Optimising non-viral gene delivery in a tumour spheroid model
Abstract
Background Our current understanding of how the unique tumour microenvironment influences the efficacy of gene delivery is limited. The current investigation systematically examines the efficiency of several non-viral gene transfer agents to transfect multicellular tumour spheroids (MCTS), an in vitro model that displays a faithful three-dimensional (3D) representation of solid tumour tissue. Methods Using a luciferase reporter assay, gene transfer to MCTS was optimised for 22 kDa linear and 25 kDa branched polyethyleneimine (PEI), the cationic lipids Lipofectamine(trade mark) and DCChol : DOPE, and the physical approach of tissue electroporation. Confocal microscopy was used to take optical tissue slices to identify the tissue localisation of green fluorescent protein (GFP) reporter gene expression and the distribution of fluorescently labelled complexes. A MCTS model of quiescent tumour regions was used to establish the influence of cellular proliferation status on gene transfer efficiency. Results Of the polyplexes tested, 22 kDa linear PEI provided optimal gene delivery, with gene expression peaking at 46 h. Despite being the optimal vector tested, PEI-mediated transfection was limited to cells at the MCTS periphery. Using fluorescent PEI, it was found that complexes could only penetrate the outer 3-5 proliferating cell layers of the MCTS, sparing the deeper quiescent cells. Gene delivery in an MCTS model comprised entirely of quiescent cells demonstrated that in addition…
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Quiescent cells located at the center of 3D structures are often refractory to transfection.
“In addition, quiescent cells that are located at the center of 3D structures are often refractory to transfection (10).”
From Direct transfection of clonal organoids in Matrigel microbeads: a promising approach toward organoid-based genetic screens (2018) · cites this paper as [10]
Direct 3D transfection on already formed organoids remains challenging.
“However, direct 3D transfection on already formed organoids remains challenging.”
From Direct transfection of clonal organoids in Matrigel microbeads: a promising approach toward organoid-based genetic screens (2018) · cites this paper as [10]
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- Authors
- Mellor, H. R., Davies, L. A., Caspar, H., Pringle, C. R., Hyde, S. C., Gill, D. R., & Callaghan, R.
- Affiliations
- John Radcliffe Hospital (GB)
- Corresponding author
- Callaghan, R.
- Venue
- The Journal of Gene Medicine
- Publisher
- Wiley
- Published
- 2006-06-29
- Volume
- 8
- Issue
- 9
- Pages
- 1160-1170
- ISSN
- 1099-498X (ISSN-L)
- 1521-2254
- OpenAlex
- W2058584700
- Cited by
- 49
- References
- 44
- Citations by year
- 2023: 1 · 2022: 3 · 2021: 4 · 2020: 5 · 2019: 1 · 2018: 4 · 2017: 6 · 2016: 1 · 2015: 2 · 2014: 1 · 2013: 4 · 2012: 2
- FWCI
- 1.36
- Type
- journal article
- Language
- en
- Open access
- closed
- Locations
- The Journal of Gene Medicine (publishedVersion)
- PubMed (publishedVersion)
- Funders
- Cancer Research UK
- Topics
- Life Sciences › Biochemistry, Genetics and Molecular Biology › Molecular Biology › RNA Interference and Gene Delivery
- Physical Sciences › Materials Science › Biomaterials › Nanoparticle-Based Drug Delivery
- Life Sciences › Biochemistry, Genetics and Molecular Biology › Genetics › Virus-based gene therapy research
- Keywords
- non-viral gene delivery, tumor spheroids, polyethyleneimine, polyplexes, cationic lipids, lipofectamine, electroporation, tumor microenvironment, transfection efficiency, 3D tumor models
- MeSH terms
- Humans, Lipids, Neoplasms, Polyethyleneimine, Recombinant Proteins, Transfection, Genetic Therapy, Genes, Reporter, Gene Transfer Techniques, Electroporation, Spheroids, Cellular, Cell Line, Tumor, Cell Proliferation, Green Fluorescent Proteins
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