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Internalisation of RGD-Engineered Extracellular Vesicles by Glioblastoma Cells

SIMPLE SUMMARY: Glioblastoma multiforme (GBM) is the most aggressive and malignant type of central nervous system (CNS) tumour. Although several treatment options are available, patients generally succumb within 14 months after diagnosis. With the rapid progression of exosome bioengineering technolo...

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Detalles Bibliográficos
Autores principales: Gečys, Dovydas, Kazlauskas, Arūnas, Gečytė, Emilija, Paužienė, Neringa, Kulakauskienė, Deimantė, Lukminaitė, Indrė, Jekabsone, Aistė
Formato: Online Artículo Texto
Lenguaje:English
Publicado: MDPI 2022
Materias:
Acceso en línea:https://www.ncbi.nlm.nih.gov/pmc/articles/PMC9598886/
https://www.ncbi.nlm.nih.gov/pubmed/36290387
http://dx.doi.org/10.3390/biology11101483
Descripción
Sumario:SIMPLE SUMMARY: Glioblastoma multiforme (GBM) is the most aggressive and malignant type of central nervous system (CNS) tumour. Although several treatment options are available, patients generally succumb within 14 months after diagnosis. With the rapid progression of exosome bioengineering technologies, novel therapy opportunities are emerging for GBM treatment. The surface of GBM cells is characterised by the overexpression of transmembrane receptor integrins, which are essential for cell interactions with several proteins in the extracellular matrix. Therefore, integrin-binding drug delivery vehicles have been proposed as a potential strategy for glioblastoma therapy. Small extracellular vesicles possess several attractive characteristics for drug delivery: small size, biocompatibility, ability to cross the blood–brain barrier and capacity to be loaded with exogenous materials. Current bioengineering technologies further increase extracellular vesicle capabilities by loading them with anticancer drugs and incorporating targeting ligands. This study explored the capacity of Arginylglycylaspartic acid (RGD, or Arginine–Glycine–Aspartate)-polypeptide-engineered extracellular vesicles to internalise and deliver loaded cargo in GBM cells. The results demonstrate that introducing the RGD ligand to extracellular vesicles could significantly increase their internalisation by GBM cells and hence improve drug delivery efficacy. ABSTRACT: Glioblastoma multiforme (GBM) is the most aggressive CNS tumour with no efficient treatment, partly due to the retention of anticancer drugs by the blood–brain barrier (BBB) and their insufficient concentration in tumour cells. Extracellular vesicles (EVs) are attractive drug carriers because of their biocompatibility and ability to cross the BBB. Additional efficiency can be achieved by adding GBM-cell-specific ligands. GBM cells overexpress integrins; thus, one of the most straightforward targeting strategies is to modify EVs with integrin-recognising molecules. This study investigated the therapeutic potential of genetically engineered EVs with elevated membrane levels of the integrin-binding peptide RGD (RGD-EVs) against GBM cells in vitro. For RGD-EV production, stable RGD-HEK 293FT cells were generated by using a pcDNA4/TO-Lamp2b-iRGD-HA expression vector and performing antibiotic-based selection. RGD-EVs were isolated from RGD-HEK 293FT-cell-conditioned medium and characterised by size (Zetasizer), specific markers (ELISA) and RGD expression (Western Blot). Internalisation by human GBM cells HROG36 and U87 MG and BJ-5ta human fibroblasts was assessed by fluorescent EV RNA labelling. The effect of doxorubicin-loaded RGD-EVs on GBM cells was evaluated by the metabolic PrestoBlue viability assay; functional GAPDH gene knockdown by RGD-EV-encapsulated siRNA was determined by RT-qPCR. RGD-EVs had 40% higher accumulation in GBM cells (but not in fibroblasts) and induced significantly stronger toxicity by loaded doxorubicin and GAPDH silencing by loaded siRNA compared to unmodified EVs. Thus, RGD modification substantially increases the specific delivery capacity of HEK 293FT-derived EVs to GBM cells.