Recurrent Germline DLST Mutations in Individuals with Multiple Pheochromocytomas and Paragangliomas

Am J Hum Genet. 2019 Apr 4;104(4):651-664. doi: 10.1016/j.ajhg.2019.02.017.

Laura Remacha ¹, David Pirman ², Christopher E Mahoney ², Javier Coloma ³, Bruna Calsina ¹, Maria Currás-Freixes ¹, Rocío Letón ¹, Rafael Torres-Pérez ¹, Susan Richter ⁴, Guillermo Pita ⁵, Belén Herráez ⁵, Giovanni Cianchetta ², Emiliano Honrado ⁶, Lorena Maestre ⁷, Miguel Urioste ⁸, Javier Aller ⁹, Óscar García-Uriarte ¹⁰, María Ángeles Gálvez ¹¹, Raúl M Luque ¹², Marcos Lahera ¹³, Cristina Moreno-Rengel ¹⁴, Graeme Eisenhofer ⁴, Cristina Montero-Conde ¹, Cristina Rodríguez-Antona ¹⁵, Óscar Llorca ³, Gromoslaw A Smolen ², Mercedes Robledo ¹⁵, Alberto Cascón ¹⁶

Affiliations

1 Hereditary Endocrine Cancer Group, Spanish National Cancer Research Centre, Madrid, Madrid 28029, Spain.
2 Agios Pharmaceuticals, 88 Sidney Street, Cambridge, MA 02139, USA.
3 Structural Biology Programme, Spanish National Cancer Research Centre, Madrid, Madrid 28029, Spain.
4 Institute of Clinical Chemistry and Laboratory Medicine, University Hospital Carl Gustav Carus, Medical Faculty Carl Gustav Carus, Technische Universität Dresden, Dresden, Freistaat Sachsen 01069, Germany.
5 Human Genotyping Unit-CeGen, Human Cancer Genetics Programme, Spanish National Cancer Research Centre, Madrid, Madrid 28029, Spain.
6 Anatomical Pathology Service, Hospital of León, León, Castilla y León 24071, Spain.
7 Monoclonal Antibodies Unit, Biotechnology Programme, Spanish National Cancer Research Centre, Madrid, Madrid 28029, Spain.
8 Familial Cancer Clinical Unit, Spanish National Cancer Research Centre, Madrid, Madrid 28029, Spain.
9 Department of Endocrinology, University Hospital Puerta de Hierro, Majadahonda, Madrid 28222, Spain.
10 Nephrology Department, University Hospital of Araba, Vitoria, País Vasco 01009, Spain.
11 Service of Endocrinology and Nutrition, University Hospital Reina Sofía, Córdoba, Andalucía 14004, Spain; Maimónides Institute of Biomedical Research of Cordoba, Córdoba, Andalucía 14004, Spain.
12 Hormones and Cancer Group, Maimónides Institute of Biomedical Research of Córdoba, Córdoba, Andalucía 14004, Spain.
13 Endocrinology and Nutrition Department, La Princesa University Hospital, Madrid, Madrid 28006, Spain.
14 Department of Endocrinology and Nutrition, University Hospital of Basurto, Bilbao 48013, Spain.
15 Hereditary Endocrine Cancer Group, Spanish National Cancer Research Centre, Madrid, Madrid 28029, Spain; Centro de Investigación Biomédica en Red de Enfermedades Raras, Madrid, Madrid 28029, Spain.
16 Hereditary Endocrine Cancer Group, Spanish National Cancer Research Centre, Madrid, Madrid 28029, Spain; Centro de Investigación Biomédica en Red de Enfermedades Raras, Madrid, Madrid 28029, Spain. Electronic address: acascon@cnio.es.

PMID: 30929736 DOI: 10.1016/j.ajhg.2019.02.017

Abstract

Pheochromocytomas and paragangliomas (PPGLs) provide some of the clearest genetic evidence for the critical role of metabolism in the tumorigenesis process. Approximately 40% of PPGLs are caused by driver germline mutations in 16 known susceptibility genes, and approximately half of these genes encode members of the tricarboxylic acid (TCA) cycle. Taking as a starting point the involvement of the TCA cycle in PPGL development, we aimed to identify unreported mutations that occurred in genes involved in this key metabolic pathway and that could explain the phenotypes of additional individuals who lack mutations in known susceptibility genes. To accomplish this, we applied a targeted Sequencing of 37 TCA-cycle-related genes to DNA from 104 PPGL-affected individuals with no mutations in the major known predisposing genes. We also performed omics-based analyses, TCA-related metabolite determination, and ¹³C₅-glutamate labeling assays. We identified five germline variants affecting DLST in eight unrelated individuals (∼7%); all except one were diagnosed with multiple PPGLs. A recurrent variant, c.1121G>A (p.Gly374Glu), found in four of the eight individuals triggered accumulation of 2-hydroxyglutarate, both in tumors and in a heterologous cell-based assay designed to functionally evaluate DLST variants. p.Gly374Glu-DLST tumors exhibited loss of heterozygosity, and their methylation and expression profiles are similar to those of EPAS1-mutated PPGLs; this similarity suggests a link between DLST disruption and pseudohypoxia. Moreover, we found positive DLST immunostaining exclusively in tumors carrying TCA-cycle or EPAS1 mutations. In summary, this study reveals DLST as a PPGL-susceptibility gene and further strengthens the relevance of the TCA cycle in PPGL development.

Keywords

DLST; TCA cycle; cancer susceptibility gene; paraganglioma; pheochromocytoma.

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