Aberrant localization of beta-catenin correlates with overexpression of its target gene in human papillary thyroid cancer

Alterations of the Wnt/beta-catenin signaling pathway are known to occur in mutations of the component genes such as APC, Axin, and beta-catenin, and play a pathogenetic role in tumorigenesis. Activated Wnt signaling stabilizes beta-catenin, which associates with T cell factor, resulting in transactivation of the downstream target genes including c-myc and cyclin D1. To investigate the involvement of Wnt/beta-catenin signaling pathway in thyroid tumorigenesis, we analyzed its activation and localization in 5 human thyroid cancer cell lines and 132 thyroid tumor tissue samples. Dislocalization of beta-catenin was observed in all cell lines. Constitutive activation of T cell factor in two of four thyroid cancer cell lines was observed using reporter gene assay. Furthermore, high expression levels of c-Myc and cyclin D1 were observed in cell lines that showed cytoplasmic or nuclear accumulation of beta-catenin. In 132 paraffin-embedded thyroid carcinoma tissue samples, cytoplasmic beta-catenin was immunohistochemically observed in 52 out of 78 (67%) papillary thyroid cancers, but only in 3 of 34 (9%) follicular adenomas and 5 of 20 (25%) follicular cancers. Cytoplasmic localization of beta-catenin significantly correlated with overexpression of cyclin D1 in papillary carcinomas. Our results suggest that aberrant activation of Wnt/beta-catenin signaling is strongly involved in thyroid tumorigenesis.     

Nuclear cyclin D1 overexpression is a critical event associated with cell proliferation and invasive growth in gallbladder carcinogenesis

Cyclin D1 overexpression is remarkably frequent in several human carcinomas and is believed to be a critical event in oncogenesis. We examined cyclin D1 expression, p53 expression, and the Ki-67 labeling index by immunostaining in human gallbladder mucosa in conditions varying from normal to malignant tissue. We also examined K-ras codon 12 mutations in these tissues with a two-step polymerase chain reaction. Nuclear cyclin D1 overexpression was observed in 48% of carcinomas occurring independently of adenoma, but not in adenomas, carcinomas arising in adenomas, or nonneoplastic lesions. Cytoplasmic cyclin D1 overexpression was observed in about 15% of abnormal specimens, irrespective of the type of epithelial abnormality. Carcinomas showing nuclear cyclin D1 overexpression had significantly higher Ki-67 labeling indexes than those with no overexpression. Moderately to poorly differentiated adenocarcinomas showed a higher incidence of nuclear cyclin D1 overexpression than papillary to well differentiated carcinomas. Specimens with cyclin D1 overexpression showed a high incidence of lymph permeation, venous permeation, and lymph node metastasis. We conclude that nuclear cyclin D1 overexpression is a critical event importantly associated with cell proliferation and invasive growth in gallbladder carcinogenesis, and that cyclin D1 immunostaining may become a useful marker for evaluating gallbladder carcinomas. Received: March 9, 1999 / Accepted: July 23, 1999     

Low level of cyclin D1 protein expression in thyroid microcarcinomas from an autopsy series

In a recent epidemiological screening study in an autopsy series, we found a high prevalence of microcarcinomas (MCs) (21/443 = 4.74%). We found no iodine intake-, gender-, or age-dependent differences in the prevalence of MCs. The results suggest a different and benign behavior of MCs compared to clinical cancer. The role of cyclin D1 overexpression in the pathogenesis of thyroid tumors is not known clearly; however, overexpression of this protein was reported in well-differentiated papillary cancers and in incidentally found metastasizing MCs. To date, cyclin D1 expression has not been investigated in autopsy-derived thyroid MCs. Eight MCs were available for immunostaining and comparison with 15 clinically detected papillary thyroid cancers. Fourteen out of 15 clinical carcinomas expressed cyclin D1 (93.3%), while in the MCs this ratio was 1 out of 8 (12.5%) (p = 0.0001). The only cyclin D1-positive MC was multifocal (both lobes of the gland were affected). We concluded that the benign behavior of most autopsy-derived MCs may be associated with the lack of cyclin D1 overexpression.     

BRAF(V599E) mutation is the leading genetic event in adult sporadic papillary thyroid carcinomas

Activating mutations of BRAF have been identified in a variety of human cancers, most notably melanomas and papillary thyroid carcinomas (PTCs). The aim of the present study was to disclose the role of BRAF mutations in thyroid carcinoma development. Seventy-two thyroid tumors, including 60 PTCs, six follicular adenomas, five follicular carcinomas, and one anaplastic carcinoma, were studied. BRAF mutation screening focused on exon 15 and exon 11 of the gene by single-stranded conformational polymorphism and sequence analysis. Search of RET/PTC expression was conducted with the RT-PCR technique. The molecular genetic study of the BRAF gene showed the presence of a missense thymine to adenine transversion at nucleotide 1796, resulting in the V599E substitution, in 24 of 60 PTCs (40%), none of six follicular adenomas, and none of five follicular carcinomas or one anaplastic carcinoma. Moreover, nine of 60 PTCs (15%) presented RET/PTC expression. A genetico-clinical association analysis showed a statistically significant correlation between BRAF mutation and development of PTCs of the classic papillary histotype (P = 0.038). On the contrary, no link could be detected between expression of BRAF(V599E) and age at diagnosis, gender, dimension, and local invasiveness of the primary cancer, presence of lymph node metastases, tumor stage, and multifocality of the disease. These data clearly confirm that BRAF(V599E) is the more common genetic alteration found to date in adult sporadic PTCs, that it is unique for this thyroid cancer histotype, and that it might drive the development of PTCs of the classic papillary subtype.     

Thyroid cancer and thyroiditis in the goitrous region of Salta, Argentina, before and after iodine prophylaxis

OBJECTIVE The importance of iodine intake and thyroiditis in the pathogenesis of thyroid cancer remains controversial. We have investigated the natural history of thyroid cancer and thyroiditis in a goitrous region before and after iodine prophylaxis over a 31-year period. DESIGN For the analysis of thyroid cancer the material was divided in two periods. The first 15 years (59 cases), including 5 years before prophylaxis, was compared with the second 16 years (85 cases), a period well after iodine supplementation of salt. Histological diagnosis of the tumours was based on the WHO system. Moderate to severe thyroiditis in the non-tumoral surrounding thyroid from female patients was recorded. For this, the material was analysed in the two periods In relation to the introduction of iodine prophylaxis in 1963, taking account of the age of the patients. RESULTS Papillary carcinomas formed the largest group of tumours in both periods, with nearly twice as many in the second period as the first, while the numbers of follicular and medullary carcinomas remained about the same. The ratio of papillary to follicular carcinoma rose from 1.7:1 in the first period to 3.1:1 in the second. All three thyroid lymphomas were of the non-Hodgkin's type, and all occurred in the second period in females aged over 50. A severe lymphoid thyroiditis was present in the two cases with assessable background thyroid tissue. The frequency of lymphoid infiltrate in females rose from 8% 11/12) before 1963 to 25% (18172) after prophylaxis in the whole series. After salt prophylaxis, thyroiditis was more frequent in patients with papillary carcinoma in general (31%), and clinically significant papillary carcinomas in particular (35%), than in those with non-papillary tumours (6%) (X², P < 0.05 and P < 0.025, respectively). CONCLUSIONS Our observations indicate that a high dietary intake of iodine may be associated with a high frequency of papillary carcinoma and thyroiditis, and that thyroiditis is more commonly associated with papillary carcinoma than with other thyroid tumours. The occurrence of non-Hodgkin's lymphomas only in the post-prophylaxis period may be linked to an increase in thyroiditis.     

Differential expression of IgG Fc binding protein (FcgammaBP) in human normal thyroid tissue,thyroid adenomas and thyroid carcinomas

The genetic events involved in thyroid carcinogenesis are still incompletely understood. Several rearrangements and mutations of oncogenes have been implicated in the development of thyroid papillary carcinomas, follicular adenomas and carcinomas. However, none of these molecular alterations is suitable either as a general marker for the diagnosis of thyroid carcinomas or to differentiate between thyroid follicular adenomas and carcinomas. In order to identify new genes with altered expression which could serve as such markers, we analyzed RNA from thyroid tumor and normal tissue using a novel technique called restriction-mediated differential display. Several differentially expressed genes were identified, including the gene for IgG Fc binding protein (FcgammaBP). Differential expression of FcgammaBP was confirmed by quantitative real-time RT-PCR. Our experiments showed that IgG Fc binding protein (FcgammaBP) is differentially expressed in normal thyroid tissue, thyroid adenomas and thyroid carcinomas. While the FcgammaBP gene is constitutively expressed in normal thyroid tissue, its expression is significantly increased in follicular thyroid adenomas and significantly decreased in papillary and follicular thyroid carcinomas. Thus, measurement of the expression levels of FcgammaBP in thyroid biopsies might help to make the otherwise difficult distinction between a thyroid follicular adenoma and a follicular carcinoma.     

Immunostaining for Met/HGF receptor may be useful to identify malignancies in thyroid lesions classified suspicious at fine-needle aspiration biopsy.

The receptor for hepatocyte growth factor (Met) is not expressed in the normal thyroid but it is overexpressed in most thyroid carcinomas. We evaluated whether Met immunostaining of cytological smears from fine-needle aspiration biopsy (FNAB) may be useful for the preoperative diagnosis of thyroid cancer. Notably, routine cytological examination often fails to distinguish well-differentiated follicular carcinomas and a proportion of papillary carcinomas (low-grade papillary carcinomas and papillary carcinomas follicular variant [FVPTC]) from benign lesions: all these lesions are usually classified as suspicious. We examined 80 thyroid lesions diagnosed as suspicious at cytology that had subsequently undergone surgery. The histologic diagnosis had been: papillary carcinomas (n = 14), FVPTC (n = 11), follicular carcinomas (n = 25), atypical follicular adenomas (n = 5), follicular adenomas (n = 20), and nodular goiters (n = 5). We also studied typical papillary carcinomas (n = 30) and nodular goiters (n = 10), all correctly diagnosed at cytology. In lesions classified suspicious at routine cytology, Met immunostaining was positive in 12 of 14 (85.7%) papillary carcinomas, 8 of 11 (72.7%) FVPTC, 7 of 25 (28%) follicular carcinomas, and 5 of 5 atypical adenomas. In contrast, none of the 25 lesions cytologically suspicious but benign at histology were positive. These data suggest that Met immunostaining of suspicious cytological smears are useful for identifying malignant lesions, especially those with a papillary histotype.     

Expression of c-erbB-2 oncoprotein in different types of thyroid tumors: an immunohistochemical study

AIMS: c-erbB-2 expression has been found to be an important prognostic parameter in various types of tumors, but the role played by this oncoprotein in thyroid tumors is still controversial. This study is designed to investigate c-erbB-2 expression in different types of thyroid tumors. METHODS: c-erbB-2 protein over-expression was studied immunohistochemically (IHC) in 68 paraffin-embedded thyroid tumors (12 follicular adenomas, 20 papillary carcinomas, 20 follicular carcinomas, eight medullary carcinomas, eight anaplastic carcinomas), diagnosed at the Pathology Department of Cukurova University. RESULTS: No expression of c-erbB-2 was found in anaplastic carcinomas, but 50% of the papillary carcinomas showed a cytoplasmic staining pattern and 45% had a mixed staining pattern (cytoplasmic and membranous patterns). Eight (40%) of the 20 follicular carcinomas showed a mixed cytoplasmic/membranous staining pattern and three (15%) showed only cytoplasmic reactivity. Cytoplasmic immunoreactivity was detectable for c-erbB-2 in six out of eight (75%) medullary carcinomas. Finally, seven cases of 12 (58%) follicular adenomas showed cytoplasmic staining. CONCLUSIONS: Investigation of the exact role of c-erbB-2 in thyroid tumors requires further studies, and the different patterns of immunostaining may be helpful for determination of prognosis as is the case in breast carcinoma.     

Galectin-3: presurgical marker of thyroid follicular epithelial cell-derived carcinomas

Preoperative follicular lesion characterisation represents an unsolved diagnostic problem in thyroid nodular disease. Although fine-needle aspiration biopsy is the most reliable preoperative diagnostic procedure, it shows inherent limitations in differentiating adenoma from follicular carcinoma and, sometimes, follicular variants of papillary carcinoma. Galectin-3 cytoplasmic neoexpression has been proposed as a peculiar feature of thyroid malignant cells, easily detectable in cytological and histological samples. The aim of this study was to re-evaluate the galectin-3 expression in a large sample of thyroid lesions using an immunohistocytochemical biotin-free detection system and a specific anti-human-galectin-3 monoclonal antibody in order to avoid the interference of technical factors, a cause of conflicting results recently reported by some authors. We analysed galectin-3 expression of 39 follicular carcinomas, 26 papillary carcinomas, and 105 adenomas in both cell-block samples and their histological counterparts. All cell-block and histological papillary carcinoma samples showed high levels of galectin-3 immunoreactivity. Thirty-four follicular carcinomas were positive, whereas 5 were negative in cell-blocks but positive in their histological counterparts. Twelve out of 105 adenomas expressed galectin-3 in cell-blocks and histological samples. The diagnostic accuracy of preoperative galectin-3 evaluation in adenomas vs follicular carcinomas was 90.0%. Galectin-3 expression was also investigated in 22 minimally-invasive follicular carcinomas. All of them showed galectin-3 immunoreactivity in both cytological and histological specimens with the exception of two cases, where galectin-3 positivity was observed only in the surgical material. The routine correct use of galectin-3, by increasing the diagnostic accuracy of conventional cytology, improves the management of thyroid nodules and can lead to a sensitive reduction of useless thyroid surgeries.     

Telomerase activity in thyroid malignancy.

Telomerase activity seems to play a role in the development and pathogenesis of thyroid carcinoma. Its incidence of expression and its application as a tumor marker remain to be elucidated. Thyroid tissues obtained during thyroidectomy from 1996-1998 were rapidly frozen and stored at -80 degrees C until processed. Telomerase activity was determined using telomeric repeat amplification protocol (TRAP). Histology of the tissue examined (67 benign and 59 malignant) was reviewed. Telomerase activity was detected in 15 of 52 papillary carcinomas (29%); 1 of 1 thyroid lymphoma (100%); 1 of 2 anaplastic carcinomas (50%); and 2 of 16 lymphocytic thyroiditis specimens (13%). Telomerase activity was not detectable in 35 normal thyroid, 9 follicular adenoma, 7 nodular hyperplasia, 2 follicular carcinoma, and 2 medullary carcinoma. Lymphocytic thyroiditis was detected in 8 of 37 (22%) apparently normal thyroid tissues adjacent to papillary thyroid carcinoma and telomerase activity was present in 2 of these 8 specimens (25%). In conclusion, telomerase does not appear to be frequently activated in papillary thyroid carcinoma. The association of lymphocytic thyroiditis with papillary thyroid carcinoma may limit its clinical usefulness as a tumor marker.     

Localization and expression of thyroid hormone receptors normal and neoplastic human thyroid

The aim of this study was to investigate the regional expression of thyroid hormone nuclear receptor forms (TR(alpha) and TR(beta)) and isoform (TR(alpha1) and TR(beta2)) mRNAs in normal and neoplastic (benignant and malignant) human thyroid tissue. Tumor specimens from patients with thyroid carcinomas (papillary: 5 cases; follicular: 5 cases; anaplastic: 2 cases), thyroid follicular adenomas (7 cases) and tissue from normal thyroid glands (12 cases) were analyzed by in situ hybridization and semiquantitative RT-PCR for the expression of TR(alpha1) and beta, as well as for the isoform alpha2 that does not bind the hormone. In normal tissues, TR(alpha2) was expressed at lower levels compared to TR(alpha1) (alpha1/alpha2 = 4.3). In papillary and follicular carcinomas, the expression of TR(alpha1) and TR(beta) did not change as compared with normal thyroid tissue and adenomas (0.87 +/- 0.15 SD vs 0.89 +/- 0.17 densitometric units, DU, and 0.15 +/- 0.02 vs 0.14 +/- 0.03 DU, respectively). However, the expression of TR(alpha2) was significantly higher in differentiated carcinomas compared to normal thyroid tissue and adenomas (0.47 +/- 0.05 vs 0.20 +/- 0.05 DU, p < 0.05) with alpha1/alpha2 = 1.4. In anaplastic carcinoma all TRs were absent. We concluded that both normal and pathological thyroid tissues, with the exception of anaplastic carcinoma, express all TRs in thyreocites and that differentiated thyroid carcinomas are associated in enhancing the expression of TR(alpha2) mRNA.     

Wnt/beta-catenin signaling mediates antineoplastic effects of imatinib mesylate (gleevec) in anaplastic thyroid cancer

CONTEXT: Dysregulation of Wnt signaling is a key step in neoplastic thyrocyte proliferation. However, it is unclear whether the selective tyrosine kinase (TK) inhibitor, imatinib mesylate, is linked to the Wnt/beta-catenin cascade and is able to modulate the pathway. OBJECTIVE: Conflicting data are reported on the therapeutic effects of imatinib in anaplastic thyroid carcinomas (ATCs), but the molecular mechanism of action is unclear. Here, we further delineated the antitumor effects and the potential efficacy of imatinib in dedifferentiated thyroid carcinomas. RESULTS: Tissue microarray of histologically proven ATCs (n = 12) demonstrated that six of 12 tumors expressed at least one of the imatinib-sensitive TKs. Similarily, imatinib-sensitive TKs were detected in seven of 10 thyroid cancer cell lines derived from metastatic papillary, follicular, and ATCs. Coimmunoprecipitation in ARO cells demonstrated a direct link between c-abl and beta-catenin. Imatinib (10 microM for 48 h) drastically reduced beta-catenin expression and redistributed it from the nucleus to the cell membrane. It stabilized adherens junctions by increasing beta-catenin/E-cadherin binding and reduced the invasive potential of thyroid cancer. Furthermore, imatinib (10 microM for 48 h) attenuated T cell factor/lymphoid enhancer factor activity, reduced cyclin D1 levels and dose-dependently suppressed thyrocyte proliferation by half without affecting apoptosis. CONCLUSION: Our data provide a molecular mechanism for the antitumor activity of imatinib that may help to develop it as a therapeutic option in a subset of ATC patients.     

Transforming events in thyroid tumorigenesis and their association with follicular lesions

Thyroid tumors comprise a broad spectrum of neoplastic phenotypes, and distinct molecular events have been implicated in their pathogenesis. Pituitary tumor transforming gene, originally isolated from GH(4) pituitary cells, is tumorigenic in vivo, regulates basic fibroblast growth factor secretion, and is homologous to a securin inhibitor of chromatid separation. Pituitary tumor transforming gene 1 is expressed at low levels in several normal human tissues and is abundantly expressed in neoplasms, including colorectal carcinoma, where pituitary tumor transforming gene expression correlated highly with tumor invasiveness. As pituitary tumor transforming gene is regulated by E and as thyroid cancer shows a strong female preponderance, we examined pituitary tumor transforming gene 1 expression and action in human thyroid tumors and in normal human and rat thyroid cells. Increased pituitary tumor transforming gene 1 expression was evident early in thyroid tumors and was most abundantly expressed in a subset of thyroid hyperplasia, follicular adenomas, and follicular carcinomas (1.8-fold; P < 0.0001). Pituitary tumor transforming gene 1 overexpression in rat FRTL5 thyroid cells and in primary human thyroid cell cultures causes in vitro transformation and produces a dedifferentiated neoplastic phenotype. As pituitary tumor transforming gene 1 was abundantly overexpressed in follicular adenoma and follicular carcinoma, we propose that pituitary tumor transforming gene overexpression may play a role in the early molecular events leading to divergent development of follicular and papillary carcinoma.     

Determination of galectin-3 messenger ribonucleic Acid overexpression in papillary thyroid cancer by quantitative reverse transcription-polymerase chain reaction

Galectin-3, a lectin-family protein that appears to be involved in malignant transformation, has been reported to be an accurate immunohistochemical marker for thyroid cancer. However, immunohistochemistry is a subjective method that can be difficult to apply to cytologic specimens. Therefore, we sought to develop an objective and quantitative assay to measure galectin-3 mRNA in thyroid tissue to enhance potential clinical use of galectin-3 in the molecular analysis of thyroid nodules. In this study, total RNA from 37 snap-frozen thyroid tissue specimens was isolated from eight papillary and nine follicular thyroid cancers, six follicular adenomas, seven adenomatoid nodules, and seven normal thyroid lobes from patients undergoing thyroidectomy. Normalized levels of galectin-3 mRNA, expressed as picograms per nanogram GAPDH mRNA, were higher in papillary carcinomas (3327 pg/ng) and follicular adenomas (1314 pg/ng) than in thyroid normal tissue (426 pg/ng; P = 0.0012 and 0.032, respectively). Galectin-3 mRNA levels were also higher in papillary cancers than in adenomatoid nodules (P = 0.0012). However, galectin-3 mRNA levels were not statistically greater in follicular carcinomas than either normal tissue or follicular adenomas (P = 0.068 and 0.12, respectively). In summary, in comparison to galectin-3 immunohistochemistry, quantitative measurement of galectin-3 mRNA appears useful in the identification of papillary thyroid cancers (PTCs) but does not appear to be useful in distinguishing follicular carcinomas from follicular adenomas.     

Altered expression of cyclins and cell cycle inhibitors in papillary thyroid cancer: prognostic implications.

Currently we lack biochemical or molecular markers that predict recurrence and metastases in thyroid cancer. Recent studies in a number of other human malignancies indicate that expression and/or subcellular localization of certain cell cycle regulators has prognostic utility. We have investigated the expression of cyclins D1 and E and of cyclin-dependent kinase inhibitor's p21 and p27 in papillary thyroid cancer (PTC) and correlated this with clinical/histological stage at diagnosis and with clinical outcome. PTCs were compared to normal thyroid, adenomas, and undifferentiated thyroid cancers (UTCs). Our studies indicate that PTCs and UTCs demonstrate low nuclear expression of cyclin E and p27, allowing a clear distinction between adenomas and these carcinomas (p < 0.004). A pattern of low nuclear expression of all four markers was observed in stage IV PTCs and UTCs, while stage I PTCs had low D1 and E accompanied by high p21 or p27. Expression of cytoplasmic cyclin D1 was significantly lower in stage IV PTCs and UTCs than in stage I-III PTC's (p 相似文献   

Molecular profiling distinguishes papillary carcinoma from benign thyroid nodules

Recently we identified a molecular basis for differentiating benign and malignant follicular thyroid tumors. The purpose of these studies was to determine whether molecular analysis can be used to differentiate papillary thyroid carcinomas from benign thyroid nodules. Gene expression patterns of 14 papillary thyroid carcinomas and 21 benign tumors were analyzed by oligonucleotide array analysis. The carcinomas included seven classical papillary thyroid carcinomas (PTC) and seven follicular variant of PTC (FVPTC), and the benign tumors included 14 follicular adenomas and seven hyperplastic nodules. A hierarchical clustering analysis was performed to examine the groups for potential differences. The combined PTC and FVPTC groups had a distinct gene expression profile compared with the benign lesions. The sensitivity for a diagnosis of carcinoma was 93%, with a 100% specificity (one FVPTC clustered with the benign nodules). Cancer gene profiles contained both known (Met and galectin-3) and previously unidentified genes. Gene profiling is a reliable means of distinguishing PTC, FVPTC, and benign tumors of the thyroid. These gene profiles may provide insight into the pathogenesis of papillary thyroid carcinoma and may ultimately enhance the preoperative diagnosis of thyroid nodules on a molecular basis.