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Human Cancer Genetics Programme

Hereditary Endocrine Cancer Group

Group Leader:  Mercedes Robledo
Research highlights
New mutation in a Krebs cycle-related gene in pheochromocytoma (PCC) or paraganglioma (PGL)

Mutations in Krebs cycle genes are frequently found in patients with PCC/PGL. Disruption of SDH, FH or MDH2 enzymatic activities leads to the accumulation of specific metabolites, which give rise to epigenetic changes in the genome that cause a characteristic hypermethylated phenotype. Tumours showing this phenotype, but no alterations in the known predisposing genes, could harbour mutations in other Krebs cycle genes. We used downregulation and methylation of RBP1, as a marker of a hypermethylation phenotype, to select PCCs and PGLs for targeted exome sequencing of a panel of Krebs cycle-related genes. Methylation profiling, metabolite assessment and additional analyses were also performed in selected cases. Following this rationale, a germline GOT2 variant, c.357A>T, was found in a patient with multiple tumours and metastasis. The presence of this variant was associated with higher tumour mRNA and protein expression levels, increased GOT2 enzymatic activity in lymphoblastic cells, and altered metabolite ratios both in tumours and in GOT2 knockdown HeLa cells transfected with the variant. Thus, we propose GOT2 as a new susceptibility gene related to metastatic PGL. This study further attests to the relevance of the Krebs cycle in the development of these tumours, and points to this central metabolic pathway as being targetable in metastatic PCC/PGL patients.

MDH2 mutations in pheochromocytoma and paraganglioma

MDH2 has recently been proposed by our Group as a novel PCC/ PGL susceptibility gene. We aimed to determine the prevalence of MDH2 mutations among PPGL patients and to establish the associated phenotype. Through a worldwide network we recruited close to 800 PCC/PGL patients. In this study we needed to implement a multidisciplinary approach with up to five functional assays and twenty in silico predictions (FIGURE), including MDH2 enzymatic activity and affinity, an immunofluorescence assay to evaluate MDH2 localisation, and a molecular dynamics (MD) simulation approach to examine the potential protein structure changes of the most controversial variants. With this initiative we were able to ascertain the prevalence of MDH2 mutations in PPGL in less than 1% and to highlight the importance of including this gene in the routine genetic screening of the disease, especially in metastatic, noradrenergic-secreting, multiple tumours and in young patients.

Novel CNVs in pharmacogenes and mTOR pathway mutations to predict outcomes in cancer therapies

Personalised cancer treatment is of enormous clinical and social relevance since it can lead to safer and more efficient therapies. This year we focused on: i) understanding the effect of copy number variants (CNVs) on adverse drug reactions (ADRs); and ii) discovering biomarkers predictive of mTOR inhibitor response. ADRs cause around 6.5% of admissions to hospitals, accounting for 5-10% of the annual hospital costs. Genetic factors are responsible for many ADRs and these could be prevented by genetic tests. We performed the first systematic assessment of the CNV landscape in pharmacogenes by integrating data from 2,504 whole genomes and 59,898 exomes. We described novel exonic deletions and duplications in 97% of the genes analysed. Novel deletion frequencies ranged from singletons up to 1%, and accounted for >5% of all loss-of-function alleles in 42% of studied genes. Thus, CNVs are an additional source of pharmacogenetic variability with important implications for drug response and personalised therapy. Regarding mTOR inhibitors, by next generation sequencing (NGS) on tumour tissues, we found that those patients with somatic mutations directly activating the pathway (in MTOR, TSC1, TSC2) had improved responses. Furthermore, multiregion NGS allowed us to determine that when these mutations were acquired early on during tumour development they resulted in extraordinary responses to these drugs.