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Abstract Disclosure: L.S. Santana: None. A.G. Guimaraes: None. F. Freitas-Castro: None. A.W. Maciel: None. G.F. Fagundes: None. M.A. Pereira: None. L.F. Drager: None. L.A. Bortolotto: None. M.C. Fragoso: None. B.B. Mendonca: None. A. Latronico: None. M.Q. Almeida: None. Primary aldosteronism (PA) is the most common cause of endocrine hypertension, with an estimated prevalence of 10-20% in patients referred to tertiary hospitals. The most common causes of PA are bilateral adrenal hyperplasia (BAH) and aldosteronomas. Significant advances in elucidating the pathogenesis of PA have been made over the past decade. Pathogenic/likely pathogenic germline allelic variants in KCNJ5, CACNA1D, CACNA1H, CLCN2, PDE2A, and PDE3B genes have already been documented in individuals diagnosed with PA. However, most of the cases still lack a defined genetic etiology. Genomic investigation of these individuals through whole-exome sequencing (WES) would enable the identification of causal monogenic genotype-phenotype associations in genes not yet linked to bilateral PA. In our study, thirty-one patients were selected, including 29 probands and 2 relatives, with a confirmed diagnosis of bilateral PA by adrenal venous sampling. All probands were previously investigated for KCNJ5 pathogenic/likely pathogenic germline variants. Additionally, all cases also had negative genetic screening for glucocorticoid-remediable PA. WES was performed using BGI Genomics technology, DNA nanoball sequencing, on the DNBSEQ platform. Raw data were processed (bioinformatics) using in-house pipelines. The 31 sequenced samples showed excellent coverage metrics, with coverage ranging from 200-467X and %>10X at 99%. Each patient presented an average of 150,000 variants. After the gene/allelic prioritization workflow, 4 heterozygous SNVs were listed as candidates in 3 out of 29 patients (10.34%). Two of the variants were identified in rare but previously documented germline causes of PA: CACNA1H and CACNA1D (patients 13 and 29, respectively). Patient 4 (hypertension diagnosed at 36 years of age) presented 2 variants in possible new candidate genes, CASZ1 and STRN. The NM_001079843.3(CASZ1):c.1912G>C/p.(Asp638His) variant caught our attention not only for its absence in population genomic databases (gnomAD v.4), but also due to the association between CASZ1 locus and PA. The zinc finger transcription factor CASZ1 has CACNA1D as one of its targets and participates in key biological processes for PA pathogenesis, such as: co-repression of mineralocorticoid receptor transcriptional activity; mediation of tissue response to aldosterone and differential expression in aldosterone-producing cell clusters (APCC) and in zona glomerulosa; its hypoexpression promotes aldosterone-dependent mineralocorticoid receptor transcriptional activity and hyperexpression suppresses aldosterone biosynthesis by adrenal cells. In conclusion, an extremely rare CASZ1 variant was identified in a woman with bilateral PA. The functional characterization of the CASZ1 p.(Asp638His) variant might reinforce its role as a new monogenic cause of PA. Presentation: 6/3/2024

Abstract Disclosure: L.S. Santana: None. A.G. Guimaraes: None. F. Freitas-Castro: None. A.W. Maciel: None. G.F. Fagundes: None. M.A. Pereira: None. L.F. Drager: None. L.A. Bortolotto: None. M.C. Fragoso: None. B.B. Mendonca: None. A. Latronico: None. M.Q. Almeida: None. Primary aldosteronism (PA) is the most common cause of endocrine hypertension, with an estimated prevalence of 10-20% in patients referred to tertiary hospitals. The most common causes of PA are bilateral adrenal hyperplasia (BAH) and aldosteronomas. Significant advances in elucidating the pathogenesis of PA have been made over the past decade. Pathogenic/likely pathogenic germline allelic variants in KCNJ5, CACNA1D, CACNA1H, CLCN2, PDE2A, and PDE3B genes have already been documented in individuals diagnosed with PA. However, most of the cases still lack a defined genetic etiology. Genomic investigation of these individuals through whole-exome sequencing (WES) would enable the identification of causal monogenic genotype-phenotype associations in genes not yet linked to bilateral PA. In our study, thirty-one patients were selected, including 29 probands and 2 relatives, with a confirmed diagnosis of bilateral PA by adrenal venous sampling. All probands were previously investigated for KCNJ5 pathogenic/likely pathogenic germline variants. Additionally, all cases also had negative genetic screening for glucocorticoid-remediable PA. WES was performed using BGI Genomics technology, DNA nanoball sequencing, on the DNBSEQ platform. Raw data were processed (bioinformatics) using in-house pipelines. The 31 sequenced samples showed excellent coverage metrics, with coverage ranging from 200-467X and %>10X at 99%. Each patient presented an average of 150,000 variants. After the gene/allelic prioritization workflow, 4 heterozygous SNVs were listed as candidates in 3 out of 29 patients (10.34%). Two of the variants were identified in rare but previously documented germline causes of PA: CACNA1H and CACNA1D (patients 13 and 29, respectively). Patient 4 (hypertension diagnosed at 36 years of age) presented 2 variants in possible new candidate genes, CASZ1 and STRN. The NM_001079843.3(CASZ1):c.1912G>C/p.(Asp638His) variant caught our attention not only for its absence in population genomic databases (gnomAD v.4), but also due to the association between CASZ1 locus and PA. The zinc finger transcription factor CASZ1 has CACNA1D as one of its targets and participates in key biological processes for PA pathogenesis, such as: co-repression of mineralocorticoid receptor transcriptional activity; mediation of tissue response to aldosterone and differential expression in aldosterone-producing cell clusters (APCC) and in zona glomerulosa; its hypoexpression promotes aldosterone-dependent mineralocorticoid receptor transcriptional activity and hyperexpression suppresses aldosterone biosynthesis by adrenal cells. In conclusion, an extremely rare CASZ1 variant was identified in a woman with bilateral PA. The functional characterization of the CASZ1 p.(Asp638His) variant might reinforce its role as a new monogenic cause of PA. Presentation: 6/3/2024

Abstract Disclosure: F. Freitas-Castro: None. L.S. Santana: None. G.F. Fagundes: None. A.F. Afonso: None. E.C. Lobato: None. F.L. Ledesma: None. I.C. Soares: None. B.B. Mendonca: None. M.C. Fragoso: None. A. Latronico: None. C.A. Stratakis: None. M.Q. Almeida: None. Background: Carney-Stratakis syndrome (CSS, OMIM #606864), also reported as “paraganglioma and gastrointestinal stromal tumor syndrome” or Carney-Stratakis dyad, is an autosomal dominant rare syndrome with incomplete penetrance, characterized by the association of paragangliomas (PGL) and/or pheochromocytomas (PHEO) and gastrointestinal stromal tumors (GIST). CSS is mainly caused by germline heterozygous pathogenic variants (PVs) in the succinate dehydrogenase subunit genes (SDHB, SDHC, SDHD), with SDHB and SDHD being the most frequently affected. Aim: To investigate a novel genetic etiology for CSS not associated with germline SDHx defects. Methods: Genetic investigation using SANGER sequencing and multiplex ligation-dependent probe amplification (MLPA) rule out germline defects for SDHx in a 59-year-old woman diagnosed with a 9 cm left PHEO, 4.8 cm PGL and 9.3 cm GIST. Thus, we performed whole exome sequencing of germline DNA (paired with tumor DNA from PHEO, PGL, and GIST) and applied a targeted analysis with the enrichment of 3,485 potential neuroendocrine tumors susceptibility genes associated with cellular response to hypoxia, mitochondrion, Krebs cycle, and tumorigenesis (MAPK, mTOR, ERK, and Wnt signaling). Additionally, we performed RT-qPCR to determine SLC25A11 expression levels in PHEO and PGL samples, stratified into three groups: tumors from the case harboring the SLC25A11 variant, Cluster 1 (n= 4), and Cluster 2 (n= 8). Tumor DNA methylation status was assessed using immunostaining for 5-hydroxymethylcytosine (5hmC). Results: Exome sequencing revealed a new heterozygous germline variant (c.293G>A / p.Arg98His) in the mitochondrial 2-oxoglutarate/malate carrier gene (SLC25A11). This variant, located in a highly conserved residue of the SLC25A11 mitochondrial carrier domain, is predicted to be deleterious in silico (REVEL score= 0.81). Exome sequencing of the PHEO and PGL did not reveal somatic PVs in genes previously associated with PHEO and PGL. Moreover, somatic c-KIT and PDGFRA PVs were not identified in GIST. Notably, a significant downregulation of SLC25A11 expression was observed in tumor samples harboring the SLC25A11 p.Arg98His variant (0.57 ± 0.13) compared with tumors from cluster 1 (1.39 ± 0.45; p = 0.025) and cluster 2 (1.79 ± 0.71; p < 0.001). Interestingly, immunostaining for 5hmC was negative in all tumors (PHEO, PGL and GIST), indicating tumor hypermethylation. Conclusion: A rare germline deleterious variant in SLC25A11 was identified in a patient with CSS. Moreover, the absence of somatic drivers, hypermethylation status and loss of SLC25A11 expression in the CSS tumors support the involvement of this gene with CSS. Support: Sao Paulo Research Foundation (FAPESP) grant 2019/15873-6 (to M.Q.A.), and FAPESP post-doctoral fellowship 2021/11240-9 (to F.F-C.). Presentation: 6/1/2024

Abstract Disclosure: F. Freitas-Castro: None. L.S. Santana: None. G.F. Fagundes: None. A.F. Afonso: None. E.C. Lobato: None. F.L. Ledesma: None. I.C. Soares: None. B.B. Mendonca: None. M.C. Fragoso: None. A. Latronico: None. C.A. Stratakis: None. M.Q. Almeida: None. Background: Carney-Stratakis syndrome (CSS, OMIM #606864), also reported as “paraganglioma and gastrointestinal stromal tumor syndrome” or Carney-Stratakis dyad, is an autosomal dominant rare syndrome with incomplete penetrance, characterized by the association of paragangliomas (PGL) and/or pheochromocytomas (PHEO) and gastrointestinal stromal tumors (GIST). CSS is mainly caused by germline heterozygous pathogenic variants (PVs) in the succinate dehydrogenase subunit genes (SDHB, SDHC, SDHD), with SDHB and SDHD being the most frequently affected. Aim: To investigate a novel genetic etiology for CSS not associated with germline SDHx defects. Methods: Genetic investigation using SANGER sequencing and multiplex ligation-dependent probe amplification (MLPA) rule out germline defects for SDHx in a 59-year-old woman diagnosed with a 9 cm left PHEO, 4.8 cm PGL and 9.3 cm GIST. Thus, we performed whole exome sequencing of germline DNA (paired with tumor DNA from PHEO, PGL, and GIST) and applied a targeted analysis with the enrichment of 3,485 potential neuroendocrine tumors susceptibility genes associated with cellular response to hypoxia, mitochondrion, Krebs cycle, and tumorigenesis (MAPK, mTOR, ERK, and Wnt signaling). Additionally, we performed RT-qPCR to determine SLC25A11 expression levels in PHEO and PGL samples, stratified into three groups: tumors from the case harboring the SLC25A11 variant, Cluster 1 (n= 4), and Cluster 2 (n= 8). Tumor DNA methylation status was assessed using immunostaining for 5-hydroxymethylcytosine (5hmC). Results: Exome sequencing revealed a new heterozygous germline variant (c.293G>A / p.Arg98His) in the mitochondrial 2-oxoglutarate/malate carrier gene (SLC25A11). This variant, located in a highly conserved residue of the SLC25A11 mitochondrial carrier domain, is predicted to be deleterious in silico (REVEL score= 0.81). Exome sequencing of the PHEO and PGL did not reveal somatic PVs in genes previously associated with PHEO and PGL. Moreover, somatic c-KIT and PDGFRA PVs were not identified in GIST. Notably, a significant downregulation of SLC25A11 expression was observed in tumor samples harboring the SLC25A11 p.Arg98His variant (0.57 ± 0.13) compared with tumors from cluster 1 (1.39 ± 0.45; p = 0.025) and cluster 2 (1.79 ± 0.71; p < 0.001). Interestingly, immunostaining for 5hmC was negative in all tumors (PHEO, PGL and GIST), indicating tumor hypermethylation. Conclusion: A rare germline deleterious variant in SLC25A11 was identified in a patient with CSS. Moreover, the absence of somatic drivers, hypermethylation status and loss of SLC25A11 expression in the CSS tumors support the involvement of this gene with CSS. Support: Sao Paulo Research Foundation (FAPESP) grant 2019/15873-6 (to M.Q.A.), and FAPESP post-doctoral fellowship 2021/11240-9 (to F.F-C.). Presentation: 6/1/2024

Abstract Disclosure: L.M. José: None. M. Ataide: None. F. Aratani: None. S. Beeby: None. G.F. Fagundes: None. F. Coelho: None. F. Ledesma: None. A. Latronico: None. B.B. Mendonca: None. M.C. Fragoso: None. M.Q. Almeida: None. Background: Obstructive sleep apnea (OSA) represents an independent risk factor for cardiovascular disease, and it is associated with a wide range of conditions, such as resistant hypertension and primary aldosteronism (PA). Patients with sleep apnea have a higher sympathetic activity during wakefulness due to chronic hypoxia. We present the case of pseudopheochromocytoma caused by severe OSA in an obese patient with RH and PA.Clinical Case: A 58-year-old female Brazilian patient was referred to an endocrinologist to investigate a 1.8 cm right adrenal nodule at computed tomography (CT) performed to investigate abdominal pain. Her medical history includes resistant hypertension diagnosed at 27 years of age, type 2 diabetes, dyslipidemia, Class II obesity, a previous acute myocardial infarction, and severe OSA (apnea-hypopnea index 61.6/hour). She has used irregularly the continuous positive airway pressure (CPAP) therapy at night. She is currently using carvedilol 50 mg bid, chlorthalidone 25 mg, olmesartan 40 mg, amlodipine 10 mg, spironolactone 50 mg, clonidine 0.2 mg tid, hydralazine 50 mg every 6 hours, acetylsalicylic acid 100 mg, atorvastatin 80 mg, and metformin XR 1000 mg. At physical examination, she had BMI 37 kg/m2, BP 150 x 90 mmHg, HR 64 bpm, and no signs of Cushing syndrome. Abdominal CT revealed a right adrenal nodule measuring 1.8 cm with 18 Hounsfield unit (HU) in the pre-contrast phase, reaching a density of 117 HU at the arterial phase and showing an absolute washout of 65% and a relative washout of 56%. A nodular thickening of 1.4 cm (-4 HU) at the left adrenal gland was also displayed at CT scan. Hormone evaluation showed basal cortisol 7 µg/dL, ACTH 15.7 pg/mL, DHEAS 1152 ng/mL (189-2050 ng/mL), 1 mg overnight dexamethasone suppression test 2.3 ug/dL, aldosterone 9.9 ng/dL, direct renin concentration 36.8 uIU/mL, plasma metanephrines 0.3 nmol/L (<0.5 nmol/L) and plasma normetanephrines 1.3 nmol/L (<0.9 nmol/L). It was not possible to change anti-hypertensive medications to investigate PA. A 0.3 mg clonidine suppression test did not suppress normetanephrine levels, confirming the biochemical diagnosis of pheochromocytoma. Then, alpha-blockade with doxazosin 4 mg was started and the patient underwent right adrenalectomy. Pathological examination revealed an adenoma with immunohistochemistry positive for CYP11B2. BP control was achieved with only two medications and aldosterone decreased to 3 ng/dL confirming PA. Plasma normetanephrine levels remained elevated post-surgery and only normalized after the CPAP regular use.Clinical lessons: Chronic activation of sympathetic autonomic drive in OSA can increase norepinephrine and normetanephrine levels. OSA is a very prevalent disease and it is frequently associated with RH. Therefore, OSA should be considered a possible cause of pseudopheochromocytoma in hypertensive patients. Presentation: 6/3/2024

Abstract Disclosure: F. Freitas-Castro: None. G.F. Fagundes: None. L.S. Santana: None. A.F. Afonso: None. F.L. Ledesma: None. I.C. Soares: None. B.B. Mendonca: None. A. Latronico: None. M.Q. Almeida: None. Background: Pheochromocytomas and paragangliomas (PPGLs) are associated with germline genetic defects in 30-50% of cases and are categorized into three transcriptional clusters. Cluster 1, subdivided into Clusters 1A and 1B, involves genes associated with cellular pseudohypoxia. Cluster 1A encompasses mutations in genes encoding Krebs cycle enzymes (e.g., succinate dehydrogenase complex subunits SDHA, SDHB, SDHC, and SDHD), leading to the accumulation of oncometabolites such as succinate and pyruvate, which increase the stabilization of hypoxia-inducible factor 2-alpha (HIF-2α). Aim: To investigate novel genetic etiologies in patients with PPGLs. Methods: Whole-exome sequencing (WES) of paired germline and tumor DNA was performed on 50 patients with PPGLs who lacked germline defects in previously known susceptibility genes. A targeted analysis enriched for 3,485 genes involved in cellular responses to hypoxia, mitochondrial function, the Krebs cycle, and tumorigenesis was conducted. Results: A germline c.280A>T (p.Lys94*) variant in the Pyruvate Dehydrogenase Phosphatase Catalytic Subunit 2 (PDP2) gene was identified in a 43-year-old female diagnosed with a 10.5 cm abdominal PGL. This stop-codon variant is absent from population databases and has been classified as a variant of uncertain significance. Tumor WES revealed no oncogenic variants. Additionally, immunohistochemistry for SDHB was positive in the tumor. The patient had no family history of cancer. Among her six siblings, one was tested for the variant, with a negative result. She has no children, and her parents are deceased. PDP2 gene encodes an enzyme responsible for dephosphorylating and reactivating the E1 alpha subunit of the pyruvate dehydrogenase complex (PDC), a critical regulator of mitochondrial metabolism. While the PDC has been implicated in glucose metabolism and tumor aggression in other cancers, such as neuroblastoma, it has not been previously associated with PPGLs. No specific phenotype has been associated with this gene. Conclusion: We report, for the first time, a rare loss-of-function PDP2 variant in a patient with abdominal PGL, which may contribute to HIF-2α stabilization and the pathogenesis of PPGL. Ongoing functional studies aim to further investigate this hypothesis. Support: Sao Paulo Research Foundation (FAPESP) grant 2019/15873-6 (to M.Q.A.), and FAPESP post-doctoral fellowship 2021/11240-9 (to F.F-C.). Presentation: Sunday, July 13, 2025

Disclosure: T. Castanheira de Freitas: None. A.W. Maciel: None. G.F. Fagundes: None. J. Petenuci: None. L. Santana: None. A. Guimaraes: None. F. Freitas-Castro: None. V. Srougi: None. F. Tanno: None. J. Chambo: None. M. Pereira: None. F. Coelho: None. L.P. Brito: None. A. Cavalcante: None. B. Pilan: None. F. Carnevale: None. A. Pio-Abreu: None. L. Bortolotto: None. A. Latronico: None. M.C. Fragoso: None. L. Drager: None. B.B. Mendonca: None. M.Q. Almeida: None. Background: Confirmatory tests represent a fundamental step in primary aldosteronism (PA) diagnosis, but they are laborious and often require a hospital environment due to the risks involved. The most common tests are the saline infusion test (SIT), captopril chalenging test (CCT), fludrocortisone supression test (FST) and furosemide upright test (FUT). The sodium overloading tests are the most frequently used tests in clinical practice, but are not tolerated in patients with severe hypertension, hypervolemia and hypokalemia. The use of oral furosemide test as a new confirmatory test to simplify the confirmation of PA diagnosis was not investigated. Methods: We prospectively evaluated the diagnostic performance of oral 80 mg furosemide in 64 PA patients and in 22 with primary hypertension (controls). Direct renin concentration (DRC) was measured before, 2h and 3h after the oral furosemide. In addition, the oral furosemide test was compared with two other confirmatory tests: FUT and SIT or CCT in all PA patients. Results: The cut-off of 7.6 µU/mL for DRC at 2h after oral furosemide had a sensitivity of 92%, a specificity of 82% and an accuracy of 90% for PA diagnosis. The cut-off of 10 µU/mL at 2h had a higher sensitivity of 95.3%, but a lower specificity of 73%. In five out of six controls with low-renin hypertension, which might represent a PA spectrum, renin remained suppressed. Excluding these six controls with low-renin hypertension, the DRC cut-off of 10 µU/mL at 2h after oral furosemide had a sensitivity of 95.3%, a specificity of 93.7% and an accuracy of 95% for PA diagnosis. DRC after 3h of oral furosemide did not improve diagnostic performance. Using the cut-off of 10 µU/mL, the oral furosemide test and the FUT were concordant in 62 out of 64 (97%) PA patients. Only 4 out of 64 PA cases (6.4%) ended the oral furosemide test with potassium <3.5 mEq/L. Hypotension was not evidenced in any PA patient during the test. Conclusion: The oral furosemide test was safe, well-tolerated and represents an effective strategy for PA investigation.Support: Sao Paulo Research Foundation (FAPESP) grant 2019/15873-6 (to M.Q. Almeida) and by the Coordination of Superior Level Staff Improvement (CAPES) fellowship (to TCF). Presentation: Friday, June 16, 2023

Disclosure: A.W. Maciel: None. T. Freitas: None. D.L. Danilovic: None. G.F. Fagundes: None. F. Freitas-Castro: None. L. Santana: None. A. Guimaraes: None. A. Pio-Abreu: None. J. V. Silveira: None. F. Consolim-Colombo: None. L. Bortolotto: None. M.C. Fragoso: None. A. Latronico: None. L. Drager: None. B.B. Mendonca: None. A.O. Hoff: None. M.Q. Almeida: None. Introduction: Aldosterone excess can cause oxidative stress leading to DNA damage in vitro and in vivo. Single case reports demonstrated a coincidence of primary aldosteronism (PA) with different malignancies. A higher prevalence of thyroid nodules and non-toxic multinodular goiter was described in patients with PA compared to those with essential hypertension (HT). A single study showed an association between PA and papillary thyroid cancer (PTC), but without a paired control group. Objective: To assess PA prevalence in a transversal cohort of patients with PTC and HT compared to a paired control group with HT. Methods: In this cross-sectional case-control study, PA was investigated in all patients with PTC and HT (n= 114), regardless of HT severity, under active surveillance at a cancer institute from 2019 to 2022. The control group included 228 (2:1) age-, sex- and body mass index (BMI)-matched individuals from a retrospective cohort of HT previously investigated for PA from 2011 to 2022. Serum aldosterone and plasma direct renin concentrations were measured by a chemiluminescent immunoassay. A positive PA screening was defined by aldosterone ≥10 ng/dL and aldosterone to renin ratio ≥2 ng/dL/μUI/mL. Results: Age, sex and BMI were not statistically different between PTC and control groups, respectively (age 59.8 ± 12 vs. 58.9 ± 12.3 yrs, p= 0.67; 79% vs. 81% women, p= 0.67; BMI 30.7 ± 5.8 vs. 30.8 ± 6.5 Kg/m2, p= 0.98). PA was diagnosed in 35 out of 114 PTC patients with HT. The prevalence of PA in the PTC group (30.5%, confidence interval (IC) 22.6%-40.1%) was significantly higher when compared to the paired control group with HT (11.84%, CI 8.08%-16.93%; p< 0.0001). Although PA prevalence was higher in the PTC group, only 20.2% had stage 3/resistant HT (vs. 38% in the control group, p= 0.003). The number of anti-hypertensive medications was lower in the PTC group compared to controls (2 drugs, 1 to 3 vs. 4 drugs, 3 to 5, respectively; p< 0.001). When analyzing only PA patients in both groups, frequency of stage 3/resistant HT and number of medications were lower in the PTC group (p< 0.001 and p< 0.001, respectively). Although HT was more severe in PA patients without PTC, aldosterone and renin levels were not different in PA patients from PTC and control groups, respectively (p= 0.15 and p= 0.34). Conclusion: PA prevalence was strikingly high among patients with PTC and HT, supporting the recommendation of PA screening in this patient group, regardless of HT severity. Presentation: Thursday, June 15, 2023

Abstract Context Paragangliomas (PGLs) are rare tumors in adrenal and extra-adrenal locations. Metastasis are found in approximately 5% to 35% of PGLs, and there are no reliable predictors of metastatic disease. Objective This work aimed to develop a prognostic score of metastatic potential in PGLs. Methods A retrospective analysis was conducted of clinical data from a cohort with PGLs and tumor histological assessment. Patients were divided into metastatic PGL (presence of metastasis) and nonmetastatic PGL (absence of metastasis ≥96 months of follow-up) groups. Univariate and multivariable analysis were performed to identify predictors of metastatic potential. A prognostic score was developed based on coefficients of multivariable analysis. Kaplan-Meier curves were generated to estimate disease-specific survival (DSS). Results Out of 263 patients, 35 patients had metastatic PGL and 110 patients had nonmetastatic PGL. In multivariable analysis, 4 features were independently related to metastatic disease and composed the Prognostic Score of Paragangliomas (PSPGL): presence of central or confluent necrosis (33 points), more than 3 mitosis/10 high-power field (HPF) (28 points), extension into adipose tissue (20 points), and extra-adrenal location (19 points). A PSPGL of 24 or greater showed similar sensitivity with higher specificity than the Pheochromocytoma of the Adrenal Gland Scaled Score (PASS) and Grading System for Adrenal Pheochromocytoma and Paraganglioma (GAPP). PSPGL less than or equal to 20 was associated with a risk of metastasis of approximately 10%, whereas a PSPGL of 40 or greater was associated with approximately 80%. The presence of metastasis and Ki-67 of 3% or greater were related to lower DSS. Conclusion The PSPGL, composed of 4 easy-to-assess parameters, demonstrated good performance in predicting metastatic potential and good ability in estimating metastasis risk.

Pheochromocytomas and paragangliomas (PPGLs) are neuroendocrine tumors derived from chromaffin cells. At least 30% of PPGL patients have hereditary predisposition. PPGLs in children are more often hereditary, multiple and extra-adrenal. To date, more than 14 tumor-susceptibility genes have been reported: Cluster 1 or hypoxic (VHL, SDHB, SDHD, SDHC, SDHA, SDHAF2, FH, ENGL1 and HIF2A) and cluster 2 (RET, NF1, TMEM127 and MAX). The aim of this study was to evaluate clinical and molecular aspects of a Brazilian cohort of pediatric patients with PPGLs. Out of 262 patients with PPGLs, 26 (9 %) were diagnosed before 19 yrs of age (16 males and 10 females), with a median age of 14.5 yrs (range, 4 to 18). Genetic investigation was performed in 19 patients: 14 by automated Sanger sequencing (VHL, SDHB, SDHD and RET genes) and 5 by a custom next-generation sequencing (NGS) panel including all genes previously associated with germline mutations in PPGLs. Median tumor size was 5.5 cm (1.7 to 16). Pheochromocytomas (PHEOs), paragangliomas (PGLs) or both were diagnosed in 46%, 31% and 23% of the patients, respectively. Bilateral PHEOs were diagnosed in 61% of the cases, most of them asynchronous (75%). Genetic diagnosis was confirmed in 14 out of 19 (74%) patients and all variants were found in heterozygous state: 8 VHL missense mutations from 6 kindreds (p.R167W in 2 kindreds, p.R167Q in one and p.G114S in 3); 3 SDHB mutations (p.C98Y, c.201-2A>G and p.L180L); 2 SDHD mutations (p.Y144_H145del and p.Q121*); and one RET mutation (p.C634R). All 8 VHL patients had bilateral PHEOs and 3 of them had also abdominal PGLs. All patients with SDHB mutations had abdominal PGLs. Two patients with SDHD mutation had head and neck paraganglioma (one of them had unilateral PHEO). Genetic investigation by NGS Panel was negative in all 5 cases: 2 malignant PPGLs (one PHEO and one PGL) and 3 PHEOs. Four out of 26 (15%) pediatric PPGLs were malignant: 2 with SDHB mutation and 2 with negative screening (one PHEO and one PGL). In conclusion, the majority of pediatric PPGLs (74%) were hereditary and almost exclusively caused by mutations in hypoxic genes. VHL (PHEOs) and SDHB (only PGLs) were the most frequent affected genes in this cohort of pediatric PPGLs. Support: CAPES grant to Petenuci J.