Gelatin-based nanoparticles as DNA delivery systems: Synthesis, physicochemical and biocompatible characterization

Abstract

The rapidly rising demand for therapeutic grade DNA molecules requires associated improvements in encapsulation and delivery technologies. One of the challenges for the efficient intracellular delivery of therapeutic biomolecules after their cell internalization by endocytosis is to manipulate the non-productive trafficking from endosomes to lysosomes, where degradation may occur. The combination of the endosomal acidity with the endosomolytic capability of the nanocarrier can increase the intracellular delivery of many drugs, genes and proteins, which, therefore, might enhance their therapeutic efficacy. Among the suitable compounds, the gelification properties of gelatin as well as the strong dependence of gelatin ionization with pH makes this compound an interesting candidate to be used to the effective intracellular delivery of active biomacromolecules. In the present work, gelatin (either high or low gel strength) and protamine sulfate has been selected to form particles by interaction of oppositely charged compounds. Particles in the absence of DNA (binary system) and in the presence of DNA (ternary system) have been prepared. The physicochemical characterization (particle size, polydispersity index and degree of DNA entrapment) have been evaluated. Cytotoxicity experiments have shown that the isolated systems and the resulting gelatin-based nanoparticles are essentially non-toxic. The pH-dependent hemolysis assay and the response of the nanoparticles co-incubated in buffers at defined pHs that mimic extracellular, early endosomal and late endo-lysosomal environments demonstrated that the nanoparticles tend to destabilize and DNA can be successfully released. It was found that, in addition to the imposed compositions, the gel strength of gelatin is a controlling parameter of the final properties of these nanoparticles. The results indicate that these gelatin-based nanoparticles have excellent properties as highly potent and non-toxic intracellular delivery systems, rendering them promising DNA vehicles to be used as non-viral gene delivery systems.

Recent Advances in Nanomaterials for Gene Delivery—A Review

“…The researchers effectively utilized the psi(RFP)-tGel NPs to induce gene silencing in RFP/B16F10 melanoma cells, which was demonstrated by an 80% reduction in RFP mRNA expression relative to the control [69]. Moran et al [70] utilized gelatin B and protamine sulfate to deliver DNA. The use of gelatin B was an innovative choice, because it has an isoelectric point (pI) of 4.8–5.2, which makes it negatively charged at physiological pH and allows interactions with oppositely charged molecules [71].…”

“…Protamine sulfate (PS) is a highly positively charged molecule that can bind DNA, offering a mechanism to trap the DNA inside the gelatin B–PS complex for efficient gene delivery. Moran et al [70] demonstrated that DNA release is dependent upon the initial DNA concentration and the gel strength of the gelatin. The maximum amount of DNA released was reported to be between 12 and 17 μg·mL −1 depending on the exact gelatin configuration.…”

DNA Vaccines—How Far From Clinical Use?

“…Proteins constitute a favorable material for NC generation since they are biocompatible, biodegradable and typically show low cytotoxicity [ 181 ]. A good choice for a gene delivery system is gelatin B combined with protamine sulfate [ 182 ]. The overall negative charge of the protein switches in the endosome leading to a release of cargo.…”

Engineered Nanodelivery Systems to Improve DNA Vaccine Technologies

“…There has yet to be in vivo results to validate this delivery platform. However, in vitro results have demonstrated that gelatin-B-protamine-DNA complexes display low cytotoxicity and appear to be safe from a cellular standpoint [127].…”