Mitochondrial hyperfusion via metabolic sensing of regulatory amino acids

The relationship between nutrient starvation and mitochondrial dynamics is poorly understood. We find that cells facing amino acid starvation display clear mitochondrial fusion as a means to evade mitophagy. Surprisingly, further supplementation of glutamine (Q), leucine (L), and arginine (R) did not reverse, but produced stronger mitochondrial hyperfusion. Interestingly, the hyperfusion response to Q + L + R was dependent upon mitochondrial fusion proteins Mfn1 and Opa1 but was independent of MTORC1. Metabolite profiling indicates that Q + L + R addback replenishes amino acid and nucleotide pools. Inhibition of fumarate hydratase, glutaminolysis, or inosine monophosphate dehydrogenase all block Q + L + R-dependent mitochondrial hyperfusion, which suggests critical roles for the tricarboxylic acid (TCA) cycle and purine biosynthesis in this response. Metabolic tracer analyses further support the idea that supplemented Q promotes purine biosynthesis by serving as a donor of amine groups. We thus describe a metabolic mechanism for direct sensing of cellular amino acids to control mitochondrial fusion and cell fate

The impact of AMPK signalling and clinical therapeutics on cancer metabolism

Cancer metabolism is intricately re-wired to support tumour growth. Despite the initial discovery that cancers depend on aerobic glycolysis, it is well appreciated that cancer cells exploit numerous metabolic pathways, including those linked to mitochondrial metabolism, to help fuel tumorigenesis. Metabolic gene programs are controlled by transcriptional complexes, such as the peroxisome proliferator-activated gamma coactivator 1 (PGC-1) / estrogen-related receptor alpha (ERR) axis, which act as master orchestrators of metabolism. The activity of the PGC-1α/ERRα axis is upregulated by the AMP-activated protein kinase (AMPK), a central metabolic regulator that is triggered in response to energetic stress. The work in this thesis demonstrates that the AMPK/PGC-1α/ERRα axis increases the bioenergetic functions of cancer cells, but inhibits one-carbon metabolism and purine biosynthesis, resulting in improved sensitivity to methotrexate (MTX), a chemotherapeutic drug widely used in the clinic. MTX treatment promotes bioenergetic functions and has antiproliferative effects that are dependent on AMPK. As a result, the combination of MTX with AMPK activators can improve chemotherapeutic response. Recently, there is increased interest in repurposing metabolic drugs to treat cancer. We show that the antidiabetic drug canagliflozin decreases cancer cell proliferation and reduces the activity of the citric acid cycle through perturbation of glutamine metabolism. The work in this thesis unravels the role of the AMPK/PGC-1α/ERRα pathway in controlling antifolate response and reinforces the premise of pharmacologically targeting cancer metabolism to impede tumourigenesis.Le métabolisme du cancer s'adapte de manière complexe pour soutenir la croissance tumorale. Outre leur dépendance à la glycolyse aérobie, tel que découvert initialement, les cellules cancéreuses exploitent de nombreuses autres voies métaboliques, incluant celle du métabolisme mitochondrial, afin d'alimenter la tumorigenèse. Les gènes métaboliques sont contrôlés par des complexes transcriptionnels tels que l'axe du peroxisome proliferator-activated gamma coactivator 1 (PGC-1) / estrogen-related receptor alpha (ERR), des facteurs de transcription bien connus dans l'orchestration du métabolisme. La régulation de cet axe est activée par l'AMPK, un acteur métabolique central qui est induit en réponse à un stress énergétique. Les travaux présentés dans cette thèse montrent que l'activation de l'axe AMPK/PGC-1α/ERRα augmente la fonction bioénergétique tout en diminuant le métabolisme du folate et la biosynthèse des purines, rendant ainsi ces cellules cancéreuses sensibles au méthotrexate (MTX), un agent chimiotherapeutique communément utilisé chez des patients atteints de cancer. Les mécanismes d'action du MTX se traduisent par une promotion des fonctions bioénergétiques tout en ayant des effets antiprolifératifs dépendants de l'AMPK. Par conséquent, la combinaison du MTX et d'un activateur de l'AMPK pourrait améliorer la réponse chimiothérapeutique, rendant ainsi le traitement plus efficace. L'utilisation de médicaments métaboliques suscite de plus en plus un grand intérêt dans le traitement du cancer. En effet, nous avons mis en évidence que la canagliflozine, un médicament anti-diabétique, induit une diminution de la prolifération des cellules cancéreuses et une réduction de l'activité du cycle de Krebs, due à la perturbation du métabolisme de la glutamine. Ainsi les travaux présentés dans cette thèse mettent en lumière le rôle de la voie AMPK/PGC-1α/ERRα dans le contrôle de la réponse anti-folate et le potentiel de la thérapie pharmacologique ciblant le métabolisme du cancer comme stratégie pour bloquer la tumorigenèse

The PGC-1α/ERRα Axis Represses One-Carbon Metabolism and Promotes Sensitivity to Anti-folate Therapy in Breast Cancer

Reprogramming of cellular metabolism plays a central role in fueling malignant transformation, and AMPK and the PGC-1α/ERRα axis are key regulators of this process. The intersection of gene-expression and binding-event datasets for breast cancer cells shows that activation of AMPK significantly increases the expression of PGC-1α/ERRα and promotes the binding of ERRα to its cognate sites. Unexpectedly, the data also reveal that ERRα, in concert with PGC-1α, negatively regulates the expression of several one-carbon metabolism genes, resulting in substantial perturbations in purine biosynthesis. This PGC-1α/ERRα-mediated repression of one-carbon metabolism promotes the sensitivity of breast cancer cells and tumors to the anti-folate drug methotrexate. These data implicate the PGC-1α/ERRα axis as a core regulatory node of folate cycle metabolism and further suggest that activators of AMPK could be used to modulate this pathway in cancer

PGC-1α supports glutamine metabolism in breast cancer

Background: Glutamine metabolism is a central metabolic pathway in cancer. Recently, reductive carboxylation of glutamine for lipogenesis has been shown to constitute a key anabolic route in cancer cells. However, little is known regarding central regulators of the various glutamine metabolic pathways in cancer cells. Methods: The impact of PGC-1α and ERRα on glutamine enzyme expression was assessed in ERBB2+ breast cancer cell lines with quantitative RT-PCR, chromatin immunoprecipitation, and immunoblotting experiments. Glutamine flux was quantified using 13C-labeled glutamine and GC/MS analyses. Functional assays for lipogenesis were performed using 14C-labeled glutamine. The expression of glutamine metabolism genes in breast cancer patients was determined by bioinformatics analyses using The Cancer Genome Atlas. Results: We show that the transcriptional coactivator PGC-1α, along with the transcription factor ERRα, is a positive regulator of the expression of glutamine metabolism genes in ERBB2+ breast cancer. Indeed, ERBB2+ breast cancer cells with increased expression of PGC-1α display elevated expression of glutamine metabolism genes. Furthermore, ERBB2+ breast cancer cells with reduced expression of PGC-1α or when treated with C29, a pharmacological inhibitor of ERRα, exhibit diminished expression of glutamine metabolism genes. The biological relevance of the control of glutamine metabolism genes by the PGC-1α/ERRα axis is demonstrated by consequent regulation of glutamine flux through the citric acid cycle. PGC-1α and ERRα regulate both the canonical citric acid cycle (forward) and the reductive carboxylation (reverse) fluxes; the latter can be used to support de novo lipogenesis reactions, most notably in hypoxic conditions. Importantly, murine and human ERBB2+ cells lines display a significant dependence on glutamine availability for their growth. Finally, we show that PGC-1α expression is positively correlated with that of the glutamine pathway in ERBB2+ breast cancer patients, and high expression of this pathway is associated with reduced patient survival. Conclusions: These data reveal that the PGC-1α/ERRα axis is a central regulator of glutamine metabolism in ERBB2+ breast cancer. This novel regulatory link, as well as the marked reduction in patient survival time associated with increased glutamine pathway gene expression, suggests that targeting glutamine metabolism may have therapeutic potential in the treatment of ERBB2+ breast cancer

Copper bioavailability is a KRAS-specific vulnerability in colorectal cancer.

Despite its importance in human cancers, including colorectal cancers (CRC), oncogenic KRAS has been extremely challenging to target therapeutically. To identify potential vulnerabilities in KRAS-mutated CRC, we characterize the impact of oncogenic KRAS on the cell surface of intestinal epithelial cells. Here we show that oncogenic KRAS alters the expression of a myriad of cell-surface proteins implicated in diverse biological functions, and identify many potential surface-accessible therapeutic targets. Cell surface-based loss-of-function screens reveal that ATP7A, a copper-exporter upregulated by mutant KRAS, is essential for neoplastic growth. ATP7A is upregulated at the surface of KRAS-mutated CRC, and protects cells from excess copper-ion toxicity. We find that KRAS-mutated cells acquire copper via a non-canonical mechanism involving macropinocytosis, which appears to be required to support their growth. Together, these results indicate that copper bioavailability is a KRAS-selective vulnerability that could be exploited for the treatment of KRAS-mutated neoplasms

Mitochondrial hyperfusion via metabolic sensing of regulatory amino acids

The relationship between nutrient starvation and mitochondrial dynamics is poorly understood. We find that cells facing amino acid starvation display clear mitochondrial fusion as a means to evade mitophagy. Surprisingly, further supplementation of glutamine (Q), leucine (L), and arginine (R) did not reverse, but produced stronger mitochondrial hyperfusion. Interestingly, the hyperfusion response to Q + L + R was dependent upon mitochondrial fusion proteins Mfn1 and Opa1 but was independent of MTORC1. Metabolite profiling indicates that Q + L + R addback replenishes amino acid and nucleotide pools. Inhibition of fumarate hydratase, glutaminolysis, or inosine monophosphate dehydrogenase all block Q + L + R-dependent mitochondrial hyperfusion, which suggests critical roles for the tricarboxylic acid (TCA) cycle and purine biosynthesis in this response. Metabolic tracer analyses further support the idea that supplemented Q promotes purine biosynthesis by serving as a donor of amine groups. We thus describe a metabolic mechanism for direct sensing of cellular amino acids to control mitochondrial fusion and cell fate

Copper bioavailability is a KRAS-specific vulnerability in colorectal cancer

The oncogene KRAS is frequently mutated in cancer, including colorectal cancer. Here, using a cell-surface proteomics approach, KRAS-mutated colorectal cancer cells are shown to express high levels of the copper transporter ATP7A, which has an essential roles in cancer cell survival and proliferation

PRDM15 is a key regulator of metabolism critical to sustain B-cell lymphomagenesis

PRDM (PRDI-BF1 and RIZ homology domain containing) family members are sequence-specific transcriptional regulators involved in cell identity and fate determination, often dysregulated in cancer. The PRDM15 gene is of particular interest, given its low expression in adult tissues and its overexpression in B-cell lymphomas. Despite its well characterized role in stem cell biology and during early development, the role of PRDM15 in cancer remains obscure. Herein, we demonstrate that while PRDM15 is largely dispensable for mouse adult somatic cell homeostasis in vivo, it plays a critical role in B-cell lymphomagenesis. Mechanistically, PRDM15 regulates a transcriptional program that sustains the activity of the PI3K/AKT/mTOR pathway and glycolysis in B-cell lymphomas. Abrogation of PRDM15 induces a metabolic crisis and selective death of lymphoma cells. Collectively, our data demonstrate that PRDM15 fuels the metabolic requirement of B-cell lymphomas and validate it as an attractive and previously unrecognized target in oncology