Poster Presentation 5th International Symposium on Phaeochromocytoma and Paraganglioma 2017

Deciphering genetic aberrations through metabolic profiling: 2-Hydroxyglutarate elevations in phaeochromocytoma/paraganglioma (#76)

Susan Richter 1 , Laura Gieldon 2 , Anastasios Mangelis 1 , Mirko Peitzsch 1 , Felix Beuschlein 3 , Henri Timmers 4 , Massimo Mannelli 5 , Karel Pacak 6 , Daniela Aust 7 , Alberto Cascon 8 , Mercedes Robledo 8 , Klink Barbara 2 , Eisenhofer Graeme 1
  1. Department of Clinical Chemistry and Laboratory Medicine, Uniklinikum Dresden, The Dresden University of Technology, Dresden, Germany
  2. Institute for Clinical Genetics, Medical Faculty Carl Gustav Carus, Technische Universität Dresden, Dresden, Germany
  3. Medizinische Klinik and Poliklinik IV, Ludwig-Maximilians-Universität München, München, Germany
  4. Department of Medicine, Radboud University Nijmegen Medical Centre, Nijmegen, the Netherlands
  5. Department of Experimental and Clinical Biomedical Sciences "Mario Serio", University of Florence and Istituto Toscano Tumori, Florence, Italy
  6. Eunice Kennedy Shriver National Institute of Child Health and Human Development, National Institutes of Health, Bethesda, USA
  7. Institute of Pathology and UCC Tumor and Normal tissue bank, Uniklinikum Dresden, The Dresden University of Technology, Dresden, Deutschland
  8. Hereditary Endocrine Cancer Group, CNIO, Madrid, Spain

D- and L-2-hydroxyglutarate (D/L-2HG) are metabolites normally present in low concentrations. Elevations of D-2HG are attributed to gain-of-function mutations in isocitrate dehydrogenase 1 or 2 leading to excessive formation of D-2HG from α-ketoglutarate (αKG). Other reports identified that promiscuous activity of lactate dehydrogenase-A, malate dehydrogenases and phosphoglycerate dehydrogenase contribute to L-2HG production. Both enantiomers are able to inhibit αKG-dependent enzymes leading to pseudohypoxia and changes in genome methylation through effects on prolyl hydroxylases and histone/DNA demethylases. In phaeochromocytoma/paraganglioma (PPGL) only two patients with somatic IDH1 mutations have been described so far.

This study investigated the frequency of abnormal tissue formation of both 2HG enantiomers in PPGL. We established a database of central carbon metabolites for 397 PPGLs using LC-MS/MS. Outlier analysis of normalised 2HG values resulted in the top 31 samples being marked as outliers. These 31 PPGLs had various mutational backgrounds. Nine PPGLs with the highest 2HG levels were investigated in more detail (5 of these had unknown mutational status). Four of nine PPGLs showed not only high 2HG but also high αKG (#1,3,5,6). Tumour #1 (IDH1-mutation) and #2 had strong elevations of D-2HG relative to L-2HG (127- and 901-times, respectively), confirming the functionality of the IDH1 mutation. Tumour #3 and #4 showed 13-times higher L-2HG. In PPGL #3 the highest levels of glutamine were measured. Tumour #5 and #6 had the highest αKG to citrate ratios of the set.

Genetic testing by next generation sequencing using a panel of known cancer-associated genes and Krebs cycle related genes was used to relate identified metabolic profiles to genetic aberrations. Consistent with high D-2HG levels, tumour #2 carried an IDH2 mutation (c.514A>G, p.Arg172Gly). To our knowledge, the first one reported in PPGL.

Inexpensive metabolite profiling is a valid tool to guide mutation testing and screen for variants with potentially unknown significance.