4.6 Article

In Vitro Evaluation of Lipopolyplexes for Gene Transfection: Comparing 2D, 3D and Microdroplet-Enabled Cell Culture

期刊

MOLECULES
卷 25, 期 14, 页码 -

出版社

MDPI
DOI: 10.3390/molecules25143277

关键词

Lipopolyplex; gene transfection; multicellular tumor spheroid; single-cell microdroplets; in vitro evaluation

资金

  1. Microfluidic Layer-by-layer Assembly of Cationic Liposome -Nucleic Acid Nanoparticles for Gene Delivery project - FCT [032520]
  2. ERDF through COMPETE2020
  3. Ramon Areces Foundation [BEVP30A5827]
  4. FCT [SFRH/BD/148091/2019]
  5. Fundação para a Ciência e a Tecnologia [SFRH/BD/148091/2019] Funding Source: FCT

向作者/读者索取更多资源

Complexes combining nucleic acids with lipids and polymers (lipopolyplexes) show great promise for gene therapy since they enable compositional, physical and functional versatility to be optimized for therapeutic efficiency. When developing lipopolyplexes for gene delivery, one of the first evaluations performed is an in vitro transfection efficiency experiment. Many different in vitro models can be used, and the effect of the model on the experiment outcome has not been thoroughly studied. The objective of this work was to compare the insights obtained from three different in vitro models, as well as the potential limitations associated with each of them. We have prepared a series of lipopolyplex formulations with three different cationic polymers (poly-l-lysine, bioreducible poly-l-lysine and polyethyleneimine), and assessed their in vitro biological performance in 2D monolayer cell culture, 3D spheroid culture and microdroplet-based single-cell culture. Lipopolyplexes from different polymers presented varying degrees of transfection efficiency in all models. The best-performing formulation in 2D culture was the polyethyleneimine lipopolyplex, while lipoplexes prepared with bioreducible poly-l-lysine were the only ones achieving any transfection in microdroplet-enabled cell culture. None of the prepared formulations achieved significant gene transfection in 3D culture. All of the prepared formulations were well tolerated by cells in 2D culture, while at least one formulation (poly-l-lysine polyplex) delayed 3D spheroid growth. These results highlight the need for selecting the appropriate in vitro model depending on the intended application.

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