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Abstract Background Phaeochromocytomas and paragangliomas (PPGLs) are rare neuroendocrine tumours. Pathogenic variants have been identified in more than 15 susceptibility genes; associated tumours are grouped into three Clusters, reinforced by their transcriptional profiles. Cluster 1A PPGLs have pathogenic variants affecting enzymes of the tricarboxylic acid cycle, including succinate dehydrogenase. Within inherited PPGLs, these are the most common. PPGL tumours are known to undergo epigenetic reprograming, and here, we report on global histone post-translational modifications and DNA methylation levels, alongside clinical phenotypes. Results Out of the 25 histone post-translational modifications examined, Cluster 1A PPGLs were distinguished from other tumours by a decrease in hyper-acetylated peptides and an increase in H3K4me2. DNA methylation was compared between tumours from individuals who developed metastatic disease versus those that did not. The majority of differentially methylated sites identified tended to be completely methylated or unmethylated in non-metastatic tumours, with low inter-sample variance. Metastatic tumours by contrast consistently had an intermediate DNA methylation state, including the ephrin receptor EPHA4 and its ligand EFNA3. Gene expression analyses performed to identify genes involved in metastatic tumour behaviour pin-pointed a number of genes previously described as mis-regulated in Cluster 1A tumours, as well as highlighting the tumour suppressor RGS22 and the pituitary tumour-transforming gene PTTG1. Conclusions Combined transcriptomic and DNA methylation analyses revealed aberrant pathways, including ones that could be implicated in metastatic phenotypes and, for the first time, we report a decrease in hyper-acetylated histone marks in Cluster 1 PPGLs.

VHL disease is an inherited and autosomal dominant disorder affecting 1 in 36,0000 individuals worldwide. It is caused by von Hippel-Lindau (VHL) gene mutations and can affect both genders and all ethnic backgrounds (Nordstrom-O’Brien et al., 2009; Maher, 2004). Here, we generated and characterised two iPSC lines derived from patients with histopathologically confirmed clear cell renal cell carcinoma (ccRCC) and VHL Type 1 enrolled in the TRACERx Renal (TRAcking Renal Cell Carcinoma Evolution Through Therapy (Rx)). PBMCs were reprogrammed to pluripotency using a genome non-integrating Sendai virus (SeV) vectors protocol. Both human iPSC lines displayed normal morphology, expressed markers associated with stemness and differentiated into the three germ layers. The iPSC lines could be used as a disease-specific cellular model to understand further the inherited disorder of Type 1 von Hippel-Lindau (VHL) disease.

IntroductionThe percentage of patients diagnosed with pheochromocytoma and paraganglioma (altogether PPGL) carrying known germline mutations in one of the over fifteen susceptibility genes identified to date has dramatically increased during the last two decades, accounting for up to 35-40% of PPGL patients. Moreover, the application of NGS to the diagnosis of PPGL detects unexpected co-occurrences of pathogenic allelic variants in different susceptibility genes.MethodsHerein we uncover several cases with dual mutations in NF1 and other PPGL genes by targeted sequencing. We studied the molecular characteristics of the tumours with co-occurrent mutations, using omic tools to gain insight into the role of these events in tumour development.ResultsAmongst 23 patients carrying germline NF1 mutations, targeted sequencing revealed additional pathogenic germline variants in DLST (n=1) and MDH2 (n=2), and two somatic mutations in H3-3A and PRKAR1A. Three additional patients, with somatic mutations in NF1 were found carrying germline pathogenic mutations in SDHB or DLST, and a somatic truncating mutation in ATRX. Two of the cases with dual germline mutations showed multiple pheochromocytomas or extra-adrenal paragangliomas - an extremely rare clinical finding in NF1 patients. Transcriptional and methylation profiling and metabolite assessment showed an “intermediate signature” to suggest that both variants had a pathological role in tumour development.DiscussionIn conclusion, mutations affecting genes involved in different pathways (pseudohypoxic and receptor tyrosine kinase signalling) co-occurring in the same patient could provide a selective advantage for the development of PPGL, and explain the variable expressivity and incomplete penetrance observed in some patients.

Context: Differentiated thyroid cancer (DTC) is usually treated by thyroidectomy followed by radioiodine ablation and generally has a good prognosis. It may now be possible to limit the amount of treatment without impacting on efficacy. It is not known whether coexistent thyroiditis impacts on radioiodine uptake or on its potential efficacy, but this could provide a rationale for modification to current therapeutic protocols. Design: This was a retrospective cohort study of radioiodine uptake on imaging after radioiodine ablation for DTC in patients with and without concurrent thyroiditis. All patients with histologically confirmed DTC treated with radioiodine ablation after thyroidectomy in a single centre from 2012 to 2015 were included. The primary outcome assessed was the presence of low or no iodine uptake on post-ablation scan, as reported by a nuclear medicine physician blinded to the presence or absence of thyroiditis. Results: One hundred thirty patients with available histopathology results were included. Thyroiditis was identified in 42 post-operative specimens and 15 of these patients had low or no iodine uptake on post-ablation scan, compared to only 2 of 88 patients without thyroiditis (P < 0.0001) with further data analysis dividing the groups by ablation activity received (1100 MBq or 3000 MBq). Conclusions: Concurrent thyroiditis may impair the uptake of radioactive iodine in management of DTC. Given that patients with DTC and thyroiditis already have a good prognosis, adopting a more selective approach to this step in therapy may be indicated. Large, longitudinal studies would be required to determine if omitting radioactive iodine therapy from those patients with concurrent thyroiditis has a measurable impact on mortality from thyroid cancer.