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Introduction Von Hippel-Lindau disease (VHL) is a rare cause of hereditary bilateral Pheochromocytomas (PHEO). Traditionally, treatment has been total adrenalectomy due to a lifetime risk of developing new tumors. Limited data exists on the surgical management of bilateral PHEO in children with VHL. We reviewed our experience with laparoscopic partial adrenalectomy for bilateral PHEO. Methods A retrospective review was performed of patients undergoing adrenalectomy for PHEO in children with VHL from 2004 to 2019. Results Eight children with VHL diagnosed with bilateral PHEO underwent 16 adrenalectomies (10 synchronous, 5 metachronous, 1 for recurrence). Median age at diagnosis was 13 [range 8–17] years with a median tumor size of 2.3 [range 0.5–7.7] cm. Of 16 adrenalectomies, all were performed laparoscopically, 14 were partial adrenalectomies; 2 patients required a contralateral total adrenalectomy due to size and diffuse multinodularity. There were no postoperative complications. No patients required corticosteroid replacement at the end of the study period. Two patients had new ipsilateral tumors identified after a median follow up of 5 [range 4–6] years with one undergoing repeat partial adrenalectomy. There were no mortalities in the study period. Conclusion Partial adrenalectomy for bilateral PHEO in patients with VHL is safe and does not compromise outcomes. When technically feasible, laparoscopic partial adrenalectomy should be considered as a primary surgical approach for children with VHL. Level of Evidence Level IV - Case series with no comparison group

BACKGROUND: Patients with von Hippel–Lindau (VHL) disease have a high incidence of renal cell carcinoma owing to VHL gene inactivation and constitutive activation of the transcription factor hypoxia-inducible factor 2α (HIF-2α). METHODS: In this phase 2, open-label, single-group trial, we investigated the efficacy and safety of the HIF-2α inhibitor belzutifan (MK-6482, previously called PT2977), administered orally at a dose of 120 mg daily, in patients with renal cell carcinoma associated with VHL disease. The primary end point was objective response (complete or partial response) as measured according to the Response Evaluation Criteria in Solid Tumors, version 1.1, by an independent central radiology review committee. We also assessed responses to belzutifan in patients with non–renal cell carcinoma neoplasms and the safety of belzutifan. RESULTS: After a median follow-up of 21.8 months (range, 20.2 to 30.1), the percentage of patients with renal cell carcinoma who had an objective response was 49% (95% confidence interval, 36 to 62). Responses were also observed in patients with pancreatic lesions (47 of 61 patients [77%]) and central nervous system hemangioblastomas (15 of 50 patients [30%]). Among the 16 eyes that could be evaluated in 12 patients with retinal hemangioblastomas at baseline, all (100%) were graded as showing improvement. The most common adverse events were anemia (in 90% of the patients) and fatigue (in 66%). Seven patients discontinued treatment: four patients voluntarily discontinued, one discontinued owing to a treatment-related adverse event (grade 1 dizziness), one discontinued because of disease progression as assessed by the investigator, and one patient died (of acute toxic effects of fentanyl). CONCLUSIONS: Belzutifan was associated with predominantly grade 1 and 2 adverse events and showed activity in patients with renal cell carcinomas and non–renal cell carcinoma neoplasms associated with VHL disease. (Funded by Merck Sharp and Dohme and others; MK-6482-004 ClinicalTrials.gov number, NCT03401788.)

ContextSomatic gene mutations have been identified in only about half of cortisol-producing adenomas (CPAs). Affected genes include PRKACA, GNAS, PRKAR1A, and CTNNB1.ObjectiveThis work aims to expand our understanding of the prevalence of somatic mutations in CPAs from patients with overt Cushing syndrome (OCS) and “subclinical” mild autonomous cortisol excess (MACE), with an immunohistochemistry (IHC)‒guided targeted amplicon sequencing approach using formalin-fixed paraffin-embedded (FFPE) tissue.MethodsWe analyzed FFPE adrenal tissue from 77 patients (n = 12 men, 65 women) with either OCS (n = 32) or MACE (n = 45). Using IHC for 17α-hydroxylase/17,20-lyase (CYP17A1) and 3β-hydroxysteroid dehydrogenase (HSD3B2), we identified 78 CPAs (32 OCS CPAs and 46 MACE CPAs). Genomic DNA was isolated from the FFPE CPAs and subjected to targeted amplicon sequencing for identification of somatic mutations.ResultsSomatic mutations were identified in 71.8% (56/78) of the CPAs. While PRKACA was the most frequently mutated gene in OCS CPAs (14/32, 43.8%), somatic genetic aberrations in CTNNB1 occurred in 56.5% (26/46) of the MACE CPAs. Most GNAS mutations were observed in MACE CPAs (5/7, 71.4%). No mutations were observed in PRKAR1A. In addition to the known mutations, we identified one previously unreported mutation in PRKACA. Two patients with MACE harbored 2 adjacent tumors within the same adrenal gland - one patient had 2 CPAs, and the other patient had a CPA and an aldosterone-producing adenoma (identified by IHC for aldosterone synthase).ConclusionA comprehensive FFPE IHC-guided gene-targeted sequencing approach identified somatic mutations in 71.8% of the CPAs. OCS CPAs demonstrated a distinct mutation profile compared to MACE CPAs.

This manuscript is the result of the North American Neuroendocrine Tumor Society consensus conference on the medical management and surveillance of metastatic and unresectable pheochromocytoma and paraganglioma held on October 2 and 3, 2019. The panelists consisted of endocrinologists, medical oncologists, surgeons, radiologists/nuclear medicine physicians, nephrologists, pathologists, and radiation oncologists. The panelists performed a literature review on a series of questions regarding the medical management of metastatic and unresectable pheochromocytoma and paraganglioma as well as questions regarding surveillance after resection. The panelists voted on controversial topics, and final recommendations were sent to all panel members for final approval.

Context Adrenal tumors in noncancer patients are common. Objective Evaluate performance of 18F-fluorodeoxyglucose positron emission tomography computed tomography (18F-FDG-PET/CT) in distinguishing between benign and malignant adrenal tumors. Design Retrospective chart review 2010-2019. Setting Academic institution. Patients One hundred and seventeen noncancer patients, defined as having no history of cancer or with cancer in remission for ≥5 years, completed 18F-FDG-PET/CT to evaluate adrenal masses, with pathologic diagnoses or imaging follow-up (≥12 months). Intervention 18F-FDG-PET/CT of 117 indeterminate adrenal masses. Main Outcome Measures Receiver operator characteristic curve of the ratios of adrenal lesion standardized uptake value (SUV)max to liver SUVmean and of adrenal lesion SUVmax to aortic arch blood pool SUVmean were constructed. Results Seventy benign and 47 malignant masses (35 adrenocortical carcinomas [ACCs], 12 adrenal metastases) were identified. Malignant masses had higher median liver SUV and blood pool SUV ratios than benign masses (6.2 and 7.4 vs 1.4 and 2.0, P < .001). Median liver and blood pool SUV ratios of ACC (6.1 and 7.3, respectively) and metastases (6.7 and 7.7, respectively) were higher than those of than adenomas (1.4 and 2.2, P < .05 for all comparisons). Optimal liver SUV ratio to discern between benign and malignant masses was 2.5, yielding 85% sensitivity, 90% specificity, and 7 false negative results (including 3 ACCs). Optimal blood pool SUV ratio was 3.4, yielding 83% sensitivity, 90% specificity, and 8 false negative results (including 4 ACCs). Conclusion When used in conjunction with other clinical assessments, 18F-FDG-PET/CT can be a valuable tool in evaluating adrenal masses in noncancer patients.

Contains fulltext : 245825.pdf (Publisher’s version ) (Closed access) BACKGROUND: Phaeochromocytoma or paraganglioma (collectively known as PPGL) in pregnant women can lead to severe complications and death due to associated catecholamine excess. We aimed to identify factors associated with maternal and fetal outcomes in women with PPGL during pregnancy. METHODS: We did a multicentre, retrospective study of patients with PPGL and pregnancy between Jan 1, 1980, and Dec 31, 2019, in the International Pheochromocytoma and Pregnancy Registry and a systematic review of studies published between Jan 1, 2005, and Dec 27, 2019 reporting on at least five cases. The inclusion criteria were pregnancy after 1980 and PPGL before or during pregnancy or within 12 months post partum. Eligible patients from the retrospective study and systematic review were included in the analysis. Outcomes of interest were maternal or fetal death and maternal severe cardiovascular complications of catecholamine excess. Potential variables associated with these outcomes were evaluated by logistic regression. FINDINGS: The systematic review identified seven studies (reporting on 63 pregnancies in 55 patients) that met the eligibility criteria and were of adequate quality. A further 197 pregnancies in 186 patients were identified in the International Pheochromocytoma and Pregnancy Registry. After excluding 11 pregnancies due to potential overlap, the final cohort included 249 pregnancies in 232 patients with PPGL. The diagnosis of PPGL was made before pregnancy in 37 (15%) pregnancies, during pregnancy in 134 (54%), and after delivery in 78 (31%). Of 144 patients evaluated for genetic predisposition for phaeochromocytoma, 95 (66%) were positive. Unrecognised PPGL during pregnancy (odds ratio 27·0; 95% CI 3·5-3473·1), abdominal or pelvic tumour location (11·3; 1·5-1440·5), and catecholamine excess at least ten-times the upper limit of the normal range (4·7; 1·8-13·8) were associated with adverse outcomes. For patients diagnosed during pregnancy, α-adrenergic blockade therapy was associated with fewer adverse outcomes (3·6; 1·1-13·2 for no α-adrenergic blockade vs α-adrenergic blockade), whereas surgery during pregnancy was not associated with better outcomes (0·9; 0·3-3·9 for no surgery vs surgery). INTERPRETATION: Unrecognised and untreated PPGL was associated with a substantially higher risk of either maternal or fetal complications. Appropriate case detection and counselling for premenopausal women at risk for PPGL could prevent adverse pregnancy-related outcomes. FUNDING: US National Institutes of Health.

Purpose: Minimal data exist regarding the efficacy of screening protocols for individuals with SDHx germline pathogenic variants with Hereditary Paraganglioma-Pheochromocytoma Syndrome. This study aimed to evaluate the SDHx-related tumor detection rate in individuals undergoing clinical screening protocols. Methods: A multi-center retrospective longitudinal observational study was conducted. Individuals with germline SDHx pathogenic variants underwent clinical whole-body imaging and biochemical testing. Results: 263 individuals with SDHx germline pathogenic variants completed 491 imaging screens. Individuals with SDHB germline pathogenic variants were most common (n=188/263, 72%), followed by SDHD (n=35/263, 13%) and SDHC (n=28/263, 11%). SDHx-related tumors were found in 17% (n=45/263) of the cohort. Most SDHx-related tumors were identified on baseline imaging screen (n=39/46, 85%). Individuals with SDHD pathogenic variants had the highest tumor detection rate (n=14/35, 40%). Of imaging screens identifying SDHx-related paraganglioma/pheochromocytoma, 29% (n=12/41) had negative biochemical testing. Secondary actionable findings were identified in 15% (n=75/491) of imaging screens. Conclusion: Current SDHx screening protocols are effective at identifying SDHx-related tumors. Tumor detection rates vary by SDHx gene and screening has the potential to uncover actionable secondary findings. Imaging is an essential part of the screening process as biochemical testing alone does not detect all disease.

Purpose This work aimed to evaluate genotype-phenotype associations in individuals carrying germline variants of transmembrane protein 127 gene (TMEM127), a poorly known gene that confers susceptibility to pheochromocytoma (PHEO) and paraganglioma (PGL). Design Data were collected from a registry of probands with TMEM127 variants, published reports, and public databases. Main Outcome Analysis Clinical, genetic, and functional associations were determined. Results The cohort comprised 110 index patients (111 variants) with a mean age of 45 years (range, 21-84 years). Females were predominant (76 vs 34, P < .001). Most patients had PHEO (n = 94; 85.5%), although PGL (n = 10; 9%) and renal cell carcinoma (RCC, n = 6; 5.4%) were also detected, either alone or in combination with PHEO. One-third of the cases had multiple tumors, and known family history was reported in 15.4%. Metastatic PHEO/PGL was rare (2.8%). Epinephrine alone, or combined with norepinephrine, accounted for 82% of the catecholamine profiles of PHEO/PGLs. Most variants (n = 63) occurred only once and 13 were recurrent (2-12 times). Although nontruncating variants were less frequent than truncating changes overall, they were predominant in non-PHEO clinical presentations (36% PHEO-only vs 69% other, P < .001) and clustered disproportionately within transmembrane regions (P < .01), underscoring the relevance of these domains for TMEM127 function. Integration of clinical and previous experimental data supported classification of variants into 4 groups based on mutation type, localization, and predicted disruption. Conclusions Patients with TMEM127 variants often resemble sporadic nonmetastatic PHEOs. PGL and RCC may also co-occur, although their causal link requires further evaluation. We propose a new classification to predict variant pathogenicity and assist with carrier surveillance.

Driver somatic mutations for aldosterone excess have been found in ≈90% of aldosterone-producing adenomas (APAs) using an aldosterone synthase (CYP11B2)-guided sequencing approach. In the present study, we identified a novel somatic CACNA1H mutation (c.T4289C, p.I1430T) in an APA without any currently known aldosterone-driver mutations using CYP11B2 immunohistochemistry-guided whole exome sequencing. The CACNA1H gene encodes a voltage-dependent T-type calcium channel alpha-1H subunit. Germline variants in this gene are known as a cause of familial hyperaldosteronism IV. Targeted next-generation sequencing detected identical CACNA1H variants in 2 additional APAs in a cohort of the University of Michigan, resulting in a prevalence of 4% (3/75) in APAs. We tested the functional effect of the variant on adrenal cell aldosterone production and CYP11B2 mRNA expression using the human adrenocortical HAC15 cell line with a doxycycline-inducible CACNA1H I1430T mutation. Doxycycline treatment increased CYP11B2 mRNA levels as well as aldosterone production, supporting a pathological role of the CACNA1H p.I1430T mutation on the development of primary aldosteronism. In conclusion, somatic CACNA1H mutation is a genetic cause of APAs. Although the prevalence of this mutation is low, this study will provide better understanding of molecular mechanism of inappropriate aldosterone production in APAs.

Context Adrenocortical carcinoma (ACC) is a rare endocrine malignancy that affects patients across the age spectrum. Although the overall survival in patients with ACC is poor, there is significant heterogeneity in terms of outcomes, presentation, and underlying genetic drivers. Evidence Acquisition This review is based on the evidence collected from primary research studies, expert reviews, and published guidelines. The studies were identified through PubMed search with key words “adrenocortical carcinoma,” “prognosis,” “pathology,” and “genetics.” The PubMed search was complemented by authors’ expertise, research, and clinical experience in the field of ACC. Evidence Synthesis Identification of biomarkers has been critical to gain better insight into tumor behavior and to guide therapeutic approach to patients. Tumor stage, resection status, and Ki67 are pathological tumor characteristics that have been identified as prognosticators in patients with ACC. Cortisol excess also correlates with worse prognosis. Clinical and histopathological characteristics help stratify patient outcomes, yet still up to 25% of patients have a different outcome than predicted. To bridge this gap, comprehensive genomic profiling studies have characterized additional profiles that correlate with clinical outcomes. In addition, studies of clinically applicable molecular markers are under way to further stratify outcomes in patients with ACC tumors. Conclusions Clinical predictors in combination with pathological markers play a critical role in the approach to patients with ACC. Recent advances in genetic prognosticators will help extend the stratification of these tumors and contribute to a personalized therapeutic approach to patients with ACC.

At least 30% of all pheochromocytomas (PCCs)/paragangliomas (PGLs) arise in patients with a germline predisposition syndrome. Variants in succinate dehydrogenase subunits A, B, C, and D (SDHA, SDHB, SDHC, and SDHD) are the most common pathogenic germline alterations. Few pathogenic variants have been reported in succinate dehydrogenase assembly factor 2 (SDHAF2). Here, we describe a 30-year-old female patient who presented with a left-sided neck mass, which was later characterized as a carotid body PGL. Genetic testing revealed a likely pathogenic SDHAF2 variant (c.347G>A;p.W116X). Two sisters carried the same pathologic variant, and screening protocols were recommended. Whole-body MRI revealed thyroid nodules; this testing was followed by fine-needle aspiration, which confirmed papillary thyroid carcinoma in one sister and a follicular adenoma in the other. The two sisters then underwent hemithyroidectomy and total thyroidectomy, respectively. Because evidence for pathogenic variants in SDHAF2 causing predisposition to PCC/PGL is limited, we discuss the challenges in mutational variant interpretation and decision making regarding screening for associated tumors.

Purpose: Adrenocortical carcinoma (ACC) is a rare, aggressive malignancy with few therapies; however, patients with locoregional disease have variable outcomes. The Cancer Genome Atlas project on ACC (ACC-TCGA) identified that cancers of patients with homogeneously rapidly recurrent or fatal disease bear a unique CpG island hypermethylation phenotype, “CIMP-high.” We sought to identify a biomarker that faithfully captures this subgroup. Experimental Design: We analyzed ACC-TCGA data to characterize differentially regulated biological processes, and identify a biomarker that is methylated and silenced exclusively in CIMP-high ACC. In an independent cohort of 114 adrenocortical tumors (80 treatment-naive primary ACC, 22 adrenocortical adenomas, and 12 non-naive/nonprimary ACC), we evaluated biomarker methylation by a restriction digest/qPCR-based approach, validated by targeted bisulfite sequencing. We evaluated expression of this biomarker and additional prognostic markers by qPCR. Results: We show that CIMP-high ACC is characterized by upregulation of cell cycle and DNA damage response programs, and identify that hypermethylation and silencing of G0S2 distinguishes this subgroup. We confirmed G0S2 hypermethylation and silencing is exclusive to 40% of ACC, and independently predicts shorter disease-free and overall survival (median 14 and 17 months, respectively). Finally, G0S2 methylation combined with validated molecular markers (BUB1B-PINK1) stratifies ACC into three groups, with uniformly favorable, intermediate, and uniformly dismal outcomes. Conclusions: G0S2 hypermethylation is a hallmark of rapidly recurrent or fatal ACC, amenable to targeted assessment using routine molecular diagnostics. Assessing G0S2 methylation is straightforward, feasible for clinical decision-making, and will enable the direction of efficacious adjuvant therapies for patients with aggressive ACC.

Adrenal steroidogenesis is a robust process, involving a series of enzymatic reactions that facilitate conversion of cholesterol into biologically active steroid hormones under the stimulation of angiotensin II, adrenocorticotropic hormone and other regulators. The biosynthesis of mineralocorticoids, glucocorticoids, and adrenal-derived androgens occur in separate adrenocortical zones as a result of the segregated expression of steroidogenic enzymes and cofactors. This mini review provides the principles of adrenal steroidogenesis, including the classic and under-appreciated 11-oxygenated androgen pathways. Several adrenal diseases result from dysregulated adrenal steroid synthesis. Herein, we review growing evidence that adrenal diseases exhibit characteristic modifications from normal adrenal steroid pathways that provide opportunities for the discovery of biomarker steroids that would improve diagnosis and monitoring of adrenal disorders.

Somatic mutations have been identified in aldosterone-producing adenomas (APAs) in genes that include KCNJ5 , ATP1A1 , ATP2B3 , and CACNA1D . Based on independent studies, there appears to be racial differences in the prevalence of somatic KCNJ5 mutations, particularly between East Asians and Europeans. Despite the high cardiovascular disease mortality of blacks, there have been no studies focusing on somatic mutations in APAs in this population. In the present study, we investigated genetic characteristics of APAs in blacks using a CYP11B2 (aldosterone synthase) immunohistochemistry-guided next-generation sequencing approach. The adrenal glands with adrenocortical adenomas from 79 black patients with primary aldosteronism were studied. Seventy-three tumors from 69 adrenal glands were confirmed to be APAs by CYP11B2 immunohistochemistry. Sixty-five of 73 APAs (89%) had somatic mutations in aldosterone-driver genes. Somatic CACNA1D mutations were the most prevalent genetic alteration (42%), followed by KCNJ5 (34%), ATP1A1 (8%), and ATP2B3 mutations (4%). CACNA1D mutations were more often observed in APAs from males than those from females (55% versus 29%, P =0.033), whereas KCNJ5 mutations were more prevalent in APAs from females compared with those from males (57% versus 13%, P CACNA1D . Determination of racial differences in the prevalence of aldosterone-driver gene mutations may facilitate the development of personalized medicines for patients with primary aldosteronism.

Somatic variants in genes that regulate intracellular ion homeostasis have been identified in aldosterone-producing adenomas (APAs). Although the mechanisms leading to increased aldosterone production in APA cells have been well studied, the molecular events that cause cell proliferation and tumor formation are poorly understood. In the present study, we have performed whole-exome sequencing (WES) to characterize the landscape of somatic alterations in a homogeneous series of APA with pathogenic KCNJ5 variants. In the WES analysis on 11 APAs, 84 exonic somatic events were called by 3 different somatic callers. Besides the KCNJ5 gene, only two genes (MED13 and ZNF669) harbored somatic variants in more than one APA. Unlike adrenocortical carcinomas, no chromosomal instability was observed by the somatic copy-number alteration and loss of heterozygosity analyses. The estimated tumor purity ranged from 0.35 to 0.67, suggesting a significant proportion of normal cell infiltration. Based on the results of PureCN analysis, the KCNJ5 variants appear to be clonal. In conclusion, in addition to KCNJ5 somatic pathogenic variants, no significant somatic event that would obviously explain proliferation or tumor growth was observed in our homogeneous cohort of KCNJ5-mutated APA. The molecular mechanisms causing APA growth and tumorigenesis remain to be elucidated.

Background Patterns and prognostic implications of recurrent adrenocortical carcinoma are poorly understood. In this study, we aim to describe temporal and spatial patterns of adrenocortical carcinoma recurrence. Methods This is a retrospective review of 576 patients with adrenocortical carcinoma evaluated at a single institution. Clinicopathologic and follow-up data were collected longitudinally. Results A total of 354 patients underwent resection of stage I-III adrenocortical carcinoma. We found that 249 (70%) patients developed disease recurrence. The median recurrence-free interval after primary resection was 11 months. The most common sites of initial recurrence were lung and tumor bed. The shortest time to recurrence was associated with lung or multiple site metastases. We found that 142 of 249 patients developed one or more additional sites of recurrence (median 5 months), most commonly involving the lungs. A total of 20 patients developed a third site of recurrence. We found that 100 patients underwent one or more reoperations or metastasectomies and 79 recurred again after reoperation. Same organ or site recurrence was common after reoperation (67%). Although lung metastases occurred early, recurrences to the peritoneal cavity or to multiple sites were associated with worse survival. Metastasectomy beyond three total operations did not improve overall survival. Conclusion Survival varies according to site of recurrence and other clinicopathologic factors. Knowledge of patterns of recurrence may assist in anticipating disease course and lead to better informed selection of treatment.

Introduction We sought to characterize the prevalence and factors characteristic of head and neck paragangliomas (HNPGLs) that secrete catecholamines to inform best practices for diagnosis and management. Methods This was a retrospective cohort study from 2000 to 2020 at a single‐institution tertiary centre. One‐hundred fifty‐two patients (182 tumours) with HNPGLs with at least one measurement of urine or plasma catecholamines and/or catecholamine metabolite levels prior to treatment were included. We differentiated and characterized those patients with increased level(s) of any nature and those with ‘clinically significant’ versus ‘clinically insignificant’ catecholamine production. Results Thirty‐one (20.4%) patients had increased catecholamine and/or catecholamine metabolite levels. In most patients, these levels were ≤5‐fold above the upper limit of the reference range. Four of these 31 patients with increased levels were ultimately found to have an additional catecholamine secreting mediastinal paraganglioma or pheochromocytoma. Fourteen of 31 patients with HNPGL were deemed clinically significant secretors of catecholamines based on hyper‐adrenergic symptoms and/or profound levels of normetanephrines. This cohort was enriched for patients with paragangliomas of the carotid body or cervical sympathetic chain and those with SDHB genetic mutations. Ultimately, the prevalence of clinically significant catecholamine secreting Hangs was determined to be 9.2% and 7.7% based on a per‐patient and per‐tumour basis, respectively. Conclusions The rate of catecholamine excess in the current cohort of patients with HNPGLs was higher than previously reported. Neuroendocrine tumours of any anatomic subsite may secrete catecholamines, although not all increased laboratory level(s) are indicative of clinically significant catecholamine secretion causing symptoms or warranting adrenergic blockade., The prevalence of clinically significant catecholamine secreting head and neck paragangliomas was determined to be 7.7%–9.2% based on a per‐patient and per‐tumour basis, respectively. The rate of catecholamine excess in the current cohort of patients with HNPGLs was higher than previously reported. Neuroendocrine tumours of any anatomic subsite may secrete catecholamines, though not all increased laboratory level(s) are indicative of clinically significant catecholamine secretion causing symptoms or warranting adrenergic blockade.

Objective A large proportion of head and neck paragangliomas (HNPGLs) arise in patients with a genetic predisposition due to pathogenic variants in succinate dehydrogenase ( SDHx ) genes. Contemporary practice guidelines recommend consideration of referral for genetic testing for all patients with HNPGLs. We sought to assess adherence to these recommendations, factors associated with referral, and temporal trends in referral patterns by otolaryngologists over the past 2 decades. Study Design Retrospective cohort study. Setting Single tertiary care center. Methods All patients with newly diagnosed HNPGLs treated at a single academic center between 2000 and 2019 were included. Bivariable association of specific features of referral for genetic testing by treating surgeons were tested with χ 2 and Wilcoxon rank-sum tests. Logistic regression was used to assess temporal trends in referral patterns overall and for specific clinical subgroups over time. Results Of 221 patients included, only 77 (34.8%) were referred for genetic testing. Factors associated with referral included young age, family history of paraganglioma, more recent year of diagnosis (ie, closer to study end date), tumor subsite (all P < .0001), and treatment by an otolaryngologist (vs vascular surgeon or neurosurgeon, P = .009). Overall, referral rates increased over time ( P = .0002), but even in the most recent 5 years, only 51% of newly diagnosed patients were referred. Conclusion Our analysis suggests that referral rates for genetic testing in patients with HNPGLs are growing yet are still largely based on young age, family history, and tumor subsite.

Endogenous Cushing syndrome (CS) is an endocrine disorder marked by excess cortisol production rendering patients susceptible to visceral obesity, dyslipidemia, hypertension, osteoporosis and diabetes mellitus. Adrenal CS is characterized by autonomous production of cortisol from cortisol-producing adenomas (CPA) via adrenocorticotropic hormone-independent mechanisms. A limited number of studies have quantified the steroid profiles in sera from patients with CS. To understand the intratumoral steroid biosynthesis, we quantified 19 steroids by mass spectrometry in optimal cutting temperature compound (OCT)-embedded 24 CPA tissue from patients with overt CS (OCS, n = 10) and mild autonomous cortisol excess (MACE, n = 14). Where available, normal CPA-adjacent adrenal tissue (AdjN) was also collected and used for comparison (n = 8). Immunohistochemistry (IHC) for CYP17A1 and HSD3B2, two steroidogenic enzymes required for cortisol synthesis, was performed on OCT sections to confirm the presence of tumor tissue and guided subsequent steroid extraction from the tumor. LC-MS/MS was used to quantify steroids extracted from CPA and AdjN. Our data indicated that CPA demonstrated increased concentrations of cortisol, cortisone, 11-deoxycortisol, corticosterone, progesterone, 17OH-progesterone and 16OH-progesterone as compared to AdjN (p < 0.05). Compared to OCS, MACE patient CPA tissue displayed higher concentrations of corticosterone, 18OH-corticosterone, 21-deoxycortisol, progesterone, and 17OH-progesterone (p < 0.05). These findings also demonstrate that OCT-embedded tissue can be used to define intra-tissue steroid profiles, which will have application for steroid-producing and steroid-responsive tumors.

The COVID-19 pandemic has prompted the rapid transition of in-person outpatient care to telemedicine, and clinical training to remote learning. The endocrinology fellows at the University of Michigan maintain their own continuity-of-care clinics and rotate in the Ann Arbor Veterans Affairs (VA) Healthcare System. For these clinics, we sought to preserve patient staffing with expert attending physicians and continue the clinical training experience in a remote setting.We have adapted the online conferencing platform, Zoom, to integrate learners into a virtual teaching clinic environment. By using the Zoom “breakout room” feature, fellows are able to match staffing attending physicians to different patient cases, according to attending physicians’ areas of specialty. Similar to the traditional teaching clinic environment, our remote staffing strategy has ensured that fellows continue to provide excellent patient care and fulfill educational aims across our University and VA facilities.Outpatient clinics in other University of Michigan departments and other academic centers have inquired about or have begun utilizing our method. Even beyond COVID-19, our paradigm potentially provides a convenient virtual staffing platform to serve patient populations with geographic or transportation challenges. Following implementation, stakeholders can regularly evaluate the approach to continually improve both patient care and medical education.

Objective: The aim of this Disease State Clinical Review is to provide a practical approach to patients with newly diagnosed adrenocortical carcinoma, as well as to follow-up and management of patients with persistent or recurrent disease. Methods: This is a case-based clinical review. The provided recommendations are based on evidence available from randomized prospective clinical studies, cohort studies, cross-sectional and case-based studies, and expert opinions. Results: Adrenocortical carcinoma is a rare malignancy, often with poor outcomes. For any patient with an adrenal mass suspicious for adrenocortical carcinoma, the approach should include prompt evaluation with detailed history and physical exam, imaging, and biochemical adrenal hormone assessment. In addition to adrenal-focused imaging, patients should be evaluated with chest-abdomen-pelvis cross-sectional imaging to define the initial therapy plan. Patients with potentially resectable disease limited to the adrenal gland should undergo en bloc open surgery by an expert surgeon. For patients presenting with advanced or recurrent disease, a multidisciplinary approach considering curative repeat surgery, local control with surgery, radiation therapy or radiofrequency ablation, or systemic therapy with mitotane and/or cytotoxic chemotherapy is recommended. Conclusion: As most health care providers will rarely encounter a patient with adrenocortical carcinoma, we recommend that patients with suspected adrenocortical carcinoma be evaluated by an expert multidisciplinary team which includes clinicians with expertise in adrenal tumors, including endocrinologists, oncologists, surgeons, radiation oncologists, pathologists, geneticists, and radiologists. We recommend that patients in remote locations be followed by the local health care provider in collaboration with a multidisciplinary team at an expert adrenal tumor program. Abbreviations: ACC = adrenocortical carcinoma; ACTH = adrenocorticotropic hormone; BRACC = borderline resectable adrenocortical carcinoma; CT = computed tomography; DHEAS = dehydroepiandrosterone sulfate; EDP = etoposide, doxorubicin, cisplatin; FDG = 18F-fluorodeoxyglucose; FNA = fine-needle aspiration; HU = Hounsfield units; IVC = inferior vena cava; LFS = Li-Fraumeni syndrome; MEN1 = multiple endocrine neoplasia type 1; MRI = magnetic resonance imaging; OAC = oncocytic adrenocortical carcinoma; PC = palliative care; PET = positron emission tomography

Somatic mutations driving aldosterone production have been identified in approximately 90% of aldosterone-producing adenomas (APAs) using an aldosterone synthase (CYP11B2) immunohistochemistry (IHC)-guided DNA sequencing approach. In the present study, using CYP11B2-guided whole-exome sequencing (WES) and targeted amplicon sequencing, we detected 2 somatic variants in CLCN2 in 2 APAs that were negative for currently known aldosterone-driver mutations. The CLCN2 gene encodes the voltage-gated chloride channel ClC-2. CLCN2 germline variants have previously been shown to cause familial hyperaldosteronism type II. Somatic mutations in CLCN2 were identified in 2 of 115 APAs, resulting in a prevalence of 1.74%. One of the CLCN2 somatic mutations (c.G71A,p.G24D) was identical to a previously described germline variant causing early-onset PA, but was present only as a somatic mutation. The second CLCN2 mutation, which affects the same region of the gene, has not been reported previously (c.64-2_74del). These findings prove that WES of CYP11B2-guided mutation-negative APAs can help determine rarer genetic causes of sporadic PA.

Lack of routine fresh or frozen tissue is a barrier to widespread transcriptomic analysis of adrenal cortical tumors and an impediment to translational research in endocrinology and endocrine oncology. Our group has previously pioneered the use of targeted amplicon-based next-generation sequencing for archival formalin-fixed paraffin-embedded (FFPE) adrenal tissue specimens to characterize the spectrum of somatic mutations in various forms of primary aldosteronism. Herein, we developed and validated a novel 194-amplicon targeted next-generation RNA sequencing (RNAseq) assay for transcriptomic analysis of adrenal tumors using clinical-grade FFPE specimens. Targeted RNAseq-derived expression values for 27 adrenal cortical tumors, including aldosterone-producing adenomas (APA; n=8), cortisol-producing adenomas (CPA; n=11), and adrenal cortical carcinomas (ACC; n=8), highlighted known differentially-expressed genes (DEGs; i. e., CYP11B2, IGF2, etc.) and tumor type-specific transcriptional modules (i. e., high cell cycle/proliferation transcript expression in ACC, etc.), and a subset of DEGs was validated orthogonally using quantitative reverse transcription PCR (qRT-PCR). Finally, unsupervised hierarchical clustering using a subset of high-confidence DEGs revealed three discrete clusters representing APA, CPA, and ACC tumors with corresponding unique gene expression signatures, suggesting potential clinical utility for a transcriptomic-based approach to tumor classification. Overall, these data support the use of targeted amplicon-based RNAseq for comprehensive transcriptomic profiling of archival FFPE adrenal tumor material and indicate that this approach may facilitate important translational research opportunities for the study of these tumors.

Primary hyperparathyroidism (pHPT) is associated with familial syndromes such as multiple endocrine neoplasia type 1 (MEN1), 2A (MEN2A), MEN-like syndromes (CDKN1B), and CDC73-related disorder (hyperparathyroidism – jaw tumor syndrome (HPJT)). Familial hypocalciuric hypercalcemia (FHH) caused by CASR variants is an important differential diagnosis for pHPT. In order to evaluate the contribution of hereditary causes to pHPT in patients encountered in a specialized clinic, we conducted a retrospective study on patients with pHPT that underwent germline genetic testing. We evaluated 46 patients referred to a Cancer Genetics Clinic. Reasons for referral were young age (age

18F-fluorodeoxyglucose positron emission tomography computed tomography (FDG PET/CT) is frequently used for evaluation of indeterminate adrenal nodules. The objective of this study was to evaluate the diagnostic performance of FDG PET/CT in distinguishing benign adrenocortical adenomas (ACA) from adrenocortical carcinoma (ACC). We identified 139 patients in our institution who completed both FDG PET/CT and adrenal biopsy or adrenalectomy from 2001 to 2019 using the institutional Electronic Medical Record Search Engine (EMERSE). Patients with adrenal pathology revealing non-adrenocortical tumors were excluded. The imaging characteristics of ACAs were compared with ACCs. Sixteen patients with ACAs and 50 patients with ACC were identified; of the latter, 30 were excluded, because FDG PET/CT imaging was completed after adrenalectomy. The average age of the ACC vs. ACA group was 48±14 and 52±17 years, respectively, with a gender composition of 35% and 25% men, respectively. All 20 patients with ACC had FDG avid lesions, defined by imaging report and increased ratio of adrenal tumor maximum standardized uptake value (SUVmax) to hepatic parenchymal average SUV (SUVmean) of greater than 2.5. Among those with ACAs, 11 had positive, 1 had mildly positive, 3 had indeterminate, and 1 had negative FDG PET/CT findings according to the report. Ratio of adrenal SUVmax to hepatic SUVmean was greater in ACC compared to ACA (13.5 versus 1.6, respectively, p = 0.13), but did not reach statistical significance, likely due to the small sample size and unavailability of SUV ratios for all lesions. Our study identified a large number of ACAs that were deemed FDG avid but without adrenal SUVmax to hepatic SUVmean ratio of greater than 2.5. Although the sensitivity, and therefore the negative predictive value, of FDG PET/CT scan is excellent for the diagnosis of ACC, our data does not allow for the calculation of specificity. This is due to the selection bias inherent in our patient population referred for evaluation of unusual adrenal masses, and the fact that ACAs with negative FDG PET/CT often do not undergo surgery or biopsy, and therefore pathology results are not available. Despite this limitation, our findings show that calculation of the ratio of adrenal SUVmax to hepatic SUVmean, rather than lesion SUVmax alone, helps for characterization of adrenal lesions as malignant vs. benign, and that a negative FDG PET/CT is valuable in excluding ACC.

Background: Aldosterone-producing adenoma (APA) is a major subtype of primary aldosteronism (PA) which is the most common cause of endocrine-related hypertension. The Endocrine Society clinical practice guideline suggests that young patients (< 35 years old) with a CT-detected adrenocortical adenoma and typical phenotype of PA may not need adrenal venous sampling prior to adrenalectomy. In recent years, aldosterone-driver somatic mutations have been identified in APA, and prevalence studies suggest potential effects of patient age and sex. However, the rare nature of early-onset PA has prevented a detailed study of the histologic characteristics and aldosterone-driver somatic mutations in adrenal tumors from these patients. Objective: To determine histologic and somatic mutation profile in early-onset APA. Methods: Fifty-five formalin-fixed paraffin-embedded (FFPE) adrenals from patients at the age of 35 years old or younger who underwent adrenalectomy at the participating centers were studied (45 women, 9 men, and 1 unknown sex). CYP11B2 immunohistochemistry (IHC)-guided tumor capturing was used to selectively obtain DNA from APA. Mutation status was determined either by Sanger sequencing or targeted next-generation sequencing. Results: CYP11B2 IHC identified APAs in all adrenal specimens. Solitary APAs were found in 53 adrenals. One adrenal had multiple APAs and one had a dominant CYP11B2-negative tumor and a smaller APA. In total, DNA from 57 APAs were sequenced. Two APAs were excluded from the analysis due to low sample quality. In 52 of the 55 APAs, somatic mutations were identified in one of the aldosterone-driver genes or CTNNB1 gene, encoding β-catenin. The most common genetic alteration was seen in KCNJ5 (37/55, 67%), followed by CACNA1D (7/55, 13%), ATP1A1 (3/55, 5%), CTNNB1 (3/55, 5%), and ATP2B3 (2/55, 4%). No sex difference in the prevalence of KCNJ5 mutation was observed in this age group. Conclusion: The majority of adrenals from early-onset PA patients had a solitary APA. Regardless of sex, the most common genetic cause of early-onset APA was somatic mutations in KCNJ5.

Most ovarian steroid cell tumors arise sporadically. However, they can also be observed as a rare manifestation of von Hippel Lindau disease. Here, we present a clinical, pathological and molecular characterization of a steroid cell tumor in a VHL patient. A 14 year old girl with molecularly confirmed diagnosis of VHL developed hirsutism and amenorrhea. Initial clinical hormonal evaluation was notable for elevated 17-OHP of 406ng/dl, androstenedione 275ng/dl, and testosterone 102ng/dl. In order to exclude congenital adrenal hyperplasia as a common cause of hirsutism in adolescents, ACTH stimulation was performed, but no increase in 17-OHP was observed. Anti Muellerian hormone, inhibin (INH) A and INH B were normal. Imaging revealed a bilobed 6cm left adnexal mass. The mass was resected en bloc via a left oophorectomy. Pathological evaluation showed multinodular steroid cell tumor with clear cytoplasm and delicate vascular meshwork. Immunoprofiling demonstrated positivity for inhibin and calretinin; while renal cell carcinoma markers were negative. All laboratory values normalized post-surgery. In addition to clinical measurements pre- and post-surgery, steroid profiles were evaluated by LC-MS/MS. Quantitative RT-PCR analysis showed robust tumor expression of enzymes facilitating the production of androgens, but not estrogens. Further preliminary analysis by exome sequencing confirmed the known germline pathogenic variant in VHL, but no additional obvious somatic driver mutations were identified. Interestingly, the NGS analysis of different specimens from the same tumor revealed multiple different single base pair variants in the VHL gene as a second hit. In summary, hirsutism in VHL patients should raise the suspicion for unusual ovarian tumors. In contrary to the usual theory of a monoclonal expansion after loss of the wt VHL allele, this tumor appeared to be oligoclonal as evidenced by different somatic VHL mutations. This could be either explained by initial parallel occurrence of several clones or that the VHL second hit is not an initial event, but the mutation instead supports tumor expansion following initial steps of tumorigenesis.

Adrenocortical carcinoma (ACC) is a rare, aggressive cancer. Up to 75% of patients develop incurable metastatic disease, highlighting an urgent need for novel medical therapies. We recently identified a rapidly progressive ACC subtype characterized by CpG island hypermethylation (CIMP-high), sustained Wnt/β-catenin signaling, steroidogenic differentiation, and cell cycle activation. CIMP-high status alone accounts for 40% of ACC, but predicts 70% of recurrences and >50% of deaths. Intriguingly, hypermethylated CpG islands in CIMP-high ACC are unmethylated in fetal and adult adrenal cortex, suggesting DNA methylation is supported by cancer-specific mechanisms. We therefore sought to investigate how aberrant epigenetic programming contributes to ACC biology. In embryonic stem cells, the Polycomb repressive complex 2 (PRC2) represses differentiation programs through EZH2-mediated histone H3 lysine 27 trimethylation (H3K27me3) deposition in promoter CpG islands free of DNA methylation. Gain or loss of EZH2/PRC2 function prevails in a variety of human cancers, enabling proliferation in a tissue-specific manner. Here, we identify that CIMP-high ACC exhibit high expression of EZH2/H3K27me3, but paradoxically bear DNA hypermethylation in annotated PRC2 target regions. To determine if DNA methylation of PRC2 targets disrupts or is controlled by EZH2, we characterized EZH2’s role in CIMP-high ACC cell line NCI-H295R at baseline and in response to EZH2 inhibition (EZH2i). EZH2-directed IP-MS revealed EZH2 interacts with PRC2 members and DNA methylation-sensitive accessory proteins, but no DNA methyltransferase machinery. ChIP-seq revealed EZH2 and H3K27me3 colocalize in repressive domains genome-wide, but DNA methylation and H3K27me3 are mutually exclusive. EZH2i induced H3K27 demethylation and loss of viability, but with no effect on CIMP-high DNA methylation. These data suggest PRC2 target DNA methylation in CIMP-high ACC is maintained independently of EZH2, enabling EZH2/PRC2 to coordinate alternative programs required for cell survival. We then measured the consequences of EZH2i on the NCI-H295R transcriptome (RNA-seq), EZH2/H3K27me3 deposition genome-wide (ChIP-seq), and chromatin accessibility landscape (ATAC-seq). EZH2i led to global downregulation of cell cycle, Wnt/β-catenin transcriptional programming, and steroidogenic differentiation, partially explained by EZH2i-induced offloading of EZH2 from H3K27me3 domains to accessible promoters genome-wide. Taken together, our studies illustrate how aberrant CpG island hypermethylation in CIMP-high ACC participates in a targetable repressive epigenetic cascade that reinforces oncogenic adrenocortical transcriptional programs. Ultimately, we hope to illuminate novel strategies for tissue-specific disruption of the aberrant epigenetic wiring that defines CIMP-high ACC.

Secondary hypertension is a common condition with a broad differential diagnosis. Identification of the true cause of hypertension can be critical for guiding appropriate management. Here, we review hereditary conditions underlying the most common cause of secondary hypertension, primary aldosteronism, as well as other disorders impacting various levels of mineralocorticoid action. Recently, several pathogenic variants of ion channels have been described as etiologies of familial aldosteronism. Defects in steroid hormone synthesis cause hypertension in 11β-hydroxylase deficiency and 17α-hydroxylase deficiency, two types of congenital adrenal hyperplasia. Inappropriate activation of mineralocorticoid receptors underlies the syndrome of apparent mineralocorticoid excess and constitutive activation of the mineralocorticoid receptor. Finally, Liddle syndrome and pseudohypoaldosteronism type 2 are disorders impacting the function of renal sodium channels, the endpoint of mineralocorticoid action. We discuss the pathophysiology, clinical presentation, diagnosis and management of these low renin hypertension states that ultimately result in apparent excess mineralocorticoid activity.

Primary aldosteronism (PA) is the most common cause of secondary hypertension with a high prevalence amongst patients with resistant hypertension. Despite the recent discovery of somatic variants in aldosterone-producing adenoma (APA)-associated PA, causes for PA due to bilateral aldosterone production (bilateral hyperaldosteronism; BHA) remain unknown. Herein, we identified rare gene variants in ATP2B4, in a cohort of patients with BHA. ATP2B4 belongs to the same family of Ca-ATPases as ATP2B3, which is involved in the pathogenesis of APA. Endogenous ATP2B4 expression was characterized in adrenal tissue and the gene variants were functionally analyzed for effects on aldosterone synthase (CYP11B2) expression, steroid production in basal and agonist-stimulated conditions, and for changes in biophysical properties of channel properties. Knock down of ATP2B4 in HAC15 exhibited reduced Angiotensin II-stimulation in one of four shRNA clones. Stable HAC15 cell lines with doxycycline (dox) – inducible wildtype and variant forms of ATP2B4, were generated, and dox-induced upregulation of ATP2B4 mRNA and protein was confirmed. However, ATP2B4 variants did not alter basal or agonist-stimulated CYP11B2 expression. Whole cell recordings in HAC15 cells indicated robust endogenous ATP2B4 conductance in native cells, but reduced conductance with over-expressed WT and variant ATP2B4. The previously defined PA-causing ATP2B3 variant served as a positive control and exhibited elevated CYP11B2 mRNA. In conclusion, while this study did not confirm a pathogenic role for ATP2B4 variants in BHA, we describe the sequencing analysis for familial and sporadic BHA and outline a template for the thorough in vitro characterization of gene variants.

The XAF1-E134* variant increases the cancer risk for carriers of the TP53-R337H allele., Cancer risk is highly variable in carriers of the common TP53-R337H founder allele, possibly due to the influence of modifier genes. Whole-genome sequencing identified a variant in the tumor suppressor XAF1 (E134*/Glu134Ter/rs146752602) in a subset of R337H carriers. Haplotype-defining variants were verified in 203 patients with cancer, 582 relatives, and 42,438 newborns. The compound mutant haplotype was enriched in patients with cancer, conferring risk for sarcoma (P = 0.003) and subsequent malignancies (P = 0.006). Functional analyses demonstrated that wild-type XAF1 enhances transactivation of wild-type and hypomorphic TP53 variants, whereas XAF1-E134* is markedly attenuated in this activity. We propose that cosegregation of XAF1-E134* and TP53-R337H mutations leads to a more aggressive cancer phenotype than TP53-R337H alone, with implications for genetic counseling and clinical management of hypomorphic TP53 mutant carriers.

Hereditary leiomyomatosis and renal cell carcinoma (HLRCC) is an autosomal dominant condition caused by highly penetrant pathogenic variants in the fumarate hydratase (FH) gene.1 Fumarate hydratase is an enzyme responsible for the conversion of fumarate to malate in the Krebs cycle and is thought to be a tumor suppressor gene.1 Classically, HLRCC has a predisposition to the development of cutaneous leiomyomas, uterine leiomyomas (fibroids), and renal cell carcinoma (RCC).1 However, clinical presentation and disease phenotypes vary widely. We present a case in which the incidental observation of a few pink papules led to the diagnosis of HLRCC.

Context: Quality of life (QoL) is impaired in patients with chronic adrenal insufficiency (AI) despite standard glucocorticoid (GC) replacement therapy. Current evidence on the determinants of QoL is scarce and limited in very few European countries, and how it relates to AI subtypes remains underexplored. Objective: We conducted the first survey study in patients with AI in the USA to determine the correlations between clinical parameters, adverse outcomes, patient education, socioeconomic factors, and QoL in different subtypes of AI. Design, Setting and Participants: Cross-sectional survey study of 529 patients with AI between 2015 and 2020, at three tertiary centers in the USA. Intervention: Patient-centered questionnaires. Main Outcome Measures: QoL scores using Short-Form 36. Results: Of 529 participants, 223 (42.2%) had primary AI (PAI), 190 (35.9%) had secondary AI (SAI), and 116 (21.9%) had glucocorticoid induced AI (GIAI). Median age at the time of survey was 58 years (IQR: 43–68), 342 (64.8%) were women and 483 (91.3%) were Caucasians. Median duration of AI was 6 years (IQR: 3–14.5), longest in patients with PAI (11 vs 4 years in SAI and GIAI, p=0.0001). Overall, Physical Composite Summary (PCS) score was lower than the Mental Composite Summary (MCS) (38.1±12.9 vs 46.5±11.8). Across the eight dimensions, each individual decade- and sex-adjusted Z-score (using the normative data of USA population) in patients with PAI was significantly higher than that of the other two AI subtypes (all p25 mg hydrocortisone equivalent daily (OR: 2.3, CI 95%: 1.2–4.6), had more comorbidities related to GC excess (OR: 2.3, CI 95%: 1.3–4.0), reported higher financial burden due to AI (OR: 2.1, CI 95%: 1.3–3.6), and reported difficulties with AI management (OR: 2.5, CI 95%: 1.2–5.2). Women (OR: 2.1, CI 95%: 1.08–4.0), shorter duration of AI (OR: 2.4, CI 95%: 1.4–4.3), higher financial burden due to AI (OR: 2.3, CI 95%: 1.3–4.0), reporting difficulties with AI management (OR: 2.6, CI 95%: 1.4–4.9), and lack of family support during adrenal crisis (OR: 9.1, CI 95%: 2.3–33.3), were predictors of a worse MCS score ( Conclusions: Patients with AI have substantially impaired QoL despite GC replacement therapy. Certain determinants of QoL are modifiable and achievable, such as avoiding GC over-replacement, offering detailed hands-on education in self-management, more comprehensive insurance coverage, and more robust domestic support. Our study calls for a multidimensional effort from patients, clinicians, and society to improve QoL in this vulnerable patient population.

ObjectiveGastroenteropancreatic neuroendocrine tumours (GEP-NETs) encompass a diverse group of neoplasms that vary in their secretory products and in their location within the gastrointestinal tract. Their prevalence in the USA is increasing among all adult age groups.AimTo identify the possible derivation of GEP-NETs using genome-wide analyses to distinguish small intestinal neuroendocrine tumours, specifically duodenal gastrinomas (DGASTs), from pancreatic neuroendocrine tumours.DesignWhole exome sequencing and RNA-sequencing were performed on surgically resected GEP-NETs (discovery cohort). RNA transcript profiles available in the Gene Expression Omnibus were analysed using R integrated software (validation cohort). Digital spatial profiling (DSP) was used to analyse paraffin-embedded GEP-NETs. Human duodenal organoids were treated with 5 or 10 ng/mL of tumor necrosis factor alpha (TNFα) prior to qPCR and western blot analysis of neuroendocrine cell specification genes.ResultsBoth the discovery and validation cohorts of small intestinal neuroendocrine tumours induced expression of mesenchymal and calcium signalling pathways coincident with a decrease in intestine-specific genes. In particular, calcium-related, smooth muscle and cytoskeletal genes increased in DGASTs, but did not correlate with MEN1 mutation status. Interleukin 17 (IL-17) and tumor necrosis factor alpha (TNFα) signalling pathways were elevated in the DGAST RNA-sequencing. However, DSP analysis confirmed a paucity of immune cells in DGASTs compared with the adjacent tumour-associated Brunner’s glands. Immunofluorescent analysis showed production of these proinflammatory cytokines and phosphorylated signal transducer and activator of transcription 3 (pSTAT3) by the tumours and stroma. Human duodenal organoids treated with TNFα induced neuroendocrine tumour genes, SYP, CHGA and NKX6.3.ConclusionsStromal–epithelial interactions induce proinflammatory cytokines that promote Brunner’s gland reprogramming.

Objectives Paragangliomas of the head and neck and cranial base are typically benign, slow-growing tumors arising within the jugular foramen, middle ear, carotid bifurcation, or vagus nerve proper. The objective of this study was to provide a comprehensive characterization of our institutional experience with clinical management of these tumors and posit an algorithm for diagnostic evaluation and treatment. Methods This was a retrospective cohort study of patients undergoing treatment for paragangliomas of the head and neck and cranial base at our institution from 2000–2017. Data on tumor location, catecholamine levels, and specific imaging modalities employed in diagnostic work-up, pre-treatment cranial nerve palsy, treatment modality, utilization of preoperative angiographic embolization, complications of treatment, tumor control and recurrence, and hereditary status (ie, succinate dehydrogenase mutations) were collected and summarized. Results The mean (SD) age of our cohort was 51.8 (±16.1) years with 123 (63.4%) female patients and 71 (36.6%) male patients. Catecholamine-secreting lesions were found in nine (4.6%) patients. Fifty-one patients underwent genetic testing, with mutations identified in 43 (20 SDHD, 13 SDHB, 7 SDHD, 1 SDHA, SDHAF2, and NF1). Observation with serial imaging, surgical extirpation, radiation, and stereotactic radiosurgery were variably employed as treatment approaches across anatomic subsites. Conclusion An algorithmic approach to clinical management of these tumors, derived from our longitudinal institutional experience and current empiric evidence, may assist otolaryngologists, radiation oncologists, and geneticists in the care of these complex neoplasms. Level of Evidence 4

The recognition of hereditary causes of primary hyperparathyroidism (pHPT) is important because clinical care and surveillance differ significantly between sporadic and hereditary pHPT. In addition, the increasing number of genetic tests poses a challenge to classify mutations as benign or pathogenic. Functional work-up of variants remains a mainstay to provide evidence for pathogenicity. We describe a 52-year-old male patient with recurrent pHPT since age 35 years. Despite several neck surgeries with complete parathyroidectomy, he experienced persistent pHPT, necessitating repeated surgery for a forearm autotransplant, which finally resulted in unmeasurable parathyroid hormone (PTH) levels. Genetic testing revealed a new CDC73 variant (c.238-8G>A [IVS2-8G>A]), initially classified as a variant of uncertain significance. Parathyroid tissue from the initial surgeries showed loss of heterozygosity. Using an RT-PCR approach, we show that the mutation leads to the use of a cryptic splice site in peripheral mononuclear cells. In addition, a minigene approach confirms the use of the cryptic splice site in a heterologous cell system. The novel c.238-8G>A CDC73 variant activates a cryptic splice site, and the functional data provided justify the classification as a likely pathogenic variant. Our results underscore the importance of functional work-up for variant classification in the absence of other available data, such as presence in disease-specific databases, other syndromic clinical findings, or family history. In addition, the presented case exemplifies the importance to consider a hereditary condition in young patients with pHPT, particularly those with multi-gland involvement. © 2017 American Society for Bone and Mineral Research.

A concerted effort has been made in the past decade to better differentiate benign from malignant adrenocortical tumors. Of those tumors found to be adrenocortical carcinomas (ACCs) and through the use of multiple modalities including biochemical, radiologic, and genomic analysis, significant strides have been made in understanding what drives ACC development, how various treatments may result in different outcomes, which ACCs are more likely to respond to various treatments, and overall prognosis. While most patients will have recurrence of their ACC and succumb to their disease, the disease course is highly variable; it is therefore imperative that each patient is treated with individualized attention paid to their particular ACC. This article highlights and discusses specific, important, and many times subtle features that may impact the evaluation, management, treatment selection, and outcome of patients with ACC. Abbreviations: ACC = adrenocortical carcinoma; CT = computed tomography; XRT = external beam radiation therapy

Adrenocortical carcinoma (ACC) is a rare, often fatal disease, that may be seen sporadically or with hereditary predisposition syndromes. Patients with ACC are usually girls under the age of seven who present signs of excess production of adrenal glucocorticoids and androgens, with the diagnosis being confirmed by imaging. Here we reproduce and examine what we believe to be the first autopsy case report of a child with ACC, reported by Dr. Henry Sampson in Philosophical Transactions, published by The Royal Society of London in 1697. The paper describes the autopsy of a girl with severe virilization and profound signs of Cushing syndrome who died at age six, strongly suggesting ACC. She apparently had extensive pulmonary metastases, and may have had liver involvement. The report indicates her disease arose from her left kidney and there is no indication of an adrenal origin, perhaps because the adrenal gland was not generally known as a separate organ at that time. This classic example of an early case report is particularly instructive in the context of medical knowledge and understanding in the 17th century compared to current knowledge.

Adrenocortical carcinoma (ACC) is a rare malignancy with frequently dismal prognosis and few therapies. Patients with locoregional ACC routinely receive surgery and adjuvant mitotane, but >50% recur with metastases even after complete resection. Histological grade (Ki67>10% or >20 mitoses/50 HPF) is the strongest clinical predictor of recurrence, but time to recurrence in patients with high grade disease is variable and patients with low grade disease frequently recur. Recent molecular profiling studies suggest risk stratification by DNA methylation may better identify patients with homogenously dismal outcomes; in The Cancer Genome Atlas study on ACC (ACC-TCGA), we similarly identified that patients with tumors bearing CpG island hypermethylation (CIMP-high) exhibit rapid recurrence and early death. However, clinical translation of this complex molecular signature remains challenging. Here, we reanalyzed ACC-TCGA data and show that CIMP-high ACC is characterized by upregulation of cell cycle and DNA damage response programs, and identify that hypermethylation and silencing of G0S2 distinguishes this subgroup. We evaluated G0S2 methylation in a multi-institutional retrospective cohort of treatment-naive primary ACC (n=80) and adrenocortical adenomas (n=22) using a straightforward, overnight restriction digest/qPCR-based assay, validated by targeted bisulfite sequencing. We identified that G0S2 hypermethylation is exclusive to a subset of ACC (40%). G0S2 hypermethylation is associated with decreased disease-free survival (median DFS=14 mo., HR=6.91, p

Several somatic mutations specific to aldosterone-producing adenomas (APAs) have been described. A small proportion of adrenocortical carcinomas (ACCs) are associated with hyperaldosteronism, either primary aldosteronism or hyperreninemic hyperaldosteronism. However, it is unknown whether they harbor mutations of the same spectrum as APAs. The objective of this study is to describe the clinical phenotype and molecular genotype of ACCs with hyperaldosteronism, particularly the analysis for common APA-associated genetic changes. Patients were identified by retrospective chart review at a specialized referral center and by positive staining for CYP11B2 of tissue microarrays. Twenty-five patients with ACC and hyperaldosteronism were initially identified by retrospective chart review, and tissue for further analysis was available on 13 tumors. Seven patients were identified by positive staining for CYP11B2 in a tissue microarray, of which two were already identified in the initial chart review. Therefore, a total number of 18 patients with a diagnosis of ACC and features of either primary aldosteronism or hyperreninemic hyperaldosteronism were therefore included in the final study. Mutational status for a select list of oncogenes, tumor suppressor genes and genes known to carry mutations in APAs were analyzed by next-generation sequencing. Review of clinical data suggested autonomous aldosterone production in the majority of cases, while for some cases, hyperreninemic hyperaldosteronism was the more likely mechanism. The mutational landscape of ACCs associated with hyperaldosteronism was not different from ACCs with a different hormonal phenotype. None of the ACCs harbored mutations of known APA-associated genes, suggesting an alternative mechanism conferring aldosterone production.

Context Adrenocortical carcinoma (ACC) is a rare malignancy with high rates of recurrence and poor prognosis. The role of radiotherapy (RT) in localized ACC has been controversial, and RT is not routinely offered. Objective To evaluate the benefit of adjuvant RT on outcomes in ACC. Design This is a retrospective propensity-matched analysis. Setting All patients were seen through the University of Michigan’s Endocrine Oncology program, and all those who underwent RT were treated at the University of Michigan. Participants Of 424 patients with ACC, 78 were selected; 39 patients underwent adjuvant radiation. Intervention Adjuvant RT to the tumor bed and adjacent lymph nodes. Main Outcomes Measures Time to local failure, distant failure, or death. Results Median follow-up time was 4.21 years (95% CI, 2.79 to 4.94). The median radiation dose was 55 Gy (range, 45 to 60). The 3-year overall survival estimate for patients improved from 48.6% for patients without RT (95% CI, 29.7 to 65.2) to 77.7% (95% CI, 56.3 to 89.5) with RT, with a hazard ratio (HR) of 3.59 (95% CI, 1.60 to 8.09; P = 0.002). RT improved local recurrence-free survival (RFS) from 34.2% (95% CI, 18.8 to 50.3) to 59.5% (95% CI, 39.0 to 75.0), with an HR of 2.67 (95% CI, 1.38 to 5.19; P = 0.0035). RT improved all RFS from 18.3% (95% CI, 6.7 to 34.3) to 46.7% (95% CI, 26.9 to 64.3), with an HR 2.59 (95% CI, 1.40 to 4.79; P = 0.0024). Conclusions In the largest single institution study to date, adjuvant RT after gross resection of ACC improved local RFS, all RFS, and overall survival in this propensity-matched analysis. Adjuvant RT should be considered a part of multidisciplinary management for patients with ACC.

© 2019 Elsevier Ltd. All rights reserved., Background: Multiple endocrine neoplasia type 2B is a rare syndrome caused mainly by Met918Thr germline RET mutation, and characterised by medullary thyroid carcinoma, phaeochromocytoma, and extra-endocrine features. Data are scarce on the natural history of multiple endocrine neoplasia type 2B. We aimed to advance understanding of the phenotype and natural history of multiple endocrine neoplasia type 2B, to increase awareness and improve detection. Methods: This study was a retrospective, multicentre, international study in patients carrying the Met918Thr RET variant with no age restrictions. The study was done with registry data from 48 centres globally. Data from patients followed-up from 1970 to 2016 were retrieved from May 1, 2016, to May 31, 2018. Our primary objectives were to determine overall survival, and medullary thyroid carcinoma-specific survival based on whether the patient had undergone early thyroidectomy before the age of 1 year. We also assessed remission of medullary thyroid carcinoma, incidence and treatment of phaeochromocytoma, and the penetrance of extra-endocrine features. Findings: 345 patients were included, of whom 338 (98%) had a thyroidectomy. 71 patients (21%) of the total cohort died at a median age of 25 years (range

PURPOSE DICER1 syndrome is a recently described inherited cancer predisposition syndrome caused by pathogenic variants in DICER1. With the recent increase in integrative clinical sequencing for pediatric patients with cancer, our understanding of the DICER1 syndrome continues to evolve, as new and rare pathogenic variants are reported. As the frequency of integrative clinical sequencing increases, discussions regarding challenges encountered in the interpretation of sequencing results are essential to continue to advance the field of cancer predisposition. The purpose of this work was to identify patients with somatic and/or germline DICER1 variants in our patient population and to discuss sequencing interpretation and the clinical recommendations that result from the integrative clinical sequencing results. METHODS Patients were enrolled in the PEDS-MIONCOSEQ study. This integrative clinical sequencing study includes paired tumor/normal whole-exome sequencing and tumor transcriptome sequencing. Patients identified as having DICER1 variants were included. RESULTS We report a DICER1 variant of unknown clinical significance in a patient with a highly unusual response to therapy. Two patients had diagnoses clarified once the integrative clinical sequencing revealing a DICER1 variant was available. We also discovered a patient with low-level DICER1 mosaicism and the challenges encountered in the sequencing interpretation for this patient. In addition to the sequencing data and result interpretation, this work also highlights testing and screening recommendations made to patients with DICER1 variants and their families on the basis of these results. CONCLUSION This work serves to extend the DICER1 phenotype and advance the utility of clinical integrative sequencing in the fields of pediatric oncology and cancer genetic predisposition.

4555 Background: Inactivation of VHL leads to aberrant stabilization and accumulation of HIF-2α, which drives tumor growth. Patients (pts) with VHL disease are at risk for ccRCC, pancreatic neuroendocrine tumors (pNETs), and hemangioblastomas. Repeated surgeries are often needed to control ccRCC and other VHL disease manifestations. Prior results of this ongoing open-label phase 2 study (NCT03401788) showed activity with belzutifan in VHL disease. Updated results are presented. Methods: Adults with germline VHL alterations, measurable and localized/nonmetastatic ccRCC, no prior systemic anticancer therapy, and ECOG PS 0 or 1 received belzutifan 120 mg once daily until progression, intolerable toxicity, or decision to withdraw. The primary end point is ORR of VHL-associated ccRCC tumors per RECIST v1.1 by independent review committee (IRC). Secondary end points include DOR, time to response (TTR), PFS, and safety. Results: As of June 1, 2020, 61 pts enrolled. Most pts (82%) had ECOG PS 0, and the median number of prior tumor reduction procedures (eg, partial nephrectomy, craniotomy, radiation therapy) per pt was 5 (range, 0-15). Lesions outside the kidney (non-RCC tumors) evaluable by IRC included pNETs (33%) and CNS hemangioblastomas (82%). Median follow-up was 69 wk (range, 18-105), median duration of treatment was 68 wk (range, 8-105), and 56 pts (92%) remain on therapy. There were 22 confirmed responses (ORR, 36% [95% CI, 24-49]) and 7 (11%) unconfirmed responses (documented at 1 time point, to be confirmed at subsequent time point); all were PRs. In pts with confirmed PR, median DOR was not reached (range, 12+ to 62+ wk), median TTR was 31 wk (range, 12-61), and 56 pts (92%) had some reduction in the sum of all target lesion diameters. PFS rate at 52 wk was 98% (95% CI, 89-100). For non-RCC tumors, ORR was 80% (16/20; 1 CR) in pNETs and 32% (16/50; 1 CR) in CNS hemangioblastomas. Of 16 pts with evaluable retinal hemangioblastomas at baseline, 11 (69%) showed improvement per IRC. In those 16 pts, 29 eyes were monitored for retinal hemangioblastomas: 16 eyes (55%) showed improvement, 12 (41%) were stable, and no evaluation was available for 1 eye (3%). All 61 pts (100%) had at least one AE. The most common all-cause AE was anemia (90%), which is considered an on-target toxicity. Treatment-related AEs (TRAE) were reported by 60 pts (98%), and 8 pts (13%) had a grade 3 TRAE. No pts had grade 4/5 TRAEs. One pt discontinued treatment because of a TRAE (grade 1 dizziness). Conclusions: Belzutifan demonstrates clinical benefit and has a favorable safety profile in patients with VHL disease–associated ccRCC, pNETs, and hemangioblastomas. Clinical trial information: NCT03401788.

333 Background: Patients (pts) with VHL disease are at risk of developing benign and malignant tumors, including ccRCC, pancreatic lesions tumors, CNS hemangioblastomas, and retinal lesions. Inactivation of VHL leads to stabilization of HIF-2α, which drives tumor growth. In a phase 1/2 study, MK-6482, a potent, selective, oral small molecule HIF-2α inhibitor, demonstrated favorable safety and antitumor activity in advanced ccRCC. We present results of the open-label phase 2 study of MK-6482 for VHL disease–associated ccRCC (NCT03401788). Methods: Adults with germline VHL alterations, measurable, localized/non-metastatic ccRCC, no prior systemic anticancer therapy, and ECOG PS 0/1 received MK-6482 120 mg once daily until progression, intolerable toxicity, or decision to withdraw. Primary end point: ORR of VHL-associated ccRCC tumors per RECIST v1.1 by independent review committee (IRC). Secondary end points: DOR, time to response (TTR), PFS, and safety. Results: As of June 1, 2020, 61 pts enrolled. The majority (82%) of pts had ECOG PS 0, and median number of prior surgeries per pt was 5 (range, 1-15). Lesions outside the kidney (non-RCC tumors) evaluable by IRC included pancreatic lesions (100%) and CNS hemangioblastomas (70%). Median duration of treatment was 68 wk (range, 8-105), and 92% of pts remain on therapy. There were 22 confirmed responses (ORR, 36% [95% CI, 24%-49%]) and 7 (11%) unconfirmed (documented at 1 time point, to be confirmed at subsequent time point) responses; all PRs. In pts with confirmed PR, median DOR was not reached (range, 12-62 wk) and median TTR was 31 wk (range, 12-61); 14 (64%) pts had response duration of ≥26 wk. 56 pts (92%) had any decrease in size of target lesions. PFS rate at 52 wk was 98% (95% CI, 89%-100%). Overall, pretreatment median linear growth rate of ccRCC tumors was +3.6 mm/y (range, −3.4 to +33.1), compared with −4.5 mm/y (range, −12.8 to +5.1) while on treatment. For non-RCC tumors, ORR in pancreatic lesions was 64% (39/61, including 4 CRs) and in CNS hemangioblastomas was 30% (13/43, including 5 CRs). Median (range) TTR was 35 wk (11-60) and 12 wk (10-60) for pancreatic lesions and CNS hemangioblastomas, respectively. 11/16 (69%) pts with evaluable retinal lesions at baseline showed improvement. Of those 16 pts, 29 eyes were followed for retinal lesions; 16 eyes (55%) showed improvement, 12 (41%) remained stable, and no follow-up evaluation was available for 1 (3%) eye. Treatment-related AEs (TRAEs) occurred in 98% of pts, none grade 4/5. Most common TRAE was anemia (87%), considered to be an on-target toxicity. One pt discontinued treatment due to a TRAE (grade 1 dizziness). As of data cutoff, 1 pt (2%) required surgery for ccRCC tumors after treatment. Conclusions: MK-6482 is an active and well-tolerated therapy for VHL disease–associated ccRCC, pancreatic lesions, as well as CNS and retinal hemangioblastomas. Clinical trial information: NCT03401788 .

Objective Co-secretion of cortisol and aldosterone can be observed in adrenal adenomas. The aim of this study was to investigate the molecular characteristics of a co-existing aldosterone- and a cortisol-producing adenoma (CPA) in the same patient. Design and methods Two different adenomas within the same adrenal gland from a 49-year-old female patient with primary aldosteronism (PA) and Cushing's syndrome (CS) were studied. Multiple formalin-fixed paraffin-embedded tumor blocks were used for the analysis. Immunohistochemistry (IHC) was performed using a specific antibody against aldosterone synthase (CYP11B2). DNA and RNA were isolated separately from CYP11B2-positive and -negative tumor regions based on CYP11B2 IHC results. Results CYP11B2 IHC clearly demonstrated that three pieces from one adenoma were positive for CYP11B2 and the remaining three from the other adenoma were negative for CYP11B2. In quantitative real-time RT-PCR, CYP11B2 mRNA was upregulated in CYP11B2-positive tumor specimens (219-fold vs CYP11B2-negative tumor specimens). Targeted next-generation sequencing (NGS) detected novel KCNJ5 gene mutations (p.T148I/T149S, present in the same reads) and a PRKACA gene hotspot mutation (p.L206R) in the CYP11B2-positive and -negative tumors, respectively. Sanger sequencing of DNA from each tumor specimen (CYP11B2-positive tumor, n=3; CYP11B2-negative tumor, n=3) showed concordant results with targeted NGS. Conclusion Our findings illustrate the co-existence of two different adrenocortical adenomas causing the concurrent diagnosis of PA and CS in the same patient. Molecular analysis was able to demonstrate that the two diseases resulted from independent somatic mutations seen in double adrenocortical adenomas.

We describe a comprehensive genomic characterization of adrenocortical carcinoma (ACC). Using this dataset, we expand the catalogue of known ACC driver genes to include PRKAR1A, RPL22, TERF2, CCNE1, and NF1. Genome wide DNA copy-number analysis revealed frequent occurrence of massive DNA loss followed by whole-genome doubling (WGD), which was associated with aggressive clinical course, suggesting WGD is a hallmark of disease progression. Corroborating this hypothesis were increased TERT expression, decreased telomere length, and activation of cell-cycle programs. Integrated subtype analysis identified three ACC subtypes with distinct clinical outcome and molecular alterations which could be captured by a 68-CpG probe DNA-methylation signature, proposing a strategy for clinical stratification of patients based on molecular markers.