Technological challenges in the preclinical development of an HIV nanovaccine candidate.

Despite a very active research in the field of nanomedicine, only a few nano-based drug delivery systems have reached the market. The "death valley" between research and commercialization has been partially attributed to the limited characterization and reproducibility of the nanoformulations. Our group has previously reported the potential of a peptide-based nanovaccine candidate for the prevention of SIV infection in macaques. This vaccine candidate is composed of chitosan/dextran sulfate nanoparticles containing twelve SIV peptide antigens. The aim of this work was to rigorously characterize one of these nanoformulations containing a specific peptide, following a quality-by-design approach. The evaluation of the different quality attributes was performed by several complementary techniques, such as dynamic light scattering, nanoparticle tracking analysis, and electron microscopy for particle size characterization. The inter-batch reproducibility was validated by three independent laboratories. Finally, the long-term stability and scalability of the manufacturing technique were assessed. Overall, these data, together with the in vivo efficacy results obtained in macaques, underline the promise this new vaccine holds with regard to its translation to clinical trials. Graphical abstract.

New generation of hybrid poly/oligosaccharide nanoparticles as carriers for the nasal delivery of macromolecules.

We have recently reported a new generation of polysaccharide nanoparticles consisting of chitosan (CS) and cyclodextrin (CD) derivatives, which exhibit a number of advantages when compared to the classical CS nanoparticles. In the present work our goal was to explore the potential of these hybrid CS/CD nanoparticles as carriers for the nasal delivery of macromolecules. First, we evaluated the effect of the amount and type of CD (sulfobutylether-beta-CD or carboximethyl-beta-CD) on the physicochemical properties of the nanocarriers. Second, we investigated the interaction of CS/CD nanoparticles with the nasal epithelium by studying their ability to modulate the tight junctions between epithelial cells (Calu-3 cell model) as well as their capacity to overcome mucosal barriers (nasal epithelium of rats). Finally, we loaded two selected nanocarriers with insulin and studied their potential for enhancing the nasal transport of insulin in rabbits. The results showed that CS/CD nanoparticles caused a reversible reduction in the transepithelial resistance of the cell monolayer, thus increasing the membrane permeability. Moreover, the results obtained following the in vivo administration of fluorescent CS/CD nanoparticles to rats evidenced their capability to overcome the nasal mucosal barrier. Finally, the in vivo evaluation in conscious rabbits revealed that insulin-loaded nanoparticles (association efficiencies > 88%) were able to significantly decrease plasma glucose levels (more than 35% reduction). Overall, these results suggest that these new nanoparticles work as nasal carriers and, therefore, have a potential for enhancing the transport of complex molecules across the nasal barrier.

Chitosan/cyclodextrin nanoparticles can efficiently transfect the airway epithelium in vitro.

The main goal of the present study was to investigate the potential of a new generation of hybrid polysaccharide nanocarriers, composed of chitosan (CS) and anionic cyclodextrins (CDs), for gene delivery to the airway epithelium. More specifically, these nanocarriers were investigated with regard to their ability to enter epithelial cells and promote gene expression in the Calu-3 cell culture model. In the search for the most suitable nanocarrier composition for gene delivery, the effect of CS molecular weight (Mw) on the nanocarriers characteristics and their ability to transfect cells was investigated. Thus, hybrid CS/CD nanoparticles were prepared with two different CS Mw, medium (110 kDa) and low (10 kDa), and loaded with pSEAP (plasmid DNA model that encodes the expression of secreted alkaline phosphatase). The resulting nanoparticles presented an adequate size range (100-200 nm, depending on CS Mw), a positive surface charge (+22 to +35 mV) and very high DNA association efficiency values (>90%). Cellular uptake studies showed that the nanoparticles were effectively internalized by the cells, providing a good indication of their potential as gene carriers. The transfection efficiency of the different formulations, measured by the concentration of secreted gene product (SEAP), indicated that all the nanoparticles were able to elicit a significantly higher response than the naked DNA (control), the transfection efficiency being more important for low MwCS nanoparticles than for those composed of medium MwCS. Overall, this report is the first evidence of the potential of a new generation of safe polysaccharide nanocarriers for gene delivery to the airway epithelium.