein production.CONCLUSIONS: Stable Sf9 cell pools represent a suitable system for shortening bioprocess development times and accelerating vaccine production.

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Section: Discussionmentioning

confidence: 99%

Stable Sf9 cell pools as a system for rapid HIV‐1 virus‐like particle production

Puente‐Massaguer

Grau-Garcia

Strobl

et al. 2021

J of Chemical Tech & Biotech

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“…Sf9‐derived EVs were isolated using a previously reported method [22]. Briefly, 4.0 × 10 5 Sf9 cells·mL −1 were maintained in Sf‐900 III SFM medium (Invitrogen) overnight at 27 °C.…”

Section: Methodsmentioning

confidence: 99%

“…Recombinant baculoviruses encoding PD-1 mutant or Connexin 43 (Cx43)-enhanced green fluorescence protein (EGFP) were constructed using the Bac-to-Bac Baculovirus Expression System, as described previously [22]. Briefly, pFastBac1 plasmid carrying the membrane protein sequence of interest was transformed into DH10Bac Escherichia coli containing Bacmid and a helper plasmid encoding a transposase gene.…”

Section: Construction Of Recombinant Baculovirusesmentioning

confidence: 99%

“…influenza hemagglutinin and human T‐cell leukemia virus envelope protein) [21]. We recently developed Sf9 insect cell‐derived EVs on which functional membrane proteins were presented using a baculovirus‐expression system [22]. Programmed cell death 1 (PD‐1) membrane proteins were expressed on the EV surface while maintaining the ability to bind to ligand proteins and ligand‐expressing cancer cells.…”

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confidence: 99%

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Development and single‐particle analysis of hybrid extracellular vesicles fused with liposomes using viral fusogenic proteins

et al. 2022

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Extracellular vesicles (EVs) have potential biomedical applications, particularly as a means of transport for therapeutic agents. There is a need for rapid and efficient EV‐liposome membrane fusion that maintains the integrity of hybrid EVs. We recently described Sf9 insect cell‐derived EVs on which functional membrane proteins were presented using a baculovirus‐expression system. Here, we developed hybrid EVs by membrane fusion of small liposomes and EVs equipped with baculoviral fusogenic proteins. Single‐particle analysis of EV‐liposome complexes revealed controlled introduction of liposome components into EVs. Our findings and methodology will support further applications of EV engineering in biomedicine.

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“…It was reported that RVG-vectorized EVs loaded with glyceraldehyde 3-phosphate dehydrogenase (GAPDH) siRNA successfully delivered the therapeutic siRNA in wild-type mice to the cells of neurovascular unit and caused efficient knock down of BACE1, which represents a therapeutic target in Alzheimer's disease [114]. Ishikawa et al [148] reported vectorization of EVs to cancer tissues with programmed cell death ligand 1 (PD-L1). To modify EVs surface, the insect cell line Spodoptera frugiperda 9 was infected with recombinant baculoviruses incorporating the PD-1 mutant gene to express the target membrane protein.…”

Section: Challenges Of Insufficient Targeting Efficiency Of Evs Formulations To Disease Tissuesmentioning

confidence: 99%

Extracellular Vesicle-Based Therapeutics: Preclinical and Clinical Investigations

Klyachko

Arzt

et al. 2020

Pharmaceutics

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Drug nanoformulations hold remarkable promise for the efficient delivery of therapeutics to a disease site. Unfortunately, artificial nanocarriers, mostly liposomes and polymeric nanoparticles, show limited applications due to the unfavorable pharmacokinetics and rapid clearance from the blood circulation by the reticuloendothelial system (RES). Besides, many of them have high cytotoxicity, low biodegradability, and the inability to cross biological barriers, including the blood brain barrier. Extracellular vesicles (EVs) are novel candidates for drug delivery systems with high bioavailability, exceptional biocompatibility, and low immunogenicity. They provide a means for intercellular communication and the transmission of bioactive compounds to targeted tissues, cells, and organs. These features have made them increasingly attractive as a therapeutic platform in recent years. However, there are many obstacles to designing EV-based therapeutics. In this review, we will outline the main hurdles and limitations for therapeutic and clinical applications of drug loaded EV formulations and describe various attempts to solve these problems.

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Preparation of engineered extracellular vesicles with full-length functional PD-1 membrane proteins by baculovirus expression system

Cited by 7 publications

References 26 publications

Stable Sf9 cell pools as a system for rapid HIV‐1 virus‐like particle production

Stable Sf9 cell pools as a system for rapid HIV‐1 virus‐like particle production

Development and single‐particle analysis of hybrid extracellular vesicles fused with liposomes using viral fusogenic proteins

Extracellular Vesicle-Based Therapeutics: Preclinical and Clinical Investigations

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