Neuroendocrine Tumor Markers

Frontiers in Neuroendocrinology 22, 309 –339 (2001) doi:10.1006/frne.2001.0218, available online at http://www.idealibrary.com on

Neuroendocrine Tumor Markers Steven W. J. Lamberts, Leo J. Hofland, and Frank R. E. Nobels 1 Department of Medicine, Erasmus University, Rotterdam, The Netherlands

Tumor markers used in the diagnosis and follow-up of patients with neuroendocrine tumors are in most instances not specific for a given tumor and circulate under normal conditions in the serum, making their use as an early diagnostic tool difficult (low sensitivity). By combining hormone measurements with tissue responsiveness, demonstrations of inappropriate secretions of PTH, insulin, and gastrin during hypercalcemia, hypoglycemia, and hyperacidity, respectively, become highly sensitive and specific diagnostic tests. The application of polyclonal antibodies in RIAs of hormones, such as ACTH, insulin, and gastrin, increase the diagnostic level of hormone measurements in patients with neuroendocrine tumors. Other markers, such as chromogranin A, neuron-specific enolase, and ␣-subunit, as well as peptide receptor visualization, are of increasing importance in the diagnosis and follow-up of neuroendocrine and non-neuroendocrine tumors. KEY WORDS: neuroendocrine tumors; hormones; chromogranin A; neuron-specific enolase; ␣-subunit; somatostatin receptor scintigraphy; tumor markers. © 2001 Academic Press

INTRODUCTION

The perfect tumor marker is the one that is produced solely by a given type of tumor and is secreted in measurable amounts into the circulation. It should be detectable only in the presence of the tumor, and it should identify a malignant cell proliferation before it has spread beyond a localized site. Its quantitative amount in the blood should reflect the bulk of tumor, and changes in the serum concentration of the marker should reflect responses to treatment, as well as progressive disease (46, 91, 69). Unfortunately, no tumor markers have been recognized yet which fulfill all five requirements: sensitivity and specificity for screening and a prognostic indicator for size, growth, and spread. Tumor markers used in cancer diagnosis and treatment, such as ␣-fetoprotein, mucinous glycoproteins, such as CA 15-3, CA 19-9, and CA-125, and neuron-specific enolase (NSE) have in common that they are not actively secreted by cancer cells, but enter the circulation as a result of tumor lysis (69). The term “neuroendocrine” (NE) defines cells by their secretory products and some cytoplasmic proteins, rather than their localization or embryological Address correspondence and reprint requests to Steven Lamberts, Department of Medicine, University Hospital Dijkzigt, 40 Dr. Molewaterplein, 3015 GD Rotterdam, The Netherlands. Fax: 31-10-463.4937. E-mail: [email protected]. 1 Present address: OLV Ziekenhuis, Moorselbaan 164, 9300 Aalst, Belgium. 309

0091-3022/01 $35.00 Copyright © 2001 by Academic Press All rights of reproduction in any form reserved.

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derivation. The NE cell system includes all endocrine and neuronal cells, which share a common phenotype characterized by the simultaneous expression of certain marker proteins (general NE markers) and cell type-specific hormonal products. NE cells form small organs, distinct cell clusters within other tissues, or a network of cells dispersed in the thymus, thyroid, lung, and gut. Tumors originating from these cells reproduce many of the functional features of their cell(s) of origin and may give rise to characteristic syndromes (117, 40, 88, 12). Assessment of the anatomical spread and progression of disease in patients with NE tumors has become an essential part of disease management, but was until recently in most patients difficult to measure. Surgical resection of primary lesions, together with debulking of locoregional and/or limited liver metastases, is indicated in most patients. Also, new antitumor therapies (somatostatin analogs, interferon-␣) are considered more and more against aggressive chemotherapy and/or cytoreductive surgery for distant metastases in patients with advanced disease, especially because the survival and quality of life is often better than those in patients with solid cancers (64, 67, 68). Apart from the measurement of serum concentrations of NE tumor markers which are actively secreted by NE tumors, the in vivo visualization of peptide receptors on NE tumor cells also contributes to the clinical decision-making in patients with NE tumors, allowing an adequate assessment of the localization and extent of tumor burden, as well as its growth over time (67). In this review we aim to define the current thoughts on the optimal use of NE tumor markers in the diagnosis and treatment of patients with NE tumors.

GENERAL FEATURES OF NEUROENDOCRINE CELLS

NE cells have uniform nuclei and abundant granular or faintly stained (clear) cytoplasm. If they are gland-forming they present in solid clusters or show a trabecular pattern. If they are dispersed among other cells they are often difficult to recognize with the light microscope. Among the staining techniques which help to identify these cells the most important are the argentaffin and argyrophilic methods. At the ultrastructural level, they are characterized by the presence of membrane-bound dense-core secretory granules in the cytoplasm (40, 12, 116).

Peptides and Amines

The various cell types of the NE cell system can secrete specific hormonal products, such as peptides and biogenic amines. Peptide hormones are synthesized as precursors, which are cleaved in a sequence-specific and tissuespecific manner to yield the biologically active peptides. Before being secreted, however, these prohormones are processed via subsequent glycosylation, amidation, phosphorylation, and/or sulfatation (118). NE cells separate prohor-

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mones from other proteins by sorting and trafficking by a complex and highly efficient process. Sorting is usually over 98% efficient in normal NE cells, but is often defective in NE tumor cells. This inefficiency of tumor cells in secreting mature, biologically active peptide hormones can be used in the diagnosis and follow-up of patients with NE tumors (118). Well-known examples of prohormones secreted by such tumors include proopiomelanocortin and its cleavage products (␤-lipotropin, ␤-endorphin, ␣-melanotropin stimulating hormone (␣MSH), proinsulin and C-peptide, proglucagon, and progastrin). The detection of high-molecular-weight forms of ACTH, insulin, glucagon, and gastrin in the serum often indicates the presence of a NE tumor (118). This has major consequences for the choice of the assays used to measure these hormones (see below).

General Neuroendocrine Markers

A number of other components specific for neuronal and NE cells have been identified which can serve as tumor markers. The clinical significance of these markers is that they can be used independently of the specific peptides and/or amines produced by a given NE tumor. These markers are associated with secretory granules or small vesicles or are present in the cytosol of NE (tumor) cells.

Secretory Granule-Associated Markers Chromogranins A, B, and C form a group of acidic monomeric soluble proteins which are localized in the electron-dense core secretory granules, where they are costored and cosecreted with the locally present peptides (99). Although their biological role has not yet been firmly established, several functions have been postulated: they might exert regulatory activities in the packaging and processing of peptide hormones and in the modulation of NE secretion (19, 45). Chromogranin A (CgA) was originally discovered in the chromaffin cells of the adrenal medulla (8), which was also the source of most currently used immunoassays to measure serum CgA concentrations. CgA, CgB, and to a lesser extern CgC are powerful universal serum markers for NE tumors (19, 97, 117).

Cytosolic Markers and Oncogenic Proteins NSE is a neuron-specific isomer of the ubiquitous glycolytic enzyme 2-phospho-D-glycerate hydrolase or enolase (100). This isomer is present in neurons and NE cells and can serve as a serum marker for tumors derived from these

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cells (2). Carcinoembryonic antigen (CEA) is an oncogenic protein, which is not specific for NE cancers, but is frequently synthesized by medullary thyroid cancers (69).

Peptide Membrane Receptors

In recent years it has become evident that tumors with NE characteristics often express membrane receptors for small peptides, such as somatostatin, vasoactive intestinal peptide (VIP), bombesin, cholecystokinin, and/or Substance P (91). The demonstration of peptide receptors on tumors by ligand binding studies, autoradiography, immunohistochemistry, or in situ hybridization techniques has extended the number of NE markers which can be used in the pathological characterization of tumors (40, 12). However, apart from these in vitro investigations, peptide receptor expression by NE tumors can also be demonstrated in vivo after the administration of small amounts of peptides coupled to radionuclides (55, 56, 65, 67). This technique of peptide receptor scintigraphy has been developed successfully for the visualization of somatostatin receptors on NE tumors. After the intravenous administration of [ IIIIn-DTPA]octreotide most primary NE tumors, and often their previously unrecognized metastases, can be visualized (67, 56). The technique of somatostatin receptor scintigraphy (SRS) is an addition to the measurement of different circulating NE tumor markers, which gives information about the spread of the disease, but often also predicts a beneficial effect of therapy with somatostatin analogs (66, 67).

Molecular Diagnosis of DNA Mutations

Although not a NE tumor marker per se, the molecular diagnosis of welldefined mutations in the RET oncogene DNA sequence in children born in families with multiple endocrine neoplasia (MEN) types 2A and 2B can now be considered “a NE tumor marker,” indicating the necessity of preventive total thyroidectomy and/or adrenalectomy at a very young age (70). The demonstration of specific mutations, notably in codons 609, 611, 618, 620, and 634 in the case of MEN 2A, and in codon 981 in the case of MEN 2B of the RET oncogene sequence, makes the use of the pentagastrin stimulation test of calcitonin secretion less important (54). The molecular diagnosis of mutations in the MEN 1 gene and the Von Hippel Lindau gene are currently not routinely used as “NE tumor markers,” but are also important in identifying individuals who should be screened on a regular basis for increases in circulating NE tumor marker levels (54). Schematic representations of the different NE tumor markers in a given NE tumor cell are presented in Fig. 1 and Table 1.

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FIG. 1. A schematic representation of the pathways of protein/hormone secretion in a neuroendocrine cell, as well as a description of potential tumor markers. CgA, chromogranin A; NSE, neuron-specific enolase; CEA, carcinoembryonic antigen.

THE PRACTICAL USE OF NE TUMOR MARKERS: GENERAL ASPECTS Inappropriate Hormone Secretion

Classic, simple endocrine paradigms are of great help in the diagnosis of some NE tumors: hypercalciemia associated with an elevated serum parathyroid hormone (PTH) concentration, hypoglycemia associated with inappropriately elevated serum insulin concentrations, and elevated gastrin levels assoTABLE 1 Tumor Markers Used in the Diagnosis and Treatment of Patients with Neuroendocrine Tumors 1. Molecular diagnosis of mutations in the RET oncogene (MEN 2A and B) 2. (Pro)hormones and amines Peptides and glycoproteins Clinically functioning neuroendocrine tumors e.g., (pro)ACTH, (pro)gastrin, (pro)insulin, etc. Clinically nonfunctioning neuroendocrine tumors e.g., pancreatic polypeptide (PP), ␣-subunit, ␤-human chorion gonadotropin (␤-HCG) Amines: Serotonin, Substance P, tachykinins 3. Secretory granin proteins: CgA, CgB 4. Enzymes: neuron-specific enolase (NSE) 5. Oncogenic proteins: Carcinoembryonic antigen (CEA) 6. Membrane receptors: Somatostatin, VIP, Substance P

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ciated with a high acid output by the stomach are indicative of primary hyperparathyroidism, insulinoma, and gastrinoma, respectively (102). In contrast, hypercalcemia with virtually undetectable PTH levels should raise the suspicion of the presence of a tumor secreting PTH-related peptide, hypoglycemic attacks with undetectable insulin levels might point to a mesenchymal tumor secreting insulin-like growth factor-II, and elevated serum gastrin levels in patients with low acid production frequently indicate the presence of atrophic gastritis or the use of drugs lowering gastric acid production.

The Choice of the Assay

When studying the physiological role of a regulatory peptide, the antiserum used to measure the peptide concentration in the serum should preferably not cross-react with precursor forms, metabolites, or other related peptides (110, 115). However, when an antiserum is used to monitor peptide concentrations as NE tumor markers, it should preferably cross-react with as many molecular forms of the peptide family in question as possible, since tumors in individual patients may result in increases in different molecular forms of the peptide (115). For example it is well known that high-molecular-weight forms of ACTH are present in tumors from patients with nonpituitary tumors, of which some give rise to the “ectopic” ACTH syndrome (82). Stewart et al. (105) developed two antibody “sandwich-type” immunoassays that quantify and distinguish between ACTH precursors and ACTH. It was demonstrated that ACTH precursor molecules are present in increased concentrations and that the ratio of pro-ACTH molecules to ACTH is high in plasma from patients with cancerrelated Cushing’s syndrome (Fig. 2). In contrast in patients with pituitarydependent Cushing’s disease the conversion efficiency of pro-ACTH to ACTH is high and the pro-ACTH concentration is much lower (105). This example of ACTH demonstrates how important it is for the clinician to know what type of assay is used for hormone measurements in the laboratory: a sensitive assay for a given “native” hormone is very helpful in identifying the levels of the biologically active hormone, but such an assay might not be optimal or even be misleading in situations where there are high levels of precursors. Typical other clinical examples where this problem arises are insulin/proinsulin/C-peptide measuring assays, PTH assays which also measure parts of the molecule that circulates in high concentrations in renal insufficiency, and gastrin assays which also measure progastrin that, for example, can be secreted by colorectal carcinomas (6).

The Use of Stimulation Tests and Selective Catheterization

Many techniques have been introduced in order to increase the diagnostic value of hormone measurements in the diagnosis of NE tumors. Some of these

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FIG. 2. The circulating ACTH concentrations in patients with the ectopic ACTH syndrome (a), Cushing’s disease (b), and controls (c). Plasma concentrations of native ACTH 1–39 (o) and ACTH precursors (●) were measured using specific monoclonal-based immunoradiometric assays at 0900 h (adapted from 105).

tests, however, have become outdated or should be used in exceptional situations only (110). In the diagnosis of pancreatic endocrine tumors the 72-h fasting test to demonstrate inappropriate insulin secretion during hypoglycemia remains central in the diagnosis of insulinoma (102). However, standardized meal stimulation tests to differentiate between normal and tumoral secretion patterns of pancreatic polypeptide secretion (103), secretin stimulation tests of gastrin secretion to diagnose a gastrinoma, pentagastrin stimulation tests of calcitonin release to diagnose medullary thyroid cancer, and calcium infusion tests to diagnose primary hyperparathyroidism now seem hardly ever to be necessary in daily practice. Selective catheterization of the arterial tree of the gastrointestinal tract during angiography in order to take blood samples for hormone determination to localize a NE tumor seems also largely outdated since peptide receptor scintigraphy with three-dimensional SPECT images became available for routine use. Only the selective catheterization of the inferior petrosal sinus for CRH-stimulated ACTH measurement in the differential diagnosis of Cushing’s syndrome remains of frequent benefit (41). General Markers

The role of CgA as a universal NE tumor marker has some shortcomings. A major problem for the diagnostic use of CgA measurements is that CgA is

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processed in a tissue- and tumor-specific manner into a multitude of different fragments (76). Depending on the antiserum raised, only CgA assays recognizing the most carboxy terminal part of the molecule appear useful in the diagnosis of NE tumors. For example, in a carcinoid tumor the CgA concentration varied from 0.5 to 34.0 nmol/g of tissue depending on the specificity of the CgA assay, with the lowest concentration measured with assays specific for the NH 2-terminus of CgA. The epitope specificity of CgA antisera thus profoundly influences diagnostic sensitivity (78). The variable processing of the CgA molecule by different tumors, on the other hand, might necessitate the use of a library of sequence-specific CgA immunoassays for an optimal use as a NE tumor marker. Eriksson et al. (23) developed a polyclonal antiserum against CgA and CgB and compared it with an antiserum directed against CgA only. Cg determinations with the former antiserum appeared to be a much more sensitive marker than those with the latter one, both immunohistochemically and in the circulation. The concentrations of circulating “granins” were about 10 times higher with the CgA ⫹ B antiserum than with the CgA antiserum. Assays using combinations of such antisera might substantially improve the sensitivity of the detection of small tumors. CgA is a relatively stable peptide. Serum CgA levels drawn on 2 consecutive days correlated closely (3). The circulating plasma “pool” of CgA is, however, substantially greater than that of most peptide hormones and as a result is more difficult to acutely perturb. Because of the many potential tissue sources of the peptide, and the relatively high baseline level in normal individuals, small NE tumors secreting CgA cannot be detected. A certain size of the tumor and the ensuing increased contribution of this tumor CgA to the plasma “pool” eventually allows the detection of such a tumor. In many studies serum CgA levels have been reported to assess tumor size of different NE tumors accurately, but again for small tumors CgA measurements demonstrate a low sensitivity (32). To complicate matters further the kidney excretes CgA and mild renal insufficiency is already accompanied by an elevation of serum CgA levels (44). For example, the utility of CgA measurements during chemotherapeutic treatment of NE tumors with streptozotocin is not possible, as the nephrotoxicity of the drug itself results in increased CgA levels, even if tumor mass shrinks (81). Also, liver function disturbances sometimes cause increased CgA levels (81). Apart from these shortcomings serum CgA levels can be successfully used as a predictive tumor marker in the follow-up of most patients with NE tumors (Fig. 3; 77, 5).

Peptide Receptor Scintigraphy

Somatostatin receptors have been shown to remain present on a variety of tumors which arise in tissues which also contain these receptors in the normal state. Large numbers of high-affinity somatostatin receptors have been found on most growth hormone secreting pituitary adenomas, as well as on most

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FIG. 3. Serum concentrations of chromogranin A (CgA) in patients with neuroendocrine tumors and in controls with nonendocrine tumors. Individual levels are presented as dots and median levels as solid lines. The dashed line represents the upper cutoff level of normal values. The results are plotted logarithmically to accommodate extreme values. MTC, medullary thyroid carcinoma; pheo, pheochromocytoma; SCLC, small cell lung carcinoma; NF, nonfunctioning (adapted from 77). Nonendocrine tumors include a great number of patients with mostly adenocarcinomas. Interestingly, a portion of them demonstrated increased CgA levels (see text).

metastatic endocrine pancreatic tumors and carcinoids (91). In parallel, chronic therapy with the somatostatin analog octreotide eliminates clinical symptomatology, as well as the biochemical abnormalities in most acromegalic patients: both the hypersecretion of growth hormone and the elevated circulating levels of insulin-like growth factor I virtually normalize in most instances (64). Hormonal hypersecretion from metastatic endocrine pancreatic tumors and carcinoids is also well controlled during octreotide treatment of most patients, while in parallel the clinical symptomatology greatly improves. Interestingly, evidence of control of tumor growth during somatostatin analog treatment has been observed in some of these patients (58). These results led to an instant improvement in the quality of life of these patients, making the clinical introduction of octreotide a major breakthrough in the treatment of these endocrine cancers (68). The technique which allows the detection of somatostatin receptor-positive tumors in vivo after the administration of a radioactive iodine-labeled analog might be interpretated as an “in vivo autoradiography” of somatostatin receptor-positive tumors (66). Tyr 3-octreotide is a somatostatin analog with tyrosine in position 3, where phenylalanine is present at that place in octreotide. The biological activities of octreotide and Tyr 3-octreotide are similar. Initially Tyr 3octreotide coupled to 123I was used in patients who were suspected of having somatostatin receptor-positive tumors, while planar or ECT (emissioncomputed tomography) images were made with a gamma camera (65). The [ 123I-Tyr 3]octreotide scanning procedure revealed the localization of the pri-

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TABLE 2 Results of Somatostatin Receptor Scintigraphy with [ IIIIn-DTPA]Octreotide in Patients with Neuroendocrine Tumors. Endocrine pancreatic tumors Gastrinoma Insulinoma Glucagonoma Nonfunctioning Carcinoids Small cell lung cancer Medullary thyroid cancer Paraganglioma Pheochromocytoma

12/12 14/23 3/3 16/18 69/72 34/34 20/28 33/33 12/14

mary tumor and/or its previously unknown metastases in 18 of 21 patients with endocrine pancreatic tumors (86%). In a group of 5 of these positive tumors we subsequently investigated the surgically removed tumor (65). There was a close relationship between the in vitro detection of somatostatin receptors in these tumors using autoradiography and the gamma camera pictures obtained after injection of [ 123I-Tyr 3]octreotide in vivo (66). This indicates that the ligand binding to the tumor in vivo represents specific binding to somatostatin receptors. In addition, we carried out preoperative in vivo and in vitro experiments with cultured tumor cells. Again there was a close parallel between the presence of somatostatin receptors on these tumors and the in vivo and in vitro effects of octreotide on hormonal secretion by these tumors (66). This means that a positive scan predicts a beneficial effect of therapy with octreotide on hormonal hypersecretion by these tumors. In 30 of 31 patients with metastatic carcinoids we visualized the primary and secondary tumors, while these “positive” patients were subsequently shown to respond beneficially to therapy with octreotide (65, 67). [ 123I-Tyr 3]Octreotide has two major drawbacks: it has a very short half-life and there is a high background of radioactivity in the abdomen. Therefore, an alternative was developed by coupling DTPA (diethylenetriaminopentaacetic acid) to octreotide. [ IIIIn-DTPA°]Octreotide has a longer half-life and is excreted via the kidneys. This new compound turned out to be able to visualize somatostatin receptor-positive tumors at least as clearly as [ 123ITyr 3]octreotide (56). Apart from endocrine pancreatic tumors and carcinoids we extended our studies to the use of the somatostatin receptor imaging technique using [ IIIInDTPA°]octreotide in other tumors with neuroendocrine characteristics (Table 2). In many instances multiple tumor localizations, as well as metastatic disease could be demonstrated. In subsequent years it was demonstrated that somatostatin receptor scintigraphy (SRS) was not entirely specific for the visualization of NE tumors. Most well-differentiated human brain tumors, like meningiomas and low-

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grade astrocytomas, contain somatostatin receptors, while undifferentiated brain tumors mainly contain epidermal growth factor receptors (92–94). All 11 meningiomas and 4 low-grade astrocytomas investigated could be clearly visualized with somatostatin receptor imaging techniques, while 3 glioblastomas were negative (67). Other tumors which express enough somatostatin receptors to allow their in vivo visualization include part of the neuroblastomas, Merkel cell tumors, Hodgkin and non-Hodgkin lymphomas, as well as ⬎50% of primary breast cancers (56). Also, most granulomatous and autoimmune diseases, such as sarcoidosis, tuberculosis, Wegener’s granulomatosis, DeQuervain’s thyroiditis, aspergillosis, Graves’ hyperthyroidism, and Graves’ ophthalmopathy are visualized by SRS (63, 89, 112). At SRS, normal scintigraphic features include visualization of the normal thyroid, spleen, liver, kidneys, and in some patients the pituitary gland (Fig. 4). The visualization of the pituitary gland, thyroid, and spleen occurs because of receptor binding. Uptake in the kidneys is for the most part from reabsorption of the radiolabeled peptide in the renal tubular cells after glomerular filtration, while somatostatin receptors have been demonstrated in human renal tubular cells and vasa recta (95). “False-positive” results of SRS have been reported. In virtually all cases the term “false-positive” is a misnomer because somatostatin receptor-positive lesions are present that are not related to the pathology for which the investigation is performed (63). Examples include the visualization of the gallbladder, thyroid abnormalities, accessory spleens, activity at the site of a recent surgical incision, and other conditions (30). In some patients the coexistence of two different somatostatin receptor-positive disease processes should also be considered. SRS in virtually all cases visualizes somatostatin receptor-positive processes: its sensitivity is very high, but because of the highly variable expression of somatostatin receptors on both NE and non-NE tumors and immune diseases its specificity is low.

NEUROENDOCRINE TUMORS Endocrine Pancreatic Tumors

Most of these tumors, which originate from pancreatic islet cells, secrete multiple peptide hormones simultaneously. Clinical syndromes associated with these tumors indicate in at least two thirds of the patients which hormonal marker should be measured in a given patient. In order of frequency, these syndromes include the insulinoma syndrome (hypoglycemic attacks), the gastrinoma syndrome (Zollinger-Ellison syndrome), the watery diarrhea hypokalemia achlorhydria syndrome (Vipoma), the glucagonoma syndrome (necrolytic skin disease, thrombosis), and the somatostatinoma syndrome (steatorrhea, gallstones, secondary diabetes). In about one third of patients with endocrine pancreatic tumors no specific clinical symptomatology is recognized.

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FIG. 4. Somatostatin receptor imaging, 24 h after injection; anterior images of the head (top), chest (middle), and abdomen (bottom) showing normal uptake in the thyroid, liver, spleen, and kidneys and also some bowel activity. Clear uptake in a residual nonfunctioning neuroendocrine pancreatic tumor (bottom), with abdominal (bottom) and skeletal metastases in the chest (middle) (courtesy of Dr. D. J. Kwekkeboom).

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These clinically “nonfunctioning” tumors are diagnosed by chance on radiological examination of the abdomen or when they cause local compression of the pancreatic or bile ducts. In very rare cases “ectopic” production of ACTH or growth hormone releasing hormone by an endocrine pancreatic tumor causes Cushing’s syndrome or acromegaly, respectively. Prolonged (72-h) fast is the classic diagnostic test for an insulinoma (102). Blood glucose levels below 2.2 mmol/liter, with insulin levels being 6 ␮U/ml (36 pmol/liter) or higher in a polyclonal assay, or 3 ␮U/ml or higher in an immunochemoluminometric assay are indicative for an insulinoma. Criteria for hyperinsulinemia measuring proinsulin or C-peptide also exist (102). Recently, a 48-h fast was demonstrated to be of virtually equal value as the 72-h fast in the diagnosis of insulinomas (43). Localization procedures of insulinomas have a low sensitivity. Echoendoscopy but especially intraoperative ultrasound are the best techniques for localizing these, in most instances, small pancreatic tumors. Elevated gastrin levels are in most patients associated with hypo- or achlorhydria, which is related to atrophic gastritis or the use of acid lowering drugs. The presence of an elevated serum gastrin concentration together with a high basal acid output (⬎10 mmol/h) is diagnostic of a gastrinoma (28). In 112 consecutive patients with the Zollinger-Ellison syndrome the sensitivity for the presence of gastrinoma tissue was higher for CgA than for fasting serum gastrin concentrations (92% vs 80%; p ⫽ 0.02) (31). However, the specificity of CgA was 67%. Further, gastrinoma growth was readily detected by following serum CgA concentrations. In the initial diagnosis of gastrinomas elevated serum CgA levels seem, however, not to reflect tumor size, but the degree of endocrine cell hyperplasia in the stomach (77). Primary duodenal gastrinomas are best detected by duodenoscopy, while the localization of primary pancreatic gastrinomas and their metastases are best visualized by SRS (29). SRS was superior to any other single imaging study in 122 consecutive gastrinoma patients and altered management in 47% of them (109). The Vipoma, glucagonoma, and somatostatinoma syndromes are in most patients clinically well recognizable, and the measurement of the respective hormones in the serum, together with the demonstration of the primary tumor and the often already present metastases by SRS, completes the diagnosis. Serum CgA concentrations are elevated in most patients as well and thus represent for many clinicians an easier alternative serum tumor marker in the follow-up. Clinically “nonfunctioning” pancreatic tumors often secrete pancreatic polypeptide (PP) (87). PP is secreted by normal pancreatic islet cells in response to the presence of nutrients in the gut lumen. The function of the peptide has not yet been elucidated. Serum PP concentrations are elevated in 15 to 74% of patients with endocrine pancreatic tumors, most frequently in combination with other peptide hormones (87, 90). In most patients, however, the effects of these concomitantly secreted peptides dominate the clinical picture. When PP is the sole hormonal product of the tumor, no endocrine clinical syndrome develops, even when serum concentrations exceeding 1000

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TABLE 3 Serum Levels of Chromogranin A, Neuron-Specific Enolase, and ␣-Subunit as well as the Result of Somatostatin Receptor Scintigraphy in 13 Patients with Clinically Nonfunctioning Tumors of the Endocrine Pancreas (adapted from Nobels et al., 1997) Patient

CgA (␮g/liter)

NSE (␮g/liter)

␣-Subunit (␮g/liter)

SRS

1 2 3 4 5 6 7 8 9 10 11 12 13

969 a 306 a 999 a 14750 a 94 641 a 207 a 99 910 a 211 a 85 126 567 a

11.4 4.6 5.6 10.3 12.6 6.8 15.5 a 11.0 90.8 a 8.1 5.3 15.6 a 8.0

1.9 a 1.7 1.1 2.1 2.0 1.3 a 1.0 1.0 1.1 1.0 0.7 1.2 a 0.7

⫹ ⫹ ⫹ ⫹ ⫹ ⫹ ⫹ ⫹ ⫹ ⫹ ⫹ ⫹ ⫹

Note. Upper cutoff values: CgA, 175 ␮g/liter in men and premenopausal women, 220 in postmenopausal women; NSE, 12.5 ␮g/liter, ␣-SU, 1.1 ␮g/liter in men, 2.3 ␮g/liter in premenopausal women, and 4.0 ␮g/liter in postmenopausal women. a Elevated levels.

times the normal level are measured (1). In a recent study, the presence of a markedly elevated fasting serum PP level in patients with MEN1 was 95% sensitive and 88% specific for the presence of radiographically detectable pancreatic islet cell tumors (74). In patients with clinically “nonfunctioning” endocrine pancreatic tumors serum NSE, ␤-HCG, and ␣-subunit concentrations are also frequently elevated. In a series of 13 patients serum CgA levels were increased in 9 (69%), but all 13 tumors were positive on SRS as well (Table 3; 77). These data suggest that for clinically “nonfunctioning” endocrine pancreatic tumors the measurement of serum PP (if available) and CgA levels, together with SRS, is currently the most optimal diagnostic approach. An important, as yet unresolved, question is whether this diagnostic approach should also be used in the differential diagnosis of pancreatic duct cancers. In none of 26 patients with pancreatic duct cancer was SRS positive (25). In a follow-up study of 62 patients with pancreatic duct cancer, 12 patients were alive for more than 3 years. Seven patients were free of metastases. Five of these 12 patients were known to have metastases. SRS was positive in all 5 patients and revision of the tumor tissue in all instances demonstrated NE positivity with different staining techniques. This study suggests that SRS might be of use in the preoperative differential diagnosis between pancreatic duct cancer and clinically “nonfunctioning” endocrine pancreatic tumors (25). In Table 4 an overview is presented of the results of the measurement of a number of specific and general tumor markers in a total of 165 patients with

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TABLE 4 Tumor Secretory Products Elevated at First Evaluation in Patients with Endocrine Pancreatic Tumors and Carcinoids Endocrine pancreatic tumors (%) (Reference and number of patients)

Tumor marker Specific Insulin Gastrin VIP Calcitonin Glucagon Somatostatin 5-HIAA UFC PTH-rP General Chromogranin A Chromogranin A⫹B Pancreatic polypepeptide Neurotensin ␤-HCG ␣-Subunit NSE Neuropeptide K

Eriksson and Oberg (24) n ⫽ 28

46 62 23 42 34 21

Carcinoids (%) (Reference and number of patients) Baudin et al. (7)

Baudin et al. (7) n ⫽ 38

Nobels et al. (17) n ⫽ 43

Foregut n ⫽ 36

Midgut n ⫽ 33

Norheim et al. (79) n ⫽ 103

Nobels et al. (77) n ⫽ 59

15 13

0 10 10 0

28

3

9 44 9 3

0 85 0 0

4

88

46

80

94 74 67 30 41

21 26 45

14 37

17 30 42

0 0 42

43 12 28

39 47

66

Note. UFC, urinary free cortisol. The number of patients with elevated tumor secretory products at first elevation are presented as a percentage of the total number of patients per study.

endocrine pancreatic tumors from three different series (24, 7, 77). From this overview it becomes evident that most of these tumors simultaneously secrete multiple hormones and general tumor markers. An optimal choice of what to measure in a specific patient primarily depends on clinical symptomatology. Once the clinical and/or biochemical diagnosis of an endocrine pancreatic tumor has been made, visualization of the primary as well as metastatic tumors can be made by endoscopic ultrasound, CT, MRI, or SRS. The role of SRS in the different tumor types is demonstrated in Table 2.

Carcinoids

While in two thirds of the patients with endocrine pancreatic tumors specific hormones related to a clinical syndrome can be relatively easily detected and used as tumor marker, this is not the case in carcinoids. Their clinical expression differs according to their origin from fore-, mid-, or hindgut. The typical carcinoid syndrome, consisting of diarrhea, flushing, and bronchospasm, is

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caused by midgut carcinoids (27). Unfortunately, since their secretions drain into the portal circulation and are degraded by the liver, such tumors usually become symptomatic only after substantial hepatic metastases have developed. In rare cases, when carcinoid tissue is present in the retroperitoneum or the ovaries, which drain directly into the systemic venous circulation, the syndrome can manifest itself in the absence of liver metastases (27). Serotonin (5-hydroxytryptamine) is one of the major secretion products of midgut carcinoids (86). The measurement of the serotonin metabolite 5-hydroxyindolacetic acid (5-HIAA) in the urine is the most frequently used parameter in the diagnosis and follow-up of midgut carcinoids. Using an upper cutoff value of 8 mg/24 h the sensitivity in the diagnosis of metastasized carcinoids is 85– 88%, with a specificity of 100% (Table 3). Direct measurement of plasma serotonin concentrations, even when determined in platelet-rich plasma, does not provide better results (26). Although the carcinoid syndrome originally was considered due to the effects of serotonin secretion, midgut carcinoid tumors are now known to secrete a mixture of bioactive amines and peptides, including histamine, protaglandins, tachykinins (most notably substance P and neuropeptide K), neurotensin, kallikreins, and bradykinin-like peptides (98, 71, 34, 51). These secretion products may actually be responsible for a large proportion of the clinical effects that previously have been attributed to increased levels of serotonin. Unfortunately, the serum concentrations of these peptides are not universally elevated in patients with the carcinoid syndrome, and they can also be produced in other situations where flushing occurs (medullary thyroid carcinoma, Vipoma, idiopathic flushing) (53, 113). Therefore, it is only recommended to screen for some of the above peptides if urinary-5HIAA levels are normal, in the presence of firm clinical suspicion of carcinoid syndrome. The use of provocative tests generally does not add to the information provided by the basal measurements. As demonstrated in Table 4, CgA is a very suitable serum tumor marker for carcinoids; it is a more stable and thus more easily manageable marker than 5HIAA measured in a 24-h urine sample (78). In one series a close correlation was reported between serum CgA concentrations and tumor size (77). While 5-HIAA and CgA are the most sensitive tumor markers for midgut carcinoids, foregut carcinoids less frequently secrete serotonin (Table 4, 79). In foregut carcinoids, CgA, but also NSE (sensitivity 42%) are the preferred tumor markers. Very few series of hindgut carcinoids have been reported, so the current advice is to use a series of tumor markers (including CgA, NSE, and ␣-subunit) in such patients (7). Most carcinoids express large numbers of somatostatin receptors, making them good candidates for visualization with SRS. In a large series of 22 patients with carcinoid tumors the sensitivity of SRS in visualizing the primary as well as metastatic carcinoids was 96% (56, 62). Data from our own series so far indicate a similar homogeneous and dense pattern of SS receptor expression over primary and metastatic tumors without important heterogeneity.

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325

Medullary Thyroid Carcinoma (MTC)

MTC is a neuroendocrine tumor arising from the calcitonin (CT)-producing parafollicular C-cells in the thyroid. CT is an excellent tumor marker for MTC. Routine measurement of serum CT concentrations in 1385 consecutive patients with nodular thyroid disease allowed a preoperative diagnosis of unsuspected sporadic MTC in 8 cases (0.57%) (83). Apart from being a specific and sensitive marker of MTC, the actual serum CT concentration is predictive for the size of the MTC (13). As MTC cells dedifferentiate, they express higher amounts of the oncogenic protein carcino embryonic antigen (CEA). Previous studies have demonstrated a variable loss of CT synthesis and secretion by MTCs exhibiting more aggressive malignant behavior (72). This is recognized by a discrepant steeper increase in serum CEA concentrations than in those of CT (11). In the past, stimulation tests, mainly using pentagastrin, were used to detect paradoxical rises in serum CT concentrations. Those were indicative of early malignancy of C-cells. Candidates for such studies were individuals known to belong to families in which MEN 2A, B, or familial medullary thyroid cancer occur. However, such individuals can now be identified much more easily and earlier by molecular diagnosis of specific mutations in the RET oncogene, necessitating preventive thyroidectomy (70). In this sense DNA sequence analysis of the RET oncogene can be considered a NE tumor marker predictive for subsequent tumor development. MTC can best be visualized by ultrasound or MRI, SRS being less reliable because distant MTC metastases sometimes dedifferentiate, losing somatostatin receptor expression (60). The measurement of CgA and/or NSE does not provide additional information; the sensitivity of CgA was 50% and of NSE 42%, respectively, in 26 consecutive patients with metastatic MTC (77). Primary Hyperparathyroidism

The presence of hypercalcemia, associated with an elevated serum PTH concentration, measured in a modern two-site assay is usually sufficient to make the diagnosis. In all 19 patients with different hyperparathyroid states (primary, secondary, as well as tertiary hyperparathyroidism) serum CgA levels were increased (80). No additional value of serum CgA measurements in the diagnosis of parathyroid disease has been demonstrated as yet. If additive preoperative information concerning the localization of the affected parathyroid(s) is required, 99Technetium-Sestamibi scanning is successful, with a sensitivity of 80 –100% and specificity of 77–100% (47, 73). Parathyroid glands do not express somatostatin receptors (67). Pheochromocytomas

The measurement of plasma or urinary excretion of catecholamines or their metabolites remains the basis for the biochemical diagnosis of pheochromocy-

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tomas. Metabolites in the form of vanillylmandelic acid (sensitivity 47–90%; specificity 87–100%) or metanephrine/normetanephrine excretion in the 24-h urine (sensitivity 97–100%; specificity 84 –98%), as well as the measurement of plasma concentrations of epinephrine, norepinephrine, and dopamine (sensitivity 85–100%; specificity 72–99.5%), are most commonly used (9, 84). Medication, such as labetolol, tricyclic antidepressants, and methyl- and levodopa sometimes interfere with these measurements, however (9). In a recent study we confirmed in 87 patients with pheochromocytomas that basal plasma catecholamine levels were increased in 81 cases (sensitivity 93%) (39). Of 6 cases with normal circulating catecholamine levels, medullary hyperplasia was found in 4 patients. Fourteen patients developed metastases. Patients with higher levels of dopamine and norepinephrine had significantly shorter metastases-free intervals. Bilateral, hereditary tumors were less productive than their unilateral counterparts (39). Catecholamines and CgA are coreleased from the adrenal medulla and sympathetic neuronal vesicles. Consequently, serum CgA concentrations are elevated in virtually all pheochromocytoma patients (sensitivity 83 and 89%, respectively in two series, with a specificity of 100%) (19, 27, 80). Also, serum CgA concentrations closely correlated with pheochromocytoma sizes (19). Selecting the appropriate approach for resection and follow-up of pheochromocytomas is highly dependent upon reliable localization and exclusion of multifocal, bilateral, or metastatic disease. Metaiodobenzylguanidine (MIBG) scintigraphy is used for functional localization of catecholamine-secreting tissues. We performed 123I-MIBG scans in a total of 75 patients with pheochromocytomas, in 70 cases prior to resection of the primary tumors, and in 5 cases because of recurrent disease (38). The overall detection rate was 83.3% for pheochromocytomas greater than 1.0 cm in diameter, respectively. 123I-MIBG uptake correlates with greater size of the pheochromocytomas ( p ⫽ 0.008). 123 I-MIBG-negative pheochromocytomas (n ⫽ 14) had significantly smaller diameters than 123I-MIBG-positive tumors ( p ⫽ 0.01). SRS was successful in visualizing only 25% (8/32) of primary benign pheochromocytomas. In 14 patients with pheochromocytoma metastases, 123I-MIBG visualized tumors in 8 cases. SRS, however, was able to detect metastases in 7/8 cases, including 3 123 I-MIBG-negative metastatic cases (38). In conclusion, 123I-MIBG uptake depends upon the size of pheochromocytomas. Its role in visualizing small bilateral and MEN-2-related pheochromocytomas seems limited. In cases of recurrent elevation of catecholamines, localization of metastases and/or recurrence should be attempted with 123I-MIBG scintigraphy, but SRS should be considered an additional technique (38).

Paragangliomas

Paragangliomas are extraadrenal tumors arising from the sympathetic and parasympathetic autonomic nervous system. Histologically they demonstrate a positive chromaffin staining. More than 75% of them are nonfunctional.

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Evidence of hypersecretion of catecholamines was reported in about 1% of cases. In 25 patients with paragangliomas serum NSE, ␣-subunit and CgA concentrations were elevated in 9 (36%), 3 (12%), and 2 (8%) of cases, respectively (Table 4, 77). Virtually all paragangliomas express somatostatin receptors. SRS visualized 50/53 paragangliomas (94%) (61); SRS demonstrated unexpected multiple localizations of paragangliomas in 9 of 25 patients (36%). This finding is of special interest since multicentricity and distant metastases each may occur in 10% of patients with paragangliomas (35). One of the major advantages of SRS is that it provides information on potential tumor sites in the whole body. SRS can thus be used as a screening test, to be followed by CT scanning, MRI, or ultrasound of the sites at which abnormalities are found (56).

Small Cell Lung Cancer (SCLC)

Elevated levels of NSE are found in 60 – 80% of patients with SCLC. This makes it a useful diagnostic test in patients with suspected pulmonary lesions. However, the specificity of NSE measurements remains low, as also foregut carcinoids (⫾50%) and non-small cell lung cancers (20%) cause elevated serum NSE concentrations (15). NSE levels are higher in patients with extensive disease, but there is considerable overlap with limited disease (48). NSE can be used as a prognostic factor to monitor tumor size during active treatment. Serum CgA concentrations were elevated above twice the upper limit of normal in 121/154 patients with untreated SCLC and were in one study better than NSE in detecting early recurrence (48, 22). In our own study (77) in 23 as yet untreated patients with small cell lung cancer, NSE levels were elevated in 74% of cases, while CgA and ␣-subunit levels were each elevated in about one third of cases (Table 5). SRS revealed the primary tumors and their metastases (e.g., in the brain) in all 34 patients with small cell lung cancer (sensitivity 100%). However, the specificity was very low, as also all primary non-small cell lung cancers in 36 patients were visualized, and bronchial carcinoids can also be visualized by SRS (63).

Pituitary Adenomas

Pituitary adenomas can be classified according to the clinical syndrome related to hormonal hypersecretion. Prolactinomas are diagnosed by the measurement of a single sample for PRL, in combination with MRI and ophthalmological investigation. Also, nonpituitary and non-PRL secreting pituitary tumors can induce hyperprolactinemia via compression of the pituitary stalk. Although not yet proven to be entirely specific scintigraphy with 123I-labeled benzoxamide (a dopamine receptor antagonist) is helpful in the differential diagnosis between pituitary and nonpituitary tumors (42). Acromegaly is di-

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TABLE 5 A Comparison between the Diagnostic Value of Serum Measurements of Chromogranin A (CgA), Neurospecific Enolase (NSE), and ␣-Subunit in Patients with Neuroendocrine Tumors (from 77) Tumor Insulinoma (n ⫽ 21) Gastrinoma (n ⫽ 9) Nonfunctioning endocrine pancreatic tumor (n ⫽ 13) Carcinoid (n ⫽ 59) Medullary thyroid carcinoma (n ⫽ 26) Paraganglioma (n ⫽ 25) Pheochromocytoma (n ⫽ 9) Neuroblastoma (n ⫽ 3) Small cell lung cancer (n ⫽ 23) Merkel cell tumor (n ⫽ 4) Clinically nonfunctioning pituitary adenoma (n ⫽ 10) Growth hormone secreting pituitary adenoma (n ⫽ 6)

Elevated CgA (%)

Elevated NSE (%)

Elevated ␣-SU (%)

10 100 69 80 50 8 89 33 39 25 20 0 50

38 44 31 47 42 36 44 67 74 50 0 0 43

0 33 23 39 19 12 11 33 35 0 20 0 24

agnosed by the measurement of serum total IGF-I concentration in one single blood sample. The biochemical basis of the diagnosis of Cushing’s syndrome rests on the demonstration of an increased excretion of urinary free cortisol and a disturbance in the overnight 1-mg dexamethasone suppression test. Subsequently Cushing’s disease (ACTH secreting pituitary microadenoma) can be diagnosed by selective sampling of the inferior petrosal sinus (41). Serum CgA levels were markedly elevated in 7 of 10 patients with NE tumors with ectopic ACTH secretion and in none of 5 patients with Cushing’s syndrome caused by adrenal tumors (76). However, elevated serum CgA measurements in patients with Cushing’s syndrome have a low specificity, as slightly elevated CgA levels were also found in 3 of 15 patients with Cushing’s disease (76). The use of SRS in the detection of small NE tumors secreting ACTH ectopically was successful only in 4 of 12 patients (108). In addition, SRS did not offer greater sensitivity than detailed CT/MRI imaging in 18 patients (111). Therefore, although helpful in selected cases SRS has no significant advantage over conventional imaging for ectopic ACTH secreting tumors. Clinically nonfunctioning pituitary adenomas comprise about 30% of pituitary tumors. Unlike other types of pituitary tumor, most nonfunctioning pituitary adenomas present with pressure symptoms and/or hypopituitarism rather than with clinical syndromes related to hormone hypersecretion. Nonfunctioning adenomas and gonadotropinomas are almost certainly derived from the same gonadotrope cell lineage. They form a part of a spectrum of phenotypes, ranging from the truly null cell adenoma to the gonadotropinoma, which may be associated with (sometimes markedly) elevated serum gonadotripin levels (37).

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Some gonadotropinomas and clinically nonfunctioning tumors secrete intact LH and/or FSH, but most commonly they secrete biologically inactive ␤-subunits and/or the glycoprotein ␣-subunit (59). Elevated levels of intact FSH may be detected in about 20% of patients with nonfunctioning pituitary adenomas (52). Intravenous TRH injection may elicit a rise in gonadotropin and/or subunit secretion from nonfunctioning adenomas. Because about 40 – 68% of patients with these tumors exhibit this paradoxical gonadotropin response to TRH stimulation, the clinical use of this test has been suggested as an important way of recognizing the pituitary nature of an intrasellar mass and as a marker to monitor the effectiveness of therapy (17). We tested whether CgA could serve as a serum marker for clinically nonfunctioning pituitary adenomas (75). Immunohistochemical staining for CgA was positive in 15 of 18 clinically nonfunctioning adenomas and negative in all 20 control tumors (craniopharyngeomas, germinomas, astrocytomas, meningeomas). CgA was present in the culture medium of 16 of 18 nonfunctioning adenomas in vitro. In 3 adenomas it was present in the absence of detectable amounts of gonadotropins or ␣-subunit. Basal serum levels of gonadotropins and/or ␣-subunit were elevated in 7 of 22 patients, while basal CgA levels were elevated in 2 of them. Significant increases in serum gonadotropin and/or ␣-subunit levels in response to TRH occurred in 14 of 21 patients with clinically nonfunctioning adenomas, while a significant increase in CgA levels after TRH was demonstrated in 6 patients (72). Although virtually all clinical nonfunctioning pituitary adenomas synthesize and secrete CgA, and normal values are too high, tumor mass is in general too small to allow CgA measurement to be used as a tumor maker. Another important aspect of our study was that the paradoxical increase in gonadotropin levels in response to TRH also occurred in 13 of 20 patients with other pituitary tumors (PRL and/or GH secreting tumors and nonendocrine tumors), demonstrating that this test is not specific for clinically nonfunctioning adenomas (72).

NEUROENDOCRINE DIFFERENTIATION OF CANCER

Increasingly, in recent years it has been recognized that human malignancies often show mixed or divergent differentiation. Occult NE features are frequently demonstrated at submicroscopic evaluation, with NE markers of many different types of tumors. Most cancers demonstrate a high degree of cellular heterogeneity. Heterogeneity is evident among different cells of the same tumor at a given time (intratumor heterogeneity), as well as at different points in time (tumor progression). The selection of different tumor cell clones, which express NE markers during the process of dedifferentiation, is a common feature of prostate, breast, and colon cancers. Elevated CgA levels are therefore not entirely specific for NE tumors. Slightly elevated levels have been identified in patients with different nonendocrine tumors (50, 20, 104, 77, 107, 16). Immunohistochemical studies reveal

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that many normal nonendocrine tissues contain NE cells, and these cells are also present in most tumors of nonendocrine origin (18, 21, 85, 101, 106). They are either diffusely scattered throughout the tumor or multifocally located in small nests. They are also present in metastatic tumor tissue (4). It is hypothesized that these cells produce peptides which affect the growth of tumor cells via an autocrine or paracrine interaction (49). The origin of these cells is not known. They are either cells of the diffuse NE system that are attracted by the tumor and are stimulated to proliferate or cells of the tumor itself that undergo NE differentiation (10, 14). The observation by Aprikian et al. (3) that NE cells in prostatic adenocarcinoma coexpress CgA and prostate-specific antigen (PSA) suggests a common malignant precursor cell. Moreover, these NE cells often show morphological changes such as nuclear hyperchromasia and pleomorphism, also indicating a malignant origin (36). Preliminary data from some (16, 21), but not all (57) investigators suggest that prostatic adenocarcinomas containing NE cells are more resistant to hormonal treatment (16, 21, 57). Similarly, patients with colorectal adenocarcinoma containing numerous NE tumor cells seem to have a worse prognosis (18, 36, 107). On the other hand, patients with pancreatic adenocarcinoma (85) or with non-small cell lung carcinoma (106) whose cancer contains many NE cells seem to have a better prognosis. In an as yet undetermined group of patients with adenocarcinomas, slightly elevated serum CgA concentrations are found (Fig. 3) Although NE markers are more and more used in the pathological classification of non-NE cancers, the use of serum NE markers is not yet extensively applied, except in patients with small cell lung cancers and neuroblastomas. Especially in this last disease the use of serum CgA measurements is a valuable (sensitive and specific) diagnostic tool which correlates with tumor size and is useful as a predictor of survival (44, 114).

THE DIFFERENTIATION STATE OF NE TUMORS

Virtually all NE tumors are potentially malignant, regardless of where they originate. There are two notable exceptions, pituitary adenomas and parathyroid adenomas, which are benign in most cases. Even if they demonstrate local invasiveness, they only seldom metastasize. A major shortcoming of the pathological examination of most NE tumors, such as carcinoids, islet cell tumors, pargangliomas, and pheochromocytomas, is that neither macro- nor microscopic characteristics of these tumors reliably predict whether they will behave aggressively. Neuroendocrine tumors commonly exhibit multiple lines of divergent differentiation, ultimately making them candidates for malignant expansion. In clinical practice some NE tumor markers can be used in predicting the degree of malignancy. Local invasive growth and infiltration of adjacent organs by NE tumors is generally accompanied by a high prevalence of immunohistochemically demonstrable ␤-HCG

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TABLE 6 A Tentative Proposal for the Optimal Use of Neuroendocrine Tumor Markers in the Diagnosis, Therapeutic Choice, and Follow-up of Patients with Neuroendocrine Tumors Initial diagnosis Endocrine pancreatic tumors Insulinoma

Choice of therapy

72-h fast glucose/insulin ratio ⫹C peptide or proinsulin

(Intraoperative)

Gastrin ⫹ basal/maximal acid output VIP Glucagon Somatostatin

SRS ⫹ endoscopy

Clinically nonfunctioning Carcinoids Foregut Midgut Hindgut

PP, CgA, NSE, SRS

SRS

CgA, NSE, SRS 5 HIAA, CgA, SRS Wide combination, SRS

SRS SRS SRS

Medullary thyroid cancer Pheochromocytoma

CT, CEA, DNA sequence RET oncogene Metanefrine, VMA, CgA, plasma cats. SRS, NSE, catecholamines NSE, CgA

CT, MRI, Ultrasound MIBG

Gastrin

Vipoma Glucagonoma Somatostatinoma

Paraganglioma Small cell lung cancer Pituitary adenomas Prolactinoma Acromegaly Cushing’s disease Clinically nonfunctioning

PRL GH/IGF-I ACTH, dex suppr., UFC, sampling ␣-Subunit, LH, LH␤, and/or FSH

Ultrasound

SRS SRS SRS

SRS

Follow-up

Fasting glucose/insulin ⫹C peptide or proinsulin CgA, gastrin, SRS

VIP, CgA, SRS Glucagon, CgA, SRS Somatostatin, CgA, SRS PP, CgA, NSE, SRS

CgA, NSE, SRS 5-HIAA, CgA, SRS Wide combinations, SRS CT, CEA Metaneph., CgA, cats., MIBG, SRS SRS

Radiological staging (SRS)

NSE, CgA

MRI ⫹ vis fields MRI ⫹ vis fields MRI ⫹ vis fields

PRL GH, IGF-I UFC

MRI ⫹ vis fields

␣-Subunit, LH, LH␤, and/or FSH

Note. Abbreviations used: SRS, somatostatin receptor scintigraphy; CgA, chromogranin A; VIP, vasoactive intestinal peptide; cats., plasma catecholamines; UFC, urinary cortisol; dex. suppr., overnight 1 mg dexamethasone suppression test; sampling, sinus petrosus sampling visual fields: vis fields, visual field determination.

and ␣-subunit expression, which often can also be demonstrated by increased concentrations of these glycopeptides in the circulation (33). In medullary thyroid cancer an increasing expression of CEA often reflects a more aggressive behavior of the tumor. Simultaneously with increasing serum CEA concentrations, a variable stabilization and/or decrease in serum calcitonin levels indicates more malignant behavior.

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LAMBERTS, HOFLAND, AND NOBELS CONCLUSIONS

In a variety of malignancies, including leukemia, prostate cancer, neuroblastoma, and hepatocellular carcinoma, PCR-based detection of tumor- or tissuespecific mRNA species can now be used to detect tumor cells in blood. Such assays can even detect 1 cancer cell among 10 5–10 6 whole blood cells. The detection of recurrent thyroid cancer after total thyreoidectomy can also be facilitated by looking for circulating thyroid cells, using a PCR-based assay for thyroglobulin mRNA. This is a more sensitive marker than an immunoassay for serum thyroglobulin (96). In the diagnosis of NE tumors, however, we are far from such a sophisticated approach. Most NE tumor markers are not specific for a given tumor and circulate under normal conditions in the serum in appreciable concentrations, giving them a low sensitivity for easy detection of small tumors. None of the NE tumor markers discussed fulfil the criteria of high specificity and high sensitivity for screening procedures. The sensitivity of three general tumor markers in the diagnosis of the most common NE tumors is indicated in Table 5. However, by combining hormone measurements with tissue responsiveness, the demonstration of inappropriate secretion of PTH, insulin, and gastrin during hypercalcemia, hypoglycemia, and hyperacidity in the stomach, these markers become highly sensitive and specific diagnostic tests. The application of polyclonal antibodies in radioimmunoassays of a number of hormones such as ACTH, insulin, and gastrin, increases the level of suspicion for the detection of primary NE and non-NE tumors. Finally, the use of CgA RIAs using a combination of C-terminal antibodies is very informative in the initial diagnosis and especially in the follow-up of patients with NE tumors. In Table 6 an overview is presented of the use of optimal tumor markers in the diagnosis of NE tumors, as well as in the follow-up of these patients after treatment. Also, tentative conclusions with regard to the optimal procedure of the visualization of the primary tumor and their often unknown metastases are presented.

REFERENCES 1. 2.

3.

4.

5.

Adrian TE, Uttenthal LO, Williams SJ, Bloom SR. Secretion of pancreatic polypeptide in patients with pancreatic endocrine tumors. N Engl J Med 1986; 315: 287–291. Akoun GM, Scarna HM, Milleron BJ, Benichou MP, Herman DP. Serum neuron-specific enolase. A marker for disease extent and response to therapy for small-cell lung cancer. Chest 1985; 87: 39 – 43. Aprikian AG, Cordon-Cardo C, Fair WR, Reuter VE. Characterization of neuroendocrine differentiation in human benign prostate and prostatic adenocarcinoma. Cancer 1993; 71: 3952–3965. Aprikian AG, Cordon-Cardo C, Fair WR, Zhang ZF, Bazinet M, Hamdy SM, Reuter VE. Neuroendocrine differentiation in metastatic prostatic adenocarcinoma. J Urol 1994; 151: 914 –919. Bajetta E, Ferrari L, Martinetti A, Celio L, Procopio G, Artale S, Zilembo N, Di Bartolomeo M, Seregni E, Bombardieri E. Chromogranin A, neuron specific enolase, carcinoembryonic

NEUROENDOCRINE TUMOR MARKERS

333

antigen, and hydroxyindole acetic acid evaluation in patients with neuroendocrine tumors. Cancer 1999; 86: 858 – 865. 6.

Baldwin GS, Shulkes A. Gastrin, gastrin receptors and colorectal carcinoma. Gut 1998; 42: 581–584.

7.

Baudin E, Bidart JM, Rougier P, Lazar V, Ruffie P, Ropers J, Ducreux M, Troalen F, Sabourin JC, Comoy E, Lasser P, DeBaere T, Schlumberger. Screening for multiple endocrine neoplasia type 1 and hormonal production in apparently sporadic neuroendocrine tumors. J Clin Endocrinol Metab 1999; 84: 69 –75.

8.

Blaschko H, Comline RS, Schneider FH, Silver M, Smith. Secretion of a chromaffin granule protein, chromogranin, from the adrenal gland after splanchnic stimulation. Nature 1967; 215: 58 –59.

9.

Bravo EL. Evolving concepts in the pathophysiology, diagnosis, and treatment of pheochromocytoma. Endocr Rev 1994; 15: 356 –368.

10.

Burchardt T, Burchardt M, Chen MW, Cao Y, de la Taille A, Shabsigh A, Hayek O, Dorai T, Buttyan R. Transdifferentiation of prostate cancer cells to a neuroendocrine cell phenotype in vitro and in vivo. J Urol. 1999; 162: 1800 –1805.

11.

Busnardo B, Girelli ME, Simioni N, Nacamulli D, Busetto R. Nonparallel patterns of calcitonin and carcinoembryonic antigen levels in the follow-up of medullary thyroid carcinoma. Cancer 1984; 53: 278 –285.

12.

Capella C, Heitz PU, Hofler H, Solcia E, Kloppel G. Revised classification of neuroendocrine tumors of the lung, pancreas and gut. Digestion 1994; 55: 11–23.

13.

Cohen R, Campos JM, Salaun C, Heshmati HM, Kraimps JL, Proye C, Sarfati E, Henry JF, Niccoli-Sire P, Modigliani E. Preoperative calcitonin levels are predictive of tumor size and postoperative calcitonin normalization in medullary thyroid carcinoma. Groupe d’Etudes des Tumeurs a Calcitonine (GETC). J Clin Endocrinol Metab 2000; 85: 919 –922.

14.

Cox ME, Deeble PD, Lakhani S, Parsons SJ. Acquisition of neuroendocrine characteristics by prostate tumor cells is reversible: Implications for prostate cancer progression. Cancer Res 1999; 59: 3821–3830.

15.

Cunningham RT, Johnston CF, Irvine GB, Buchanan KD. Serum neuron-specific enolase levels in patients with neuroendocrine and carcinoid tumours. Clin Chim Acta 1992; 212: 123–131.

16.

Cussenot O, Villette JM, Valeri A, Cariou G, Desgrandchamps F, Cortesse A, Meria P, Teillac P, Fiet J, Le Duc A. Plasma neuroendocrine markers in patients with benign prostatic hyperplasia and prostatic carcinoma. J Urol 1996; 155: 1340 –1343.

17.

Daneshdoost L, Gennarelli TA, Bashey HM, Savino PJ, Sergott RC, Bosley TM, Snyder PJ. Recognition of gonadotroph adenomas in women. N Engl J Med 1991; 324: 589 –594.

18.

DeBruine AP, Wiggers T, Beek C, Volovics A, von Meyenfeldt M, Arends JW, Bosman FT. Endocrine cells in colorectal adenocarcinomas: incidence, hormone profile and prognostic relevance. Int J Cancer 1993; 54: 765–771.

19.

Deftos LJ. Chromogranin A: Its role in endocrine function and as an endocrine and neuroendocrine tumor marker. Endocr Rev 1991; 12: 181–187.

20.

Deftos LJ, Nakada S, Burton DW, di Sant’Agnese PA, Cockett AT, Abrahamsson PA. Immunoassay and immunohistology studies of chromogranin A as a neuroendocrine marker in patients with carcinoma of the prostate. Urology 1996; 48: 58 – 62.

21.

Di Sant’Agnese PA. Neuroendocrine differentiation in carcinoma of the prostate. Diagnostic, prognostic and therapeutic implications. Cancer 1992; 70: 254 –268.

22.

Drivsholm L, Paloheimo LI, Osterlind K. Chromogranin A, a significant prognostic factor in small cell lung cancer. Br J Cancer 1999; 81: 667– 671.

23.

Eriksson B, Arnberg H, Oberg K, Hellman U, Lundqvist G, Wernstedt C, Wilander E. A polyclonal antiserum against chromogranin A and B—A new sensitive marker for neuroendocrine tumours. Acta Endocrinol 1990; 122: 145–155.

334

LAMBERTS, HOFLAND, AND NOBELS

24.

Eriksson B, Oberg K. Peptide hormones as tumor markers in neuroendocrine gastrointestinal tumors. Acta Oncol 1991; 30: 477– 483.

25.

Eijck CH van, Lamberts SW, Lemaire LC, Jeekel H, Bosman FT, Reubi JC, Bruining HA, Krenning EP. The use of somatostatin receptor scintigraphy in the differential diagnosis of pancreatic duct cancers and islet cell tumors. Ann Surg 1996; 224: 119 –124.

26.

Feldman JM, O’Dorisio TM. Role of neuropeptides and serotonin in the diagnosis of carcinoid tumors. Am J Med 1986; 81: 41.

27.

Feldman JM. Carcinoid tumors and syndrome. Semin Oncol. 1987; 14: 237–246.

28.

Fishbeyn VA, Norton JA, Benya RV, Pisegna JR, Venzon DJ, Metz DC, Jensen RT. Assessment and prediction of long-term cure in patients with the Zollinger-Ellison syndrome: The best approach. Ann Intern Med 1993; 119: 199 –206.

29.

Gibril F, Reynolds JC, Doppman JL, Chen CC, Venzon DJ, Termanini B, Weber HC, Stewart CA, Jensen RT. Somatostatin receptor scintigraphy: Its sensitivity compared with that of other imaging methods in detecting primary and metastatic gastrinomas. A prospective study. Ann Intern Med 1996; 125: 26 –34.

30.

Gibril F, Reynolds JC, Chen CC, Yu F, Goebel SU, Serrano J, Doppman JL, Jensen RT. Specificity of somatostatin receptor scintigraphy: A prospective study and effects of falsepositive localizations on management in patients with gastrinomas. J Nucl Med 1999; 40: 539 –553.

31.

Goebel SU, Serrano J, Yu F, Gibril F, Venzon DJ, Jensen RT. Prospective study of the value of serum chromogranin A or serum gastrin levels in the assessment of the presence, extent, or growth of gastrinomas. Cancer 1999; 85: 1470 –1483.

32.

Granberg D, Stridsberg M, Seensalu R, Eriksson B, Lundqvist G, Oberg K, Skogseid B. Plasma chromogranin A in patients with multiple endocrine neoplasia type 1. J Clin Endocrinol Metab 1999; 84: 2712–2717.

33.

Grossmann M, Trautmann ME, Poertl S, Hoermann R, Berger P, Arnold R, Mann K. Alpha-subunit and human chorionic gonadotropin-beta immunoreactivity in patients with malignant endocrine gastroenteropancreatic tumours. Eur J Clin Invest 1994; 24: 131–136.

34.

Grouzmann E, Comoy E, Bohuon C. Plasma neuropeptide Y concentrations in patients with neuroendocrine tumors. J Clin Endocrinol Metab 1989; 68: 808 – 813.

35.

Grufferman S, Gillman MW, Pasternak LR, Peterson CL, Young WG Jr. Familial carotid body tumors: Case report and epidemiologic review. Cancer 1980; 46: 2116 –2122.

36.

Hamada Y, Oishi A, Shoji T, Takada H, Yamamura M, Hioki K, Yamamoto. Endocrine cells and prognosis in patients with colorectal carcinoma. Cancer 1992; 69: 2641–2646.

37.

Harris PE. Biochemical markers for clinically non-functioning pituitary tumours. Clin Endocrinol 1998; 49: 163–164.

38.

Harst E van der, Herder WW de, Bruining HO, Bonjer HJ, Krijger RR de, Lamberts SWJ, Meiracker AH van de, Boomsma F, Stijnen T, Krenning EP, Bosman FT, Kwekkeboom DJ. 123 I-MIBG and IIIIn-Octreotide uptake in benign and malignant phaeochromocytomas. J Clin Endocrinol Metab 2001; 86: 685– 694.

39.

Harst E van der, Krijger RR de, Bruining HA, Herder WW de, Bonjer HJ, Lamberts SWJ, Meiracker AH van den, Koper JW, Stijnen T, Bosman FT, Boomsma F. The value of plasma markers for the clinical behaviour of phaeochromocytomas. Clin Endocrinol 2001, in press.

40.

Heitz PhU, Roth J, Zuber C, Komminoth P. Markers for neural and endocrine cells in pathology; In: Gratzl M, Langley L, Eds. Markers for Neural and Endocrine Cells. Weinheim:VCH, 1991: 203–216.

41.

Herder WW de, Uitterlinden P, Pieterman H, Tanghe HL, Kwekkeboom DJ, Pols HA Singh R, van de Berge JH, Lamberts SW. Pituitary tumour localization in patients with Cushing’s disease by magnetic resonance imaging: Is there a place for petrosal sinus sampling? Clin Endocrinol 1994; 40: 87–92.

NEUROENDOCRINE TUMOR MARKERS

335

42.

Herder WW de, Reijs AEM, de Swart J, Kaandorp Y, Lamberts SWJ, Krenning EP, Kwekkeboom DJ. Comparison of iodine-123 epidepride and iodine-123 IBZM for dopamine D2 receptor imaging in clinically non-functioning pituitary macroadenomas and macroprolactinomas. Eur J Nucl Med 1999; 26: 46 –50.

43.

Hirshberg B, Livi A, Bartlett DL, Libutti SK, Alexander HR, Doppman JL, Skarulis MC, Gorden P. Forty-eight-hour fast: The diagnostic test for insulinoma. J Clin Endocrinol Metab 2000; 85: 3222–3226.

44.

Hsiao RJ, Seeger RC, Yu AL, O’Connor DT. Chromogranin A in children with neuroblastoma. Serum concentration parallels disease stage and predicts survival. J Clin Invest 1990; 85: 1555–1559.

45.

Iacangelo AL, Eiden LE. Chromogranin A: Current status as a precursor for bioactive peptides and a granulogenic/sorting factor in the regulated secretory pathway. Regul Pept 1995; 58: 65– 88.

46.

Jacobs EL, Haskell CM. Clinical use of tumor markers in oncology. Curr Probl Cancer 1991; 15: 299 –360.

47.

James C, Starks M, MacGillivray DC, White. The use of imaging studies in the diagnosis and management of thyroid cancer and hyperparathyroidism. Surg Oncol Clin N Am 1999; 8: 145–169.

48.

Johnson PW, Joel SP, Love S, Butcher M, Pandian MR, Squires L, Wrigley PF, Slevin ML. Tumour markers for prediction of survival and monitoring of remission in small cell lung cancer. Br J Cancer 1993; 67: 760 –766.

49.

Jongsma J, Oomen MH, Noordzij MA, Romijn JC, van der Kwast TH, Schroder FH, van Steenbrugge GJ. Androgen-independent growth is induced by neuropeptides in human prostate cancer cell lines. Prostate 2000; 42: 34 – 44.

50.

Kadmon D, Thompson TC, Lynch GR, Scardino PT. Elevated plasma chromogranin-A concentrations in prostatic carcinoma. J Urol 1991; 146: 358 –361.

51.

Kapuscinski M, Shulkes A, Read D, Hardy KJ. Expression of neurotensin in endocrine tumors. J Clin Endocrinol Metab 1990; 70: 100 –106.

52.

Katznelson L, Alexander JM, Klibanski A. Clinical review 45: Clinically nonfunctioning pituitary adenomas. J Clin Endocrinol Metab 1993; 76: 1089 –94.

53.

Koch TR, Michener SR, Go VL. Plasma vasoactive intestinal polypeptide concentration determination in patients with diarrhea. Gastroenterology 1991; 100: 99 –106.

54.

Koper JW, Lamberts SW. Sporadic endocrine tumours and their relationship to the hereditary endocrine neoplasia syndromes. Eur J Clin Invest 2000; 30: 493–500.

55.

Krenning EP, Bakker WH, Breeman WA, Koper JW, Kooij PP, Ausema L, Lameris JS, Reubi JC, Lamberts SW. Localisation of endocrine-related tumours with radioiodinated analogue of somatostatin. Lancet 1989; 1: 242–244.

56.

Krenning EP, Kwekkeboom DJ, Bakker WH, Breeman WA, Kooij PP, Oei HY, van Hagen M, Postema PT, de Jong M, Reubi JC, Visser TJ, Reijs AEM, Hofland LJ, Koper JW, Lamberts SWJ. Somatostatin receptor scintigraphy with [111In-DTPA-D-Phe1]-and [1231-Tyr3]-octreotide: The Rotterdam experience with more than 1000 patients. Eur J Nucl Med 1993; 20: 716 –731.

57.

Krijnen JL, Bogdanowicz JF, Seldenrijk CA, Mulder PG, van der Kwast TH. The prognostic value of neuroendocrine differentiation in adenocarcinoma of the prostate in relation to progression of disease after endocrine therapy. J Urol 1997; 158: 171–174.

58.

Kvols LK, Moertel CG, O’Connell MJ, Schutt AJ, Rubin J, Hahn RG. Treatment of the malignant carcinoid syndrome. Evaluation of a long-acting somatostatin analogue. N Engl J Med 1986; 315: 663– 666.

59.

Kwekkeboom DJ, de Jong FH, Lamberts SW. Gonadotropin release by clinically nonfunctioning and gonadotroph pituitary adenomas in vivo and in vitro: Relation to sex and effects

336

LAMBERTS, HOFLAND, AND NOBELS of thyrotropin-releasing hormone, gonadotropin-releasing hormone, and bromocriptine. J Clin Endocrinol Metab 1989; 68: 1128 –1135.

60.

Kwekkeboom DJ, Reubi JC, Lamberts SW, Bruining HA, Mulder AH, Oei HY, Krenning EP. In vivo somatostatin receptor imaging in medullary thyroid carcinoma. J Clin Endocrinol Metab 1993; 76: 1413–1417.

61.

Kwekkeboom DJ, van Urk H, Pauw BK, Lamberts SW, Kooij PP, Hoogma RP, Krenning EP. Octreotide scintigraphy for the detection of paragangliomas. J Nucl Med 1993; 34: 873– 878.

62.

Kwekkeboom DJ, Krenning EP, Bakker WH, Oei HY, Kooij PP, Lamberts SW. Somatostatin analogue scintigraphy in carcinoid tumours. Eur J Nucl Med 1993; 20: 283–292.

63.

Kwekkeboom D, Krenning EP, de Jong. Peptide receptor imaging and therapy. J Nucl Med 2000; 41: 1704 –1713.

64.

Lamberts SW, Uitterlinden P, Verschoor L, van Dongen KJ, del Pozo E. Long-term treatment of acromegaly with the somatostatin analogue SMS 201-995. N Engl J Med 1985; 313: 1576 –1580.

65.

Lamberts SW, Bakker WH, Reubi JC, Krenning EP. Somatostatin-receptor imaging in the localization of endocrine tumors. N Engl J Med 1990; 323: 1246 –1249.

66.

Lamberts SW, Hofland LJ, van Koetsveld PM, Reubi JC, Bruining HA, Bakker WH, Krenning EP. Parallel in vivo and in vitro detection of functional somatostatin receptors in human endocrine pancreatic tumors: Consequences with regard to diagnosis, localization, and therapy. J Clin Endocrinol Metab 1990; 71: 566 –574.

67.

Lamberts SW, Krenning EP, Reubi. The role of somatostatin and its analogs in the diagnosis and treatment of tumors. Endocr Rev 1991; 12: 450 – 482.

68.

Lamberts SW, van der Lely AJ, de Herder WW, Hofland LJ. Octreotide. N Engl J Med 1996; 334: 246 –254.

69.

Lindblom A, Liljegren A. Regular review: Tumour markers in malignancies. Br Med 2000; 320: 424 – 427.

70.

Lips CJ, Landsvater RM, Hoppener JW, Geerdink RA, Blijham G, van Veen JM, van Gils AP, de Wit MJ, Zewald RA, Berends MJ. Clinical screening as compared with DNA analysis in families with multiple endocrine neoplasia type 2A. N Engl J Med 1994; 331: 828 – 835.

71.

Lucas KG, Feldman JM. Flushing in the carcinoid syndrome and plasma kalikrein. Cancer 1986; 58: 2290.

72.

Mendelsohn G, Wells SA Jr, Baylin SB. Relationship of tissue carcinoembryonic antigen and calcitonin to tumor virulence in medullary thyroid carcinoma. An immunohistochemical study in early, localized, and virulent disseminated stages of disease. Cancer 1984; 54: 657– 662.

73.

Mitchell BK, Merrell RC, Kinder BK. Localization studies in patients with hyperparathyroidism. Surg Clin North Am 1995; 75: 483– 498.

74.

Mutch MG, Frisella MM, DeBenedetti MK, Doherty GM, Norton JA, Wells SA Jr, Lairmore TC. Pancreatic polypeptide is a useful plasma marker for radiographically evident pancreatic islet cell tumors in patients with multiple endocrine neoplasia type 1. Surgery 1997; 122: 1012–1019.

75.

Nobels FR, Kwekkeboom DJ, Coopmans W, Hoekstra R, De Herder WW, Bouillon R, Lamberts SW. A comparison between the diagnostic value of gonadotropins, alpha-subunit, chromogranin-A and their response to thyrotropin-releasing hormone in clinically nonfunctioning, alpha-subunit-secreting, and gonadotroph pituitary adenomas. J Clin Endocrinol Metab 1993; 77: 784 –789.

76.

Nobels FR, de Herder WW, Kwekkeboom DJ, Coopmans W, Mulder A, Bouillon R, Lamberts SW. Serum chromogranin A in the differential diagnosis of Cushing’s syndrome. Eur J Endocrinol 1994; 131: 589 –593.

77.

Nobels FR, Kwekkeboom DJ, Coopmans W, Schoenmakers CH, Lindemans J, De Herder WW, Krenning EP, Bouillon R, Lamberts SW. Chromogranin A as serum marker for

NEUROENDOCRINE TUMOR MARKERS

337

neuroendocrine neoplasia: comparison with neuron-specific enolase and the alpha-subunit of glycoprotein hormones. J Clin Endocrinol Metab 1997; 82: 2622–2628. 78.

Nobels FR, Kwekkeboom DJ, Bouillon R, Lamberts SW. Chromogranin A: Its clinical value as marker of neuroendocrine tumours. Eur J Clin Invest 1998; 28: 431– 440.

79.

Norheim I, Oberg K, Theodorsson-Norheim E, Lindgren PG, Lundqvist G, Magnusson A, Wide L, Wilander E. Malignant carcinoid tumors. An analysis of 103 patients with regard to tumor hormone production, and survival. Ann Surg 1987; 206: 115–125.

80.

O’Connor DT, Deftos LJ. Secretion of chromogranin A by peptide-producing endocrine neoplasms. N Engl J Med 1986; 314: 1145–1151.

81.

O’Connor DT, Pandlan MR, Carlton E, Cervenka JH, Hsiao. Rapid radioimmunoassay of circulating chromogranin A: In vitro stability, exploration of the neuroendocrine character of neoplasia, and assessment of the effects of organ failure. Clin Chem 1989; 35: 1631–1637.

82.

Odell WD. Endocrine/metabolic syndromes of cancer. Semin Oncol 1997; 24: 299 –317.

83.

Pacini F, Fontanelli M, Fugazzola L, Elisei R, Romei C, Di Coscio G, Miccoli P, Pinchera A. Routine measurement of serum calcitonin in nodular thyroid diseases allows thepreoperative diagnosis of unsuspected sporadic medullary thyroid carcinoma. J Clin Endocrinol Metab 1994; 78: 826 – 829.

84.

Pauker SG, Kopelman RI. Interpreting hoofbeats: Can Bayes help clear the haze? N Engl J Med 1992; 327: 1009 –1013.

85.

Pour PM, Permert J, Mogaki M, Fujii H, Kazakoff K. Endocrine aspects of exocrine cancer of the pancreas. Their patterns and suggested biologic significance. Am J Clin Pathol 1993; 100: 223–230.

86.

Pernow B, Waldenstrom J. Determination of 5-hydroxytryptamine, 5-hydroxy indoloacetic acid and histamine in 33 cases of carcinoid tumor (argentaffinoma). Am J Med 1957; 23: 16 –25.

87.

Polak JM, Adrian TE, Bryant MG, Bloom SR, Heitz PH, Pearse AGE. Pancreatic polypeptide in insulinoma, gastrinomas, vipomas and glucogonomas. Lancet 1978; 1: 328 –330.

88.

Polak JM. Diagnostic Histopathology of Neuroendocrine Tumours. Edinburgh: ChurchillLivingstone, 1993.

89.

Postema PT, Krenning EP, Wijngaarde R, Kooy PP, Oei HY, van den Bosch WA, Reubi JC, Wiersinga WM, Hooijkaas H, van der Loos T. [111ln-DTPA-D-Phe1] Octreotide scintigraphy in thyroidal and orbital Graves’ disease: A parameter for disease activity? J Clin Endocrinol Metab 1994; 79: 1845–1851.

90.

Prinz RA, Bermes EW Jr, Kimmel JR, Marangos PJ. Serum markers for pancreatic islet cell and intestinal carcinoid tumors: A comparison of neuron-specific enolase, beta-human chorionic gonadotropin and pancreatic polypeptide. Surgery 1983; 94: 1019 –1023.

91.

Reubi JC. The role of peptides and their receptors as tumor markers. Endocrinol Metab Clin North Am 1993; 22: 917–939.

92.

Reubi JC, Maurer R, Klijn JG, Stefanko SZ, Foekens JA, Blaauw G, Blankenstein MA, Lamberts SW. High incidence of somatostatin receptors in human meningiomas: Biochemical characterization. J Clin Endocrinol Metab 1986; 63: 433– 438.

93.

Reubi JC, Lang W, Maurer R, Koper JW, Lamberts SW. Distribution and biochemical characterization of somatostatin receptors in tumors of the human central nervous system. Cancer Res 1987; 47: 5758 –5764.

94.

Reubi JC, Horisberger U, Lang W, Koper JW, Braakman R, Lamberts SW. Coincidence of EGF receptors and somatostatin receptors in meningiomas but inverse, differentiationdependent relationship in glial tumors. Am J Pathol 1989; 134: 337–344.

95.

Reubi JC, Horisberger U, Studer UE, Waser B, Laissue JA. Human kidney as target for somatostatin: High affinity receptors in tubules and vasa recta. J Clin Endocrinol Metab 1993; 77: 1323–1328.

338

LAMBERTS, HOFLAND, AND NOBELS

96.

Ringel MD, Ladenson PW, Levine MA. Molecular diagnosis of residual and recurrent thyroid cancer by amplification of thyroglobulin messenger ribonucleic acid in peripheral blood. J Clin Endocrinol Metab 1998; 83: 4435– 4442.

97.

Rosa P, Gerdes HH. The granin protein family: Markers for neuroendocrine cells and tools for the diagnosis of neuroendocrine tumors. J Endocrinol Invest 1994; 17: 207–225.

98.

Sandler M, Karim SMM, William ED. Prostaglandin in amine-peptide-secreting tumors. Lancet 1968; 2: 1053.

99.

Scammell JG. Granins. Markers of the regulated secretory pathway. Trends Endocrinol Metab 1993; 4: 14 –18.

100.

Schmechel D, Marangos PJ, Brightman M. Neurone-specific enolase is a molecular marker for peripheral and central neuroendocrine cells. Nature 1978; 276: 834 – 836.

101.

Scopsi L, Andreola S, Pilotti S, Testori A, Baldini MT, Leoni F, Lombardi L, Hutton JC, Shimizu F, Rosa P. Argyrophilia and granin (chromogranin/secretogranin) expression in female breast carcinomas. Their relationship to survival and other disease parameters. Am J Surg Pathol 1992; 16: 561–576.

102.

Service FJ. Hypoglycemic disorders. N Engl J Med 1995; 332: 1144 –52.

103.

Skogseid B, Oberg K, Benson L, Lindgren PG, Lorelius LE, Lundquist G, Wide L, Wilander E. A standardized meal stimulation test of the endocrine pancreas for early detection of pancreatic endocrine tumors in multiple endocrine neoplasia type 1 syndrome: Five years experience. J Clin Endocrinol Metab 1987; 64: 1233–1240.

104.

Sobol RE, O’Connor DT, Addison J, Suchocki K, Royston I, Deftos Elevated serum chromogranin A concentrations in small-cell lung carcinoma. Ann Intern Med 1986; 105: 698 –700.

105.

Stewart PM, Gibson S, Crosby SR, Penn R, Holder R, Ferry D, Thatcher N, Phillips P, London DR, White A. ACTH precursors characterize the ectopic ACTH syndrome. Clin Endocrinol 1994; 40: 199 –204.

106.

Sundaresan V, Reeve JG, Stenning S, Stewart S, Bleehen NM. Neuroendocrine differentiation and clinical behaviour in non-small cell lung tumours. Br J Cancer 1991; 64: 333–338.

107.

Syversen U, Halvorsen T, Marvik R, Waldum HL. Neuroendocrine differentiation in colorectal carcinomas. Eur J Gastroenterol Hepatol 1995; 7: 667– 674.

108.

Tabarin A, Valli N, Chanson P, Bachelot Y, Rohmer V, Bex-Bachellerie V, Catargi B, Roger P, Laurent F. Usefulness of somatostatin receptor scintigraphy in patients with occult ectopic adrenocorticotropin syndrome. J Clin Endocrinol Metab 1999; 84: 1193–1202.

109.

Termanini B, Gibril F, Reynolds JC, Doppman JL, Chen CC, Stewart CA, Sutliff VE, Jensen RT. Value of somatostatin receptor scintigraphy: A prospective study in gastrinoma of its on clinical management. Gastroenterology 1997; 112: 335–347.

110.

Theodorsson E. Regulatory peptides as tumour markers. Acta Oncol 1989; 28: 319 –324.

111.

Torpy DJ, Chen CC, Mullen N, Doppman JL, Carrasquillo JA, Chrousos GP, Niema LK. Lack of utility of (111)In-pentetreotide scintigraphy in localizing ectopic ACTH producing tumors: Follow-up of 18 patients. J Clin Endocrinol Metab 1999; 84: 1186 –1192.

112.

Vanhagen PM, Krenning EP, Reubi JC, Kwekkeboom DJ, Bakker WH, Mulder AH, Laissue I, Hoogstede HC, Lamberts SW. Somatostatin analogue scintigraphy in granulomatous diseases. Eur J Nucl Med 1994; 21: 497–502.

113.

Vinik AI, Gonin J, England BG, Jackson T, McLeod MK. Cho Plasma substance-P in neuroendocrine tumors and idiopathic flushing: The value of pentagastrin stimulation tests and the effects of somatostatin analog. J Clin Endocrinol Metab 1990; 70: 1702–1709.

114.

Wassberg E, Stridsberg M, Christofferson R. Plasma levels of chromogranin A are directly proportional to tumour burden in neuroblastoma. J Endocrinol 1996; 151: 225–230.

115.

White A, Gibson S. ACTH precursors: Biological significance and clinical relevance. Clin Endocrinol 1998; 48: 251–255.

116.

Wick MR. Neuroendocrine neoplasia. Am J Clin Pathol 2000; 113: 331–335.

NEUROENDOCRINE TUMOR MARKERS 117.

339

Wiedenmann B, Huttner WB. Synaptophysin and chromogranins/secretogranins—Widespread constituents of distinct types of neuroendocrine vesicles and new tools in tumor diagnosis. Virchows Arch B Cell Pathol Incl Mol Pathol 1989; 58: 95–121. 118. Wilson HE, White A. Prohormones: Their clinical relevance. Trends Endocrinol Metab 1998; 9: 396 – 402.