Genotype-specific Differences in the Expression of Markers of Warburg Effect in Pheochromocytoma and Paraganglioma

Program: Abstracts - Orals, Featured Poster Presentations, and Posters
Session: MON 37-82-Pheochromocytoma & Paraganglioma
Monday, June 17, 2013: 1:45 PM-3:45 PM
Expo Halls ABC (Moscone Center)

Poster Board MON-45
Jyotsna Upendra Rao*1, Tuna Demir2, Benno Kusters3, Egbert Oosterwijk3, Ron Wevers3, Ad R.M.M. Hermus4, Arjen Mensenkamp5, Henricus PM Kunst4, Fred CG J Sweep1 and Henri JLM Timmers3
1Radboud Univ Nijmegen Med Ctr, Nijmegen, Netherlands, 2Radboud University Nijmegen Medical Centre, Nijmegen, Netherlands, 3Radboud University Nijmegen Medical Center, Nijmegen, Netherlands, 4Radboud University Nijmegen Medical Centre, Nijmegen, 5Radboud University Nijmegen Medical Center
Background: Pheochromocytomas and paragangliomas (PGLs) are neuroendocrine tumors of sympathetic and parasympathetic paraganglia. At least 30% of the PGLs are caused by germline mutations in ten identified tumor susceptibility genes viz., RET, NF1, VHL, SDHA/B/C/D/AF-2 (succinate dehydrogenase subunits A, B, C, D and assembly factor 2), TMEM127 and MAX. In tumors with SDHx mutations, the ability of cells for oxidative phosphorylation is compromised and there is accumulation of succinate resulting in stabilization of hypoxia-inducible factors (HIF) -1α and -2α. Upon nuclear translocation, together with HIF-1β, they form an active HIF complex that induces the expression of genes with hypoxia response elements which include genes involved in glycolysis and angiogenesis. Thus, in these tumors, the pseudo-hypoxic drive is hypothesized to mediate an increase in aerobic glycolysis, also known as Warburg effect.

Objective: The aim of the study was to examine if genotype specific differences exist in the expression of Hexokinases 1, 2 and 3, GLUT-1 and -3, vascular endothelial growth factor (VEGF) and carbonic anhydrase 9 (CA-9).

Methods: 30 primary PGLs collected from patients with hereditary mutations in SDHB (n=2), SDHD (n=4), VHL (n=2), RET (n=5), NF1 (n=1), MAX (n=1) and sporadic patients (n=15) were investigated. Patients whose genotype was tested negative for SDHA/B/C/D/AF-2, RET and VHL were considered as sporadics. Paraffin embedded sections of these tumors were investigated using immunohistochemical methods with diaminobenzidine as read-out. The staining pattern was assessed by an experienced pathologist and a score was given to each sample after multiplication of percentage of positively stained area with staining intensity (scale:0-4 from 0-absent to 4-strong).

Results: Hexokinase 1 and 2 showed a granular staining pattern typical of mitochondria associated proteins while Hexokinase 3 and VEGF showed cytoplasmic staining. GLUT-1 and -3 also showed a predominantly cytoplasmic staining with around 5-20% of the cells showing membrane staining. Further, increased expression of Hexokinase 2 and VEGF were observed in SDH related tumors when compared to sporadic and RET, NF1 and MAX tumors. No evidence for the expression of CA-9 was observed in the PGLs examined except in one VHL tumor where a membrane staining was observed.

Conclusions: Results indicate an evidence for increased glycolysis and angiogenesis at molecular level in the SDH related tumors.

Nothing to Disclose: JU, TD, BK, EO, RW, ARMMH, AM, HPK, FCJS, HJT

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Sources of Research Support: European Union Seventh Framework Programme (FP7/2007-2013) under grant agreement no. 259735 (ENSAT CANCER)