Nanotechnology strategies to address challenges in topical and cellular delivery of siRNAs in skin disease therapy.

Gene therapy is one of the most advanced therapies in current medicine. In particular, interference RNA-based therapy by small interfering RNA (siRNA) has gained attention in recent years as it is a highly versatile, selective and specific therapy. In dermatological conditions, topical delivery of siRNA offers numerous therapeutic advantages, mainly by inhibiting the expression of target transcripts directly in the skin. However, crossing the stratum corneum and overcoming intracellular barriers is an inherent challenge. Substantial efforts by scientists have moved towards the use of multimodal and multifunctional nanoparticles to overcome these barriers and achieve greater bioavailability in their site of action, the cytoplasm. In this review the most innovative strategies based on nanoparticle and physical methods are presented, as well as the design principles and the main factors that contribute to the performance of these systems. This review also highlights the synergistic contributions of medicine, nanotechnology, and molecular biology to advancing translational research into siRNA-based therapeutics for skin diseases.

Solid Lipid-Polymer Hybrid Nanoplatform for Topical Delivery of siRNA: In Vitro Biological Activity and Permeation Studies.

Small interfering RNA (siRNA) molecules have limited transfection efficiency and stability, necessitating the use of delivery systems to be effective in gene knockdown therapies. In this regard, lipid-polymeric nanocarriers have emerged as a promising class of nanoparticles for siRNA delivery, particularly for topical applications. We proposed the use of solid lipid-polymer hybrid nanoparticles (SLPHNs) as topical delivery systems for siRNA. This approach was evaluated by assessing the ability of SLPHNs-siRNA complexes to internalize siRNA molecules and both to penetrate skin layers in vitro and induce gene knocking down in a skin cell line. The SLPHNs were formed by a specific composition of solid lipids, a surfactant polymer as a dispersive agent, and a cationic polymer as a complexing agent for siRNA. The optimized nanocarriers exhibited a spherical shape with a smooth surface. The average diameter of the nanoparticles was found to be 200 nm, and the zeta potential was measured to be +20 mV. Furthermore, these nanocarriers demonstrated excellent stability when stored at 4 °C over a period of 90 days. In vitro and in vivo permeation studies showed that SLPHNs increased the cutaneous penetration of fluorescent-labeled siRNA, which reached deeper skin layers. Efficacy studies were conducted on keratinocytes and fibroblasts, showing that SLPHNs maintained cell viability and high cellular uptake. Furthermore, SLPHNs complexed with siRNA against Firefly luciferase (siLuc) reduced luciferase expression, proving the efficacy of this nanocarrier in providing adequate intracellular release of siRNA for silencing specific genes. Based on these results, the developed carriers are promising siRNA delivery systems for skin disease therapy.

TNFα siRNA delivery by nanoparticles and photochemical internalization for psoriasis topical therapy.

Psoriasis is a chronic inflammatory skin disease that presents increased expression of tumor necrosis factor α (TNFα), a proinflammatory cytokine. The discovery of RNA interference (RNAi), mediated by short interfering RNA (siRNA), made it possible for the expression of some genes to be eliminated. However, for its application, it is necessary to use carriers that can protect siRNA and release it in the target cells. Herein, we developed a delivery system for siRNA based on hybrid polymer-lipid nanoparticles (PLNs) and combined this system with photochemical internalization (PCI), photoactivating the photosensitizer TPPS2a, to optimize the endosomal escape of TNFα siRNA in the cytoplasm, aiming to use the system as a topical formulation to treat psoriasis. The PLNs composed of 2.0% of Compritol® 888 ATO (lipid), 1.5% of poloxamer 188 and 0.1% of the cationic polymer poly(allylamine hydrochloride) showed an average nanoparticle size of 142 nm, a zeta potential of +25 mV, and the ability to efficiently coencapsulate TPPS2a and complexed siRNA. In addition, these materials did not present cellular toxicity and showed high cellular uptake. In vitro delivery studies using porcine skin model revealed that the PLNs delivered siRNA and TPPS2a into the skin. The efficacy was verified using an in vivo psoriasis animal (hairless mouse) model induced by imiquimod (IMQ) cream. The results revealed that PLN-TPPS2a-TNFα siRNA combined with PCI resulted in a decrease in the levels of TNFα, showing the efficiency of the treatment to silence this cytokine in psoriatic lesions, which was accompanied by a reduction in the redness and scaling of the mouse skin. The results showed the potential of the developed PLNs in combined silencing gene therapy and PCI for topical treatment of psoriasis.

Ex vivo model of human skin (hOSEC) for assessing the dermatokinetics of the anti-melanoma drug Dacarbazine.

Alternative models to replace animals in experimental studies remain a challenge in testing the effectiveness of dermatologic and cosmetic drugs. We proposed a model of human organotypic skin explant culture (hOSEC) to assess the profile of cutaneous drug skin distribution, adopting dacarbazine as a model, and respective new methodologies for dermatokinetic analysis. The viability tests were evaluated in primary keratinocytes and fibroblasts, and skin by MTT and TTC assays, respectively. Then, dacarbazine was applied to the culture medium, and the hOSEC method was applied to verify the dynamics of skin distribution of dacarbazine and determine its dermatokinetic profile. The results of cell and tissue viability showed that both were considered viable. The dermatokinetic results indicated that dacarbazine can be absorbed through the skin, reaching a concentration of 36.36 µg/mL (18,18%) of the initial dose (200 µg/mL) after 12 h in culture. Histological data showed that the skin maintained its structure throughout the tested time that the hOSEC method was applied. No apoptotic cells were observed in the epidermal and dermal layers. No visible changes in the dermo-epidermal junction and no inflammatory processes with the recruitment of defense cells were observed. Hence, these findings suggest that the hOSEC concept as an alternative ex vivo model for assessing the dynamics of skin distribution of drugs, such as dacarbazine, and determining their respective dermatokinetic profiles.

Cytotoxic and chemosensitizing effects of glycoalkaloidic extract on 2D and 3D models using RT4 and patient derived xenografts bladder cancer cells.

Glycoalkaloids have been widely demonstrated as potential anticancer agents. However, the chemosensitizing effect of these compounds with traditional chemotherapeutic agents has not been explored yet. In a quest for novel effective therapies to treat bladder cancer (BC), we evaluated the chemosensitizing potential of glycoalkaloidic extract (GE) with cisplatin (cDDP) in RT4 and PDX cells using 2D and 3D cell culture models. Additionally, we also investigated the underlying molecular mechanism behind this effect in RT4 cells. Herein, we observed that PDX cells were highly resistant to cisplatin when compared to RT4 cells. IC50 values showed at least 2.16-folds and 1.4-folds higher in 3D cultures when compared to 2D monolayers in RT4 cells and PDX cells, respectively. GE + cDDP inhibited colony formation (40%) and migration (28.38%) and induced apoptosis (57%) in RT4 cells. Combination therapy induced apoptosis by down-regulating the expression of Bcl-2 (p < 0.001), Bcl-xL (p < 0.001) and survivin (p < 0.01), and activating the caspase cascade in RT4 cells. Moreover, decreased expression of MMP-2 and 9 (p < 0.01) were observed with combination therapy, implying its effect on cell invasion/migration. Furthermore, we used 3D bioprinting to grow RT4 spheroids using sodium alginate-gelatin as a bioink and evaluated the effect of GE + cDDP on this system. Cell viability assay showed the chemosensitizing effect of GE with cDDP on bio-printed spheroids. In summary, we showed the cytotoxicity effect of GE on BC cells and also demonstrated that GE could sensitize BC cells to chemotherapy.

Advances in lyotropic liquid crystal systems for skin drug delivery.

INTRODUCTION: Lyotropic liquid crystals (LLCs) are organized mesophases with intermediate properties between liquids and solids. The LLC and its liquid crystalline nanoparticles (LCNPs) have attracted great interest from the scientific community in recent years as potential drug delivery systems due to the high internal ordering and symmetry with a wide interfacial area. AREAS COVERED: This article aims to gather information and to provide a description of the highly organized structures of LLCs. Updates on production methods and new insights for LCNPs optimization and physico-chemical and morphological caracterization techniques were discussed. We also discussed why these systems proved to be a platform for the design of nanocarrier drug delivery, with an emphasis on topical and transdermal applications. EXPERT OPINION: Drug delivery platforms are of particular importance to improve the biopharmaceutical aspects of therapies topically. Although several systems can be used, LLC or LCNPs appear to be favored due to their similarity to the lipid structure of the skin. The highly ordered structure and the possibility of chemical modifications make it possible to obtain better clinical responses. The results of several studies support the innovations in this field and predict that these systems can innovate the market of technologies for the treatment of cutaneous diseases and cosmetology.

Therapeutic applications and delivery systems for triptolide.

Triptolide (TPL) is a natural compound and active component of Tripterygium wilfordii Hook F., an Asian native woody vine widely used for over 200 years in Chinese medicine. Hot water, ethanol-ethyl acetate, and chloroform-methanol extracts are the first reported TPL preparations in the literature, and since then, several studies for application in inflammation processes and cancer are described due to the antitumor, anti-inflammatory, and immunosuppressive characteristics of the molecule. However, physicochemical properties such as poor solubility and bioavailability are the main concerns regarding the TPL safety and efficacy in clinical studies since trials have reported adverse side effects alongside the excellent TPL therapeutic effects. Here, we review the main TPL applications and issues related to the drug usage, and a comprehensive summary of diseases is provided. Special emphasis is given to drug delivery systems designed to overcome the TPL physicochemical characteristics such as poor drug solubility, and how to increase efficacy and obtain a safe drug profile. Graphical abstract.

Liquid crystalline nanodispersion functionalized with cell-penetrating peptides improves skin penetration and anti-inflammatory effect of lipoic acid after in vivo skin exposure to UVB radiation.

In this study, the development and the performance of a new targeted liquid crystalline nanodispersion (LCN) by the attachment of cell-penetrating peptides (CPP) onto their surfaces to improve skin delivery of lipoic acid (LA) were evaluated. For that, the synthesis and characterization of this new platform as well as its spatiotemporal analysis from in vitro and in vivo topical application were explored and extensively discussed in this paper. The TAT or D4 peptides were chosen as CPP due to specific target strategies by the charge grouping on the skin surface or target the overexpressed epidermal growth factor receptor (EGFR) of cell membrane of keratinocytes, respectively. Thus, the nanoparticle characterization results when taken together suggested that designed LCNs maintained their hexagonal phase structure, nanoscale particle size, and low polydispersity index even after drug, lipopolymers, and peptide additions, which are proved to be favorable for topical skin delivery. There were no statistical differences among the LCNs investigated, except for superficial charge of LCN conjugated with TAT which may have altered the LCN zeta potential due to cationic charge of TAT amino acid sequence compared with D4. The cumulative amounts of LA retained into the skin were determined to be even higher coming from the targeted LCNs. Moreover, the exogenous antioxidant application of the LA from the LCNs can prevent ROS damage, which was demonstrated by this study with the less myeloperoxidase (MPO) activity and decrease in cytokine levels (TNF-alpha and IL-1ß) generated by the oxidative stress modulation. Together, the data presented highlights the potential of these targeted LCNs, and overall, opens new frontiers for preclinical trials.

Topical delivery of siRNA into skin using ionic liquids.

Skin diseases such as lupus, cancer, psoriasis, and hyperhidrosis can potentially be treated effectively by suppressing allele-specific genes using small interfering RNA (siRNA). Injections of siRNA into skin, though effective, are painful and cover small surface areas and thus are not suitable as a long-term treatment option. Topical delivery of siRNA is an attractive alternative option to mediate RNA interference (RNAi). However, the barrier function of the epidermis impedes effective permeation of siRNA into the skin. Herein, we describe topical delivery of siRNA using ionic liquids (ILs) capable of complexing with siRNA non-covalently and delivering it effectively. Using complementary and synergistic strategies of ionic liquids, we report delivery of effective doses of siRNA into skin. The first strategy involved the use of hydrophobic cations to robe the siRNA and the second strategy involved the use of choline-geranic acid ionic liquid (CAGE) to enhance its dermal penetration. In vitro studies in porcine skin confirmed the synergistic effect of these strategies in enhancing epidermal and dermal penetration. In vivo application of siRNA formulation to SKH-1E hairless mice significantly suppressed GAPDH expression with no clinical evidence of toxicity. This is a simple, personalized, and scalable platform for effective topical delivery of siRNA for treating genetic skin diseases.

Microemulsion co-delivering vitamin A and vitamin E as a new platform for topical treatment of acute skin inflammation.

In this study, we developed a water-in-oil microemulsion containing vitamin A (retinol) and vitamin E (α-tocopherol), which serves as a multifunctional nanosystem that co-delivers antioxidants and displayed additive effect against acute skin inflammation. Microemulsion (ME) was prepared by mixing a surfactant blend (Tween 80 and propylene glycol, 5:1) with isopropyl myristate and water (ratio of 50:40:10, respectively). Vitamin A (0.05% w/w concentration) and/or vitamin E (0.1% w/w concentration) were incorporated into the surfactant mixture of ME by stirring with a magnetic stirrer for 30 min. This multifunctional ME displayed physical stability, with low cytotoxicity in 3T3 cell line, as well as cellular internalization into the cytosol. In vivo treatments using ME delivering α-tocopherol reduced dermal expression of TNF-α by 1.3-fold (p < 0.01), when compared to unloaded ME treatment group. When retinol was added into the ME containing α-tocopherol, it further reduced TNF-α expression by 2-fold (p < 0.001), suggesting the additive effect of vitamin E and vitamin A in the treatment against skin inflammation. In conclusion, we successfully developed the use of water-in-oil ME to pack both vitamin E and vitamin A, and demonstrated for the first time its anti-inflammatory potential when applied topically to TPA-induced inflamed skin.

Nanotechnology approaches in the current therapy of skin cancer.

Skin cancer is a high burden disease with a high impact on global health. Conventional therapies have several drawbacks; thus, the development of effective therapies is required. In this context, nanotechnology approaches are an attractive strategy for cancer therapy because they enable the efficient delivery of drugs and other bioactive molecules to target tissues with low toxic effects. In this review, nanotechnological tools for skin cancer will be summarized and discussed. First, pathology and conventional therapies will be presented, followed by the challenges of skin cancer therapy. Then, the main features of developing efficient nanosystems will be discussed, and next, the most commonly used nanoparticles (NPs) described in the literature for skin cancer therapy will be presented. Subsequently, the use of NPs to deliver chemotherapeutics, immune and vaccine molecules and nucleic acids will be reviewed and discussed as will the combination of physical methods and NPs. Finally, multifunctional delivery systems to codeliver anticancer therapeutic agents containing or not surface functionalization will be summarized.

Nanostructured lipid carrier co-delivering tacrolimus and TNF-α siRNA as an innovate approach to psoriasis.

Since psoriasis is an immuno-mediated skin disease, long-term therapies are necessary for its treatment. In clinical investigations, tacrolimus (TAC), a macrolide immunosuppressive inhibitor of calcineurin, arises as an alternative for the treatment of psoriasis, acting in some cytokines involved in the pathogenesis of the disease. Here, we aim to study the psoriasis treatment with TAC and siRNA for one of most cytokines expressed in psoriasis, the TNF-α. A multifunctional nanostructure lipid carrier (NLC) was developed to co-delivery TAC and siRNA. Results showed that the particle size and zeta potential were around 230 nm and + 10 mV, respectively. The release study demonstrated a controlled release of TAC, and the permeation and retention profile in the skin tissue show to be promising for topical application. The cell viability and uptake in murine fibroblast presented low toxicity associated to uptake of NLC in 4 h, and finally, the in vivo animal model demonstrates the efficiency of the NLC multifunctional, exhibiting a reduction of the cytokine TNF-α expression about 7-fold and presenting a synergic effect between the TAC and TNF-α siRNA. The developed system was successfully to treat in vivo psoriatic animal model induced by imiquimod and the synergic combination was reported here for the first time. Graphical abstract.

Current Non-viral siRNA Delivery Systems as a Promising Treatment of Skin Diseases.

BACKGROUND: Gene therapy is a new approach to discover and treat many diseases. It has attracted considerable attention from researchers in the last decades. The gene therapy through RNA interference has been considered one of the most recent and revolutionary approaches used in individualized therapy. In the last years, we have witnessed the rapid development in the field of the gene silencing and knockdown by topical siRNA. Its application in gene therapy has become an attractive alternative for drug development. METHODS: This article will address topical delivery of siRNA as a promising treatment for skin disorders. An update on the advances in siRNA-based nanocarriers as a powerful therapeutic strategy for several skin diseases will be discussed giving emphasis on in vitro evaluations. RESULTS: Through the in-depth review of the literature on the use of siRNAs for skin diseases we realize how widespread this use is. We have also realized that nanoparticles as non-viral vectors are increasingly being explored. Skin diseases where the use of siRNA has been explored most are skin cancer (melanoma and nonmelanoma), psoriasis, vitiligo, dermatitis and leprosy. But we also report here other diseases where the use of siRNA has been growing as acne, alopecia areata, cutaneous leishmaniasis, mycoses, herpes, epidermolysis bullosa and oculocutaneous albinism. Also highlighted, the first clinical trial of siRNA for cutaneous diseases, aimed at Pathyounychia Congenita. CONCLUSION: The treatment of skin diseases based on topical delivery of siRNA, which act by inhibiting the expression of target transcripts, offers many potential therapeutic advantages for suppressing genes into the skin.

In Vitro TyRP-1 Knockdown Based on siRNA Carried by Liquid Crystalline Nanodispersions: an Alternative Approach for Topical Treatment of Vitiligo.

PURPOSE: Vitiligo is a skin disease characterized by depigmentation and the presence of white patches that are associated with the loss of melanocytes. The most common explanation for the cause of this condition is that it is an autoimmune condition. TyRP-1 is involved in melanin pigment synthesis but can also function as a melanocyte differentiation antigen. This protein plays a role in the autoimmune destruction of melanocytes, which results in the depigmentation, characteristic of this disease. In this study, we evaluated liquid crystalline nanodispersions as non-viral vectors to deliver siRNA-TyRP-1 as an alternative for topical treatment of vitiligo. METHODS: Liquid crystalline nanodispersions were obtained and characterized with respect to their physical-chemical parameters including size, PdI and zeta potential, as well as Small Angle X-ray Scattering and complexing to siRNA. The effects of the liquid crystalline nanodispersions on the cellular viability, cell uptake and levels of the knockdown target TyRP-1 were evaluated in melan-A cells after 24 h of treatment. RESULTS: The liquid crystalline nanodispersions demonstrated adequate physical-chemical parameters including nanometer size and a PdI below 0.38. These systems promoted a high rate of cell uptake and an impressive TyRP-1 target knockdown (> 80%) associated with suitable loading of TyRp-1 siRNA. CONCLUSIONS: We demonstrated that the liquid crystalline nanodispersions showed promising alternative for the topical treatment of vitiligo due to their physical parameters and ability in knockdown the target protein involved with autoimmune destruction of melanocytes.

Ketoprofen Microemulsion for Improved Skin Delivery and In Vivo Anti-inflammatory Effect.

We have designed a microemulsion (ME) containing Ketoprofen (KET) for anti-inflammatory effect evaluated using the rat paw edema model. The ME was prepared by adding propylene glycol (PG) loaded with 1% KET/water (3:1, w/w), to a mixture of sorbitan monooleate and polysorbate 80 (47.0%) at 3:1 (w/w) and canola oil (38.0%). The physicochemical characterization of KET-loaded ME involved particle size and zeta potential determination, entrapment efficiency, calorimetric analysis, and in vitro drug release. The in vivo anti-inflammatory study employed male Wistar rats. Measurement of the foot volume was performed using a caliper immediately before and 2, 4, and 6 h after injection of Aerosil. KET-loaded ME showed particle size around 20 nm, with zeta potential at -16 mV and entrapment efficiency at 70%. Moreover, KET was converted to the amorphous state when loaded in the formulation and it was shown that the drug was slowly released from the ME. Finally, the in vivo biological activity was similar to that of the commercial gel, but ME better controlled edema at 4 h. These results demonstrated that the ME formulation is an alternative strategy for improving KET skin permeation for anti-inflammatory effect. Furthermore, our findings are promising considering that the developed ME was loaded with only 1% KET, and the formulation was able to keep a similar release profile and in vivo effect compared to the commercial gel with 2.5% KET. Therefore, the KET-loaded developed herein ME is likely to have a decreased side effect compared with that of the commercial gel, but both presented the same efficacy.

Avaliação da citotoxicidade da microemulsão contendo o fotossensibilizador ZNPCSO4, sobre cultura de célula de linhagem de câncer de mama / Evaluación de la citotoxicidad de la microemulsión que contiene el fotosensibilizador ZNPCSO4, en la cultura celular del cáncer de mama / Cytotoxicity evaluation of microemulsion containing the photosensibilizer ZNPCSO4, on breast cancer cell culture line

Introdução: O câncer de mama tem alta incidência e mortalidade dentre todos os tipos de câncer e, apesar de todos os avanços tecnológicos recentes,mantém-se ainda como um problema mundial de saúde pública. Novas estratégias terapêuticas, como a Terapia Fotodinâmica, têm sido usadas para tratarvários tipos de tumores de tecidos moles e hiperplasia. Objetivo: Caracterizar o efeito citotóxico da microemulsão contendo zinco ftalocianina tetrasulfonada(ZnPcSO4). Material e Método: A microemulsão (ME) foi preparada por adição da fase aquosa (15%) composta de propilenoglicol e água (3:1), em umamistura de fase oleosa composta pelos surfactantes monooleato de sorbitanol e polissorbato 80 (47%) e óleo de canola (38%). Para o teste de citotoxicidadefoi realizado o ensaio de MTT com as células MCF-7 de adenocarcinoma de mama humana. Resultados: Os baixos valores de condutividade obtidos (0,63e 0,53 para ME com e sem ZnPcSO4, respectivamente) indicaram que a ME era de tipo água/óleo. O diâmetro interno das partículas foi na ordem de 60nmna presença e ausência do ZnPcSO4 sugerindo que a presença do fotossensibilizante não alterou as propriedades do sistema nanoestruturado. O índicede polidispersividade foi menor do que 0,10, enquanto que a carga superfi cial da nanopartícula foi negativa, em concordância com as característicasdos compostos utilizados para o preparo da ME. A atividade metabólica das células não sofreu interferências com as MEs não demonstrando toxicidade...

Introduction: Breast cancer has a high incidence and mortality among all types of cancer and, despite all the recent technological advances, still remains aworldwide public health problem. New therapeutic strategies, such as Photodynamic Therapy, have been used to treat various types of soft tissue tumorsand hyperplasia. Objective: To characterize the cytotoxic effect of the zinc-containing microemulsion containing tetrasulf...

Introducción: El cáncer de mama tiene alta incidencia y mortalidad entre todos los tipos de cáncer y, a pesar de todos los avances tecnológicos recientes,sigue siendo un problema mundial de salud pública. Nuevas estrategias terapéuticas, como la Terapia Fotodinámica, se han utilizado para tratar varios tiposde tumores de tejidos blandos e hiperplasia. Objetivo: Caracterizar el efecto citotóxico de la microemulsión que contiene el cinc ftalocianina tetrasulfonada(ZnPcSO4). Material y método: La microemulsión (ME) fue preparada por adición de la fase acuosa (15%) compuesta de propilenglicol y agua (3: 1), enuna mezcla de fase oleosa compuesta por los surfactantes monooleato de sorbitanol y polisorbato 80 (47%), Y el aceite de canola (38%). Para la pruebade citotoxicidad se realizó el ensayo de MTT con las células MCF-7 de adenocarcinoma de mama humana. Resultados: Los bajos valores de conductividadobtenidos (0,63 y 0,53 para ME con y sin ZnPcSO4, respectivamente)...

Comparação do perfil de liberação de nitrato de isoconazol em formas farmacêuticas semissólidas usando modelo matemático de diferença (f1) e semelhança (f2) / Comparación de perfiles de liberación del nitrato isoconazol en formas farmacéuticas semisólido utilizando el modelo de diferencia matemática (f1) y semejanza (f2) / Comparison of isoconazole nitrate release profile in pharmaceutical semisolid forms using f1 (differential) and f2 (similarities) mathematical models

Introdução: A comprovação da eficácia e segurança de produtos é dada por meio dos estudos de equivalência farmacêutica e bioequivalência. Abioequivalência entre os dois produtos farmacêuticos pode ser demonstrada por meio de estudos diferentes: farmacocinético, farmacodinâmico,clínicos comparativos ou ensaios in vitro. Objetivo: Comparar o perfil de liberação in vitro do nitrato de isoconazol a partir de medicamentosreferência e genéricos, aplicando dois diferentes modelos estatísticos para a análise dos resultados baseados nos fatores F1 e F2. Material eMétodo: Os perfis de liberação do nitrato de isoconazol foram determinados em condições previamente otimizadas em célula de difusão verticalcontendo solução 0,01M de fosfato de sódio (pH 7,4) como meio receptor (7 mL) e membrana sintética de acetato de celulose (com área dedifusão de 1,77 cm2). Após 1, 2, 3, 4, 6 e 8 horas de experimento, exatos 1,0 mL da solução receptora foi coletada e a quantidade de nitratode isoconazol presente na solução foi analisada por método validado de Cromatografia Líquida de Alta Eficiência. O fluxo de entrega do nitratode isoconazol foi obtido com base no perfil de liberação utilizando a porção linear da curva. Os valores de diferença e semelhança, F1 e F2,respectivamente, obtidos entre as curvas de liberação de nitrato de isoconazol, foram aplicados como uma ferramenta de análise da equivalênciafarmacêutica de formas semissólidas. Resultados: Houve equivalência farmacêutica entre os medicamentos referência e genérico, considerando osnúmeros de lotes utilizados nestes ensaios...

Introduction: Evidence of efficacy and safety of products is given through pharmaceutical equivalence and bioequivalence studies. Bioequivalencebetween the two pharmaceutical products can be demonstrated by different studies: pharmacokinetics, pharmacodynamics, comparative clinicaltrials or in vitro assay. Objective: To compare the in vitro release profile of isoconazole nitrate from reference and generic drugs using twodifferent statistical models based on factors F1 and F2. Material and Method: The release profiles of isoconazole nitrate were determined underpre-optimized conditions in a vertical diffusion cell containing 0.01M sodium phosphate solution (pH 7.4) as receptor medium (7mL) and syntheticcellulose acetate membrane (with diffusion area of 1.77 cm2). After 1, 2, 3, 4, 6 and 8 hours of permeation, exactly 1.0 mL of the receptor solutionwas collected and the amount of isoconazole nitrate was analyzed by validated High Performance Liquid Chromatography method. The isoconazolenitrate delivery flow was obtained based on the release profile using the linear portion of the curve. The values of difference and similarity, F1 andF2 respectively, obtained between the release curves of isoconazole nitrate from generic and reference products, were applied as a tool for analysisof the pharmaceutical equivalence of semisolid forms. Results: There was a pharmaceutical equivalence between reference and generic drugs,considering the batches number...

Introducción: La prueba de la eficacia y seguridade del producto está dada por los estudios de equivalência farmacéutica y bioequivalência. Labioequivalencia entre las dos productos farmacêuticos puede ser demostrada en diversos estudios: farmacocinética, farmacodinámica, clínicos yencomparación in vitro. Objetivo: Comparar el perfil de liberación in vitro de nitrato de isoconazol de medicamentos genéricos y de referencia,utilizando dos diferentes modelos estadísticos para el análisis de los resultados en función de los factores F1 y F2. Material y Método: Los perfilesde liberación de isoconazol han sido determinados en condiciones de nitrato isoconazol previamente optimizados en la célula de difusión verticalque contiene solución de fosfato de sodio 0,01 M (pH 7,4) como medio receptor (7 ml) y membrana sintética (con 1,77 cm2 área de difusión).Después de 1, 2, 3, 4, 6 y 8 horas de experimento se recogieronexactamente 1,0 ml de solución de receptor y la cantidad de nitrato isoconazolen lasolución se analizó por lacromatografía líquida de alto rendimiento. Se obtuvo el flujo de entrega isoconazol nitrato basado en el perfil deliberación usando la parte lineal de la curva. Las diferencias y semejanza de los valores, F1 y F2, respectivamente, obtenidos a partir de las curvasde liberación de nitrato isoconazol, se aplicaron como una herramienta de análisis de equivalência farmacéutica de formas semisólidas. Resultados:Se demostró la equivalência farmacéutica entre lós...

Development and characterization of novel 1-(1-Naphthyl)piperazine-loaded lipid vesicles for prevention of UV-induced skin inflammation.

1-(1-Naphthyl)piperazine (1-NPZ) has shown promising effects by inhibiting UV radiation-induced immunosuppression. Ultradeformable vesicles are recent advantageous systems capable of improving the (trans)dermal drug delivery. The aim of this study was to investigate 1-NPZ-loaded transethosomes (NPZ-TE) and 1-NPZ-loaded vesicles containing dimethyl sulfoxide (NPZ-DM) as novel delivery nanosystems, and to uncover their chemopreventive effect against UV-induced acute inflammation. Their physicochemical properties were evaluated as follows: vesicles size and zeta potential by dynamic and electrophoretic light scattering, respectively; vesicle deformability by pressure driven transport; rheological behavior by measuring viscosity and I-NPZ entrapment yield by HPLC. In vitro topical delivery studies were performed in order to evaluate the permeation profile of both formulations, whereas in vivo studies sought to assess the photoprotective effect of the selected formulation on irradiated hairless mice by measuring myeloperoxidase activity and the secretion of proinflammatory cytokines. Either NPZ-TE or NPZ-DM exhibited positive results in terms of physicochemical properties. In vitro data revealed an improved permeation of 1-NPZ across pig ear skin, especially by NPZ-DM. In vivo studies demonstrated that NPZ-DM exposure was capable of preventing UVB-induced inflammation and blocking mediators of inflammation in mouse skin. The successful results here obtained encourage us to continue these studies for the management of inflammatory skin conditions that may lead to the development of skin cancers.

Quantification of lipoic acid from skin samples by HPLC using ultraviolet, electrochemical and evaporative light scattering detectors.

Lipoic acid (LA) is an endogenous organosulfur compound with potent antioxidant property. LA is often used as a drug for the treatment of skin disorders. For the accomplishment of topical applications of LA appropriate drug quantification methods are essential. Thus far, no HPLC methods have been reported for the measurement of LA extracted from skin. In this article we report on the development and validation of three sensitive and specific HPLC methods for LA and dihydrolipoic acid (DHLA) using ultraviolet (UV), electrochemical (EC) or evaporative light scattering (ELS) detection. These methods demonstrate different linearity ranges. The chromatographic separations were performed by RP-HPLC (250 × 4 mm, 5 µm) with isocratic elution using an acidic mobile phase for the three detection techniques. The lower limits of detection and quantification were 0.04 and 0.08 ng LA, respectively, for HPLC coupled to ELS, an innovative detector for LA with high sensitivity. The extraction of LA from skin samples showed recoveries greater than 71%. The recovered LA concentrations from stratum corneum and epidermis+dermis layers were: 5.41 ± 0.56 and 4.92 ± 0.33 µg/mL, respectively for HPLC/UV and 6.52 ± 0.49 and 5.01 ± 0.41 µg/mL, respectively, for HPLC/EC for the added LA concentration (6.67 µg/mL), and 8.88 ± 0.46 and 8.95 ± 0.08 µg/mL, respectively, for HPLC/ELS for the added LA concentration (10 µg/mL). These three optimized HPLC methods allowed for a simple, rapid and reliable determination of LA in human skin. They should be useful for the development of drug delivery systems for topical applications of LA.

Optimization of protoporphyrin IX skin delivery for topical photodynamic therapy: Nanodispersions of liquid-crystalline phase as nanocarriers.

Nanodispersions of liquid-crystalline phases (NLPs) composed of monoolein and oleic acid were chosen as nanocarriers to improve the topical retention of the photosensitizer protoporphyrin IX (PpIX) and thereby optimize photodynamic therapy (PDT) using this photosensitizer. The nanodispersions were characterized by polarized light microscopy, small-angle X-ray diffraction and dynamic light scattering. The stability and encapsulation efficiency (EE%) of the nanodispersions were also evaluated. In vitro and in vivo skin penetration studies were performed to determine the potential of the nanodispersions for cutaneous application. In addition, skin penetration and skin irritancy (in an animal model) after in vivo application were visualized by fluorescence light microscopy. The nanodispersion obtained was characterized as a monodisperse system (~150.0 nm) of hexagonal liquid-crystalline phase, which provided a high encapsulation efficiency of PpIX (~88%) that remained stable over 90 days of investigation. Skin penetration studies demonstrated that the nanodispersion enhanced PpIX skin uptake 11.8- and 3.3-fold (in vitro) and 23.6- and 20.8-fold (in vivo) compared to the PpIX skin uptake of control formulations, respectively. In addition, the hexagonal phase nanodispersion did not cause skin irritation after application for two consecutive days. Overall, the results show that the nanocarrier developed is suitable for use in topical PDT with PpIX.