Submitted by Nivaldo Melo (nivaldo.melo@ufvjm.edu.br) on 2015-11-30T16:45:15Z No. of bitstreams: 2 marcos_vinicius_barbosa.pdf: 2721412 bytes, checksum: 173504d75a25f2de0569ca85af50f0df (MD5) license_rdf: 23898 bytes, checksum: e363e809996cf46ada20da1accfcd9c7 (MD5) / Approved for entry into archive by Rodrigo Martins Cruz (rodrigo.cruz@ufvjm.edu.br) on 2015-12-03T16:06:21Z (GMT) No. of bitstreams: 2 marcos_vinicius_barbosa.pdf: 2721412 bytes, checksum: 173504d75a25f2de0569ca85af50f0df (MD5) license_rdf: 23898 bytes, checksum: e363e809996cf46ada20da1accfcd9c7 (MD5) / Made available in DSpace on 2015-12-03T16:06:22Z (GMT). No. of bitstreams: 2 marcos_vinicius_barbosa.pdf: 2721412 bytes, checksum: 173504d75a25f2de0569ca85af50f0df (MD5) license_rdf: 23898 bytes, checksum: e363e809996cf46ada20da1accfcd9c7 (MD5) Previous issue date: 2014 / Funda??o de Amparo ? Pesquisa do estado de Minas Gerais (FAPEMIG) / Conselho Nacional de Desenvolvimento Cient?fico e Tecnol?gico (CNPq) / Rede Mineira de Pesquisas em Nanobiotecnologia (Nanobiomg) / Coordena??o de Aperfei?oamento de Pessoal de N?vel Superior (Capes) / O paclitaxel (PTX) ? um agente quimioter?pico usado no tratamento de v?rios tipos de tumores s?lidos como c?ncer de pr?stata, mama e ov?rio. Devido a sua baixa solubilidade aquosa, o PTX (Taxol?) ? formulado em um ve?culo constitu?do por Cremophor EL? e ?lcool desidratado. No entanto, rea??es de hipersensibilidade aguda, nefrotoxicidade e neurotoxicidade, al?m de problemas de instabilidade f?sico-qu?mica t?m sido relatados com o uso do Taxol?. Diante disso, o objetivo desse estudo foi desenvolver um novo sistema lipossomal multifuncionalizado para veicular o PTX e avaliar a citotoxicidade em linhagens tumorais de c?ncer de mama (MCF-7 e MDA-MB-231). Inicialmente foi desenvolvido e validado um m?todo anal?tico para quantifica??o do PTX por espectrofotometria derivada no ultravioleta. O m?todo apresentou linearidade adequada e permitiu a quantifica??o do PTX na formula??o com adequada precis?o, exatid?o e especificidade. As formula??es propostas foram compostas por dioleilfosfatidiletanolamina (DOPE), hemisuccinato de colesterila (CHEMS), e diestearoilfosfatidiletanolamina associado ao polietilenoglicol 2000 (DSPE-PEG2000); ou por fosfatidilcolina de soja (SPC), DOPE, CHEMS e DSPE-PEG e preparadas pelo m?todo de hidrata??o do filme lip?dico. A influ?ncia da composi??o lip?dica qualitativa e quantitativa (10 e 20 mM) e da concentra??o do PTX (0,5; 1,0 ou 20 mg/mL) nas caracter?sticas f?sico-qu?micas dos lipossomas foi avaliada. Uma redu??o no teor de encapsula??o foi verificada na formula??o composta por DOPE:CHEMS:DSPE-PEG a medida que a concentra??o de PTX foi aumentada (p<0,05), rela??o que n?o foi observada para a formula??o com SPC. A associa??o com o PTX promoveu aumento significativo no di?metro m?dio das ves?culas em compara??o com a formula??o sem PTX e o aumento na concentra??o do f?rmaco tamb?m levou a um aumento no di?metro. O potencial zeta de todas as formula??es apresentou valores pr?ximos da neutralidade. Essa prepara??o apresentou boa estabilidade, com manuten??o do di?metro m?dio, teor de encapsula??o e potencial zeta durante 100 dias de armazenamento. Modifica??es da organiza??o supramolecular da DOPE indicativas de mudan?as de fase lamelar para n?o-lamelar em fun??o da redu??o do pH foram observadas nas prepara??es compostas por DOPE:CHEMS:DSPE-PEG. A inclus?o da SPC na bicamada, alterou a pH-sensibilidade do sistema e maior reten??o do PTX foi observada em pH mais ?cido quando comparada ? formula??o sem a SPC (p<0,05). A an?lise morfol?gica demonstrou a exist?ncia de part?culas esf?ricas regulares com di?metros heterog?neos para as formula??es contendo ou n?o PTX. A atividade citot?xica in vitro mostrou que o PTX associado aos lipossomas com folato (LpHSF-PTX) foi mais ativo que o PTX encapsulado nos lipossomas sem associa??o ao folato (LpHS-PTX) e que o PTX livre. Em suma, esses resultados sugerem que os LpHSF-PTX representam uma alternativa promissora para entrega intracelular do PTX no tratamento do c?ncer de mama. / Disserta??o (Mestrado) ? Programa de P?s-gradua??o em Ci?ncias Farmac?uticas, Universidade Federal dos Vales do Jequitinhonha e Mucuri, 2014. / ABSTRACT Paclitaxel (PTX) is a chemotherapeutic agent used in the treatment against various solid tumors such as prostate cancer, breast and epithelial ovarian carcinoma. Due to its low aqueous solubility, PTX is prepared in a vehicle composed of Cremophor EL? and dehydrated ethanol (Taxol?). However, acute hypersensitivity reactions, nephrotoxicity and neurotoxicity, as well as problems of physical-chemical instability have been reported with use of Taxol?. Thus, the purpose of this study was to develop a multifunctional liposomal system to carrier PTX, and to evaluate its cytotoxicity in tumor cell lines breast cancer (MCF-7 and MDA-MB-231). Firstly, an analytical method for quantification of PTX by derivative ultraviolet spectrophotometry was developed and validated. The method showed adequate linearity and allowed the quantification of PTX formulation with adequate precision, accuracy and sensitivity. The proposed formulations were composed by dioleoylphosphatidylethanolamine (DOPE), cholesteryl hemisuccinate (CHEMS) and distearoylphosphatidylethanolamine-polyethyleneglycol 2000 (DSPE-PEG) or soy phosphatidylcholine (SPC), DOPE, CHEMS and DSPE-PEG and prepared by lipid film hydration method. The influence of the qualitative and quantitative lipid composition (10 and 20mM), and the PTX concentration (0.5, 1.0 and 2.0 mg/mL) on the physicochemical characteristics of liposomes was evaluated. A decrease in the encapsulation percentage was observed in formulation composed DOPE:CHEMS:DSPE-PEG as the PTX concentration was increased (p<0,05), however, this relation was not observed for formulations with SPC. The PTX association caused a significant increase in the mean diameter of the vesicle compared to the formulation without PTX and the increase in drug concentration also led to an increase in mean diameter. The zeta potential for all formulations showed values near neutrality. This preparation showed good stability, maintaining the average diameter, encapsulating percentage and zeta potential over 100 days of storage. Alterations in the supramolecular organization of DOPE indicative of changes in lamellar to non-lamellar phase due to pH reduction in formulation made up DOPE:CHEMS:DSPE-PEG could be observed. The inclusion of SPC in the bilayer decreased the pH sensitivity of this system and increased of PTX retention in more acidic pH was observed as compared to the formulation without SPC (p<0,05). Morphological analysis evidenced the existence of regular spherical vesicles with heterogeneous diameters in formulations with and without PTX. The in vitro cytotoxicity assay showed the PTX associated with liposomes folate-targeted (LpHSF-PTX) was more active in PTX encapsulated than in non-targeted liposomes (LpHS-PTX) and PTX free. These results suggest that the LpHSF-PTX represent a promising alternative for intracellular PTX delivery in the treatment of breast cancer.

Made available in DSpace on 2014-12-17T15:42:03Z (GMT). No. of bitstreams: 1 NiveaNM_DISSERT.pdf: 4430545 bytes, checksum: 80673edc56d71303c3e6a5cd7b2b1996 (MD5) Previous issue date: 2012-07-12 / Coordena??o de Aperfei?oamento de Pessoal de N?vel Superior / The present study describes the stability and rheological behavior of suspensions of poly (N-isopropylacrylamide) (PNIPAM), poly (N-isopropylacrylamide)-chitosan (PNIPAMCS), and poly (N-isopropylacrylamide)-chitosan-poly (acrylic acid) (PNIPAM-CS-PAA) crosslinked particles sensitive to pH and temperature. These dual-sensitive materials were simply obtained by one-pot method, via free-radical precipitation copolymerization with potassium persulfate, using N,N -methylenebisacrylamide (MBA) as a crosslinking agent. Incorporation of the precursor materials into the chemical networks was confirmed by elementary analysis and infrared spectroscopy. The influence of external stimuli such as pH and temperature, or both, on particle behavior was investigated through rheological measurements, visual stability tests and analytical centrifugation. The PNIPAM-CS particles showed higher stability in acid and neutral media, whereas PNIPAM-CS-PAA particles were more stable in neutral and alkaline media, both below and above the LCST of poly (Nisopropylacrylamide) (stability data). This is due to different interparticle interactions, as well as those between the particles and the medium (also evidenced by rheological data), which were also influenced by the pH and temperature of the medium. Based on the results obtained, we found that the introduction of pH-sensitive polymers to crosslinked poly (Nisopropylacrylamide) particles not only produced dual-sensitive materials, but allowed particle stability to be adjusted, making phase separation faster or slower, depending on the desired application. Thus, it is possible to adapt the material to different media / Esse estudo descreve a estabilidade e o comportamento reol?gico de suspens?es aquosas de part?culas reticuladas sensitivas de poli (N-isopropilacrilamida) (PNIPAM), poli (N-isopropilacrilamida)-quitosana (PNIPAM-QS) e poli (N-isopropilacrilamida)-quitosanapoli (?cido acr?lico) (PNIPAM-CS-PAA). A obten??o dos materiais foi realizada por polimeriza??o por precipita??o, via radicais livres, utilizando persulfato de pot?ssio como iniciador e N,N -metilenobisacrilamida (MBA) como agente reticulante. A incorpora??o dos precursores nas part?culas reticuladas foi confirmada por an?lise elementar e espectroscopia na regi?o do infravermelho. A influ?ncia de est?mulos externos, como pH, temperatura, ou ambos, no comportamento das part?culas, foi investigado por medidas reol?gicas, estabilidade visual e centrifuga??o anal?tica. As part?culas de PNIPAM-QS apresentaram maior estabilidade em meios ?cido e neutro, enquanto que as part?culas de PNIPAM-QS-PAA foram mais est?veis em meios neutro e alcalino, tanto abaixo quanto acima da LCST da PNIPAM (dados de estabilidade). Isso ocorreu devido ?s diferentes intera??es interpart?culas e entre as part?culas e o meio (tamb?m evidenciadas pelos dados reol?gicos), que foi influenciada pelo pH e temperatura do meio. Baseando-se nos resultados obtidos, observou-se que a introdu??o de pol?meros sens?veis ao pH nas part?culas reticuladas de poli (N-isopropilacrilamida), resultou n?o apenas na s?ntese de copol?meros reticulados com dupla sensibilidade, mas em part?culas pass?veis de ajuste de estabilidade, ou seja, que podem sofrer separa??o de fases mais ou menos r?pida, dependendo da aplica??o desejada. Dessa forma, ? poss?vel adaptar as part?culas sintetizadas aos mais diversos meios