Cellular energy metabolism is one of the main processes affected during the transition from normal to cancer cells, and it is a crucial determinant of cell proliferation or cell death. As a support for rapid proliferation, cancer cells choose to use glycolysis even in the presence of oxygen (Warburg effect) to fuel macromolecules for the synthesis of nucleotides, fatty acids, and amino acids for the accelerated mitosis, rather than fuel the tricarboxylic acid cycle and oxidative phosphorylation. Mitochondria biogenesis is also reprogrammed in cancer cells, and the destiny of those cells is determined by the balance between energy and macromolecule supplies, and the efficiency of buffering of the cumulative radical oxygen species. In glioblastoma, the most frequent and malignant adult brain tumor, a metabolic shift toward aerobic glycolysis is observed, with regulation by well known genes as integrants of oncogenic pathways such as phosphoinositide 3-kinase/protein kinase, MYC, and hypoxia regulated gene as hypoxia induced factor 1. The expression profile of a set of genes coding for glycolysis and the tricarboxylic acid cycle in glioblastoma cases confirms this metabolic switch. An understanding of how the main metabolic pathways are modified by cancer cells and the interactions between oncogenes and tumor suppressor genes with these pathways may enlighten new strategies in cancer therapy. In the present review, the main metabolic pathways are compared in normal and cancer cells, and key regulations by the main oncogenes and tumor suppressor genes are discussed. Potential therapeutic targets of the cancer energetic metabolism are enumerated, highlighting the astrocytomas, the most common brain cancer.

A heterogeneous population of uncommon neoplasms of the central nervous system (CNS) cause significant morbidity and mortality. To explore their genetic origins, we sequenced the exomes of 12 pleomorphic xanthoastrocytomas (PXA), 17 non-brainstem pediatric glioblastomas (PGBM), 8 intracranial ependymomas (IEP) and 8 spinal cord ependymomas (SCEP). Analysis of the mutational spectra revealed that the predominant single base pair substitution was a C:G>T:A transition in each of the four tumor types. Our data confirm the critical roles of several known driver genes within CNS neoplasms, including TP53 and ATRX in PGBM, and NF2 in SCEPs. Additionally, we show that activating BRAF mutations play a central role in both low and high grade glial tumors. Furthermore, alterations in genes coding for members of the mammalian target of rapamycin (mTOR) pathway were observed in 33% of PXA. Our study supports the hypothesis that pathologically similar tumors arising in different age groups and from different compartments may represent distinct disease processes with varied genetic composition.

Angiogenesis, induced by the vascular endothelial growth factor A through its ligation to the vascular endothelial growth receptor 2, has been described as a crucial point in high-grade glioma development. The aim of this study was to evaluate the influence of VEGFA–2578C/A, −2489C/T, −1154G/A, −634G/C, and −460C/T, and KDR–604T/C, −271G/A, +1192G/A, and +1719A/T single-nucleotide polymorphisms on risk and clinicopathological aspects of high-grade glioma. This case–control study enrolled 205 high-grade glioma patients and 205 controls. Individuals with VEGFA–2578 CC or CA, VEGFA–1154 GG, VEGFA–634 GC or CC, and VEGFA–460 CT or TT genotypes were under 2.56, 1.53, 1.54, and 1.84 increased risks of high-grade glioma, compared to others, respectively. And 1.61, 2.66, 2.52, 2.53, and 2.02 increased risks of high-grade glioma were seen in individuals with VEGFA–2578 CC plus VEGFA–1154 GG, VEGFA–2578 CC or CA plus VEGFA–634 GC or CC, VEGFA–2578 CC or CA plus VEGFA–460 CT or TT, VEGFA–1154 GG or GA plus VEGFA–634 GC or CC, and VEGFA 634 GC or CC plus VEGFA–460 CT or TT combined genotypes, respectively, when compared to others. The “CAGT” haplotype of KDR single-nucleotide polymorphisms was more common in patients with grade IV than in those with grade III tumors, and individuals carrying this haplotype were at 1.76 increased risk of developing grade IV tumors than others. We present, for the first time, preliminary evidence that VEGFA–2578C/A and VEGFA–1154G/A single-nucleotide polymorphisms increases high-grade glioma risk, and “CAGT” haplotype of the KDR gene alters high-grade glioma aggressiveness and risk of grade IV tumors in Brazil.

Angiogenic switch is a hallmark for tumor growth, and strategies to inhibit this process have been pursued for cancer therapy, but with limited long-term efficacy. Recently, targeting endothelial cell (EC) metabolism emerged as an alternative to regulate angiogenesis. In fact, glutamine metabolism is essential for EC sprouting in vitro, and the consumption/secretion rate measurements revealed that proliferating ECs consumed glutamine more than any other amino acid. Glutamine is transported into the cells and converted into α-ketoglutarate to enter the tricarboxylic acid cycle through glutaminolysis, which is upregulated in cancer. Glutaminase (coded by GLS), the main regulator of this pathway, presents two isoforms: GLSiso1 and GLSiso2. We have analyzed the expression level of both GLS isoforms in different grades of astrocytoma, with a gradual increase of their expressions in parallel to the malignancy. GLSiso1 was highly expressed in normal brain samples, in contrast to low GLSiso2 expression, indicating GLSiso2 as a more eligible therapeutic target. GLSiso2-specific transcript inhibition by siRNA in GBM U87MG cells lead to significant decrease of tumor cell proliferation, and a temozolomide-sensitizing effect, relative to NTC-controls. The RNASeq transcriptome analysis of these GLSiso2-silenced cells revealed a differential expression of genes related to blood vessel development, extracellular matrix organization, angiogenesis and tube development with FDR of 4.95e-04, 5.25e-06, 5.92e-05, 8.6e-06 and 1.0e-04, respectively. The immunohistochemistry analysis in human astrocytoma FFPE samples demonstrated GLS positivity on tumor cells and blood vessels, with higher protein expression in more malignant astrocytoma. Such observation corroborated the transcriptomic finding of GLSiso2 participation in the EC compartment. Moreover, the system biology analysis revealed an interesting net among GLS, TP53, ITB1, TGFBI, TGFBR1/2, COL1A1/2, COL5A2, LUM, ENG, ANGPT1 connecting genes related to metabolism, extracellular matrix and angiogenesis. In the search for the most suitable EC line to study the impact of GLS isoforms on the vessel compartment, we observed that HUVEC immortalized umbilical EC line presented higher expression of GLSiso2 than GLSiso1. Chemical blockade of the total GLS by BPTES and 968 was tested in HUVEC cells and a significant proliferation and migration decrease were observed. Further GLSiso2 specific silencing of ECs might clarify the crosstalk between tumor cells and ECs, and will allow the identification of key targets in their metabolic reprogramming during tumor growth. Citation Format: Yollanda E. Franco, Roseli S. Soares, Marina T. Lima, Antonio M. Lerario, Sueli M. Shinjo, Suely K. Marie. Understanding the link between glutamine metabolism and angiogenesis in astrocytoma [abstract]. In: Proceedings of the Annual Meeting of the American Association for Cancer Research 2020; 2020 Apr 27-28 and Jun 22-24. Philadelphia (PA): AACR; Cancer Res 2020;80(16 Suppl):Abstract nr 3739.

Glioblastoma (GBM) is the most common and aggressive malignant brain tumor in adults and the standard treatment consists of surgical resection of the tumor followed by radiation and chemotherapy with temozolomide. Among the genes with increased expression in GBMs compared to pilocytic astrocytomas, we have previously identified MELK (Marie et al., 2008), which codes for Maternally Embryonic Leucine Zipper Kinase with a role in various cellular processes such as proliferation and cell cycle, apoptosis, gene expression control, hematopoiesis, and oncogenesis. Further microarray and proteomic analyses were performed to identify proteins associated with MELK pathway in astrocytomas. These studies showed high MELK and STMN1 expression levels in astrocytoma of different malignant grades, and identified STMN1 downstream MELK pathway (Marie et al., 2016). Stathmin 1 (coded by STMN1), also known as oncoprotein 18, is an important cytosolic protein that destabilizes microtubules and plays a critical role in mitosis by regulating microtubule dynamics. Also, STMN1 is involved in a variety of other biologic processes, such as cell cycle progression and cell migration, through the phosphorylation of its four serines (Ser16, Ser25, Ser38, and Ser 63). Specific phosphorylation of each serine causes the activation of STMN1, weakening its binding to the tubulin molecules. When the tubulins are free in the cytoplasm, they associate to form microtubules, and participate in biologic processes fundamental to tumor progression, such as formation of mitotic spindle (cell cycle progression) and cytoskeleton remodeling for cell migration. Amongst several proteins that phosphorylate STMN1, FANCC (Fanconi anemia complementation group C) phosphorylates STMN1 at Ser16 and Ser38. Thus, FANCC participates in the regulation of cellular division and is involved in a signaling network that assures the safeguard of chromosomes. The aim of this study was to analyze MELK, STMN1, and FANCC expression levels in our GBM cohort. Additionally, these data were validated in silico in larger GBM cohorts of The Cancer Genome Atlas (TCGA) and The Repository of Molecular Brain Neoplasia Data (Rembrandt) database. We have previously described decrease of STMN1 expression level when MELK was knocked down with siRNA. Interestingly, FANCC expression was also diminished in this condition, indicating a possible involvement of FANCC in MELK pathway. FANCC have higher expression levels in diffusely infiltrative astrocytomas compared to non-neoplastic brain tissue samples. MELK and STMN1 expressions are positively correlated in our GBM series (r=0.678 and p<0.0001 – Spearman-rho test). MELK and FANCC expression levels also correlated (r=0.340 and p=0.0006 – Spearman-rho test). Nonetheless, no correlation was found for expression levels of STMN1 and FANCC. The correlations of MELK and STMN1 and MELK and FANCC were further corroborated on TCGA database (r=0.213 and p=<0.0059, r=0.689 and p=<0.0001, respectively) and Rembrandt database (r=0.17 and p=<0.0083, r=0.414 and p=<0.0001, respectively). In summary, our results suggest that the three genes, STMN1, MELK, and FACC, participate in a signaling pathway related to cytoskeleton remodeling involved in cell cycle, and cell migration, both biologic functions relevant to tumor progression. Further functional studies are worthwhile to search for novel therapeutic targets in GBM. Citation Format: Fernanda Oliveira Serachi, Sr., Sueli Mieko Oba Shinjo, Sr. Role of MELK, STMN1, and FANCC in human astrocytoma [abstract]. In: Proceedings of the AACR International Conference held in cooperation with the Latin American Cooperative Oncology Group (LACOG) on Translational Cancer Medicine; May 4-6, 2017; São Paulo, Brazil. Philadelphia (PA): AACR; Clin Cancer Res 2018;24(1_Suppl):Abstract nr B68.