Proefschrift Universiteit van Amsterdam. / Auteursnaam op omslag: Jet Gisolf. Met bibliogr., lit. opg. - Met samenvatting in het Nederlands.

PEREIRA, Juliana Fernandes. Efeito in vitro do saquinavir isolado e em combinação com o itraconazol frente a cepas de histoplasma capsulatum var. capsulatum. 2012. 96 f. Dissertação (Mestrado em Ciências Médicas) - Universidade Federal do Ceará. Faculdade de Medicina, Fortaleza, 2012. / Submitted by denise santos (denise.santos@ufc.br) on 2015-03-23T13:14:03Z No. of bitstreams: 1 2012_dis_jfpereira.pdf: 994358 bytes, checksum: 841297998e1d9cd11701f0d79cbb1e7c (MD5) / Approved for entry into archive by denise santos(denise.santos@ufc.br) on 2015-03-23T13:15:27Z (GMT) No. of bitstreams: 1 2012_dis_jfpereira.pdf: 994358 bytes, checksum: 841297998e1d9cd11701f0d79cbb1e7c (MD5) / Made available in DSpace on 2015-03-23T13:15:27Z (GMT). No. of bitstreams: 1 2012_dis_jfpereira.pdf: 994358 bytes, checksum: 841297998e1d9cd11701f0d79cbb1e7c (MD5) Previous issue date: 2012 / Histoplasmosis is a systemic infectious disease caused by the dimorphic fungus Histoplasma capsulatum, which has three varieties: H. capsulatum var. capsulatum, H. capsulatum var. duboisii and H. capsulatum var. farciminosum. This fungus can occur in different clinical modalities and mimic other diseases, being primarily associated with frames of immunosuppression. Despite the existence of effective therapies for the treatment of this disease, there is a need to search for new therapeutic approaches for this mycosis. Studies have shown that after the introduction of highly active antiretroviral therapy, the mortality and morbidity due to a wide variety of opportunistic infections, including mycoses, have decreased among HIV-infected patients, which may show the impact of antiretroviral therapy in these opportunistic infections. Some studies have also shown that antiretroviral drugs in the class of protease inhibitors may affect the virulence of certain fungi. Thus, this study aimed to evaluate the in vitro inhibitory effect of protease inhibitor saquinavir (SQV), alone and in combination with antifungal agents: amphotericin B (AMB), fluconazole (FLU), itraconazole (ITC), voriconazole (VRC) and caspofungin (CAS) against the H. capsulatum. It was used 30 strains of H. capsulatum in the filamentous phase and 10 strains of H. capsulatum in the yeast phase to the test of the drugs isolated. To test the combination of SAQ with ITC, 30 strains in the filamentous phase and 20 strains in the yeast phase were used. The study was conducted by broth microdilution assay, described in document M27-A2, standardized by the Clinical Laboratory Standards Institute (CLSI). The interaction of drugs was analyzed by calculating the Fractional Inhibitory Concentration Index (FICI), defined as the sum of relations between the minimum inhibitory concentration (CIM) of each drug in combination and CIM of the same drug alone, whereas values less than or equal to 0.5 suggestive synergism between the drugs. Regarding the results, SQV inhibited the growth of all strains of H. capsulatum, with CIMs ranging from 0.122 to 0.977 μg / mL for both phases. In relation to combination of SQV with antifungal agents tested, only the combination of SQV with the ITR showed synergism, with values of FICI less than 0.5, where there was a significant reduction of the CIMs of both drugs. The results suggest that SQV can act as an adjuvant therapy in vitro, increasing the inhibitory effect on fungal growth, being able to lower the CIM values of antifungal drugs. Given the above, additional studies to identify the mechanism of action of SQV against H. capsulatum are needed, as well as its interaction with the ITR antifungal. / A histoplasmose é uma doença infecciosa sistêmica, causada pelo fungo dimórfico Histoplasma capsulatum o qual possui três variedades: H. capsulatum var. capsulatum, H. capsulatum var. duboisii e H. capsulatum var. farciminosum,, e pode se apresentar de diversas modalidades clínicas e mimetizar outras doenças, estando principalmente associada a quadros de imunossupressão. Apesar da existência de terapias eficazes para o tratamento da desta doença, existe a necessidade de uma busca por novas opções terapêuticas para essa micose. Estudos comprovaram que após a instituição da terapia antirretroviral altamente ativa a mortalidade e a morbidade em decorrência de uma ampla variedade de infecções oportunistas, inclusive fúngicas, têm diminuído entre os pacientes infectados pelo HIV. Alguns estudos também têm demonstrado que fármacos antirretrovirais da classe dos inibidores de proteases podem afetar a virulência de certos fungos. Assim, o presente estudo visou avaliar o efeito inibitório in vitro do inibidor de protease saquinavir (SQV) isolado e em combinação com os antifúngicos anfotericina B (AMB), fluconazol (FLU), itraconazol (ITC), voriconazol (VRC) e caspofungina (CAS) frente ao H. capsulatum. Foram utilizadas 30 cepas de H. capsulatum na fase filamentosa e 10 cepas de H. capsulatum na fase leveduriforme para o teste das drogas isoladas e 20 cepas de H. capsulatum na fase filamentosa e 10 na fase leveduriforme para o teste da combinação do SQV com o ITR. O estudo foi realizado por meio de ensaio de microdiluição em caldo, descrito no documento M27-A2, padronizado pelo Clinical Laboratory Standards Institute (CLSI). A interação das drogas foi analisada através do cálculo do Índice da Concentração Inibitória Fracionária (FICI), definido como a soma das relações entre a concentração inibitória mínima (CIM) de cada droga em combinação e a CIM da mesma droga isolada, considerando os valores menores ou iguais a 0,5 sugestivos de sinergismo entre as drogas. Com relação aos resultados, o SQV inibiu o crescimento de todas as cepas de H. capsulatum, com CIM variando de 0,122 a 0,977μg/mL para ambas as fases. E, em relação a combinação do SQV com os antifúngicos testados, somente a combinação do SQV com o ITR apresentou sinergismo, com valores de FICI menores que 0,5, onde observou-se uma redução significativa da CIM de ambas as drogas. Os resultados obtidos sugerem que o SQV pode agir como adjuvante terapêutico in vitro, incrementando o efeito inibitório sobre o crescimento fúngico, sendo capaz de diminuir os valores de CIM de drogas antifúngicas. Diante do exposto, estudos adicionais para identificar o mecanismo de ação do SQV frente ao H. capsulatum são necessários, bem como sua interação com o ITR.

No description available.

Saquinavir é um medicamento antirretroviral desenvolvido para inibir a protease do vírus da imunodeficiência humana. Este fármaco apresenta várias limitações em função dos seus efeitos colaterais, baixa biodisponibilidade e sensorial não agradável. A nanoencapsulação de fármacos é uma técnica promissora para superar esses problemas. O objetivo deste trabalho é desenvolver suspensões aquosas de nanocápsulas poliméricas contendo saquinavir utilizando uma mistura de poli-ε-caprolactona (PCL) e poli- ε-caprolactone triol (PCL-T) visando uma formulação líquida adequada para o tratamento de crianças infectadas. As nanocápsulas foram preparadas pela técnica de deposição interfacial de polímeros pré-formados. A análise de diâmetro médio de partícula foi realizada por diferentes técnicas, difração a laser, dispersão de luz dinâmica e análise de rastreamento de nanopartículas. O potencial zeta foi determinado por mobilidade eletroforética, o pH por potenciometria e o teor de fármaco e a taxa de encapsulação por cromatografia líquida de alta eficiência com detecção no ultravioleta. O ensaio de liberação in vitro foi realizado com membrana de diálise e coleta das amostras nos intervalos de 0-120 horas, a 37°C. A estabilidade foi verificada nos tempos 0, 15, 30, 60 e 90 dias, ao abrigo da luz, em temperatura ambiente e com o monitoramento do pH, potencial zeta, diâmetro, teor de fármaco e através da análise de retroespalhamento de luz. A morfologia foi observada por microscopia eletrônica de transmissão. A citotoxicidade em linfócitos T humano foi determinada pelo teste de exclusão de azul de tripan e o mascaramento do sabor de saquinavir pela língua eletrônica. Todas as formulações apresentaram tamanho nanométrico entre 172 ± 18 e 225 ± 4 nm e uma boa distribuição de tamanho. A taxa de encapsulação de saquinavir foi maior que 99%. As nanocápsulas apresentaram potencial zeta negativo, valores de pH levemente ácido e morfologia esférica. Os parâmetros avaliados na estabilidade mantiveram-se constantes ao longo do tempo analisado. O ensaio in vitro mostrou uma liberação controlada de saquinavir. As formulações não apresentaram toxicidade in vitro e foram capazes de mascarar o sabor do saquinavir. Portanto, a associação de polímeros é eficaz para a encapsulação de saquinavir, apresentando características nanotecnológicas adequadas para a sua administração. / Saquinavir is an antiretroviral drug designed to inhibit the human immunodeficiency virus protease. This drug presents several limitations regarding side effects, very low bioavailability and sensory not pleasant. The drug nanoencapsulation is a promising technique to overcome these issues. The aim of this work is to develop aqueous suspensions of polymeric nanocapsules containing saquinavir using poly-ε-caprolactone (PCL) and poly-ε-triol caprolactone (PCL-T) towards a liquid formulation suitable for the treatment of infected children. The nanocapsules were prepared using the technique of interfacial deposition of preformed polymers. The characterization study was conducted by examining the average particle diameter by three techniques, the laser diffraction, the dynamic light scattering and the nanoparticle tracking analysis. The zeta potential was evaluated by electrophoretic mobility, the pH by potentiometry and drug content and encapsulation rate by high performance liquid chromatography with UV detection. The in vitro release assay was carried out in a dialysis membrane with analysis at 0-120 hours, at 37 °C. The stability was checked at 0, 15, 30, 60 and 90 days, protected from light, at room temperature and by monitoring the pH, zeta potential, diameter, drug content and by multiple light scattering analysis. The morphology was observed by transmission electron microscopy. The cytotoxicity in human T lymphocytes was determined by exclusion of trypan blue test and the taste masking by the electronic tongue. All formulations presented nanometric size between 172 ± 18 and 225 ± 4 nm and a good size distribution. The drug encapsulation rate was higher than 99%. The nanocapsules presented negative zeta potential, pH values slightly acidic and spherical morphologies. The parameters evaluated in stability remained constant over time analyzed. The in vitro assay showed a controlled drug release from nanocapsules formulations. The developed formulations showed no toxicity and were able to mask the taste of saquinavir. The association of polymers is effective for saquinavir nanoencapsulation presenting suitable nanotechnological features for drug administration.

Saquinavir é um medicamento antirretroviral desenvolvido para inibir a protease do vírus da imunodeficiência humana. Este fármaco apresenta várias limitações em função dos seus efeitos colaterais, baixa biodisponibilidade e sensorial não agradável. A nanoencapsulação de fármacos é uma técnica promissora para superar esses problemas. O objetivo deste trabalho é desenvolver suspensões aquosas de nanocápsulas poliméricas contendo saquinavir utilizando uma mistura de poli-ε-caprolactona (PCL) e poli- ε-caprolactone triol (PCL-T) visando uma formulação líquida adequada para o tratamento de crianças infectadas. As nanocápsulas foram preparadas pela técnica de deposição interfacial de polímeros pré-formados. A análise de diâmetro médio de partícula foi realizada por diferentes técnicas, difração a laser, dispersão de luz dinâmica e análise de rastreamento de nanopartículas. O potencial zeta foi determinado por mobilidade eletroforética, o pH por potenciometria e o teor de fármaco e a taxa de encapsulação por cromatografia líquida de alta eficiência com detecção no ultravioleta. O ensaio de liberação in vitro foi realizado com membrana de diálise e coleta das amostras nos intervalos de 0-120 horas, a 37°C. A estabilidade foi verificada nos tempos 0, 15, 30, 60 e 90 dias, ao abrigo da luz, em temperatura ambiente e com o monitoramento do pH, potencial zeta, diâmetro, teor de fármaco e através da análise de retroespalhamento de luz. A morfologia foi observada por microscopia eletrônica de transmissão. A citotoxicidade em linfócitos T humano foi determinada pelo teste de exclusão de azul de tripan e o mascaramento do sabor de saquinavir pela língua eletrônica. Todas as formulações apresentaram tamanho nanométrico entre 172 ± 18 e 225 ± 4 nm e uma boa distribuição de tamanho. A taxa de encapsulação de saquinavir foi maior que 99%. As nanocápsulas apresentaram potencial zeta negativo, valores de pH levemente ácido e morfologia esférica. Os parâmetros avaliados na estabilidade mantiveram-se constantes ao longo do tempo analisado. O ensaio in vitro mostrou uma liberação controlada de saquinavir. As formulações não apresentaram toxicidade in vitro e foram capazes de mascarar o sabor do saquinavir. Portanto, a associação de polímeros é eficaz para a encapsulação de saquinavir, apresentando características nanotecnológicas adequadas para a sua administração. / Saquinavir is an antiretroviral drug designed to inhibit the human immunodeficiency virus protease. This drug presents several limitations regarding side effects, very low bioavailability and sensory not pleasant. The drug nanoencapsulation is a promising technique to overcome these issues. The aim of this work is to develop aqueous suspensions of polymeric nanocapsules containing saquinavir using poly-ε-caprolactone (PCL) and poly-ε-triol caprolactone (PCL-T) towards a liquid formulation suitable for the treatment of infected children. The nanocapsules were prepared using the technique of interfacial deposition of preformed polymers. The characterization study was conducted by examining the average particle diameter by three techniques, the laser diffraction, the dynamic light scattering and the nanoparticle tracking analysis. The zeta potential was evaluated by electrophoretic mobility, the pH by potentiometry and drug content and encapsulation rate by high performance liquid chromatography with UV detection. The in vitro release assay was carried out in a dialysis membrane with analysis at 0-120 hours, at 37 °C. The stability was checked at 0, 15, 30, 60 and 90 days, protected from light, at room temperature and by monitoring the pH, zeta potential, diameter, drug content and by multiple light scattering analysis. The morphology was observed by transmission electron microscopy. The cytotoxicity in human T lymphocytes was determined by exclusion of trypan blue test and the taste masking by the electronic tongue. All formulations presented nanometric size between 172 ± 18 and 225 ± 4 nm and a good size distribution. The drug encapsulation rate was higher than 99%. The nanocapsules presented negative zeta potential, pH values slightly acidic and spherical morphologies. The parameters evaluated in stability remained constant over time analyzed. The in vitro assay showed a controlled drug release from nanocapsules formulations. The developed formulations showed no toxicity and were able to mask the taste of saquinavir. The association of polymers is effective for saquinavir nanoencapsulation presenting suitable nanotechnological features for drug administration.

Saquinavir é um medicamento antirretroviral desenvolvido para inibir a protease do vírus da imunodeficiência humana. Este fármaco apresenta várias limitações em função dos seus efeitos colaterais, baixa biodisponibilidade e sensorial não agradável. A nanoencapsulação de fármacos é uma técnica promissora para superar esses problemas. O objetivo deste trabalho é desenvolver suspensões aquosas de nanocápsulas poliméricas contendo saquinavir utilizando uma mistura de poli-ε-caprolactona (PCL) e poli- ε-caprolactone triol (PCL-T) visando uma formulação líquida adequada para o tratamento de crianças infectadas. As nanocápsulas foram preparadas pela técnica de deposição interfacial de polímeros pré-formados. A análise de diâmetro médio de partícula foi realizada por diferentes técnicas, difração a laser, dispersão de luz dinâmica e análise de rastreamento de nanopartículas. O potencial zeta foi determinado por mobilidade eletroforética, o pH por potenciometria e o teor de fármaco e a taxa de encapsulação por cromatografia líquida de alta eficiência com detecção no ultravioleta. O ensaio de liberação in vitro foi realizado com membrana de diálise e coleta das amostras nos intervalos de 0-120 horas, a 37°C. A estabilidade foi verificada nos tempos 0, 15, 30, 60 e 90 dias, ao abrigo da luz, em temperatura ambiente e com o monitoramento do pH, potencial zeta, diâmetro, teor de fármaco e através da análise de retroespalhamento de luz. A morfologia foi observada por microscopia eletrônica de transmissão. A citotoxicidade em linfócitos T humano foi determinada pelo teste de exclusão de azul de tripan e o mascaramento do sabor de saquinavir pela língua eletrônica. Todas as formulações apresentaram tamanho nanométrico entre 172 ± 18 e 225 ± 4 nm e uma boa distribuição de tamanho. A taxa de encapsulação de saquinavir foi maior que 99%. As nanocápsulas apresentaram potencial zeta negativo, valores de pH levemente ácido e morfologia esférica. Os parâmetros avaliados na estabilidade mantiveram-se constantes ao longo do tempo analisado. O ensaio in vitro mostrou uma liberação controlada de saquinavir. As formulações não apresentaram toxicidade in vitro e foram capazes de mascarar o sabor do saquinavir. Portanto, a associação de polímeros é eficaz para a encapsulação de saquinavir, apresentando características nanotecnológicas adequadas para a sua administração. / Saquinavir is an antiretroviral drug designed to inhibit the human immunodeficiency virus protease. This drug presents several limitations regarding side effects, very low bioavailability and sensory not pleasant. The drug nanoencapsulation is a promising technique to overcome these issues. The aim of this work is to develop aqueous suspensions of polymeric nanocapsules containing saquinavir using poly-ε-caprolactone (PCL) and poly-ε-triol caprolactone (PCL-T) towards a liquid formulation suitable for the treatment of infected children. The nanocapsules were prepared using the technique of interfacial deposition of preformed polymers. The characterization study was conducted by examining the average particle diameter by three techniques, the laser diffraction, the dynamic light scattering and the nanoparticle tracking analysis. The zeta potential was evaluated by electrophoretic mobility, the pH by potentiometry and drug content and encapsulation rate by high performance liquid chromatography with UV detection. The in vitro release assay was carried out in a dialysis membrane with analysis at 0-120 hours, at 37 °C. The stability was checked at 0, 15, 30, 60 and 90 days, protected from light, at room temperature and by monitoring the pH, zeta potential, diameter, drug content and by multiple light scattering analysis. The morphology was observed by transmission electron microscopy. The cytotoxicity in human T lymphocytes was determined by exclusion of trypan blue test and the taste masking by the electronic tongue. All formulations presented nanometric size between 172 ± 18 and 225 ± 4 nm and a good size distribution. The drug encapsulation rate was higher than 99%. The nanocapsules presented negative zeta potential, pH values slightly acidic and spherical morphologies. The parameters evaluated in stability remained constant over time analyzed. The in vitro assay showed a controlled drug release from nanocapsules formulations. The developed formulations showed no toxicity and were able to mask the taste of saquinavir. The association of polymers is effective for saquinavir nanoencapsulation presenting suitable nanotechnological features for drug administration.

The employment of HIV-1 protease (PR) inhibitors (PIs) in antiviral therapy has been successful in reducing mortality of HIV/AIDS patients. However, the long-term efficacy of PIs is challenged by the rapid emergence of drug-resistant mutants of PR. To understand the underlying mechanism of drug resistance, structures and activities of HIV-1 PR and its drug resistant mutants have been extensively studied. Here, PR mutants PRR8Q, PRD30N, PRI47V, PRI50V, PRI54M, PRV82A, and PRN88D/S bearing single substitutions have been investigated by crystallography and kinetics. GRL-0519 is a potent new antiviral inhibitor of HIV-1 PR that possesses tris-tetrahydrofuran (tris-THF) as the P2 ligand. The crystal structures of GRL-0519 were determined at resolutions of 1.06-1.49 Å in complex with the mutants PRR8Q, PRD30N, PRI50V, PRI54M, and PRV82A. I50V lost its interaction with inhibitor while V82Aand I54M compensated for the mutation through the main chain shift and flexibility of 80’s loop (residues 78-82), respectively. The structural changes may account for the worst inhibition of GRL-0519 for PRI50V (60-fold decrease relative to wild-type enzyme)and moderate inhibition for PRI54M and PRV82A (6-7-fold decrease). The large tris-THF group at P2 provides a good fit in the S2 subsite and may be effective against resistant virus with mutations of residues in this subsite. SQV and DRV are two clinical inhibitors that were designed to target the wild type PR and its drug resistant mutants, respectively. The crystal structures of PR mutants PRI47V, PRN88D/s in complex with DRV and mutants PRI47V and PRN88D in complex with SQV with resolutions of 1.13-1.72 Å were also analyzed. Mutation I47V gained more hydrophobic interactions with DRV and SQV. Interestingly, the structural changes did not affect the inhibition of both inhibitors for PRI47V (relative Ki is 0.7 and 1 for DRV and SQV, respectively). DRV and SQV showed 8-fold increase in Ki for PRN88D and only very subtle local changes have been observed on the structures. DRV induced 0.3 fold reduction in Ki for PRN88S and the distal structural changes have been transferred to the active site. This study provided fundamental information for understanding drug resistance and future design of potential antiviral drugs.

Sucrose acetate isobutyrate (SAIB), a randomly substituted sucrose approximating sucrose diacetate hexaisobutyrate, is produced by Eastman Chemical Company for a variety of applications. SAIB is widely used in the food industry as a weighting agent to disperse flavoring oils in primarily citrus-based soft drink beverages. Additionally, SAIB is currently being marketed by another company as a parenteral drug delivery system. The studies reported here focused on investigating SAIB as an excipient, or delivery vehicle, for use in oral delivery of several drugs, including ibuprofen, saquinavir, and clarithromycin. Dissolution experiments were conducted using both ibuprofen and caffeine, and results suggest that SAIB can be used in dosage forms to control release rate. Pharmacokinetic studies in which laboratory rats were dosed with formulations containing drugs such as ibuprofen, saquinavir, and clarithromycin suggest that SAIB may act to reduce animal-to-animal variability in drug concentration profiles in some cases, and that it may also enhance gastroretention of the dosage forms. Finally, dosage form imaging studies suggest but do not reliably confirm that SAIB may aid in promoting gastric retention, which would make its use in dosage form formulation beneficial for administration of drugs whose action is intended to occur in the stomach.

O presente trabalho é composto por 4 capítulos distintos. No primeiro intitulado \"Ciclodextrinas: tecnologia para melhoria da solubilidade de fármacos pouco solúveis\" aborda-se uma revisão das ciclodextrinas e de seus derivados, como também a formação dos complexos de inclusão. No segundo capítulo, \"Obtenção e caracterização de complexos ternários de saquinavir, β-ciclodextrina e polivinilpirrolidona\", foram avaliadas condições que poderiam influenciar na obtenção dos complexos, tais como: tempo de agitação na formação dos complexos, proporção equimolar de fármaco e ciclodextrina, solvente para solubilização do fármaco e porcentagem de polímero hidrossolúvel, concluindo-se que as melhores condições para obtenção dos complexos são 48 horas de agitação, razão equimolar 1:2 e 1:4, água para solubilização do fármaco e acréscimo de1% de polímero ao sistema. O terceiro capítulo, \"Influência do método de secagem na obtenção de complexos ternários de saquinavir\", teve o objetivo de obter e caracterizar complexos ternários de saquinavir, β-ciclodextrina e polivinilpirrolidona pelos métodos de secagem em estufa e liofilização, bem como, comparar a influência dos métodos de obtenção na solubilidade do fármaco, chegando-se ao aumento de 44 vezes da solubilidade do fármaco quando na forma de complexo SAQ:βCD:PVP de proporção equimolar 1:2 obtido por liofilização. O quarto, \"Perfil de dissolução de cápsulas e comprimidos contendo complexos ternários de saquinavir, ciclodextrina e polivinilpirrolidona\", teve o objetivo de comparar o perfil de dissolução de cápsulas de gelatina dura e comprimidos contendo sistemas ternários SAQ:βCD:PVP, obtidos pelos métodos de secagem em estufa e liofilização e o produto referência (Fortovase®). Os resultados indicam que os perfis de dissolução das formas farmacêuticas contendo complexos liofilizados apresentaram melhores resultados de eficiência de dissolução e que as cápsulas liberam mais prontamente o ativo que os comprimidos. / The present study has 4 captions. The first is called \"Cyclodextrins: Technology to improve the solubility of the little soluble drugs\" show a review of the cyclodextrins and its derivates, as well the formations of inclusion complexes. At the second caption, \"Preparation and characterization of the ternary complexes of the saquinavir, &#946-cyclodextrin and polyvinylpyrrolidone\", were evaluated some conditions that could influence in an obtention of the complexes like stirring time at the formation of the complexes, equimolar ratio of the drug and cyclodextrin, solvent for the solubility of the drug and percentage of the hydrosoluble polymer concluding that the best conditions for the formation of the complexes are: 48 hours of the stirring, equimolar ratio 1:2 and 1:4, water to solubilize the drug and addition of 1% of the polymer in the system. The third caption, \"Influence of the drying method at the formation of the ternary complexes of the saquinavir\", had the aim to prepare and characterize ternary complexes of the saquinavir, &#946-cyclodextrin and polyvinylpyrrolidone by the method of drying in equipment and lyophilization, as well to compare the influence of the methods of the obtention at the solubility of the drug, reaching the increasing of 44 times the solubility of the drug when at the form of the ternary inclusion complex (SAQ:βCD:PVP) at the equimolar ratio 1:2 obtained by lyophilization. The fourth and the last caption, \"Dissolution rate of the capsules and tablets of ternary complex of the saquinavir, cyclodextrin and polyvinylpyrrolidone, had the aim to compare the dissolution profile of capsules of the hard gelatine and tablets of ternary complexes SAQ:βCD:PVP obtained by the method of drying in equipment and lyophilization and the reference product (Fortovase®). The results indicate that the dissolution profiles of the pharmaceutical forms of lyophilized complexes show better results of the efficience of the dissolution and that the capsules liberate more efficient the active than the tablets.

Referat: Background: Human African Trypanosomiasis (HAT) also called sleeping sickness is an infectious disease of humans caused by an extracellular protozoan parasite. The disease, if left untreated, results in 100% mortality. However, the available drugs are full of severe drawbacks and fail to escape the fast development of trypanosoma resistance. Due to the probable similarities in cell metabolism among tumor and trypanosoma cells, some of the current registered drugs against HAT were derived from cancer chemotherapeutic research. Here too, for the first time, we have demonstrated that the simple ester, ethyl pyruvate, comprises such properties. On the other hand initial studies have confirmed the efficacy of protease inhibitors in treatment of Trypanosoma cruzi, Plasmodium falciparum and Leishmania major. However, studies on efficacy and specific proteases inhibition using HIV-1 protease inhibitors on T. brucei cells remain untouched. Methodology/Principal findings: The current study covers efficacy and corresponding target evaluation of ethyl pyruvate and HIV-1 protease inhibitors (ritonavir and saquinavir) on T. brucei cell lines using a combination of biochemical techniques including cell proliferation assays, enzyme kinetics, zymography, phase contrast microscopic video imaging and ex vivo drug toxicity tests. We have shown that ethyl pyruvate effectively kills trypanosomes most probably by net ATP depletion through inhibition of pyruvate kinase (Ki=3.0±0.29 mM). The potential of this compound as an anti-trypanosomal drug is also strengthened by its fast acting property, killing cells within three hours post exposure. This was demonstrated using video imaging of live cells as well as concentration and time dependency experiments. Most importantly, this drug produced minimal side effects in human erythrocytes and is known to easily cross the blood-brain-barrier (BBB) which makes it a promising candidate for effective treatment of the two clinical stages of sleeping sickness. Trypanosome drug resistance tests indicate irreversible killing of cells and a low chance of drug resistance development under applied experimental conditions. In addition to ethyl pyruvate our experimental study on HIV-1 protease inhibitors showed that both ritonavir (RTV) (IC50=12.23 µM) and saquinavir (SQV) (IC50=11.49 µM) effectively inhibited T. brucei cells proliferation. The major proteases identified in these cells were the cysteine- (~29kDa Mr) and metallo- (~66kDa Mr) proteases. Their proteolytic activity was, however, not hampered by either of these two protease inhibitors. Conclusion/Significance: Our results present ethyl pyruvate as a safe and fast acting drug. Hence, because of its predefined property to easily cross the BBB, it can probably be a new candidate agent to treat the heamolymphatic as well as neurological stages of sleeping sickness. Similarly, HIV-1 protease inhibitors, SQV and RTV, exhibited their antitrypanosomal potential but require further anlysis to identify their specific targets.