Hermine Mohr1,2, Simone Ballke3, Nicole Bechmann4,5, Sebastian Gulde1,2, Jaber Malekzadeh-Najafabadi6, Mirko Peitzsch5, Vasilis Ntziachristos6,7, Katja Steiger3, Tobias Wiedemann1,2, Natalia S Pellegata1,2. 1. Institute for Diabetes and Cancer, Helmholtz Centre Munich, Ingolstaedter Landstr.1, 85764 Neuherberg, Germany. 2. Joint Heidelberg-IDC Translational Diabetes Program, Heidelberg University Hospital, 69120 Heidelberg, Germany. 3. Institute of Pathology, School of Medicine, Technical University Munich, Trogerstr. 18, 81675 Munich, Germany. 4. Department of Medicine III, University Hospital Carl Gustav Carus, Technical University Dresden, Fetscherstrasse 74, 01307 Dresden, Germany. 5. Institute of Clinical Chemistry and Laboratory, University Hospital Carl Gustav Carus, Technical University Dresden, Fetscherstrasse 74, 01307 Dresden, Germany. 6. Chair of Biological Imaging, Technical University of Munich, Ismaninger Straße 22, 81675 Munich, Germany. 7. Institute for Biomedical Imaging, Helmholtz Centre Munich, Ingolstaedter Landstr.1, 85764 Neuherberg, Germany.
Abstract
BACKGROUND: Pseudohypoxic tumors activate pro-oncogenic pathways typically associated with severe hypoxia even when sufficient oxygen is present, leading to highly aggressive tumors. Prime examples are pseudohypoxic pheochromocytomas and paragangliomas (p-PPGLs), neuroendendocrine tumors currently lacking effective therapy. Previous attempts to generate mouse models for p-PPGLs all failed. Here, we describe that the rat MENX line, carrying a Cdkn1b (p27) frameshift-mutation, spontaneously develops pseudohypoxic pheochromocytoma (p-PCC). METHODS: We compared rat p-PCCs with their cognate human tumors at different levels: histology, immunohistochemistry, catecholamine profiling, electron microscopy, transcriptome and metabolome. The vessel architecture and angiogenic potential of pheochromocytomas (PCCs) was analyzed by light-sheet fluorescence microscopy ex vivo and multi-spectral optoacoustic tomography (MSOT) in vivo. RESULTS: The analysis of tissues at various stages, from hyperplasia to advanced grades, allowed us to correlate tumor characteristics with progression. Pathological changes affecting the mitochrondrial ultrastructure where present already in hyperplasias. Rat PCCs secreted high levels of norepinephrine and dopamine. Transcriptomic and metabolomic analysis revealed changes in oxidative phosphorylation that aggravated over time, leading to an accumulation of the oncometabolite 2-hydroxyglutarate, and to hypermethylation, evident by the loss of the epigenetic mark 5-hmC. While rat PCC xenografts showed high oxygenation, induced by massive neoangiogenesis, rat primary PCC transcriptomes possessed a pseudohypoxic signature of high Hif2a, Vegfa, and low Pnmt expression, thereby clustering with human p-PPGL. CONCLUSION: Endogenous rat PCCs recapitulate key phenotypic features of human p-PPGLs. Thus, MENX rats emerge as the best available animal model of these aggressive tumors. Our study provides evidence of a link between cell cycle dysregulation and pseudohypoxia.
BACKGROUND:Pseudohypoxic tumors activate pro-oncogenic pathways typically associated with severe hypoxia even when sufficient oxygen is present, leading to highly aggressive tumors. Prime examples are pseudohypoxic pheochromocytomas and paragangliomas (p-PPGLs), neuroendendocrine tumors currently lacking effective therapy. Previous attempts to generate mouse models for p-PPGLs all failed. Here, we describe that the rat MENX line, carrying a Cdkn1b (p27) frameshift-mutation, spontaneously develops pseudohypoxic pheochromocytoma (p-PCC). METHODS: We compared rat p-PCCs with their cognate humantumors at different levels: histology, immunohistochemistry, catecholamine profiling, electron microscopy, transcriptome and metabolome. The vessel architecture and angiogenic potential of pheochromocytomas (PCCs) was analyzed by light-sheet fluorescence microscopy ex vivo and multi-spectral optoacoustic tomography (MSOT) in vivo. RESULTS: The analysis of tissues at various stages, from hyperplasia to advanced grades, allowed us to correlate tumor characteristics with progression. Pathological changes affecting the mitochrondrial ultrastructure where present already in hyperplasias. Rat PCCs secreted high levels of norepinephrine and dopamine. Transcriptomic and metabolomic analysis revealed changes in oxidative phosphorylation that aggravated over time, leading to an accumulation of the oncometabolite 2-hydroxyglutarate, and to hypermethylation, evident by the loss of the epigenetic mark 5-hmC. While ratPCC xenografts showed high oxygenation, induced by massive neoangiogenesis, rat primary PCC transcriptomes possessed a pseudohypoxic signature of high Hif2a, Vegfa, and low Pnmt expression, thereby clustering with humanp-PPGL. CONCLUSION: Endogenous rat PCCs recapitulate key phenotypic features of humanp-PPGLs. Thus, MENX rats emerge as the best available animal model of these aggressive tumors. Our study provides evidence of a link between cell cycle dysregulation and pseudohypoxia.
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