Genetic, histologic, and molecular characterisation of pheochromocytomas and paragangliomas : an integrated approach

Paragangliomas and pheochromocytomas (PPGL) are neuroendocrine tumours arising from extra-adrenal paraganglia tissues and adrenal glands. They often release catecholamines, which are at the origin of the main symptoms. Half of them are due to a germline mutation in one of the known genes, and somatic mutations have been identified in 30% additional PPGL in different genes. One issue is malignancy, since there is no reliable marker to predict the tumour behaviour and no established treatment. This work had two main objectives: search for somatic mutations in known genes, and for genetic markers to predict metastatic dissemination. We identified somatic mutations in half of the sequenced tumours, and a minority of tumours carrying a large number of mutations. In a second time, we observed a consistent overexpression of Contactin 4 in malignant PPGL using different techniques. We also showed that the overexpression of Contactin 4 gives a survival benefit to a cell line derived from a human PGL, hPheo1. In conclusion, comprehensive characterisation of PPGL should continue to lead to more personalised care.(BIFA - Sciences biomédicales et pharmaceutiques) -- UCL, 201

ADDED VALUE OF CATECHOLAMINE PHENOTYPING AND GENETIC SCREENING FOR THE CHARACTERIZATION OF PHEOCHROMOCYTOMA: A BELGIAN MULTICENTRE COHORT

Objective: Pheochromocytomas/paragangliomas (PPGL) are rare neuroendocrine tumors arising from chromaffin cells of the adrenal medulla or from neural-crest derived sympathetic tissue. They secrete catecholamines in varying amounts, which accounts for their symptomatology. Hypertension is the most common sign, found in approximately 95% of patients. Nowadays, biochemical testing remains the simplest and most widely available method to assess PPGL in first intention. Identification of mutations in known susceptibility genes may help further refining tumor characterization and prognosis assessment. The objective of this study was to analyze clinical characteristics of PPGL from Belgium according to their secretory and genetic profiles. Design and method: We retrospectively analyzed a cohort of 120 cases of non-syndromic PPGL diagnosed in 19 Belgian centres. Clinical characteristics were correlated with three catecholamine phenotypes based on urinary metanephrines (noradrenergic, adrenergic and silent) and with the results of genetic screening. Results: Our cohort included a majority of women (59%). The mean age at diagnosis was 47 ± 7 years. We documented the prevalence of pediatric (7.5%), extra-adrenal (13.3%), bilateral (2.9%), multifocal (5.1%), recurrent (8.5%), metastatic (7.6%) and familial (5.9%) cases. While the yield of positive genetic screening was low (15%), the presence of at least one of these criteria was associated with a 4-fold increased prevalence of mutation (30.3 vs 8.5%; p = 0.03). In the subset in whom metanephrines values were available (n = 62), the prevalence of adrenergic, noradrenergic and silent phenotypes was 58%, 34% and 8% respectively. Patients belonging to these three subsets differed by age at diagnosis (51 ± 14; 38 ± 17; 42 ± 13 years; p = 0.009), the proportion of extra-adrenal tumors (2.8%; 9.5%; 40%; p = 0.016), pediatric (2.8%; 19%; 0%; p = 0.074) and familial (0%; 18.8%; 0%; p = 0.041) cases. Finally, an excess of dopamine secretion was associated with a higher prevalence of metastatic disease (25% vs 1.9%; p = 0.005). Conclusions: Secretory phenotypes and genetic profiles deserve to be integrated in the preoperative characterization of PPGL and may help orienting management and follow-up. Simple clinical features are associated with an increased probability of inherited tumor. Identification of an increased dopamine secretion should raise the suspicion of metastatic PPGL

Toward an improved definition of the genetic and tumor spectrum associated with SDH germ-line mutations

The tricarboxylic acid, or Krebs, cycle is central to the cellular metabolism of sugars, lipids, and amino acids; it fuels the mitochondrial respiratory chain for energy generation. In the past decade, mutations in the Krebs-cycle enzymes succinate dehydrogenase, fumarate hydratase, and isocitrate dehydrogenase have been documented to be causally involved in carcinogenesis. This review is focused on the relationship between SDH mutations and the carcinogenic phenotype. The succinate dehydrogenase complex catalyzes the oxidation of succinate to fumarate; mutations in its subunits SDHA, SDHB, SDHC, and SDHD, and in the assembly factor SDHAF2, result in syndromes with distinct tumor types, including pheochromocytoma/paraganglioma, gastrointestinal stromal tumor, and, less often, renal-cell carcinoma and pituitary adenoma. In this study we collected all previously reported SDH mutations with the aim of defining their nature and tumor spectrum. In addition, genotype-phenotype correlations as well as mechanisms of biallelic inactivation were analyzed in the SDH-deficient setting. Finally, we performed bioinformatics analysis using SIFT, Polyphen2, and Mutation Assessor to predict the functional impact of nonsynonymous mutations. The prediction of the latter was further compared with available SDHA and/or SDHB immunohistochemistry data

STK39 and WNK1 Are Potential Hypertension Susceptibility Genes in the BELHYPGEN Cohort

The serine/threonine kinase With-No-Lysine (K) Kinase 1 (WNK1) activates the thiazide-sensitive Na/Cl cotransporter through phosphorylation of STE20/SPS1-related proline/alanine-rich kinase, another serine/threonine kinase encoded by STK39. The aim of this study was to look for association between WNK1 and STK39 gene variants, and blood pressure (BP) and hypertension.Seven hundred seventy-nine Caucasian hypertensive patients (HYP) recruited in 6 academic centers from Belgium, and 906 normotensive (NT) controls were genotyped for 5 single nucleotide polymorphisms-rs3754777, rs6749447, rs35929607 (STK39), rs1468326, and rs765250 (WNK1)-using the Snapshot method.The rare TT genotype at the rs3754777 locus (STK39) was overrepresented in HYP versus NT (7.3% vs 3.0%, P = 0.0002). In the whole study population, the multivariable-adjusted odds ratio (OR) for having hypertension associated with the TT genotype was 5.9 (95% confidence interval: 2.2-15.6), and systolic BP was 10 mm Hg higher in TT compared with wild-type subjects (140.1 vs 130.4 mm Hg, P = 0.002). Similarly, the AA genotype at the rs1468326 locus (WNK1) was twice as frequent in HYP versus NT (5.5% vs 2.3%, P < 0.0001), and associated with an increased adjusted OR of hypertension (4.1; 1.5-11.7) and a higher systolic BP (139.8 vs 130.1 mm Hg, P = 0.003). In the whole cohort, a dose-dependent increase in systolic BP was observed according to the number of at-risk genotypes (0: 129.8 mm Hg; 1: 133.0 mm Hg; 2: 149.3 mm Hg, P = 0.02).Single nucleotide polymorphisms rs3754777 (STK39) and rs1468326 (WNK1) were associated with hypertension and BP in our multicenter Belgian case-control study, which supports the role of STK39 and WNK1 as potential hypertension susceptibility genes. Replication in different clinical settings and study of other candidate loci belonging to the same molecular pathway is warranted.status: publishe

STK39 and WNK1 Are Potential Hypertension Susceptibility Genes in the BELHYPGEN Cohort

The serine/threonine kinase With-No-Lysine (K) Kinase 1 (WNK1) activates the thiazide-sensitive Na+/Cl− cotransporter through phosphorylation of STE20/SPS1-related proline/alanine-rich kinase, another serine/threonine kinase encoded by STK39. The aim of this study was to look for association between WNK1 and STK39 gene variants, and blood pressure (BP) and hypertension. Seven hundred seventy-nine Caucasian hypertensive patients (HYP) recruited in 6 academic centers from Belgium, and 906 normotensive (NT) controls were genotyped for 5 single nucleotide polymorphisms—rs3754777, rs6749447, rs35929607 (STK39), rs1468326, and rs765250 (WNK1)—using the Snapshot method. The rare TT genotype at the rs3754777 locus (STK39) was overrepresented in HYP versus NT (7.3% vs 3.0%, P = 0.0002). In the whole study population, the multivariable-adjusted odds ratio (OR) for having hypertension associated with the TT genotype was 5.9 (95% confidence interval: 2.2–15.6), and systolic BP was 10 mm Hg higher in TT compared with wild-type subjects (140.1 vs 130.4 mm Hg, P = 0.002). Similarly, the AA genotype at the rs1468326 locus (WNK1) was twice as frequent in HYP versus NT (5.5% vs 2.3%, P < 0.0001), and associated with an increased adjusted OR of hypertension (4.1; 1.5–11.7) and a higher systolic BP (139.8 vs 130.1 mm Hg, P = 0.003). In the whole cohort, a dose-dependent increase in systolic BP was observed according to the number of at-risk genotypes (0: 129.8 mm Hg; 1: 133.0 mm Hg; 2: 149.3 mm Hg, P = 0.02). Single nucleotide polymorphisms rs3754777 (STK39) and rs1468326 (WNK1) were associated with hypertension and BP in our multicenter Belgian case-control study, which supports the role of STK39 and WNK1 as potential hypertension susceptibility genes. Replication in different clinical settings and study of other candidate loci belonging to the same molecular pathway is warranted

Severity of COVID-19 among hospitalized patients : Omicron remains a severe threat for immunocompromised hosts

The Omicron variant of severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) emerged in the general population in the context of a relatively high immunity gained through the early waves of coronavirus disease 19 (COVID-19), and vaccination campaigns. Despite this context, a significant number of patients were hospitalized, and identifying the risk factors associated with severe disease in the Omicron era is critical for targeting further preventive, and curative interventions. We retrospectively analyzed the individual medical records of 1501 SARS-CoV-2 positive hospitalized patients between 13 December 2021, and 13 February 2022, in Belgium, of which 187 (12.5%) were infected with Delta, and 1036 (69.0%) with Omicron. Unvaccinated adults showed an increased risk of moderate/severe/critical/fatal COVID-19 (crude OR 1.54; 95% CI 1.09–2.16) compared to vaccinated patients, whether infected with Omicron or Delta. In adults infected with Omicron and moderate/severe/critical/fatal COVID-19 (n = 323), immunocompromised patients showed an increased risk of in-hospital mortality related to COVID-19 (adjusted OR 2.42; 95% CI 1.39–4.22), compared to non-immunocompromised patients. The upcoming impact of the pandemic will be defined by evolving viral variants, and the immune system status of the population. The observations support that, in the context of an intrinsically less virulent variant, vaccination and underlying patient immunity remain the main drivers of severe disease

9B.09: Identification of markers predictive for malignant behaviour of Pheochromocytomas and paragangliomas

OBJECTIVE: Pheochromocytomas and paragangliomas (PPGL) are relatively rare and mostly benign tumours. Approximately 10% of PPGL are malignant, as defined by the presence of metastases, i.e chromaffin tissue at a location that usually does not contain chromaffin cells. However, up to 35% of tumours in patients carrying an SDHB mutation appears to be malignant. Nowadays, no reliable marker allows to predict whether a PPGL is, or will become malignant. In addition, there are no curative treatments if metastases occur. The aim of the present study was to dentify genetic markers allowing to distinguish benign from malignant tumours. DESIGN AND METHOD: An mRNA expression array was performed on benign and malignant PPGL. The genes showing a different expression between the benign and malignant tumours were selected to be confirmed and validated by qRT-PCR. Finally, the remaining genes were stained by immunohistochemistry on Tissue MicroArray including a large series of PPGL. RESULTS: Forty benign and 12 malignant PPGL were investigated for differences in mRNA expression with Affymetrix arrays. Expression data were normalized according to Affymetrix recommendations. Then, using Pomelo II (http://pomelo2.bioinfo.cnio.es/), a Limma t-test was performed, to assess which genes were differentially expressed between benign and malignant PPGL. First, a non-clustered analysis was performed and 10 genes with a False Discovery Rate (FDR) below 0.05 and a relative overexpression ratio of at least 4 were found, including Interleukin 13 Receptor alpha 2 (IL13RA2) and Monooxygenase DBH-like 1 (MOXD1). Secondly, a supervised cluster analysis was performed (based on HIF target genes), resulting in 2 groups, which were both investigated for differences in mRNA expression between benign and malignant tumours. Five genes showed an FDR below 0.01 and were overexpressed in malignant tumours with a ratio higher than 4, including Contactin 4 (CNTN4), Iroquois Homeobox 3 (IRX3), and Sulfatase 2 (SULF2). These genes were further investigated using qRT-PCR, and immunohistochemistry on Tissue Micro Array including 91 benign and 12 malignant PPGL. CONCLUSIONS: Significant overexpression of Contactin 4 was shown in malignant compared to benign tumours, and may therefore contribute to distinguish malignant from benign PPGL