Optimization of Polyplex Formation between DNA Oligonucleotide and Poly(ʟ-Lysine): Experimental Study and Modeling Approach

Abstract

The polyplexes formed by nucleic acids and polycations have received a great attention owing to their potential application in gene therapy. In our study, we report experimental results and modeling outcomes regarding the optimization of polyplex formation between the double-stranded DNA (dsDNA) and poly(l-Lysine) (PLL). The quantification of the binding efficiency during polyplex formation was performed by processing of the images captured from the gel electrophoresis assays. The design of experiments (DoE) and response surface methodology (RSM) were employed to investigate the coupling effect of key factors (pH and N/P ratio) affecting the binding efficiency. According to the experimental observations and response surface analysis, the N/P ratio showed a major influence on binding efficiency compared to pH. Model-based optimization calculations along with the experimental confirmation runs unveiled the maximal binding efficiency (99.4%) achieved at pH 5.4 and N/P ratio 125. To support the experimental data and reveal insights of molecular mechanism responsible for the polyplex formation between dsDNA and PLL, molecular dynamics simulations were performed at pH 5.4 and 7.4.

The Multifaceted Histidine-Based Carriers for Nucleic Acid Delivery: Advances and Challenges

“…As previously discussed, NMR showed that histidines/imidazoles formed hydrogen bonds with nucleic acids [ 12 ], and further studies should be able to define the specific functional groups on the nucleotide which form the hydrogen bonds. Alternatively, molecular dynamic simulations have been done to understand the interactions between polymers [ 108 , 109 , 110 ], including PEI and polylysine, with oligonucleotides. Similar studies could readily be applied to histidine-rich polymers and oligonucleotides.…”

Smart Supra- and Macro-Molecular Tools for Biomedical Applications

“…For example, at full PLL protonation (which takes place at pH 5.4), the distance between macromolecules drops in a couple of nanoseconds (from roughly 40 Å to below 20 Å). At a smaller degree of protonation (50%, attainable at pH 7.4), the same distance between macromolecules is reached after roughly 30 s [110].…”

“…The intricate transport mechanism is not completely understood, and it is customarily narrowed down to a restrained collection of computational and experimental parameters [110]. The complexity of this puzzle is further enhanced by a wide variety of vector architectures and, thus, the ideal, both efficient (transport and safety) and easy-to-manufacture non-viral carrier cannot be fully described and designed.…”

“…Theoretical molecular dynamics simulations and experimental studies explain their success by the variable ability to neutralize the negative charges of DNA through pH-dependent, electrostatic binding and high density of amine functionalities susceptible to protonation in endolysosomal-specific pH conditions (pH 5.0-7.4) [105,110,115,116].…”

Aptamer Functionalized Upconversion Nanotheranostic Agent With Nuclear Targeting as the Highly Localized Drug-Delivery System of Doxorubicin

“…Furthermore, NIR as the excitation light of UCNPs is coincident with the biological transparency window; thus, greater tissue penetration depth and reduced background autofluorescence would be obtained ( Dong et al, 2017 ). Therefore, UCNPs have shown outstanding merits in in vitro fluorescence detection ( Tian et al, 2015 ), multimodal imaging in small animals, and drug carriers ( Vasiliu et al, 2017 ). In this work, we used UCNPs as the luminescence core and then coated them with a layer of cationic polymer, poly- D -lysine (PDL), as a gene carrier which has been frequently used due to its low gene transfection and the easy binding with nucleic acids ( Song et al, 2017 ).…”