Poly-glutamic dendrimer-based conjugates for cancer vaccination - a computational design for targeted delivery of antigens.

Computational techniques are useful to predict interaction models and molecular properties for the design of drug delivery systems, such as dendrimers. This work evaluated the impact of surface modifications of mannosamine-conjugated multifunctional poly(glutamic acid) (PG)-dendrimers as nanocarriers of the tumour associated antigens (TAA) MART-1, gp100:44 and gp100:209. Molecular dynamics simulations and docking studies were performed. Nitrobenzoxadiazole (NBD)-PG-G4-dendrimer displayed 64 carboxylic groups, however, the Frontier Molecular Orbital Theory study evidenced that only 32 of those were available to form covalent bonds. When the number of mannosamines conjugated to dendrimer was increased from 16 to 32, the dendrimer interacted with the receptor with higher affinity. However, 16 mannosamines-NBD-PG-G4-dendrimer was chosen to conjugate TAA for added functionality as no carboxylic end groups were available for further conjugation in the 32 mannosamines-dendrimer. Docking results showed that the majority of TAA-conjugated NBD-PG-G4-dendrimer demonstrated a favourable interaction with mannosamine binding site on mannose receptor, thus constituting a promising tool for TAA targeted delivery. Our in silico approach effectively narrows down the selection of the best candidates for the synthesis of functionalised PG-dendrimers with desired functionalities. These results will significantly reduce the time and efforts required to experimentally synthesise modified dendrimers for optimal antigen delivery.

EMT blockage strategies: Targeting Akt dependent mechanisms for breast cancer metastatic behaviour modulation.

Epithelial Mesenchymal Transition (EMT) is an event where epithelial cells acquire mesenchymal-like phenotype. EMT can occur as a physiological phenomenon during tissue development and wound healing, but most importantly, EMT can confer highly invasive properties to epithelial carcinoma cells. The impairment of E-cadherin expression, an essential cell-cell adhesion protein, together with an increase in the expression of mesenchymal markers, such as N-cadherin, vimentin, and fibronectin, characterize the EMT process and are usually correlated with tumor migration, and metastization. A wide range of micro-environmental and intracellular factors regulate tumor development and progression. The dynamic cross-talk between the adhesion-related proteins such as E-cadherin and the EMT-related transcription factors, with special focus on TWIST, will be discussed here, with the aim of finding a suitable biological pathway to be used as potential target for cancer therapy. Emerging concepts such as the role of the PI3K/AKT/TWIST pathway in the regulation of the E-cadherin expression will be highlighted, since it seems to be consistently involved in cells EMT. The well-known efficacy of the RNA interference as a tool to silence the expression of specific proteins has come into focus as a strategy to control different tumor sub-populations. Despite the oligonucleotides enormous sensitivity and low in vivo stability, new (nano)technological solutions are expected to enable RNAi clinical application in cancer therapy.

Incorporation of tocopherol acetate-containing particles in acrylic bone cement.

Acrylic bone cement (BC) is used in orthopaedic surgery to anchor cemented prostheses to bone. Association of antioxidant molecules to BC may suppress reactive species injury which contributes to implant failure. Tocopherol acetate (ATA)-loaded polymethylmethacrylate (PMMA) particles (ATA(PMMA)) were prepared by single emulsion solvent evaporation technique and were incorporated into BC. An encapsulation efficiency of 84% (w/w) was obtained and drug release studies showed distinct ATA release profiles and mechanisms before and after particle incorporation into BC. Experimental data, analysed using first-order, Higuchi and Korsmeyer-Peppas models revealed that ATA was released from particles by a Fickian diffusion mechanism while a non-Fickian transport was observed upon particle incorporation in BC. There were no changes in the mechanical properties of BC specimens containing ATA(PMMA) particles, in contrast to what was observed when ATA was loaded directly into BC. Overall, ATA(PMMA) particles are potential carriers for the incorporation of an antioxidant drug into BC.

Surface modified polymeric nanoparticles for immunisation against equine strangles.

The successful development of particulate vaccines depends on the understanding of their physicochemical and biological characteristics. Therefore, the main purpose of this study was to develop and characterise stable surface modified poly(lactic acid) (PLA) nanoparticles, using polyvinyl alcohol (PVA), alginate (ALG) and glycolchitosan (GCS) containing a Streptococcus equi enzymatic extract adsorbed onto the surface. The characterisation of the preparations and a physicochemical study of the adsorption process were performed. The adsorption of S. equi proteins is a rapid process reaching, within 1h, maximum adsorption efficiency values of 75.2+/-1.9% (w/w) for PLA-PVA, 84.9+/-0.2% (w/w) for PLA-GCS and 78.1+/-0.4% (w/w) for PLA-ALG nanoparticles. No protein degradation was detected throughout the formulation procedures. As expected from a complex mixture of proteins, adsorption data suggest a Freundlich-type of equilibrium with regression coefficients (r(2)) of 0.9958, 0.9839 and 0.9940 for PLA-PVA, PLA-GCS and PLA-ALG, respectively. Desorption studies revealed a burst release within the first 6h, for all formulations, followed by a sustained release profile. Nanoparticle surface modification with GCS improved the sustained release profile, as 20% of protein remained attached to the particle surface after 30 days. The results show that adsorption is an alternative method for the production of S. equi antigen carriers for vaccination purposes.

New approach on the development of a mucosal vaccine against strangles: Systemic and mucosal immune responses in a mouse model.

Streptococcus equi subspecies equi affects animals of Equidae family and is the causative agent of strangles, an acute, extremely contagious and deadly disease. Prolonged periods of protection associated to absence of serious adverse reactions were not yet achieved. Thus, this experimental work is focused on the study of mucosal, humoral and cellular immune responses developed in a mouse model, after the intranasal administration of S. equi antigens associated by adsorption or encapsulation to poly(lactic acid) nanospheres, modified by mucoadhesive polymers and absorption enhancers. Particles fitted the nanometer range and proteins integrity and antigenicity were not affected. PLA nanospheres induced a mixed Th1 and Th2 response, being therefore potential carriers for the delivery of S. equi antigens.

The enhancement of the immune response against S. equi antigens through the intranasal administration of poly-epsilon-caprolactone-based nanoparticles.

Strangles is a bacterial infection of the Equidae family that affects the nasopharynx and draining lymph nodes, caused by Streptococcus equi subspecies equi. This agent is responsible for 30% of all worldwide equine infections and is quite sensitive to penicillin and other antibiotics. However, prevention is still the best option because the current antibiotic therapy and vaccination is often ineffective. As S. equi induces very strong systemic and mucosal responses in convalescent horses, an effective and economic strangles vaccine is still a priority. In this study the humoral, cellular and mucosal immune responses to S. equi antigens encapsulated or adsorbed onto poly-epsilon-caprolactone nanospheres were evaluated in mice. Particles were produced by a double (w/o/w) emulsion solvent evaporation technique and contained mucoadhesive polymers (alginate or chitosan) and absorption enhancers (spermine, oleic acid). Their intranasal administration, particularly those constituted by the mucoadhesive polymers, increased the immunogenicity and mucosal immune responses (SIgA) to the antigen. The inclusion of cholera toxin B subunit in the formulations successfully further activated the paths leading to Th1 and Th2 cells. Therefore, those PCL nanospheres are potential carriers for the delivery of S.equi antigens to protect animals against strangles.

Streptococcus equi antigens adsorbed onto surface modified poly-epsilon-caprolactone microspheres induce humoral and cellular specific immune responses.

Streptococcus equi subsp. equi is the causative agent of Strangles, which is one of the most costly and widespread infectious diseases, affecting the respiratory tract of Equidae. In this work, polyvinyl alcohol, alginate and chitosan were used in formulations of surface modified poly-epsilon-caprolactone microspheres which were evaluated after adsorption of S.equi enzymatic extract for physicochemical characteristics and in vivo immune responses in mice. After subcutaneous immunisation, the formulations induced higher lymphokines levels, in accordance with cellular and humoral immune responses, as compared to the free antigen, successfully activating the paths leading to Th1 and Th2 cells. The obtained results highlight the role of these microspheres as an adjuvant and their use to protect animals against strangles.