Metastatic Melanoma: A Preclinical Model Standardization and Development of a Chitosan-Coated Nanoemulsion Containing Temozolomide to Treat Brain Metastasis.

Melanoma is the most aggressive type of skin cancer. Brain metastasis is the worst scenario in metastatic melanoma and the treatment options for these patients are limited. Temozolomide (TMZ) is a chemotherapy agent used to treat primary central nervous system tumors. Our objective was to develop chitosan-coated nanoemulsion containing temozolomide (CNE-TMZ) for nasal route administration to melanoma brain metastasis treatment. A preclinical model of metastatic brain melanoma was standardized, and the efficiency of the developed formulation was further determined in vitro and in vivo. The nanoemulsion was done by spontaneous emulsification method and the formulation was characterized by size, pH, polydispersity index, and zeta potential. Culture assessments to determine cell viability were done in the A375 human melanoma cell line. To determine the safety of formulation, healthy C57/BL6 mice were treated with a nanoemulsion without TMZ. The model in vivo used B16-F10 cells implanted by stereotaxic surgery in C57/BL6 mice brains. The results demonstrate that the preclinical model used showed to be useful to analyze the efficiency of new candidate drugs to treat melanoma brain metastasis. The chitosan-coated nanoemulsions with TMZ showed the expected physicochemical characteristics and demonstrated safety and efficacy, reducing around 70% the tumor size compared to control mice, and presenting a tendency in mitotic index reduction, becoming an interesting approach to treat melanoma brain metastasis.

Nasal administration of a temozolomide-loaded thermoresponsive nanoemulsion reduces tumor growth in a preclinical glioblastoma model.

Glioblastoma (GB) is the worst and most common primary brain tumor. Temozolomide (TMZ), an alkylating agent, is widely used for treating primary and recurrent high-grade gliomas. However, at least 50% of TMZ treated patients do not respond to TMZ and the development of chemoresistance is a major problem. Here, we designed a lipid nanoemulsion containing a thermoresponsive polymer (poloxamer 407) aiming to improve TMZ release into the brain via nasal delivery. Increasing amounts of poloxamer 407 were added to preformed nanoemulsions (250 nm-range) obtained by spontaneous emulsification. The influence of the polymer concentration (from 2.5% to 12.5%) and temperature on viscosity was clearly evidenced. Such effect was also noticed on the mucoadhesiveness of formulations, as well as TMZ release rate and retention/permeation through nasal porcine mucosa using Franz-type diffusion cells. From these results, a formulation containing 10% of poloxamer (NTMZ-P10) was selected for further experiments by nasal route. A significantly higher TMZ amount was observed in the brain of rats from NTMZ-P10 in comparison with controls. Finally, our results show that formulation reduced significantly tumor growth by three-fold: 103.88 ± 43.67 mm3 (for NTMZ-P10) and 303.28 ± 95.27 mm3 (control). Overall, these results suggest the potential of the thermoresponsive formulation, administered by the non-invasive nasal route, as a future effective glioblastoma treatment.

Chitosan-coated rosmarinic acid nanoemulsion nasal administration protects against LPS-induced memory deficit, neuroinflammation, and oxidative stress in Wistar rats.

Rosmarinic acid (RA) lipid-nanotechnology-based delivery systems associate with mucoadhesive biopolymers for nasal administration has arisen as a new promising neuroprotective therapy for neurodegenerative disorders (ND). We have previously demonstrated the glioprotective effect of chitosan-coated RA nanoemulsions (RA CNE) against lipopolysaccharide (LPS)-induced damage in rat astrocyte primary culture. Here, we further investigate the protective effect of RA CNE nasal administration on LPS-induced memory deficit, neuroinflammation, and oxidative stress in Wistar rats, since these in vivo studies were crucial to understand the impact of developed delivery systems in the RA neuroprotective effects. The animals were treated through nasal route with RA CNE (2 mg·mL-1), free RA (2 mg·mL-1), blank CNE, and saline (control and LPS groups) administrations (n.a., 100 µL per nostril) twice a day (7 a.m./7 p.m.) for six days. On the sixth day, the animals received the last treatments and LPS was intraperitoneally (i.p.) administrated (250 µg·kg-1). Overall results, proved for the first time that the RA CNE nasal administration elicits a neuroprotective effect against LPS-induced damage, which was associated with increased 1.6 times recognition index, decreased 5.0 and 1.9 times in GFAP+ cell count and CD11b expression, respectively, as well as increased 1.7 times SH in cerebellum and decreased 3.9 times TBARS levels in cerebral cortex in comparison with LPS group. RA CNE treatment also facilitates RA bioavailability in the brain, confirmed by RA quantification. Free RA also demonstrates a protective effect in some studied parameters, although no RA was quantified in the brain.

Solid Dispersion of Kaempferol: Formulation Development, Characterization, and Oral Bioavailability Assessment.

Kaempferol (KPF), an important flavonoid, has been reported to exert antioxidant, anti-inflammatory, and anticancer activity. However, this compound has low water solubility and hence poor oral bioavailability. This work aims to prepare a solid dispersion (SD) of KPF using Poloxamer 407 in order to improve the water solubility, dissolution rate, and pharmacokinetic properties KPF. After optimization, SDs were prepared at a 1:5 weight ratio of KPF:carrier using the solvent method (SDSM) and melting method (SDMM). Formulations were characterized by Fourier transform infrared spectroscopy (FTIR), X-ray diffractometry (XRD) analysis, differential scanning calorimetry (DSC), and scanning electron microscopy (SEM). The solubility in water of carried-KPF was about 4000-fold greater than that of free KPF. Compared with free KPF or the physical mixture, solid dispersions significantly increased the extent of drug release (approximately 100% within 120 min) and the dissolution rate. Furthermore, after oral administration of SDMM in rats, the area under the curve (AUC) and the peak plasma concentration (Cmax) of KPF from SDMM were twofold greater than those of free KPF (p < 0.05). In conclusion, SD with Poloxamer 407 is a feasible pharmacotechnical strategy to ameliorate the dissolution and bioavailability of KPF.

Effects of Surface Characteristics of Polymeric Nanocapsules on the Pharmacokinetics and Efficacy of Antimalarial Quinine.

INTRODUCTION: The surface charge of nanoparticles, such as nanospheres (NS) and nanocapsules (NC), has been studied with the purpose of improving the in vivo performance of drugs. The aim of this study was to develop, characterize, and evaluate the in vitro antimalarial efficacy of NCP80 and NSP80 (polysorbate coated) or NCEUD and NSEUD (prepared with Eudragit RS 100) loading quinine (QN). METHODS: Formulations were prepared by the nanoprecipitation method, followed by wide physicochemical characterization. Antimalarial activity in Plasmodium berghei-infected mice and populational pharmacokinetics (PopPK) in rats were evaluated. RESULTS: The formulations showed a nanometric range (between 138 ± 3.8 to 201 ± 23.0 nm), zeta potential (mV) of -33.1 ± 0.7 (NCP80), -30.5 ± 1 (UNCP80), -25.5 ± 1 (NSP80), -20 ± 0.3 (UNSP80), 4.61 ± 1 (NCEUD), 14.1 ± 0.9 (UNCEUD), 2.86 ± 0.3 (NSEUD) and 2.84 ± 0.6 (UNSEUD), content close to 100%, and good QN protection against UVA light. There was a twofold increase in the penetration of QN into infected erythrocytes with NC compared to that with NS. There was a significant increase in t1/2 for all NC evaluated compared to that of Free-QN, due to changes in Vdss. PopPK analysis showed that NCP80 acted as a covariate to Q (intercompartmental clearance) and V2 (volume of distribution in the peripheral compartment). For NCEUD, V1 and Q were modified after QN nanoencapsulation. Regarding in vivo efficacy, NCEUD increased the survival of mice unlike Free-QN. CONCLUSION: Cationic nanocapsules modified the pharmacology of QN, presenting a potential alternative for malaria treatment.

Optimization of Curcuma Oil/Quinine-Loaded Nanocapsules for Malaria Treatment.

Quinine, a treatment used in chloroquine-resistant falciparum malaria, was loaded into poly(ɛ-caprolactone) or Eudragit® RS100 nanocapsules using Curcuma oil as the oil-based core. Until now, the effect of cationic nanocapsules on malaria has not been reported. A 24 factorial design was adopted using, as independent variables, the concentration of Curcuma oil, presence of quinine, type of polymer, and aqueous surfactant. Diameter, zeta potential, and pH were the responses studied. The formulations were also evaluated for drug content, encapsulation efficiency, photostability, and antimalarial activity against Plasmodium berghei-infected mice. The type of polymer influenced all of the responses studied. Quinine-loaded Eudragit® RS100 (F13) and PCL nanocapsules (F9), both with polysorbate 80 coating, showed nanometric particle size, positive zeta potential, neutral pH, high drug content, and quinine photoprotection ability; thus, these nanocapsules were selected for in vivo tests. Both formulations showed lower levels of parasitemia from the beginning of the experiment (5.78 ± 3.60 and 4.76 ± 3.46% for F9 and F13, respectively) and highest survival mean time (15.3 ± 2.0 and 14.9 ± 5.6 days for F9 and F13, respectively). F9 and F13 showed significant survival curve compared to saline, thus demonstrating that nanoencapsulation improved bioefficacy of QN and co-encapsulated curcuminoids, regardless of the surface charge.

The use of Brazilian vegetable oils in nanoemulsions: an update on preparation and biological applications

ABSTRACT Vegetable oils present important pharmacological properties, which gained ground in the pharmaceutical field. Its encapsulation in nanoemulsions is considered a promising strategy to facilitate the applicability of these natural compounds and to potentiate the actions. These formulations offer several advantages for topical and systemic delivery of cosmetic and pharmaceutical agents including controlled droplet size, protection of the vegetable oil to photo, thermal and volatilization instability and ability to dissolve and stabilize lipophilic drugs. For these reasons, the aim of this review is to report on some characteristics, preparation methods, applications and especially analyze recent research available in the literature concerning the use of vegetable oils with therapeutic characteristics as lipid core in nanoemulsions, specially from Brazilian flora, such as babassu (Orbignya oleifera), aroeira (Schinus molle L.), andiroba (Carapa guaianiensis), casca-de-anta (Drimys brasiliensis Miers), sucupira (Pterodon emarginatus Vogel) and carqueja doce (Stenachaenium megapotamicum) oils.

Clozapine-Loaded Polysorbate-Coated Polymeric Nanocapsules: Physico-Chemical Characterization and Toxicity Evaluation in Caenorhabditis elegans Model.

The aim of this work was to develop and characterize clozapine loaded polysorbate-coated polymeric nanocapsules and assess their toxicity in Caenorhabditis elegans, an invertebrate animal model. Formulations were prepared by nanoprecipitation method and characterized by particle size, zeta potential, pH, drug loading, entrapment efficiency and in vitro drug release. All nanocapsules prepared presented diameter around 140 nm, pH slightly acid and negative zeta potential. In vitro studies showed biphasic drug release from nanocapsules with decreasing of the release rate on nanoencapsulation. The t(1/2)beta of clozapine was 7.23 +/- 0.73 and 2.23 +/- 0.97 h for nanoencapsulated and free drug, respectively (p < 0.05), in pH 1.2 medium. Similar results were obtained in pH 6.8 buffer. Regarding toxicity evaluation, worms exposed to clozapine-loaded nanocapsules did not show the same mortality rate compared to others formulations, as the survival was significantly higher than the free drug treated-group. In addition, we observed that free clozapine decreased egg laying at the first reproductive day, whereas nanoencapsulated clozapine did not depict significant change of this parameter. Longevity assay showed no significant difference, demonstrating that the toxicological effects of clozapine observed in C. elegans are acute. In addition, we proved that free and nanoencapsulated clozapine were orally uptake by the worms, as determined by fluorescein-labeled nanocapsules. Then, the use of nanocapsules delayed the drug release and minimized the toxic effects of clozapine in worms, which can be used as a new animal model to evaluate the nanotoxicity of drug delivery systems.

Educational Activities for Rural and Urban Students to Prevent Skin Cancer in Rio Grande do Sul, Brazil.

BACKGROUND: Excessive exposure to the sun during childhood is strongly associated with the development of skin cancer in the future. The only way to prevent the development of skin cancer is to protect against ultraviolet radiation, which can be achieved through strategic awareness during childhood and adolescence. OBJECTIVE: The aim of this work was to evaluate the impact of educational activities for rural and urban students to promote the use of sunscreens and prevent skin cancer. MATERIALS AND METHODS: This study was carried out with students (9-12 years) of rural (n=70) and urban (n=70) schools in Rio Grande do Sul state, Brazil. The educational interventions were lectures and games. The impact of this strategy was evaluated through the application of a questionnaire before and after the interventions. RESULTS: Before the intervention, it was found around 50% of rural and urban students were not aware of the damage caused by sun exposure, often exposing themselves to UV radiation without use sunscreen ( ~ 25 %) and at the most critical times of the day/year. After the lectures we observed an improvement in the behavior of the students with regard to sun exposure and knowledge about skin cancer. CONCLUSIONS: The results of this study emphasize the importance of prevention strategies for skin cancer and promoting the use of sunscreens based educational strategies. The interventions were of great value in relation to disseminating knowledge on the subject.

Effect of quinine-loaded polysorbate-coated nanocapsules on male and female reproductive systems of rats.

Quinine is an antimalarial drug; however, its use is limited by its narrow therapeutic index and elevated side effects. The nanosystems are promising delivery vehicles of antimalarial drugs, enhancing their therapeutic potential. This study aimed to compare the toxicity of quinine and quinine loaded nanocapsules (Q-NC) on the reproductive system of male and female rats. The animals received quinine or Q-NC orally at the same dose of 25 mg kg-1 for 7 days (real period of quinine therapy in humans). 24 hours after the last administration, the rats were euthanized and the ovarian and testicular tissues were removed for histological and biochemical analyses. The groups treated with quinine presented ovarian and testicular damage, evidenced by the increase of reactive species and malondialdehyde levels, the decrease of 17ß-hydroxysteroid dehydrogenase activity and alterations on total antioxidant capacity. The females presented a decrease of follicular viability and the males presented a decrease of spermatozoa membrane integrity, as well as moderated histological alterations on testis after the exposure to quinine. After the treatment with Q-NC, the males presented decreased reactive species levels and total antioxidant capacity at control levels, as well as spermatozoa with 100% of membrane integrity. The females treated with Q-NC presented reactive species levels, total antioxidant capacity, 17ß-hydroxysteroid dehydrogenase activity and follicular viability at control levels, and decreased malondialdehyde levels when compared to quinine, but not at control levels. This study demonstrated that loading polymeric nanocapsules with quinine decreased the deleterious effects induced by quinine on ovaries and partially on testicles.