Molecular tumor board: molecularly adjusted therapy upon identification and functional validation of a novel ALK resistance mutation in a case of lung adenocarcinoma.

We report a case of a long-term surviving patient with EML4/ALK translocated non-small cell adenocarcinoma of the lung in UICC8 stage IVA. During recurrence under continuous crizotinib therapy, a hitherto insufficiently characterized missense mutation in the ALK gene (Arg1181His) was identified through targeted sequencing. The aforementioned EML4/ALK translocation could still be detected in this situation. Employing a 3D reconstruction of the ALK tertiary structure, considering its interaction with various ALK inhibitors at the molecular binding site, our analysis indicated the presence of a mutation associated with crizotinib resistance. To validate the biological relevance of this previously unknown mutation, we carried out an in vitro validation approach in cell culture in addition to the molecular diagnostics accompanied by the molecular tumor board. The tumor scenario was mimicked through retroviral transfection. Our comparative in vitro treatment regimen paired with the clinical trajectory of the patient, corroborated our initial clinical and biochemical suspicions. Our approach demonstrates preclinical, in silico, and clinical evidence of a novel crizotinib resistance mutation in ALK as well as sensitivity toward brigatinib and potentially lorlatinib. In future cases, this procedure represents an important contribution to functional diagnostics in the context of molecular tumor boards.

Responses to systemic therapy in metastatic pheochromocytoma/paraganglioma: a retrospective multicenter cohort study.

OBJECTIVE: The therapeutic options for metastatic pheochromocytomas/paragangliomas (mPPGLs) include chemotherapy with cyclophosphamide/vincristine/dacarbazine (CVD), temozolomide monotherapy, radionuclide therapies, and tyrosine kinase inhibitors such as sunitinib. The objective of this multicenter retrospective study was to evaluate and compare the responses of mPPGLs including those with pathogenic variants in succinate dehydrogenase subunit B (SDHB), to different systemic treatments. DESIGN: This is a retrospective analysis of treatment responses of mPPGL patients (n = 74) to systemic therapies. METHODS: Patients with mPPGLs treated at 6 specialized national centers were selected based on participation in the ENSAT registry. Survival until detected progression (SDP) and disease-control rates (DCRs) at 3 months were evaluated based on imaging reports. RESULTS: For the group of patients with progressive disease at baseline (83.8% of 74 patients), the DCR with first-line CVD chemotherapy was 75.0% (n = 4, SDP 11 months; SDHB [n = 1]: DCR 100%, SDP 30 months), with somatostatin peptide receptor-based radionuclide therapy (PPRT) 85.7% (n = 21, SDP 17 months; SDHB [n = 10]: DCR 100%, SDP 14 months), with 131I-meta-iodobenzylguanidine (131I-MIBG) 82.6% (n = 23, SDP 43 months; SDHB [n = 4]: DCR 100%, SDP 24 months), with sunitinib 100% (n = 7, SDP 18 months; SDHB [n = 3]: DCR 100%, SDP 18 months), and with somatostatin analogs 100% (n = 4, SDP not reached). The DCR with temozolomide as second-line therapy was 60.0% (n = 5, SDP 10 months; SDHB [n = 4]: DCR 75%, SDP 10 months). CONCLUSIONS: We demonstrate in a real-life clinical setting that all current therapies show reasonable efficacy in preventing disease progression, and this is equally true for patients with germline SDHB mutations.

Regulation of epinephrine biosynthesis in HRAS-mutant paragangliomas.

The biochemical phenotype of paragangliomas (PGLs) is highly dependent on the underlying genetic background and tumor location. PGLs at extra-adrenal locations usually do not express phenylethanolamine N-methyltransferase (PNMT), the enzyme required for epinephrine production, which was explained by the absence of glucocorticoids. PGLs with pathogenic variants (PVs) in Harvey rat sarcoma viral oncogene homolog (HRAS) can occur in or outside of the adrenal, but always synthesize epinephrine independently of the localization. Here, we characterize the signaling pathways through which PVs in HRAS influence PNMT expression. Catecholamines, cortisol, and transcriptional features of PGL tissues with known genetic background were analyzed. Genetically modified rat pheochromocytoma cells carrying PVs in Hras were generated and analyzed for regulation of Pnmt expression. Elevated epinephrine contents in PGLs with PVs in HRAS were accompanied by enrichment in mitogen-activated protein kinase (MAPK) signaling compared to PGLs with PVs in genes that activate hypoxia pathways. In vitro, Hras PVs increased Pnmt expression and epinephrine biosynthesis through increased phosphorylation of stimulatory protein 1 via MAPK signaling. Here, we provide a molecular mechanism that explains the PV-dependent epinephrine production of PGLs.

Molecularly Stratified Treatment Options in Primary Refractory DLBCL/HGBL with MYC and BCL2 or BCL6 Rearrangements (HGBL, NOS with MYC/BCL6).

BACKGROUND: There is growing evidence supporting multidisciplinary molecular tumor boards (MTB) in solid tumors whereas hematologic malignancies remain underrepresented in this regard. OBJECTIVE: The present study aimed to assess the clinical relevance of MTBs in primary refractory diffuse large B-cell lymphomas/high-grade B-cell lymphomas with MYC and BCL2 rearrangements (prDLBCL/HGBL-MYC/BCL2) (n = 13) and HGBL, not otherwise specified (NOS), with MYC and BCL6 rearrangements (prHGBL, NOS-MYC/BCL6) (n = 6) based on our previously published whole-exome sequencing (WES) cohort. PATIENTS AND METHODS: For genomic analysis, the institutional MTB WES pipeline (University Cancer Center Schleswig-Holstein: UCCSH), certified for routine clinical diagnostics, was employed and supplemented by a comprehensive immunohistochemical work-up. Consecutive database research and annotation according to established evidence levels for molecularly stratified therapies was performed (NCT-DKTK/ESCAT). RESULTS: Molecularly tailored treatment options with NCT-DKTK evidence level of at least m2A were identified in each case. We classified mutations in accordance with biomarker/treatment baskets and detected a heterogeneous spectrum of targetable alterations affecting immune evasion (IE; n = 30), B-cell targets (BCT; n = 26), DNA damage repair (DDR; n = 20), tyrosine kinases (TK; n = 13), cell cycle (CC; n = 7), PI3K-MTOR-AKT pathway (PAM; n = 2), RAF-MEK-ERK cascade (RME; n = 1), and others (OTH; n = 11). CONCLUSION: Our virtual MTB approach identified potential molecularly targeted treatment options alongside targetable genomic signatures for both prDLBCL/HGBL-MYC/BCL2 and prHGBL, NOS-MYC/BCL6. These results underline the potential of MTB consultations in difficult-to-treat lymphomas early in the treatment sequence.

Metastatic Pheochromocytoma and Paraganglioma: Somatostatin Receptor 2 Expression, Genetics, and Therapeutic Responses.

CONTEXT: Pheochromocytomas and paragangliomas (PPGLs) with pathogenic mutations in the succinate dehydrogenase subunit B (SDHB) are associated with a high metastatic risk. Somatostatin receptor 2 (SSTR2)-dependent imaging is the most sensitive imaging modality for SDHB-related PPGLs, suggesting that SSTR2 expression is a significant cell surface therapeutic biomarker of such tumors. OBJECTIVE: Exploration of the relationship between SSTR2 immunoreactivity and SDHB immunoreactivity, mutational status, and clinical behavior of PPGLs. Evaluation of SSTR-based therapies in metastatic PPGLs. METHODS: Retrospective analysis of a multicenter cohort of PPGLs at 6 specialized Endocrine Tumor Centers in Germany, The Netherlands, and Switzerland. Patients with PPGLs participating in the ENSAT registry were included. Clinical data were extracted from medical records, and immunohistochemistry (IHC) for SDHB and SSTR2 was performed in patients with available tumor tissue. Immunoreactivity of SSTR2 was investigated using Volante scores. The main outcome measure was the association of SSTR2 IHC positivity with genetic and clinical-pathological features of PPGLs. RESULTS: Of 202 patients with PPGLs, 50% were SSTR2 positive. SSTR2 positivity was significantly associated with SDHB- and SDHx-related PPGLs, with the strongest SSTR2 staining intensity in SDHB-related PPGLs (P = .01). Moreover, SSTR2 expression was significantly associated with metastatic disease independent of SDHB/SDHx mutation status (P < .001). In metastatic PPGLs, the disease control rate with first-line SSTR-based radionuclide therapy was 67% (n = 22, n = 11 SDHx), and with first-line "cold" somatostatin analogs 100% (n = 6, n = 3 SDHx). CONCLUSION: SSTR2 expression was independently associated with SDHB/SDHx mutations and metastatic disease. We confirm a high disease control rate of somatostatin receptor-based therapies in metastatic PPGLs.

Primary refractory plasmablastic lymphoma: A precision oncology approach.

Introduction: Hematologic malignancies are currently underrepresented in multidisciplinary molecular-tumor-boards (MTB). This study assesses the potential of precision-oncology in primary-refractory plasmablastic-lymphoma (prPBL), a highly lethal blood cancer. Methods: We evaluated clinicopathological and molecular-genetic data of 14 clinically annotated prPBL-patients from initial diagnosis. For this proof-of-concept study, we employed our certified institutional MTB-pipeline (University-Cancer-Center-Schleswig-Holstein, UCCSH) to annotate a comprehensive dataset within the scope of a virtual MTB-setting, ultimately recommending molecularly stratified therapies. Evidence-levels for MTB-recommendations were defined in accordance with the NCT/DKTK and ESCAT criteria. Results: Median age in the cohort was 76.5 years (range 56-91), 78.6% of patients were male, 50% were HIV-positive and clinical outcome was dismal. Comprehensive genomic/transcriptomic analysis revealed potential recommendations of a molecularly stratified treatment option with evidence-levels according to NCT/DKTK of at least m2B/ESCAT of at least IIIA were detected for all 14 prPBL-cases. In addition, immunohistochemical-assessment (CD19/CD30/CD38/CD79B) revealed targeted treatment-recommendations in all 14 cases. Genetic alterations were classified by treatment-baskets proposed by Horak et al. Hereby, we identified tyrosine-kinases (TK; n=4), PI3K-MTOR-AKT-pathway (PAM; n=3), cell-cycle-alterations (CC; n=2), RAF-MEK-ERK-cascade (RME; n=2), immune-evasion (IE; n=2), B-cell-targets (BCT; n=25) and others (OTH; n=4) for targeted treatment-recommendations. The minimum requirement for consideration of a drug within the scope of the study was FDA-fast-track development. Discussion: The presented proof-of-concept study demonstrates the clinical potential of precision-oncology, even in prPBL-patients. Due to the aggressive course of the disease, there is an urgent medical-need for personalized treatment approaches, and this population should be considered for MTB inclusion at the earliest time.

Genomic and immune landscape Of metastatic pheochromocytoma and paraganglioma.

The mechanisms triggering metastasis in pheochromocytoma/paraganglioma are unknown, hindering therapeutic options for patients with metastatic tumors (mPPGL). Herein we show by genomic profiling of a large cohort of mPPGLs that high mutational load, microsatellite instability and somatic copy-number alteration burden are associated with ATRX/TERT alterations and are suitable prognostic markers. Transcriptomic analysis defines the signaling networks involved in the acquisition of metastatic competence and establishes a gene signature related to mPPGLs, highlighting CDK1 as an additional mPPGL marker. Immunogenomics accompanied by immunohistochemistry identifies a heterogeneous ecosystem at the tumor microenvironment level, linked to the genomic subtype and tumor behavior. Specifically, we define a general immunosuppressive microenvironment in mPPGLs, the exception being PD-L1 expressing MAML3-related tumors. Our study reveals canonical markers for risk of metastasis, and suggests the usefulness of including immune parameters in clinical management for PPGL prognostication and identification of patients who might benefit from immunotherapy.

Recurrent Disease in Patients With Sporadic Pheochromocytoma and Paraganglioma.

CONTEXT: Long-term follow-up has been recommended for patients with pheochromocytoma or paraganglioma (PPGL) due to potential for recurrent disease. However, the need to follow patients with sporadic PPGL has recently become controversial. OBJECTIVE: To investigate the prevalence of recurrence among patients with sporadic compared with hereditary PPGL and to identify predictors of recurrence for sporadic disease. METHODS: This multicenter study included retrospective data from 1127 patients with PPGL. In addition to sex and age at primary tumor diagnosis, clinical information included location, size, and catecholamine phenotype of primary tumors, genetic test results, and subsequent development of recurrent and/or metastatic disease. Patients with sporadic PPGL were defined as those with negative genetic test results. RESULTS: Prevalence of recurrence among patients with sporadic PPGL (14.7%) was lower (P < 0.001) than for patients with pathogenic variants that activate pseudohypoxia pathways (47.5%), but similar to those with variants that activate kinase pathways (14.9%). Among patients with sporadic recurrent PPGL, 29.1% and 17.7% were respectively diagnosed at least 10 and 15 years after first diagnosis. Multivariable regression analysis showed that a noradrenergic/dopaminergic phenotype (HR 2.73; 95% CI, 1.553-4.802; P < 0.001), larger size (HR 1.82; 95% CI, 1.113-2.962; P = 0.017) and extra-adrenal location (HR 1.79; 95% CI, 1.002-3.187; P = 0.049) of primary tumors were independent predictors of recurrence in sporadic PPGL. CONCLUSION: Patients with sporadic PPGL require long-term follow-up, as supported by the 14.7% prevalence of recurrent disease, including recurrences at more than 10 years after first diagnosis. The nature of follow-up could be individualized according to tumor size, location, and biochemical phenotype.

Co-occurrence of mutations in NF1 and other susceptibility genes in pheochromocytoma and paraganglioma.

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

Succinate Mediates Tumorigenic Effects via Succinate Receptor 1: Potential for New Targeted Treatment Strategies in Succinate Dehydrogenase Deficient Paragangliomas.

Paragangliomas and pheochromocytomas (PPGLs) are chromaffin tumors associated with severe catecholamine-induced morbidities. Surgical removal is often curative. However, complete resection may not be an option for patients with succinate dehydrogenase subunit A-D (SDHx) mutations. SDHx mutations are associated with a high risk for multiple recurrent, and metastatic PPGLs. Treatment options in these cases are limited and prognosis is dismal once metastases are present. Identification of new therapeutic targets and candidate drugs is thus urgently needed. Previously, we showed elevated expression of succinate receptor 1 (SUCNR1) in SDHB PPGLs and SDHD head and neck paragangliomas. Its ligand succinate has been reported to accumulate due to SDHx mutations. We thus hypothesize that autocrine stimulation of SUCNR1 plays a role in the pathogenesis of SDHx mutation-derived PPGLs. We confirmed elevated SUCNR1 expression in SDHx PPGLs and after SDHB knockout in progenitor cells derived from a human pheochromocytoma (hPheo1). Succinate significantly increased viability of SUCNR1-transfected PC12 and ERK pathway signaling compared to control cells. Candidate SUCNR1 inhibitors successfully reversed proliferative effects of succinate. Our data reveal an unrecognized oncometabolic function of succinate in SDHx PPGLs, providing a growth advantage via SUCNR1.

Metastatic pheochromocytoma and paraganglioma: signs and symptoms related to catecholamine secretion.

BACKGROUND: The presence or future development of metastatic pheochromocytomas or paragangliomas (mPPGLs) can be difficult to diagnose or predict at initial presentation. Since production of catecholamines from mPPGLs is different from non-metastatic tumors (non-mPPGLs), this study aimed to clarify whether presenting catecholamine-related signs and symptoms (cSS) might also differ. METHODS: The study included 249 patients, 43 with mPPGL and 206 with non-mPPGL. Clinical data at the time of biochemical diagnosis (i.e. at entry into the study) were used to generate a cumulative score of cSS for each patient. RESULTS: Patients with mPPGL were significantly younger (43.3 ± 14 vs. 48.9 ± 16.1 years) and included a lower proportion of females (39.5% vs. 60.7%) than patients with non-mPPGLs. Frequencies of signs and symptoms did not differ between the two groups. Patients with mPPGLs had lower (P < 0.001) urinary excretion of epinephrine (3.5 (IQR, 1.9-6.5) µg/day) than those with non-mPPGLs (19.1 (IQR, 4.3-70.2) µg/day). There was no difference in urinary excretion of norepinephrine. In patients with mPPGLs a high cSS score was associated with high urinary excretion of norepinephrine and normetanephrine. In contrast, in patients with non-mPPGLs, a high cSS was associated with high urinary excretion of epinephrine and metanephrine. CONCLUSION: Although presenting signs and symptoms were associated with production of norepinephrine in patients with mPPGLs and of epinephrine in patients with non-mPPGLs, there were no differences in signs and symptoms between the two groups. Therefore, consideration of signs and symptoms does not appear helpful for distinguishing patients with and without mPPGLs.

Ganglioneuromas across age groups: Systematic review of individual patient data.

BACKGROUND: Ganglioneuromas are very rare tumours of the sympathetic nervous system. Clinical and pathological knowledge is currently based on largely incomparable registries and case series that focus on paediatric or adrenal cases. To comprehensively characterize the full clinical spectrum across ages and locations, a meta-analysis was performed where amenable and complemented by systematic literature review of individual patient data (IPD). DESIGN: Articles containing "ganglioneuroma" in English on humans, published from 1/1/1995-6/27/2018, were identified from PubMed. Aggregate data from 10 eligible patient series on 19 variables were considerably inhomogeneous, restricting meta-analysis to age and gender distribution. To determine basic disease characteristics across ages and locations, IPD were retrieved from case reports and small case series (PROSPERO CRD42018010247). RESULTS: Individual patient data representing 364 cases revealed that 65.7% (60.6%-70.4%) were diagnosed in adults, more frequently in females (62%, 56.9%-66.9%). 24.5% (20.3%-39.1%) were discovered incidentally. Most often, ganglioneuromas developed in abdomen/pelvis (66.2, 32.1% adrenal). With age, the proportion of ganglioneuroma localizations with high post-surgical complication rate (35.6% head/neck and 16.3% thorax) decreased. Contrarily, the diagnosis of adrenal ganglioneuromas (<1% post-surgical complications) increased with age. Hormone production, hypertension or coincidence with another non-neuroblastic neural-crest-derived tumour component was more common for adrenal location. Recurrence and metastatic spread have not been reported for ganglioneuromas without secondary tumour component. CONCLUSIONS: This work summarizes characteristics of the currently largest number of international GN patients across all ages. The data confirm a benign nature of GN, independent of age. Age-related differences in predominant tumour location, associated post-surgical complications and hormone production suggest case-centred management strategies.

Plasma metanephrines and prospective prediction of tumor location, size and mutation type in patients with pheochromocytoma and paraganglioma.

Objectives: Plasma free metanephrines are commonly used for diagnosis of pheochromocytoma and paraganglioma (PPGLs), but can also provide other information. This multicenter study prospectively examined whether tumor size, location, and mutations could be predicted by these metabolites. Methods: Predictions of tumor location, size, and mutation type, based on measurements of plasma normetanephrine, metanephrine, and methoxytyramine were made without knowledge of disease in 267 patients subsequently determined to have PPGLs. Results: Predictions of adrenal vs. extra-adrenal locations according to increased plasma concentrations of metanephrine and methoxytyramine were correct in 93 and 97% of the respective 136 and 33 patients in who these predictions were possible. Predicted mean tumor diameters correlated positively (p<0.0001) with measured diameters; predictions agreed well for pheochromocytomas but were overestimated for paragangliomas. Considering only patients with mutations, 51 of the 54 (94%) patients with NF1 or RET mutations were correctly predicted with those mutations according to increased plasma metanephrine, whereas no or minimal increase in metanephrine correctly predicted all 71 patients with either VHL or SDHx mutations; furthermore, among the latter group increases in methoxytyramine correctly predicted SDHx mutations in 93% of the 29 cases for this specific prediction. Conclusions: Extents and patterns of increased plasma O-methylated catecholamine metabolites among patients with PPGLs allow predictions of tumor size, adrenal vs. extra-adrenal locations and general types of mutations. Predictions of tumor location are, however, only possible for patients with clearly increased plasma methoxytyramine or metanephrine. Where possible or clinically relevant the predictions are potentially useful for subsequent clinical decision-making.

HIF2α supports pro-metastatic behavior in pheochromocytomas/paragangliomas.

Mutations that drive the stabilization of hypoxia inducible factor 2α (HIF2α) and downstream pseudohypoxic signaling are known to predispose to the development of pheochromocytomas and paragangliomas (PPGLs). However, any role of HIF2α in predisposition to metastatic disease remains unclear. To assess such a role we combined gene-manipulations in pheochromocytoma cell lines with retrospective analyses of patient data and gene expression profiling in tumor specimens. Among 425 patients with PPGLs identified with mutations in tumor-susceptibility genes, those with tumors due to activation of pseudohypoxic pathways had a higher frequency of metastatic disease than those with tumors due to activation of kinase-signaling pathways, even without inclusion of patients with mutations in SDHB (18.6% vs 4.3% in, P < 0.0001). Three out of nine (33%) patients with gain-of-function mutations in HIF2α had metastatic disease. In cell line studies, elevated expression of HIF2α enhanced cell proliferation and led to increased migration and invasion capacity. Moreover, HIF2α expression in HIF2α-deficient cells resulted in increased cell motility, diffuse cluster formation and emergence of pseudopodia indicating changes in cell adhesion and cytoskeletal remodeling. In a mouse liver metastasis model, Hif2a enhanced the metastatic load. Transcriptomics data revealed alterations in focal adhesion and extracellular matrix-receptor interactions in HIF2α-mutated PPGLs. Our translational findings demonstrate that HIF2α supports pro-metastatic behavior in PPGLs, though other factors remain critical for subsequent transition to metastasis. We identified LAMB1 and COL4A2 as new potential therapeutic targets for HIF2α-driven PPGLs. Identified HIF2α downstream targets might open a new therapeutic window for aggressive HIF2α-expressing tumors.

Sino-European Differences in the Genetic Landscape and Clinical Presentation of Pheochromocytoma and Paraganglioma.

CONTEXT: Pheochromocytomas and paragangliomas (PPGLs) are characterized by distinct genotype-phenotype relationships according to studies largely restricted to Caucasian populations. OBJECTIVE: To assess for possible differences in genetic landscapes and genotype-phenotype relationships of PPGLs in Chinese versus European populations. DESIGN: Cross-sectional study. SETTING: 2 tertiary-care centers in China and 9 in Europe. PARTICIPANTS: Patients with pathologically confirmed diagnosis of PPGL, including 719 Chinese and 919 Europeans. MAIN OUTCOME MEASURES: Next-generation sequencing performed in tumor specimens with mutations confirmed by Sanger sequencing and tested in peripheral blood if available. Frequencies of mutations were examined according to tumor location and catecholamine biochemical phenotypes. RESULTS: Among all patients, higher frequencies of HRAS, FGFR1, and EPAS1 mutations were observed in Chinese than Europeans, whereas the reverse was observed for NF1, VHL, RET, and SDHx. Among patients with apparently sporadic PPGLs, the most frequently mutated genes in Chinese were HRAS (16.5% [13.6-19.3] vs 9.8% [7.6-12.1]) and FGFR1 (9.8% [7.6-12.1] vs 2.2% [1.1-3.3]), whereas among Europeans the most frequently mutated genes were NF1 (15.9% [13.2-18.6] vs 6.6% [4.7-8.5]) and SDHx (10.7% [8.4-13.0] vs 4.2% [2.6-5.7]). Among Europeans, almost all paragangliomas lacked appreciable production of epinephrine and identified gene mutations were largely restricted to those leading to stabilization of hypoxia inducible factors. In contrast, among Chinese there was a larger proportion of epinephrine-producing paragangliomas, mostly due to HRAS and FGFR1 mutations. CONCLUSIONS: This study establishes Sino-European differences in the genetic landscape and presentation of PPGLs, including ethnic differences in genotype-phenotype relationships indicating a paradigm shift in our understanding of the biology of these tumors.

Glucocorticoid Excess in Patients with Pheochromocytoma Compared with Paraganglioma and Other Forms of Hypertension.

CONTEXT: Catecholamines and adrenocortical steroids are important regulators of blood pressure. Bidirectional relationships between adrenal steroids and catecholamines have been established but whether this is relevant to patients with pheochromocytoma is unclear. OBJECTIVE: This study addresses the hypothesis that patients with pheochromocytoma and paraganglioma (PPGL) have altered steroid production compared with patients with primary hypertension. DESIGN: Multicenter cross-sectional study. SETTING: Twelve European referral centers. PATIENTS: Subjects included 182 patients with pheochromocytoma, 36 with paraganglioma and 270 patients with primary hypertension. Patients with primary aldosteronism (n = 461) and Cushing syndrome (n = 124) were included for additional comparisons. INTERVENTION: In patients with PPGLs, surgical resection of tumors. OUTCOME MEASURES: Differences in mass spectrometry-based profiles of 15 adrenal steroids between groups and after surgical resection of PPGLs. Relationships of steroids to plasma and urinary metanephrines and urinary catecholamines. RESULTS: Patients with pheochromocytoma had higher (P < .05) circulating concentrations of cortisol, 11-deoxycortisol, 11-deoxycorticosterone, and corticosterone than patients with primary hypertension. Concentrations of cortisol, 11-deoxycortisol, and corticosterone were also higher (P < .05) in patients with pheochromocytoma than with paraganglioma. These steroids correlated positively with plasma and urinary metanephrines and catecholamines in patients with pheochromocytoma, but not paraganglioma. After adrenalectomy, there were significant decreases in cortisol, 11-deoxycortisol, corticosterone, 11-deoxycorticosterone, aldosterone, and 18-oxocortisol. CONCLUSIONS: This is the first large study in patients with PPGLs that supports in a clinical setting the concept of adrenal cortical-medullary interactions involving an influence of catecholamines on adrenal steroids. These findings could have implications for the cardiovascular complications of PPGLs and the clinical management of patients with the tumors.

Overnight/first-morning urine free metanephrines and methoxytyramine for diagnosis of pheochromocytoma and paraganglioma: is this an option?

OBJECTIVE: Sympathoadrenal activity is decreased during overnight rest. This study assessed whether urinary-free normetanephrine, metanephrine and methoxytyramine in overnight/first-morning urine collections might offer an alternative to measurements in 24-h collections or plasma for diagnosis of pheochromocytoma and paraganglioma (PPGL). DESIGN AND METHODS: Prospective multicenter cross-sectional diagnostic study involving 706 patients tested for PPGL, in whom tumors were confirmed in 79 and excluded in 627 after follow-up. Another 335 age- and sex-matched volunteers were included for reference purposes. Catecholamines and their free O-methylated metabolites were measured in 24-h collections divided according to waking and sleeping hours and normalized to creatinine. Plasma metabolites from blood sampled after supine rest were measured for comparison. RESULTS: Urinary outputs of norepinephrine, normetanephrine, epinephrine and metanephrine in the reference population were respectively 50 (48-52)%, 35 (32-37)%, 76 (74-78)% and 15 (12-17)% lower following overnight than daytime collections. Patients in whom PPGLs were excluded showed 28 (26-30)% and 6 (3-9)% day-to-night falls in normetanephrine and metanephrine, while patients with PPGLs showed no significant day-to-night falls in metabolites. Urinary methoxytyramine was consistently unchanged from day to night. According to receiver-operating characteristic curves, diagnostic accuracy of metabolite measurements in overnight/first-morning urine samples did not differ from measurements in 24-h urine collections, but was lower for both than for plasma. Using optimized reference intervals, diagnostic specificity was higher for overnight than daytime collections at similar sensitivities. CONCLUSIONS: Measurements of urinary-free catecholamine metabolites in first-morning/overnight urine collections offer an alternative for diagnosis of PPGL to 24-h collections but remain less accurate than plasma measurements.

Pheochromocytoma and paraganglioma: clinical feature-based disease probability in relation to catecholamine biochemistry and reason for disease suspicion.

OBJECTIVE: Hypertension and symptoms of catecholamine excess are features of pheochromocytomas and paragangliomas (PPGLs). This prospective observational cohort study assessed whether differences in presenting features in patients tested for PPGLs might assist establishing likelihood of disease. DESIGN AND METHODS: Patients were tested for PPGLs because of signs and symptoms, an incidental mass on imaging or routine surveillance due to previous history or hereditary risk. Patients with (n = 245) compared to without (n = 1820) PPGLs were identified on follow-up. Differences in presenting features were then examined to assess the probability of disease and relationships to catecholamine excess. RESULTS: Hyperhidrosis, palpitations, pallor, tremor and nausea were 30-90% more prevalent (P < 0.001) among patients with than without PPGLs, whereas headache, flushing and other symptoms showed little or no differences. Although heart rates were higher (P < 0.0001) in patients with than without PPGLs, blood pressures were not higher and were positively correlated to BMI, which was lower (P < 0.0001) in patients with than without PPGLs. From these differences in clinical features, a score system was established that indicated a 5.8-fold higher probability of PPGLs in patients with high than low scores. Higher scores among patients with PPGLs were associated, independently of tumor size, with higher biochemical indices of catecholamine excess. CONCLUSIONS: This study identifies a complex of five signs and symptoms combined with lower BMI and elevated heart rate as key features in patients with PPGLs. Prevalences of these features, which reflect variable tumoral catecholamine production, may be used to triage patients according to likelihood of disease.

A high rate of modestly elevated plasma normetanephrine in a population referred for suspected PPGL when measured in a seated position.

OBJECTIVE: Determine rate of high plasma normetanephrine or metanephrine (PNM-PMN) in a large sample of patients according to PNM-PMN posture and age-adjusted references. DESIGN: Retrospective re-analysis of PNM-PMN from a Canadian reference laboratory (n = 5452), 2011-2015; most were in seated position (n = 5112) rather than supine (n = 340). An international PPGL database demonstrated expected distribution of supine PNM-PMN in PPGL patients. METHODS: All PNM-PMN from a tertiary referral laboratory were reviewed. Any PNM-PMN result greater than 2× upper reference limit (URL) was considered likely true PPGL. Results 1-2× URL were uncertain, requiring additional testing/follow-up despite most being false positive given the rarity of PPGL. The rate of results in the 1-2× URL category were calculated for each group according to collection posture and differing published URL: seated, supine or supine age adjusted. RESULTS: When collected and interpreted by seated URL, 19.6% of PNM required additional testing; only 4.6% being >2× URL. For patients over age 50 years, the abnormal rate was 24.9%. When collected supine, interpreted by supine age-adjusted URL, only 5.3% of PNM were mildly elevated. Possible false positives may be even lower when considering PMN or plasma methoxytyramine which were commonly high in true PPGL despite mild PNM elevations. CONCLUSIONS: In a general medical population, seated PNM has a high rate of abnormal results, far exceeding expected prevalence. Supine measurement with supine, age-adjusted interpretation is strongly preferred prior to costly or invasive PPGL investigations. SUMMARY: Review of 5452 plasma normetanephrine measurements showed 20% to be high, likely false positives for most. Supine, age-adjusted measures were half as likely to be elevated.

Adrenomedullary function, obesity and permissive influences of catecholamines on body mass in patients with chromaffin cell tumours.

BACKGROUND: Obesity-associated activation of sympathetic nervous outflow is well documented, whereas involvement of dysregulated adrenomedullary hormonal function in obesity is less clear. This study assessed relationships of sympathoadrenal function with indices of obesity and influences of circulating catecholamines on body mass. METHODS: Anthropometric and clinical data along with plasma and 24-h urine samples were collected from 590 volunteers and 1368 patients tested for phaeochromocytoma and paraganglioma (PPGL), among whom tumours were diagnosed in 210 individuals. RESULTS: Among patients tested for PPGL, those with tumours less often had a body mass index (BMI) above 30 kg/m2 (12 vs. 31%) and more often a BMI under 25 kg/m2 (56 vs. 32%) than those without tumours (P < 0.0001). Urinary outputs of catecholamines in patients with PPGL were negatively related to BMI (r = -0.175, P = 0.0133). Post-operative weight gain (P < 0.0001) after resection of PPGL was positively related to presurgical tumoural catecholamine output (r = 0.257, P = 0.0101). Higher BMI in men and women and percent body fat in women of the volunteer group were associated with lower plasma concentrations and urinary outputs of adrenaline and metanephrine, the former indicating obesity-related reduced adrenaline secretion and the latter obesity-related reduced adrenomedullary adrenaline stores. Daytime activity was associated with substantial increases in urinary adrenaline and noradrenaline excretion, with blunted responses in obese subjects. CONCLUSIONS: The findings in patients with PPGL support an influence of high circulating catecholamines on body weight. Additional associations of adrenomedullary dysfunction with obesity raise the possibility of a permissive influence of the adrenal medulla on the regulation of body weight.