Graeme Eisenhofer | Technische Universität Dresden (original) (raw)

Papers by Graeme Eisenhofer

Genetics in medicine : official journal of the American College of Medical Genetics, Jan 27, 2018

Metabolic aberrations have been described in neoplasms with pathogenic variants (PV) in the Krebs... more Metabolic aberrations have been described in neoplasms with pathogenic variants (PV) in the Krebs cycle genes encoding succinate dehydrogenase (SDH), fumarate hydratase (FH) and isocitrate dehydrogenase (IDH). In turn, accumulation of oncometabolites succinate, fumarate, and 2-hydroxyglutarate can be employed to identify tumors with those PV . Additionally, such metabolic readouts may aid in genetic variant interpretation and improve diagnostics. Using liquid chromatography-mass spectrometry, 395 pheochromocytomas and paragangliomas (PPGLs) from 391 patients were screened for metabolites to indicate Krebs cycle aberrations. Multigene panel sequencing was applied to detect driver PV in cases with indicative metabolite profiles but undetermined genetic drivers. Aberrant Krebs cycle metabolomes identified rare cases of PPGLs with germline PV in FH and somatic PV in IDHx and SDHx, including the first case of a somatic IDH2 PV in PPGL. Metabolomics also reliably identified PPGLs with SDH...

Purpose: MDH2 (malate dehydrogenase 2) has recently been proposed as a novel potential pheochromo... more Purpose: MDH2 (malate dehydrogenase 2) has recently been proposed as a novel potential pheochromocytoma/paraganglioma (PPGL) susceptibility gene, but its role in the disease has not been addressed. This study aimed to determine the prevalence of MDH2 pathogenic variants among PPGL patients and determine the associated phenotype. Methods: Eight hundred thirty patients with PPGLs, negative for the main PPGL driver genes, were included in the study. Interpretation of variants of unknown significance (VUS) was performed using an algorithm based on 20 computational predictions, by implementing cell-based enzymatic and immuno-fluorescence assays, and/or by using a molecular dynamics simulation approach.

The Journal of molecular diagnostics : JMD, Jul 25, 2017

Genetic diagnosis is recommended for all pheochromocytoma and paraganglioma (PPGL) cases, as driv... more Genetic diagnosis is recommended for all pheochromocytoma and paraganglioma (PPGL) cases, as driver mutations are identified in approximately 80% of the cases. As the list of related genes expands, genetic diagnosis becomes more time-consuming, and targeted next-generation sequencing (NGS) has emerged as a cost-effective tool. This study aimed to optimize targeted NGS in PPGL genetic diagnostics. A workflow based on two customized targeted NGS assays was validated to study the 18 main PPGL genes in germline and frozen tumor DNA, with one of them specifically directed toward formalin-fixed, paraffin-embedded tissue. The series involved 453 unrelated PPGL patients, of whom 30 had known mutations and were used as controls. Partial screening using Sanger had been performed in 275 patients. NGS results were complemented with the study of gross deletions. NGS assay showed a sensitivity ≥99.4%, regardless of DNA source. We identified 45 variants of unknown significance and 89 pathogenic mu...

The Journal of Clinical Endocrinology & Metabolism, 2014

Context: Mutations of succinate dehydrogenase A/B/C/D genes (SDHx) increase susceptibility to dev... more Context: Mutations of succinate dehydrogenase A/B/C/D genes (SDHx) increase susceptibility to development of pheochromocytomas and paragangliomas (PPGLs), with particularly high rates of malignancy associated with SDHB mutations. Objective: We assessed whether altered succinate dehydrogenase product-precursor relationships, manifested by differences in tumor ratios of succinate to fumarate or other metabolites, might aid in identifying and stratifying patients with SDHx mutations. Design, Setting and Patients: PPGL tumor specimens from 233 patients, including 45 with SDHx mutations, were provided from eight tertiary referral centers for mass spectrometric analyses of Krebs cycle metabolites. Main outcome measure: Diagnostic performance of the succinate:fumarate ratio for identification of pathogenic SDHx mutations. Results: SDH-deficient PPGLs were characterized by 25-fold higher succinate and 80% lower fumarate, cis-aconitate and isocitrate tissue levels than PPGLs without SDHx mutations. Receiveroperating characteristic curves for use of ratios of succinate to fumarate or to cis-aconitate and isocitrate to identify SDHx mutations indicated areas under curves of 0.94 to 0.96; an optimal cutoff of 97.7 for the succinate:fumarate ratio provided a diagnostic sensitivity of 93% at a specificity of 97% to identify SDHX-mutated PPGLs.

International Journal of Cancer, 2014

Pheochromocytomas and paragangliomas (PPGLs) are catecholamine-producing chromaffin cell tumors w... more Pheochromocytomas and paragangliomas (PPGLs) are catecholamine-producing chromaffin cell tumors with diverse phenotypic features reflecting mutations in numerous genes, including MYC-associated factor X (MAX). To explore whether phenotypic differences among PPGLs reflect a MAX-mediated mechanism and opposing influences of hypoxia-inducible factor (HIF)s HIF2α and HIF1α, we combined observational investigations in PPGLs and gene-manipulation studies in two pheochromocytoma cell lines. Among PPGLs from 140 patients, tumors due to MAX mutations were characterized by gene expression profiles and intermediate phenotypic features that distinguished these tumors from other PPGLs, all of which fell into two expression clusters: one cluster with low expression of HIF2α and mature phenotypic features and the other with high expression of HIF2α and immature phenotypic features due to mutations stabilizing HIFs. Max-mutated tumors distributed to a distinct subcluster of the former group. In cell lines lacking Max, re-expression of the gene resulted in maturation of phenotypic features and decreased cell cycle progression. In cell lines lacking Hif2α, overexpression of the gene led to immature phenotypic features, failure of dexamethasone to induce differentiation and increased proliferation. HIF1α had opposing actions to HIF2α in both cell lines, supporting evolving evidence of their differential actions on tumorigenic processes via a MYC/MAX-related pathway. Requirement of a fully functional MYC/MAX complex to facilitate differentiation explains the intermediate phenotypic features in tumors due to MAX mutations. Overexpression of HIF2α in chromaffin cell tumors due to mutations affecting HIF stabilization explains their proliferative features and why the tumors fail to differentiate even when exposed locally to adrenal steroids.

Clinical cancer research : an official journal of the American Association for Cancer Research, Jan 30, 2015

Pheochromocytoma and paraganglioma (PPGL) are rare neuroendocrine tumors, associated with highly ... more Pheochromocytoma and paraganglioma (PPGL) are rare neuroendocrine tumors, associated with highly variable post-operative evolution. The scarcity of reliable PPGL prognostic markers continues to complicate patient management. In this study, we explored genome-wide DNA methylation patterns in the context of PPGL malignancy to identify novel prognostic markers. We retrospectively investigated DNA methylation patterns in PPGL with and without metastases utilizing high-throughput DNA methylation profiling data (Illumina 27K) from two large, well-characterized discovery (n=123; 24 metastatic) and primary validation (n=154; 24 metastatic) series. Additional validation of candidate CpGs was performed by bisulfite pyrosequencing in a second independent set of 33 paraffin-embedded PPGLs (19 metastatic). Of the initial 86 candidate CpGs, we successfully replicated fifty-two (47 genes), associated with metastatic PPGL. Of these, 48 CpGs showed significant associations with time to progression e...

The Journal of steroid biochemistry and molecular biology, 2015

Steroid profiling for diagnosis of endocrine disorders featuring disordered production of steroid... more Steroid profiling for diagnosis of endocrine disorders featuring disordered production of steroid hormones is now possible from advances in liquid chromatography with tandem mass spectrometry (LC-MS/MS). Adrenal venous (AV) measurements of aldosterone and cortisol are a standard practice in the clinical work-up of primary aldosteronism, but do not yet take advantage of steroid profiling. A novel LC-MS/MS based method was developed for simultaneous measurement of 15 adrenal steroids: aldosterone, corticosterone, 11-deoxycorticosterone, progesterone, pregnenolone, cortisone, cortisol, 11-deoxycortisol, 17-hydroxyprogesterone, androstenedione, dehydroepiandrosterone, dehydroepiandrosterone-sulfate, 21-deoxycortisol, 18-oxocortisol and 18-hydroxycortisol. These were compared in peripheral venous (pV) and AV plasma from 70 patients undergoing AV sampling with and without cosyntropin stimulation. Aldosterone and cortisol levels measured by LC-MS/MS were compared with those measured by imm...

Nature reviews. Endocrinology, 2014

A promising grading system to predict metastasis in patients with phaeochromocytoma and paragangl... more A promising grading system to predict metastasis in patients with phaeochromocytoma and paraganglioma assigns risk according to selected histological and other criteria. Such risk stratification might be useful for personalized management and screening programmes, as it could limit the costs of follow-up and reduce the risk of disseminated disease.

Journal of the autonomic nervous system, Jan 13, 1997

After norepinephrine (NE) is deaminated by monoamine oxidase (MAO), the aldehyde formed is either... more After norepinephrine (NE) is deaminated by monoamine oxidase (MAO), the aldehyde formed is either metabolized to 3,4-dihydroxy-mandelic acid (DHMA) by aldehyde dehydrogenase or is converted to 3,4-dihydroxyphenylglycol (DHPG) by aldehyde or aldose reductase. The present study examined the effects of inhibition of aldehyde and aldose reductase on production of DHPG and DHMA in rats. Mean (+/- S.E.) baseline plasma concentrations of DHPG (4.73 +/- 0.21 pmol/ml) were 60-fold higher than those of DHMA (0.08 +/- 0.01 pmol/ml). Inhibition of aldose and aldehyde reductase reduced plasma DHPG concentrations to 1.88 +/- 0.14 pmol/ml and increased plasma DHMA to 4.43 +/- 0.29 pmol/ml; additional inhibition of MAO reduced plasma DHPG to 0.16 +/- 0.06 pmol/ml and DHMA to 0.19 +/- 0.02 pmol/ml. Inhibition of aldehyde and aldose reductase also increased brain tissue levels of DHMA from 8 +/- 2 to 384 +/- 47 pmol/g and decreased levels of DHPG from 70 +/- 9 to 44 +/- 5 pmol/g. The results show tha...

Journal of Heart and Lung Transplantation, 2001

dilol in patients with end stage CHF accepted for transplantation (TX). Methods: The study was de... more dilol in patients with end stage CHF accepted for transplantation (TX). Methods: The study was designed as a double-blind, randomized, placebo-controlled trial and conducted in 3 German transplant centers. Study duration was 12 months. Patients accepted for TX with advanced left ventricular (LV) systolic dysfunction categorized as NYHA IV at least once since onset of CHF were eligible. The primary endpoint was the absolute change from baseline to latest available LV ejection fraction (EF) measurement determined by radionuclide ventriculography between Carvedilol and placebo. Results: The trial prospectively randomized 118 patients with CHF of ischemic (nϭ44) or non-ischemic etiology, a mean ϮSD [median] age of 53.3Ϯ9.8 [55.5] years, and a mean LVEF at baseline of 19.9Ϯ6.6 [20.1]%. Mean ϮSD [median] absolute change of LVEF from baseline was ϩ6.0Ϯ9.3 [ϩ3.9]% in the Carvedilol group versus ϩ0.7Ϯ7.1 [0.0]% in the placebo treated patients (pϽ0.008 [Ͻ0.006, Wilcoxon, 2-sided]) in the intention-to-treat population (nϭ74) and ϩ8.2Ϯ10.3 [ϩ6.0]% in Carvedilol versus ϩ1.4Ϯ7.9 [0.0]% on placebo (pϽ0.011 [Ͻ0.013, Wilcoxon,) in the per-protocol population (nϭ52). Serious adverse events were experienced by 33/60 patients (13 deaths) on placebo and 29/58 patients (9 deaths) on Carvedilol. Conclusion: Even in patients with endstage CHF accepted for TX pharmacological therapy may significantly improve cardiac function over a prolonged period of time. Carvedilol appears to be an appropriate and safe drug also in this severely ill patient population. Thus, the EFICAT trial extends the observations made in COPERNICUS.

Hypertension with renal artery stenosis is associated with both an activated renin-angiotensin sy... more Hypertension with renal artery stenosis is associated with both an activated renin-angiotensin system and elevated sympathetic activity. Therefore, in this condition it may be favorable to use a therapeutic modality that does not reflexly increase heart rate, renin secretion, and sympathetic nervous activity. The purpose of the present study was to assess overall, renal, and muscle sympathetic activity after short-term

Journal of Chromatography B: Biomedical Sciences and Applications, 1994

Several modifications of an HPLC-electrochemical assay method for plasma levels of norepinephrine... more Several modifications of an HPLC-electrochemical assay method for plasma levels of norepinephrine (NE), epinephrine (EPI), dopamine (DA), dihydroxyphenylglycol (DHPG), dihydroxyphenylalanine (DOPA) and dihydroxyphenylacetic acid (DOPAC) that improve the accuracy and reliability of DHPG, DOPA, and DOPAC measurements are described. In batch alumina extractions, increasing the amount of alumina decreased analytical recoveries of DHPG, DOPA, and especially DOPAC, and increasing the strength of the eluting acid increased recoveries of these catechols, without affecting recoveries of the amines NE, EPI and DA. Refrigeration (4°C) until injection stabilized DOPAC in aqueous solution and therefore improved the reproducibility of plasma DOPAC measurements.

Journal of the National Cancer Institute, 2015

Disruption of the Krebs cycle is a hallmark of cancer. IDH1 and IDH2 mutations are found in many ... more Disruption of the Krebs cycle is a hallmark of cancer. IDH1 and IDH2 mutations are found in many neoplasms, and germline alterations in SDH genes and FH predispose to pheochromocytoma/paraganglioma and other cancers. We describe a paraganglioma family carrying a germline mutation in MDH2, which encodes a Krebs cycle enzyme. Whole-exome sequencing was applied to tumor DNA obtained from a man age 55 years diagnosed with multiple malignant paragangliomas. Data were analyzed with the two-sided Student's t and Mann-Whitney U tests with Bonferroni correction for multiple comparisons. Between six- and 14-fold lower levels of MDH2 expression were observed in MDH2-mutated tumors compared with control patients. Knockdown (KD) of MDH2 in HeLa cells by shRNA triggered the accumulation of both malate (mean ± SD: wild-type [WT] = 1±0.18; KD = 2.24±0.17, P = .043) and fumarate (WT = 1±0.06; KD = 2.6±0.25, P = .033), which was reversed by transient introduction of WT MDH2 cDNA. Segregation of t...

1. Urinary excretion of dopamine (DA) increases during dietary salt loading. The majority of urin... more 1. Urinary excretion of dopamine (DA) increases during dietary salt loading. The majority of urinary DA is derived from circulating dihydroxyphenylalanine (dopa). Whether the increase in urinary DA excretion during salt loading results from increased efficiency of uptake of dopa by proximal tubular cells of the kidney, facilitation of intracellular conversion of dopa to DA, or increased delivery of dopa to tubular uptake sites, has been unknown. 2. In 10 inpatient normal volunteers on a constant diet, daily excretion of dopa and DA was assessed during normal sodium intake (109 mmol/day) for 1 week, low sodium intake (9 mmol/day) for 1 week and high sodium intake (249 mmol/day) for 1 week. 3. Urinary DA excretion exceeded urinary dopa excretion by about tenfold, and the excretion of both DA and dopa increased by about twofold between the low and high salt diets, with similar proportionate changes. Plasma dopa was unchanged by dietary salt manipulation. 4. The results indicate that increases in urinary DA excretion during dietary salt loading can be accounted for by increased delivery of dopa to sites of uptake by proximal tubular cells. Since dopa is released into the bloodstream by sympathetic nerve endings and by the brain, and since interference with decarboxylation of dopa attenuates natriuretic responses, dopa may function indirectly as a neurohormone involved in homoeostatic regulation of sodium balance.

1. The source and physiological significance of dopamine (DA) sulphate, which exists in plasma at... more 1. The source and physiological significance of dopamine (DA) sulphate, which exists in plasma at much higher concentrations than free DA, have long been a puzzle. The present article reviews how the convergence of modern molecular and traditional clinical approaches is shedding new light on the origins and meaning of DA sulphate. 2. The sulphotransferase isoenzyme responsible for production of DA sulphate in humans (SULT1A3) has been cloned and shown to be expressed in large quantities in the gastro-intestinal tract, but not in liver. No orthologue of SULT1A3 has yet been identified in other species, consistent with the greater importance of sulphate conjugation of DA in humans than in most animals. 3. Diet has a major impact on plasma DA sulphate, with dramatic increases after ingestion of meals and foods rich in biogenic amines; however, substantial amounts of DA sulphate remaining after prolonged fasting indicate the presence of a mainly endogenous source. The lack of influence of acute or chronic changes in sympathetic outflow or of sympathoneural degeneration on plasma DA sulphate indicates that DA sulphate does not derive from sympathetic nerve. Relatively low rates of production from intravenously infused DA indicate that very little DA sulphate (< 2%) derives from metabolism of circulating DA, such as in red cells or platelets. 4. Consistent increments in DA sulphate from arterial to the outflowing venous plasma draining mesenteric organs, without increments across other organs or tissues (e.g., heart, lungs, liver), indicate that the gastrointestinal tract is a major source of more than 75% of DA sulphate produced in the body. The gastro-intestinal tract is also the site of a novel DA autocrine/paracrine system that produces nearly 50% of the DA in the body. Therefore, production of DA sulphate appears to reflect an enzymatic 'gut-blood' barrier for detoxifying dietary biogenic amines and delimiting autocrine/paracrine effects of endogenous DA generated in a novel 'third catecholamine system'.

1. Muscle sympathetic nerve activity (MSA) was recorded in the peroneal nerve at the knee by micr... more 1. Muscle sympathetic nerve activity (MSA) was recorded in the peroneal nerve at the knee by microneurography in ten healthy subjects and determinations were made simultaneously of intra-arterial blood pressure, and whole-body and cardiac noradrenaline spillover to plasma. Measurements were made at rest, during isometric handgrip at 30% of maximum power and during stress induced by forced mental arithmetic. 2. At rest there were significant positive correlations between spontaneous MSA (expressed as number of sympathetic bursts min-1) and both spillover of noradrenaline from the heart and concentration of noradrenaline in coronary sinus venous plasma. 3. Both isometric handgrip and mental arithmetic led to sustained increases of blood pressure, heart rate and MSA. Plasma concentrations of noradrenaline and spillover of noradrenaline (total body and cardiac) increased. In general the effects were more pronounced during handgrip than during stress. 4. When comparing effects during handgrip and stress the ratio between the fractional increases of MSA and cardiac noradrenaline spillover were significantly greater during handgrip. 5. The data suggest (a) that there are proportional interindividual differences in the strength of resting sympathetic activity to heart and skeletal muscle which are determined by a common mechanism and (b) that handgrip and mental stress are associated with differences in balance between sympathetic outflows to heart and skeletal muscle.

Modern pathology : an official journal of the United States and Canadian Academy of Pathology, Inc, Jan 27, 2015

Despite the established role of SDHB/SDHA immunohistochemistry as a valuable tool to identify pat... more Despite the established role of SDHB/SDHA immunohistochemistry as a valuable tool to identify patients at risk for familial succinate dehydrogenase-related pheochromocytoma/paraganglioma syndromes, the reproducibility of the assessment methods has not as yet been determined. The aim of this study was to investigate interobserver variability among seven expert endocrine pathologists using a web-based virtual microscopy approach in a large multicenter pheochromocytoma/paraganglioma cohort (n=351): (1) 73 SDH mutated, (2) 105 non-SDH mutated, (3) 128 samples without identified SDH-x mutations, and (4) 45 with incomplete SDH molecular genetic analysis. Substantial agreement among all the reviewers was observed either with a two-tiered classification (SDHB κ=0.7338; SDHA κ=0.6707) or a three-tiered classification approach (SDHB κ=0.6543; SDHA κ=0.7516). Consensus was achieved in 315 cases (89.74%) for SDHB immunohistochemistry and in 348 cases (99.15%) for SDHA immunohistochemistry. Amon...

Journal of Clinical Endocrinology & Metabolism, 2006

Adrenal and extraadrenal paragangliomas are tumors of chromaffin cells that are usually benign bu... more Adrenal and extraadrenal paragangliomas are tumors of chromaffin cells that are usually benign but that may also develop into malignant disease. Mutations of the gene for succinate dehydrogenase subunit B (SDHB) are associated with a high risk of malignancy, but establishing the precise contribution requires relatively large numbers of patients with well-defined malignancy. We assessed the prevalence of SDHB mutations in a series of patients with malignant paraganglioma. SDHB mutation testing was carried out in 44 consecutive patients with malignant paraganglioma. Clinical characteristics of patients with malignant disease due to SDHB mutations were compared with those without mutations. Pathogenic SDHB mutations were found in 13 of the 44 patients (30%). Close to one third of patients had metastases originating from an adrenal primary tumor, compared with a little over two thirds from an extraadrenal tumor. Among the latter patients, the frequency of SDHB mutations was 48%. This study establishes that missense, nonsense, frameshift, and splice site mutations of the SDHB gene are associated with about half of all malignancies originating from extraadrenal paragangliomas. The high frequency of SDHB germline mutations among patients with malignant disease, particularly when originating from an extraadrenal paraganglioma, may justify a high priority for SDHB germline mutation testing in these patients.