Exosome-mediated RNAi of PAK4 prolongs survival of pancreatic cancer mouse model after loco-regional treatment
Autor: | Khuloud T. Al-Jamal, Farid N. Faruqu, Yau M. Lim, Paul Lavender, Kee Y. Lim, Revadee Liam-or, Adam A. Walters, Claire M. Wells, Lizhou Xu, Julie Tzu-Wen Wang, David J. Fear |
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Rok vydání: | 2020 |
Předmět: |
Biophysics
Bioengineering 02 engineering and technology Exosomes Exosome Flow cytometry Biomaterials 03 medical and health sciences Mice In vivo Cell Line Tumor medicine Animals 030304 developmental biology 0303 health sciences Gene knockdown medicine.diagnostic_test Chemistry Electroporation fungi 021001 nanoscience & nanotechnology In vitro 3. Good health Pancreatic Neoplasms p21-Activated Kinases Mechanics of Materials Ceramics and Composites NSG mouse Cancer research RNA Interference 0210 nano-technology Ex vivo |
Zdroj: | Biomaterials. 264 |
ISSN: | 1878-5905 |
Popis: | With a dismal survival rate, pancreatic cancer (PC) remains one of the most aggressive and devastating malignancies, predominantly due to the absence of a valid biomarker for diagnosis and limited therapeutic options for advanced diseases. Exosomes (Exo) as cell-derived vesicles, are widely used as natural nanocarriers for drug delivery. P21-activated kinase 4 (PAK4) is oncogenic when overexpressed, promoting cell survival, migration and anchorage-independent growth. Herein we validated PAK4 as a therapeutic target in an in vivo PC tumour mouse model using Exo-mediated RNAi following intra-tumoural administration. PC derived Exo were firstly isolated by ultracentrifugation on sucrose cushion and characterised for their surface marker expression, size, number, purity and morphology. SiRNA was encapsulated into Exo via electroporation and dual uptake of Exo and siRNA was investigated by flow cytometry and confocal microscopy. In vitro siPAK4 silencing in PC cells following uptake was assessed by flow cytometry, western blotting, and in vitro scratch assay. In vivo efficacy (tumour growth delay and mouse survival) of siPAK4 was evaluated in PC bearing NSG mouse model. Ex vivo tumours were examined using Haematoxylin and eosin (H&E) staining and immunohistochemistry. Results showed high quality PC-derived PANC-1 Exo were obtained. SiRNA was incorporated in Exo with 16.5% encapsulation efficiency. In vitro imaging confirmed Exo and siRNA co-localisation in cells. PAK4 knockdown was successful with 30 nM Exo-siPAK4 at 24 h post incubation in vitro. Intra-tumoural administration of Exo-siPAK4 (0.03 mg/kg siPAK4 and 6.1 × 1011 Exo, each dose, two doses) reduced PC tumour growth in vivo and enhanced mice survival (p |
Databáze: | OpenAIRE |
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