Absence of Neurofibromin Induces an Oncogenic Metabolic Switch via Mitochondrial ERK-Mediated Phosphorylation of the Chaperone TRAP1.

Mutations in neurofibromin, a Ras GTPase-activating protein, lead to the tumor predisposition syndrome neurofibromatosis type 1. Here, we report that cells lacking neurofibromin exhibit enhanced glycolysis and decreased respiration in a Ras/ERK-dependent way. In the mitochondrial matrix of neurofibromin-deficient cells, a fraction of active ERK1/2 associates with succinate dehydrogenase (SDH) and TRAP1, a chaperone that promotes the accumulation of the oncometabolite succinate by inhibiting SDH. ERK1/2 enhances both formation of this multimeric complex and SDH inhibition. ERK1/2 kinase activity is favored by the interaction with TRAP1, and TRAP1 is, in turn, phosphorylated in an ERK1/2-dependent way. TRAP1 silencing or mutagenesis at the serine residues targeted by ERK1/2 abrogates tumorigenicity, a phenotype that is reverted by addition of a cell-permeable succinate analog. Our findings reveal that Ras/ERK signaling controls the metabolic changes orchestrated by TRAP1 that have a key role in tumor growth and are a promising target for anti-neoplastic strategies.

Metabolic reprogramming identifies the most aggressive lesions at early phases of hepatic carcinogenesis.

Metabolic changes are associated with cancer, but whether they are just bystander effects of deregulated oncogenic signaling pathways or characterize early phases of tumorigenesis remains unclear. Here we show in a rat model of hepatocarcinogenesis that early preneoplastic foci and nodules that progress towards hepatocellular carcinoma (HCC) are characterized both by inhibition of oxidative phosphorylation (OXPHOS) and by enhanced glucose utilization to fuel the pentose phosphate pathway (PPP). These changes respectively require increased expression of the mitochondrial chaperone TRAP1 and of the transcription factor NRF2 that induces the expression of the rate-limiting PPP enzyme glucose-6-phosphate dehydrogenase (G6PD), following miR-1 inhibition. Such metabolic rewiring exclusively identifies a subset of aggressive cytokeratin-19 positive preneoplastic hepatocytes and not slowly growing lesions. No such metabolic changes were observed during non-neoplastic liver regeneration occurring after two/third partial hepatectomy. TRAP1 silencing inhibited the colony forming ability of HCC cells while NRF2 silencing decreased G6PD expression and concomitantly increased miR-1; conversely, transfection with miR-1 mimic abolished G6PD expression. Finally, in human HCC patients increased G6PD expression levels correlates with grading, metastasis and poor prognosis. Our results demonstrate that the metabolic deregulation orchestrated by TRAP1 and NRF2 is an early event restricted to the more aggressive preneoplastic lesions.

Inhibition of glucose-6-phosphate dehydrogenase sensitizes cisplatin-resistant cells to death.

The mechanisms of cisplatin resistance, one of the major limitations of current chemotherapy, has only partially been described. We previously demonstrated that cisplatin-resistant ovarian cancer cells (C13), are characterized by reduced mitochondrial activity and higher glucose-dependency when compared to the cisplatin-sensitive counterpart (2008). In this work we further characterized the role of metabolic transformation in cisplatin resistance. By using transmitochondrial hybrids we show that metabolic reprogramming of cisplatin-resistant cell is not caused by inherent mtDNA mutations. We also found that C13 cells not only present an increased glucose-uptake and consumption, but also exhibit increased expression and enzymatic activity of the Pentose Phosphate pathway (PPP) enzyme Glucose-6-Phosphate Dehydrogenase (G6PDH). Moreover, we show that cisplatin-resistant cells are more sensitive to G6PDH inhibition. Even if the metabolomic fingerprint of ovarian cancer cells remains to be further elucidated, these findings indicate that PPP offers innovative potential targets to overcome cisplatin resistance.

Aerobic glycolysis tunes YAP/TAZ transcriptional activity.

Increased glucose metabolism and reprogramming toward aerobic glycolysis are a hallmark of cancer cells, meeting their metabolic needs for sustained cell proliferation. Metabolic reprogramming is usually considered as a downstream consequence of tumor development and oncogene activation; growing evidence indicates, however, that metabolism on its turn can support oncogenic signaling to foster tumor malignancy. Here, we explored how glucose metabolism regulates gene transcription and found an unexpected link with YAP/TAZ, key transcription factors regulating organ growth, tumor cell proliferation and aggressiveness. When cells actively incorporate glucose and route it through glycolysis, YAP/TAZ are fully active; when glucose metabolism is blocked, or glycolysis is reduced, YAP/TAZ transcriptional activity is decreased. Accordingly, glycolysis is required to sustain YAP/TAZ pro-tumorigenic functions, and YAP/TAZ are required for the full deployment of glucose growth-promoting activity. Mechanistically we found that phosphofructokinase (PFK1), the enzyme regulating the first committed step of glycolysis, binds the YAP/TAZ transcriptional cofactors TEADs and promotes their functional and biochemical cooperation with YAP/TAZ. Strikingly, this regulation is conserved in Drosophila, where phosphofructokinase is required for tissue overgrowth promoted by Yki, the fly homologue of YAP. Moreover, gene expression regulated by glucose metabolism in breast cancer cells is strongly associated in a large dataset of primary human mammary tumors with YAP/TAZ activation and with the progression toward more advanced and malignant stages. These findings suggest that aerobic glycolysis endows cancer cells with particular metabolic properties and at the same time sustains transcription factors with potent pro-tumorigenic activities such as YAP/TAZ.

VEGF-targeted therapy stably modulates the glycolytic phenotype of tumor cells.

Anti-VEGF therapy perturbs tumor metabolism, severely impairing oxygen, glucose, and ATP levels. In this study, we investigated the effects of anti-VEGF therapy in multiple experimental tumor models that differ in their glycolytic phenotypes to gain insights into optimal modulation of the metabolic features of this therapy. Prolonged treatments induced vascular regression and necrosis in tumor xenograft models, with highly glycolytic tumors becoming treatment resistant more rapidly than poorly glycolytic tumors. By PET imaging, prolonged treatments yielded an increase in both hypoxic and proliferative regions of tumors. A selection for highly glycolytic cells was noted and this metabolic shift was stable and associated with increased tumor aggressiveness and resistance to VEGF blockade in serially transplanted mice. Our results support the hypothesis that the highly glycolytic phenotype of tumor cells studied in xenograft models, either primary or secondary, is a cell-autonomous trait conferring resistance to VEGF blockade. The finding that metabolic traits of tumors can be selected by antiangiogenic therapy suggests insights into the evolutionary dynamics of tumor metabolism.

Cancer stem cells from epithelial ovarian cancer patients privilege oxidative phosphorylation, and resist glucose deprivation.

We investigated the metabolic profile of cancer stem cells (CSC) isolated from patients with epithelial ovarian cancer. CSC overexpressed genes associated with glucose uptake, oxidative phosphorylation (OXPHOS), and fatty acid ß-oxidation, indicating higher ability to direct pyruvate towards the Krebs cycle. Consistent with a metabolic profile dominated by OXPHOS, the CSC showed higher mitochondrial reactive oxygen species (ROS) production and elevated membrane potential, and underwent apoptosis upon inhibition of the mitochondrial respiratory chain. The CSC also had a high rate of pentose phosphate pathway (PPP) activity, which is not typical of cells privileging OXPHOS over glycolysis, and may rather reflect the PPP role in recharging scavenging enzymes. Furthermore, CSC resisted in vitro and in vivo glucose deprivation, while maintaining their CSC phenotype and OXPHOS profile. These observations may explain the CSC resistance to anti-angiogenic therapies, and indicate this peculiar metabolic profile as a possible target of novel treatment strategies.

Inhibition of succinate dehydrogenase by the mitochondrial chaperone TRAP1 has anti-oxidant and anti-apoptotic effects on tumor cells.

TRAP1 is a mitochondrial chaperone highly expressed in many tumor types; it inhibits respiratory complex II, down-modulating its succinate dehydrogenase (SDH) enzymatic activity. SDH inhibition in turn leads to a pseudohypoxic state caused by succinate-dependent HIF1α stabilization and promotes neoplastic growth. Here we report that TRAP1 inhibition of SDH also shields cells from oxidative insults and from the ensuing lethal opening of the mitochondrial permeability transition pore. This anti-oxidant activity of TRAP1 protects tumor cells from death in conditions of nutrient paucity that mimic those encountered in the neoplasm during the process of malignant accrual, and it is required for in vitro tumorigenic growth. Our findings demonstrate that SDH inhibition by TRAP1 is oncogenic not only by inducing pseudohypoxia, but also by protecting tumor cells from oxidative stress.

The mitochondrial chaperone TRAP1 promotes neoplastic growth by inhibiting succinate dehydrogenase.

We report that the mitochondrial chaperone TRAP1, which is induced in most tumor types, is required for neoplastic growth and confers transforming potential to noncancerous cells. TRAP1 binds to and inhibits succinate dehydrogenase (SDH), the complex II of the respiratory chain. The respiratory downregulation elicited by TRAP1 interaction with SDH promotes tumorigenesis by priming the succinate-dependent stabilization of the proneoplastic transcription factor HIF1α independently of hypoxic conditions. These findings provide a mechanistic clue to explain the switch to aerobic glycolysis of tumors and identify TRAP1 as a promising antineoplastic target.