Somatostatin receptor scintigraphy versus chromogranin A assay in the management of patients with neuroendocrine tumors of different types: clinical role

M. Cimitan1,+, A. Buonadonna2, R. Cannizzaro3, V. Canzonieri4, E. Borsatti1, R. Ruffo1 and L. De Apollonia5

1 Nuclear Medicine Unit, 2 Medical Oncology B Division, 3 Gastroenterology Unit, 4 Pathology Division, 5 Blood Transfusion, Immunohematology and Clinical Chemistry Unit, National Cancer Institute (CRO–IRCCS), Aviano, Italy

Received 18 February 2003; revised 27 February 2003; accepted 10 March 2003


    Abstract
 Top
 Abstract
 Introduction
 Patients and methods
 Results
 Discussion
 Conclusions
 References
 
Background:

Current diagnosis and staging of neuroendocrine tumors (NETs) are significantly improved by the introduction of the chromogranin A (CgA) assay in plasma or serum as a tumor marker, and by the use of somatostatin receptor scintigraphy (SRS) for tumor localization. However, the clinical role of CgA assay compared with SRS in the management of NETs has not been well elucidated.

Patients and methods:

Sixty-three consecutive patients with a histological diagnosis of NET underwent plasma CgA assay and SRS for tumor staging (23 cases), evaluation of tumor response (18 cases) and evaluation of tumor recurrence on follow-up (22 cases). Twenty-one patients had well-differentiated neuroendocrine tumors (WDNETs: 18 gastroenteropancreatic tumors and three lung NETs); 22 patients had well-differentiated neuroendocrine carcinomas (WDNECs: 17 gastroenteropancreatic carcinomas, two lung neuroendocrine carcinomas and three neuroendocrine carcinomas of unknown origin) and 20 patients had poorly differentiated neuroendocrine carcinomas (PDNECs: 14 extra-pulmonary small-cell carcinomas and six Merkel cell carcinomas). Almost all (58 of 63) NETs were non-functioning. The quantitative determination of CgA was performed in plasma using an enzyme immunoassay with a cut-off value fixed at 34 U/l. Scintigraphies with indium 111-DTPA-octreotide (111In-pentetreotide) included whole-body images and single photon emission computed tomography (SPECT) scans of the chest and abdomen.

Results:

SRS results were compared with CgA findings and final clinical data. The overall sensitivity of SRS and CgA, based on the final clinical data, was 77% and 55%, respectively, whereas the specificity of both SRS and CgA was 94%. Concerning tumor type, SRS accuracy was 95% for WDNETs, 86% for WDNECs and 60% for PDNECs; CgA accuracy was 76% for WDNETs, 68% for WDNECs and 50% for PDNECs. With regard to disease extent, SRS sensitivity was 100% for limited disease and 72% for advanced disease; CgA sensitivity was 43% for limited disease and 57% for advanced disease.

Conclusions:

In our NET series, SRS proved to be more sensitive than CgA, with equivalent specificity. Tumor differentiation influences the sensitivity of SRS and CgA analysis. In addition, the plasma CgA level is related to tumor secretory activity. Nevertheless both SRS and CgA should be considered useful tools in the diagnostic work-up of NET patients.

Key words: chromogranin A, 111In-pentetreotide, neuroendocrine tumors, somatostatin receptor scintigraphy


    Introduction
 Top
 Abstract
 Introduction
 Patients and methods
 Results
 Discussion
 Conclusions
 References
 
Neuroendocrine tumors (NETs) represent a heterogeneous group of neoplastic entities derived from cell populations of the diffuse neuroendocrine system; they encompass aggressive tumors such as neuroendocrine small-cell cancer, low-growing tumors such as carcinoid and tumors with intermediate aggressiveness such as malignant carcinoid [1, 2]. They are usually divided into functioning or non-functioning tumors according to their ability to produce, or not, hormones and biologically active substances responsible for clinical syndromes. Although these tumors often have a good prognosis, they are generally diagnosed late because of their versatile presentation. Current diagnosis and staging of NETs are significantly improved by the introduction of the chromogranin A (CgA) assay in plasma or serum, as a tumor marker [36], and by the use of somatostatin receptor scintigraphy (SRS) for tumor localization [711]. Human CgA is a glycoprotein belonging to the family of chromogranins, also known as secretogranins, which is present in the secretory granules storing peptide hormones and catecholamine throughout the neuroendocrine system. Elevated plasma CgA levels have been found in a variety (80–100%) of functional and non-functional NETs and the circulating CgA levels also reflect the tumor extent [12, 13].

SRS using indium 111-DTPA-octreotide (111In-pentetreotide) is widely used to localize NET since octreotide is able to bind to somatostatin receptor subtypes 2 and 5, and almost all (80%) NETs contain the somatostatin receptor subtype 2 [7, 14, 15].

In a previous study, Kalkner et al. [16] found that positive 111In-pentetreotide scintigraphy correlates with elevated levels of plasma CgA in patients with carcinoid tumors. However, the role of CgA assay compared with SRS has not been well elucidated. CgA production seems to depend on tumor type or tumor differentiation and the sensitivity of CgA assay as a tumor marker may be low in moderately or poorly differentiated neuroendocrine carcinomas [13]. On the other hand, the sensitivity of 111In-pentetreotide scintigraphy may be of limited value in tumors <0.5 cm and in tumors lacking somatostatin receptor subtype 2 and 5 expression [14]. The aim of our study was to investigate the impact of SRS versus plasma CgA assay in patients with different types of NETs histologically diagnosed according to the new World Health Organization (WHO) clinicopathological formulation. We specifically compared the results of SRS and those of plasma CgA with regard to tumor staging, tumor response evaluation and follow-up.


    Patients and methods
 Top
 Abstract
 Introduction
 Patients and methods
 Results
 Discussion
 Conclusions
 References
 
Patients
Sixty-three consecutive patients (32 men and 31 women, aged between 18 and 76 years) with a histological diagnosis of NETs underwent plasma CgA analysis and SRS for tumor staging (23 cases), evaluation of tumor response to chemotherapy and/or somatostatin analogs (18 cases) and evaluation of tumor recurrence on follow-up (22 cases). All patients gave informed consent to participation in the study. According to the WHO pathological formulation, the histological diagnosis included 21 well-differentiated neuroendocrine tumors (WDNETs: 18 gastroenteropancreatic tumors and three lung NETs), 22 well-differentiated neuroendocrine carcinomas (WDNECs: 17 gastroenteropancreatic carcinomas, two lung neuroendocrine carcinomas and three neuroendocrine carcinomas of unknown origin), and 20 poorly differentiated neuroendocrine carcinomas (PDNECs: 14 extra-pulmonary small-cell carcinomas and six Merkel cell carcinomas). Patients with small-cell lung carcinoma, in whom spiral computed tomography (CT) is generally used as sensitive diagnostic procedure, were excluded from the study.

With regard to disease extent, the disease was defined as ‘limited disease’ (a tumor <2.5 cm or a tumor >2.5 cm without lymph-node metastases), ‘locally advanced disease’ (a tumor with lymph-node metastases) and ‘metastatic disease’. Among the 63 patients investigated, only five (8%) were syndromic.

Immunoassay
The quantitative determination of CgA was performed in plasma using an enzyme immunoassay (DAKO Chromogranin A ELISA Kit; DAKO A/S Produktionsvej 42, DK-2600 Glostrup, Denmark). It is a simplified double antibody sandwich assay where samples and peroxidase-conjugated antibody to CgA are incubated simultaneously in microwells coated with antibody to CgA. After a washing step, the chromogenic substrate is added. Color development results from the oxidation of the substrate through an enzyme-catalyzed reaction. The assay uses rabbit antibodies to a 23 kDa C-terminal fragment of human CgA. CgA concentrations determined with the kit are given as U/l. Total coefficients of variation (within and between assay) are 7.3% for mean levels of 30.5 U/l (n = 36), 8.6% for mean levels of 195.4 U/l (n = 30) and 5.8% for mean levels of 364.4 U/l (n = 30). In our laboratory, the normal CgA values obtained in healthy subjects (58 men aged 20–78 years and 57 women aged 20–75 years) ranged between 0 and 33.39 U/l (mean value ± standard deviation = 10.41 ± 7.66 U/l) with a cut-off value fixed at 34 U/l.

Somatostatin receptor scintigraphy
SRS was performed with 111In-pentetreotide, a [111In-DTPA-D-Phe-] conjugate of octreotide, a somatostatin analog, which binds to somatostatin receptor subtypes 2 and 5 (OctreoScan; Mallinckrodt Medical, Petten, The Netherlands). Planar or whole-body (WB) images, as well as single photon emission computed tomography (SPECT) of chest and abdomen were performed in all patients 4–6 h and 24 h after intravenous (i.v.) injection of 200–220 MBq of 111In-pentetreotide (seldom continued for 48–72 h), using a rectangular large field of view gamma camera (Sophicamera DSX, Sopha; BUC, France). Planar images were acquired for 15–20 min/image, using a 256 x 256 word matrix; WB images, anterior and posterior, were acquired into a 2048 x 512 word matrix with a speed of 8 cm/min. All SPECT studies were performed using a 128 x 128 matrix, 64 projections with a rotation of 360° and 44–55 s acquisition time per projection, and SPECT row data were prefiltered using a Butterworth filter (order 8 and cut-off 0.15). The data were then reconstructed by a maximum activity projection algorithm using a ramp filter without attenuation correction.

Data interpretation
All patients with plasma CgA values >34 U/l were considered positive for neuroendocrine disease. Plasma CgA assays were obtained within 1–4 weeks of scintigraphy in all cases.

SRS was interpreted independently of the result of any prior investigation. To define the tumor as visualized during SRS, a simple yes-or-no system was used. Any focal activity higher than the surrounding background activity was considered a positive result if it was present on the images of both early 4–6 h and delayed 24–48 h studies.

SRS and CgA results were rated true-positive (TP) or true-negative (TN) based on the final data of surgery (n = 39) and/or of other diagnostic procedures such as spiral CT, magnetic resonance, ultrasonography and endoscopy (n = 24).


    Results
 Top
 Abstract
 Introduction
 Patients and methods
 Results
 Discussion
 Conclusions
 References
 
Clinical data, SRS results and plasma CgA values of all 63 patients are summarized in Table 1. At initial diagnosis (before SRS and plasma CgA investigations), 21 (33%) patients had limited disease, eight (13%) locally advanced disease and 34 (54%) metastatic disease. Fourteen of 21 patients with limited disease, and two of eight patients with locally advanced disease at initial diagnosis, were disease-free based on the final clinical data.


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Table 1. Clinical data, somatostatin receptor scintigraphy (SRS) results and chromogranin A (CgA) levels
 
Of the 63 patients examined, 37 (59%) had scintigraphic lesions and 27 (43%) had plasma CgA levels above the normal limit (>34 U/l).

Concordance between SRS and CgA results
For all patients, the concordance between SRS and CgA results was 75%; 24 patients had positive scan findings and elevated CgA levels, and 23 had negative scan findings and CgA levels in the normal range (Table 2). Concerning tumor type, SRS and CgA results agreed in 15 (71%) of the 21 WDNETs, in 16 (73%) of the 22 WDNECs and in 16 (80%) of the 20 PDNECs. Discrepancies, such as positive SRS with normal CgA levels, were noted in six of 21 cases of WDNETs, in four of 22 cases of WDNECs and in three of 20 cases of PDNECs, whereas negative SRS with high CgA levels was seen in only two cases of WDNECs and in one case of PDNECs. Concordant results between SRS and CgA were seen in 74% of patients studied at staging, in 78% of patients evaluated during treatment and in 77% of patients studied on follow-up.


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Table 2. Concordance between somatostatin receptor scintigraphy (SRS) results and plasma chromogranin A (CgA) levels
 
Overall results for SRS and CgA assay based on final clinical data
We found one false-positive and 11 false-negative SRS results based on the final clinical data, as well as one false-positive and 21 false-negative CgA analyses (Table 3). The overall sensitivity (TP/TP + FN) of SRS and CgA was 77% and 55%, respectively; the specificity (TN/TN + FP) of both SRS and CgA was 94%. With regard to the histological classification, SRS sensitivity was 100% for WDNETs, 84% for WDNECs and 50% for PDNECs, whereas CgA sensitivity was 58% for WDNETs, 68% for WDNECs and 37% for PDNECs.


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Table 3. Overall results for somatostatin receptor scintigraphy (SRS) and plasma chromogranin A (CgA) assay based on the final clinical data
 
With regard to disease extent, SRS sensitivity was 100% for limited disease and 72% for_advanced disease (locally advanced disease and metastatic disease); CgA sensitivity was 43% for limited disease and 57% for advanced disease.

On the whole, SRS accuracy (TP + TN/TP + TN + FP + FN) was 81% and CgA accuracy was 65%. SRS provided additional detection sites compared with conventional imaging in only one case.


    Discussion
 Top
 Abstract
 Introduction
 Patients and methods
 Results
 Discussion
 Conclusions
 References
 
In this study, we have investigated the clinical role of SRS and circulating CgA in the management of patients with NETs with respect to stage and type of neoplasm according to the revised WHO clinicopathological classification [1, 2, 17].

We found that positive SRS correlated with elevated levels of plasma CgA, as reported by Kalkner et al. [16]; however, SRS proved to be more sensitive than CgA with equivalent specificity (Figure 1).



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Figure 1. A man aged 48 years at post-surgical staging for carcinoid tumor in the blind gut. Chromogranin A levels were in the normal range (13 U/l). Conversely, somatostatin receptor scintigraphy showed a focal activity in the abdomen (arrow), on early (A) and delayed (B) planar images. Three-dimensional single photon emission computed tomography imaging (C, D) was useful in localizing the tumor site which was a lymph-node metastasis on surgical exploration, not seen by other imaging procedures.

 
In previous reports, the specificity of SRS (expressed on a per patient basis) for NETs of the gastroenteropancreatic tract was 81% [18], and the specificity of circulating CgA ranged between 68% and 95% depending on the upper threshold of the normal range [19, 20]. The sensitivity of SRS in any NETs varied between 78% and 100% [7, 1518], whereas CgA levels depended on tumor type or disease extent and presence of functioning tumors [13, 1921].

Our results showed an overall specificity of both SRS and CgA for any NETs as high as 94%. We observed only one false-positive SRS result in a patient with limited disease, studied on follow-up (Table 1). Similarly, only one false-positive CgA result was registered in a patient with WDNEC suffering from essential hypertension and arrhythmia (Table 1). False-positive SRS results could be limited by having a thorough knowledge of other diseases or circumstances that can result in a false-positive response, as suggested by Gibril et al. [22], and the accurate definition of the normal range of CgA values obtained in our healthy control group (n = 115) has increased the CgA specificity [19].

In our study, the overall sensitivity of SRS (expressed on a per patient basis) was 77%. Moreover, SRS sensitivity was higher (100%) in the WDNET subgroup than in WDNECs (84%) and PDNECs (50%). The sensitivity of SRS was also higher (100%) in patients with limited disease than in patients with advanced disease (72%). This indicates that somatostatin receptor expression is more likely present in highly differentiated and low aggressive NETs.

Conversely, a low overall CgA sensitivity of 55% was found. It is interesting to note that in our patient series, almost all (58 of 63) NETs were non-functioning, and a low CgA sensitivity was observed both in patients with limited disease (43%) and in patients with advanced disease (57%). However, our data showed that the accuracy of CgA was more elevated in patients with low-growing NETs (76%) than in patients with intermediate aggressive neuroendocrine carcinomas (68%) or with poorly differentiated neuroendocrine carcinomas (50%), as reported in the literature [20, 23].


    Conclusions
 Top
 Abstract
 Introduction
 Patients and methods
 Results
 Discussion
 Conclusions
 References
 
In our NET series, which included many cases of highly aggressive (20 cases) and intermediate aggressive (22 cases) tumors, SRS showed a better sensitivity than CgA with equivalent high specificity. Our results confirm that tumor differentiation influences the sensitivity of SRS and CgA analysis. In addition, plasma CgA level is related to tumor secretory activity. Nevertheless, both SRS and CgA should be considered useful tools for the management of NET patients.


    Footnotes
 
+ Correspondence to: Dr M. Cimitan, Nuclear Medicine Unit, National Cancer Institute (CRO–IRCCS), Via Pedemontana Occidentale 12, I-33081 Aviano (PN), Italy. Tel: +39-0434-659337; Fax: +39-0434-652182; E-mail: mcimitan{at}cro.it Back


    References
 Top
 Abstract
 Introduction
 Patients and methods
 Results
 Discussion
 Conclusions
 References
 
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