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Dipartimento di Medicina Sperimentale e Clinica (G.C.), Università Magna Græcia, 88100 Catanzaro, Italy; Unità Operativa Semplice Dipartimentale di Endocrinologia (D.M., D.B.), Ospedale di Bentivoglio, 40010 Bologna, Italy; Dipartimento di Scienze Cliniche (C.D., M.M., S.F.), Università degli Studi di Roma La Sapienza, 00161 Rome, Italy; Divisione di Medicina Interna (M.N.), Ospedale S. Giovanni di Dio, 88900 Crotone, Italy; Dipartimento di Scienze Biomediche (S.T.), Università di Catania, 95123 Catania, Italy; Divisione di Endocrinologia (U.C., M.T.), Istituto di Ricovero e Cura a Carattere Scientifico, Casa Sollievo della Sofferenza, 71013 San Giovanni Rotondo, Italy; and Divisione di Endocrinologia (M.A.), Ospedale Cervello, 90146 Palermo, Italy
Address all correspondence and requests for reprints to: Sebastiano Filetti, M.D., 2a Clinica Medica, Dipartimento di Scienze Cliniche, Università degli Studi di Roma La Sapienza, Viale le del Policlinico, 155, 00161 Rome, Italy. E-mail: sebastiano.filetti{at}uniroma1.it.
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Abstract |
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Objective: The objective of this study was to evaluate the diagnostic accuracy of systematic CT measurement in non-multiple endocrine neoplasia type 2 patients with nodular thyroid disease.
Settings: This study was conducted at a national healthcare system hospital (outpatient and inpatient sectors).
Subjects: Consecutive patients with nodular thyroid disease (n = 5817) were studied.
Main Outcome Measures: Serum CT levels were measured under basal conditions, and when basal values were more than or equal to 20 and less than 100 pg/ml, testing was repeated after pentagastrin stimulation. Basal or stimulated levels more than 100 pg/ml were indication for surgery.
Results: Fifteen cases of MTC and seven of C cell hyperplasia (CCH) were identified. MTCs were diagnosed in all patients with basal CT more than 100 pg/ml. The four patients with basal CT more than or equal to 50 and less than 100 pg/ml included two diagnosed with MTC and two with CCH. In 10 patients with basal levels more than or equal to 20 and less than 50 pg/ml, histology confirmed the presence of MTC in four, four others had CCH, and the remaining two were negative for thyroid malignancy. Positive predictive values for basal CT levels in the preoperative diagnosis of MTC were: 23.1% for values more than or equal to 20 pg/ml, 100% for values more than 100 pg/ml, 25% for levels more than or equal to 50 and less than 100 pg/ml, and 8.3% for values more than or equal to 20 and less than 50 pg/ml. Positive predictive values for the pentagastrin test (>100 pg/ml) were 40% in the entire series.
Conclusions: CT screening of thyroid nodules is a highly sensitive test for early diagnosis of MTC, but confirmatory stimulation testing is necessary in most cases to identify true positive increases.
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Introduction |
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TopAbstractIntroductionPatients and MethodsResultsDiscussionReferences |
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There is evidence that routine measurement of serum CT in patients with thyroid nodules can result in earlier detection of MTC and also C cell hyperplasia (CCH). The latter is considered a preneoplastic lesion in familial forms of MTC (2), but it has never been demonstrated that sporadic forms of CCH undergo malignant transformation. Several studies have been conducted to assess the value of this type of screening (6, 7, 8, 9, 10, 11, 12, 13), but the issue remains controversial (14, 15, 16). The American Thyroid Association has declined to recommend for or against this practice, citing among other things the unresolved issue of cost-effectiveness. They also note that, in most studies, the specificity of CT screening has been increased by confirmatory testing after stimulation with pentagastrin (PG), which is no longer marketed in the United States (17). In contrast, in the European Thyroid Association’s recently published consensus statement, serum CT measurement is recommended in the initial diagnostic evaluation of thyroid nodules (18), although no indications are offered regarding assay methods, interpretation of results, or use of stimulation testing.
We recently evaluated the diagnostic accuracy of a protocol based on serum CT measurement for preoperative identification of CCH or MTC in a large sample of 5817 patients with thyroid nodular disease. The strategy tested involves first level screening of basal CT levels and confirmatory retesting of suspicious cases after PG stimulation.
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Patients and Methods |
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The study population included a total of 5817 patients (1111 males, 4706 females; age range, 11–72 yr; mean age, 49.7 ± 16.6 yr) consecutively diagnosed with thyroid nodules between January 1, 2001 and December 31, 2004. Each provided written informed consent to all study procedures. The majority came from areas of Italy classified as mildly to moderately iodine-deficient (19, 20). Exclusion criteria were renal failure, persistent or recurrent MTC, or a family history of MTC. Based on the results of thyroid sonography, circulating levels of free thyroid hormones and TSH, and titers of thyroperoxidase and thyroglobulin autoantibodies (21), clinical diagnoses were: euthyroid nodular/multinodular goiter (n = 4894), Hashimoto’s thyroiditis with nodules (n = 436), autonomously functioning thyroid nodules (n = 276), and toxic nodular goiter (n = 211). Thyroid scans were performed in patients with toxic nodular or multinodular goiter and those suspected of having autonomously functioning thyroid nodules. Fine-needle aspiration biopsy (FNAB) was proposed for all nonautonomous nodules exceeding 10 mm in diameter; in patients with multinodular goiters, only the largest palpable nodule was biopsied. The pathologist examining the specimens was not aware of the patient’s serum CT values.
Serum CT assay
Serum CT was measured with a commercially available chemiluminescence assay (Nichols Advantage Calcitonin Chemiluminescence assay, San Juan Capistrano, CA) with analytical sensitivity of 1 pg/ml. The upper limit of the normal range was 10 pg/ml. Intraassay coefficients of variation determined by the manufacturer using serial dilutions of pooled human serum were: 6.2% at 12.1 pg/ml, 3.5% at 95.5 pg/ml, and 3.5% at 544 pg/ml. Interassay coefficients of variation were 8.8% at 10.5 pg/ml, 7.3% at 85.7 pg/ml, and 4.5% at 527 pg/ml. In assays of synthetic CT, a hook effect has been observed at more than 500,000 pg/ml.
Clinical protocol
Basal serum CT levels were measured in all 5817 patients upon enrollment, and the results were classified as follows. 1) Levels less than or equal to 10 pg/ml were considered negative for MTC or CCH. 2) Levels more than 10 and less than 20 pg/ml were considered indeterminate, and patients in this gray zone underwent longitudinal follow-up, which included yearly measurement of basal CT and FNAB at least once every 4 yr. 3) Levels more than or equal to 20 and less than 100 pg/ml (at baseline or during follow up) were followed up with confirmatory PG stimulation testing. Serum CT was measured 2 and 5 min after iv injection of PG (0.5 µg/kg), and results were considered indicative of MTC when peak CT levels exceeded 100 pg/ml. 4) Basal levels more than or equal to 100 pg/ml were considered strongly suggestive of MTC, and confirmatory testing was omitted in these cases.
In all patients diagnosed with MTC, basal and PG-stimulated CT levels were measured postoperatively to evaluate the possibility of residual tumor. Findings were considered negative when both levels were within normal limits for the assay used (
10 pg/ml). This cutoff was validated in 12 volunteers who had undergone total thyroidectomy for multinodular goiters, all of whom presented PG-stimulated CT levels less than 10 pg/ml. Postthyroidectomy, PG-stimulated CT levels were considered positive for residual disease when they were more than or equal to 30 pg/ml (i.e. three times higher than the assay’s upper normal limit) and indeterminate when they were more than 10 and less than 30 pg/ml. Follow-up in the latter cases included yearly PG stimulation testing.
Surgery was proposed for all patients with any of the following: 1) FNAB indicative or suggestive of thyroid malignancy, 2) multinodular autonomous or toxic goiters, 3) large euthyroid goiters causing compression, or 4) serum CT levels (basal or PG-stimulated) more than 100 pg/ml.
Histological evaluation
Diagnosis of MTC was based on typical histological characteristics (tumor cells arranged in trabecular, insular, or sheet-like patterns, with or without stromal amyloid deposits) and immunohistochemical findings (positive staining for CT and chromogranin, negative staining for thyroglobulin). CCH was diagnosed as described by Guyetant (22). Serial sections (5 mm thick) were used for routine staining and immunohistochemistry. The immunostained sections were screened for C cells, and CCH was diagnosed when there were at least three low-power fields containing more than or equal to 50 C cells.
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Results |
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TopAbstractIntroductionPatients and MethodsResultsDiscussionReferences |
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As shown in Table 1
, 5535 of the 5817 patients we examined (prevalence, 95.2%) were unequivocally negative in level 1 screening (basal CT < 10 pg/ml), and 216 others (prevalence, 3.71%; 216 of 5817) had indeterminate basal levels (>10 and <20 pg/ml) and were referred for follow-up. The remaining 66 of 5817 (1.13%) had values more than or equal to 20 pg/ml. The nine patients in this group with basal CT levels exceeding 100 pg/ml (prevalence, 0.15%) were referred for surgery without PG test confirmation.
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The only adverse effects of PG testing were occasional brief (<1 min) bouts of nausea, headache, and/or flushing. There were no significant gender-related differences in either basal or PG-stimulated CT levels.
Final diagnosis in patients with abnormal CT levels
Based on surgical histology, 15 patients were diagnosed with MTC (prevalence, 0.26%; 15 of 5817), and seven had CCH (prevalence, 0.12%). The combined prevalence of CCH-MTC in this series was thus 0.38% (22 of 5817).
All nine patients with basal CT levels more than 100 pg/ml underwent surgery and were definitively diagnosed with MTC. In six (66.6%), the preoperative FNAB had also been suggestive of MTC.
Surgery was also recommended for all 16 patients whose PG tests were positive. This group included four patients with basal CT levels in the more than or equal to 50 and less than 100 pg/ml range. Surgical histology revealed MTC in two and CCH in the other two. Of the 12 patients with basal CT levels of more than or equal to 20 and less than 50 pg/ml, two (with peak PG-stimulated CT levels of 158 and 172 pg/ml) refused surgery and were followed for 1 and 3 yr, respectively, with no change in basal or PG-stimulated CT levels. The remaining 10 had surgery, which revealed MTC in four, CCH in four, and no evidence of neoplastic disease in two. Three of the four patients with final diagnoses of MTC had a preoperative FNAB, which was negative for MTC in two cases.
Follow-up of patients who did not undergo surgery
Forty-five of the 216 patients with indeterminate basal CT levels (>10 and <20 pg/ml) at baseline refused follow-up. The 171 who were followed (duration, 2–4 yr) included 170 whose basal levels remained less than 20 pg/ml, and one who experienced an increase (to 33 pg/ml) after 2 yr of follow-up. He underwent PG testing with positive results (peak level, 317 pg/ml) and was subjected to surgery, which demonstrated the presence of CCH.
The 41 patients with basal CT elevations (
20 and <100 pg/ml) and negative confirmatory tests at baseline have been followed with yearly PG tests, and thus far, no positive results have been observed. This group includes four patients (all followed for >4 yr) whose basal levels were in the more than or equal to 50 and less than 100 pg/ml range.
Diagnostic value of CT screening
We calculated the positive predictive value (PPV) of basal CT levels for the diagnosis of CCH, MTC, and CCH or MTC (Table 2). In both analyses, we considered only 65 of 67 patients with basal CT levels more than or equal to 20 pg/ml. The remaining two (both PG-positive) had refused surgery and were excluded from PPV analyses due to the absence of histological diagnoses. The 22 patients with histological diagnoses of CCH or MTC were considered true positives. The false-positive group (n = 43) included two PG-positive patients whose nodules were histologically diagnosed as benign and 41 with negative PG tests and FNABs that showed no cytological evidence of malignancy (naturally, none of these 41 had surgery).
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Basal CT levels displayed a PPV of 33.8% for CCH/MTC, and even lower values emerged when each diagnosis was considered alone. When the analysis was repeated for subgroups defined by basal CT levels, we found that levels more than 100 pg/ml exhibited a PPV of 100% for MTC. For levels of more than or equal to 50 and less than 100 pg/ml and more than or equal to 20 and less than 50 pg/ml, the PPVs were, respectively, 50 and 18.7% for CCH/MTC, 25 and 8.3% for MTC alone, and 25 and 10.5% for CCH alone.
In calculating the PPV for elevations of basal CT + PG confirmation, we excluded the two PG-positive patients who had refused surgery. Of the remaining 15, only 13 had surgical diagnoses of CCH (n = 7) or MTC (n = 6). PG-confirmed CT elevations thus had a PPV of 86.6% for diagnosis of CCH/MTC, but this value dropped to 40% when we considered only the diagnosis of MTC (Table 2
). In the four patients with basal CT levels more than or equal to 50 and less than 100 pg/ml, the addition of PG confirmation resulted in PPVs of 100% for CCH/MTC, 50% for MTC, and 50% for CCH alone. More limited improvement was seen in the 11 cases with basal CT levels more than or equal to 20 and less than 50 (10 at baseline, one during follow-up): PPVs of 81.8% for CCH/MTC, 36.4% for MTC, and 45.5% for CCH.
As shown in Fig. 1, all patients with MTC diagnoses presented peak PG-stimulated CT levels more than 1000 pg/ml (range, 1078–4385; median, 1990). Corresponding levels for patients diagnosed with CCH were markedly lower (range, 170–629 pg/ml; median, 300 pg/ml). Therefore, stimulated CT levels more than 1000 pg/ml had a PPV for diagnosis of MTC of 100%, and levels ranging from more than 100 to less than 1000 pg/ml had a PPV of 80% for CCH diagnosis.
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Overall, 11 of the 15 MTCs were found in clinically relevant nodules, i.e. more than or equal to 10-mm diameter. The other four were microcarcinomas that were discovered at surgery in smaller, nonbiopsied nodules within multinodular goiters. In all four cases, biopsies of the clinically relevant nodule within the goiters were benign. Tumor stages based on the classification of malignant tumors (TNM) (23) for the 15 MTC patients are shown in Table 3. Only 40% (six of 15) had stage 1 MTCs. Stage 2 tumors were found in one third of the patients (five of 15), three (20%) had stage 3 disease, and one tumor was very advanced (stage 4) at the time of diagnosis. We found no relation between basal CT levels and TNM stage.
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) indicated the presence of residual disease in five of the 15 MTC patients. This group included four of the nine MTC patients with preoperative basal CT levels more than 100 pg/ml. Their postoperative CT levels varied widely: basal CT 13–580 pg/ml with PG-stimulated peaks of 170-2800 pg/ml. All four were found to have lymph node metastases, and one also had distant metastases. The fifth patient presented a basal CT level of 17 pg/ml and PG-stimulated peak of 84 pg/ml after surgery, but we could find no evidence of either lymph node or distant metastases (T2NxMx). His preoperative results were: basal CT, 52 pg/ml; and PG-stimulated CT, 4383 pg/ml.
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Discussion |
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TopAbstractIntroductionPatients and MethodsResultsDiscussionReferences |
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In our experience, the percentage of cases identified as suspicious in first level screening (basal CT > 20 pg/ml) was very low (67 of 5817; 1.15%), and this figure would probably have been lower still if we had excluded thyroid nodule patients who also had Hashimoto’s thyroiditis, which is reportedly associated with nonneoplastic CT elevations (24). Patients with this condition represented 7.5% of our study population and 13.3% of those with suspicious basal CT levels, which is in keeping with previous reports (24).
Dual-level screening (based on basal plus PG-stimulated CT levels) allowed us to identify 15 MTCs and seven cases of CCH. The prevalence of CCH/MTC in our population was thus 0.38%, and this figure dropped to 0.26% when we considered only cases of MTC. Similar results have recently emerged from a small study (24) based on the same CT assay we used, which has been proven to be highly sensitive and specific (25), but higher prevalence rates (0.4–1.3%) have been found in the majority of published studies (6, 7, 8, 9, 10, 11, 12, 13). The discrepancy might be related to our deliberate exclusion of patients with past medical and family histories suggestive of multiple endocrine neoplasia type 2 (MEN2) disease or familial MTC (although one enrollee was ultimately identified as an index case of MEN2B with MTC). Moreover, a substantial percentage of our participants lived in regions characterized by mild to moderate iodine deficiency and, consequently, a high prevalence of benign thyroid nodules (20). Both factors could have contributed to the relatively low prevalence rate we found. On the other hand, it is conceivable that there were additional undetected cases of CCH or MTC in the 216 patients whose basal CT levels fell within the indeterminate range (>10 to <20 pg/ml). In fact, PG tests were not performed in these cases. However, the patients were followed for up to 4 yr with yearly assays of basal CT, and only one additional case (CCH) emerged. The probability of undetected cases thus appears limited, although we cannot exclude the possibility that one or more slowly progressive, early stage forms of C cell proliferative disease (or even MTC itself) might have been detected in a much longer follow-up.
As reported by others (6, 9, 13), the overall sensitivity of CT screening in our series was remarkably higher than that of FNAB. Nonetheless, basal elevations alone had limited value in predicting a diagnosis of CCH or MTC, and the PPV for diagnosis of MTC was less than 25%. Values more than 100 pg/ml were highly predictive of MTC, and, in our opinion, they should be considered a clear indication for surgery. Below this cutoff, however, the frequency of false-positivity increased progressively and diminished the PPV even for the diagnosis of CCH/MTC. On the other hand, in terms of absolute numbers, there were twice as many CCH or MTC diagnoses in the group with basal CT more than or equal to 20 and less than 50 pg/ml compared with the group with basal levels of more than or equal to 50 and less than 100 pg/ml.
The PG stimulation test thus played a major role in establishing a diagnosis of CCH or MTC when basal CT levels were less than 100 pg/ml. It also allowed preoperative differentiation of the two conditions, as noted by others (26, 27). PG is not available in the United States, but provocative testing can also be done with a short iv infusion of calcium (28, 29, 30). In our experience, this method is just as effective as PG stimulation for identifying patients with CCH or MTC (data not shown). Furthermore, in these patients, and in normal subjects as well, the stimulated CT levels measured after a 30-sec infusion of calcium gluconate (dose, 2.5 mg/kg) are very similar to those recorded after PG administration (Bianchi, D., unpublished data).
In the present study, PG test confirmation of basal increases between 20 and 100 pg/ml had a PPV of close to 90% for diagnosis of CCH or MTC, but it was much less helpful in identifying MTC alone (PPV, 40%). Over half of the patients with PG test positivity had CCH rather than MTC. Although CCH has clearly been shown to progress to MTC in patients with germline RET mutations (31), it is important to recall that there is no convincing evidence that this occurs in patients with sporadic disease. The goal of CT screening must, therefore, be restricted to diagnosis of MTC. In this setting, a PG-stimulated CT level more than 1000 pg/ml in a patient whose basal elevation was less than 100 pg/ml is strongly indicative of MTC, and we feel that it should probably be considered a clear indication for surgery. Less marked increases in response to PG (i.e. CT peak >100 to <1000 pg/ml) are fairly reliable predictors of CCH (PPV of 80%), but values in this range also led to surgery for benign nodules in two of our patients.
What are the clinical benefits of CT screening? In our series, almost half of the MTCs associated with basal CT levels more than 100 pg/ml were diagnosed at a stage when surgical treatment was unable to eliminate all neoplastic tissue. In contrast, the rate of persistent disease was substantially lower (less than 20%) among the six MTC patients with preoperative basal CT less than 100 pg/ml. It is noteworthy that four tumors in the latter group were micro-MTCs found at surgery in smaller, nonbiopsied nodules making up multinodular goiters. The larger nodules in these lesions had been biopsied and found to be benign, so these four tumors would undoubtedly have been missed if CT screening had not been carried out. In our opinion, this finding is not sufficient to justify routine CT screening for all patients with thyroid nodules. For one thing, there is no reason to assume that the frequency of micro-MTCs in this patient subset is any higher than it is in the general population. One could then argue for CT screening of the general population, but this approach would dramatically increase the cost/benefit ratio. In addition, there is no definitive evidence that these tiny tumors inevitably evolve into macro-MTCs.
In conclusion, our experience strengthens the view that circulating CT levels are the most sensitive marker available for early diagnosis of CCH and MTC (6, 7, 8, 9, 10, 11, 12, 13). If we agree that all instruments capable of improving MTC-related morbidity and mortality rates must be employed, then CT screening should clearly be included in the routine diagnostic work-up of patients with nodular thyroid disease. Our data indicate, however, that the improved sensitivity of currently available CT assays is associated with an increase in the rate of false-positive results that diminishes the cost-effectiveness of this approach. According to a generally accepted principle in clinical epidemiology, the PPV of a test can be improved by increasing its specificity (which unfortunately implies a compensatory loss of sensitivity) or by increasing the prevalence of the target phenomenon in the population to be screened, in other words, by restricting the screening process to a smaller higher-risk population. In terms of the early cost-effective diagnosis of MTC, this would mean limiting basal and PG-stimulated serum CT screening to a select subset of thyroid nodule patients at higher risk for this type of malignancy. Future prospective studies should focus on the identification of factors defining this subset (e.g. nodules associated with pain and/or localized in the upper third of the lobe, indeterminate FNAB results, and/or U.S. findings suggestive of possible malignancy).
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Footnotes |
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G.C., D.M., C.D., D.B., M.N., S.T., U.C., M.A., M.M., M.T., and S.F. have nothing to declare.
First Published Online November 21, 2006
1 G.C. and D.M. contributed equally to this study. 
Abbreviations: CCH, C cell hyperplasia; CT, calcitonin; FNAB, fine-needle aspiration biopsy; MTC, medullary thyroid carcinoma; PG, pentagastrin; PPV, positive predictive value; TNM, classification of malignant tumors.
Received July 21, 2006.
Accepted November 14, 2006.
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References |
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TopAbstractIntroductionPatients and MethodsResultsDiscussionReferences |
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