Reproduction de : Thèse doctorat : Pharmacie. Biomolécules et dynamique cellulaire : Reims : 2006. / Titre provenant de l'écran-titre. Bibliogr. p.137-172.

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No description available.

Diss. (sammanfattning) Stockholm : Karol. inst., 2003. / Härtill 5 uppsatser.

Cette thèse concerne le développement et l'évaluation des nanoparticules pour le traitement du cancer et plus particulièrement pour le cancer de la prostate.Le manuscrit comprend une revue de la littérature sur l'application de la nano médecine pour le traitement du cancer de la prostate. Dans la première partie expérimentale, des nanoparticules d'or fonctionnalisées ont été caractérisées et chargées avec le docétaxel par adsorption non covalente.Ces nanoparticules d'or ont montré un effet cytotoxique in vitro prolongé contre les cellules cancéreuses de la prostate. La deuxième partie expérimentale de cette thèse décrit une étude de synthèse et une nano-précipitation de polyesters pour la co-délivrance de deux médicaments chimio-thérapeutiques, le docétaxel (DOC) et la mitoxantrone (MIT). Les polycaprolactone, poly(acide lactique) et poly(lactide-co-glycolide) ont été synthétisés par polymérisation par ouverture de cycle avec des poids moléculaires différents de polyéthylène glycol. Des nanoparticules monodisperses d’un diamètre d’environ 80 nm ont été obtenues et se sont avérées être efficaces contre les cellules cancéreuses de la prostate quand cela est chargé en MIT et DOC. De plus, un effet synergique a été observé en utilisant des combinaisons de ces nanoparticules. Par conséquent, ces nanoparticules, à base de polyester, ont de potentielles applications cliniques. / This thesis concerns the development and evaluation of nanoparticles for cancer treatment, and in particular to prostate cancer. The manuscript includes a literature review on the application of nanomedicine to the treatment of prostate cancer. In the first experimental part, functionalized gold nanoparticles were characterized and loaded with docetaxel by non covalent adsorption. These gold nanoparticles showed a sustained cytotoxic effect in vitro against prostate cancer cells. The second experimental part of this thesis describes a study of synthesis and nanoprecipitation of polyesters for the co-delivery of two chemotherapeutic drugs, docetaxel (DOC) and mitoxantrone (MIT). Polycaprolactone, poly(lactic acid) and poly (lactide-co-glycolide) were synthesized by ring-opening polymerization with different molecular weights of polyethylene glycol. Monodisperse nanoparticles with diameters of less than 80 nm were produced and were shown to be effective against prostate cancer cells when loaded with MIT and DOC. Moreover, a synergistic effect was observed using combinations of these nanoparticles. Therefore, these polyester based nanoparticles have potential clinical applications.

A mitoxantrona (MXT) é um antineoplásico utilizado no tratamento de tumores como leucemias, linfoma não-Hodgkin e câncer de mama e próstata. Ela é classificada como uma antracenodiona, sendo um análogo estrutural das antraciclinas, como a doxorrubicina (DOX), cujo mecanismo de ação é baseado na inibição da enzima topoisomerase II (Topo II), através da formação dos complexos estabilizados Topo II-DNA. As antraciclinas e a MXT também são capazes de formar lesões do tipo adutos, pontes intercadeias de DNA (interstrand crosslink – ICL) e espécies reativas de oxigênio (ERO). Estudos têm demonstrado que a via de reparo por excisão de nucleotídeos (Nucleotide Excision Repair – NER) está envolvido na remoção de lesões no DNA induzidas pela DOX. Considerando as similaridades estruturais e de mecanismo de ação entre a MXT e a DOX, o objetivo deste trabalho foi avaliar a influência da via NER na citotoxicidade da MXT, a fim de elucidar possíveis mecanismos envolvidos na resistência tumoral a esta droga. Os resultados encontrados demonstraram que células deficientes na via NER (XPA, XPD, XPC e CSB) apresentam elevada sensibilidade a MXT comparadas a células proficientes em reparo (MRC5). Apesar disso, células CSB (deficientes na subvia associada à transcrição - Transcription coupled – TCR-NER) são mais sensíveis a MXT que células XPC (deficientes na subvia de reparo global do genoma – Global genome repair – GGR-NER) e também apresentam diferenças no perfil de ciclo celular, síntese de DNA e formação dos complexos Topo II-DNA após tratamento com MXT. Células XPC, da mesma forma que as células proficientes MRC5 apresentam parada de ciclo celular em G2/M, recuperação da síntese de DNA e sinal semelhante para formação dos complexos Topo II-DNA, enquanto células CSB apresentam acúmulo de células na fase S, diminuição na síntese de DNA e sinal mais intenso para formação dos complexos Topo II-DNA. Além disso, a complementação das células CSB com a proteína CSB recuperou a resistência das células a MXT e também diminuiu a intensidade do sinal dos complexos Topo II-DNA. Estes resultados indicaram que a via NER está envolvida na resistência das células ao tratamento com MXT e que a proteína CSB ou a subvia TCR-NER tem um papel chave no processamento dos complexos Topo II-DNA. / Mitoxantrone (MXT) is an antineoplastic drug used in treatment of tumors like leukemia, non-Hodgkin lymphoma and breast and prostate cancer. It is classified as an anthracenedione, being a structural analogue of anthracyclines, like doxorubicin (DOX), which action mechanism is based on topoisomerase II (Topo II) inhibition and formation of stabilized Topo II-DNA complexes. Anthracyclines and MXT also can form lesions like DNA adducts, interstrand crosslinks (ICL) and reactive oxygen species (ROS). Studies have shown that nucleotide excision repair (NER) pathway is involved in removal of lesions induced by DOX. Due to structural and action mechanism similarities between MXT and DOX, the aim of this work was to evaluate the influence of NER pathway in cytotoxicity of MXT, in order to elucidate possible mechanisms involved in tumor resistance to this drug. The results demonstrated that NER-deficient cells (XPA, XPD, XPC and CSB) show high sensitivity to MXT compared to repair proficient cells (MRC5). However, CSB cells (deficient in Transcription coupled repair – TCR) were more sensitive to MXT than XPC cells (deficient in Global genome repair – GGR) and also showed differences in cell cycle, DNA synthesis and Topo II-DNA complexes formation upon MXT treatment. XPC cells, in the same way as MRC5 proficient cells present G2/M cell cycle arrest, DNA synthesis recovery and similar signal for Topo II-DNA complexes formation, while CSB cells present accumulation of cells in S phase, reduced DNA synthesis and a more intense signal for Topo II-DNA complexes formation. Moreover, CSB cells complementation recovery MXT-resistance and also diminished Topo II-DNA complexes signal intensity. These results indicate that NER pathway is involved in cells resistance to MXT treatment and that CSB protein or TCR-NER sub pathway has a key role in processing of MXT induced Topo II-DNA complexes.

The focus of this investigation is about DNA topoisomerases, the molecular targets of clinically important chemotherapy, and mechanisms of drug resistance in human myeloma and leukemia cell lines. The ultimate goal of this investigation was to identify mechanism(s) of drug resistance to anticancer agents so that a strategy to overcome drug resistance could be conceived. We established an in vitro cell model by using human leukemia and myeloma cell lines to investigate possible mechanisms of drug resistance that are observed in confluent cells. Plateau cell densities demonstrated de novo drug resistance to commonly used chemotherapeutic agents that was independent of altered drug transport. We established that cellular drug resistance in these cells is a function of topo IIalpha subcellular localization and further demonstrate that topo IIalpha translocates to the cytoplasm in a cell-density dependent manner. We provide experimental data that supports the nuclear export of topo IIalpha as the most likely event contributing to drug resistance to topoisomerase II inhibitors, which occurs when transformed cells transition from log to plateau cell density. We provided a plausible nuclear export pathway for topo IIalpha, by identifying two Leptomycin B sensitive nuclear export signals, which are homologous to the binding sites recognized by the nuclear export receptor, exportin-1. Thus, topo IIalpha is likely to be exported from the nucleus at plateau cell densities when exportin-1 binds topo IIalpha. We confirmed that the nuclear export signals identified in topo IIalpha are functional when expressed in human myeloma cells transfected with an epitope-tagged topo IIalpha gene. Furthermore we demonstrate that the nuclear export signals can be abolished by site-directed mutagenesis of specific amino acids residues found in the nuclear export signal. Our data may have clinical relevance because plasma cells obtained from bone marrow aspirates of patients with multiple myeloma contain a cytoplasmic distribution of topo IIalpha. The potential implications of a functioning nuclear enzyme located in the cytoplasm of cells and theoretical mechanisms for overcoming the observed drug resistance are considered.

A mitoxantrona (MXT) é um antineoplásico utilizado no tratamento de tumores como leucemias, linfoma não-Hodgkin e câncer de mama e próstata. Ela é classificada como uma antracenodiona, sendo um análogo estrutural das antraciclinas, como a doxorrubicina (DOX), cujo mecanismo de ação é baseado na inibição da enzima topoisomerase II (Topo II), através da formação dos complexos estabilizados Topo II-DNA. As antraciclinas e a MXT também são capazes de formar lesões do tipo adutos, pontes intercadeias de DNA (interstrand crosslink – ICL) e espécies reativas de oxigênio (ERO). Estudos têm demonstrado que a via de reparo por excisão de nucleotídeos (Nucleotide Excision Repair – NER) está envolvido na remoção de lesões no DNA induzidas pela DOX. Considerando as similaridades estruturais e de mecanismo de ação entre a MXT e a DOX, o objetivo deste trabalho foi avaliar a influência da via NER na citotoxicidade da MXT, a fim de elucidar possíveis mecanismos envolvidos na resistência tumoral a esta droga. Os resultados encontrados demonstraram que células deficientes na via NER (XPA, XPD, XPC e CSB) apresentam elevada sensibilidade a MXT comparadas a células proficientes em reparo (MRC5). Apesar disso, células CSB (deficientes na subvia associada à transcrição - Transcription coupled – TCR-NER) são mais sensíveis a MXT que células XPC (deficientes na subvia de reparo global do genoma – Global genome repair – GGR-NER) e também apresentam diferenças no perfil de ciclo celular, síntese de DNA e formação dos complexos Topo II-DNA após tratamento com MXT. Células XPC, da mesma forma que as células proficientes MRC5 apresentam parada de ciclo celular em G2/M, recuperação da síntese de DNA e sinal semelhante para formação dos complexos Topo II-DNA, enquanto células CSB apresentam acúmulo de células na fase S, diminuição na síntese de DNA e sinal mais intenso para formação dos complexos Topo II-DNA. Além disso, a complementação das células CSB com a proteína CSB recuperou a resistência das células a MXT e também diminuiu a intensidade do sinal dos complexos Topo II-DNA. Estes resultados indicaram que a via NER está envolvida na resistência das células ao tratamento com MXT e que a proteína CSB ou a subvia TCR-NER tem um papel chave no processamento dos complexos Topo II-DNA. / Mitoxantrone (MXT) is an antineoplastic drug used in treatment of tumors like leukemia, non-Hodgkin lymphoma and breast and prostate cancer. It is classified as an anthracenedione, being a structural analogue of anthracyclines, like doxorubicin (DOX), which action mechanism is based on topoisomerase II (Topo II) inhibition and formation of stabilized Topo II-DNA complexes. Anthracyclines and MXT also can form lesions like DNA adducts, interstrand crosslinks (ICL) and reactive oxygen species (ROS). Studies have shown that nucleotide excision repair (NER) pathway is involved in removal of lesions induced by DOX. Due to structural and action mechanism similarities between MXT and DOX, the aim of this work was to evaluate the influence of NER pathway in cytotoxicity of MXT, in order to elucidate possible mechanisms involved in tumor resistance to this drug. The results demonstrated that NER-deficient cells (XPA, XPD, XPC and CSB) show high sensitivity to MXT compared to repair proficient cells (MRC5). However, CSB cells (deficient in Transcription coupled repair – TCR) were more sensitive to MXT than XPC cells (deficient in Global genome repair – GGR) and also showed differences in cell cycle, DNA synthesis and Topo II-DNA complexes formation upon MXT treatment. XPC cells, in the same way as MRC5 proficient cells present G2/M cell cycle arrest, DNA synthesis recovery and similar signal for Topo II-DNA complexes formation, while CSB cells present accumulation of cells in S phase, reduced DNA synthesis and a more intense signal for Topo II-DNA complexes formation. Moreover, CSB cells complementation recovery MXT-resistance and also diminished Topo II-DNA complexes signal intensity. These results indicate that NER pathway is involved in cells resistance to MXT treatment and that CSB protein or TCR-NER sub pathway has a key role in processing of MXT induced Topo II-DNA complexes.

A mitoxantrona (MXT) é um antineoplásico utilizado no tratamento de tumores como leucemias, linfoma não-Hodgkin e câncer de mama e próstata. Ela é classificada como uma antracenodiona, sendo um análogo estrutural das antraciclinas, como a doxorrubicina (DOX), cujo mecanismo de ação é baseado na inibição da enzima topoisomerase II (Topo II), através da formação dos complexos estabilizados Topo II-DNA. As antraciclinas e a MXT também são capazes de formar lesões do tipo adutos, pontes intercadeias de DNA (interstrand crosslink – ICL) e espécies reativas de oxigênio (ERO). Estudos têm demonstrado que a via de reparo por excisão de nucleotídeos (Nucleotide Excision Repair – NER) está envolvido na remoção de lesões no DNA induzidas pela DOX. Considerando as similaridades estruturais e de mecanismo de ação entre a MXT e a DOX, o objetivo deste trabalho foi avaliar a influência da via NER na citotoxicidade da MXT, a fim de elucidar possíveis mecanismos envolvidos na resistência tumoral a esta droga. Os resultados encontrados demonstraram que células deficientes na via NER (XPA, XPD, XPC e CSB) apresentam elevada sensibilidade a MXT comparadas a células proficientes em reparo (MRC5). Apesar disso, células CSB (deficientes na subvia associada à transcrição - Transcription coupled – TCR-NER) são mais sensíveis a MXT que células XPC (deficientes na subvia de reparo global do genoma – Global genome repair – GGR-NER) e também apresentam diferenças no perfil de ciclo celular, síntese de DNA e formação dos complexos Topo II-DNA após tratamento com MXT. Células XPC, da mesma forma que as células proficientes MRC5 apresentam parada de ciclo celular em G2/M, recuperação da síntese de DNA e sinal semelhante para formação dos complexos Topo II-DNA, enquanto células CSB apresentam acúmulo de células na fase S, diminuição na síntese de DNA e sinal mais intenso para formação dos complexos Topo II-DNA. Além disso, a complementação das células CSB com a proteína CSB recuperou a resistência das células a MXT e também diminuiu a intensidade do sinal dos complexos Topo II-DNA. Estes resultados indicaram que a via NER está envolvida na resistência das células ao tratamento com MXT e que a proteína CSB ou a subvia TCR-NER tem um papel chave no processamento dos complexos Topo II-DNA. / Mitoxantrone (MXT) is an antineoplastic drug used in treatment of tumors like leukemia, non-Hodgkin lymphoma and breast and prostate cancer. It is classified as an anthracenedione, being a structural analogue of anthracyclines, like doxorubicin (DOX), which action mechanism is based on topoisomerase II (Topo II) inhibition and formation of stabilized Topo II-DNA complexes. Anthracyclines and MXT also can form lesions like DNA adducts, interstrand crosslinks (ICL) and reactive oxygen species (ROS). Studies have shown that nucleotide excision repair (NER) pathway is involved in removal of lesions induced by DOX. Due to structural and action mechanism similarities between MXT and DOX, the aim of this work was to evaluate the influence of NER pathway in cytotoxicity of MXT, in order to elucidate possible mechanisms involved in tumor resistance to this drug. The results demonstrated that NER-deficient cells (XPA, XPD, XPC and CSB) show high sensitivity to MXT compared to repair proficient cells (MRC5). However, CSB cells (deficient in Transcription coupled repair – TCR) were more sensitive to MXT than XPC cells (deficient in Global genome repair – GGR) and also showed differences in cell cycle, DNA synthesis and Topo II-DNA complexes formation upon MXT treatment. XPC cells, in the same way as MRC5 proficient cells present G2/M cell cycle arrest, DNA synthesis recovery and similar signal for Topo II-DNA complexes formation, while CSB cells present accumulation of cells in S phase, reduced DNA synthesis and a more intense signal for Topo II-DNA complexes formation. Moreover, CSB cells complementation recovery MXT-resistance and also diminished Topo II-DNA complexes signal intensity. These results indicate that NER pathway is involved in cells resistance to MXT treatment and that CSB protein or TCR-NER sub pathway has a key role in processing of MXT induced Topo II-DNA complexes.

abstract: Cancer is the second leading cause of death in the United States and novel methods of treating advanced malignancies are of high importance. Of these deaths, prostate cancer and breast cancer are the second most fatal carcinomas in men and women respectively, while pancreatic cancer is the fourth most fatal in both men and women. Developing new drugs for the treatment of cancer is both a slow and expensive process. It is estimated that it takes an average of 15 years and an expense of $800 million to bring a single new drug to the market. However, it is also estimated that nearly 40% of that cost could be avoided by finding alternative uses for drugs that have already been approved by the Food and Drug Administration (FDA). The research presented in this document describes the testing, identification, and mechanistic evaluation of novel methods for treating many human carcinomas using drugs previously approved by the FDA. A tissue culture plate-based screening of FDA approved drugs will identify compounds that can be used in combination with the protein TRAIL to induce apoptosis selectively in cancer cells. Identified leads will next be optimized using high-throughput microfluidic devices to determine the most effective treatment conditions. Finally, a rigorous mechanistic analysis will be conducted to understand how the FDA-approved drug mitoxantrone, sensitizes cancer cells to TRAIL-mediated apoptosis. / Dissertation/Thesis / Ph.D. Chemical Engineering 2013