Paragangliomas and phaeochromocytomas are neuroendocrine tumours whose pathogenesis and progression are very strongly influenced by genetics. A germline mutation in one of the susceptibility genes ...identified so far explains ∼40% of all cases; the remaining 60% are thought to be sporadic cases. At least one-third of these sporadic tumours contain a somatic mutation in a predisposing gene. Genetic testing, which is indicated in every patient, is guided by the clinical presentation as well as by the secretory phenotype and the immunohistochemical characterization of the tumours. The diagnosis of an inherited form drives clinical management and tumour surveillance. Different 'omics' profiling methods have provided a neat classification of these tumours in accordance with their genetic background. Transcriptomic studies have identified two main molecular pathways that underlie development of these tumours, one in which the hypoxic pathway is activated (cluster 1) and another in which the MAPK and mTOR (mammalian target of rapamycin) signalling pathways are activated (cluster 2). DNA methylation profiling has uncovered a hypermethylator phenotype in tumours related to SDHx genes (a group of genes comprising SDHA, SDHB, SDHC, SDHD and SDHAF2) and revealed that succinate acts as an oncometabolite, inhibiting 2-oxoglutarate-dependent dioxygenases, such as hypoxia-inducible factor prolyl-hydroxylases and histone and DNA demethylases. 'Omics' data have suggested new therapeutic targets for patients with a malignant tumour. In the near future, new 'omics'-based tests are likely to be transferred into clinical practice with the goal of establishing personalized medical management for affected patients.
Paragangliomas are neuroendocrine tumors frequently associated with mutations in RET, NF1, VHL, and succinate dehydrogenase (SDHx) genes. Methylome analysis of a large paraganglioma cohort identified ...three stable clusters, associated with distinct clinical features and mutational status. SDHx-related tumors displayed a hypermethylator phenotype, associated with downregulation of key genes involved in neuroendocrine differentiation. Succinate accumulation in SDH-deficient mouse chromaffin cells led to DNA hypermethylation by inhibition of 2-OG-dependent histone and DNA demethylases and established a migratory phenotype reversed by decitabine treatment. Epigenetic silencing was particularly severe in SDHB-mutated tumors, potentially explaining their malignancy. Finally, inactivating FH mutations were identified in the only hypermethylated tumor without SDHx mutations. These findings emphasize the interplay between the Krebs cycle, epigenomic changes, and cancer.
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•SDH and FH mutations establish a hypermethylator phenotype in PGL/PCC•Succinate inhibits DNA and histone demethylases in SDH-deficient chromaffin cells•DNA hypermethylation silences key genes involved in neuroendocrine differentiation•A stronger hypermethylator phenotype may explain malignancy of SDHB-mutated tumors
Low-grade oncocytic renal tumor (LOT) is an emerging provisional entity, described as rare solid renal oncocytic/eosinophilic tumor sharing diffuse CK7 and negative CD117 immunoprofile. The links ...between LOT and other eosinophilic chromophobe like-renal cell carcinomas (RCC) are currently discussed. We sequenced tumoral DNA with a next generation sequencing panel for kidney cancer and carried out immunohistochemical analyses with CK7, CD117, SDHB, 4EBP1-P, S6K-P, and FOXI1 antibodies in a series of ten cases of LOT (9 females, 1 male; mean age at surgery: 66 years, 42.3 to 83.4) retrospectively diagnosed from a cohort of 272 tumors initially classified as chromophobe RCC (CHRCC). All LOT were single, without known hereditary predisposition, classified stage pT1 (70%), pT2 (20%) or pT3a (10%). Morphological features were similar to previous descriptions and clinical behavior was indolent for the six cases with available follow-up. We identified genetic variations in mTOR pathway related genes in 80% of cases, MTOR (7 cases) or TSC1 (1 case). Expression of FOXI1 was absent in all cases. In 9 LOT, 4EBP1-P and S6K-P were overexpressed, suggesting mTOR pathway activation.Our data highlights the major role of mTOR pathway in tumorigenesis of LOT mostly due to activating MTOR gene variations. Absence of FOXI1 expression is a strong argument to distinguish LOT from eosinophilic CHRCC and to bring them closer to other recently described FOXI1 negative eosinophilic-CHRCC like with MTOR/TSC mutations. Altogether, our data argue to consider LOT as a distinct entity with a favorable clinical outcome. However, in case of metastasis, an accurate diagnosis of LOT would be essential for the patient's management and could allow targeted therapy.
The tricarboxylic acid (TCA) cycle is a central metabolic pathway responsible for supplying reducing potential for oxidative phosphorylation and anabolic substrates for cell growth, repair and ...proliferation. As such it thought to be essential for cell proliferation and tissue homeostasis. However, since the initial report of an inactivating mutation in the TCA cycle enzyme complex, succinate dehydrogenase (SDH) in paraganglioma (PGL), it has become clear that some cells and tissues are not only able to survive with a truncated TCA cycle, but that they are also able of supporting proliferative phenotype observed in tumours. Here, we show that loss of SDH activity leads to changes in the metabolism of non-essential amino acids. In particular, we demonstrate that pyruvate carboxylase is essential to re-supply the depleted pool of aspartate in SDH-deficient cells. Our results demonstrate that the loss of SDH reduces the metabolic plasticity of cells, suggesting vulnerabilities that can be targeted therapeutically.
Pheochromocytomas and paragangliomas (PPGLs) are rare neuroendocrine tumors. Whereas most PPGLs are benign, up to 20% may become metastatic with
- and
-mutated tumors showing the higher risk. We ...aimed at determining the contribution of immortalization mechanisms to metastatic progression.
Immortalization mechanisms were investigated in 200 tumors. To identify telomerase (+) tumors, we analyzed genomic alterations leading to transcriptional activation of
comprising promoter mutations, hypermethylation and gain copy number. To identify tumors that activated the alternative lengthening of telomere (ALT) mechanism, we combined analyses of telomere length by slot blot, telomere heterogeneity by telomere FISH, and
mutations by next-generation sequencing. Univariate/multivariate and metastasis-free survival (MFS) and overall survival (OS) analyses were carried out for assessment of risk factors and clinical outcomes.
Only 37 of 200 (18.5%) tumors achieved immortalization. Telomerase activation occurred in 12 metastatic tumors and was prevalent in
-mutated paragangliomas (
= 2.42e-09). ALT features were present in 25 tumors, mostly pheochromocytomas, regardless of metastatic status or molecular group (
= 0.169), yet
mutations were found preferentially in
-mutated metastatic tumors (
= 0.0014). Telomerase activation and
mutations were independent factors of poor prognosis: MFS (hazard ratio, 48.2 and 33.1;
= 6.50E-07 and 1.90E-07, respectively); OS (hazard ratio, 97.4 and 44.1;
= 4.30E-03 and 2.00E-03, respectively) and were associated with worse MFS and OS (log-rank tests
< 0.0001).
Assessment of telomerase activation and
mutations could be used to identify metastatic PPGLs, particularly in tumors at high risk of progression.
Succinate dehydrogenase (SDH) inhibitors (SDHIs) are used worldwide to limit the proliferation of molds on plants and plant products. However, as SDH, also known as respiratory chain (RC) complex II, ...is a universal component of mitochondria from living organisms, highly conserved through evolution, the specificity of these inhibitors toward fungi warrants investigation. We first establish that the human, honeybee, earthworm and fungal SDHs are all sensitive to the eight SDHIs tested, albeit with varying IC50 values, generally in the micromolar range. In addition to SDH, we observed that five of the SDHIs, mostly from the latest generation, inhibit the activity of RC complex III. Finally, we show that the provision of glucose ad libitum in the cell culture medium, while simultaneously providing sufficient ATP and reducing power for antioxidant enzymes through glycolysis, allows the growth of RC-deficient cells, fully masking the deleterious effect of SDHIs. As a result, when glutamine is the major carbon source, the presence of SDHIs leads to time-dependent cell death. This process is significantly accelerated in fibroblasts derived from patients with neurological or neurodegenerative diseases due to RC impairment (encephalopathy originating from a partial SDH defect) and/or hypersensitivity to oxidative insults (Friedreich ataxia, familial Alzheimer's disease).
Pheochromocytomas and paragangliomas are neuroendocrine tumors that occur in the context of inherited cancer syndromes in ∼30% of cases and are linked to germline mutations in the VHL, RET, NF1, ...SDHA, SDHB, SDHC, SDHD, SDHAF2 and TMEM127 genes. Although genome-wide expression studies have revealed some of the mechanisms likely to be involved in pheochromocytoma/paraganglioma tumorigenesis, the complete molecular distinction of all subtypes of hereditary tumors has not been solved and the genetic events involved in the generation of sporadic tumors are unknown. With these purposes in mind, we investigated 202 pheochromocytomas/paragangliomas, including 75 hereditary tumors, using expression profiling, BAC array comparative genomic hybridization and somatic mutation screening. Gene expression signatures defined the hereditary tumors according to their genotype and notably, led to a complete subseparation between SDHx- and VHL-related tumors. In tumor tissues, the systematic characterization of somatic genetic events associated with germline mutations in tumor suppressor genes revealed loss of heterozygosity (LOH) in a majority of cases, but also detected point mutations and copy-neutral LOH. Finally, guided by transcriptome classifications and LOH profiles, somatic mutations in VHL or RET genes were identified in 14% of sporadic pheochromocytomas/paragangliomas. Overall, we found a germline or somatic genetic alteration in 45.5% (92/202) of the tumors in this large series of pheochromocytomas/paragangliomas. Regarding mutated genes, specific molecular pathways involved in tumorigenesis mechanisms are identified. Altogether, these new findings suggest that somatic mutation analysis is likely to yield important clues for personalizing molecular targeted therapies.
Comprehensive genetic analyses have identified germline
and
gene mutations as predominant causes of metastatic paraganglioma and pheochromocytoma. However, some suspicious cases remain unexplained. ...In this study, we performed whole-exome sequencing of a paraganglioma exhibiting an
-like molecular profile in the absence of
or
mutations and identified a germline mutation in the
gene, which encodes the mitochondrial 2-oxoglutarate/malate carrier. Germline
mutations were identified in six other patients, five of whom had metastatic disease. These mutations were associated with loss of heterozygosity, suggesting that
acts as a tumor-suppressor gene. Pseudohypoxic and hypermethylator phenotypes comparable with those described in
- and
-related tumors were observed both in tumors with mutated
and in
immortalized mouse chromaffin knockout cells generated by CRISPR-Cas9 technology. These data show that
is a novel paraganglioma susceptibility gene for which loss of function correlates with metastatic presentation.
A gene encoding a mitochondrial carrier is implicated in a hereditary cancer predisposition syndrome, expanding the role of mitochondrial dysfunction in paraganglioma.
.
The development of pheochromocytomas and paragangliomas is strongly linked to the presence of germline mutations in more than 15 predisposing genes. Among them, germline and somatic VHL mutations ...account for ~10% of all cases. In contrast with SDHA and SDHB immunohistochemistries that are routinely used to validate SDHx gene mutations, there is no such tool available for VHL mutations. The aim of this study was to evaluate whether CA9 immunostaining could be used as a tool to predict the presence or validate the pathogenicity of VHL gene mutations in paraganglioma. Immunohistochemistry for CA9 was performed on 207 tumors. A retrospective series of 100 paragangliomas with known mutation status for paraganglioma susceptibility genes was first investigated. Then, a prospective series of 107 paragangliomas was investigated for CA9 immunostaining followed by germline and/or somatic genetic testing of all paraganglioma susceptibility genes by next-generation sequencing. Cytosolic CA9 protein expression was heterogeneous in the different samples. However, we observed that a membranous CA9 staining was almost exclusively observed in VHL-related cases. Forty two of 48 (88%) VHL-mutated samples showed a CA9 membranous immunostaining. Positive cells were either isolated, varying from 1 or 2 cells (5% of cases) to 10-20 cells per tumor block (35% of cases), grouped in areas of focal positivity representing between 1 and 20% of the tissue section (35% of cases), or widely distributed on 80-100% of the tumor sections (25% of samples). In contrast, 142/159 (91%) of non-VHL-mutated tumors presented no membrane CA9 localization. Our results demonstrate that VHL gene mutations can be predicted or validated reliably by an easy-to-perform and low-cost immunohistochemical procedure. CA9 immunohistochemistry on paragangliomas will improve the diagnosis of VHL-related disease, which is important for the surveillance and therapeutic management of paraganglioma patients, and in case of germline mutation, their family members.