No abstract

In current times, DNA vaccines are seen as a promising approach to treat and prevent diseases, such as virus infections and cancer. Aiming at the production of a functional and effective plasmid DNA (pDNA) delivery system, four chitosan polymers, differing in the molecular weight, were studied using the design of experiments (DoE) tool. These gene delivery systems were formulated by ionotropic gelation and exploring the chitosan and TPP concentrations as DoE inputs to maximize the nanoparticle positive charge and minimize their size and polydispersity index (PDI) as DoE outputs. The obtained linear and quadratic models were statistically significant (p-value < 0.05) and non-significant lack of fit, with suitable coefficient of determination and the respective optimal points successfully validated. Furthermore, morphology, stability and cytotoxicity assays were performed to evaluate the endurance of these systems over time and their further potential for future in vitro studies. The subsequent optimization process was successful achieved for the delivery systems based on the four chitosan polymers, in which the smallest particle size was obtained for the carrier containing the 5 kDa chitosan (~82 nm), while the nanosystem prepared with the high molecular weight (HMW) chitosan displayed the highest zeta potential (~+26.8 mV). Delivery systems were stable in the formulation buffer after a month and did not exhibit toxicity for the cells. In this sense, DoE revealed to be a powerful tool to explore and tailor the characteristics of chitosan/pDNA nanosystems significantly contributing to unraveling an optimum carrier for advancing the DNA vaccines delivery field.

Nucleic acid vaccines have become a revolutionary technology to give a fast, safe, cost-effective and efficient response against viral infections, such as SARS-CoV-2 or Human papillomavirus (HPV). However, to ensure their effectiveness, the development of adequate methods to protect, carry, and deliver nucleic acids is fundamental. In this work, nanoparticles (NPs) of chitosan (CS)-tripolyphosphate (TPP)-plasmid DNA (pDNA) were thoroughly modulated and characterized, by measuring the charge and size through dynamic light scattering (DLS) and morphology by scanning electron microscopy (SEM). Stability, cytotoxicity and cellular uptake of NPs were also evaluated. Finally, the effect of polyplexes on the expression of HPV E7 antigen in human fibroblast and RAW cells was investigated through polymerase chain reaction (PCR) and real-time PCR. The results showed NPs with a spherical/oval shape, narrow size distribution <180 nm and positive zeta potentials (>20 mV) and good stability after one month of storage at 4 °C in formulation buffer or when incubated in culture medium and trypsin. In vitro studies of NPs cytotoxicity revealed that the elimination of formulation buffers led to an improvement in the rate of cell viability. The E7 antigen transcription was also increased for NPs obtained with high pDNA concentration (60 μg/mL). The analyzed CS-TPP-pDNA polyplexes can offer a promising vehicle for nucleic acid vaccines, not only in the prevention or treatment of viral infections, but also to fight emergent and future pathogens.

Intramuscular delivery is one of the main routes for DNA vaccines administration. However, it requires large amounts of the DNA to be administered and external stimulation to encourage the internalization of the DNA. In this work, we consider alternative routes for less invasive administration and develop drug delivery systems (DDS) for intranasal administration. Chitosan polyplexes using sodium tripolyphosphate (TPP) as a crosslinker were prepared using the ionic gelation method. Our method allowed preparation of nanoparticles with the size below 50 nm, which is at least two times lower than previously reported sizes. Moreover, despite the small sizes, we obtained DNA encapsulation efficiencies of about 70%. Parameters that may affect the encapsulation efficiency were investigated, including different TPP-chitosan ratios and concentrations of DNA. We found that encapsulation efficiency of DNA inside the particles decreases with the increasing TPP-chitosan ratio. Moreover, increasing the DNA concentration leads to a higher encapsulation efficiency. Small (<50 nm) chitosan nanoparticles hold enormous potential as DNA carriers due to their physiological barriers and subsequent internalization.

Paraffin has been identified as a promising alternative for use in propellant grains for hybrid rocket motors. Studies have been conducted using grain as fuel paraffin or liquid hidroxylated polybutadiene polymer (HTPB) due to the high thrust generated during firing. However, in consequences the low mechanical properties of pure paraffin, this study aims to evaluate the kinetics of degradation of a mixture composed of paraffin particles (PP) and HTPB compared with grain fuel pure paraffin and pure HTPB. For this we used a thermal analysis technique thermogravimetric in conjunction with the application of kinetic methods based on ASTM E1641 and Vyazovkin theory. Based on these results we can say the activation energy results are in agreement with the literature and the mixture PP / HTPB presents intermediate activation energy values as expected.