In vitro interaction of polyethylene glycol-block-poly(D,L-lactide) nanocapsule devices with host cardiomyoblasts and Trypanosoma cruzi-infective forms.

Chagas disease, caused by the protozoan Trypanosoma cruzi, is an important public health problem in Latin America. Nanoencapsulation of anti-T. cruzi drugs has significantly improved their efficacy and reduced cardiotoxicity. Thus, we investigated the in vitro interaction of polyethylene glycol-block-poly(D,L-lactide) nanocapsules (PEG-PLA) with trypomastigotes and with intracellular amastigotes of the Y strain in cardiomyoblasts, which are the infective forms of T. cruzi, using fluorescence and confocal microscopy. Fluorescently labeled nanocapsules (NCs) were internalized by non-infected H9c2 cells toward the perinuclear region. The NCs did not induce significant cytotoxicity in the H9c2 cells, even at the highest concentrations and interacted equally with infected and non-infected cells. In infected cardiomyocytes, NCs were distributed in the cytoplasm and located near intracellular amastigote forms. PEG-PLA NCs and trypomastigote form interactions also occurred. Altogether, this study contributes to the development of engineered polymeric nanocarriers as a platform to encapsulate drugs and to improve their uptake by different intra- and extracellular forms of T. cruzi, paving the way to find new therapeutic strategies to fight the causative agent of Chagas disease.

Repositioning of Tamoxifen in Surface-Modified Nanocapsules as a Promising Oral Treatment for Visceral Leishmaniasis.

Standards of care for human visceral leishmaniasis (VL) are based on drugs used parenterally, and oral treatment options are urgently needed. In the present study, a repurposing strategy was used associating tamoxifen (TMX) with polyethylene glycol-block-polylactide nanocapsules (NC) and its anti-leishmanial efficacy was reported in vivo. Stable surface modified-NC (5 mg/mL of TMX) exhibited 200 nm in size, +42 mV of zeta potential, and 98% encapsulation efficiency. Atomic force microscopy evidenced core-shell-NC. Treatment with TMX-NC reduced parasite-DNA quantified in liver and spleen compared to free-TMX; and provided a similar reduction of parasite burden compared with meglumine antimoniate in mice and hamster models. Image-guided biodistribution showed accumulation of NC in liver and spleen after 30 min post-administration. TMX-NC reduced the number of liver granulomas and restored the aspect of capsules and trabeculae in the spleen of infected animals. TMX-NC was tested for the first time against VL models, indicating a promising formulation for oral treatment.

Nanostructured lipid carriers as a novel tool to deliver sclareol: physicochemical characterisation and evaluation in human cancer cell lines

Sclareol (SC) is arousing great interest due to its cytostatic and cytotoxic activities in several cancer cell lines. However, its hydrophobicity is a limiting factor for its in vivo administration. One way to solve this problem is through nanoencapsulation. Therefore, solid lipid nanoparticles (SLN-SC) and nanostructured lipid carriers (NLC-SC) loaded with SC were produced and compared regarding their physicochemical properties. NLC-SC showed better SC encapsulation than SLN-SC and was chosen to be compared with free SC in human cancer cell lines (MDA-MB-231 and HCT-116). Free SC had slightly higher cytotoxicity than NLC-SC and produced subdiploid DNA content in both cell lines. On the other hand, NLC-SC led to subdiploid content in MDA-MB-231 cells and G2/M checkpoint arrest in HCT-116 cells. These findings suggest that SC encapsulation in NLC is a way to allow the in vivo administration of SC and might alter its biological properties

Development of Long-Circulating and Fusogenic Liposomes Co-encapsulating Paclitaxel and Doxorubicin in Synergistic Ratio for the Treatment of Breast Cancer.

BACKGROUND: The co-encapsulation of paclitaxel (PTX) and doxorubicin (DXR) in liposomes has the potential to offer pharmacokinetic and pharmacodynamic advantages, providing delivery of both drugs to the tumor at the ratio required for synergism. OBJECTIVE: To prepare and characterize long-circulating and fusogenic liposomes co-encapsulating PTX and DXR in the 1:10 molar ratio (LCFL-PTX/DXR). METHODS: LCFL-PTX/DXR was prepared by the lipid film formation method. The release of PTX and DXR from liposomes was performed using a dialysis method. Studies of cytotoxicity, synergism, and cellular uptake were also carried out. RESULTS: The encapsulation percentage of PTX and DXR was 74.1 ± 1.8 % and 89.6 ± 12.3%, respectively, and the mean diameter of the liposomes was 244.4 ± 28.1 nm. The vesicles remained stable for 30 days after their preparation. The drugs were simultaneously released from vesicles during 36 hours, maintaining the drugs combination in the previously established ratio. Cytotoxicity studies using 4T1 breast cancer cells showed lower inhibitory concentration 50% (IC50) value for LCFL-PTX/DXR treatment (0.27 ± 0.11 µm) compared to the values of free drugs treatment. In addition, the combination index (CI) assessed for treatment with LCFL-PTX/DXR was equal to 0.11 ± 0.04, showing strong synergism between the drugs. Cell uptake studies have confirmed that the molar ratio between PTX and DXR is maintained when the drugs are administered in liposomes. CONCLUSION: It was possible to obtain LCFL-PTX/DXR suitable for intravenous administration, capable of releasing the drugs in a fixed synergistic molar ratio in the tumor region.

Cloxacillin benzathine-loaded polymeric nanocapsules: Physicochemical characterization, cell uptake, and intramammary antimicrobial effect.

The present work shows the development and evaluation of the veterinary antibiotic cloxacillin benzathine (CLOXB) loaded into poly-ε-caprolactone (PCL) nanocapsules (NC), as a potential new treatment strategy to manage bovine intramammary infections, such as mastitis. Staphylococcus aureus-induced mastitis is often a recurrent disease due to the persistence of bacteria within infected cells. CLOXB-PCL NC were prepared by interfacial deposition of preformed biodegradable polymer followed by solvent displacement method. The mean diameter of NC varied from 241 to 428 nm and from 326 to 375 nm, when determined by dynamic light scattering and by atomic force microscopy, respectively. The zeta potential of NC was negative and varied from -28 to -51 mV. In vitro release studies from the NC were performed in two media under sink conditions: PBS with 1% polyethylene glycol or milk. A reversed-phase HPLC method was developed to determine the NC entrapment efficiency and kinetics of CLOXB release from the NC. Free CLOXB dissolution occurred very fast in both media, while drug release from the NC was slower and incomplete (below 50%) after 9 h. CLOXB release kinetics from polymeric NC was fitted with the Korsmeyer-Peppas model indicating that CLOXB release is governed by diffusion following Fick's law. The fluorescence confocal microscopy images of macrophage-like J774A.1 cells reveal NC uptake and internalization in vitro. In addition, antimicrobial effect of the intramammary administration of CLOXB-PCL NC in cows with mastitis resulted in no clinical signs of toxicity and allowed complete pathogen elimination after treatment. The in vivo results obtained in this work suggest that CLOXB-PCL NC could be a promising formulation for the treatment of intramammary infections in cattle, considering their physicochemical properties, release profiles and effects on bovine mastitis control.

Phthalocyanine photosensitizer in polyethylene glycol-block-poly(lactide-co-benzyl glycidyl ether) nanocarriers: Probing the contribution of aromatic donor-acceptor interactions in polymeric nanospheres.

For best photosensitizer activity phthalocyanine dyes used in photodynamic therapy should be molecularly dispersed. Polyethylene glycol-block-polylactide derivatives presenting benzyl side-groups were synthesized to encapsulate a highly lipophilic phthalocyanine dye (AlClPc) and evaluate the effect of π-π interactions on the nanocarrier colloidal stability and dye dispersion. Copolymers with 0, 1, 2 and 6 mol% of benzyl glycidyl ether (BGE) were obtained via polyethylene glycol initiated ring-opening copolymerization of D,l-lactide with BGE. The block copolymers formed stable, monodisperse nanospheres with low in vitro cytotoxicity. AlClPc loading increased the nanosphere size and affected their colloidal stability. The photo-physical properties of the encapsulated dye, studied in batch and after separation by field flow fractionation, demonstrated the superiority of plain PEG-PLA over BGE-containing copolymers in maintaining the dye in its monomeric (non-aggregated) form in aqueous suspension. High dye encapsulation and sustained dye release suggest that these nanocarriers are good candidates for photodynamic therapy.

Mechanisms of interaction of biodegradable polyester nanocapsules with non-phagocytic cells.

The interaction of polymer nanocapsules (NC) prepared from four biodegradable polyesters with variable polymer hydrophobicity (PCL, PLA, PLGA and PLA-PEG) was investigated in the non-phagocytic Vero, Caco-2 and HepG2 cell lines. The NC, labeled with the highly lipophilic fluorescent indocarbocyanine dye DIL, had very similar sizes (approx. 140 nm) and negative zeta-potentials. Asymmetric flow field-flow fractionation evidenced NC colloidal stability and negligible transfer of the dye to serum proteins in the incubation medium. The cytotoxicity of the NC was evaluated via MTT assay over a large polymer concentration range (1-1000 µg/mL) and time of exposure (2, 24 and 48 h). The NC were safe in vitro up to a concentration of approx. 100 µg/mL or higher, depending on the cell line and nature of the polymer. Vero cells were more sensitive to the NC, in particular NC of the more hydrophobic polymer. The cells were exposed to endocytosis inhibitors, incubated with NC, and the cell-associated fluorescence was quantified by spectrofluorometry. HepG2 cells presented a 1.5-2-fold higher endocytic capacity than Caco-2 and Vero cells. The main mechanism of NC uptake was caveolin-mediated endocytosis in HepG2 and Vero cells, and macropinocytosis in Caco-2 cells. Polymer hydrophobicity had an effect on the level of NC associated to HepG2 cells and to a lesser extent on the endocytosis mechanisms in Vero and Caco-2 cells. The NC uptake levels and endocytosis mechanisms differed significantly between cell lines tested.

α- Tocopherol succinate loaded nano-structed lipid carriers improves antitumor activity of doxorubicin in breast cancer models in vivo.

Combination-based chemotherapies have been the standard treatment for multiple solid tumors since the 1960s. Combined therapies where both agents have toxicity results in dose-limiting effects. α- tocopherol succinate (TS) is an analogue of vitamin E that exhibits antitumor properties in the absence of toxicity. Hence, its combination with a frontline chemotherapy, doxorubicin (DOX) is an alternative to increase antitumor efficacy. Therefore, the aim of this work was to evaluate the antitumor activity of nanostructed lipid carriers (NLC) loaded with TS and DOX. The NLC-TS-DOX were prepared, characterized and radiolabeled with technetium-99m. Cytotoxicity studies were performed in vitro, using two breast cancer cell lines, MDA-MB-231 and 4T1. Biodistribution and antitumor activity were evaluated in 4T1 tumor-bearing mice. The results showed that NLC-TS-DOX had a small diameter (85 nm) and a long blood clearance (T1/2ß = 1107.71 min) that consequently resulted in a higher tumor uptake compared to contralateral muscle for up to 48 h. Drug combination studies in MDA-MB-231 and 4T1 cells showed a combination index below 0.8 at ED50-90 for both cell lines. Interestingly, a high synergism was found at ED90. Antitumor activity showed a better control of tumor growth for animals treated with NLC-ST-DOX. The small particle size, along with the EPR effect and the controlled release of DOX from the particle, associated with the synergic combination between TS and DOX led to an increase of the antitumor efficacy. Therefore, NLC-TS-DOX can be considered a plausible alternative to improve antitumor efficacy in DOX therapeutic regimens.

A new nanoemulsion formulation improves antileishmanial activity and reduces toxicity of amphotericin B.

This work aimed to optimise a new nanoemulsion (NE) formulation loaded with Amphotericin B (AmB) and to evaluate its in vivo antileishmanial activity and in vitro haemolytic toxicity. The influence of gradual increases in pressure, using a high-pressure homogeniser, was evaluated. The NE was characterised for droplet size, polydispersity index, zeta potential and encapsulation efficiency (EE). For antileishmanial activity studies, AmB-NE was administered intravenously in mice infected by Leishmania infantum chagasi, which causes Visceral Leishmaniasis (VL). When the NE was submitted to gradual increases in pressure, the PI values and droplet size decreased. The droplet size (∼145 nm) was lower than that obtained in previous studies. The zeta potential was negative and the EE was almost 100%. The haemolytic toxicity, evaluated on human red blood cells, for AmB-loaded NE was lower than that observed for the conventional AmB (C-AmB). C-AmB at 2 mg/kg was very toxic. In contrast, administration of the AmB-loaded NE, at same dose, did not result in any sign of acute toxicity, promoting a significant reduction in parasite burden as compared to the C-AmB. These findings suggest that this new AmB-loaded NE constitutes an attractive alternative for the treatment of VL due to improved efficacy and lower toxicity.

Increased Body Exposure to New Anti-Trypanosomal Through Nanoencapsulation.

Lychnopholide, a lipophilic sesquiterpene lactone, is efficacious in mice at the acute and chronic phases of Chagas disease. Conventional poly-ε-caprolactone (PCL) and long-circulating poly(D,L-lactide)-block-polyethylene glycol (PLA-PEG) nanocapsules containing lychnopholide were developed and characterized. Lychnopholide presented high association efficiency (>90%) with the nanocapsules. A new, fast and simple HPLC-UV-based bioanalytical method was developed, validated in mouse plasma and applied to lychnopholide quantification in in vitro release kinetics and pharmacokinetics. The nanocapsules had mean hydrodynamic diameters in the range of 100-250 nm, negative zeta potentials (-30 mV to -57 mV), with good physical stability under storage. Atomic force microscopy morphological analysis revealed spherical monodispersed particles and the absence of lychnopholide crystallization or aggregation. Association of lychnopholide to PLA-PEG nanocapsules resulted in a 16-fold increase in body exposure, a 26-fold increase in plasma half-life and a dramatic reduction of the lychnopholide plasma clearance (17-fold) in comparison with free lychnopholide. The improved pharmacokinetic profile of lychnopholide in long-circulating nanocapsules is in agreement with the previously reported improved efficacy observed in Trypanosoma cruzi-infected mice. The present lychnopholide intravenous dosage form showed great potential for further pre-clinical and clinical studies in Chagas disease and cancer therapies.

Impact of dose and surface features on plasmatic and liver concentrations of biodegradable polymeric nanocapsules.

The effect of polymeric nanocapsule dose on plasmatic and liver concentrations 20min after intravenous administration in mice was evaluated. Nanocapsules were prepared with different polymers, namely, poly(D,L-lactide) (PLA), polyethylene glycol-block-poly(D,L-lactide) (PLA-PEG), and PLA with chitosan (PLA-Cs) and compared with a nanoemulsion. These formulations were labelled with a phthalocyanine dye for fluorescent detection. The nanostructures had narrow size distributions upon separation by asymmetric flow field flow fractionation with static and dynamic light scattering detection, with average hydrodynamic diameters in the 130-300nm range, negative zeta potentials, except PLA-Cs nanocapsules, which had a positive zeta potential. Flow cytometry revealed uptake mostly by monocytes and neutrophils in mice and human blood. PLA nanocapsules and the nanoemulsion showed dose-dependent plasma concentrations, where the percentage of plasmatic fluorescence increased with increasing administered dose. In contrast, PLA-PEG nanocapsules led to a dose-independent plasmatic profile. PLA-Cs nanocapsules showed the lowest plasmatic and liver levels of fluorescence at all administered doses and significant intravenous toxicity in mice. This work demonstrates the importance of considering the nanocarrier dose when evaluating pharmacokinetic and biodistribution data and emphasizes the role of surface features in determining the plasmatic and liver concentrations of a poorly soluble lipophilic encapsulated compound.

Correlation between swallowing-related quality of life and videofluoroscopy after head and neck cancer treatment. / Associação entre os achados do questionário de disfagia M. D. Anderson e a videofluoroscopia da deglutição após tratamento do câncer de cabeça e pescoço.

INTRODUCTION: The use of symptom-specific questionnaires on head and neck cancer (HNC), together with objective swallowing measures, can be sensitive to changes in quality of life (QoL) resulting from dysphagia, but this tool is not broadly used as a complement to clinical evaluations. PURPOSE: To analyze the correlation between the M. D. Anderson Dysphagia Inventory (MDADI) questionnaire and videofluoroscopy (VF) in patients treated for head and neck cancer. METHODS: This is a retrospective study with review of clinical data, VF and MDADI results. The study sample was composed of adult patients (>18 y.o.) treated for tumors at the oral cavity, oropharynx, hypopharynx, and larynx, regardless of treatment type. For the VF examination, swallowing of 5 and 20 ml of nectar-thick liquids were considered. The Mann-Whitney nonparametric test was applied to evaluate the correlations between the MDADI and VF. RESULTS: Thirty-nine patients, mostly men (87.18%), with mean age of 61 years participated in the study. Most patients (16) presented oral cavity tumors (41.03%). Twenty-two patients were in advanced clinical stage (IV). Surgery was the most prevalent treatment (41.03%). Approximately half of the participants (20) received oral feeding. The total mean (TM) on the MDADI was 63.36. Comparison between VF and MDADI data showed significant correlation between TM, emotional domain (ED), and physical domain (PD) with penetration during the swallowing of 5 ml. Penetration and aspiration with 20 ml determined worse QoL on the global (p=0.018 and p=0.0053), emotional (p=0.0012 and p=0.027) and physical (p=0.0002 and p=0.0051) domains, and TM (p=0.0023 and p=0.0299), respectively. The presence of stasis did not determine worse QoL. CONCLUSION: Patients treated for HNC who presented penetration/aspiration showed worse QoL on the emotional and physical domains of the MDADI.

Associação entre os achados do questionário de disfagia M. D. Anderson e a videofluoroscopia da deglutição após tratamento do câncer de cabeça e pescoço / Correlation between swallowing-related quality of life and videofluoroscopy after head and neck cancer treatment

RESUMO Introdução A utilização de questionários sintoma-específicos no câncer de cabeça e pescoço (CCP) em conjunto com avaliações objetivas da deglutição pode ser sensível às mudanças na qualidade de vida (QV) decorrentes da disfagia, porém é uma ferramenta pouco utilizada como complemento de avaliações clínicas. Objetivo analisar a associação entre o questionário de disfagia M. D. Anderson (MDADI) com a videofluoroscopia (VF) da deglutição em pacientes tratados do CCP. Método Estudo retrospectivo, com revisão de prontuários, dados da VF e do questionário de disfagia MDADI. Foram incluídos indivíduos maiores de 18 anos, tratados do câncer de cavidade oral, orofaringe, hipofaringe e laringe, independentemente do tratamento curativo. Para o exame de VF, foram consideradas as deglutições de 5 e 20 ml na consistência néctar. O teste não paramétrico de Mann-Whitney foi utilizado para avaliar a associação entre o questionário MDADI e a VF. Resultados Casuística de 39 indivíduos, predomínio de homens, 34 (87,18%), e média de idade de 61 anos. Prevalência de câncer de cavidade oral, 16 (41,03%). Vinte e dois (56,4%) possuíam estádio clínico IV. Cirurgia isolada foi o tratamento mais prevalente, 16 (41,03%). Vinte indivíduos (51,28%) se alimentavam por via oral. A média total (MT) do MDADI foi de 63,36. Na correlação da VF com o MDADI, observou-se associação significante entre MT, domínio emocional (DE) e domínio físico (DFis) com penetração para 5 ml. Penetração e aspiração com 20 ml determinou prejuízo para questão global (p=0,018 e p=0,0053), DE (p=0,0012 e p=0,027), DFis (p=0,0002 e p=0,0051) e MT (p=0,0023 e p=0,0299), respectivamente. A presença de estase não determinou piora da QV. Conclusão Pacientes tratados do CCP que apresentam penetração/aspiração demonstram impacto na qualidade de vida nos DE e DFis.

ABSTRACT Introduction The use of symptom-specific questionnaires on head and neck cancer (HNC), together with objective swallowing measures, can be sensitive to changes in quality of life (QoL) resulting from dysphagia, but this tool is not broadly used as a complement to clinical evaluations. Purpose To analyze the correlation between the M. D. Anderson Dysphagia Inventory (MDADI) questionnaire and videofluoroscopy (VF) in patients treated for head and neck cancer. Methods This is a retrospective study with review of clinical data, VF and MDADI results. The study sample was composed of adult patients (>18 y.o.) treated for tumors at the oral cavity, oropharynx, hypopharynx, and larynx, regardless of treatment type. For the VF examination, swallowing of 5 and 20 ml of nectar-thick liquids were considered. The Mann-Whitney nonparametric test was applied to evaluate the correlations between the MDADI and VF. Results Thirty-nine patients, mostly men (87.18%), with mean age of 61 years participated in the study. Most patients (16) presented oral cavity tumors (41.03%). Twenty-two patients were in advanced clinical stage (IV). Surgery was the most prevalent treatment (41.03%). Approximately half of the participants (20) received oral feeding. The total mean (TM) on the MDADI was 63.36. Comparison between VF and MDADI data showed significant correlation between TM, emotional domain (ED), and physical domain (PD) with penetration during the swallowing of 5 ml. Penetration and aspiration with 20 ml determined worse QoL on the global (p=0.018 and p=0.0053), emotional (p=0.0012 and p=0.027) and physical (p=0.0002 and p=0.0051) domains, and TM (p=0.0023 and p=0.0299), respectively. The presence of stasis did not determine worse QoL. Conclusion Patients treated for HNC who presented penetration/aspiration showed worse QoL on the emotional and physical domains of the MDADI.

Polarity-sensitive nanocarrier for oral delivery of Sb(V) and treatment of cutaneous leishmaniasis.

There is a great need for orally active drugs for the treatment of the neglected tropical disease leishmaniasis. Amphiphilic Sb(V) complexes, such as 1:3 Sb-N-octanoyl-N-methylglucamide complex (SbL8), are promising drug candidates. It has been previously reported that SbL8 forms kinetically stabilized nanoassemblies in water and that this simple dispersion exhibits antileishmanial activity when given by oral route to a murine model of visceral leishmaniasis. The main objective of the present work was to interfere in the structural organization of these nanoassemblies so as to investigate their influence on the oral bioavailability of Sb, and ultimately, optimize an oral formulation of SbL8 for the treatment of cutaneous leishmaniasis. The structural organization of SbL8 nanoassemblies was manipulated through addition of propylene glycol (PG) to the aqueous dispersion of SbL8. The presence of 50% (v/v) PG resulted in the loss of hydrophobic microenvironment, as evidenced by fluorescence probing. However, nanostructures were still present, as demonstrated by dynamic light scattering, small-angle X-ray scattering, and atomic force microscopy (AFM). A remarkable property of these nanoassemblies, as revealed by AFM analysis, is the flexibility of their supramolecular organization, which showed changes as a function of the solvent and substrate polarities. The formulation of SbL8 in 1:1 water:PG given orally to mice promoted significantly higher and more sustained serum levels of Sb, when compared to SbL8 in water. The new formulation, when given as repeated doses (200 mg Sb/kg/day) to BALB/c mice infected with Leishmania amazonensis, was significantly more effective in reducing the lesion parasite burden, compared to SbL8 in water, and even, the conventional drug Glucantime(®) given intraperitoneally at the same dose. In conclusion, this work introduces a new concept of polarity-sensitive nanocarrier that was successfully applied to optimize an oral formulation of Sb(V) for treating cutaneous leishmaniasis.

Polymeric nanocapsules prevent oxidation of core-loaded molecules: evidence based on the effects of docosahexaenoic acid and neuroprostane on breast cancer cells proliferation.

BACKGROUND: Nanocapsules, as a delivery system, are able to target drugs and other biologically sensitive molecules to specific cells or organs. This system has been intensively investigated as a way to protect bioactives drugs from inactivation upon interaction with the body and to ensure the release to the target. However, the mechanism of improved activity of the nanoencapsulated molecules is far from being understood at the cellular and subcellular levels. Epidemiological studies suggest that dietary polyunsaturated fatty acids (PUFA) can reduce the morbidity and mortality from breast cancer. This influence could be modulated by the oxidative status of the diet and it has been suggested that the anti-proliferative properties of docosahexaenoic acid (DHA) are enhanced by pro-oxidant agents. METHODS: The effect of encapsulation of PUFA on breast cancer cell proliferation in different oxidative medium was evaluated in vitro. We compared the proliferation of the human breast cancer cell line MDA-MB-231 and of the non-cancer human mammary epithelial cell line MCF-10A in different experimental conditions. RESULTS: DHA possessed anti-proliferative properties that were prevented by alpha-tocopherol (an antioxidant) and enhanced by the pro-oxidant hydrogen peroxide that confirms that DHA has to be oxidized to exert its anti-proliferative properties. We also evaluated the anti-proliferative effects of the 4(RS)-4-F4t-neuroprostane, a bioactive, non-enzymatic oxygenated metabolite of DHA known to play a major role in the prevention of cardiovascular diseases. DHA-loaded nanocapsules was less potent than non-encapsulated DHA while co-encapsulation of DHA with H2O2 maintained the inhibition of proliferation. The nanocapsules slightly improves the anti-proliferative effect in the case of 4(RS)-4-F4t-neuroprostane that is more hydrophilic than DHA. CONCLUSION: Overall, our findings suggest that the sensitivity of tumor cell lines to DHA involves oxidized metabolites. They also indicate that neuroprostane is a metabolite participating in the growth reducing effect of DHA, but it is not the sole. These results also suggest that NC seek to enhance the stability against degradation, enhance cellular availability, and control the release of bioactive fatty acids following their lipophilicities.

Experimental design of a liposomal lipid system: A potential strategy for paclitaxel-based breast cancer treatment.

Paclitaxel (PTX) is widely used as a first-line treatment for patients with metastatic breast cancer; however, its poor water solubility represents a major challenge for parenteral administration. The encapsulation of the PTX in drug-delivery systems with high affinity for tumor sites could improve the uptake and increase its therapeutic efficacy. In this work, long-circulating and pH-sensitive PEG-coated (SpHL-PTX) and PEG-folate-coated liposomes containing PTX (SpHL-FT-PTX) were prepared, and the physicochemical properties and in vitro cytotoxic activity were evaluated. Both formulations presented adequate physicochemical properties, including a mean diameter smaller than 200 nm, zeta potential values near the neutral range, and an encapsulation percentage higher than 93%. Moreover, SpHL-FT-PTX showed a good stability after storage for 100 days at 4 °C. The viability studies on breast cancer cell lines (MDA-MB-231 and MCF-7) demonstrated cytotoxic activity more pronounced for SpHL-FT-PTX than for SpHL-PTX or free drug for both tumor cell lines. This activity was reduced to a rate comparable to SpHL-PTX when the cells were previously treated with folic acid in order to saturate the receptors. In contrast, in the normal cell line (L929), cell viability was decreased only by free or liposomal PTX in the highest concentrations. A significantly higher selectivity index was obtained after SpHL-FT-PTX treatment compared to SpHL-PTX and free PTX. Therefore, the results of the present work suggest that SpHL-FT-PTX can be a promising formulation for the treatment of metastatic breast cancer.

Improved nonclinical pharmacokinetics and biodistribution of a new PPAR pan-agonist and COX inhibitor in nanocapsule formulation.

We report the in vitro release profile and comparative pharmacokinetics and biodistribution of a new peroxisome proliferator-activated receptor-γ agonist and cyclooxygenase inhibitor (Lyso-7) free or associated to poly(D,L-lactic acid) nanocapsules (NC) after intravenous administration in mice. Lyso-7 pertains to the class of insulin-sensitizing agents that shows potential beneficial effects in diabetes therapy. Monodispersed Lyso-7 NC with a mean diameter of 273 nm with high encapsulation efficiency (83%) were obtained. Lyso-7 dissolution rate was reduced (2.6-fold) upon loading in NC. The pharmacokinetic parameters were determined using a non-compartmental approach. In comparison with Lyso-7 in solution, the plasma-AUC increased 14-fold, the mean residence time 2.6-fold and the mean half-life (t1/2) 1.5-fold for Lyso-7-NC; the Lyso-7 plasma clearance, distribution volume and elimination rate were reduced 13, 10 and 1.4 fold, respectively, which indicates higher retention of encapsulated Lyso-7 in the blood compartment. Upon association with NC, organ exposure to Lyso-7 was higher in the heart (3.6-fold), lung (2.8-fold), spleen (2.3-fold), kidney (2-fold) and liver (1.8-fold) compared to Lyso-7 in solution. The analysis of whole data clearly indicates that body exposure to Lyso-7 was enhanced and the general toxicity reduced upon nanoencapsulation, allowing further evaluation of Lyso-7 in nonclinical and clinical studies.

A reduction of viral mRNA, proteins and induction of altered morphogenesis reveals the anti-HTLV-1 activity of the labdane-diterpene myriadenolide in vitro.

BACKGROUND: Human T-lymphotropic virus 1 (HTLV-1) has been associated with leukemia/lymphoma (ATL) and myelopathy/tropical spastic paraparesis (HAM/TSP), in addition to other inflammatory diseases as well as infection complications. Therapeutic approaches for HTLV-1-related pathologies are limited. The labdane diterpene myriadenolide (AMY) is a natural product that exhibit biological activities, such as anti-inflammatory and antiviral activity as reported for HIV and herpesvirus. RESULTS: We demonstrated that this natural product was able to inhibit the expression of gag-pol mRNA and substantially reduced the expression of the structural proteins p19 and gp46. Comparison of treated and untreated cells shows that AMY alters both the morphology and the release of viral particles. The Atomic Force Microscopy assay showed that the AMY treatment reduced the number of particles on the cell surface by 47%. CONCLUSION: We demonstrated that the labdane diterpene myriadenolide reduced the expression of the structural proteins and the budding of viral particles, besides induces altered morphogenesis of HTLV-1, conferring on AMY a new antiviral activity that may be useful for the development of new compounds with specific anti-HTLV-1 activity.

Investigation of the structural organization of cationic nanoemulsion/antisense oligonucleotide complexes.

Atomic force microscopy image analysis and energy dispersive X-ray diffraction experiments were used to investigate the structural organization of cationic nanoemulsion/oligonucleotide complexes. Oligonucleotides targeting topoisomerase II gene were adsorbed on cationic nanoemulsions obtained by means of spontaneous emulsification procedure. Topographical analysis by atomic force microscopy allowed the observation of the nanoemulsion/oligonucleotide complexes through three-dimensional high-resolution images. Flattening of the oil droplets was observed, which was reduced in the complexes obtained at high amount of adsorbed oligonucleotides. In such conditions, complexes exhibit droplet size in the 600nm range. The oligonucleotides molecules were detected on the surface of the droplets, preventing their fusion during aggregation. A lamellar structure organization was identified by energy dispersive X-ray diffraction experiments. The presence of the nucleic acid molecules led to a disorganization of the lipid arrangement and an expansion in the lattice spacing, which was proportional to the amount of oligonucleotides added.

Chloroaluminium phthalocyanine polymeric nanoparticles as photosensitisers: photophysical and physicochemical characterisation, release and phototoxicity in vitro.

Nanoparticles of poly(d,l-lactide-co-glycolide), poly(d,l-lactide) and polyethylene glycol-block-poly(d,l-lactide) were developed to encapsulate chloroaluminium phthalocyanine (AlClPc), a new hydrophobic photosensitiser used in photodynamic therapy (PDT). The mean nanoparticle size varied from 115 to 274 nm, and the encapsulation efficiency ranged from 57% to 96% due to drug precipitation induced by different types of polymer. All nanoparticle formulations presented negative zeta potential values (-37 mV to -59 mV), explaining their colloidal stability. The characteristic photophysical parameters were analysed: the absorption spectrum profile, fluorescence quantum yield and transient absorbance decay, with similar values for free and nanoparticles of AlClPc. The time-resolved spectroscopy measurements for AlClPc triplet excited state lifetimes indicate that encapsulation in nanocapsules increases triplet lifetime, which is advantageous for PDT efficiency. A sustained release profile over 168 h was obtained using external sink method. An in vitro phototoxic effect higher than 80% was observed in human fibroblasts at low laser light doses (3 J/cm(2)) with 10 µM of AlClPc. The AlClPc loaded within polymeric nanocapsules presented suitable physical stability, improved photophysical properties, sustained released profile and suitable activity in vitro to be considered a promising formulation for PDT.