Um dos principais desafios no tratamento quimioterápico em seres humanos e animais é a resistência que as células neoplásicas apresentam, sendo esse mecanismo responsável por falhas no tratamento e recidivas da doença. A resistência pode ser intrínseca ou adquirida e ocorre em função da expressão de transportadores de membrana ABC, como a glicoproteína P (ABCB1/MDR), proteínas de resistência a múltiplas drogas (ABCC1/MRP) e proteína de resistência do câncer de mama (ABCG2/BCRP). O linfoma é a neoplasia hematopoiética mais comum em cães, altamente responsiva à quimioterapia, mas que recidiva durante o tratamento antineoplásico, sendo a resistência das células neoplásicas aos quimioterápicos um fator responsável pela alta taxa de recidiva e óbito dos animais. Neste estudo avaliou-se a expressão de genes relacionados à resistência a múltiplas drogas em cães com linfoma, no diagnóstico e na recidiva da doença, em três diferentes protocolos quimioterápicos utilizados na rotina clínica. A expressão dos genes ABCB1, ABCC1, ACBG2 foi determinada por RT-PCR (PCR em tempo real) em 25 animais naturalmente acometidos pela doença, divididos aleatoriamente em 3 grupos tratados com os protocolos quimioterápicos COP, VCM e Short-Madison, além de um "pool" controle constituído por linfonodos normais de oito animais. A expressão dos genes foi detectada em todas as amostras, tanto de linfonodos normais quanto de animais com linfoma. No diagnóstico da doença, a expressão do gene ABCC1 foi relacionada negativamente com idade (p=0,008) e positivamente com duração da remissão (p=0,027) e sobrevida (p=0,007), entretanto para os genes ABCB1 e ABCG2 não houve diferença estatística significante. Na recidiva, a expressão dos genes não sofreu variação estatística significante em função do tipo e duração da remissão e sobrevida. Não houve variação na expressão dos genes ABCB1, ACBC1 e ABCG2 no momento da recidiva quando comparado ao protocolo quimioterápico utilizado. / One of the main challenges of the chemotherapy treatment in human and animals is the resistance of the neoplasic cells, being this mechanism responsible for failures in the treatment and relapse of the disease. The resistance could be intrinsic or acquired and it occurs due to the expression of ABC membrane transporters, such as p-glycoprotein (ABCB1/MDR), resistance protein to multiple drugs (ABCC1/MRP) and resistance protein of breast cancer (ABCG2/BCRP). Lymphoma is the most common hematopoietic cancer disease in dogs, highly responsive to chemotherapy, but relapse during chemotherapy treatment, being the resistance of neoplastic cells to chemotherapy drugs the responsible factor for the high rate of relapse and death of animals. In this study, genes expression related to multiples drugs resistance it was evaluated in dogs with lymphoma, in the diagnosis and in the relapse of the disease in three different chemotherapy protocols used in the clinical routine. The genes expression ABCB1, ABCC1, ACBG2 was determined by RT-PCR (real time PCR) in 25 animals naturally undertaken by the illness, randomly divided into 3 groups treated with the chemotherapy protocols COP, VCM and Short-Madison, besides a "pool" control constituted by normal lymph node of eight animals. The genes expression was detected in all the samples, both in the normal lymph node and in the animals with lymphoma. In the diagnosis of the disease, the gene expression ABCC1 was negatively related with age (p=0,008) and positively with the duration of remission (p=0,027) and survival (p=0,007); however, for ABCB1 and ABCG2 there was no statiscally significant difference. In the relapse, the genes expression had no statiscally significant difference due to the type and duration of remission and survival. There was no variation in the genes expression ABCB1, ACBC1 and ABCG2 in the moment of relapse when compared to the chemotherapy protocol used.

Um dos principais desafios no tratamento quimioterápico em seres humanos e animais é a resistência que as células neoplásicas apresentam, sendo esse mecanismo responsável por falhas no tratamento e recidivas da doença. A resistência pode ser intrínseca ou adquirida e ocorre em função da expressão de transportadores de membrana ABC, como a glicoproteína P (ABCB1/MDR), proteínas de resistência a múltiplas drogas (ABCC1/MRP) e proteína de resistência do câncer de mama (ABCG2/BCRP). O linfoma é a neoplasia hematopoiética mais comum em cães, altamente responsiva à quimioterapia, mas que recidiva durante o tratamento antineoplásico, sendo a resistência das células neoplásicas aos quimioterápicos um fator responsável pela alta taxa de recidiva e óbito dos animais. Neste estudo avaliou-se a expressão de genes relacionados à resistência a múltiplas drogas em cães com linfoma, no diagnóstico e na recidiva da doença, em três diferentes protocolos quimioterápicos utilizados na rotina clínica. A expressão dos genes ABCB1, ABCC1, ACBG2 foi determinada por RT-PCR (PCR em tempo real) em 25 animais naturalmente acometidos pela doença, divididos aleatoriamente em 3 grupos tratados com os protocolos quimioterápicos COP, VCM e Short-Madison, além de um "pool" controle constituído por linfonodos normais de oito animais. A expressão dos genes foi detectada em todas as amostras, tanto de linfonodos normais quanto de animais com linfoma. No diagnóstico da doença, a expressão do gene ABCC1 foi relacionada negativamente com idade (p=0,008) e positivamente com duração da remissão (p=0,027) e sobrevida (p=0,007), entretanto para os genes ABCB1 e ABCG2 não houve diferença estatística significante. Na recidiva, a expressão dos genes não sofreu variação estatística significante em função do tipo e duração da remissão e sobrevida. Não houve variação na expressão dos genes ABCB1, ACBC1 e ABCG2 no momento da recidiva quando comparado ao protocolo quimioterápico utilizado. / One of the main challenges of the chemotherapy treatment in human and animals is the resistance of the neoplasic cells, being this mechanism responsible for failures in the treatment and relapse of the disease. The resistance could be intrinsic or acquired and it occurs due to the expression of ABC membrane transporters, such as p-glycoprotein (ABCB1/MDR), resistance protein to multiple drugs (ABCC1/MRP) and resistance protein of breast cancer (ABCG2/BCRP). Lymphoma is the most common hematopoietic cancer disease in dogs, highly responsive to chemotherapy, but relapse during chemotherapy treatment, being the resistance of neoplastic cells to chemotherapy drugs the responsible factor for the high rate of relapse and death of animals. In this study, genes expression related to multiples drugs resistance it was evaluated in dogs with lymphoma, in the diagnosis and in the relapse of the disease in three different chemotherapy protocols used in the clinical routine. The genes expression ABCB1, ABCC1, ACBG2 was determined by RT-PCR (real time PCR) in 25 animals naturally undertaken by the illness, randomly divided into 3 groups treated with the chemotherapy protocols COP, VCM and Short-Madison, besides a "pool" control constituted by normal lymph node of eight animals. The genes expression was detected in all the samples, both in the normal lymph node and in the animals with lymphoma. In the diagnosis of the disease, the gene expression ABCC1 was negatively related with age (p=0,008) and positively with the duration of remission (p=0,027) and survival (p=0,007); however, for ABCB1 and ABCG2 there was no statiscally significant difference. In the relapse, the genes expression had no statiscally significant difference due to the type and duration of remission and survival. There was no variation in the genes expression ABCB1, ACBC1 and ABCG2 in the moment of relapse when compared to the chemotherapy protocol used.

No description available.

Indiana University-Purdue University Indianapolis (IUPUI) / Triple-negative breast cancer (TNBC) is a highly aggressive form of breast cancer with a dismal prognosis. Targeted therapies for breast cancer with expression of estrogen receptor (ER), progesterone receptor (PR), and human epidermal growth factor receptor 2 (HER2) are currently available; however, due to the lack of ER, PR, and HER2 in TNBC, targeted therapies are limited. While surgery and traditional chemotherapy remain the standard of care, development of a new treatment strategy for TNBC is needed to improve clinical outcomes. Fatty acid synthase (FASN) has been implicated as a metabolic oncogene and has given cancer cells a survival advantage by increasing NHEJ repair. Recently, it has been shown that FDA-approved proton pump inhibitors, used for the treatment of acid related digestive diseases, have antitumor effects. Here, I show that a metabolite of lansoprazole, 5-hydroxy lansoprazole sulfide, has increased potency over parent compound lansoprazole. 5-hydroxy lansoprazole sulfide inhibits FASN, leading to a decrease in PARP and NHEJ DNA repair activity in TNBC. Ultimately, this leads to an increase in DNA damage and cell death via apoptosis. These findings suggest that 5-hydroxy lansoprazole sulfide, as a metabolite of lansoprazole, may have better activity in suppressing TNBC cells and that 5-hydroxy lansoprazole sulfide may be developed as a therapeutic for TNBC treatment. Furthermore, due to the role of FASN in increasing NHEJ repair, we hypothesized that FASN played a role in resistance to CC-115, a dual mTOR/DNA-PK inhibitor currently in clinical trials, by increasing DNA-PK activity. However, it was found that ABCG2, an ATP-binding cassette transporter, and not FASN, has a role in CC-115 resistance. ABCG2 effluxes CC-115 from cancer cells, increasing resistance to treatment. Inhibition of ABCG2 by FTC or PZ39C8 led to accumulation of CC-115 within cells and sensitization to treatment. Therefore, ABCG2 status should be assessed to stratify patients into treatment groups, increasing the efficacy of CC-115 treatment. / 2020-02-21

No description available.

A introdução do Mesilato de Imatinibe (MI) como primeiro inibidor específico de BCR-ABL1 na prática clínica revolucionou o tratamento da Leucemia Mieloide Crônica (LMC), tornando-se a terapia padrão para o tratamento desta doença. Porém, cerca de 30% dos pacientes com LMC não respondem à terapia com MI e um número substancial destes casos de resistência não tem causa conhecida. O MI interage com transportadores de membrana ABCG2 e SLCO1A2. Este estudo teve como objetivo investigar a relação da expressão gênica de ABCG2 e de SLCO1A2 com marcadores de resposta ao tratamento com MI, em indivíduos com LMC e avaliar a influência dos polimorfismos ABCG2 c.421C>A e ABCG2 c.-19-99G>A na resposta ao MI. Foram incluídos 118 pacientes com LMC os quais foram classificados em dois grupos: Grupo Respondedor, constituído por 70 pacientes com resposta citogenética completa com a dose padrão de MI (400 mg/dia) por até 18 meses e, Grupo não Respondedor constituído por 48 pacientes sem resposta citogenética completa à dose inicial de 400 mg/dia de MI ou que perderam esta resposta ao longo do tratamento e foram reescalonados para doses de 600 ou 800 mg/dia. A resposta ao tratamento foi avaliada segundo os critérios da European LeukemiaNet. Foram excluídos pacientes com alterações citogenéticas diferentes do cromossomo Ph e mutações no gene BCR-ABL1. Amostras de sangue periférico foram utilizadas para: extração do RNA total para quantificação dos transcritos BCR-ABL1 e expressão gênica de ABCG2 e SLCO1A2; extração de DNA e análise citogenética de banda G. A expressão do gene ABCG2 e SLCO1A2 e as análises dos polimorfismos foram feitas por PCR em tempo real. A expressão de ABCG2 foi maior no grupo de não respondedores ao MI (P=0,028). Este resultado foi influenciado pelos pacientes com resistência primária (N= 34 P=0,029), mas não pelos que apresentaram resistência secundária (N=14 P=0,249) quando comparado com respondedores (N=70). A elevada expressão do gene ABCG2 foi também associada àqueles pacientes que não tiveram resposta molecular maior (número de transcritos BCR-ABL1 ≤ 0,1%) (P=0,027) quando todos os pacientes foram analisados. O gene estudado não foi associado com a resposta molecular completa (número de transcritos BCR-ABL1 ≤ 0,032%). Com relação ao gene SLCO1A2 não foi possível determinar sua expressão devido à baixa concentração do RNA obtido. Os polimorfismos c.421C>A e c.-19-99G>A não foram associados com a expressão do gene ABCG2 e a resposta ao MI. A RMC (no grupo de respondedores) foi associada com o genótipo 421CC e houve tendência a maior frequencia de portadores do genótipo -19-99GG neste mesmo grupo. Portadores do genótipo -19-99AA apresentaram tendência ao risco de ter LMC. Os resultados deste estudo nos permitem concluir que a maior expressão de ABCG2 está associada com a resistência primária ao MI podendo então ser um mediador da resistência ao MI. Os polimorfismos do gene ABCG2 não influenciaram na expressão gênica de ABCG2, mas impactaram na RMC no grupo respondedor ao MI. / The introduction of imatinib mesylate (IM) as the first specific inhibitor of BCR-ABL1 in clinical practice has revolutionized the treatment of chronic myeloid leukemia (CML), becoming the standard therapy for this disease. However, about 20% of CML patients do not respond to therapy with IM and a substantial number of these cases of resistance have no known cause. The MI interacts with membrane transporters ABCG2 and SLCO1A2. The aim of this study was to investigate the relationship of ABCG2 and SLCO1A2 gene expression with markers of response to MI in individuals with CML and evaluate the influence of polymorphisms ABCG2 c.421C> A and c. ABCG2-19-99G> A in response to the MI. One hundred and eighteen patients in chronic phase of CML were studied and classified in two groups: Responder Group comprised 70 patients who had a complete cytogenetic response within 18 months of treatment. The non-responder group comprised 48 patients who did not have a complete cytogenetic response with the initial dose (400 mg/day) of IM or who relapsed during treatment and were submitted to higher doses of 600 or 800 mg/day. Criteria of failed response to treatment were established by European LeukemiaNet. Patients with cytogenetic patterns other than the Philadelphia chromosome and patients with mutations in the BCR-ABL1 gene were excluded from this study. Blood samples were obtained for: total RNA extraction for quantification of BCR-ABL1 and gene expression of ABCG2 and SLCO1A2; genomic DNA extraction and band G cytogenetic analysis. The gene expression and the analysis of the polymorphisms were performed by real time PCR. Expression of ABCG2 in non-responder group was higher than in responder group (P=0.028). This result was influenced by patients with primary resistance (n= 34 P=0.029) but not secondary resistance (n=14 P=0.249) when compared with responders (n=70). The higher expression of ABCG2 gene was also associated with those patients who had major molecular response (number of BCR-ABL1 . 0.1%) (P=0.027) when all patients were analyzed. The studied gene was not associated with the complete molecular response (number of BCR-ABL1 .0.0032). Regarding to the gene SLCO1A2 was not possible to determine its expression due to low concentration of RNA obtained. The c.421C>A e c.-19-99G>A were not associated neither with the ABCG2 gene expression and MI response. CMR in responders group was associated with the 421CC genotype ant there was a trend for higher frequency of carriers of genotype -19-99GG in the same group. Carriers of 19-99AA genotype tended to the risk of having CML. The results of this study allow us to conclude that the higher expression of ABCG2 is associated with primary resistance to IM and may be a mediator of resistance to IM. The ABCG2 polymorphisms did not influence the gene expression of ABCG2 but impacted in CMR of the responders to IM.

A introdução do Mesilato de Imatinibe (MI) como primeiro inibidor específico de BCR-ABL1 na prática clínica revolucionou o tratamento da Leucemia Mieloide Crônica (LMC), tornando-se a terapia padrão para o tratamento desta doença. Porém, cerca de 30% dos pacientes com LMC não respondem à terapia com MI e um número substancial destes casos de resistência não tem causa conhecida. O MI interage com transportadores de membrana ABCG2 e SLCO1A2. Este estudo teve como objetivo investigar a relação da expressão gênica de ABCG2 e de SLCO1A2 com marcadores de resposta ao tratamento com MI, em indivíduos com LMC e avaliar a influência dos polimorfismos ABCG2 c.421C>A e ABCG2 c.-19-99G>A na resposta ao MI. Foram incluídos 118 pacientes com LMC os quais foram classificados em dois grupos: Grupo Respondedor, constituído por 70 pacientes com resposta citogenética completa com a dose padrão de MI (400 mg/dia) por até 18 meses e, Grupo não Respondedor constituído por 48 pacientes sem resposta citogenética completa à dose inicial de 400 mg/dia de MI ou que perderam esta resposta ao longo do tratamento e foram reescalonados para doses de 600 ou 800 mg/dia. A resposta ao tratamento foi avaliada segundo os critérios da European LeukemiaNet. Foram excluídos pacientes com alterações citogenéticas diferentes do cromossomo Ph e mutações no gene BCR-ABL1. Amostras de sangue periférico foram utilizadas para: extração do RNA total para quantificação dos transcritos BCR-ABL1 e expressão gênica de ABCG2 e SLCO1A2; extração de DNA e análise citogenética de banda G. A expressão do gene ABCG2 e SLCO1A2 e as análises dos polimorfismos foram feitas por PCR em tempo real. A expressão de ABCG2 foi maior no grupo de não respondedores ao MI (P=0,028). Este resultado foi influenciado pelos pacientes com resistência primária (N= 34 P=0,029), mas não pelos que apresentaram resistência secundária (N=14 P=0,249) quando comparado com respondedores (N=70). A elevada expressão do gene ABCG2 foi também associada àqueles pacientes que não tiveram resposta molecular maior (número de transcritos BCR-ABL1 ≤ 0,1%) (P=0,027) quando todos os pacientes foram analisados. O gene estudado não foi associado com a resposta molecular completa (número de transcritos BCR-ABL1 ≤ 0,032%). Com relação ao gene SLCO1A2 não foi possível determinar sua expressão devido à baixa concentração do RNA obtido. Os polimorfismos c.421C>A e c.-19-99G>A não foram associados com a expressão do gene ABCG2 e a resposta ao MI. A RMC (no grupo de respondedores) foi associada com o genótipo 421CC e houve tendência a maior frequencia de portadores do genótipo -19-99GG neste mesmo grupo. Portadores do genótipo -19-99AA apresentaram tendência ao risco de ter LMC. Os resultados deste estudo nos permitem concluir que a maior expressão de ABCG2 está associada com a resistência primária ao MI podendo então ser um mediador da resistência ao MI. Os polimorfismos do gene ABCG2 não influenciaram na expressão gênica de ABCG2, mas impactaram na RMC no grupo respondedor ao MI. / The introduction of imatinib mesylate (IM) as the first specific inhibitor of BCR-ABL1 in clinical practice has revolutionized the treatment of chronic myeloid leukemia (CML), becoming the standard therapy for this disease. However, about 20% of CML patients do not respond to therapy with IM and a substantial number of these cases of resistance have no known cause. The MI interacts with membrane transporters ABCG2 and SLCO1A2. The aim of this study was to investigate the relationship of ABCG2 and SLCO1A2 gene expression with markers of response to MI in individuals with CML and evaluate the influence of polymorphisms ABCG2 c.421C> A and c. ABCG2-19-99G> A in response to the MI. One hundred and eighteen patients in chronic phase of CML were studied and classified in two groups: Responder Group comprised 70 patients who had a complete cytogenetic response within 18 months of treatment. The non-responder group comprised 48 patients who did not have a complete cytogenetic response with the initial dose (400 mg/day) of IM or who relapsed during treatment and were submitted to higher doses of 600 or 800 mg/day. Criteria of failed response to treatment were established by European LeukemiaNet. Patients with cytogenetic patterns other than the Philadelphia chromosome and patients with mutations in the BCR-ABL1 gene were excluded from this study. Blood samples were obtained for: total RNA extraction for quantification of BCR-ABL1 and gene expression of ABCG2 and SLCO1A2; genomic DNA extraction and band G cytogenetic analysis. The gene expression and the analysis of the polymorphisms were performed by real time PCR. Expression of ABCG2 in non-responder group was higher than in responder group (P=0.028). This result was influenced by patients with primary resistance (n= 34 P=0.029) but not secondary resistance (n=14 P=0.249) when compared with responders (n=70). The higher expression of ABCG2 gene was also associated with those patients who had major molecular response (number of BCR-ABL1 . 0.1%) (P=0.027) when all patients were analyzed. The studied gene was not associated with the complete molecular response (number of BCR-ABL1 .0.0032). Regarding to the gene SLCO1A2 was not possible to determine its expression due to low concentration of RNA obtained. The c.421C>A e c.-19-99G>A were not associated neither with the ABCG2 gene expression and MI response. CMR in responders group was associated with the 421CC genotype ant there was a trend for higher frequency of carriers of genotype -19-99GG in the same group. Carriers of 19-99AA genotype tended to the risk of having CML. The results of this study allow us to conclude that the higher expression of ABCG2 is associated with primary resistance to IM and may be a mediator of resistance to IM. The ABCG2 polymorphisms did not influence the gene expression of ABCG2 but impacted in CMR of the responders to IM.

Le traitement médical du cancer colorectal a connu ces dernières années d'importantes avancées avec l'arrivée notamment des thérapies ciblées anti-angiogéniques et anti-EGFR. Néanmoins ces molécules ne bénéficient pas à tous les patients et il est à ce jour impossible de bien individualiser leur utilisation. Mon travail de thèse s'est donc intéressé à d'une part à identifier de nouveaux facteurs prédictifs de réponse en particulier au bévacizumab, d'autre part à rechercher et antagoniser de nouvelles cibles ou des mécanismes participant à la résistance aux molécules disponibles notamment pour les patients non répondeurs aux anti-EGFR. Premièrement, nous avons montré que le sorafenib améliorait l'activité anti-tumorale de l'irinotecan in vitro et in vivo sur des lignées cellulaires de cancer du colon rendues résistantes au SN38 indépendamment de leur statut K-ras. Pour expliquer ce phénomène, nous avons mis en évidence que le sorafenib inhibait la pompe d'efflux ABCG2 et favorisait l'accumulation intra-cellulaire de l'irinotecan et donc sa cytotoxicité. De plus, nous avons vérifié la faisabilité d'une telle association chez l'homme et confirmé son efficacité chez des patients K-ras mutés lourdement prétraités. Deuxièmement, nous avons développé un nouveau score radiologique objectif combinant l'évolution de la taille et la densité tumorale normalisée à celle du foie qui pourrait être utilisé comme marqueur de substitution pour déterminer précocément les bons répondeurs à une chimiothérapie à base de bevacizumab chez des patients présentant des métastases hépatiques. / The medical treatment of colorectal cancer has made significant progresses in recent years including the arrival of targeted therapies: anti-angiogenic and anti-EGFR. However, these molecules don't benefit all patients and their use is not well personalized. My thesis is therefore aimed, on one hand, to identify new predictors of response especially to bevacizumab and, on the other hand, to find and antagonize new targets or mechanisms involved in chemo-resistance, especially for K-ras mutated patients. Firstly, we have showed that sorafenib improved the anti-tumoral activity of irinotecan, both in vitro and in vivo, in SN-38 resistant colon adenocarcinoma cell lines independently of their K-ras status. To explain this phenomenon, we have demonstrated that sorafenib inhibited the efflux pump ABCG2 and promoted the intracellular accumulation of irinotecan and thus its cytotoxicity. In addition, we have checked the feasibility of such an association in human and confirmed its efficacy in K-ras mutated heavily pretreated patients. Secondly, we have developed a new objective radiological score combining both tumor size and density normalized to the liver that could be used as objective surrogate markers to determine early good responders after bevacizumab-containing chemotherapy in patients with colorectal liver metastases.

La résistance à de multiples drogues anticancéreuses (Multidrug Resistance ou MDR) est actuellement un problème majeur dans le cas de nombreuses chimiothérapies. Parmi les mécanismes à l'origine de la MDR, la surexpression de protéines membranaires de type ABC est le plus étudié. Les deux protéines ABCG2 et MRP1 sont parmi les protéines membranaires impliquées. Ces transporteurs sont capables d'induire un efflux massif des agents anticancéreux hors des cellules cancéreuses, réduisant ainsi leur concentration intracellulaire et donc leur efficacité thérapeutique. Afin de contrecarrer cette chimiorésistance, notre objectif s'est concentré sur le développement de nouveaux modulateurs d'ABCG2 et de MRP1. Dans ce cadre, de nouveaux inhibiteurs d'ABCG2, dérivés de chromones, ont été conçus afin de restaurer la sensibilité des cellules cancéreuses aux agents anticancéreux. De plus, la modélisation de modèles pharmacophores nous a permis d'obtenir de nouvelles informations quant aux interactions ABCG2-ligands. Les nouveaux modulateurs de MRP1, dérivés de flavonoïdes, sont capables quant à eux d'induire un efflux massif de glutathion cellulaire via MRP1, sans être transportés eux même, entraînant l'apoptose sélective des cellules cancéreuses surexprimant le transporteur. / Resistance to chemotherapeutic agents (Multidrug Resistance or MDR) is a major hurdle for anticancer chemotherapy. Among different mechanisms involved in MDR, the overexpression of membrane proteins belonging to ABC family is the most relevant one. Among such proteins, ABCG2 and MRP1 are considered to play an important role. These transporters are able to induce a massive efflux of anticancer agents out of the cancer cells, reducing their intracellular concentration and their therapeutic potency. In order to overcome this resistance, novel modulators of ABCG2 and MRP1 were designed, synthetized and tested biologically. In this context, new derivatives of chromones as inhibitors of ABCG2 were developed in order to restore sensitivity of cancer cells to chemotherapeutic agents. In addition, molecular modelling of new pharmacophores allowed us to gather new data exploring ABCG2-ligand interactions. New modulators of MRP1, derivatives of flavonoids, are able to induce a massive efflux of intracellular glutathione that is mediated by the protein, without being transported and causing selective apoptosis of cancer cells overexpressing MRP1.

ABCG2 ou BCRP est une protéine membranaire de la famille des transporteurs ABC. Elle utilise l’énergie de l’hydrolyse de l’ATP pour exporter des composés endogènes et exogènes hors des cellules. Elle participe ainsi à la protection et la détoxication de l’organisme. En revanche, dans le cas des cellules cancéreuses, elle est surexprimée et participe donc au phénotype de résistance à de multiples médicaments (MDR : Multi Drug Resistance). En effet, lors de la surexpression de cette protéine, les agents anti-cancéreux sont exportés hors des cellules tumorales, ce qui diminue leurs concentrations intracellulaires sous leurs seuils de cytotoxicité et les rend inefficaces. De par l’importance d’ABCG2 dans la chimiorésistance, de nombreux efforts sont effectués pour concevoir des inhibiteurs afin de restaurer la sensibilité des cellules cancéreuses. Dans ce contexte, le projet de thèse vise à caractériser ABCG2 sur les plans structural et fonctionnel afin de comprendre son mécanisme d’action. La protéine ABCG2 exprimée chez E. coli, a été purifiée, sous forme stable et homogène et le rendement est de 1,5 mg de protéine par litre de culture. La caractérisation fonctionnelle de celle-ci témoigne de son repliement correct. En effet, il a été démontré qu’elle est capable de fixer différents substrats (naturels et agents anti-cancéreux) avec des affinités différentes. Des essais préliminaires de cristallisation ainsi que des observations par microscopie électronique révèlent des résultats encourageants pour la suite de la caractérisation structurale / ABCG2 or BCRP is a membrane protein that belongs to the ABC transporter family. It uses the hydrolysis energy of ATP to export endogenous and exogenous compounds out of cells. It is thus involved in the protection and detoxification of the body. However, it is overexpressed by cancer cells and participates in the multidrug resistance phenotype (MDR); in fact, anti-cancer agents are exported out of the tumor cells, which reduce their concentration below their cytotoxicity threshold and renders them ineffective. Because of its importance in chemoresistance, many efforts are made to design inhibitors to restore the sensitivity of cancer cells. In this context, the PhD project aims to characterize ABCG2 at structural and functional levels in order to understand its mechanism of action. We have succeeded in purifying ABCG2 expressed in E. coli, the protein obtained is stable and homogeneous, with a yield of 1.5 mg of protein per liter of culture. The functional characterization of ABCG2 demonstrates its correct folding. In fact, we have demonstrated that it is able to bind different substrates (natural and anti-cancer agents) with different affinities. Preliminary crystallization assays and electron microscopy observations reveal encouraging results for subsequent structural characterization