Para fármacos administrados por via oral, o controle da extensão e da velocidade de absorção depende basicamente de duas importantes etapas: solubilidade do fármaco nos líquidos fisiológicos e sua permeabilidade através das membranas biológicas. Assim, o Sistema de Classificação Biofarmacêutica (SCB) foi proposto como uma ferramenta para o desenvolvimento de novos fármacos, de novas formulações e para auxiliar nos processos de bioisenção. No entanto, outro fator relacionado à biodisponibilidade e que deve ser considerado nos estudos biofarmacêuticos é o metabolismo. Desta forma, o Sistema de Classificação Biofarmacêutica de Distribuição de Fármacos (SCBDF) foi proposto com a finalidade de classificar os fármacos de acordo com suas características de solubilidade e de metabolismo de modo que seja possível avaliar e predizer o comportamento do fármaco in vivo. O metabolismo tem sido amplamente investigado, sobretudo as enzimas do citocromo P450, as quais estão presentes também nos enterócitos. Além disso, o SCBDF oferece um suporte quanto à avaliação dos mecanismos de permeabilidade envolvidos nos processos de absorção, interações fármaco-fármaco e interações fármaco-alimento. Assim, o presente trabalho teve como objetivo elucidar os mecanismos envolvidos na permeabilidade de fármacos antirretrovirais por meio dos modelos ex vivo (câmaras de difusão vertical tipo Franz) e in vitro (PAMPA, MDCK-MDR1 e microssomas) considerando os aspectos relacionados ao metabolismo intestinal e ao efluxo destes fármacos. Dada a importância da utilização de fármacos antirretrovirais na terapia medicamentosa contra a Síndrome da Imunodeficiência Adquirida (SIDA) e que estes medicamentos são normalmente administrados cronicamente, a compreensão dos mecanismos envolvidos na permeabilidade é de suma importância, uma vez que estes não estão totalmente esclarecidos e poucas informações são encontradas na literatura. Além disso, a biodisponibilidade de fármacos como estavudina, lamivudina e zidovudina indica variação na permeabilidade, necessitando de uma investigação científica mais aprofundada dos processos absortivos. Assim, segmentos de jejuno provenientes de ratos machos Wistar foram utilizados para a avaliação da permeabilidade intestinal dos referidos antirretrovirais considerando a avaliação de efluxo pela glicoproteína-P e o metabolismo intestinal pela CYP3A. De maneira complementar, estudos in vitro com o emprego de membranas artificiais paralelas (PAMPA) e culturas celulares de MDCK-MDR1 foram realizados com a finalidade de auxiliar na elucidação dos mecanismos de permeabilidade dos fármacos antirretrovirais. Além disso, a avaliação do metabolismo dos referidos fármacos foi realizada com o emprego de microssomas a fim de verificar se tais substâncias são substratos de enzimas da família CYP3A e, assim, verificar o impacto do metabolismo intestinal na absorção. Os resultados de permeabilidade obtidos em PAMPA foram: 0,74±0,11 x 10-6 cm/s para a estavudina, 0,25±0,12 x 10-6 cm/s para a lamivudina e 1,14±0,25 x 10-6 cm/s para a zidovudina. Já no modelo ex vivo com o emprego de câmaras de difusão vertical tipo Franz, os resultados foram: 1,56±0,32 x 10-5 cm/s para a estavudina, 1,26±0,27 x 10-5 cm/s para a lamivudina e 2,54±0,49 x 10-5 cm/s para a zidovudina. Portanto, com base nos resultados obtidos a partir dos dois métodos empregados, sugere-se que 30 outro mecanismo de transporte que não envolva a permeabilidade por difusão transcelular passiva possa estar relacionado à permeabilidade dos fármacos antirretrovirais. Com relação aos estudos de efluxo, os resultados obtidos a partir dos experimentos realizados em câmaras de difusão vertical tipo Franz demonstraram o aumento significativo da permeabilidade dos três antirretrovirais quando o inibidor de P-gp foi empregado, sendo: de 15,6 x 10-6 para 42,5 x 10-6 cm/s para a estavudina, de 12,6 x 10-6 para 37,5 x 10-6 cm/s para a lamivudina e de 25,4 x 10-6 para 56,6 x 10-6 cm/s para a zidovudina. Em culturas celulares MDCK-MDR1, os resultados de permeabilidade foram utilizados para a obtenção das razões entre as direções B→A e A→B. Os valores de Papp na condição inibida para os fármacos estudados apresentaram razão menor do que 1. Já a razão B→A/A→B para cada fármaco nos ensaios sem inibidor apresentou-se igual ou maior que 2, evidenciando a interação fármaco-transportador. Com base nisso, o modelo ex vivo com o emprego de segmentos intestinais em câmaras de difusão vertical tipo Franz apresentou-se adequado na avaliação do mecanismo de efluxo dos fármacos antirretrovirais, o que foi confirmado com os estudos realizados em MDCK-MDR1. Assim, os fármacos antirretrovirais estudados apresentaram interação significativa com a P-gp. Em relação aos estudos de metabolismo realizados em câmaras de difusão vertical tipo Franz, os resultados demonstraram grande variação na permeabilidade dos três antirretrovirais quando o inibidor de CYP3A foi empregado, sendo: de 15,6 x 10-6 para 23,5 x 10-6 cm/s para a estavudina, de 12,6 x 10-6 para 27,3 x 10-6 cm/s para a lamivudina e de 25,4 x 10-6 para 40,5 x 10-6 cm/s para a zidovudina. Já no modelo que emprega microssomas, os resultados de metabolização na ausência e na presença de inibidor de CYP3A foram: de 16,56% para 19,79% para a estavudina, de 14,56% para 15,55% para a lamivudina e de 17,85% para 16,48% para a zidovudina. Com base nisso, sugerese o emprego de microssomas para a determinação de metabolismo, uma vez que o método ex vivo empregado demonstrou grande variação entre os valores obtidos. Desta forma, observou-se que, para cada fármaco, não houve influência significativa no metabolismo pré-sistêmico relacionado às enzimas do complexo CYP3A, o que indica que a absorção oral das referidas substâncias não é limitada por tais enzimas. Portanto, a utilização dos diferentes métodos empregados no desenvolvimento do presente trabalho permitiu compreender os mecanismos envolvidos no transporte dos fármacos antirretrovirais, o que se torna de grande relevância nas etapas de desenvolvimento farmacêutico de novas moléculas e na compreensão de eventos clínicos ainda não esclarecidos atualmente. / For orally administered drugs, control of the extent and rate of absorption depends on two important steps: solubility of the drug in physiological liquids and their permeability across biological membranes. Thus, the Biopharmaceutics Classification System (BCS) has been proposed as a tool for the development of new drugs, new formulations and aid in the biowaiver processes. However, another factor related to bioavailability that should be considered in biopharmaceutic studies is the metabolism. Thus, the Biopharmaceutics Drug Disposition Classification System (BDDCS) has been proposed for drug classification according to their solubility and metabolism characteristics, so it is possible to evaluate and predict the in vivo behavior of a compound. Metabolism has been extensively investigated, especially cytochrome P450 enzymes, which are also expressed in enterocytes. Besides, BDDCS provides support in evaluating the permeability mechanisms involved in the absorption processes, drug-drug interactions and drug-food interactions. Thus, the present study aimed to evaluate the mechanisms of permeability of antiretroviral drugs through the ex vivo (Franz cells) and in vitro (PAMPA, MDCK-MDR1 and microsomes) models considering aspects related to the intestinal metabolism and efflux of these drugs. Given the importance of the use of antiretroviral drugs in drug therapy against Acquired Immune Deficiency Syndrome (AIDS) and that these drugs are usually administered in a long-term way, understanding the mechanisms involved in the permeability is of a great importance, since they are not totally elucidated and no information is found in the literature. In addition, drugs as stavudine, lamivudine and zidovudine indicate variation in the permeability, which require further scientific investigation of absorptive processes. Thus, jejunum segments from rats were used to evaluate the intestinal permeability of these antiretroviral drugs, considering the evaluation of efflux by P-glycoprotein and intestinal metabolism by CYP3A. In a complementary manner, in vitro studies using parallel artificial membranes (PAMPA) and cell cultures MDCK-MDR1 were performed to aid in the elucidation of the permeability mechanisms of antiretroviral drugs. Also, the evaluation of the metabolism was carried out using microsomes to verify if such substances are substrates of CYP3A, and verify the impact of the intestinal metabolism in the absorption. The permeability results obtained in PAMPA were: 0.74±0.11x10-6 cm/s for stavudine, 0.25±0.12x10-6 cm/s for lamivudine and 1.14±0.25x10-6 cm/s for zidovudine. In ex vivo method using the intestinal segments in Franz cells, the results were: 1.56±0.32x10-5 cm/s for stavudine, 1.26±0.27x10-5 cm/s for lamivudine and 2.54±0.49x10-5 cm/s for zidovudine. Thus, based on the results obtained from these two methods, it is suggested that the antiretroviral drugs present other transport mechanism that is different from transcellular passive diffusion. For efflux studies, results obtained from experiments performed in Franz cells shown the increase of the permeability of the three antiretroviral drugs when the P-gp inhibitor was used: from 15.6x10-6 to 42,5x10-6 cm/s for stavudine, from 12.6x10-6 cm/s to 37.5x10-6 cm/s for lamivudine, and 25.4x10-6 to 56.6x10-6 cm/s for zidovudine. In MDCK-MDR1, the permeability results were used for obtaining ratio values between the directions B→A and A→B. The Papp values obtained with 33 inhibitor shown a ratio less than 1. For ratio B→A/A→B for each drug in experiments without inhibitor, the values obtained was equal or greater than 2, which shows the interaction between drug and transporter. Based on that, the ex vivo model using intestinal segments in Franz cells seems to be adequate for evaluation of efflux mechanism of antiretroviral drugs, which was confirmed by MDCK-MDR1 studies. Thus, the antiretroviral drugs presented interaction with P-gp. For metabolism studies in intestinal segments in Franz cells, a wide range of standard deviation was observed for the three antiretroviral drugs when the CYP3A inhibitor was used: from 15.6x10-6 cm/s to 23.5x10-6 cm/s for stavudine, from 12.6x10-6 cm/s to 27.3x10-6 cm/s for lamivudine, and from 25.4x10-6 cm/s to 40.5x10-6 cm/s for zidovudine. In experiments in microsomes, the results of metabolization in the absence and presence of CYP3A inhibitor were: from 16.56 to 19.79% for stavudine, from 14.56 to 15.55% for lamivudine and from 17.85 to 16.48% for zidovudine. Based on that, it is suggested the use of microsomes for metabolism evaluation, since the ex vivo method presented high variability between the results obtained. For each drug, no significative influence in pre-systemic metabolism related to CYP3A enzymes was observed, which indicates that the oral absorption of the drugs is not limited by these enzymes. The use of different methods in this work allowed to understand the mechanisms involved in the transport of antiretroviral drugs, which is of a great relevance in drug development and in the understanding of clinical events currently not clarified.

A via de administração oral é a forma favorita de administração de fármacos em função das vantagens que apresenta, dentre elas destacam-se: a adesão do paciente, conveniência e praticidade. Em função disto, a maioria dos medicamentos comercializados encontra-se disponível na forma farmacêutica de administração oral, entretanto, o sucesso de um tratamento medicamentoso por esta via requer que a absorção gastrointestinal do fármaco seja suficiente para assegurar a sua disponibilidade no local de ação (VOLPE, 2010). No entanto, a absorção do fármaco no trato gastrointestinal é complexa e pode ser influenciada por vários fatores, os quais têm impacto sobre a dissolução, solubilidade e permeabilidade do fármaco. Com o intuito de aumentar a biodisponibilidade de fármacos, que possuem absorção dificultada pela via oral, a via de administração pela mucosa bucal vem sendo uma alternativa na atualidade farmacêutica. Esta mucosa é um tecido não queratinizado, altamente vascularizado e apresenta poucas enzimas metabolizadoras. Tais características possibilitam boa absorção de fármacos sem que ocorra a metabolização pré-sistêmica, ou efeito de primeira passagem, somando-se ao fato desta apresentar fácil acessibilidade para a administração de fármacos (VRIES, M. E et al., 1991; NIELSEN, H. M &RASSING, M. R, 1999). Nesse sentido, o presente trabalho teve como objetivo avaliar a permeabilidade dos fármacos antirretrovirais (lamivudina e estavudina) por meio de modelo ex vivo em segmentos da mucosa bucal de suínos, com emprego de câmaras de difusão do tipo células de Franz. Para avaliação da permeabilidade bucal dos fármacos antirretrovirais, lamivudina e estavudina, e dos marcadores para transporte transcelular (metoprolol) e paracelular (fluoresceína sódica), empregou-se método ex-vivo, em células de Franz, com segmento de mucosa bucal de suíno ( a 37ºC, meio Ringer- Krebs- HEPES, pH 7,4), e Franz posterior análise das concentrações das substâncias permeadas (fármacos e marcadores) por cromatografia líquida de alta eficiência. Os resultados obtidos, por meio do protocolo desenvolvido, demonstram que o transporte através da via paracelular (marcador fluoresceína) foi mais expressivo que o transporte transcelular (marcador metoprolol), o que provavelmente se deve ao fato dos espaços intercelulares da mucosa bucal serem mais frouxos do que aqueles observados na mucosa intestinal (junções íntimas). Quanto à lamivudina e estavudina, os resultados de permeabilidade indicaram que estes fármacos permearam por mecanismo semelhante ao do metoprolol, isto é, por via transcelular. / The oral route of administration is the preferred mode of administration of drugs according to the advantages which features, among which stand out: patient compliance, convenience and practicality. Because of this, most of the marketed drug is available in pharmaceutical form for oral administration, however, the success of a drug therapy in this way requires that the drug intestinal absorption is sufficient to ensure their availability at the site of action (VOLPE, 2010). However, the drug absorption in the gastrointestinal tract is complex and may be influenced by various factors which have an impact on the dissolution, solubility and permeability of the drug. In order to increase the bioavailability of drugs, which have impeded absorption by oral route of administration from oral mucosa has been an alternative the pharmaceutical today. This mucosa is not keratinized tissue, highly vascular and shows metabolizing enzymes. These characteristics allow good absorption of drugs occurs without presystemic metabolism or first-pass effect, adding to the fact that this display easy accessibility for the administration of drugs (Vries, M. E et al, 1991;. NIELSEN, H. M & RASSING, M. R, 1999). In this sense, this study aimed to evaluate the permeability of antiretroviral drugs (lamivudine and stavudine) through ex vivo model segments of the oral mucosa of pigs with use of diffusion chambers type Franz cells. To evaluate the oral permeability of antiretroviral drugs, lamivudine and stavudine, and markers for transcellular transport (metoprolol) and paracellular (sodium fluorescein), used ex-vivo method, Franz cells, with buccal mucosa segment of swine ( at 37, through Krebs-Ringer- HEPES, pH 7.4) and Franz subsequent analysis of concentrations of the permeated substances (drugs and markers) by high-performance liquid chromatography. The results obtained using the protocol developed demonstrate that transport through the paracellular pathway (marker fluorescein) was higher than that transcellular transport (marker metoprolol), which probably is due to the intercellular spaces of the oral mucosa are looser than those observed in the intestinal mucosa (tight junctions). The lamivudine and stavudine, the permeability results indicate that these drugs permeated by a mechanism similar to that of metoprolol, by transcellular pathway.

Introdução: Pessoas infectadas pelo vírus da imunodeficiência humana (HIV) têm alto risco de desenvolver doença pneumocócica. Infecções invasivas pelo Streptococcus pneumoniae (pneumococo) lideram a morbidade e a mortalidade entre crianças, idosos e pessoas com doenças de base que comprometem o sistema imune ou diminuem a função esplênica. A infecção pelo HIV está associada a um aumento de até dez vezes na incidência das pneumonias bacterianas no mundo. S. pneumoniae é o agente mais comum identificado. A imunização contra S. pneumoniae tem sido recomendada para indivíduos infectados pelo HIV em vários países, incluindo o Brasil, onde a vacina de polissacarídeos antipneumococo com 23 sorotipos (PPV-23) está sendo utilizada desde 1993. A efetividade da vacina de polissacarídeos antipneumococo varia entre 60 a 80% entre adultos com função imunológica relativamente normal. Entre adultos infectados pelo HIV, os estudos realizados até o momento apresentam resultados inconclusivos ou mesmo contraditórios. Objetivo: o objetivo deste estudo é avaliar a efetividade da vacina de polissacarídeos antipneumococo 23-valente (PPV-23) em uma amostra da população adulta infectada pelo HIV, em São Paulo. Métodos: Estudo de caso controle, tipo caso-incidente, entre adultos infectados pelo HIV, em São Paulo. A exposição é ter sido vacinado com a vacina 23 valente e o desfecho é infecção invasiva causada pelo S. pneumoniae. Caso: pessoa infectada pelo HIV, >=18 anos de idade, com doença invasiva por pneumococo, diagnosticada por meio de cultura positiva (isolamento de S. pneumoniae) em material biológico obtido de qualquer fluido corporal normalmente estéril. Controle: pessoa infectada pelo HIV, com >=18 anos de idade, recebendo cuidados nas mesmas instituições, sem doença invasiva por pneumococo, com o mesmo nível de células T CD4+ no mesmo período do diagnóstico do caso, estratificadas por faixas (CD4<200; CD4>=200<=499; >=500) cels/mm3. Foram conduzidas análise univariada e regressão logística condicional. Resultados: 79 casos e 241 controles foram incluídos, média de 39 anos, 63% do sexo masculino, mais de 50% com escolaridade entre média e fundamental, 63,5% brancos, 19% vivendo em condições precárias de habitação. Idade, sexo e raça não diferiram entre casos e controles. Bacteremia foi a manifestação clínica mais freqüente. Fatores de risco associados à doença pneumocócica incluíram: uso de alcool, de drogas injetáveis, ter menor nível de escolaridade, condições precárias de habitação, contato próximo com criança <10 anos e hospitalizações prévias por pneumonia. O uso de anti-retrovirais e a vacina antipneumocócica estiveram associados com uma diminuição do risco. A efetividade da vacina foi de aproximadamente 65% (OR=0,35 IC95%:0,18-0,70). Após ajustar por alguns fatores associados à doença invasiva (uso de drogas injetáveis, hospitalização prévia por pneumonia e uso de ARV), o possível efeito protetor da vacina (OR=, EV=54%) perdeu sua significância estatística (IC95% 0,27-1,13). Amostras de 47 casos foram sorotipadas. Conclusão: Este é o primeiro estudo a avaliar a efetividade da vacina de polissacarídeos antipneumocócica entre os indivíduos infectados pelo HIV no Brasil, e a incluir dados sobre a distribuição dos sorotipos de pneumococo neste subgrupo. A vacina anti-pneumocócica e o uso de anti-retrovirais atuaram como fatores protetores contra doença pneumocócica invasiva. Estes resultados reforçam a recomendação do uso da vacina no Brasil, e indicam a necessidade de estudos com amostras maiores que possam elucidar de forma inequívoca o efeito da vacina antipneumocócica entre portadores do HIV / Background: HIV-infected individuals are at increased risk of bacterial pneumonia in general and of pneumonia due to S. pneumoniae in particular. S. pneumoniae is the leading cause of morbidity and mortality among children, elderly and those with underlying conditions that compromise the immune system or diminish the splenic function. HIV infection is associated to a tenfold increase in the incidence of bacterial pneumonias worldwide and S. pneumoniae is the most common causal agent identified. Immunization against S. pneumoniae with Polysaccharide pneumococcal vaccine is recommended for use in HIV-infected adults in Brazil. The effectiveness of PPV23 ranges from to approximately 30 to 80%, among those with normal immune function. Among HIV-infected adults, studies have suggested contradictory or inconclusive results. Objective: To assess the effectiveness of the 23-valent polysaccharide pneumococcal vaccine among HIV-infected adult patients in São Paulo, Brazil. Methods: A prospective matched case-control study among HIV-infected adults in São Paulo. Exposure is vaccination with PPV-23 and outcome is invasive disease. Case definition: HIV-infected individual over 18 years old, with invasive pneumococcal disease, defined as recovery of S. pneumoniae from a normally sterile site (e.g. such as blood, pleural fluid, spinal fluid, pericardial fluid). Controls: HIV-infected individuals over 18 years of age, with o history of documented or strong suspicion of invasive pneumococcal disease, receiving medical care at the same group of institutions, matched to the cases by level of CD4 lymphocyte cell counts, according to the following (<200; 200=500 cells/mm3), measured during the same period. Results: 79 cases and 241 controls were included; mean age of 39 years; 63% male; education level lower than high school for 50% of the sample; 63,5% whites, 19% reported living in \"sub-standard\" housing. Bacteremia was the most frequent clinical manifestation. Risk factors associated with invasie disease: alcohol use, IDU, lower level of education, \"sub-standard\" housing, close contact with child less than 10 years old and previous hospitalization with pneumonia. ARV use and pneumococcal vaccine were associated to decreased risk. Overall vaccine effectiveness was 65% (OR=0,35 IC95%:0,18-0,70). After adjustment for confounding factors (IDU, hospitalization with pneumonia, and ARV use) the point estimate for the effectiveness of 23-valent polysaccharide against all invasive pneumococcal infection was 45% (95% CI: <0% to 73%). Isolates from 47 were available for serotyping. 40 (85%) were of serotypes included in the PPV-23. All 11 isolates from previously vaccinated cases were of serotypes included in the vaccine. Conclusion To our knowledge this is the first study to evaluate vaccine effectiveness among HIV-infected persons in Brazil and the first to include data on pneumococcal serotype distribution among HIV-infected adults in Sao Paulo. Although we were not able to provide definitive answers regarding vaccine effectiveness among the specific population, we reinforce the role of anti-retroviral therapy on preventing invasive disease. Our findings support the continuity of the recommendation on immunizing HIV-infected patients with the polysaccharide vaccine at the same time that reinforces the need of more data on the subject

Magister Public Health - MPH / Background: The advent of antiretroviral therapy (ART) has significantly redefined the course of the HIV pandemic making HIV, a chronic illness rather than a death sentence. To maximize the efficacy of ART in improving survival rates of HIV/AIDS patients, lowering the incidence of opportunistic infections, reducing HIV transmission and minimizing the possibilities of developing drug resistance, long-term retention in care is critical. In South Africa, poor retention in care of 32% has been noted in women who were initiated on ART during pregnancy as compared to 13% in non-pregnant women initiated on ART. However, little is known in Swaziland about the retention in care in women who were initiated on ART during pregnancy and the factors that influence retention in care among this category of women. Aim: To determine the factors associated with poor retention in care among women initiated on ART during pregnancy at King Sobhuza II Public Health Unit (PHU) in Swaziland. Methodology: A quantitative, retrospective cohort review of 316 medical records of women who were initiated on ART during pregnancy from January 2012 to December 2013 was conducted. A data extraction sheet was used to collect data from the files of patients who were initiated on ART during pregnancy. The dataset was imported into IBM SPSS Statistic 20 Software for analysis. Bi-variate analysis was done to determine risk factors associated with retention in ART care at ART initiation and on the last ART refill visit. Kaplan-Meier analysis was used to determine retention in care at 6, 12, 24 and 36 months. Cox proportional hazards models were then used to determine factors associated with poor retention. Results: The overall retention rate of women who were initiated on ART during pregnancy at the PHU after a median duration on ART of 25.80 months [interquartile range (IQR): 16.70 – 30.98] of follow up was 74.1% (n=316). Most women initiated on ART during pregnancy (52.4%) became lost to follow up after giving birth as compared to 47.6% who became lost to follow up before giving birth. After 6 months on ART, the lost to follow up rate was 16.5% (n=316); but increased to 20.9% (n=316), 23.5% (n=243) and 26.9% (n=52) after 12, 24 and 36 months respectively. On the ART initiation visit, the factors associated with retention in care for pregnant women included being married, having the partner on ART, disclosing one’s HIV status to the partner, not drinking alcohol, being a non-smoker and reporting no financial challenges. In addition, on the last ART refill visit, the risk factors for retention in care for women initiated on ART during pregnancy were having the ART regimen changed, having regular CD4 cell count done, rise in CD4 cell count, good adherence on ART and use of contraceptive other than the condom for family planning after delivery. Conclusion: The retention in care for women who were initiated on ART during pregnancy was found to be lower than in the general adult population. However, the study findings on retention in care are similar to what has been found in other settings. The factors influencing poor retention also mirror those found in the other parts of sub-Saharan Africa. Whilst decentralisation of ART services improves ART coverage it should be coupled with strategies aimed at improving patient retention.

La majorité des infections par le VIH de l’enfant sont acquises par transmission de la mère à l’enfant en période périnatale. Le fait que le système immunitaire des nourrissons soit « immature » par certains aspects et le fait que le virus transmis ait déjà échappé au contrôle de l’environnement immunitaire maternel, génétiquement proche de celui de l’enfant, sont probablement responsables du risque important que l’évolution naturelle soit extrêmement rapide lorsque l’infection est acquise en tout début de vie. En l’absence d’un traitement antirétroviral, la réplication virale va en effet se maintenir à un niveau très élevé pendant les premières années de vie chez tous les enfants et jusque 25% d’entre eux vont évoluer jusqu’au stade SIDA ou le décès durant leur 1ère année de vie. L'utilisation des traitements antirétroviraux puissants actuellement disponibles permet de réduire drastiquement la réplication virale et a profondément modifié le pronostic de l’infection par le VIH. D’une infection inéluctablement fatale, elle est devenue une affection chronique peu ou pas évolutive pour autant que le traitement antirétroviral soit pris de façon ininterrompue. Dans ce travail, nous présentons les résultats d’une étude prospective qui incluait les patients chez qui un traitement antirétroviral efficace était débuté pour la première fois. Nous avons pu démontrer que le nombre de lymphocytes CD4+ naïfs était d’autant plus rapidement reconstitué après l’initiation d’un traitement antiviral que celui-ci avait été débuté jeune. Non seulement les patients restauraient leurs nombres de cellules CD4+, et leurs cellules CD4+ naïves en particulier, mais des tests plus fonctionnels des lymphocytes (réponses lymphoprolifératives aux mitogènes) montraient également une nette amélioration sous traitement antirétroviral. Malgré cela, certaines anomalies, et en particulier l’activation des lymphocytes CD8+ persistaient après un an de traitement.Dans une étude cross-sectionnelle ayant inclus 46 patients, nous avons ensuite démontré qu’un traitement antirétroviral de longue durée débuté lorsque l’immunodépression est déjà sévère permet de récupérer des réponses lymphoprolifératives semblable à celles des progresseurs lents, y compris contre un antigène spécifique du VIH ce qui n’est généralement pas le cas chez les adultes. Toutefois, des altérations dans la sécrétion des cytokines en réponse à un mitogène (la phytohémagglutinine) persistent, et la sécrétion de cytokines après stimulation par un antigène spécifique du VIH est biaisée vers une réponse de type Th2, montrant encore une fois que si les capacités d’immunorestauration des enfants sont très importantes, des anomalies immunitaires persistent même après un traitement de longue durée lorsque le traitement est débuté à un stade avancé dans l’évolution de la maladie.Dans le contexte de l’absence d'espoir d'atteindre l'éradication du virus par la seule utilisation des drogues antirétrovirales, la caractérisation du phénotype des patients « non progresseurs » prend tout son sens. Nous avons donc axé notre travail suivant sur la caractérisation des patients chez qui la maladie progresse lentement et qui gardent une réplication virale basse sans prendre de traitement antirétroviral. Ces patients ont une faible activation et une faible différentiation de leurs lymphocytes CD4+ et un profil particulier de réponses CD4 vis-à-vis de la protéine Gag du VIH.Enfin, la dernière partie du travail décrit les bénéfices de ces traitements lorsqu’ils sont administrés très précocement (dans les 2 mois après la naissance) et le devenir à long terme de ces enfants en se basant sur la cohorte suivie au sein du CHU Saint-Pierre. L’histoire naturelle de l’infection par le VIH de l’enfant contaminé par sa mère est en effet caractérisée par une évolution bi-modale. Trois quarts des enfants infectés vont avoir une évolution comparable aux patients infectés à l’âge adultes, mais le dernier quart aura une évolution beaucoup plus rapide menant au développement d’une affection classant l’enfant au stade SIDA ou au décès dans la première année de vie. De plus, à cet âge, il y a un large recouvrement entre les valeurs des taux de lymphocytes CD4+ des enfants qui vont évoluer rapidement ou plus lentement. Nous avons participé activement à la démonstration qu’un traitement antirétroviral très précoce est très bien supporté par les nourrissons, est capable d’empêcher la réplication virale de façon durable et réduit la quantité de DNA proviral intracellulaire prévenant de ce fait les manifestations cliniques ou biologiques de l’infection. Alors que les adultes infectés par le VIH expriment quasi toujours des anticorps contre le VIH, même lorsqu’ils sont traités très précocement par une trithérapie, les enfants traités précocement deviennent souvent séronégatifs. Ils perdent les anticorps spécifiques contre le VIH transmis durant la grossesse mais n’en sécrètent pas par la suite, alors qu’ils peuvent les synthétiser dès que le traitement antirétroviral est interrompu. Ces patients traités très précocement ont un profil particulier caractérisé par un système immunitaire non altéré par l’infection par le VIH mais aussi par l’absence de défenses spécifiques contre ce virus et un réservoir viral très faible. / Doctorat en Sciences médicales (Médecine) / info:eu-repo/semantics/nonPublished

Philosophiae Doctor - PhD / AIDS is considered a pandemic causing millions of deaths worldwide and a cure for this disease is still not available. Failure to implement early treatments due to the poor diagnostic methods and ineffective therapeutic regimens to treat HIV patients to achieve complete viral eradication from the human body has encouraged the escalation of this disease at an exponential rate. Though the current treatment regimens (High Active Antiretroviral Therapy) have aided in increasing the lifespan of HIV patients, it still suffers from some shortcomings such as adverse side effects and non-eradication of the virus. Thus, there is a need for a non-toxic therapeutic regimen to stop further infection of HIV-infected patients. Antimicrobial Peptides (AMPs) are naturally occurring peptides which are components of the first line of defence of many organisms against infections and have been proven to be promising therapeutic agents against HIV. The use of AMPs as anti-microbial agents is due to the fact that most AMPs have a net positive charge and are mostly hydrophobic molecules. These features allow AMPs to be site directed electro-statistically to the mostly negatively charged pathogens. In a previous study, a number of novel anti-HIV AMPs was identified using a predictive algorithm Profile Hidden Markov Models (HMMER). The AMP's threedimensional structures were predicted using an in-silico modelling tool I-TASSER and an insilico protein-peptide interaction study of the AMPs to HIV protein gp120 was performed using PatchDock. Five AMPs were identified to bind gp120, at the site where gp120 interacts with CD4 to prevent HIV invasion and HIV replication. Therefore, the aims of this research were to perform in-silico site-directed mutation on the parental anti-HIV AMPs to increase their binding affinity to the gp120 protein, validate the anti-HIV activity of these peptides and confirm the exclusivity of this activity by testing possible anti-bacterial and anti-cancer activities of the AMPs. Firstly, the five parental anti-HIV AMPs were used to generate mutated AMPs through insilico site-directed mutagenesis. The AMPs 3-D structures were determined using I-TASSER and the modelled AMPs were docked against the HIV protein gp120 using PatchDock. Secondly, an "in house" Lateral Flow Device (LFD) tool developed by our industrial partner, Medical Diagnostech (Pty) Ltd, was utilised to confirm the in-silico docking results. Furthermore, the ability of these AMPs to inhibit HIV-1 replication was demonstrated and additional biological activities of the peptides were shown on bacteria and cancer cell lines. In an effort to identify AMPs with increased binding affinity, the in-silico results showed that two mutated AMPs Molecule 1.1 and Molecule 8.1 bind gp120 with high affinity, at the point where gp120 bind with CD4. The molecular binding however showed that only Molecule 3 and Molecule 7 could prevent the interaction of gp120 protein and CD4 surface protein of human cells, in a competitive binding assay. Additionally, the testing of the anti-HIV activity of the AMPs showed that Molecule 7, Molecule 8 and Molecule 8.1 could inhibit HIV-1 NL4-3 with maximal effective concentration (EC₅₀) values of 37.5 μg/ml and 93.75 μg/ml respectively. The EC₅₀ of Molecule 8.1 was determined to be around 12.5 μg/ml. This result looks promising since 150 μg/ml of the AMPs could not achieve 80% toxicity of the human T cells, thus high Therapeutics Index (TI) might be obtained if 50% cytotoxic concentration (CC₅₀) is established. Further biological activity demonstrates that Molecule 3 and Molecule 7 inhibited P. aeruginosa completely after 24 hours treatment with peptide concentrations ranging from 0.5 mg/ml to 0.03125 mg/ml. Nevertheless, moderate inhibition was observed when CHO, HeLa, MCF-7 and HT-29 were treated with these peptides at peptides concentration of 100 μg/ml. The ability of these AMPs to block the entrance of HIV via the binding to CD4 of the host cells is a good concept since they pave the way for the design of anti-HIV peptide-based drugs Entry Inhibitors (FIs) or can be exploited in the production microbicide gels/films to suppress the propagation of the virus. / DST-NIC/Mintek

Les pharmacocinétiques de deux inhibiteurs de protéase, le lopinavir et l'atazanavir et celle d'un inhibiteur non nucléosidique de la transcriptase inverse, la névirapine, ont été étudiées chez l'enfant par une approche de population. Cette approche nous a permis d'étudier les différentes sources de variabilité pharmacocinétique de chaque molécule antirétrovirale. La prise en compte des relations concentration-effet précédemment établies chez l'adulte, nous a permis, concernant l'atazanavir et la névirapine, de ré-évaluer les recommandations posologiques chez l'enfant en termes d'efficacité et de toxicité. L'étude réalisée sur le lopinavir/ritonavir nous a conduits à comparer en termes de pharmacocinétique, d'efficacité et de tolérance, le changement de rythme d'administration de deux prises à une unique prise par jour. Les résultats obtenus ont mis en évidence dans les trois études une augmentation de la clairance et du volume de distribution apparents en fonction du poids. Pour l'atazanavir, il a été montré que la co-administration de ritonavir ou de ténofovir résulte respectivement en une diminution ou augmentation de sa clairance apparente. Pour la névirapine, un effet de l'âge sur sa biodisponibilité a été mis en évidence, la biodisponibilité relative augmentant avec l'âge. En conclusion, le changement de rythme d'administration du lopinavir/ritonavir s'est avéré être équivalent sur le plan pharmacocinétique, mais a résulté en une baisse de la proportion de patients présentant une charge virale indétectable. Les recommandations posologiques actuelles d'atazanavir/ritonavir pourraient mener à un sur-dosage pour l'intervalle de poids 32-50 kg. Les doses de névirapine recommandées par l'Organisation Mondiale de la Santé pourraient mener à un sous-dosage pour les enfants pesant entre 3 et 10 kg. / The pharmacokinetics of two protease inhibitors, lopinavir and atazanavir and that of a non-nucleoside reverse transcriptase inhibitors, nevirapine, has been studied in children by a population approach. This approach allowed us to study the factors affecting the pharmacokinetic variability of each antiretroviral drug. Based on adult concentration-effect relationships, we evaluated the recommended dosage of atazanavir and nevirapine in children in terms of efficacy and toxicity. The study of lopinavir led us to compare in terms of pharmacokinetics, efficacy and safety, a switch from the twice-daily to the once-daily lopinavir/ritonavir regimen. These three studies showed that the apparent clearance and the apparent volume of distribution increased allometrically with body weight. For atazanavir, it was shown that co-administration of ritonavir or tenofovir resulted respectively in a decrease or increase of its apparent clearance. For nevirapine, an effect of age on its bioavailability was pointed out, the relative bioavailability increased with age. In conclusion, the switch of lopinavir/ritonavir regimen was found to be to be equivalent in terms of pharmacokinetics, but resulted in a decrease in the proportion of patients with undetectable viral load. Current dosing recommendations for atazanavir/ritonavir may lead to over-dosage for the body weight range 32-50 kg. Doses of nevirapine recommended by the World Health Organization could lead to under-dosing for children weighing between 3 and 10 kg.

Magister Artium (Child and Family Studies) - MA(CFS) / HIV patients face an array of social and psychological problems, such as depression, which can affect their quality of life. Moreover, HIV infection is also linked to psychological distress such as anxiety. In addition, avoidant emotion-focused strategies such as acceptance, wishful thinking and self-blame are associated with higher levels of psychological distress in persons with HIV. Current health services in the city of Cairo, Egypt, are not adapted to provide advice and psychological support to people living with HIV to aid in the development of problem-solving skills to cope with the stress of living with HIV. The purpose of this study was to examine the relationship between quality of life, psychological distress and the coping strategies of persons living with HIV/AIDS in Cairo, Egypt. A quantitative methodology with a cross-sectional correlational design was adopted in this study. Data collection entailed questionnaires that consisted of four sections: Demographics, Quality of Life Enjoyment and Satisfaction Questionnaire (Q-LES-Q), Depression, Anxiety, Stress Scales (DASS) and the Cope Inventory. The sample consisted of 202 HIV/AIDS participants who access the National AIDS Program (NAP). The data were analysed using the Statistical Program for Social Science V23 (SPSS). The results are provided using descriptive and inferential statistics. The findings of the study show that in terms of the prevalence of psychological distress, the moderate scales were stress and depression, while the severe scale was anxiety. The most prevalent of coping styles was emotion-focused coping, specifically in terms of acceptance and religion. In terms of the prevalence of quality of life, the highest scores were for family and social relationships, while the lowest scores were for sexual drive and leisure time activities. The findings also show that psychological distress and certain coping styles such as substance use negatively predicted quality of life of patients with HIV/AIDS. Positive predictors included coping styles such as venting, positive reframing, humour, acceptance and religion.

[pt] O comportamento eletroquímico e o desenvolvimento de metodologias eletroanalíticas para a determinação de zidovudina (AZT), lamivudina (3TC) e zalcitabina (DDC) em medicamentos foram investigados no presente trabalhoatravés de técnicas voltamétricas. Estas substâncias possuem atividade antiretroviral, sendo utilizadas no tratamento de indivíduos portadores de HIV. AZT, 3TC e DDC sofrem redução na superfície do eletrodo de mercúrio. Ainfluência dos parâmetros eletroquímicos, tais como, velocidade de varredura, amplitude, natureza do eletrólito suporte e pH foram estudados para selecionar as melhores condições para quantificar estes princípios ativos em formulações farmacêuticas. A redução da zidovudina ocorre em apenas uma etapa, sendo esta atribuída à redução do grupo azida (-N3), originando em um pico de potencial de-970 mV vs Ag/AgCl (3 mol L-1), utilizando tampão fosfato, pH 8,0. A faixa linear encontrada foi de 0,25 a 1,25 mg L-1. O mecanismo de redução da zidovudina envolve a participação de 4 elétrons e 2 prótons. Adicionalmente novos estudos eletroquímicos foram desenvolvidos para a determinação de lamivudina e de zalcitabina através da voltametria em pulso diferencial em tampão Clark-Lubs, pH 2,0, onde se obteve um pico no potencial de -1160 e -1180 mV, respectivamente. Lamivudina obteve faixa linear de 2,28 a 8,85 mg L-1 e zalcitabina de 10,0 a 23,3 mg L-1. O mecanismo de redução destas drogas envolve a participação de números iguais de prótons e elétrons. Todas as metodologias desenvolvidas também foram validadas. / [en] The electrochemical behaviour and the development of an analytical methodology for the zidovudine (AZT), lamivudine (3TC) and zalcitabine (DDC)determination in drugs using voltammetrics techniques were the objectives of the current study. These drugs have been extensively used in the treatment of HIV patients because its antiretroviral activity. The AZT, 3TC and DDC are reduced at a hanging mercury drop electrode (HMDE). The influence of electroanalytical parameters such as, scan rate, amplitude, nature of the support electrolyte and pH in the signal was verified to select the best conditions for quantifying these drugs in pharmaceutical forms. The obtained results had showed AZT, in solution of pH 8.0 of phosphate buffer, with only one reduction in -970 mV vs Ag/AgCl (3 mol L-1). This peak is due to reduction of the azido group present on zidovudine molecule. The linear range of standards is from 0.25 to 1.25 mg L-1. The electrodic mechanism involving the participation of 4 electrons and 2 protons in the reduction of AZT molecules. News studies were development for lamivudine and zalcitabine determination by differential pulse voltammetry in solution of pH 2.0 of Clark-Lubs buffer, with only one reduction in - 1160 and -1180 mV, respectively. Lamivudine has been the linear range of standards from 2.28 to 8.85 mg L-1 and zalcitabine from 10.0 to 23.3 mg L-1. The mechanism of these drugs involving the participation of the same number of electrons and protons. All these analytical methods were validated.

Introduction: Decentralization of antiretroviral therapy (ART) services faces decreasing quality when increasing ART coverage. This study compares nurse initiated and managed patients to doctor managed patients under these circumstances, using retention in care as a crude measure of quality of care. Methods: This was an observational retrospective cohort study. A simple data abstraction tool was used to collect baseline patient data from medical records of HIV positive patients (N=871) initiating ART at Mbabane Government Hospital and four of its outreach clinics, between 1st January and 30th June 2011. Descriptive summary statistics and comparison of the two cohorts using multivariate analysis was done. Results There was no statistically significant difference in retention rates between the doctors and nurses cohorts at 69.1% and 70.9%, respectively (P was 0.56). After adjusting for sex, haemoglobin, CD4 cell count, weight and WHO stage, the odds of being retained in care were similar between the two groups, adjusted OR: 1.11(95% CI: 0.72, 1.69), with a p value of 0.64. Haemoglobin and weight were positively associated with retention in care, while male sex was negatively associated with retention in care. Discussion: The similar retention rates between the two cohorts suggest that in terms of retention in care the service provided by the nurses was comparable to that provided by doctors. This is important to ART program managers as they scale-up ART decentralization. Conclusion: Task-shifting of ART initiation from doctors to nurses is feasible as nurse initiated and managed antiretroviral therapy is comparable to doctor initiated and managed treatment. / Dissertation (MSc)--University of Pretoria, 2014. / gm2014 / Clinical Epidemiology / unrestricted