Ana Salgado

In vivo delivery of peptides and Toll-like receptor ligands by mannose-functionalized polymeric nanoparticles induces prophylactic and therapeutic anti-tumor immune responses in a melanoma model.

We hypothesized that the co-entrapment of melanoma-associated antigens and the Toll-like receptor (TLR) ligands Poly(I:C) and CpG, known to be Th1-immunopotentiators, in mannose-functionalized aliphatic polyester-based nanoparticles (NPs) could be targeted to mannose receptors on antigen-presenting cells and induce anti-tumor immune responses. High entrapment efficiencies of antigens and immunopotentiators in 150nm NPs were obtained. The co-entrapment of the model antigen ovalbumin and the TLR ligands was crucial to induce high IgG2c/IgG1 ratios and high levels of IFN-γ and IL-2. Mannose-functionalization of NPs potentiated the Th1 immune response. The nanoparticulate vaccines decreased the growth rate of murine B16F10 melanoma tumors in therapeutic and prophylatic settings. The combination of mannose-functionalized NPs containing MHC class I- or class II-restricted melanoma antigens and the TLR ligands induced the highest tumor growth delay. Overall, we demonstrate that the multifunctional properties of NPs in terms of targeting and antigen/adjuvant delivery have high cancer immunotherapeutic potential.

Development of functionalized nanoparticles for vaccine delivery to dendritic cells: a mechanistic approach

AIM Produce biodegradable nanoparticles to target antigen-presenting cells (APCs) and evaluate their potential to be used as a vaccine delivery system. MATERIALS & METHODS Untargeted PEGylated poly(d,l-lactic-co-glycolide)-based nanoparticles and mannose-grafted nanoparticles were formulated and physicochemically characterized. Immortalized and primary APCs were used to study nanoparticle internalization patterns. The endocytic pathways and intracellular trafficking followed by nanoparticles were also investigated. RESULTS & DISCUSSION Nanoparticles displayed mannose residues available for binding at the nanoparticle surface. Different nanoparticle internalization patterns by immortalized and primary APCs were verified. Macropinocytosis, clathrin-mediated endocytosis, caveolin- and lipid raft-dependent endocytosis are involved in nanoparticles internalization. Nanoparticles demonstrate both endolysosomal and cytosolic localizations and a tendency to accumulate nearby the endoplasmic reticulum. CONCLUSION The developed nanoparticles might drive antigens to be presented through MHC class I and II molecules to both CD8(+) and CD4(+) T cells, favoring a complete and coordinated immune response.

In vitro and in vivo topical delivery studies of tretinoin-loaded ultradeformable vesicles

INTRODUCTION Ultradeformable vesicles are highly promising tools to enhance the percutaneous transport of different drugs such as tretinoin across the skin barrier and also to increase the formulation stability at absorption site and reduce the drug induced irritation. METHODS Topical delivery of tretinoin-loaded ultradeformable vesicles (tretinoin-UDV) was evaluated concerning different studies, such as: the release and permeation profiles (tape stripping); skin penetration (fluorescence analysis); induced electrical changes in skin barrier properties; cytotoxicity (Trypan Blue assay) and skin irritation in in vivo conditions (Draize test). The novel formulation performance was also compared to a commercial tretinoin formulation regarding in vivo studies. RESULTS It was obtained a sustained and controlled drug release, as expected for UDV formulation. In addition, a dermal delivery was observed regarding the permeation study since it was not detected any drug amount in the receptor phase after 24h. Nile Red-UDV stained intensively mostly in the stratum corneum, corroborating the tape stripping results. Tretinoin-UDV decreased skin resistance, suggesting its ability to induce skin barrier disruption. Finally, the formulation vehicle (empty UDV) and tretinoin-UDV were not toxic under in vitro and in vivo conditions, at least, at 5×10(-3)mg/mL and 0.5mg/mL of tretinoin, respectively. CONCLUSION Tretinoin-UDV is a promising delivery system for tretinoin dermal delivery without promoting skin irritation (unlike other commercial formulations), which is quite advantageous for therapeutic purpose.

Cold processed oil-in-water emulsions for dermatological purpose: formulation design and structure analysis

Abstract The aim of this work is to develop, optimize and characterize cold process emulsions that are stable at acidic pH. The main surfactant was selected according to the hydrophilic lipophilic balance (HLB) concept and surface tension, whereas polymers were selected by viscoelastic measurements and analytical centrifugation. It was showed that the inclusion of methyl vinyl ether/maleic anhydride copolymer crosslinked with decadiene (PVM/MA) increased the storage modulus (G′) of the gels (23.9–42.1 Pa) two-fold and the droplet migration decreased from 3.66% to 0.95%/h. Cetrimide was selected as a preservative based on its microbiological results and additional contribution to the stability of the emulsions. Four emulsions were developed that differed by the co-emulsifier used (PEG-20 glyceril laurate and polyglyceryl-4-isostearate) and the glycol (2-methyl-2,4-pentanediol and ethoxydiglycol). Viscoelastic measurements and droplet size/microscopic analysis showed that the structure of PEG-20 glyceril laurate emulsion (η’ = 76.0 Pa.s at 0.01 Hz and 32.9 ± 3.7 µm, respectively) was stronger compared to polyglyceryl-4-isostearate (η’ = 37.4 Pa.s at 0.01 Hz and 37.8 ± 15.7 µm, respectively). Differential scanning calorimetry (DSC) results were in accordance with the latter and showed that PEG-20 glyceril laurate with 2-methyl-2,4-pentanediol corresponded to the strongest structure (|224.4| W °C g−1). This cold process allowed a total production savings of more than 17% when compared to the traditional hot process.

Topical gels of etofenamate: in vitro and in vivo evaluation

Abstract Non-steroid anti-inflammatory drugs (NSAIDs), such as etofenamate, are among the most prescribed drugs used for their analgesic, anti-rheumatic, antipyretic and anti-inflammatory properties. Topical formulations have the main advantage of targeted delivery. However, drugs must overcome the skin due to its role as a physical and chemical barrier against the penetration of chemicals and microorganisms. This barrier must be altered to allow the permeation of drugs at a suitable rate to the desired site of activity. Permeation modulators can intercalate the skin outer layers causing structure disruption, opening an energetically favourable route for the drug to diffuse through. The aim of this work was the development of hydroalcoholic gels containing 5.0% (w/w) of etofenamate for topical administration with anti-inflammatory activity and enhanced drug delivery. The physical and chemical characterization, in vitro release and permeation studies and in vivo anti-inflammatory activity were assessed. The gel with 30% ethanol showed in vivo anti-inflammatory activity with suitable physical chemical and microbiologic characteristics. In vitro release and permeation studies revealed that the different amounts of ethanol used influenced the release profiles of etofenamate. Moreover, it was demonstrated that this formulation is an adequate vehicle for the etofenamate skin permeation.

Safety assessment and biological effects of a new cold processed SilEmulsion for dermatological purpose.

It is of crucial importance to evaluate the safety profile of the ingredients used in dermatological emulsions. A suitable equilibrium between safety and efficacy is a pivotal concern before the marketing of a dermatological product. The aim was to assess the safety and biological effects of a new cold processed silicone-based emulsion (SilEmulsion). The hazard, exposure, and dose-response assessment were used to characterize the risk for each ingredient. EpiSkin assay and human repeat insult patch tests were performed to compare the theoretical safety assessment to in vitro and in vivo data. The efficacy of the SilEmulsion was studied using biophysical measurements in human volunteers during 21 days. According to the safety assessment of the ingredients, 1,5-pentanediol was an ingredient of special concern since its margin of safety was below the threshold of 100 (36.53). EpiSkin assay showed that the tissue viability after the application of the SilEmulsion was 92 ± 6% and, thus considered nonirritant to the skin. The human studies confirmed that the SilEmulsion was not a skin irritant and did not induce any sensitization on the volunteers, being safe for human use. Moreover, biological effects demonstrated that the SilEmulsion increased both the skin hydration and skin surface lipids.

Development, stability and in vitro permeation studies of gels containing mometasone furoate for the treatment of dermatitis of the scalp

Dermatological inflammatory diseases such as atopic dermatitis, psoriasis and seborrhoeic dermatitis often affect the scalp and the eyebrows. Although there are many dosage forms available, these are particularly critical anatomic regions for application of topical formulations because of the presence of hair. Lotions are therefore the recommended type of drug delivery system for these areas. The presence of hair may limit the application and thus the acceptability of the formulation and its compliance. Because of its low apparent viscosity, lotion application is unpleasant. Gels, given their consistency and adhesiveness, are a suitable alternative to lotions in this situation. The aim of this study was to formulate a stable gel containing mometasone furoate, which is an anti-inflammatory and anti-pruritic corticosteroid, in order to improve topical treatment of scalp dermatitis. In this study, pharmaceutical development, physical-chemical characterization, stability and in vitro permeation studies were performed. In terms of the pH, viscosity, assay and macroscopic and microbiological properties, the gel was stable over the period of study. The in vitro permeation studies allowed the characterization of the mometasone furoate permeation profile for the gel through different membranes. Mometasone furoate presented a slow permeation through the skin. This gel appears safe for topical application.

Mometasone furoate hydrogel for scalp use: in vitro and in vivo evaluation

Abstract Dermatological inflammatory diseases often affect the scalp and the eyebrows. Common dosage forms available on the market for those situations are lotions; however, the presence of hair limits their use. Gels, for their consistency and adhesiveness, are a suitable alternative to the lotions in these situations. The aim of this study was to develop a new stable gel containing mometasone furoate (MF), with anti-inflammatory activity and a controlled delivery, to improve topical treatment of scalp dermatitis. Pharmaceutical development, physical and chemical characterization, stability, in vitro release and permeation studies and in vivo anti-inflammatory activity were performed. The gel presented an acidic pH and an apparent viscosity of 35 Pa.s. The microbiological analysis showed that the results were within the established specification limits. The release and the permeation profiles suggest that the drug is mainly retained in the upper skin layers. MF gel was tested in an animal model of cutaneous inflammation and presented similar anti-inflammatory activity compared to a commercially available MF dosage form. The gel was chemically, physically and microbiologically stable. The results suggest that the developed hydrogel formulation containing MF can be of actual value for improving the clinical effectiveness in the treatment of scalp dermatitis.

Development of a Topical Formulation Containing S. Lutea Extract: Stability, In Vitro Studies and Cutaneous Permeation

Flavonoids have been widely incorporated into cosmetic and dermatological formulations, affording benefits such as antioxidant action, improved skin tone and fewer lines and wrinkles. Brazil has a huge biodiversity, with one of the richest flora in the world, and existing studies justify the quest for greater research efforts in this area. The cajazeira (Spondias lutea L.), a plant of the Anacardiaceae family from tropical America, is widely disseminated in Brazil. This plant was chosen because of the presence of flavonoids that exhibit antioxidant activity. The purpose of this research was to develop a stable topical formulation containing Spondias lutea extract with the aim of preventing skin diseases caused by UV radiation. Hydro ethanolic extract of Spondias lutea fruit was prepared and assayed for its the flavonoids content. The antioxidant activity was estimated by DPPH and superoxide assay. The physicochemical stability and skin permeation of the cream containing 8% (w/w) of extract were assessed. The results showed that the S. lutea extract possessed antioxidant activity, and that it is possible to obtain a stable cosmetic containing the extract, which is able to penetrate the skin. Thus, it is possible to use this extract to produce an anti-aging cosmetic.

Nystatin and lidocaine pastilles for the local treatment of oral mucositis

Abstract Oral mucositis (OM) is a common adverse reaction to radiotherapy and chemotherapy in oncology. Its treatment requires oral formulations that enhance therapy compliance, improve administration and ensure drug effectiveness. Solid dosage forms that act by slow dissolution, such as pastilles, are an effective alternative to mouthwashes, for their versatility, ease of administration and extended residence time in the oral cavity. The present work describes the development and stability studies of an innovative formulation of nystatin and lidocaine pastilles for the treatment of oral mucositis. Full pharmaceutical quality testing was carried out, including disintegration and dissolution testing, texture profile analysis, grittiness and an antifungal activity testing. A soft pastille formulation containing 0.25% lidocaine and 78,000 IU nystatin was obtained, presenting suitable pharmaceutical characteristics, as a disintegration time of 17 ± 2 min, dissolution rate and microbiological and physicochemical for 30 days when stored at 2–8 °C under light protection. Palatability was also evaluated, being well accepted by a panel of 38 healthy volunteers. This formulation allows an accurate drug dosing by the prescriber, while enabling the patients to control the retention time of the drugs in the oral cavity and consequently manage their pain treatment.