Charting the Genetic Landscape and Clonal Architectures of Pheochromocytoma

University dissertation from Uppsala : Acta Universitatis Upsaliensis

Abstract: Genotypic and phenotypic inter patient heterogeneity characterize pheochromocytoma and paraganglioma (PPGL). Up to 60% of PPGL are associated with either somatic or germline mutations in at least 14 established disease causing genes. Consequently, a comprehensive screening test for PPGL patients utilizing standard techniques is not feasible and in the diagnostic approach, multiple different phenotype guided gene prioritization protocols have been utilized. This may result in misdiagnosis, especially in patients with sporadic presentation. Diagnostic testing of somatic mutations in tumour material is not performed due to the lack of actionable results.The aims of this study were, (1) to investigate the use of novel sequencing techniques in a clinical application, (2) to discover novel PPGL disease causing loci using novel sequencing techniques, (3) to characterize a large cohort of PPGL for mutations in known disease causing genes and to analyse corresponding genotype-phenotype correlations, (4) to dissect the molecular and genetic landscape of MEN2 PPGL and (5) to determine the clonal architecture and heterogeneity within, and in-between matched PPGL.For these purposes we studied PPGL tumours from a total of 96 patients using targeted and/or whole exome enrichment, capillary and high throughput sequencing as well as genome wide array based genotyping. Novel bioinformatics pipelines were constructed for raw data processing and downstream interpretation. Quantitative PCR, western blot and immunohistochemistry were utilized in order to characterize molecular traits. Selected experimental findings were correlated to patient phenotype.We conclude that novel sequencing techniques could be utilized in clinical genetic screening of patients with PPGL. Somatic gain-of-function mutations in H-RAS are likely to contribute to disease pathogenesis. Analysing tumour DNA for somatic mutations in disease causing genes could provide relevant clinical information and have an impact on patient management. Concomitant mutations in PPGL may occur in exceptional cases and have a substantial impact on tumour biology and patient phenotype. And finally genetic heterogeneity is present between and within a majority of PPGL tumours.