Whilst progress has been made in reducing mortality in some cancers, mortality rates remain high in many cancers and there is a need to develop novel therapeutic strategies. In this thesis, various pharmacological strategies designed to enhance the activity of existing therapeutic drugs were evaluated. Cancer cells are dependent upon aerobic glycolysis (the Warburg effect) and glutamine uptake. Using clinically approved tyrosine kinase inhibitors and Bortezomib, significant enhancement of chemosensitivity was observed when used in combination with inhibitors of lactate dehydrogenase (Gossypol) and pyruvate kinase dehydrogenase (Dichloroacetate). In contrast, depletion of glutamine from media had to be extensive in order to induce cell death and cell death only occurred after prolonged exposure to glutamine-deprived conditions. This suggests that glutamine depletion strategies alone are unlikely to be successful but may be useful in combination with other agents targeting glutamine addiction in cancer cells. Finally, Temozolomide (TMZ) is an important drug in the treatment of glioblastomas but its activity is reduced by resistance mechanisms including O6 methyl guanine methyltransferase (MGMT) and mismatch repair (MMR). This thesis has identified analogues of TMZ (EA02-45, EA02-59, EA02-64 and EA02-65) that are MGMT and MMR independent in terms of inducing cell kill in vitro. These compounds are promising leads for future development. In conclusion, this thesis has demonstrated that interfering with the metabolic phenotype of cancer can enhance the activity of existing drugs and identified novel analogues of TMZ that circumvent drug resistance mechanisms that hamper the efficacy of TMZ.

Le métabolisme cellulaire est la source principale d’énergie et les cellules cancéreuses ont un métabolisme différent des cellules non transformées. La cellule tumorale a tendance à éviter l’activité mitochondriale et ainsi la phosphorylation oxydative, pour lui préférer la voie de la glycolyse pour la production d’énergie (Effet Warburg). Cette altération du métabolisme est si bénéfique pour les cellules en croissance que cela favorise la croissance tumorale et supprime la réponse immunitaire anticancéreuse. La spécificité de ce métabolisme en fait une cible intéressante pour le développement de thérapies anticancéreuses. Mon travail de thèse comporte deux parties. La première partie décrit que lorsque les cellules cancéreuses sont forcées à utiliser la voie mitochondriale comme source d’énergie à travers l’oxydation phosphorylative, elles initient un mécanisme antioxydant pour tolérer les effets délétères des espèces oxygénées réactives (EOR ou ROS pour reactive oxygene species) produites au cours de l’activité mitochondriale. La stimulation mitochondriale entraîne l’activation de la voie de signalisation ERK5-MEF2, et cette dernière engendre un mécanisme antioxydant de deux façons.Initialement, nous avons observé que MEF2 régule positivement l’expression de miR23a, et ce dernier inhibe l’expression de KEAP1. Cette protéine est responsable de la dégradation ubiquitine dépendante de NRF2, un régulateur clé de la réponse antioxydante cellulaire. L’inhibition de KEAP1 empêche la dégradation cytoplasmique de NRF2. Consécutivement à cela la concentration cytoplasmique en NRF2 augmente ce qui engendre sa translocation dans le noyau où il se lie à une séquence élément de réponse antioxydant (ARE) dans la région promotrice de nombreux gènes antioxydants, initiant ainsi leur transcription. Plus tard nous avons observé que l’activation de la voie ERK5-MEF2 induisait directement la synthèse de novo de NRF2, induisant sa translocation nucléaire et un mécanisme antioxydant. L’inhibition de la voie ERK5-MEF2 altère la réponse antioxydante, sensibilisant ainsi les cellules au stress oxydant.La seconde partie de mon travail a exploré les mécanismes à l’origine des effets hypolipémiants du dichloroacétate (DCA). Le DCA est une petite molécule qui inhibe la PDK1 et permet au pyruvate d’entrer dans la mitochondrie. Il a été utilisé en clinique dans le passé pour baisser les taux plasmatiques de cholestérol mais le mécanisme n’était pas clair et nous l’avons décris. Le DCA force les cellules à entrer en oxydation phosphorylative ce qui active la voie ERK5-MEF2. Cette voie augmente directement l’expression du LDLR (Low Density Lipoprotein Receptor ; récepteur aux lipoprotéines de basse densité) qui permet l’endocytose des LDL riches en cholestérol qui sont responsables de la plupart des maladies cardiovasculaires. L’inhibition de cette voie supprime l’afflux de lipides et par conséquent serait une cible intéressante pour de futures recherches puisque de hauts taux de cholestérols sont directement corrélés avec une augmentation du risque d’athérosclérose et de toutes les complications mortelles qu’il entraine.Notre prochain objectif est d’explorer les autres mécanismes cellulaires régulés par la voie ERK5-MEF2. Sur la base de nos résultats préliminaires, nous proposons que cette voie non seulement régule l’expression du LDLR mais aussi celle de nombreux autres gènes qui sont impliqués directement ou indirectement dans le métabolisme des lipides. / Cellular metabolism is the main source of energy and cancer cells has different metabolism than non-transformed cells. Tumor cell tends to avoid mitochondrial activity and oxidative phosphorylation (OXPHOS) and prefer glycolysis for energy production (Warburg effect). This alteration in metabolism is beneficial for growing cells in many ways that promote tumor growth and suppress the anti-cancer immune response. This specific metabolism is an auspicious target for the better development of cancers chemotherapies.My thesis work comprises two parts. The first portion describes that when cancer cells are forced to utilize their mitochondria in order to obtain the energy from OXPHOS they initiate an antioxidant mechanism to cope with the deleterious effects of reactive oxygen species (ROS) produced during mitochondrial activity. Mitochondrial stimulation leads to activation of ERK5-MEF2 signaling pathway, which triggers the antioxidant mechanism by at least two ways.Initially we observed that MEF2 up regulates the expression of miR23a, which inhibits KEAP1 expression. This protein is responsible for ubiquitinational degradation of NRF2, a master regulator of the antioxidant response in cells. The inhibition of KEAP1 prevents the NRF2 cytoplasmic degradation. This results in high built up of NRF2 in cytoplasm that translocates to nucleus where it binds to ARE (antioxidant response element) in the upstream promoter region of many antioxidant genes hence initiates their transcription. Latter we observed that activation of ERK5-MEF2 pathway directly results in de novo synthesis of NRF2, resulting in nuclear translocation and triggering of the antioxidative mechanism. Inhibition of ERK5-MEF2 pathway impairs the cellular antioxidant response, thus sensitizing cells towards oxidative stress.The second part of my work explored the mechanism behind the lipid lowering effects of dichloroacetate (DCA). DCA is a small molecule, which inhibits the PDK1 and enables pyruvate to enter the mitochondria. It was used clinically in past to lower the plasma cholesterol level but the underlying mechanism was not clear and we describe it here. DCA forces cells to perform OXPHOS, which activate the ERK5-MEF2 pathway. This pathway directly up-regulates the expression of Low Density Lipoprotein Receptors (LDLR) that are mainly involved in the endocytosis of cholesterol-rich low density lipoproteins, which are responsible for the majority of cardiovascular diseases. Inhibition of this pathway suppresses lipid influx and hence, it would be an interesting target of future investigation since high cholesterol level is the main cause of various life threatening diseases and the development of atherosclerosis.Our next goal is to exploit other possible cellular mechanism regulated by ERK5-MEF2 pathway. Based on our preliminary data, we propose that this pathway not only regulate the LDLR expression but many other genes, which are directly or indirectly involved in lipid metabolism.

Objective: Challenging differential diagnosis Background: B cell lymphoma constitutes 80-85% of cases of Non Hodgkin's lymphoma in the Untied States. Metabolic complications may arise from the disease itself or through its end organ involvement. Case Report: We describe a case of a diffuse large B cell lymphoma diagnosed by abdominal computed tomography after it initially presented as hypoglycemia not correctable by dextrose infusion that instead resulted in increased anion gap metabolic acidosis with elevated lactate levels. Conclusions: The case illustrates how lymphomas can present unusually with hypoglycemia and lactic acidosis, the latter being an ominous sign that can occur without liver involvement. In this regard, the case demonstrates the metabolic sequelae of lymphoma that should raise suspicion for an underlying process. This has implications for diagnosis, treatment, and patient survival. Attention should be paid especially in the primary care setting in order to minimize delays in diagnosis.

Objective: Challenging differential diagnosis Background: B cell lymphoma constitutes 80-85% of cases of Non Hodgkin's lymphoma in the Untied States. Metabolic complications may arise from the disease itself or through its end organ involvement. Case Report: We describe a case of a diffuse large B cell lymphoma diagnosed by abdominal computed tomography after it initially presented as hypoglycemia not correctable by dextrose infusion that instead resulted in increased anion gap metabolic acidosis with elevated lactate levels. Conclusions: The case illustrates how lymphomas can present unusually with hypoglycemia and lactic acidosis, the latter being an ominous sign that can occur without liver involvement. In this regard, the case demonstrates the metabolic sequelae of lymphoma that should raise suspicion for an underlying process. This has implications for diagnosis, treatment, and patient survival. Attention should be paid especially in the primary care setting in order to minimize delays in diagnosis.

京都大学 / 0048 / 新制・課程博士 / 博士(医学) / 甲第19591号 / 医博第4098号 / 新制||医||1014(附属図書館) / 32627 / 京都大学大学院医学研究科医学専攻 / (主査)教授 野田 亮, 教授 武田 俊一, 教授 小川 修 / 学位規則第4条第1項該当 / Doctor of Medical Science / Kyoto University / DFAM

No description available.

Whilst progress has been made in reducing mortality in some cancers, mortality rates remain high in many cancers and there is a need to develop novel therapeutic strategies. In this thesis, various pharmacological strategies designed to enhance the activity of existing therapeutic drugs were evaluated. Cancer cells are dependent upon aerobic glycolysis (the Warburg effect) and glutamine uptake. Using clinically approved tyrosine kinase inhibitors and Bortezomib, significant enhancement of chemosensitivity was observed when used in combination with inhibitors of lactate dehydrogenase (Gossypol) and pyruvate kinase dehydrogenase (Dichloroacetate). In contrast, depletion of glutamine from media had to be extensive in order to induce cell death and cell death only occurred after prolonged exposure to glutamine-deprived conditions. This suggests that glutamine depletion strategies alone are unlikely to be successful but may be useful in combination with other agents targeting glutamine addiction in cancer cells. Finally, Temozolomide (TMZ) is an important drug in the treatment of glioblastomas but its activity is reduced by resistance mechanisms including O6 methyl guanine methyltransferase (MGMT) and mismatch repair (MMR). This thesis has identified analogues of TMZ (EA02-45, EA02-59, EA02-64 and EA02-65) that are MGMT and MMR independent in terms of inducing cell kill in vitro. These compounds are promising leads for future development. In conclusion, this thesis has demonstrated that interfering with the metabolic phenotype of cancer can enhance the activity of existing drugs and identified novel analogues of TMZ that circumvent drug resistance mechanisms that hamper the efficacy of TMZ.

No description available.

No description available.

No description available.