Definition and validation of targets associated with the metabolic reprogramming of haematological malignancies

Abstract

[eng] Cancer cells, including leukemic cells, can react to therapeutic treatment by altering their metabolic phenotype (“metabolic reprogramming”) to keep their accelerated proliferative state, eventually becoming resistant to the treatment. There is an increasing amount of evidence indicating that metabolic reprogramming is one of the key mechanisms of acquisition of drug resistance by cancer cells. In agreement, several metabolic studies targeting leukaemia and specifically acute myeloid leukaemia (AML) and chronic myeloid leukaemia (CML), have been conducted over the last decades. However, there is still a lack of understanding the metabolic features of both AML and CML leukaemia specially in the acquisition of drug resistance, that is needed for unveiling novel and effective treatments for resistant and non-resistant patients. Therefore, the main objective of this thesis was to investigate the rewiring of cell metabolism occurring in the process of acquisition of resistance to conventional therapeutic treatments in AML and CML malignancies. Next, by revealing this metabolic rewiring, we intended to highlight potential metabolic and non-metabolic targets that could be exploited to overcome resistance to treatments. To this end, we have performed a comprehensive and comparative multi-OMIC study to analyse the links between the metabolic reprogramming and the resistance acquisition of THP-1 and HL- 60 AML cell models sensitive or resistant to cytarabine (AraC) and doxorubicin (Dox), and of KU812 CML cell model sensitive or resistant to imatinib, all under normoxic (21% O2) and hypoxic (1% O2) conditions. The results of this thesis are divided into two chapters. On the one hand, in Chapter 1, the multi-OMIC study performed in AML parental and resistant cells unveiled that the acquisition of AraC resistance causes the reprogramming of the glucose metabolism of THP-1 and HL-60 cells by increasing the glycolytic flux whereas it is not associated with an alteration in the mitochondrial respiration. Moreover, our results also exhibited a possible disfunction of ETC complex I as well as alterations in glutamine and serine-glycine-1C metabolism in AML cells that display a more active mitochondrial metabolism. Moreover, we have also identified that the acquisition of Dox resistance causes alterations in the glucose and amino acid metabolism. Importantly, we have observed an important loss of mitochondrial respiration capacity of AML cells resistant to Dox chemotherapeutic drug, which constitutes a potential metabolic vulnerability that can be exploited for the treatment of AML patients resistant to Dox. On the other hand, in Chapter 2 is shown that the acquisition of imatinib resistance causes the reprogramming of glucose metabolism by enhancing the glycolytic flux, PPP, and glycogen metabolism, thus highlighting these metabolic pathways as potential metabolic weaknesses of KU812 cells resistant to imatinib. Moreover, we have observed a high metabolic plasticity of KU812 cells resistant to imatinib which includes the orchestration of many metabolic routes associated with the amino acid metabolism. Importantly, the CML multi-OMIC study has also unveiled an enhanced mitochondrial respiration capacity, which constitutes another potential vulnerability that can be exploited to overcome imatinib resistance. Finally, both AML and CML multi-OMIC studies have allowed us to propose and/or validate different metabolic and non-metabolic targets. In this regard, in this thesis we have identified and validated a battery of single-hit inhibitions that were able to reduce the cell viability of both parental and resistant AML and CML cells. Finally, we have confirmed that the repurposing of Dox chemotherapeutic drug counteracts the imatinib resistance in the KU812 cells resistant to imatinib.

[spa] Las células cancerosas, incluyendo las leucémicas, pueden reaccionar al tratamiento terapéutico alterando su fenotipo metabólico, para evadir así el efecto de la droga y mantener su proliferación acelerada. Esta alteración del fenotipo metabólico se conoce como "reprogramación metabólica", y cada vez hay más pruebas que sugieren que esta reprogramación metabólica es uno de los mecanismos clave en la adquisición de resistencia farmacológica de las células cancerosas. A pesar del conocimiento generado en la última década, hoy en día sigue siendo un desafío el comprender las características metabólicas tanto de las células de leucemia mieloide aguda (LMA) como de leucemia mieloide crónica (LMC), que permitan desarrollar nuevos tratamientos eficaces en los pacientes que han generado resistencia a los tratamientos actuales. Por lo tanto, el objetivo principal de esta tesis fue investigar la reprogramación metabólica que se produce en las células de LMA y LMC después del proceso de adquisición de resistencia a los diferentes tratamientos terapéuticos convencionales y, en consecuencia, revelar las posibles dianas metabólicas y no metabólicas que podrían explotarse para superar la resistencia farmacológica adquirida. Para ello, se ha realizado un estudio multiómico comparativo entre los modelos celulares de LMA (THP-1 y HL-60) y LMC (KU812) sensibles o resistentes a los quimioterapéuticos convencionales de LMA (AraC y Dox) y LMC (imatinib) en condiciones normóxicas (21% O2) e hipóxicas (1% O2). Los resultados revelaron que el mecanismo de resistencia farmacológica desarrollado por las líneas celulares y los fármacos aquí estudiados se asocia con una importante reprogramación metabólica que es dependiente tanto del modelo celular de estudio como de la droga que genera la resistencia. Además, el estudio de la reprogramación metabólica nos ha permitido proponer y validar diferentes dianas metabólicas y no metabólicas. Así, en esta tesis se han propuesto varias estrategias terapéuticas basadas en inhibir una sola diana (single hit) que fueron capaces de reducir la viabilidad celular tanto de las células LMA y LMC parentales como de las resistentes. Por último, cabe destacar que hemos propuesto y confirmado una mayor eficacia del tratamiento con el quimioterapéutico doxorrubicina en células LMC KU812 resistentes a imatinib.