Orientador : Maria Palmira Daflon Gremião / Coorientador: Marco Vinícius Chaud / Banca: Victor Manuel Cardoso Figueiredo Balcão / Banca: Clovis Augusto Ribeiro / Banca: Ana Luiza Ribeiro de Souza / Banca: Beatriz Stringhetti Ferreira Cury / Resumo: Neste estudo foi proposta a utilização do processo antissolvente supercrítico (SAS), no qual o gás carbônico foi selecionado como agente antissolvente, para a obtenção de sistema de liberação controlada de fármaco. No delineamento experimental fatorial 32, a proporção entre zidovudina e poli(L-ácido lático) (AZT:PLLA) representou o fator X1, e as condições de temperatura e pressão representou o fator X2, sendo ambos variáveis independentes; o rendimento do processo e a morfologia macroscópica das partículas representaram as variáveis dependentes. No presente estudo, as variáveis dependentes determinaram os produtos dos lotes que foram selecionados para a caracterização do estado sólido, teor de AZT, tipo e cinética de liberação de AZT e permeação intestinal de AZT. A caracterização do estado sólido utilizou as análises de dispersão de raios X (DRX), microscopia eletrônica de varredura (MEV), espectroscopia de infravermelho com transformada de Fourier (FITR) e calorimetria exploratória diferencial (DSC). O teor de AZT foi determinado por metodologia analítica validada. O teste de dissolução forneceu dados para as avaliações da liberação e da cinética de liberação de AZT. O modelo de saco intestinal invertido de rato foi adotado para o estudo ex vivo de permeação intestinal do AZT. Os lotes L3 (91,54 % de rendimento e amostra com aspecto uniforme), L5 e L9 (59,06 % e 51,50 % de rendimento, respectivamente; ambos resultaram em amostra com aspecto não uniforme e sólido filamentoso) resultantes do planejamento fatorial 32 foram selecionados para os estudos analíticos. O lote L3 de proporção AZT:PLLA (1:2, m/m) resultou em rendimento de 91,54 % e teor de AZT 58,76 % elevados em comparação aos lotes produzidos e foi selecionado para o estudo de permeação intestinal. A permeabilidade do AZT a partir do lote L3 (9,87 ± 0,47 %) foi maior em relação ao AZT puro (3,84 ± 0,45 %). O AZT permaneceu ... / Abstract: In this study we proposed the use of supercritical antisolvent process (SAS), in which carbon dioxide was selected as antisolvent agent, to obtain drug controlled release system. Factorial 32 experimental design use the of zidovudine and poly (L-lactic acid) ratio (AZT:PLLA ) as X1 factor, also the temperature and pressure conditions as X2 factor, both independent variables, the process yield and particles macroscopic morphology represented the dependent variables. In this study, the dependent variables determined the product that have been selected to solid state characterization, AZT content, type, release kinetics, and intestinal permeation. Solid state characterization used X-ray diffraction (XRD) analysis, scanning electron microscopy (SEM), infrared spectroscopy with Fourier transform (FTIR), and differential scanning calorimetry (DSC). The AZT content was determined by validated analytical methodology. The dissolution test provided data for AZT release and release kinetics evaluation. The rat everted gut sac model was take on for the ex vivo study of intestinal permeation of AZT. The L3 (91.54 % yield and sample with uniform appearance), L5 and L9 lots (59.06 % and 51.50 % yield , respectively , both of which resulted in samples with non-uniform and solid filamentous appearance) resulting from factorial 32 planning were selected for analytical studies. The L3batch ratio of AZT:PLLA (1:2, w/w) resulted in a high yield of 91.54 % and 58.76 % AZT content compared to batches produced, and was selected for the study of intestinal permeation . The AZT permeability from L3 lot (9.87 ± 0.47%) was higher than pure AZT (3.84 ± 0.45 %). AZT remained in crystalline form and PLLA remained in semi - crystalline form, in lots L3, L5, and L9 produced by the SAS process, when evaluated by SEM, XRD, DSC, and FTIR. This study demonstrated that it is possible to use the SAS process to obtaining modified release systems for poorly permeable drugs ... / Doutor

AZT (3'-azido-2’, 3’-dideoxythymidine) has been shown to prolong the survival of patients infected with human immunodeficiency virus (HIV) and decrease the severity of opportunistic infections. Such studies have prompted the use of AZT to treat symptomless individuals infected with HIV in the hope of delaying or even preventing the progression to acquired immunodeficiency syndrome (AIDS). However, before chronic use of AZT in symptomless individuals is initiated, it is important to establish whether this anti-viral drug would directly alter the phenotype and functions of the cells involved in the immune system. In the current study, we observed that AZT when administered orally for 7 -14 days into DBA/2 mice at 500 - 1000 mg/kg body weight induced a dose-dependent decrease in cellularity of the thymus. AZT caused significant alterations in the thymus resulting from a significant decrease in the number of double-positive (CD4⁺CD8​​⁺) cells and an increase in the number of double-negative (CD4⁻CD8⁻) cells. Interestingly, after the i.p. administration of interleukin-2 (IL-2) simultaneously with AZT, the total cellularity of the thymus was completely reconstituted. We also observed that AZT effectively suppressed the in vivo T cell response to conaibumin and gp120 of HIV. Furthermore, the addition of AZT to in vitro cultures caused a dose-dependent decrease in T and B cell proliferative responses to mitogens at 50μM or greater concentrations. Also, AZT inhibited the generation of cytotoxic T lymphocytes when added to the culture and this inhibition was reconstituted by the addition of exogenous IL-2. Together, our studies demonstrate that AZT modulates the phenotype and function of cells of the immune system which, in turn, could have marked repercussions on immune responses of the host toward infections and cancers. Also, our data demonstrating that AZT can suppress T cell responsiveness against HIV antigens caution against chronic use of AZT in asymptomatic HIV-infected individuals. / Master of Science

Thesis (MSc.(Anatomy)--Faculty of Health Sciences)-University of Pretoria, 2008.] / Includes bibliographical references.

This study has involved the design, synthesis and evaluation of novel HIV-1 enzyme inhibitors accessed by synthetic elaboration of Baylis-Hillman adducts. Several series of complex coumarin-AZT and cinnamate ester-AZT conjugates have been prepared, in high yields, by exploiting the click reaction between appropriate Baylis-Hillman derived precursors and azidothymidine (AZT), all of which have been fully characterised using spectroscopic techniques. These conjugates, designed as potential dual-action HIV-1 inhibitors, were tested against the appropriate HIV-1 enzymes, i.e. HIV-1 reverse transcriptase and protease or HIV-1 reverse transcriptase and integrase. A number of the ligands have exhibited % inhibition levels and IC50 values comparable to drugs in clinical use, permitting their identification as lead compounds for the development of novel dual-action inhibitors. In silico docking of selected ligands into the active sites of the respective enzymes has provided useful insight into binding conformations and potential hydrogen-bonding interactions with active-site amino acid residues. A series of furocoumarin carboxamide derivatives have been synthesised in four steps starting from resorcinol and these compounds have also been tested for HIV-1 integrase inhibition activity. The structures of unexpected products isolated from Aza-Baylis-Hillman reactions of N-tosylaldimines have been elucidated by spectroscopic analysis, and confirmed by single crystal X-ray analysis. A mechanism for what appears to be an unprecedented transformation has been proposed. Microwave-assisted SeO₂ oxidation of Baylis-Hillman-derived 3-methylcoumarins has provided convenient and efficient access to coumarin-3-carbaldehydes, and a pilot study has revealed the potential of these coumarin-3-carbaldehydes as scaffolds for the construction of tricyclic compounds. The HCl-catalysed reaction of tert-butyl acrylate derived Baylis-Hillman adducts has been shown to afford 3-(chloromethyl)coumarins and α-(chloromethyl)cinnamic acids, the Zstereochemistry of the latter being established by X-ray crystallography. ¹H NMR-based experimental kinetic and DFT-level theoretical studies have been undertaken to establish the reaction sequence and other mechanistic details. Base-catalysed cyclisation on the other hand, has been shown to afford 2H-chromene rather than coumarin derivatives.

Thesis (Dr.P.H.)--University of Texas Health Science Center at Houston, School of Public Health, 1999. / Includes bibliographical references (leaves 127-136).

The overall approach in the present investigation has been to explore applications of the Morita-Baylis-Hillman (MBH) reaction in asymmetric synthesis and in the continuation of systems with medicinal potential. To this end, a series of varied camphor-derived acrylate esters was prepared to serve as chiral substrates in asymmetric Morita-Baylis- Hillman reactions. Reduction of N-substituted camphor-10-sulfonamides afforded the 3- exo-hydroxy derivatives as the major products. Acylation of the corresponding sodium alkoxides gave the desired 3-exo-acrylate esters, isolation of which was complicated by concomitant formation of hydrochlorinated and diastereomeric competition products. Bulky camphorsulfonamides containing alkyl, dialkyl, aromatic and adamantyl groups were selected as N-substituents with the view of achieving stereoselective outcome in subsequent MBH reactions. The synthesis of novel camphor-derived Morita-Baylis-Hillman adducts using various pyridine-carboxaldehydes proceeded with exceptionally high yields with diastereoselectivities ranging from 7-33 % d.e. Both 1D and 2D NMR and HRMS techniques were employed to confirm the structures and an extensive study of the electropositive fragmentation patterns of a number of camphor-derived chiral acrylate esters was conducted. Attention has also been given to the application of MBH methodology in the construction of heterocyclic ‘cinnamate-like’ AZT conjugates which were designed to serve as dualaction HIV-1 integrase-reverse transcriptase (IN-RT) inhibitors. A number of pyridine carboxaldehyde-derived MBH adducts were synthesized using methyl, ethyl and t-butyl acrylates in the presence of 3-hydroxyquinuclidine (3-HQ) as catalyst. The yields for these reactions were excellent. The resulting MBH adducts were acetylated and subjected to aza-Michael addition using propargylamine. The resulting alkylamino compounds were then used in ‘Click reactions’ to form the targeted AZT-conjugates in moderate to excellent yield. In silico docking of computer modelled AZT-conjugates into the HIV-1 integrase and reverse transcriptase enzyme-active sites and potential hydrogen-bonding interaction with active-site amino acid residues were identified. The electrospray MS fragmentations of the AZT and the novel AZT-conjugates were also investigated and common fragmentation pathways were identified.