Warburg Effect’s Manifestation in Aggressive Pheochromocytomas and Paragangliomas: Insights from a Mouse Cell Model Applied to Human Tumor Tissue

Abstract

A glycolytic profile unifies a group of pheochromocytomas and paragangliomas (PHEOs/PGLs) with distinct underlying gene defects, including von Hippel-Lindau (VHL) and succinate dehydrogenase B (SDHB) mutations. Nevertheless, their tumor aggressiveness is distinct: PHEOs/PGLs metastasize rarely in VHL-, but frequently in SDHB-patients. To date, the molecular mechanisms causing the more aggressive phenotype in SDHB-PHEOs/PGLs remain largely unknown. Recently, however, an excellent model to study aggressive PHEOs (mouse tumor tissue (MTT) cells) has been developed from mouse PHEO cells (MPC). We employed this model for a proteomics based approach to identify changes characteristic for tumor aggressiveness, which we then explored in a homogeneous set of human SDHB- and VHL-PHEOs/PGLs. The increase of glucose transporter 1 in VHL, and of hexokinase 2 in VHL and SDHB, confirmed their glycolytic profile. In agreement with the cell model and in support of decoupling of glycolysis, the Krebs cycle and oxidative phosphorylation (OXPHOS), SDHB tumors showed increased lactate dehydrogenase levels. In SDHB-PGLs OXPHOS complex activity was increased at complex III and, as expected, decreased at complex II. Moreover, protein and mRNA expression of all tested OXPHOS-related genes were higher in SDHB- than in VHL-derived tumors. Although there was no direct evidence for increased reactive oxygen species production, elevated superoxide dismutase 2 expression may reflect elevated oxidative stress in SDHB-derived PHEOs/PGLs. For the first time, we show that despite dysfunction in complex II and evidence for a glycolytic phenotype, the Warburg effect does not seem to fully apply to SDHB-PHEOs/PGLs with respect to decreased OXPHOS. In addition, we present evidence for increased LDHA and SOD2 expression in SDHB-PHEOs/PGLs, proteins that have been proposed as promising therapeutic targets in other cancers. This study provides new insight into pathogenic mechanisms in aggressive human PHEOs/PGLs, which may lead to identifying new diagnostic and prognostic markers in the near future.

Current views on cell metabolism in SDHx-related pheochromocytoma and paraganglioma

“…HIF-α enhances the glycolytic pathway by increasing target gene expression from GLUT-1, GLUT-3, through HK2 and pyruvate kinase variant M2 (PKM2) to lactate dehydrogenase-A (LDH-A) and other glycolytic and anabolic enzymes and metabolites (Osthus, et al 2000; Soga 2013). Some expression studies have not found over-expression of GLUT-1 in SDHx- related tumors (Favier et al 2009; Fliedner et al 2012; Lopez-Jimenez et al 2010). Moreover, increased expression of GLUT-3 and HK2 mRNAs observed in SDHx- related tumors (Favier et al 2009; Fliedner et al 2012) can explain the high sensitivity of [ 18 F]-FDG PET for SDHx -related tumors, mainly observed in SDHB -related PHEOs/PGLs (Taieb, et al 2009; Timmers, et al 2012; Timmers, et al 2009a; Timmers, et al 2007; Zelinka, et al 2008).…”

“…The activity of the ROS-generating enzyme Nox1 is required for vascular endothelial growth factor (VEGF), a potent stimulator of tumor angiogenesis (Dudkina, et al 2005; Pan, et al 2009; Rustin et al 2002; Slane et al 2006). Fliedner et al (Fliedner, et al 2012) detected elevated superoxide dismutase 2 expression in SDHB -derived PHEOs/PGLs that is indirect evidence for increased ROS production and may reflect elevated oxidative stress.…”

Reactive Oxygen Species in Normal and Tumor Stem Cells

“…Several recent studies have also shown that mutations in complex II subunits C and D increase O 2 •− and H 2 O 2 production as well as causing genomic instability and cancer induction (Aykin-Burns et al, 2011; Ishii et al, 1998; Owens et al, 2012; Slane et al, 2006). Furthermore, familial forms of two human cancers (paraganglioma and pheochromocytoma) have also been found to be associated with mutations in genes coding for subunits B, C, and D in complex II (Fliedner et al, 2012; Gimm, Armanios, Dziema, Neumann, & Eng, 2000). In addition, the mitochondria of malignant human tumor cells and rodent tumors have been shown to exhibit histological abnormalities characterized by unusual arrangements of mitochondrial cristae, mitochondrial hypertrophy, and mitochondrial fragmentation when compared to normal human cells (Bize, Oberley, & Morris, 1980; Springer, 1980).…”

Pheochromocytoma: Gasping for Air

“…HIF enhances the glycolytic pathway by increasing expression of target genes involved in the glycolytic and anabolic processes (glucose transporters (GLUT) 1, 2, hexokinase (HK) 2, PKM2, LDH-A) [34,140]. Although some studies have not found the GLUT1 overexpression of SDHx-related tumors [29, 135,141], there was still increased expression of GLUT3 and HK2 as well as LDH-A [135,141]. Functional analysis of PHEO/PGL tumor tissue from patients with somatic HIF2A and germline PHD1 and PHD2 mutations also showed increased activity of the HIF signaling pathway and its target gene expression, including GLUT1 and in PHD-mutated PHEOs/PGLs also LDH-A [12,16].…”