Priority Research Papers:
MYCN and HDAC5 transcriptionally repress CD9 to trigger invasion and metastasis in neuroblastoma
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Abstract
Johannes Fabian1,2, Desirée Opitz1, Kristina Althoff3, Marco Lodrini1,4, Barbara Hero5, Ruth Volland5, Anneleen Beckers6,7, Katleen de Preter6,7, Anneleen Decock6,7, Nitin Patil8, Mohammed Abba8, Annette Kopp-Schneider9, Kathy Astrahantseff4, Jasmin Wünschel1,4, Sebastian Pfeil4, Maria Ercu4, Annette Künkele4, Jamie Hu1,10, Theresa Thole1,4, Leonille Schweizer11, Gunhild Mechtersheimer12, Daniel Carter13, Belamy B. Cheung13, Odilia Popanda14, Andreas von Deimling11,15, Jan Koster16, Rogier Versteeg16, Manfred Schwab17, Glenn M. Marshall13,18, Frank Speleman6,7, Ulrike Erb19, Margot Zoeller19, Heike Allgayer8, Thorsten Simon5, Matthias Fischer5,20,21, Andreas E. Kulozik22, Angelika Eggert4, Olaf Witt1,22, Johannes H. Schulte4 and Hedwig E. Deubzer1,4,22,23
1 Clinical Cooperation Unit Pediatric Oncology, German Cancer Research Center (DKFZ) and German Consortium for Translational Cancer Research (DKTK), INF, Heidelberg, Germany
2 Phenex Pharmaceuticals AG, Waldhofer Straße, Heidelberg, Germany
3 Department of Pediatric Hematology and Oncology, University Children’s Hospital Essen, Essen, Germany
4 Department of Pediatric Hematology, Oncology and SCT, Charité - University Hospital Berlin, Campus Virchow-Klinikum, Berlin, Germany
5 Department of Pediatric Hematology and Oncology, University of Cologne, Cologne, Germany
6 Center for Medical Genetics Ghent, Ghent University, De Pintelaan, Ghent, Belgium
7 Cancer Research Institute Ghent, De Pintelaan, Ghent, Belgium
8 Department of Experimental Surgery, Medical Faculty Mannheim, University of Heidelberg, Centre for Biomedicine and Medical Technology, Mannheim, Germany
9 Department of Biostatistics, German Cancer Research Center (DKFZ), INF, Heidelberg, Germany
10 Yale University, New Haven, CT
11 Department of Neuropathology, University of Heidelberg, INF, Heidelberg, Germany
12 Department of Pathology, University of Heidelberg, INF, Heidelberg, Germany
13 Children’s Cancer Institute, UNSW, Randwick, NSW, Australia
14 Division of Epigenomics and Cancer Risk Factors, DKFZ, INF, Heidelberg, Germany
15 Clinical Cooperation Unit Neuropathology, DKFZ, INF, Heidelberg, Germany
16 Department of Oncogenomics and Emma Children’s Hospital, Academic Medical Center, University of Amster- dam, Meibergdreef, Amsterdam, the Netherlands
17 Neuroblastoma Genetics, DKFZ, INF, Heidelberg, Germany
18 Kids Cancer Centre, Sydney Children’s Hospital Randwick, Randwick, NSW, Australia
19 Experimental Surgery and Tumor Cell Biology, University of Heidelberg, INF, Heidelberg, Germany
20 Center for Molecular Medicine Cologne, University of Cologne, Cologne, Germany
21 Max Plank Institute for Metabolism Research, Cologne, Germany
22 Department of Pediatric Hematology and Oncology, Heidelberg University, INF, Heidelberg, Germany
23 Junior Neuroblastoma Research Group, Experimental and Clinical Research Center of the Max-Delbrück Center for Molecular Medicine in the Helmholtz Community and the Charité - University Medicine Berlin, Lindenberger Weg, Berlin, Germany
Correspondence to:
Hedwig E. Deubzer, email:
Keywords: antimetastatic therapy, chromatin modulation, histone deacetylases, grainyhead-like transcription factor family, tetraspanin family
Received: April 29, 2016 Accepted: August 24, 2016 Published: August 27, 2016
Abstract
The systemic and resistant nature of metastatic neuroblastoma renders it largely incurable with current multimodal treatment. Clinical progression stems mainly from the increasing burden of metastatic colonization. Therapeutically inhibiting the migration-invasion-metastasis cascade would be of great benefit, but the mechanisms driving this cycle are as yet poorly understood. In-depth transcriptome analyses and ChIP-qPCR identified the cell surface glycoprotein, CD9, as a major downstream player and direct target of the recently described GRHL1 tumor suppressor. CD9 is known to block or facilitate cancer cell motility and metastasis dependent upon entity. High-level CD9 expression in primary neuroblastomas correlated with patient survival and established markers for favorable disease. Low-level CD9 expression was an independent risk factor for adverse outcome. MYCN and HDAC5 colocalized to the CD9 promoter and repressed transcription. CD9 expression diminished with progressive tumor development in the TH-MYCN transgenic mouse model for neuroblastoma, and CD9 expression in neuroblastic tumors was far below that in ganglia from wildtype mice. Primary neuroblastomas lacking MYCN amplifications displayed differential CD9 promoter methylation in methyl-CpG-binding domain sequencing analyses, and high-level methylation was associated with advanced stage disease, supporting epigenetic regulation. Inducing CD9 expression in a SH-EP cell model inhibited migration and invasion in Boyden chamber assays. Enforced CD9 expression in neuroblastoma cells transplanted onto chicken chorioallantoic membranes strongly reduced metastasis to embryonic bone marrow. Combined treatment of neuroblastoma cells with HDAC/DNA methyltransferase inhibitors synergistically induced CD9 expression despite hypoxic, metabolic or cytotoxic stress. Our results show CD9 is a critical and indirectly druggable suppressor of the invasion-metastasis cycle in neuroblastoma.
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PII: 11662