Novel acquisitions in the diagnosis of medullary thyroid carcinoma.

INTRODUCTION: The correct identification of medullary thyroid carcinoma (MTC) has been a challenge since its first description. In the last few years, some advances in this context have been achieved. Here we aimed to review and discuss published data on the more recent acquisition in the diagnosis of MTC. EVIDENCE ACQUISITION: A literature search of the medical databases was conducted by searching papers reporting tool for diagnosis of MTC published in the last years. The search was updated until July 2016. EVIDENCE SYNTHESIS: The literature search revealed several relevant articles which focused on different topics of the diagnosis of MTC. The results are reported by four paragraphs, such as 1) fine-needle aspiration; 2) molecular analysis; 3) serum markers; 4) imaging. CONCLUSIONS: The measurement of calcitonin in FNA washout fluids is essential due to poor accuracy of conventional cytology to detect MTC. Genetic analysis can help to identify those advanced MTC with poorer prognosis who do not respond to chemotherapy. Procalcitonin may in the next future replace calcitonin as serum diagnostic marker of MTC. Recent evidence based data seem to suggest the emerging role of functional imaging in recurrent MTC in patients with calcitonin serum levels >150 pg/mL.

Galectin-3 and HBME-1 improve the accuracy of core biopsy in indeterminate thyroid nodules.

Core needle biopsy (CNB) has been recently described as an accurate second-line test in thyroid inconclusive cytology (FNA). Here we retrospectively investigated the potential improvement given by Galectin-3, Cytokeratin-19, and HBME-1 on the accuracy of CNB in thyroid nodules with prior indeterminate FNA report. The study included 74 nodules. At CNB diagnosis, 15 were cancers, 40 were benign, and 19 had uncertain/non-diagnostic CNB report. The above immunohistochemical (IHC) panel was analyzed in all cases. After surgery, 19 malignant and 55 benign lesions were found. All 15 cancers and all 40 benign nodules diagnosed at CNB were confirmed at final histology. Regarding the uncertain CNB group, 4 (21 %) were malignant and 15 (79 %) benign. When we considered all the series, the most accurate IHC combination was Galectin-3 plus HBME-1, while HBME-1 was the most sensitive marker in those nodules with uncertain CNB report. The combination of CNB plus IHC could indentify 19/19 cancers and 53/55 benign lesions. Sensitivity and specificity of CNB increased from 79 to 100 % and from 73 to 96 %, respectively, by adding IHC. CNB can diagnose the majority of thyroid nodules with previous indeterminate FNA cytology, while the accuracy of CNB is increased by adding Galectin-3, Cytokeratin-19, and HBME-1 panel. We suggest to adopt CNB as a second-line approach to indeterminate thyroid FNA, and apply IHC in those lesions with uncertain/non-diagnostic CNB report. This approach should improve the pre-surgical diagnosis of patients. These results should be confirmed in larger prospective series.

Ultrasound features of medullary thyroid carcinoma correlate with cancer aggressiveness: a retrospective multicenter study.

BACKGROUND: Poor prognosis of medullary thyroid cancer (MTC) with suspicious ultrasound (US) features has been reported. The aim of the study was to investigate the association between preoperative US presentation and aggressiveness features of MTC. Also, US features of MTC were compared with those previously reported. METHODS: Study group comprised 134 MTC from nine different centers. Based on US presentation the nodules were stratified in "at risk for malignancy" (m-MTC) or "probably benign" (b-MTC) lesions. RESULTS: Eighty nine (66.4%) m-MTC and 45 (33.6%) b-MTC were found. Metastatic lymph nodes (p = 0.0001) and extrathyroid invasiveness (p < 0.0001) were more frequent in m-MTC. There was statistically significant correlation (p = 0.0002) between advanced TNM stage and m-MTC with an Odds Ratio 5.5 (95% CI 2.1-14.4). Mean postsurgical calcitonin values were 224 ± 64 pg/ml in m-MTC and 51 ± 21 in b-MTC (p = 0.003). CONCLUSIONS: This study showed that sonographically suspicious MTC is frequently associated with features of aggressiveness, suggesting that careful preoperative US of MTC patients may better plan their surgical approach.

Clinical characteristics as predictors of malignancy in patients with indeterminate thyroid cytology: a meta-analysis.

Indeterminate thyroid nodules (ITN) constitute the gray zone of thyroid fine-needle aspiration cytology (FNAC). About 70-80 % of ITN are later diagnosed as benign; therefore, it is very important to identify the predictors of malignancy. Aim of the study was to summarize published data about clinical risk factors for malignancy in patients with ITN and thereby provide more robust estimates of the effect of these risk factors. Sources comprised studies published through December 2012. Original articles that investigated clinical parameters as potential predictors of malignancy in ITN were identified. Two authors performed the data extraction independently. A meta-analysis of 19 relevant studies was conducted that included 3,494 patients with ITN according to FNAC. The pooled prevalence of malignancy was 28 % (95 % CI 23-33), 26 % in females and 34 % in males. The pooled OR was 1.51 (95 % CI 1.2-1.83) for males and 0.68 (95 % CI 0.53-0.88) for females. Regarding the nodule's size, the pooled OR was 2.10 (95 % CI 1.26-3.50) for nodules >4 cm in diameter. Analysis of the patient age as a risk factor was not feasible because of marked difference found between the studies. In patients with indeterminate thyroid nodules diagnosed at FNAC, the pooled rate of malignancy from 19 studies was 28 %. Patients that are male and have ITN greater than 4 cm in diameter should be considered at higher risk of cancer.

Calcitonin measurement in aspiration needle washout fluids has higher sensitivity than cytology in detecting medullary thyroid cancer: a retrospective multicentre study.

OBJECTIVE: Only few studies analysed the capability of cytology in detecting medullary thyroid cancer (MTC), and they reported a low accuracy of this diagnostic technique. Recently, calcitonin (CT) measurement in aspiration needle washout (FNA-CT) of thyroid and neck lesions has been reported as a sensitive tool for MTC. The aim of this study is to compare the sensitivity of FNA-CT and cytology in detecting MTC and to assess a cut-off value of FNA-CT for clinical practice. PATIENTS: Thirty-eight MTC lesions from 36 patients were retrospectively studied, diagnosed and treated in four different centres. Furthermore, 52 nonmedullary lesions from subjects undergone biopsy following increased serum CT were collected as a control group. RESULTS: Cytology detected MTC in 21/37 lesions with 56·8% sensitivity. The median FNA-CT value was 2000 pg/ml (range 58-10 000 pg/ml) in MTC and 2·7 pg/ml (range <2-13 pg/ml) in controls (P < 0·001). Using a cut-off of 39·6 pg/ml, MTC lesions could be identified with 100% sensitivity and specificity. As the most important finding, 14 histologically proved MTC lesions could be detected by FNA-CT, despite they were cytologically diagnosed as benign or nonconclusive. CONCLUSIONS: This study shows, as the first in a multicentre series, that FNA-CT sensitivity is higher than that of cytology in diagnosing MTC. To avoid false-negative MTC by cytology, CT measurement in aspiration needle washout is to be performed in all patients undergoing biopsy following high serum CT.

Diagnostic performance of (99m)Tc-MIBI scan in predicting the malignancy of thyroid nodules: a meta-analysis.

Several studies have investigated the diagnostic performance of (99m)Tc-MIBI scan in the evaluation of thyroid nodules suspicious for malignancy with conflicting results. The aim of our study is to meta-analyze published data on this topic. A comprehensive literature search of studies published through December 2012 regarding the diagnostic performance of (99m)Tc-MIBI scan in the evaluation of thyroid nodules suspicious for malignancy was carried out. Pooled sensitivity and specificity of (99m)Tc-MIBI scan on a per lesion-based analysis and the area under the ROC curve were calculated. Pathological reports of thyroid nodules were considered as reference standard. Twenty-one studies were included in the meta-analysis. Pooled sensitivity and specificity of (99m)Tc-MIBI scan in detecting malignant thyroid nodules were 85.1 % [95 % confidence interval (95 % CI): 81.1-88.5 %] and 45.7 % (95 % CI: 42.7-48.7 %), respectively, on a per lesion-based analysis, irrespective of eventual results of previous technetium pertechnetate ((99m)TcO4) or iodine-123 ((123)I) scan. The area under the ROC curve was 0.78. A sub-analysis restricted to data on hypofunctioning nodules on (99m)TcO4 or (123)I scans was performed: pooled sensitivity and specificity of (99m)Tc-MIBI scan in these nodules were 82.1 % (95 % CI: 77.2-86.3 %) and 62.8 % (95 % CI: 58.9-66.7 %), respectively, on a per lesion-based analysis. The area under the ROC curve was 0.81. (99m)Tc-MIBI scan is a sensitive diagnostic tool in predicting the malignancy of thyroid nodules. Therefore, this imaging method could be helpful in patients with thyroid nodules in which malignancy is suspected on the basis of conventional diagnostic techniques. Higher specificity can be reached when hypofunctioning thyroid nodules are considered.

Mammalian target of rapamycin pathway activation is associated to RET mutation status in medullary thyroid carcinoma.

CONTEXT: The genetic pathways involved in medullary thyroid carcinomas (MTC), except for RET mutations, are largely unknown, as is the detailed mapping of proteins activated as a consequence of RET tyrosine kinase phosphorylation. OBJECTIVE: The present study was designed to screen for the presence of mutations in other genes downstream to RET activation and to detect the activation patterns of a panel of intracellular regulators of cell growth. DESIGN: Forty-nine cases of MTC were analyzed for mutations in RET, BRAF, N-, H-, and K-RAS, and phosphatidylinositol-3 (PI3) kinase genes. Immunohistochemical analysis was performed using antibodies against several intracellular transducers. The effect of mammalian target of rapamycin (mTOR) inhibition was assessed in vitro onto TT cells by means of methyl thiazolyl tetrazolium and Western blot assays. RESULTS: BRAF, K-, H-, and N-RAS, and PI3 kinase mutations were absent in all cases examined. Germline RET mutations were detected in 20% of cases overall, whereas somatic RET mutations represented 53% of sporadic tumors. RET mutational status was associated to age, presence of multifocal tumors, and nodal status, but not disease outcome. Protein expression of markers investigated was highly heterogeneous, with a strong association between phospho-mTOR, phospho-AKT, and phospho-p70S6K, positively correlated to the presence of germline RET mutations. Moreover, selective mTOR inhibition affected cell proliferation of RET-mutant TT cells. CONCLUSIONS: Taken together, our findings indicate that mTOR intracellular signaling pathway is functionally activated in MTC with a preferential expression in cases with germline RET mutations; genes downstream to RET tyrosine kinase such as BRAF, RAS isoforms, and PI3 kinase are not mutated in MTC.

99mTc-MIBI Imaging in the presurgical characterization of thyroid follicular neoplasms: relationship to multidrug resistance protein expression.

UNLABELLED: Recently, thyroid (99m)Tc-methoxyisobutylisonitrile ((99m)Tc-MIBI) scintiscanning has been proposed in an attempt to preoperatively identify thyroid malignancies, but discrepant results have been reported for oncocytic lesions. The aim of this study was to investigate the usefulness of visual and semiquantitative analyses of (99m)Tc-MIBI scintigraphy for preoperatively characterizing thyroid nodules with indeterminate cytologic diagnoses, segregating in advance nononcocytic variants from those that are oncocytic. This study also aimed to analyze the relationship between (99m)Tc-MIBI images and P-glycoprotein (P-gp)/multidrug resistance-associated protein-1 (MRP1) immunohistochemical expression. METHODS: Fifty-one consecutive patients with cold thyroid nodules cytologically diagnosed as nononcocytic or oncocytic follicular neoplasm were prospectively studied. Visual and semiquantitative (99m)Tc-MIBI scanning was performed and the diagnoses of the lesions were histologically proven by subsequent thyroidectomy. Immunohistochemical evaluation of P-gp and MRP1 was also performed on surgical samples. RESULTS: Visual and semiquantitative (99m)Tc-MIBI scintiscans showed a low specificity in preoperatively discriminating malignant oncocytic lesions. In nononcocytic nodules, the semiquantitative method was more accurate than the visual (94.44% and 77.78%, respectively). P-gp protein expression was negative in all thyroid lesions, whereas apical plasma membrane MRP1 expression was found in 78% of the lesions with a negative (99m)Tc-MIBI retention index, compared with 11% of lesions with a positive retention index, correlating most strongly with a negative (99m)Tc-MIBI RI in those cases with strong MRP1 apical expression. CONCLUSION: Semiquantitative (99m)Tc-MIBI scintigraphy is an adjunctive method to predict preoperatively the malignant behavior of nononcocytic follicular thyroid nodules indeterminate at fine-needle aspiration biopsy, with a potential impact on the definition of their clinical management. Moreover, the good correlation found between immunohistochemical apical expression of MRP1 and the scintigraphic findings supports the (99m)Tc-MIBI results and provides tissue information on the molecular mechanisms responsible for (99m)Tc-MIBI images in thyroid lesions.

Galectin-3-expression analysis in the surgical selection of follicular thyroid nodules with indeterminate fine-needle aspiration cytology: a prospective multicentre study.

BACKGROUND: In the USA, about 30 200 well-differentiated thyroid carcinomas were diagnosed in 2007, but the prevalence of thyroid nodules is much higher (about 5% of the adult population). Unfortunately, the preoperative characterisation of follicular thyroid nodules is still a challenge, and many benign lesions, which remain indeterminate after fine-needle aspiration (FNA) cytology are referred to surgery. About 85% of these thyroid nodules are classified as benign at final histology. We aimed to assess the diagnostic effect of galectin-3 expression analysis in distinguishing preoperatively benign from malignant follicular thyroid nodules when FNA findings were indeterminate. METHODS: 544 patients were enrolled between June 1, 2003, and Aug 30, 2006. We used a purified monoclonal antibody to galectin-3, a biotin-free immunocytohistochemical assay, and a morphological and phenotypic analysis of FNA-derived cell-block preparations. Galectin-3-expression analysis was applied preoperatively on 465 follicular thyroid proliferations that were candidates for surgery, and its diagnostic accuracy was compared with the final histology. FINDINGS: 31 patients were excluded because they had small galectin-3-negative thyroid nodules; we did not have data for 47 patients; and one patient with an oncocytic nodule was excluded. 331 (71%) of the assessable 465 preoperative thyroid FNA samples did not express galectin-3. 280 (85%) of these galectin-3-negative lesions were classified as benign at final histology. Galectin-3 expression was detected, instead, in 134 of 465 (29%) thyroid proliferations, 101 (75%) of which were confirmed as malignant. The overall sensitivity of the galectin-3 test was 78% (95% CI 74-82) and specificity was 93% (90-95). Estimated positive predictive value was 82% (79-86) and negative predictive value was 91% (88-93). 381 (88%) of 432 patients with follicular thyroid nodules who were referred for thyroidectomy were correctly classified preoperatively by use of the galectin-3 test. However, 29 (22%) of 130 cancers were missed by the galectin-3 method. INTERPRETATION: Our findings show that if the option of surgery was based theoretically on galectin-3 expression alone, only 134 thyroid operations would have been done in 465 patients; therefore a large proportion (71%) of unnecessary thyroid surgical procedures could be avoided, although a number of galectin-3-negative cancers could be potentially missed. The galectin-3 test proposed here does not replace conventional FNA cytology, but represents a complementary diagnostic method for those follicular nodules that remain indeterminate.

Absence of RET gene point mutations in sporadic thyroid C-cell hyperplasia.

Progression from C-cell hyperplasia (CCH) to medullary thyroid carcinoma (MTC) has been demonstrated to date only in familial forms, whereas in nonfamilial MTC, such hypothesis is suggested by the rare concurrence of both lesions, although no epidemiological and molecular data are available to prove or disprove this event. Therefore, the clinical management of patients with sporadic CCH is controversial. To evaluate the malignant potential of sporadic CCHs, pure laser-microdissected C-cell populations of 24 CCH cases, either reactive or associated with nonfamilial MTC, were analyzed for MTC-associated protein neural cell adhesion molecule expression and RET point mutations in exons 10, 11, 15, and 16, by using immunohistochemistry and polymerase chain reaction-single-strand conformation polymorphism/heteroduplex electrophoresis/direct sequencing, respectively. No RET mutations were found in any of the 24 CCH cases, whereas M918T mutation was detected in three concomitant MTCs. Neural cell adhesion molecule was immunoreactive in the majority of CCH associated with MTC even in the absence of morphological atypia, but not in reactive forms. The absence of RET alterations in all cases of CCH examined supports the hypothesis that the development of MTC is independent of pre-existing CCH in the nonfamilial setting; thus, sporadic CCH should not be considered a risk factor for nonfamilial MTC.

Galectin-3 and Ki-67 expression in multiglandular parathyroid lesions.

Hyperplastic and neoplastic parathyroid lesions may present overlapping morphologic features, and several markers have been proposed to distinguish benign from malignant growths. Recently, it was reported that galectin-3 is a useful marker of malignancy in uniglandular parathyroid diseases. To investigate galectin-3 and Ki-67 immunoexpression in parathyroid hyperplastic disease, 63 multiglandular lesions (13 primary, 40 secondary, and 10 tertiary hyperplasia cases) were analyzed and compared with 45 control cases of parathyroid adenomas and 24 carcinomas. Our data showed that hyperplastic lesions responsible for primary nonfamilial or tertiary hyperparathyroidism, as well as parathyroid adenomas, were negative for galectin-3, as opposed to carcinomas. In addition, secondary and familial primary hyperplasia cases were surprisingly positive for galectin-3 in approximately two thirds of cases. All hyperplastic lesions (positive or negative for galectin-3) had a low Ki-67 index. Based on these findings, secondary hyperplasia has a low proliferative potential but an unexplained galectin-3 reactivity, which reduces its diagnostic role in differentiating benign from malignant nodules in the context of multiglandular parathyroid diseases.

Galectin-3 and HBME-1 expression in oncocytic cell tumors of the thyroid.

Oncocytic cell tumors (OCTs) of the thyroid include oncocytic cell adenomas (OCAs) and oncocytic cell carcinomas (OCCs). Oncocytic variant of papillary carcinoma (OVPC) has also been described. These tumors may present similar diagnostic problems as their non-oncocytic counterparts, in both conventional histology and fine-needle aspiration biopsies. Several markers were shown able to distinguish benign from malignant thyroid follicular tumors, galectin-3 and HBME-1 being the most promising ones. Controversial data have been reported on their discriminatory potential in the small series of OCTs so far analyzed. We aimed to assess the role of galectin-3 and HBME-1 in a large series of 152 OCTs (including 50 OCAs, 70 OCCs and 32 OVPCs). The expression of PPARgamma protein was also evaluated. Using a biotin-free detection system, the sensitivity of galectin-3 was 95.1%, while that for HBME-1 was nearly 53%. The combination of galectin-3 and HBME-1 increased the sensitivity up to 99%. However, for both markers, the specificity was 88%, lower than that reported for non-oncocytic follicular tumors. PPARgamma protein overexpression was absent in all OCAs tested and present in only 10% of OCCs, confirming previous reports on the low prevalence of PAX8-PPARgamma translocations in OCT and ruling out its role as a potential diagnostic marker of malignancy.

Role of galectin-3 immunodetection in the cytological diagnosis of thyroid cystic papillary carcinoma.

OBJECTIVE: Cystic thyroid lesions can harbour an occult papillary carcinoma, which fine needle aspiration (FNA) biopsy may fail to detect. Recently, new markers such as galectin-3 lectin have been proposed to distinguish benign from malignant thyroid lesions of follicular origin. The aim of this study was to assess the role of galectin-3 immunodetection in a series of FNA cytological samples of benign and malignant thyroid cystic nodules. METHODS: We retrospectively analysed galectin-3 expression by immunoperoxidase staining on 32 cytological paraffin-embedded samples of cystic papillary carcinoma and on 12 samples of benign cysts, both obtained by FNA biopsy. Specificity, sensitivity, positive/negative predictive values, and accuracy of standard cytological examination and galectin-3 immunodetection were assessed. RESULTS: Among cystic papillary carcinomas, 29 of 32 samples were galectin-3 positive, whereas standard FNA cytology made a correct diagnosis in only 25 of 32 samples. All the benign cysts were negative for galectin-3. In comparing the sensitivity and specificity of the two methods, it appeared that both had a 100% specificity, whereas the sensitivity of cytological examination alone was 75% versus 89.3% obtained by galectin-3 immunohistochemistry. CONCLUSIONS: Galectin-3 immunostaining represents a valid pre-operative adjunct to pick up malignant cells in those cases where a very poor number of epithelial cells may lead to a cytological misdiagnosis. Therefore, we suggest that in poorly cellular FNA biopsies of simple or complex thyroid cysts, galectin-3 expression by epithelial cells is consistent with a cystic carcinoma and supports surgical treatment indication.