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Clinical Neurocardiology Section, National Institute of Neurological Disorders and Stroke (G.E., D.S.G.); the Urologic Oncology Branch, National Cancer Institute (M.M.W.); the Hypertension Endocrine Branch, National Heart Lung and Blood Institute (H.R.K.); and the Pediatric and Reproductive Endocrinology Branch (K.P.), National Institute of Child Health and Human Development, National Institutes of Health, Bethesda, Maryland 20892; the Department of Clinical Physiology (P.F.), Sahlgren’s University Hospital, S-41345 Gothenburg, Sweden; and the Department of General Internal Medicine (J.W.M.L.), St. Radboud University Hospital, 6525 GA Nijmegen, The Netherlands
Address all correspondence and requests for reprints to: Graeme Eisenhofer, Building 10, Room 6N252, NIH, 10 Center Drive, Bethesda, Maryland 20892-1620. E-mail: ge{at}box-g.nih.gov.
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Abstract |
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Introduction |
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A remaining problem, common to all biochemical tests used for diagnosis of pheochromocytoma, is that a high value for normetanephrine or metanephrine does not necessarily prove the presence of a tumor. False-positive results must be expected when the normal range for a test is set at anything less than the 100% confidence intervals of a reference population. False-positive results usually, however, occur more frequently than expected, probably due to differences in clinical characteristics of reference and patient populations and reduced control over sources of interference and sampling conditions.
In one study, the specificity of plasma free metanephrines, although higher than for plasma or urinary catecholamines or urinary fractionated metanephrines, was as low as 82% in patients with signs and symptoms suggesting pheochromocytoma (4). Another study from an independent center confirmed the high diagnostic sensitivity of plasma-free metanephrines, but noted a specificity of 85% that was less than that for urinary catecholamines and total metanephrines (5). At the usual low prevalence of pheochromocytoma among patients tested for the tumor, such values for specificity mean that false-positive results will greatly exceed true-positive results. This high frequency of false-positive results poses a diagnostic dilemma for clinicians, often requiring extensive and costly follow-up tests, and in many cases instigating unnecessary attempts to localize a suspected tumor. Before imaging studies are undertaken, it is preferable to conclusively establish the presence of a tumor by biochemical testing.
The present study examined medication-associated causes of false-positive results and the use of supplementary biochemical tests to further exclude or confirm pheochromocytoma in patients where initial tests yielded positive, but equivocal results. In particular, the study examined the utility of measurements of plasma normetanephrine responses to clonidine to distinguish positive results due to sympathetic activation from those due to a pheochromocytoma.
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Subjects and Methods |
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Patients were tested for pheochromocytoma at the NIH (Bethesda, MD), at St. Radboud University Medical Center (Nijmegen, The Netherlands), or at Sahlgren’s University Hospital (Gothenburg, Sweden). Testing was based solely on the presence of signs and symptoms suggestive of pheochromocytoma (e.g. therapy-resistant or paroxysmal hypertension and sweating, headache or palpitations) in 401 patients or on the incidental finding of an adrenal mass in 26 patients. In another 409 patients, testing was carried out as part of routine screening for hereditary pheochromocytoma or because of clinical suspicion of a tumor in patients with a family history of pheochromocytoma. Similarly, testing was carried out in a further 20 patients at high risk for pheochromocytoma because of a previous history of sporadic pheochromocytoma.
Pheochromocytoma was confirmed in 208 patients by pathological examination of surgically resected or biopsied tumor tissue or a diagnosis of inoperable metastatic disease by imaging studies. Pheochromocytoma was excluded in 648 patients by one or more of the following criteria: negative abdominal imaging studies, pathological examination of a surgically resected adrenal mass, normal biochemical results, or lack of the tumor on patient follow-up 2 yr or more after initial testing. Studies were approved by the appropriate Institutional Review Boards and all patients gave informed consent.
Biochemical tests
Samples of blood were drawn into 10-ml heparinized tubes via a forearm iv cannula with patients supine for at least 20 min before sampling. Patients were instructed to fast and abstain from caffeinated and decaffeinated beverages overnight before blood sampling. Plasma was analyzed for concentrations of catecholamines and free metanephrines using HPLC with electrochemical detection (6, 7). Twenty-four-hour urine collections were also obtained in most patients and analyzed for catecholamines or fractionated metanephrines by HPLC with electrochemical detection (8). Upper reference limits for biochemical tests were as established previously (2, 4).
Medication-associated false-positive results
All patients tested for pheochromocytoma were instructed to avoid taking acetaminophen for 5 d before blood sampling because of known analytical interference of the drug with assays of plasma free metanephrines (7). Patients did not discontinue other medications but instead were instructed to provide a listing of all medications taken at the time of initial testing. Such listings were obtained from 510 patients in whom the tumor was subsequently excluded according to the criteria outlined above.
Clonidine-suppression testing
Clonidine-suppression tests were carried out in 97 patients. Pheochromocytoma was confirmed pathologically in 48 patients, including 20 with von Hippel-Lindau syndrome and 4 with multiple endocrine neoplasia type 2. These patients were aged 34 ± 16 yr (mean ± SD) and included 24 females and 24 males. The other 49 patients without the tumor were aged 51 ± 14 yr and included 27 females and 22 males. Five had von Hippel-Lindau syndrome, and 44 underwent testing because of signs and symptoms suggesting pheochromocytoma.
Several medications, including diuretics, tricyclic antidepressants, and ß blockers, have been described to interfere with norepinephrine responses to the clonidine suppression test (9, 10, 11). Additionally, profound hypotensive responses to clonidine can be troublesome in some patients taking certain antihypertensive medications. When indicated, such medications were withdrawn for a period of 1–5 d, depending on the plasma half-life of the particular drug.
All clonidine suppression tests were carried out after an overnight fast with patients supine. A baseline blood sample was drawn into a 10-ml heparinized tube through a forearm venous cannula after at least 20 min of supine rest. Clonidine was then given orally at a dose of 0.3 mg (for a 60–80 kg subject), adjusted for body weight as necessary. A second blood sample was obtained 3 h after administration of clonidine with patients remaining supine.
There are several criteria for what constitute positive or negative results for the clonidine-suppression test. One criterion for a normal response to clonidine is a fall in plasma norepinephrine to within the normal range (12, 13, 14, 15). Another is the magnitude of the fall of plasma norepinephrine, with a fall to less than 50% of baseline values considered normal (14, 15, 16). Used alone and in combination both criteria have limitations (10, 14, 16, 17, 18, 19). In the present study, we examined a combination of the above criteria, and extended the analyses to use of plasma normetanephrine responses to clonidine.
Statistical analysis
Data are presented as means ± SD. Two-sided Student’s t tests, 
2 tests, McNemar’s test, and ANOVA with Scheffé’s test were used to compare results among groups.
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Results |
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Patients with pheochromocytoma showed more consistent elevations of plasma free metanephrines than of catecholamines (Fig. 1
). Forty-seven out of 208 (23%) patients with pheochromocytoma had normal plasma concentrations of norepinephrine (<498 ng/liter; <2.95 nmol/liter), compared with only 8 (4%) with normal levels of normetanephrine (<112 ng/liter; <0.61 nmol/liter). Similarly, 142 patients (68%) had normal plasma concentrations of epinephrine (<83 ng/liter; <0.45 nmol/liter), compared with 98 (47%) with normal levels of metanephrine (<61 ng/liter; <0.31 nmol/liter). Among the 47 patients with normal plasma levels of norepinephrine, epinephrine levels were also normal in 36 patients.
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Plasma concentrations of catecholamines and free metanephrines in patients with pheochromocytoma were higher, but showed some overlap with concentrations in patients in whom the tumor was excluded (Fig. 1
). The extent of overlap was, however, smaller for plasma free metanephrines than for catecholamines. Only 32% of patients with pheochromocytoma had plasma concentrations of normetanephrine below the highest false-positive level (400 ng/liter; 2.19 nmol/liter) observed in patients without the tumor, compared with 67% for norepinephrine. Similarly, 62% of patients with pheochromocytoma had plasma concentrations of metanephrine below the highest false-positive level (236 ng/liter; 1.20 nmol/liter), compared with 87% for epinephrine.
In contrast to the higher frequency of true-positive results for plasma metanephrines than catecholamines, there was a lower frequency of false-positive results for metanephrines than catecholamines (Fig. 1). There were 104 patients (16%) with false-positive elevations of plasma norepinephrine compared with 55 (8%) with false-positive elevations of normetanephrine, these including 38 (6%) with false-positive elevations of both amines. Other false-positive elevations were observed in 36 patients for plasma epinephrine and 31 for plasma metanephrine, including 16 with false-positive elevations of both amines.
Patients with pheochromocytoma who had elevations of plasma metanephrines or catecholamines below the highest respective levels in patients without pheochromocytoma showed different patterns of biochemical results than those observed in patients with false-positive results (Table 1). In contrast to similar plasma levels of catecholamines, concentrations of metanephrines in patients with true-positive results were 7.5- to 9.1-fold higher (P < 0.001) than in patients with false-positive results. Thus, ratios of plasma normetanephrine to norepinephrine or of metanephrine to epinephrine were 6.8- to 7.7-fold higher (P < 0.001) in patients with true-positive compared with false-positive results.
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). In contrast, normetanephrine to norepinephrine ratios were greater than 0.52 in 20 of the 64 (31%) patients with pheochromocytoma who had elevations of normetanephrine in the false-positive range (112–400 ng/liter; 0.61–2.19 nmol/liter); and metanephrine to epinephrine ratios were greater than 4.2 in 6 of 39 (15%) patients with pheochromocytoma who had elevations of metanephrine levels in the false-positive range (61–236 ng/liter; 0.31–1.20 nmol/liter).
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Phenoxybenzamine and tricyclic antidepressants were the medications most frequently associated with false-positive results, together accounting for 41–45% of all elevated plasma levels of normetanephrine and norepinephrine in patients without pheochromocytoma (Table 2). Patients taking these medications had 1.9- to 2.6-fold higher (P < 0.001) plasma concentrations or urinary outputs of normetanephrine and norepinephrine and 2.3- to 7.7-fold higher (P < 0.02) likelihoods of false-positive results compared with patients taking other drugs or no medications. These influences were restricted to norepinephrine and normetanephrine and not to epinephrine and metanephrine.
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). Also, compared with patients taking tricyclics, those taking selective serotonin reuptake blockers had lower plasma concentrations (P < 0.001) and lower rates of false-positive results for plasma norepinephrine (P < 0.001) and normetanephrine (P = 0.016).
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1-adrenoceptor blocking drugs such as doxazosin, terazosin, and prazosin were not associated with an increased frequency of false-positive results for plasma norepinephrine or normetanephrine. These medications were, however, associated with a 4-fold higher (P = 0.017) frequency of false-positive elevations of urinary norepinephrine (7% vs. 27%) but were without influence on other urinary analytes.
ß-Adrenoceptor blocking drugs, such as atenolol, metoprolol, propranolol, and including the combined - and ß-adrenoceptor blocker, labetolol, were not associated with an increased frequency of false-positive elevations of plasma normetanephrine, norepinephrine or epinephrine, as measured using the assays in this study. However, as a group these medications were associated with 60% (9/15) of all false-positive elevations of plasma metanephrine. Although the frequency of elevated plasma concentrations of metanephrine in patients taking ß-adrenoceptor blockers was not high, it was substantially higher (P < 0.001) than in patients not taking these drugs (12.5% vs. 1.6%).
ß-Adrenoceptor blockers were also associated with a 2.6-fold higher (P = 0.03) frequency of false-positive elevations of urinary norepinephrine, a 6.5-fold higher (P = 0.003) frequency of false-positive elevations of urinary epinephrine, a 2.1-fold higher (P < 0.001) frequency of false-positive elevations of urinary normetanephrine, and a 2.2-fold increase (P = 0.024) in false-positive elevations of urinary metanephrine. These influences did not appear to be associated with any specific type of ß-adrenoceptor blocker.
Calcium channel blockers were associated with a 3-fold higher (P = 0.003) frequency of false-positive elevations of plasma norepinephrine, but not plasma normetanephrine, epinephrine, or metanephrine. These drugs were also associated with a 3-fold higher (P = 0.01) frequency of false-positive results for urinary norepinephrine, a 5-fold higher (P = 0.02) frequency of false-positive elevations of urinary epinephrine but had no effects on frequencies of false-positive results for urinary normetanephrine and metanephrine. In contrast to the above antihypertensives, diuretics, angiotensin-converting enzyme inhibitors, and angiotensin-1 receptor blockers appeared to have little influence on frequencies of false-positive biochemical results for any of the analytes measured in either plasma or urine.
All of 7 patients taking sympathomimetics (e.g. pseudoephedrine) had false-positive elevations of urinary normetanephrine or metanephrine compared with only 2 of 10 patients with elevations of plasma normetanephrine or metanephrine. In three patients with highly elevated urinary outputs of metanephrine (4393, 2561, and 1064 µg/d), the cause was traced to buspirone, an anxiolytic agent known to interfere with HPLC analysis of urinary metanephrine (20). There were no effects of this drug on HPLC analysis of plasma metanephrine.
Clonidine-suppression testing
Clonidine decreased (P < 0.001) plasma norepinephrine and normetanephrine in all patients without pheochromocytoma (Fig. 4). Percent decreases in plasma norepinephrine after clonidine varied from 16–91%, and on average were larger (P < 0.001) than those of normetanephrine (65 ± 19% vs. 48 ± 15%), which varied from 4–77%. Plasma concentrations of epinephrine showed variable responses to clonidine, but on average were decreased(P < 0.001) by 33 ± 51% (Table 3). In contrast, plasma concentrations of metanephrine remained unchanged.
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). However, nine patients with pheochromocytoma had falls in norepinephrine of more than 50%, including two with parallel falls of normetanephrine (Figs. 4 and 5). Among these nine patients, two had baseline levels of norepinephrine that were moderately increased (1758 and 3397 ng/liter), 5 had slightly increased norepinephrine levels (575–761 ng/liter) and two had normal baseline levels (< 498 ng/liter). An additional seven patients with pheochromocytoma had normal plasma concentrations of norepinephrine after clonidine, with no or little change from normal baseline levels (n = 5), or a small decrease from slightly elevated baseline levels to normal levels after clonidine (n = 2). All had elevated plasma concentrations of normetanephrine that were unaffected by clonidine.
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Because clonidine-induced falls in plasma normetanephrine were less than those of norepinephrine, we defined a positive response of plasma normetanephrine to clonidine as a fall of less than 40% (compared with 50% for norepinephrine). These and other criteria of positive test results—values after clonidine that remained above the upper reference limits—were used to examine distributions of positive and negative test results as functions of the different defining criteria (Fig. 5). These distributions were then used to compare the diagnostic utility of plasma norepinephrine and normetanephrine as end-point markers of the clonidine-suppression test according to four different definitions of a positive result (Table 4).
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). The difference in sensitivity was particularly significant (96% for normetanephrine vs. 67% for norepinephrine, P < 0.001) when a positive result was defined as a lack of clonidine-induced suppression combined with a plasma concentration after clonidine remaining above the upper reference limit. This definition of a positive result also yielded high specificities (98–100%) for use of both normetanephrine and norepinephrine as end-point markers. With more relaxed definitions of a positive result, specificities decreased, test sensitivity for norepinephrine increased, but that for normetanephrine remained unchanged. Similar diagnostic sensitivities and specificities were obtained when patients with von Hippel Lindau syndrome or multiple endocrine neoplasia type 2 were excluded from analysis. For this restricted group of patients who were tested because of suspicion of sporadic pheochromocytoma, diagnostic sensitivities were 96% for responses of plasma normetanephrine and 67% for those of norepinephrine. Respective specificities were 100% and 98%.
Values for diagnostic sensitivity were, however, lower when data were restricted to patients with (n = 15) and without (n = 20) pheochromocytoma who had baseline values for plasma normetanephrine in the equivocal range (112–400 ng/liter; 0.61–2.19 nmol/liter). For these patients, the diagnostic sensitivities were 87% for responses of plasma normetanephrine and only 33% for those of norepinephrine. Respective specificities were 100% and 95%.
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Discussion |
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). More specifically, we identify medications that are commonly associated with false-positive results and show how differences in plasma free metanephrines and catecholamines can be used to distinguish false-positive from true-positive results. We further show that measuring plasma-free normetanephrine, in conjunction with the clonidine-suppression test, improves diagnosis of pheochromocytoma in difficult cases where preliminary testing yields equivocal results.
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Follow-up testing should depend primarily on findings of positive results of initial biochemical tests, which should include measurements of plasma-free metanephrines, or if unavailable, measurements of urinary fractionated metanephrines as the next best test. These recommendations are based on the high diagnostic sensitivity of measurements of plasma free or urinary fractionated metanephrines and findings of normal catecholamines and elevated metanephrines in about 20% of patients with pheochromocytoma, particularly those screened because of a hereditary syndrome or an adrenal incidentaloma (2, 3, 4, 5, 21). Such patients are usually normotensive and asymptomatic. However, as we describe here and others have reported elsewhere (22, 23, 24), there are also occasional sporadic cases of pheochromocytoma where plasma and urinary catecholamines may be normal, despite signs and symptoms of the tumor.
Normal plasma and urinary catecholamines therefore do not exclude pheochromocytoma, even when testing is done because of hypertension and symptoms suggestive of the tumor. In contrast, levels of normetanephrine, metanephrine, or both metabolites are increased in almost all patients with pheochromocytoma. Exceptions include patients with small or microscopic tumors (<1 cm) that produce only small amounts of catecholamines, who may be encountered during screening because of a hereditary predisposition or a previous history for pheochromocytoma (2, 4). In such patients, follow-up testing at a later date remains mandatory and likely will lead to a positive diagnosis as the disease progresses. Other more rare exceptions include patients with pheochromocytomas that do not produce norepinephrine or epinephrine (5). Normal plasma levels of normetanephrine and metanephrine otherwise exclude pheochromocytoma, so that no immediate further testing for the tumor should usually be necessary.
Although problems of false-negative results in the diagnosis of pheochromocytoma are virtually eliminated by measurements of plasma free metanephrines, false-positive results remain a commonly encountered and potentially time-consuming and expensive problem for follow-up. The nature of this problem is illustrated in this study by the substantial overlap of biochemical results in patients with and without pheochromocytoma. For all biochemical tests there is a gray area where positive results in patients with the tumor are difficult to distinguish from positive results in patients without the tumor.
As shown here, the gray area for measurements of plasma free metanephrines extends from the upper reference limits of normal to 400 ng/liter (2.19 nmol/liter) for normetanephrine and to 236 ng/liter (1.2 nmol/liter) for metanephrine. In another independent study, the highest false-positive result was about 467 ng/liter (2.55 nmol/liter) for normetanephrine and about 158 ng/liter (0.8 nmol/liter) for metanephrine (5). In our experience, close to 80% of patients with pheochromocytoma have plasma concentrations of normetanephrine, metanephrine or both above these equivocal ranges (4). In such patients, the probability of pheochromocytoma is so high that the immediate task is to locate the tumor. In the remaining patients with elevated levels that fall within the equivocal ranges, additional tests are required to confirm or exclude the tumor.
Medication-associated false-positive results
Before additional biochemical tests are implemented, some consideration should be given to possible causes of false-positive results, including accompanying medical conditions, medications, inappropriate sampling conditions, and diet. In our study, the latter two causes of false-positive results were minimized by adherence to an overnight fast before blood sampling, which was carried out with patients resting supine.
Biochemical testing for pheochromocytoma should ideally be carried out after discontinuation of medications known to elevate levels of catecholamines and their metabolites or interfere directly with biochemical analyses. Patient safety considerations often, however, make this impractical. It is also often difficult to identify which medications interfere with a given test, particularly as new tests and drugs are developed.
Among our patients, phenoxybenzamine and tricyclic antidepressants accounted for up to 45% of false-positive elevations of plasma or urinary norepinephrine and normetanephrine. Tricyclic antidepressants are established causes of false-positive results (25), whereas phenoxybenzamine has not until now been recognized as an important problem.
The high rates of false-positive results in patients taking tricyclic antidepressants are probably due to the primary actions of these agents to inhibit monoamine reuptake (26, 27). These drugs also profoundly suppress release of norepinephrine from sympathetic nerves so that plasma concentrations of norepinephrine are usually decreased and not increased after acute administration (26). This influence, however, lessens after chronic administration (27). The primary action of these drugs to block monoamine reuptake then predominates resulting in increased escape of norepinephrine from sympathetic nerve terminals into the bloodstream. These complex actions may be responsible for the blood pressure disturbances that can accompany use of tricyclic antidepressants (28), and which may contribute to suspicion of pheochromocytoma. As we show here, selective serotonin reuptake inhibitors are not a significant cause of false-positive results and may provide an alternative medication when biochemical testing for pheochromocytoma is necessary.
Phenoxybenzamine is a nonspecific -adrenoceptor blocker commonly used to treat patients with pheochromocytoma. Presumably, the drug elevates norepinephrine and normetanephrine by attenuating 2-adrenoceptor-mediated feedback inhibition of norepinephrine release, possibly combined with reflexive sympathetic activation. Our findings of high rates of false-positive results associated with phenoxybenzamine lead us to suggest that this drug be avoided until biochemical testing is complete. As advocated elsewhere (15, 29, 30), alternative medications for blood pressure control include calcium channel blockers and selective 1-adrenoceptor blockers such as doxazosin. As we show here, false-positive elevations of plasma metanephrines and catecholamines with selective 1-adrenoceptor blockers are not a problem and with calcium channel blockers appear restricted to norepinephrine, an effect most likely due to the reflexive sympathetic activation occurring with the short-acting agents (31).
Most other antihypertensive medications were not significant sources of false-positive elevations of plasma metanephrines or catecholamines. However, ß-adrenoceptor blockers were associated with 60% of all false-positive elevations of plasma metanephrine. Nevertheless, the false-positive rate was not high (12.5%) and does not appear to justify withdrawing these medications unless an equivocal result has been obtained and repeat testing is necessary.
Patterns of biochemical test results
After the potential confounding influence of medications or other causes of false-positive results have been eliminated, some consideration should be given to the choice of additional biochemical tests most appropriate for more firmly establishing or refuting the diagnosis of pheochromocytoma. When initial testing yields an elevated plasma metanephrine or normetanephrine, this may be corroborated by a similar pattern of results after additional measurements of urinary normetanephrine and metanephrine. Conversely, when initial testing yields a positive result for measurements of urinary normetanephrine and metanephrine, additional measurements of plasma free metanephrines are useful.
As we show here, patterns of increases in plasma-free metanephrines and catecholamines can provide additional information for confirming pheochromocytoma in patients where initial tests of plasma free metanephrines are positive but are insufficiently elevated for a firm diagnosis. More specifically, patients with pheochromocytoma usually have larger relative increases in metanephrines than catecholamines, whereas patients with false-positive results due to sympathoadrenal activation usually have larger increases in catecholamines than metanephrines.
The above differences are partly due to the substantial amounts of free metanephrines formed continuously within pheochromocytoma tumor cells and released into the circulation independently of variations in release of the parent catecholamines (32). Another factor is the large contribution of the adrenal medulla to normal circulating levels of metanephrine (91%) and normetanephrine (23%), a contribution that is again independent of catecholamine release (33). Thus, during sympathoadrenal activation, increases in plasma-free metanephrine are negligible and increases of normetanephrine are smaller than those of the respective parent amines (34).
The above explains why, contrary to usual considerations (35), a patient with an elevated plasma normetanephrine or metanephrine, but normal or slightly elevated norepinephrine or epinephrine is more likely to have a pheochromocytoma than a patient with a highly elevated norepinephrine or epinephrine and slightly elevated normetanephrine or metanephrine. As we show here, a plasma normetanephrine to norepinephrine ratio above 0.52 or a metanephrine to epinephrine ratio above 4.2 can provide confirmatory evidence of pheochromocytoma in up to 30% of patients where increases in plasma metanephrines are insufficient to conclusively prove the tumor. However, because some tumors secrete relatively large amounts of catecholamines compared with metanephrines, use of the reverse pattern to exclude pheochromocytoma is unreliable. Other tests, such as the clonidine-suppression test, remain essential.
Clonidine-suppression testing
The clonidine-suppression test was introduced by Bravo et al. (12) to address the problem of how to distinguish patients with pheochromocytoma from those with false-positive biochemical results after initial testing for the tumor. By activating 2-adrenoceptors in the brain and on sympathetic nerve endings, clonidine suppresses norepinephrine release by sympathetic nerves. Decreases in elevated plasma norepinephrine concentrations after clonidine therefore suggest sympathetic activation, whereas a lack of decrease suggests pheochromocytoma.
The normal suppression of plasma norepinephrine after clonidine in many of our patients with pheochromocytoma, but particularly those with mildly elevated or normal baseline levels, agrees with several other reports documenting similar limitations of the clonidine-suppression test (13, 14, 36, 37, 38). Presumably, normal suppression occurs because much of the norepinephrine is derived from sympathetic nerves and remains responsive to clonidine. The clonidine-suppression test is therefore recommended for patients with plasma catecholamine levels over 1000 ng/liter (5.9 nmol/liter), with a normal response defined as a fall to within the normal range (39).
The above recommendation makes it problematic to use the clonidine-suppression test in patients with normal or only mildly elevated plasma norepinephrine levels. This is particularly troublesome because such patients represent those in whom it is most difficult to conclusively diagnose pheochromocytoma (18, 36). To overcome this limitation, another criterion for a normal response has been a fall in plasma norepinephrine after clonidine of more than 50% (14, 16). Although this allows identification of additional patients with pheochromocytoma who have normal or mildly increased norepinephrine levels, the trade-off, as shown here and elsewhere (10, 14, 16, 19), is increased numbers of false-positive results and reduced diagnostic specificity. Also, the gain in diagnostic sensitivity is offset by false-negative results in patients with episodically secreting pheochromocytomas, who can show apparent suppression due to sampling after clonidine on a downward swing in norepinephrine release by a tumor.
The above limitations led us to consider whether the clonidine-suppression test might be improved by measurements of plasma normetanephrine. Because pheochromocytomas cause larger, more consistent, and less episodic increases of plasma normetanephrine than of norepinephrine (3, 32, 40), we hypothesized that the metabolite would provide a better end-point marker for the clonidine-suppression test than the parent amine, but that this would depend on the normal responsiveness of the metabolite to clonidine. Because over 90% of circulating metanephrine is normally derived from metabolism of epinephrine within adrenal chromaffin cells, a process that is independent of epinephrine release (33, 34), we did not expect plasma levels of metanephrine to decrease after clonidine. However, because about 76% of plasma normetanephrine is derived from norepinephrine released by sympathetic nerves (33, 34), we did expect this metabolite to respond to clonidine in patients without pheochromocytoma.
Our expectations were confirmed by absent decreases in plasma metanephrine and consistent decreases in normetanephrine after clonidine in patients without pheochromocytoma. Thus, similar to the findings for norepinephrine, lack of decrease of plasma normetanephrine combined with a high plasma level after clonidine establishes a high probability of pheochromocytoma. More importantly, plasma normetanephrine concentrations did not fall and remained elevated after clonidine in 96% of patients with pheochromocytoma compared with only 67% for norepinephrine. Thus, normetanephrine responses to clonidine confirmed over 40% more pheochromocytomas and enabled more reliable exclusion of the tumor than did use of norepinephrine responses.
A remaining minor limitation is that responses of plasma metanephrine to clonidine cannot be used to distinguish true- from false-positive results for this metabolite. Clonidine-induced changes of epinephrine also offer limited help (17, 41). However, our experience shows that normetanephrine is the metabolite that is invariably increased and for which confirmation of pheochromocytoma is most important. Nevertheless, rather than replacing measurements of catecholamines, we recommend that measurements of plasma normetanephrine be included to optimize performance and extend the range of the test to patients where plasma norepinephrine is insufficiently elevated for reliable use.
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Conclusions |
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TopAbstractIntroductionSubjects and MethodsResultsDiscussionConclusionsReferences |
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). As described elsewhere (4), findings of plasma levels of normetanephrine less than 112 ng/liter (0.61 nmol/liter) and of metanephrine less than 61 ng/liter (0.31 nmol/liter) virtually excludes pheochromocytoma so that no immediate further testing for the tumor should be necessary. With plasma concentrations of normetanephrine above 400 ng/liter (2.19 nmol/liter) or of metanephrine above 236 ng/liter (1.20 nmol/liter), the probability of pheochromocytoma is so high that the immediate task is to locate the tumor. The key problem until now has been conclusive diagnosis in patients with plasma concentrations of normetanephrine or metanephrine between the above levels, where numbers of false-positive results can exceed those of true-positive results. The present study illustrates that consideration of medications, particularly phenoxybenzamine, tricyclic antidepressants, and ß-adrenoceptor blockers can be useful in identifying many patients with false-positive results. Additional appropriately selected biochemical tests and patterns of test results can provide further information for firmly establishing a diagnosis. Conclusive diagnosis of pheochromocytoma in most of the remaining patients can then be achieved by coupling the clonidine-suppression test with measurements of plasma normetanephrine, in addition to standard measurements of norepinephrine. This considerably extends the range of the test to patients in whom plasma norepinephrine is not sufficiently elevated for reliable diagnosis. The above strategy should minimize delay in diagnosis of pheochromocytoma and avoid expensive, time-consuming, and imperfect imaging studies in patients who do not have the tumor.
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Acknowledgments |
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Received December 30, 2002.
Accepted March 11, 2003.
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 J. T. Adler, G. Y. Meyer-Rochow, H. Chen, D. E. Benn, B. G. Robinson, R. S. Sippel, and S. B. Sidhu Pheochromocytoma: Current Approaches and Future Directions Oncologist, July 1, 2008; 13(7): 779 - 793. [Abstract] [Full Text] [PDF]
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 R. T Peaston and S. Ball Biochemical detection of phaeochromocytoma: why are we continuing to ignore the evidence? Ann Clin Biochem, January 1, 2008; 45(1): 6 - 10. [Abstract] [Full Text] [PDF]
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 A. Algeciras-Schimnich, C. M. Preissner, W. F. Young Jr., R. J. Singh, and S. K. G. Grebe Plasma Chromogranin A or Urine Fractionated Metanephrines Follow-Up Testing Improves the Diagnostic Accuracy of Plasma Fractionated Metanephrines for Pheochromocytoma J. Clin. Endocrinol. Metab., January 1, 2008; 93(1): 91 - 95. [Abstract] [Full Text] [PDF]
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 A. Karagiannis, D. P Mikhailidis, V. G Athyros, and F. Harsoulis Pheochromocytoma: an update on genetics and management Endocr. Relat. Cancer, December 1, 2007; 14(4): 935 - 956. [Abstract] [Full Text] [PDF]
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K. Pacak Preoperative Management of the Pheochromocytoma Patient J. Clin. Endocrinol. Metab., November 1, 2007; 92(11): 4069 - 4079. [Abstract] [Full Text] [PDF]
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 J. A. Lee, R. Zarnegar, W. T. Shen, E. Kebebew, O. H. Clark, and Q.-Y. Duh Adrenal Incidentaloma, Borderline Elevations of Urine or Plasma Metanephrine Levels, and the "Subclinical" Pheochromocytoma Arch Surg, September 1, 2007; 142(9): 870 - 874. [Abstract] [Full Text] [PDF]
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M. d'Herbomez, G. Forzy, C. Bauters, C. Tierny, P. Pigny, B. Carnaille, F. Pattou, J.-L. Wemeau, and N. Rouaix An analysis of the biochemical diagnosis of 66 pheochromocytomas Eur. J. Endocrinol., May 1, 2007; 156(5): 569 - 575. [Abstract] [Full Text] [PDF]
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 J. W.M. Lenders, J. J. Willemsen, G. Eisenhofer, H. A. Ross, K. Pacak, H. J.L.M. Timmers, and C.G.J. Sweep Is Supine Rest Necessary before Blood Sampling for Plasma Metanephrines? Clin. Chem., February 1, 2007; 53(2): 352 - 354. [Abstract] [Full Text] [PDF]
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 T. Zornitzki, H. Knobler, and A. Schattner Reboxetine treatment and pseudopheochromocytoma QJM, January 1, 2007; 100(1): 61 - 62. [Full Text] [PDF]
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K. L. Brain, J. Kay, and B. Shine Measurement of Urinary Metanephrines to Screen for Pheochromocytoma in an Unselected Hospital Referral Population Clin. Chem., November 1, 2006; 52(11): 2060 - 2064. [Abstract] [Full Text] [PDF]
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N. Unger, C. Pitt, I. L. Schmidt, M. K Walz, K. W Schmid, T. Philipp, K. Mann, and S. Petersenn Diagnostic value of various biochemical parameters for the diagnosis of pheochromocytoma in patients with adrenal mass. Eur. J. Endocrinol., March 1, 2006; 154(3): 409 - 417. [Abstract] [Full Text] [PDF]
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G. Eisenhofer, J. W.M. Lenders, D. S. Goldstein, M. Mannelli, G. Csako, M. M. Walther, F. M. Brouwers, and K. Pacak Pheochromocytoma Catecholamine Phenotypes and Prediction of Tumor Size and Location by Use of Plasma Free Metanephrines Clin. Chem., April 1, 2005; 51(4): 735 - 744. [Abstract] [Full Text] [PDF]
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 G. Eisenhofer, I. J. Kopin, and D. S. Goldstein Catecholamine Metabolism: A Contemporary View with Implications for Physiology and Medicine Pharmacol. Rev., September 1, 2004; 56(3): 331 - 349. [Abstract] [Full Text] [PDF]
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L. J. Hoy, M. Emery, J. A. Wedzicha, A. G. Davison, S. L. Chew, J. P. Monson, and K. A. Metcalfe Obstructive Sleep Apnea Presenting as Pseudopheochromocytoma: A Case Report J. Clin. Endocrinol. Metab., May 1, 2004; 89(5): 2033 - 2038. [Abstract] [Full Text] [PDF]
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 D. S. Goldstein, G. Eisenhofer, J. A. Flynn, G. Wand, and K. Pacak Diagnosis and Localization of Pheochromocytoma Hypertension, May 1, 2004; 43(5): 907 - 910. [Abstract] [Full Text] [PDF]
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I. Ilias and K. Pacak Current Approaches and Recommended Algorithm for the Diagnostic Localization of Pheochromocytoma J. Clin. Endocrinol. Metab., February 1, 2004; 89(2): 479 - 491. [Full Text] [PDF]
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W. M. Manger In Search of Pheochromocytomas J. Clin. Endocrinol. Metab., September 1, 2003; 88(9): 4080 - 4082. [Full Text] [PDF]
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) Patients with equivocal elevations of normetanephrine (112–400 ng/liter; 0.61–2.19 nmol/liter) or metanephrine (61–236 ng/liter; 0.31–1.20 nmol/liter) and normal plasma concentrations of catecholamines; (
) patients with equivocal elevations of norepinephrine (498–2388 ng/liter; 2.95–14.13 nmol/liter) or epinephrine (83–424 ng/liter; 0.45–2.32 nmol/liter) and normal plasma concentrations of metanephrine; (•) Patients with equivocal elevations of metanephrines and catecholamines. Dashed horizontal lines show the highest ratio in patients with false-positive results.
