Functional role of pentose phosphate pathway and glutamine in cancer cell metabolism

Resumen

Cancer is a term that gathers up many heterogeneous sets of diseases and it is characterized by the loss of physiological control and the malignant transformation of the healthy cells. Acquirement of functional and genetic abnormalities leads non-transformed cells to tumorigenesis and tumor progression. These abnormal cells divide out of the control with a tendency to spread throughout the body and to found new colonies of cancer cells, which is termed as metastasis which is the main reason of cancer related deaths. It is vital to understand cancer cell biology in order to identify novel biomarkers for early diagnosis and design new therapeutic strategies. Metabolic reprogramming is an emerging hallmark of cancer, which means that cancer cells switch their core metabolism to meet the increased requirements of cell growth and division. Therefore, exploring metabolic reprograming that cancer cells undergo is a key strategy to identify new targets for cancer therapy and diagnosis. In this thesis, new possibilities for cancer treatment have been explored by analyzing the tumor metabolic reprogramming. In this regard, we studied and proposed pentose phosphate pathway (PPP) enzymes as putative therapeutic targets against breast and colon cancers. Also, we have explored the glutamine metabolism in breast cancer cells and the metabolic network adaptations that they undergo in order to circumvent glutamine deprivation and general mitochondrial impairment. Besides nucleotide biosynthesis and redox detoxification, PPP also has important roles in many other aspects related to the viability of cancer cells, involving proliferation. Thus, targeting PPP is the interest of investigators to use both oxidative and non-oxidative phases of this metabolic pathway as a therapeutic drug target. To test this, we inhibited ox-PPP enzymes 6PGD in breast cancer cells and G6PD in colon cancer cells. We have performed the characterization of metabolic reprogramming induced by inhibition of ox-PPP enzyme by RNA interferase (RNAi) mediated silencing, in order to explore the potential of this enzyme as a therapeutic drug target in two breast cancer cell lines. In addition, we also investigated the relation between p53 activation and PPP and how PPP is related with glutamine metabolism. We demosntrated that 6PGD inhibition resulted in decreased proliferation rate, cell cycle arrest and induction of apoptosis mediated by p53 activation. Moreover, 6PGD knockdown altered stem cell characteristics of breast cancer cells by decreasing mammosphere formation capabilities and altered central carbon metabolism by modulating Warburg phenomenan and enhancing glutamine metabolism. On the other hand, we showed the effect of G6PD inhibition on proliferation of colon cancer cells and that PPP is regulated by glutamine availability in colon cancer cells. Moreover, we characterized the metabolic adaptations that breast cancer cells undergo in the deprivation of glutamine or when mitochondria are defected. We conducted metabolic flux analysis using metabolomics and fluxomics approaches and we employed Systems Biology approaches in order to estimate a global view of flux alterations in different culture conditions. We observed an increased pyruvate cycle with glutamine deprivation, thus indicating that targeting the enzymes of this pathway such as malic enzyme could be a promising approach combined with inhibition of glutaminase enzyme. On the other hand, we observed that mimicking hypoxia by oligomycin treatment redirected breast cancer cells to increase reductive carboxylation. Considering that hypoxia is a common condition in the tumor environment, targeting reductive carboxylation mechanism could be a novel strategy to fight against cancer. Collectively, all the results provided in this thesis demosntrate the importance of metabolism in cancer cell proliferation and survival. This work also highlights the importance of Systems Biology approaches to comprehend the molecular mechanisms underlying complex multifactorial diseases in order to point out new potential therapeutic targets.