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Multi-level kinetic model of mRNA delivery via transfection of lipoplexes.

ABSTRACT: Recent work on the use of mRNA lipoplexes for gene delivery demonstrates the need for a mathematical model that simulates and predicts kinetics and transfection efficiency. The small copy numbers involved make it necessary to use stochastic models and include statistical analysis of the variation observed in the experimental data. The modeling requirements are further complicated by the multi-level nature of the problem, where mRNA molecules are contained in lipoplexes, which are in turn contained in endosomes, where each of these entities displays a behavior of its own. We have created a mathematical model that reproduces both the time courses and the statistical variance observed in recent experiments using single-cell tracking of GFP expression after transfection. By applying a few key simplifications and assumptions, we have limited the number of free parameters to five, which we optimize to match five experimental determinants by means of a simulated annealing algorithm. The models demonstrate the need for modeling of nested species in order to reproduce the shape of the dose-response and expression-level curves.

SUBMITTER: Ligon TS

PROVIDER: S-EPMC4169511 | biostudies-literature | 2014

REPOSITORIES: biostudies-literature

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Multi-level kinetic model of mRNA delivery via transfection of lipoplexes.

Ligon Thomas S TS Leonhardt Carolin C Rädler Joachim O JO

PloS one 20140919 9

Recent work on the use of mRNA lipoplexes for gene delivery demonstrates the need for a mathematical model that simulates and predicts kinetics and transfection efficiency. The small copy numbers involved make it necessary to use stochastic models and include statistical analysis of the variation observed in the experimental data. The modeling requirements are further complicated by the multi-level nature of the problem, where mRNA molecules are contained in lipoplexes, which are in turn contain ...[more]

PMID: 25237886

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Project description:Some quaternary gemini amphiphiles (GAs) were synthesized as nonviral gene delivery carriers. The critical miceller concentration values of these amphiphiles are indicative of their superior surface-active properties. All of the synthesized GAs, alone or along with lipids like cholesterol and/or dioleoylphosphatidyl ethanolamine (DOPE), were formulated as liposomes. Formulations of GAs with DOPE showed average particle diameters of 326-400 nm with positive ζ-potential (30.1-46.4 mV). The lipoplexes of theses formulations showed complete pDNA retention at the base at a N/P ratio higher than 1.0 in gel retardation study. The GAs were effective in condensing pDNA into a ψ-phase, as indicated by circular dichroism study, and provided complete protection of the pDNA against the enzyme DNase at a N/P ratio more than 1. In vitro cell line studies showed that GA liposomal formulations caused β-gal expression and offered a higher transfection efficiency than that of liposomes prepared with the help of N-[1-(2,3-dioleoyloxy)propyl]-N,N,N-trimethylammonium methyl-sulfate (DOTAP)/DOPE and dicyclocarbodiimide (DCC)/DOPE but comparable to those of Lipofectamine 2000 in A549 and HeLa cell lines. Modulation of head group polarity significantly affected the transfection efficacy of GAs. The cell viabilities of almost all of the formulations were comparable to those of the standards (DCC/DOPE and DOTAP/DOPE liposomes). Incorporation of cholesterol [GA/DOPE/cholesterol in the ratio of 1:1:1] further improved the serum compatibility of the formulations and improved the transfection efficacy when evaluated in A549 and HeLa cell lines. Fluorescence-assisted cell sorting studies showed comparable number of transfected cells to Lipofectamine 2000 in the HeLa cell line. Intracellular trafficking studies using confocal microscopy indicated transfection of the HeLa cells with the reporter gene within 30 min of lipoplex treatment. γ-Scintigraphy using 99mTc-labeled lipoplexes showed higher concentrations of the lipoplexes in vital tissues like liver, spleen, lungs, and kidneys.

Dataset Information

Multi-level kinetic model of mRNA delivery via transfection of lipoplexes.

Publications

Multi-level kinetic model of mRNA delivery via transfection of lipoplexes.

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