Most existing chemotherapeutics lack adequate specificity for transformed cells and therefore have high rates of collateral damage to normal tissue. Moreover, such therapies often depend on p53 to induce cell death and are ineffective on the large number of human cancers that have lost p53 function. The discovery of novel p53-independent cancer therapies is therefore of significant interest. The Chicken Anemia Virus protein Apoptin selectively induces apoptosis in transformed cells in a p53-independent manner while leaving normal primary cells unaffected. This selectivity is thought to be largely due to cell type-specific localization: in primary cells Apoptin is cytoplasmic, whereas in transformed cells the protein localizes to the nucleus. The basis for this cell type-specific localization remains to be determined. In this study, Apoptin is revealed to be a nucleo-cytoplasmic shuttling protein whose localization is mediated by an N-terminal nuclear export signal (NES) and a C-terminal nuclear localization signal (NLS). Both signals are required for cell type-specific localization, as Apoptin fragments containing either the NES or NLS fail to localize differently between transformed and primary cells. Significantly, cell type-specific localization can be rescued in trans by co-expression of the two separate fragments, which are able to interact through an Apoptin multimerization domain. Interestingly, this multimerization domain overlaps with the NES suggesting that these two activities may be functionally coupled in cytoplasmic retention in primary cell types. Factors present in transformed cells induce localization of Apoptin to the nucleus where a biochemically distinct, more soluble form of the protein exists. Using affinity-purification and mass spectroscopy it was found that, specifically in transformed cells, Apoptin is associated with APC1, a subunit of the anaphase-promoting complex/cyclosome (APC/C). The APC/C is required to establish a mitotic cell-cycle checkpoint, and its inhibition results in G2/M arrest and apoptosis. Expression of wild type Apoptin in transformed cells inhibits APC/C function and induces G2/M arrest and apoptosis, whereas Apoptin mutants that are unable to associate with APC1 have no effect. In p53 null cells, ablation of APC1 by RNA interference induces a G2/M arrest and apoptosis analogous to that observed following Apoptin expression. Furthermore, Apoptin was found to induce the formation of PML bodies and to recruit APC/C subunits to these nuclear structures suggesting a mechanism involving sequestration and subsequent inhibition of the APC/C. Thus, the results of this study clarify Apoptin cell type-specific localization behavior and explain the ability of Apoptin to induce apoptosis in transformed cells in the absence of p53. This study advances a newly emerging field of viral mechanisms of apoptosis involving G2/M arrest and APC/C modulation. The resultant p53-independent apoptosis suggests that the APC/C may be an attractive target for the development of anti-cancer drugs.

Made available in DSpace on 2015-07-22T20:50:31Z (GMT). No. of bitstreams: 0 Previous issue date: 2008-07-30. Added 1 bitstream(s) on 2015-08-11T03:25:42Z : No. of bitstreams: 1 Publico-10756.pdf: 198016 bytes, checksum: b1b248dd88de1aeab5859b81fa05e4c2 (MD5) / INTRODUÇÃO: Os antagonistas 5HT-3 e corticosteróides são drogas importantes no controle da emese induzida por quimioterapia Apresentam controle de 50-80% da emese aguda porém controlam menos de 50% da emese tardia induzida por quimioterapia. Em estudo prévio realizado em nossa instituição, observou-se que o controle da emese tardia poderia ser melhorado com a omissão do segundo dia dos antagonistas 5HT-3. Reportamos aqui os resultados de um estudo fase III confirmando os achados prévios PACIENTES E MÉTODOS: Estudo fase III randomizado e duplo-cego foi realizado na FMABC Santo André/Brasil. Pacientes submetidos a esquemas quimioterápicos alta e moderadamente emetogênicos pela 1ª vez receberam Ondasetron 16mg e Dexametasona 20 mg IV antes da quimioterapia no Dia 1. Foram então randomizados em dois grupos. Grupo A recebeu metoclopramida VO 10mg 8/8hs, Granisetron VO 0,5mg/dia,e dexametasona VO 8mg /dia por 2 dias, a partir do dia 2 (dias 2 e 3). Metoclopramida VO 10 mg 8/8 hs foi continuada no dia 4. Grupo B recebeu placebo no dia 2 e o mesmo esquema de drogas dia 3 e dia 4. Pacientes foram entrevistados no dia 1 e dia 6. RESULTADOS: 73 pacientes foram incluídos no estudo. Os grupos foram bem balanceados em relação as suas características clinicas com exceção ao melhor controle da emese aguda no grupo A (p=0,04). Proteção completa de náuse a e vômito tardios (do dia 2 ao 5 ), foi semelhante em ambos os grupos (30% vs. 32%; p=0.5). Em análise multivariada, tanto a proteção completa da emese aguda (p=0.001) quanto grupo de estudo (p=0.06) estavam independentemente relacionados à proteção completa da náusea e vômito tardios. Selecionando apenas os pacientes que obtiveram controle completo da emese na fase aguda, pacientes do grupo B obtiveram maior proteção da emese tardia (85% vs 50% ,p=0,02). CONCLUSÃO: A omissão da medicação antiemética no dia 2 representa uma estratégia para redução do custo da profilaxia da emese tardia induzida pela quimioterapia. / BACKGROUND: Nausea and vomiting are important symptoms observed in cancer patients. In a previous study we showed that delayed chemotherapyinduced nausea and vomiting (CINV) control could be potentially improved by skipping the administration of a 5HT3-antagonist on day 2. We report here a trial confirming our previous findings. PATIENTS/METHODS: A phase-IIIrandomized- placebo-controlled trial was conducted in which patients received (IV) ondansetron 16mg, dexamethasone 20mg and ranitidine 50mg before highly/moderately emetogenic chemotherapy (day 1).Starting on day 2, all patients received metoclopramide 10mg PO q8 hours (days 2,3 and 4, Dexamethasone 8mg QD (days 2 and 3) and Ranitidine 150mg q12 hours (days 2 and 3). Patients were randomized to receive either Granisetron 0.5mg PO (days 2 and 3) (Group A) or Placebo instead for Granisetron on day 2 and Granisetron 0.5mg on day 3 and 4 (Group B) RESULTS: 73 patients were enrolled. Groups were similar regarding clinical characteristics, despite better control during the acute phase of CINV in group A (p=0.04). Complete delayed protection from nausea/vomiting (DCPNV) from day 2 to 5 was similar in both groups. (30% vs. 32%; p=0.5). Analyzing DCPNV by logistic regression multivariate analyses, acute complete protection from nausea/vomiting (ACPNV) (p=0.001) and study group (p=0.06) were independently associated with DCPNV. Selecting patients who achieved ACPNV, we observed that group B had a superior DCPNV (85% vs 50%, p=0.02). CONCLUSION: DCPNV can be improved just by skipping day 2 of 5HT3- antagonists. Future studies should compare this inexpensive strategy with NK1- antagonists or second generation 5HT3-antagonists. Key words: delayed chemotherapy-induced nausea and vomiting, tachyphylaxis, cost- saving regimen, antiemetics, Granisetron/administration and dosage. / TEDE / BV UNIFESP: Teses e dissertações

Orientadores: João Ernesto de Carvalho, Mary Ann Foglio / Dissertação (mestrado) - Universidade Estadual de Campinas, Instituto de Biologia / Made available in DSpace on 2018-08-10T22:03:53Z (GMT). No. of bitstreams: 1 Marchetti_GabrielaMenezes_M.pdf: 1256255 bytes, checksum: 67243f8ebf02e8430f9c2bef8d87bd9a (MD5) Previous issue date: 2008 / Resumo: Durante muitos séculos, as plantas originaram um grande número de agentes terapêuticos, na qual seus compostos deram origem a medicamentos ou serviram de base para a síntese dos mesmos. Atualmente a biologia do câncer tem sido muito estudada e uma das principais linhas de pesquisas nesta área é o desenvolvimento de novos quimioterápicos. Partindo do princípio que plantas e drogas derivadas de plantas têm uma impressionante variedade de estruturas e funções, está claro que podem ser a fonte de novas drogas para a quimioterapia do câncer, pois, atualmente, mais de 60% delas são derivados de fontes naturais. Este trabalho teve como objetivo avaliar a atividade anticâncer de uma espécie do cerrado brasileira, Calea pinnatifida Banks. Os extratos brutos diclorometânico (EBD) e etanólico (EBE) das folhas frescas e secas dessa espécie foram avaliados em ensaio de citotoxicidade in vitro em cultura de células tumorais humanas. Como o EBD de folhas secas apresentou o melhor perfil de atividade in vitro, foi submetido a diversos procedimentos cromatográficos, sendo as frações obtidas biomonitoradas pelo teste de citotoxicidade. Esses procedimentos deram origem a diversas frações com atividade antiproliferativa com destaque para a fração acetato, fração A, C. D e neutra apolar. Com a finalidade de avaliar a biodisponibilidade do extrato este foi selecionado para avaliação em modelo de câncer murino, o tumor ascítico e sólido de Ehrlich. No modelo de tumor ascítico de Ehrlich, esse extrato apresentou atividade antitumoral através do aumento da sobrevida dos animais. Já no modelo de tumor sólido de Ehrlich, esse extrato apresentou atividade antitumoral sistêmica, através da inibição do crescimento tumoral. Com isso, a identificação dos princípios responsáveis por essa atividade tornou-se fundamental. Esses resultados estimulam a continuidade dos estudos com a C. pinnatifida, com o objetivo de identificar os princípios ativos, determinar o mecanismo de ação anticâncer e comprovar sua atividade em modelos experimentais de câncer in vivo / Abstract: Plants have provided a rich source of therapeutic agents for many centuries useful as themselves or the basis for synthetic drugs. Nowadays, the biology of cancer has been studied manly for the chemoprevention drug discovery. As plants and drugs based on plants have a lot of different structures and function they could be source of new drugs for cancer chemotherapy because more than 60% of the chemotherapics used today is from natural products. This work aimed the evaluation of the anticancer activity of a Brazilian specie, Calea pinnatifida Banks. Dichlorometanic (DCE) and ethanolic (ECE) crude extracts obtained from fresh and dried leaves were tested in an in vitro cytotoxicity assay against human cancer cell lines. As long as DCE from dried leaves showed the best anticancer activity profile, it was also evaluated in a murine cancer model, the Ehrlich Ascite Tumor (EAT) and Ehrlich Solid Tumor (EST). In the EAT experiment, DCE also demonstrated an anticancer activity resulted from the increase of the survival time of the animals. Whereas in the EST experiment, this extract have systemic anticancer activity by the inhibition of the tumor growth. Therefore the isolation and identification of the active principle responsible for that activity became the major focus of this work. As a result, DCE was submitted to many cromatographic procedures which were biomonitored by the anticancer assay in vitro. These chromatographic purifications originated a lot of fraction with antiproliferative activity in which the most significant are the acetate, A, C, D and non-polar fractions. These results encouraged following up on studies with C. pinnatifida, priorizing the identification of active principles, the determination of anticancer mechanism of action and to prove the anticancer activity in experimental cancer models in vivo / Mestrado / Biologia Celular / Mestre em Biologia Celular e Estrutural

O câncer de mama é a neoplasia com maior mortalidade entre as mulheres. O emprego de agentes alquilantes no tratamento desta neoplasia pode ocasionar o surgimento de instabilidades genômicas. Tais instabilidades podem estar associadas ao desenvolvimento de neoplasias secundárias como, por exemplo, leucemias. A presente tese avaliou a instabilidade de microssatélites em amostras de sangue, sedimento urinário e plasma de pacientes portadoras de carcinoma mamário ao diagnóstico, 3 e 6 meses após o início do tratamento quimioterápico. Também foi avaliada a concentração do DNA plasmático livre como possível marcador tumoral junto aos marcadores séricos CEA e CA15.3, empregados no acompanhamento do câncer de mama. Foram avaliadas as regiões de microssatélites: Tp53-ALU, Tp53.PCR15.1, BAT 40, BAT26, FMR2 e APC. Entre as 40 pacientes incluídas no presente estudo 88,57% apresentaram instabilidade de microssatélites na fração mononuclear do sangue periférico, 85,8% nas amostras de sedimento urinário e 62,5% no DNA plasmático livre. Não houve concordância significativa entre as instabilidades encontradas nos três tipos de amostra. A concentração de DNA plasmático livre das pacientes quando comparada às doadoras sadias apresentou correlação estatisticamente significativa (p>0,0001), e em paciente em regimes neoadjuvantes que responderam objetivamente à quimioterapia (p=0,02) e não houve correlação com os marcadores séricos CEA e CA15.3. / Breast cancer has the major mortality in women among all kind of cancer. The use of alkylating agents at the treatment of this disease is associated with genomic instability. This instability could be associated with the development of secondary cancer, for example, leukemia. The present thesis evaluated microsatellite instability in blood, pellet cells urinary and plasma in patients with breast cancer at diagnosis, 3 and 6 months after the beginning of chemotherapy. There were evaluated also Free Plasmatic DNA concentration as a possible tumoral marker with the serum markers CEA and CA15.3 used in breast cancer follow up. The microsatellites regions assayed were: Tp53-ALU,Tp53.PCR15.1, BAT 40, BAT26, FMR2 e APC. Among the 40 patients included at the present study 88,57% showed microsallite instability in peripheral mononuclear blood cells, 85,8% in urinary pellet cells samples and 62,5% in Free Plasmatic DNA. There werent statistical significant relationship for the instability found at the three kind of samples assayed. The Free Plasmatic DNA concentration of the patients when compared with healthy donors, showed a statistical significant relationship (p<0,0001). And among patients in neoadjuvant chemotherapy regime who reacted positively by treatment (p=0,02). And there werent statistical significant relationship between Free Plasmatic DNA and serum markers CEA and CA15.3.

Objetivo: Estimar a utilização de recursos e custos diretos relacionados à terapia endócrina (TE) versus quimioterapia (QT) no tratamento de câncer de mama avançado (CMA) receptores hormonais positivos (RH+) em pacientes pós menopáusicas,depois de pelo menos uma TE anterior. Métodos: Estudo longitudinal retrospectivo analisou pacientes pós menopáusicas com CMA em tratamento com fulvestranto ou QT entre 2006 e 2008, em um serviço público de oncologia ambulatorial. Apenas pacientes sem crise visceral e com pelo menos uma terapia anterior hormonal foram considerados elegíveis. Os prontuários foram revisados e as informações sobre diagnóstico, tratamento, e utilização de recursos foram obtidas. Resultados: As pacientes eram do sexo feminino e a idade média foi de 64,6 ± 12,6 anos. As pacientes estavam bem balanceados entre os grupos, considerando as características basais. Vinte e cinco pacientes foram incluídas no estudo, 13 pacientes receberam QT e 12 pacientes receberam fulvestranto. O esquema de QT mais usado foi o regime com paclitaxel (n = 5, 38%). O número médio de ciclos foi de 7,6 e 5,8 para fulvestranto e QT, respectivamente. O custo médio de tratamento por paciente foi de R$ 16.679 (USD11,914, 2005 índice de paridade de poder de compra 1USD = 1,4BRL) para fulvestranto e BRL 32946 (USD 23, 533) para QT. O custo médio por ciclo foi de R$ 2,199 (US$ 1,571) e BRL 5,710 (USD 4,079) para fulvestranto e QT, respectivamente, resultando em BRL 3,511(USD 2,508) de custo incremental por ciclo. Conclusões: Nossos resultados indicam que TE com fulvestranto pode ser economicamente adequada em pacientes com CMA RH + que falharam a pelo menos uma linha anterior de TE. Futuras pesquisas são necessárias para validar estes resultados em outros contextos, mas consideramos que as nossas estimativas refletem o mundo real da prática clínica no Brasil / OBJECTIVES: To estimate the resource utilization and costs related to endocrine therapy (ET) versus chemotherapy (CT) in the treatment of hormonal receptor positive (HR+), advanced breast cancer (ABC) patients, after at least one previous ET. METHODS: This retrospective longitudinal study analyzed ABC patients treatment with fulvestrant or CT between 2006 and 2008 in a public oncology outpatient service. Only patients without visceral crisis and with at least one previous hormonal therapy were considered eligible. Medical charts were reviewed by two investigators and information about diagnosis, course of treatment, and resource utilization was obtained. RESULTS: Patients were all female and the mean age was 64,6 ± 12,6 years. Patients were well matched between groups considering baseline characteristics. Twenty-five patients were enrolled in the study, 13 patients received CT and 12 patients received fulvestrant. The most common CT regimen was paclitaxel (n = 5, 38%). The mean number of cycles was 7,6 and 5,8 for fulvestrant and CT, respectively. The mean treatment cost per patient was BRL 16,679 (USD 11,914; 2005 purchasing power parity index 1USD = 1.4BRL) for fulvestrant and BRL 32,946 (USD 23,533) for CT. The mean cost per cycle was BRL 2,199 (USD 1,571) and BRL 5,710 (USD 4,079) for fulvestrant and CT, respectively, resulting in BRL 3,511 (USD 2,508) incremental cost per cycle. CONCLUSIONS: Our study results indicate that subsequent ET with fulvestrant can be economically appropriate among HR+ ABC patients. Further researches could validate these findings in other contexts, but we consider that our estimations reflect the real world clinical practice in Brazil

Submitted by Renata Lopes (renatasil82@gmail.com) on 2017-01-06T12:38:24Z No. of bitstreams: 1 julianabarrosorodriguesguedes.pdf: 1939234 bytes, checksum: ffb86b0f1beeff1e7f0b74faee06917a (MD5) / Approved for entry into archive by Diamantino Mayra (mayra.diamantino@ufjf.edu.br) on 2017-01-31T11:28:52Z (GMT) No. of bitstreams: 1 julianabarrosorodriguesguedes.pdf: 1939234 bytes, checksum: ffb86b0f1beeff1e7f0b74faee06917a (MD5) / Made available in DSpace on 2017-01-31T11:28:53Z (GMT). No. of bitstreams: 1 julianabarrosorodriguesguedes.pdf: 1939234 bytes, checksum: ffb86b0f1beeff1e7f0b74faee06917a (MD5) Previous issue date: 2016-03-30 / FAPEMIG - Fundação de Amparo à Pesquisa do Estado de Minas Gerais / A hormonioterapia no câncer de mama é fundamental para a transição do tratamento ativo aos cuidados de sobrevivência, pois melhora significativamente os resultados de sobrevida a longo prazo, além de propiciar uma melhor qualidade de vida aos doentes e de reduzir os custos de hospitalização. Porém, para se atingir resultados desejáveis no enfretamento da doença é importante a adesão e persistência ao tratamento recomendado. Sabe-se que a a adesão e persistência à hormonioterapia podem ser influenciadas por diversos fatores, relacionados ao paciente, tratamento, serviços de saúde, crenças e hábitos de vida, porém têm sido pouco explorados até à data. A condução de pesquisas capazes de dimensionar tais fenômenos e retratá-los em um contexto definido representa uma importante estratégia para comparação, compreensão e produção de evidências para os serviços de saúde, a fim de promover o sucesso da terapia empregada e a melhora da qualidade de vida do paciente. O objetivo desse estudo foi avaliar a adesão e a persistência à terapia hormonal adjuvante e suas variáveis associadas, em pacientes com câncer de mama em tratamento em um hospital de referência em oncologia da região sudeste do Brasil. Realizou-se um estudo de caráter analítico e delineamento longitudinal, a partir da coleta de dados secundários de 182 mulheres com câncer de mama, que iniciaram a hormonioterapia no ano de 2009 no Hospital do Câncer de Muriaé – MG/Fundação Cristiano Varella. As informações pertinentes ao estudo (variáveis dependentes e independentes) foram coletadas a partir do prontuário médico e do controle de dispensação de hormonioterápico, do setor da farmácia do Hospital do Câncer de Muriaé – MG/Fundação Cristiano Varella. Para o cálculo da adesão e persistência foram utilizados os métodos “Razão de Posse de Medicamentos” e “Nível Estimado de Persistência”, respectivamente. Foram consideradas aderentes ao tratamento mulheres com Razão de Posse de Medicamentos igual ou superior à 80% e persistentes àquelas que não descontinuaram o tratamento por 60 dias ou mais. Para analisar os fatores associados à adesão, foram realizadas análises bivariadas, utilizando o teste estatístico do qui-quadrado (χ2), já a probabilidade de persistência foi calculada através do método de Kaplan Meier e as variáveis associadas pelo teste de Log-rank. Para estimar o nível de significância das variáveis encontradas para a persistência, foram conduzidas análises de regressão linear múltipla e modelos de regressão uni e multivariada de Cox. O método "backward elimination" foi utilizado, removendo as variáveis que não se mantiveram significativas (p ≤ 0,05) e o teste diagnóstico de resíduos de Schoenfeld para avaliar a proporcionalidade dos modelos de Cox. A taxa de adesão entre as participantes foi de 85,2% no final do seguimento, destas 47,7% eram persistentes. Já a persistência foi modificada ao longo do tempo, sendo de 85,5% ao primeiro ano de tratamento e 43,6% no final dos 5 anos. Os resultados da análise bivariada para associação entre a taxa de adesão e variáveis independentes estudadas não se mostrou significativa para nenhuma delas. Na análise da associação dessas variáveis com a persistência, mulheres com estadiamento avançado ao diagnóstico, que não realizaram cirurgia e com um número elevado de internações exibiram maior risco de descontinuidade da hormonioterapia. Observa-se que apesar da adesão nesta coorte se apresentar elevada, à medida que o tempo avança há um aumento progressivo de pacientes não persistentes ao tratamento, muitas vezes associada às características relaciondas à gravidade da doença, aumentando o risco de resposta terapêutica inadequada e piores resultados referentes à taxa de sobrevida e recorrência da doença nessas mulheres. / Hormonetherapy in breast cancer is key to the transition from active treatment to survival of care, because it improves the long-term survival results, as well as providing a better quality of life for patients and reduce hospital costs. However, to achieve desirable results in the coping of the disease is important adherence and persistence to the recommended treatment. It is known that the adherence and persistence to hormonetherapy may be influenced by several factors related to the patient, treatment, health services, beliefs and lifestyle, but have been little explored to date. The conduct of research able to scale these phenomena and portray them in a defined context is an important strategy for comparison, comprehension and production of evidence for health services in order to promote the success of therapy used and improved quality life of the patient. The aim of this study was to evaluate the adherence and persistence adjuvant hormonal therapy and its associated variables in patients with treatment for breast cancer in a referral hospital in oncology of southeastern Brazil. We conducted an analytical study of character and longitudinal design, from the collection of secondary data from 182 women with breast cancer who started hormone therapy in 2009 at the Hospital of the Muriaé Cancer - MG/Fundação Cristiano Varella, following for 5 years. Information relevant to the study (dependent and independent variables) were collected from medical records and hormonetherapy of dispensing control, the Hospital of the Muriaé Cancer - MG/Fundação Cristiano Varella pharmacy sector. For the calculation of adherence and persistence were used methods "Medication Possession Ratio" and "Estimated level of Persistence", respectively. Were considered adherent to the treatment women with Medication Possession Ratio greater than or equal to 80% and persistent those who did not discontinue for 60 days or more. To analyze the factors associated with adherence, bivariate analyzes were performed using the statistical test Chi-square (χ2). The probability of persistence was calculated using the Kaplan Meier compared by log-rank test. To estimate hazard ratios and the level of significance of the variables associated with persistence, univariate and multivariate Cox regression analyzes were conducted. The method "backward elimination" was used by removing the variables that did not remain significant in the final model (p ≤ 0.05) and the diagnosis of Schoenfeld residuals test to assess the proportionality of the Cox models. The adherence rate among participants was 85.2% at the end of follow-up, and of these, 47.7% were persistent. The persistence was modified over time, with 85.5% in the first year of treatment and 43.6% at the end of the 5 years. The results of the bivariate analysis for association between adherence rate and independent variables studied was not significant for any of them. In the multivariate Cox model, women with advanced stage at diagnosis, who did not undergo surgery and with a high number of hospital admissions (≥ 3 hospitalizations) showed increased risk of discontinuation of hormone therapy. It is observed that despite the accession this cohort is considerable, as time progresses there is a progressive increase in non-persistent patients to treatment, influenced by characteristics related to the severity of the disease, increasing the risk of inadequate therapeutic response and worse results for the rate of survival and recurrence of the disease in these women.

15N-based nuclear magnetic resonance techniques are considered very powerful to study the molecular properties of platinum-containing anticancer agents, these properties being responsible for the efficacy of the compounds, but also for the understanding of resistance mechanisms and toxicity. Therefore, the first part of the present work aimed to develop a new method for synthesizing 15N-labeled, chiral platinum compounds. A theoretical discussion on the nucleophilic ring-opening of aziridines has also been envisaged, rationalizing an interesting regiochemistry question. Indeed, a surprising inversion of regiochemistry arose during the development of the above-mentioned synthetic pathway, and density functional theory calculation brought a rational framework to the experimental findings.<p><p>Infrared spectroscopy probes the global chemical composition of a sample and has been used to produce a snapshot of cancer cells contents after treatment with platinum coordinates. Indeed, in vitro studies focused here on the use of modern spectroscopic methods to fingerprint the cellular impact of platinum complexes. These drug signatures help to classify and select promising compounds. It makes no doubt that such systemic approaches for compound discovery are helpful technologies. Also, we made the use of the COMPARE algorithm from the NCI, which analyzes similarity between any active compounds previously tested by the NCI large scale in vitro screening program of anticancer agents. <p><p>The last chapter aimed to study the interactions between a series of platinum coordinates and DNA. Binding mode to telomeric-like sequences and binding kinetics to genomic-like sequences were assessed to investigate any differences between the compounds and to gain insight into structure-activity relationships. <p> / Doctorat en Sciences biomédicales et pharmaceutiques / info:eu-repo/semantics/nonPublished

La chimiothérapie conventionnelle est principalement constituée d’agents pro-apoptotiques. Or, de nombreux cancers sont intrinsèquement résistants à l’apoptose et cela explique en grande partie les mauvais pronostics associés à certains cancers, dont les cancers métastatiques, en raison de leur résistance aux chimiothérapies pro-apoptotiques. La thématique de ce travail s’inscrit dans la recherche de molécules présentant des modes d’actions différents afin de vaincre les mécanismes de résistance des cellules cancéreuses. <p><p>Une nouvelle série de composés di- et trivanilliques a ainsi été déclinée. La structure de ces composés est inspirée de molécules naturelles divanilliques étudiées pour leurs propriétés anti-falciformation dans la drépanocytose. Notre stratégie a été de transposer l’activité présumée de ces composés sur le cytosquelette des érythrocytes falciformes au niveau de la prolifération et de la migration des cellules cancéreuses. A cette fin, des méthodes pour analyser l’activité antitumorale de cette série de nouveaux composés ont été mises en place.<p><p>In vitro, les composés que nous qualifions d’actifs inhibent la croissance d’une dizaine de populations cellulaires cancéreuses distinctes à des valeurs inhibitrices de croissance à 50 % (indice IC50) de l’ordre de 30 µM. Les dérivés trivanilliques se classent parmi les composés les plus actifs de la série, en particulier le composé trivanillique chloré 13c qui exerce un effet cytostatique sur les lignées cellulaires cancéreuses analysées tout en présentant un certain taux de biosélectivité vis-à-vis des cellules normales. Nous avons ainsi consacré une partie de ce travail à approfondir la compréhension du mécanisme d’action du composé 13c et à le comparer à ses analogues de synthèse pour tenter d’établir une relation de type structure-activité.<p><p>Le dérivé 13c est cytostatique car il interfère avec le processus mitotique et il inhibe à plus de 75 % l’activité de 26 kinases à la concentration de 2 µM qui est donc plus de dix fois inférieure à la concentration inhibitrice IC50 moyenne de 30 µM rapportée ci-avant. Huit de ces 26 kinases sont directement impliquées dans l’organisation du cytosquelette d’actine et notamment dans les processus de cytokinèse. Parmi les kinases dont l’activité est la plus fortement modifiée par le composé 13c figurent les Aurora kinases qui sont de puissants contrôleurs de la cytokinèse. Nos résultats montrent également que l’activité cytostatique du composé 13c entraînant une désorganisation du cytosquelette d’actine pourrait être induite non seulement par l’inhibition de l’activité de certaines kinases mais aussi par des modifications induites par ce produit au niveau de l’homéostasie du calcium intracellulaire.<p><p>L’efficacité des agents chimiothérapeutiques est souvent altérée par une résistance tumorale intrinsèque appelée résistance multidrogue (le phénotype MDR). In vitro, le composé 13c s’est montré tout aussi efficace sur des lignées cancéreuses possédant cette résistance multidrogue que sur des cellules cancéreuses sensibles aux agents chimiothérapeutiques. De même, diverses lignées cellulaires résistantes aux stimuli pro-apoptotiques sont également sensibles à l’activité inhibitrice de croissance in vitro du trivanillate 13c. <p><p>Enfin, l’activité du composé 13c a été testée in vivo sur un modèle de pseudométastases pulmonaires issues du mélanome B16F10 murin. Les tumeurs pulmonaires présentent une cible intéressante pour le composé 13c au vu de son activité anti-tyrosine kinase comprenant la kinase EGFR et ses formes mutées EGFR-L858R et EGFR-T790M fréquemment retrouvées au sein des cancers du poumon non-à-petites-cellules (NSCLC). Le principe actif a été administré dans les poumons par inhalation via un dispositif endotrachéal. La voie inhalée est avantageuse par rapport à la voie systémique pour l’administration du produit 13c car cette molécule possède des liaisons esters qui sont hydrolysables par les estérases plus présentes au niveau des voies systémiques que dans les tissus pulmonaires. L’inhalation permet aussi de cibler mieux le site tumoral afin de réduire les dommages collatéraux. <p><p>Ce travail offre des perspectives au niveau de l’élaboration de formulations permettant d’augmenter la biodisponibilité de la molécule 13c. Une association d’un traitement au composé 13c et d’une chimiothérapie conventionnelle est envisagée car elle pourrait potentialiser les effets de ces deux traitements. <p> / Doctorat en Sciences biomédicales et pharmaceutiques / info:eu-repo/semantics/nonPublished

Le glioblastome est la plus fréquente et la plus agressive des tumeurs cérébrales primaires.<p>Malgré un traitement standard pluridisciplinaire (chirurgie, radiothérapie et chimiothérapie),<p>la médiane de survie des patients atteints de ce type de tumeur est de 15 mois et aucun patient<p>n’a pu être guéri à ce jour. Ce pronostic très sombre est directement lié à la capacité<p>d’invasion diffuse du parenchyme cérébral par les cellules gliales tumorales, ce qui rend<p>impossible une résection chirurgicale totale. De plus, ces cellules sont particulièrement<p>résistantes aux traitements chimiothérapeutiques de type pro-apoptotique, entrainant une<p>récidive quasi inévitable de la tumeur. De nombreuses stratégies thérapeutiques telles que<p>l’immunothérapie ou les thérapies ciblées sont actuellement explorées pour tenter de<p>combattre ces tumeurs. Cependant, leur efficacité au niveau clinique s’est avérée décevante. Il<p>devient donc indispensable d’identifier de nouveaux agents thérapeutiques afin d’améliorer la<p>survie des patients atteints de glioblastome.<p>Dans ce travail, nous avons caractérisé les propriétés anticancéreuses in vitro et in vivo de<p>deux terpénoïdes extraits de champignons, la fusicoccine A et l’ophioboline A, puis nous<p>avons caractérisé en partie leur mécanisme d’action anti-tumoral dans des cellules de<p>glioblastome.<p>Tout d’abord, nous avons montré que l’activité inhibitrice de croissance in vitro de la<p>fusicoccine A et de l’ophioboline A n’était pas dépendante du degré de sensibilité ou de<p>résistance à l’apoptose des cellules gliales tumorales. Nous avons ensuite mis en évidence que<p>la fusicoccine A était capable de diminuer l’invasion des cellules de glioblastome in vitro en<p>ciblant la kinase focale d’adhérence (FAK). Dans le même temps, nous avons démontré que<p>l’ophioboline A était capable d’induire la mort de ces cellules par paraptose en inhibant<p>l’activité du canal ionique BKCa. Ces deux cibles sont intéressantes car en plus d’être<p>surexprimées dans les glioblastomes, elles interviennent dans de nombreux processus<p>cellulaires tels que la prolifération, la migration et la survie cellulaire.<p>Pour finir, nous avons analysé le pouvoir anti-tumoral in vivo de ces deux terpénoïdes en<p>utilisant un modèle murin de mélanome métastatique, couramment utilisé dans notre<p>laboratoire. Seule l’ophioboline A, injectée en intrapéritonéal à 10 mg/kg, augmentait de<p>manière significative la survie des souris traitées avec cette molécule par rapport aux souris<p>contrôles. Dans ce premier modèle, nous n’avions pas déterminé les conditions optimales<p>9<p>pour l’évaluation in vivo de la fusicoccine A et de l’ophioboline A. Lorsque celles-ci seront<p>définies, des modèles de xénogreffes orthotopiques de glioblastomes humains implantées dans<p>le cerveau de souris immunodéficientes seront utilisés.<p>En conclusion, l’ophioboline A, pouvant être produite en quantités industrielles par culture<p>du champignon qui la synthétise, possédant un mécanisme d’action original et montrant déjà<p>un début d’activité in vivo, pourrait représenter une molécule méritant des études plus<p>approfondies en termes d’agent thérapeutique susceptible de combattre les glioblastomes. / Doctorat en Sciences biomédicales et pharmaceutiques / info:eu-repo/semantics/nonPublished

There is an urgent need to develop reliable and cost-saving methods to select pre-clinically new drug candidates with original mechanism for cancer therapy. Previous results have shown that IR spectra of cancer cells exposed to various drugs provided a global signature of all the metabolic changes induced by the treatments. In this thesis, we attempted to develop a selection criterion – based on FTIR spectroscopy – for potential antitumor compounds according to their mechanism of action. <p>In chapter III, it was demonstrated that spectral variations in IR spectra of cancer cells induced by a treatment can be correlated to the mechanism of the drug. Human prostate cancer PC-3 cells were exposed to 7 well-described anticancer drugs belonging to 3 distinct classes. Each class is characterized by a unique mode of action. Drugs known to induce similar types of metabolic disturbances appear to cluster when spectrum shapes are analyzed. Chapter IV generalized the results obtained on PC-3 cells with six other cell lines. We showed that the spectral signatures of drug effects are mainly independent of the cell line. Chapter V indicated that, while the cell cycle phase influence IR spectra of cells, the drug spectral signature was dominated by global metabolic modifications and not much by the cell cycle perturbations due to this drug. <p>Chapter VI and VII focused on lipids. While the precise identification of particular molecules is particularly complex with IR spectroscopy, we attempted to extract more precise information and to assign spectral variations to specific changes in lipids. IR spectra of lipids contain very interesting details on their nature and structure. We achieved to build a tool which quantifies five major lipid classes in complex mixtures such as total lipid cell extracts. However, based on this tool, the treatments used do not induce any variation in the lipid cell composition (for five classes).<p>Finally, in chapter VIII, we applied the method developed previously on a new potential class of anticancer molecules: the polyphenols. A global method was particularly interesting as the development of therapy using these compounds is hampered by the complexity of the multiple anticarcinogenic mechanisms of these molecules. We have noticed the similarities and discrepancies among 3 very close synthetic molecules and the observations were coherent with previous biological data. We also compared them with 3 natural molecules already in clinical phase for treatment of various cancers.<p>In conclusion, we developed an objective classifier for potential anticancer drugs based on their global effects on cancer cells. Applied to a larger scale, this method could constitute a first step in the screening method to select drugs with original mode of action.<p>There is an urgent need to develop reliable and cost-saving methods to select pre-clinically new drug candidates with original mechanism for cancer therapy. Previous results have shown that IR spectra of cancer cells exposed to various drugs provided a global signature of all the metabolic changes induced by the treatments. In this thesis, we attempted to develop a selection criterion – based on FTIR spectroscopy – for potential antitumor compounds according to their mechanism of action. <p>In chapter III, it was demonstrated that spectral variations in IR spectra of cancer cells induced by a treatment can be correlated to the mechanism of the drug. Human prostate cancer PC-3 cells were exposed to 7 well-described anticancer drugs belonging to 3 distinct classes. Each class is characterized by a unique mode of action. Drugs known to induce similar types of metabolic disturbances appear to cluster when spectrum shapes are analyzed. Chapter IV generalized the results obtained on PC-3 cells with six other cell lines. We showed that the spectral signatures of drug effects are mainly independent of the cell line. Chapter V indicated that, while the cell cycle phase influence IR spectra of cells, the drug spectral signature was dominated by global metabolic modifications and not much by the cell cycle perturbations due to this drug. <p>Chapter VI and VII focused on lipids. While the precise identification of particular molecules is particularly complex with IR spectroscopy, we attempted to extract more precise information and to assign spectral variations to specific changes in lipids. IR spectra of lipids contain very interesting details on their nature and structure. We achieved to build a tool which quantifies five major lipid classes in complex mixtures such as total lipid cell extracts. However, based on this tool, the treatments used do not induce any variation in the lipid cell composition (for five classes).<p>Finally, in chapter VIII, we applied the method developed previously on a new potential class of anticancer molecules: the polyphenols. A global method was particularly interesting as the development of therapy using these compounds is hampered by the complexity of the multiple anticarcinogenic mechanisms of these molecules. We have noticed the similarities and discrepancies among 3 very close synthetic molecules and the observations were coherent with previous biological data. We also compared them with 3 natural molecules already in clinical phase for treatment of various cancers.<p>In conclusion, we developed an objective classifier for potential anticancer drugs based on their global effects on cancer cells. Applied to a larger scale, this method could constitute a first step in the screening method to select drugs with original mode of action.<p><p>Afin d’améliorer les thérapies contre le cancer, il devient actuellement cruciale de développer une méthode pour améliorer la sélection préclinique de nouvelles molécules, potentiellement anticancéreuses. Des publications précédentes ont mis en évidence que les spectres infrarouges de cellules cancéreuses exposées à différents agents thérapeutiques fournissent une empreinte globale de l’ensemble des changements métaboliques induit par ce médicament. Dans cette thèse, nous proposons d’utiliser la spectroscopie infrarouge pour mettre au point un critère de sélection basé sur le mode d’action des agents anticancéreux. Plusieurs aspects de la technique ont été investigués. Nous avons d’abord démontré la possibilité d’utiliser les spectres infrarouges de cellules cancéreuses de prostate PC-3 traitées avec 7 drogues pour classer ces molécules selon leur mode d’action. Nous avons ensuite reproduit les résultats obtenus sur PC-3 avec 6 autres lignées cellulaires et montré que la signature spectrale obtenue était largement indépendante de la lignée. Par la suite, nous avons étudié si l’effet sur le cycle cellulaire induit par de nombreuses molécules anticancéreuses, pouvait expliquer certains changements spectraux observés suite au traitement. Nous avons pu montrer que la majorité des variations spectrales n’étaient pas liées à une perturbation du cycle cellulaire. Nous nous sommes ensuite concentrés sur une classe de molécules en particulier: les lipides. Après avoir mis en évidence l’ensemble des informations contenues dans un spectre infrarouge de lipides, nous avons mis au point un modèle permettant de quantifier 5 classes de lipides dans des mélanges complexes tels que des extraits lipidiques provenant de cellules. Néanmoins, aucune variation du contenu en ces 5 classes de lipides n’a été observée pour les traitements utilisés dans cette étude. Enfin, nous avons appliqué la méthode mise au point dans cette thèse à une classe de molécules prometteuses :les polyphénols. Une approche globale s’avère particulièrement intéressante pour ces composés étant donné qu’ils présentent des mécanismes anticancéreux multiples et complexes. Nous avons comparé 3 molécules naturelles en phase clinique pour le traitement de certains cancers et 3 molécules synthétiques présentant une structure très proche. Par notre méthode, nous avons mis en évidence certaines similarités et différences de ces 6 molécules en termes d’effets globaux sur les cellules. En conclusions, nous avons développé un outil objectif de classification pour les molécules anticancéreuses potentielles, basée sur le mécanisme global des composés. Appliquée à plus large échelle, cette méthode pourrait constituer une première étape permettant de sélectionner les molécules avec un mode d’action original. / Doctorat en Sciences agronomiques et ingénierie biologique / info:eu-repo/semantics/nonPublished