TSHR

Gene Summary

Gene:TSHR; thyroid stimulating hormone receptor
Aliases: LGR3, CHNG1, hTSHR-I
Location:14q31.1
Summary:The protein encoded by this gene is a membrane protein and a major controller of thyroid cell metabolism. The encoded protein is a receptor for thyrothropin and thyrostimulin, and its activity is mediated by adenylate cyclase. Defects in this gene are a cause of several types of hyperthyroidism. Three transcript variants encoding different isoforms have been found for this gene. [provided by RefSeq, Dec 2008]
Databases:OMIM, HGNC, Ensembl, GeneCard, Gene
Protein:thyrotropin receptor
Source:NCBIAccessed: 31 August, 2019

Ontology:

What does this gene/protein do?
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Pathways:What pathways are this gene/protein implicaed in?
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Cancer Overview

Research Indicators

Publications Per Year (1994-2019)
Graph generated 31 August 2019 using data from PubMed using criteria.

Literature Analysis

Mouse over the terms for more detail; many indicate links which you can click for dedicated pages about the topic.

  • Messenger RNA
  • Adenoma
  • DNA Mutational Analysis
  • Triiodothyronine
  • Cancer Gene Expression Regulation
  • BRAF
  • GTP-Binding Protein alpha Subunits, Gs
  • Mutation
  • Cyclic AMP
  • Papillary Carcinoma
  • Base Sequence
  • Thyroxine
  • Follicular Adenocarcinoma
  • Childhood Cancer
  • Hyperthyroidism
  • Symporters
  • Thyrotoxicosis
  • Thyroglobulin
  • Carcinoma
  • Thyroid Gland
  • Molecular Sequence Data
  • Cell Differentiation
  • Promoter Regions
  • Codon
  • p53 Protein
  • Point Mutation
  • DNA Methylation
  • Signal Transduction
  • Retinoic Acid
  • Transcription Factors
  • Thyroid Nodule
  • Chromosome 14
  • Biomarkers, Tumor
  • Polymorphism
  • Adolescents
  • Polymerase Chain Reaction
  • Receptors, Thyrotropin
  • Transcription
  • Thyroid Cancer, Papillary
  • Valine
Tag cloud generated 31 August, 2019 using data from PubMed, MeSH and CancerIndex

Specific Cancers (1)

Data table showing topics related to specific cancers and associated disorders. Scope includes mutations and abnormal protein expression.

Entity Topic PubMed Papers
-TSHR and Adenoma View Publications27

Note: list is not exhaustive. Number of papers are based on searches of PubMed (click on topic title for arbitrary criteria used).

Latest Publications: TSHR (cancer-related)

Nicolson NG, Murtha TD, Dong W, et al.
Comprehensive Genetic Analysis of Follicular Thyroid Carcinoma Predicts Prognosis Independent of Histology.
J Clin Endocrinol Metab. 2018; 103(7):2640-2650 [PubMed] Related Publications
Context: Follicular thyroid carcinoma (FTC) is classified into minimally invasive (miFTC), encapsulated angioinvasive (eaFTC), and widely invasive (wiFTC) subtypes, according to the 2017 World Health Organization guidelines. The genetic signatures of these subtypes may be crucial for diagnosis, prognosis, and treatment but have not been described.
Objective: Identify and describe the genetic underpinnings of subtypes of FTC.
Methods: Thirty-nine tumors, comprising 12 miFTCs, 17 eaFTCs, and 10 wiFTCs, were whole-exome sequenced and analyzed. Somatic mutations, constitutional sequence variants, somatic copy number alterations, and mutational signatures were described. Clinicopathologic parameters and mutational profiles were assessed for associations with patient outcomes.
Results: Total mutation burden was consistent across FTC subtypes, with a median of 10 (range 1 to 44) nonsynonymous somatic mutations per tumor. Overall, 20.5% of specimens had a mutation in the RAS subfamily (HRAS, KRAS, or NRAS), with no notable difference between subtypes. Mutations in TSHR, DICER1, EIF1AX, KDM5C, NF1, PTEN, and TP53 were also noted to be recurrent across the cohort. Clonality analysis demonstrated more subclones in wiFTC. Survival analysis demonstrated worse disease-specific survival in the eaFTC and wiFTC cohorts, with no recurrences or deaths for patients with miFTC. Mutation burden was associated with worse prognosis, independent of histopathological classification.
Conclusions: Though the number and variety of somatic variants are similar in the different histopathological subtypes of FTC in our study, mutational burden was an independent predictor of mortality and recurrence.

Shih YL, Huang YH, Lin KH, et al.
Identification of Functional Thyroid Stimulating Hormone Receptor and
Anticancer Res. 2018; 38(5):2793-2802 [PubMed] Related Publications
BACKGROUND/AIM: Extra-thyroid expression of thyroid stimulating hormone (TSH) receptor (TSHR) has been reported in normal liver tissues, but never assessed in hepatocellular carcinoma (HCC).
PATIENTS AND METHODS: Paired cancerous and non-cancerous HCC tissues were analyzed with TSHR expression assays. TSHR functional assessments and sequence analysis for the TSHR exon-10 were performed.
RESULTS: TSHR overexpression was found in 150/197 (76.1%) HCCs. Higher TSHR expression was associated with unfavorable postoperative outcomes. Immunohistochemical analysis revealed predominantly nuclei/peri-nuclei localization of TSHR in cancerous tissues but cell membrane localization in non-cancerous parts. TSH stimulation on hepatoma cells resulted in increased cyclic adenosine monophosphate levels with altered cell sensitivity to cisplatin. Gene mutations leading to TSHR truncation were detected in 8/81 (9.9%) HCC tissues.
CONCLUSION: Overexpression of TSHR was found in a great majority of HCC tissues and associated with unfavorable prognosis. Cell-based experiments and gene mutation analysis suggested that TSHR in HCCs was functional.

Mon SY, Riedlinger G, Abbott CE, et al.
Cancer risk and clinicopathological characteristics of thyroid nodules harboring thyroid-stimulating hormone receptor gene mutations.
Diagn Cytopathol. 2018; 46(5):369-377 [PubMed] Related Publications
BACKGROUND: Thyroid-stimulating hormone receptor (TSHR) gene mutations play a critical role in thyroid cell proliferation and function. They are found in 20%-82% of hyperfunctioning nodules, hyperfunctioning follicular thyroid cancers (FTC), and papillary thyroid cancers (PTC). The diagnostic importance of TSHR mutation testing in fine needle aspiration (FNA) specimens remains unstudied.
METHODS: To examine the association of TSHR mutations with the functional status and surgical outcomes of thyroid nodules, we evaluated 703 consecutive thyroid FNA samples with indeterminate cytology for TSHR mutations using next-generation sequencing. Testing for EZH1 mutations was performed in selected cases. The molecular diagnostic testing was done as part of standard of care treatment, and did not require informed consent.
RESULTS: TSHR mutations were detected in 31 (4.4%) nodules and were located in exons 281-640, with codon 486 being the most common. Allelic frequency ranged from 3% to 45%. Of 16 cases (12 benign, 3 FTC, 1 PTC) with surgical correlation, 15 had solitary TSHR mutations and 1 PTC had comutation with BRAF V600E. Hyperthyroidism was confirmed in all 3 FTC (2 overt, 1 subclinical). Of 5 nodules with solitary TSHR mutations detected at high allelic frequency, 3 (60%) were FTC. Those at low allelic frequency (3%-22%) were benign. EZH1 mutations were detected in 2 of 4 TSHR-mutant malignant nodules and neither of 2 benign nodules.
CONCLUSION: We report that TSHR mutations occur in ∼5% thyroid nodules in a large consecutive series with indeterminate cytology. TSHR mutations may be associated with an increased cancer risk when present at high allelic frequency, even when the nodule is hyperfunctioning. Benign nodules were however most strongly correlated with TSHR mutations at low allelic frequency.

Vastrad B, Vastrad C, Godavarthi A, Chandrashekar R
Molecular mechanisms underlying gliomas and glioblastoma pathogenesis revealed by bioinformatics analysis of microarray data.
Med Oncol. 2017; 34(11):182 [PubMed] Related Publications
The aim of this study was to identify key genes associated with gliomas and glioblastoma and to explore the related signaling pathways. Gene expression profiles of three glioma stem cell line samples, three normal astrocyte samples, three astrocyte overexpressing 4 iPSC-inducing and oncogenic factors (myc(T58A), OCT-4, p53DD, and H-Ras(G12V)) samples, three astrocyte overexpressing 7 iPSC-inducing and oncogenic factors (OCT4, H-Ras(G12V), myc(T58A), p53DD, cyclin D1, CDK4(RC24) and hTERT) samples and three glioblastoma cell line samples were downloaded from the ArrayExpress database (accession: E-MTAB-4771). The differentially expressed genes (DEGs) in gliomas and glioblastoma were identified using FDR and t tests, and protein-protein interaction (PPI) networks for these DEGs were constructed using the protein interaction network analysis. The GeneTrail2 1.5 tool was used to identify potentially enriched biological processes among the DEGs using gene ontology (GO) terms and to identify the related pathways using the Kyoto Encyclopedia of Genes and Genomes, Reactome and WikiPathways pathway database. In addition, crucial modules of the constructed PPI networks were identified using the PEWCC1 plug-in, and their topological properties were analyzed using NetworkAnalyzer, both available from Cytoscape. We also constructed microRNA-target gene regulatory network and transcription factor-target gene regulatory network for these DEGs were constructed using the miRNet and binding and expression target analysis. We identified 200 genes that could potentially be involved in the gliomas and glioblastoma. Among them, bioinformatics analysis identified 137 up-regulated and 63 down-regulated DEGs in gliomas and glioblastoma. The significant enriched pathway (PI3K-Akt) for up-regulated genes such as COL4A1, COL4A2, EGFR, FGFR1, LAPR6, MYC, PDGFA, SPP1 were selected as well as significant GO term (ear development) for up-regulated genes such as CELSR1, CHRNA9, DDR1, FGFR1, GLI2, LGR5, SOX2, TSHR were selected, while the significant enriched pathway (amebiasis) for down-regulated gene such as COL3A1, COL5A2, LAMA2 were selected as well as significant GO term (RNA polymerase II core promoter proximal region sequence-specific binding (5) such as MEIS2, MEOX2, NR2E1, PITX2, TFAP2B, ZFPM2 were selected. Importantly, MYC and SOX2 were hub proteins in the up-regulated PPI network, while MET and CDKN2A were hub proteins in the down-regulated PPI network. After network module analysis, MYC, FGFR1 and HOXA10 were selected as the up-regulated coexpressed genes in the gliomas and glioblastoma, while SH3GL3 and SNRPN were selected as the down-regulated coexpressed genes in the gliomas and glioblastoma. MicroRNA hsa-mir-22-3p had a regulatory effect on the most up DEGs, including VSNL1, while hsa-mir-103a-3p had a regulatory effect on the most down DEGs, including DAPK1. Transcription factor EZH2 had a regulatory effect on the both up and down DEGs, including CD9, CHI3L1, MEIS2 and NR2E1. The DEGs, such as MYC, FGFR1, CDKN2A, HOXA10 and MET, may be used for targeted diagnosis and treatment of gliomas and glioblastoma.

Khatami F, Larijani B, Heshmat R, et al.
Meta-analysis of promoter methylation in eight tumor-suppressor genes and its association with the risk of thyroid cancer.
PLoS One. 2017; 12(9):e0184892 [PubMed] Free Access to Full Article Related Publications
Promoter methylation in a number of tumor-suppressor genes (TSGs) can play crucial roles in the development of thyroid carcinogenesis. The focus of the current meta-analysis was to determine the impact of promoter methylation of eight selected candidate TSGs on thyroid cancer and to identify the most important molecules in this carcinogenesis pathway. A comprehensive search was performed using Pub Med, Scopus, and ISI Web of Knowledge databases, and eligible studies were included. The methodological quality of the included studies was evaluated according to the Newcastle Ottawa scale table and pooled odds ratios (ORs); 95% confidence intervals (CIs) were used to estimate the strength of the associations with Stata 12.0 software. Egger's and Begg's tests were applied to detect publication bias, in addition to the "Metatrim" method. A total of 55 articles were selected, and 135 genes with altered promoter methylation were found. Finally, we included eight TSGs that were found in more than four studies (RASSF1, TSHR, PTEN, SLC5A, DAPK, P16, RARβ2, and CDH1). The order of the pooled ORs for these eight TSGs from more to less significant was CDH1 (OR = 6.73), SLC5 (OR = 6.15), RASSF1 (OR = 4.16), PTEN (OR = 3.61), DAPK (OR = 3.51), P16 (OR = 3.31), TSHR (OR = 2.93), and RARβ2 (OR = 1.50). Analyses of publication bias and sensitivity confirmed that there was very little bias. Thus, our findings showed that CDH1 and SCL5A8 genes were associated with the risk of thyroid tumor genesis.

Slattery ML, Herrick JS, Mullany LE, et al.
The co-regulatory networks of tumor suppressor genes, oncogenes, and miRNAs in colorectal cancer.
Genes Chromosomes Cancer. 2017; 56(11):769-787 [PubMed] Free Access to Full Article Related Publications
Tumor suppressor genes (TSGs) and oncogenes (OG) are involved in carcinogenesis. MiRNAs also contribute to cellular pathways leading to cancer. We use data from 217 colorectal cancer (CRC) cases to evaluate differences in TSGs and OGs expression between paired CRC and normal mucosa and evaluate how TSGs and OGs are associated with miRNAs. Gene expression data from RNA-Seq and miRNA expression data from Agilent Human miRNA Microarray V19.0 were used. We focus on genes most strongly associated with CRC (fold change (FC) of ≥1.5 or ≤0.67) that were statistically significant after adjustment for multiple comparisons. Of the 74 TSGs evaluated, 22 were associated with carcinoma/normal mucosa differential expression. Ten TSGs were up-regulated (FAM123B, RB1, TP53, RUNX1, MSH2, BRCA1, BRCA2, SOX9, NPM1, and RNF43); six TSGs were down-regulated (PAX5, IZKF1, GATA3, PRDM1, TET2, and CYLD); four were associated with MSI tumors (MLH1, PTCH1, and CEBPA down-regulated and MSH6 up-regulated); and two were associated with MSS tumors (PHF6 and ASXL1 up-regulated). Thirteen of these TSGs were associated with 44 miRNAs. Twenty-seven of the 59 OGs evaluated were dysregulated: 14 down-regulated (KLF4, BCL2, SSETBP1, FGFR2, TSHR, MPL, KIT, PDGFRA, GNA11, GATA2, FGFR3, AR, CSF1R, and JAK3), seven up-regulated (DNMT1, EZH2, PTPN11, SKP2, CCND1, MET, and MYC); three down-regulated for MSI (FLT3, CARD11, and ALK); two up-regulated for MSI (IDH2 and HRAS); and one up-regulated with MSS tumors (CTNNB1). These findings suggest possible co-regulatory function between TSGs, OGs, and miRNAs, involving both direct and indirect associations that operate through feedback and feedforward loops.

Kronborg TM, Hansen JF, Rasmussen ÅK, et al.
The flame retardant DE-71 (a mixture of polybrominated diphenyl ethers) inhibits human differentiated thyroid cell function in vitro.
PLoS One. 2017; 12(6):e0179858 [PubMed] Free Access to Full Article Related Publications
BACKGROUND: Normal thyroid function is essential for general growth and metabolism, but can be affected by endocrine disrupting chemicals (EDCs). Polybrominated diphenyl ethers (PBDEs) have been used worldwide to reduce flammability in different materials and are suspected to be EDCs. The production of the commercial Penta- and OctaBDE mixtures is banned, but DecaBDEs and existing products may leak PBDEs into the environment. Our aim was to investigate the effect of the PentaBDE mixture DE-71 on human thyroid cells in vitro.
MATERIALS AND METHODS: Primary human thyroid cells were obtained as paraadenomatous tissue and cultured in monolayers. The influence of DE-71 on cyclic adenosine monophosphate (cAMP) and thyroglobulin (Tg) production was examined in the culture medium by competitive radioimmunoassay and enzyme-linked immunosorbent assay, respectively. Real-time quantitative PCR analysis of thyroid-specific genes was performed on the exposed cell cultures. PBDE concentrations were determined in cellular and supernatant fractions of the cultures.
RESULTS: DE-71 inhibited Tg-release from TSH-stimulated thyrocytes. At 50 mg/L DE-71, mean Tg production was reduced by 71.9% (range: 8.5-98.7%), and cAMP by 95.1% (range: 91.5-98.8%) compared to controls). Expression of mRNA encoding Tg, TPO and TSHr were significantly inhibited (p<0.0001, p = 0.0079, and p = 0.0002, respectively). The majority of DE-71 added was found in the cell fraction. No cytotoxicity was found.
CONCLUSIONS: DE-71 inhibited differentiated thyroid cell functions in a two phase response manner and a concentration-dependent inhibition of Tg and cAMP production, respectively, as well as expression of mRNA encoding Tg, TPO and TSHr. Our findings suggest an inhibiting effect of PBDEs on thyroid cells.

Stephen JK, Chen KM, Merritt J, et al.
Methylation markers differentiate thyroid cancer from benign nodules.
J Endocrinol Invest. 2018; 41(2):163-170 [PubMed] Related Publications
PURPOSE: The incidence of thyroid cancer (TC) is increasing. Cytology by itself cannot distinguish TC from some benign nodules especially in certain subtypes of TC. Our immediate goal is to identify DNA methylation markers for early detection of TC and to molecularly differentiate TC subtypes from benign nodules.
METHODS: Promoter methylation status of 21 candidate genes was examined on formalin-fixed paraffin-embedded tissue (FFPE) utilizing quantitative methylation-specific polymerase chain reaction (QMSP) in a retrospective cohort of 329 patients (56% white, 29% African American, 61% female) comprising 71 normal thyroid, 83 benign nodules [follicular adenomas (FA)], 90 follicular TC (FTC) and 85 papillary TC (PTC). All genes were analyzed individually (Kruskal-Wallis and Wilcoxon rank sum tests) and in combination (logistic regression models) to identify genes whose methylation levels might best separate groups.
RESULTS: Combination gene panels TPO and UCHL1 (ROC = 0.607, sensitivity 78%) discriminated FTC from FA, and RASSF1 and TPO (ROC = 0.881, sensitivity 78%) discriminated FTC from normal. Methylation of TSHR distinguished PTC from FTC (ROC = 0.701, sensitivity 84%) and PTC from FA (ROC = 0.685, sensitivity 70%). The six gene panel of TIMP3, RARB2, SERPINB5, RASSF1, TPO and TSHR, which differentiates PTC from normal thyroid, had the best combination sensitivity (91%) and specificity (81%) of the panels addressing discrimination of cancer tissue.
CONCLUSIONS: Aberrant gene methylation used in combination panels may be useful clinically in differentiating FTC and PTC from benign nodules. If confirmed in additional studies, these findings could help reduce the over diagnosis of thyroid cancer and surgeries related to over diagnosis.

Sciacchitano S, Lavra L, Ulivieri A, et al.
Comparative analysis of diagnostic performance, feasibility and cost of different test-methods for thyroid nodules with indeterminate cytology.
Oncotarget. 2017; 8(30):49421-49442 [PubMed] Free Access to Full Article Related Publications
Since it is impossible to recognize malignancy at fine needle aspiration (FNA) cytology in indeterminate thyroid nodules, surgery is recommended for all of them. However, cancer rate at final histology is <30%. Many different test-methods have been proposed to increase diagnostic accuracy in such lesions, including Galectin-3-ICC (GAL-3-ICC), BRAF mutation analysis (BRAF), Gene Expression Classifier (GEC) alone and GEC+BRAF, mutation/fusion (M/F) panel, alone, M/F panel+miRNA GEC, and M/F panel by next generation sequencing (NGS), FDG-PET/CT, MIBI-Scan and TSHR mRNA blood assay.We performed systematic reviews and meta-analyses to compare their features, feasibility, diagnostic performance and cost. GEC, GEC+BRAF, M/F panel+miRNA GEC and M/F panel by NGS were the best in ruling-out malignancy (sensitivity = 90%, 89%, 89% and 90% respectively). BRAF and M/F panel alone and by NGS were the best in ruling-in malignancy (specificity = 100%, 93% and 93%). The M/F by NGS showed the highest accuracy (92%) and BRAF the highest diagnostic odds ratio (DOR) (247). GAL-3-ICC performed well as rule-out (sensitivity = 83%) and rule-in test (specificity = 85%), with good accuracy (84%) and high DOR (27) and is one of the cheapest (113 USD) and easiest one to be performed in different clinical settings.In conclusion, the more accurate molecular-based test-methods are still expensive and restricted to few, highly specialized and centralized laboratories. GAL-3-ICC, although limited by some false negatives, represents the most suitable screening test-method to be applied on a large-scale basis in the diagnostic algorithm of indeterminate thyroid lesions.

Weinberger P, Ponny SR, Xu H, et al.
Cell Cycle M-Phase Genes Are Highly Upregulated in Anaplastic Thyroid Carcinoma.
Thyroid. 2017; 27(2):236-252 [PubMed] Free Access to Full Article Related Publications
BACKGROUND: Anaplastic thyroid carcinoma (ATC) accounts for only 3% of thyroid cancers, yet strikingly, it accounts for almost 40% of thyroid cancer deaths. Currently, no effective therapies exist. In an effort to identify ATC-specific therapeutic targets, we analyzed global gene expression data from multiple studies to identify ATC-specific dysregulated genes.
METHODS: The National Center for Biotechnology Information Gene Expression Omnibus database was searched for high-throughput gene expression microarray studies from human ATC tissue along with normal thyroid and/or papillary thyroid cancer (PTC) tissue. Gene expression levels in ATC were compared with normal thyroid or PTC using seven separate comparisons, and an ATC-specific gene set common in all seven comparisons was identified. We investigated these genes for their biological functions and pathways.
RESULTS: There were three studies meeting inclusion criteria, (including 32 ATC patients, 69 PTC, and 75 normal). There were 259 upregulated genes and 286 downregulated genes in ATC with at least two-fold change in all seven comparisons. Using a five-fold filter, 36 genes were upregulated in ATC, while 40 genes were downregulated. Of the 10 top globally upregulated genes in ATC, 4/10 (MMP1, ANLN, CEP55, and TFPI2) are known to play a role in ATC progression; however, 6/10 genes (TMEM158, CXCL5, E2F7, DLGAP5, MME, and ASPM) had not been specifically implicated in ATC. Similarly, 3/10 (SFTA3, LMO3, and C2orf40) of the most globally downregulated genes were novel in this context, while 7/10 genes (SLC26A7, TG, TSHR, DUOX2, CDH1, PDE8B, and FOXE1) have been previously identified in ATC. We experimentally validated a significant correlation for seven transcription factors (KLF16, SP3, ETV6, FOXC1, SP1, EGFR1, and MAFK) with the ATC-specific genes using microarray analysis of ATC cell lines. Ontology clustering of globally altered genes revealed that "mitotic cell cycle" is highly enriched in the globally upregulated gene set (44% of top upregulated genes, p-value <10
CONCLUSIONS: By focusing on globally altered genes, we have identified a set of consistently altered biological processes and pathways in ATC. Our data are consistent with an important role for M-phase cell cycle genes in ATC, and may provide direction for future studies to identify novel therapeutic targets for this disease.

Liu TR, Su X, Qiu WS, et al.
Thyroid-stimulating hormone receptor affects metastasis and prognosis in papillary thyroid carcinoma.
Eur Rev Med Pharmacol Sci. 2016; 20(17):3582-91 [PubMed] Related Publications
OBJECTIVE: Although endocrine therapy of papillary thyroid carcinoma (PTC) by inhibiting thyroid-stimulating hormone (TSH) has been used for many years, its mechanism of action is not clear. This study aimed to explore the expression and role of TSH receptor (TSHR) in PTC, to provide a theoretical basis for optimization of endocrine treatment options in PTC.
PATIENTS AND METHODS: Expression of TSHR was tested by immunohistochemistry of tissues from 150 cases of PTC and 21 normal thyroid tissues. Survival analysis was performed by Kaplan-Meier and log-rank analyses, and multivariate analysis was done using a Cox model. The regulatory effects of the TSH-TSHR signal transduction pathway on differentiated thyroid carcinoma cells were explored in vitro.
RESULTS: The positive expression rate of TSHR in PTC was 68% (102/150). TSHR expression was an independent factor affecting the prognosis of PTC patients aged > 45 years (p = 0.006), and TSHR might have a role in decreasing distant metastasis (p = 0.024). In vitro experiments showed that up-regulation of TSHR promoted apoptosis of thyroid cancer cells and inhibited metastasis significantly. There was no significant regulatory effect of the TSH-TSHR signal transduction pathway on the proliferation of thyroid carcinoma cells.
CONCLUSIONS: TSHR expression is an independent factor that affects the prognosis of PTC patients, and might decrease distant metastasis in patient aged > 45 years. Up-regulation of TSHR could inhibit metastasis and promote apoptosis in PTC cells.

Jung SH, Kim MS, Jung CK, et al.
Mutational burdens and evolutionary ages of thyroid follicular adenoma are comparable to those of follicular carcinoma.
Oncotarget. 2016; 7(43):69638-69648 [PubMed] Free Access to Full Article Related Publications
Follicular thyroid adenoma (FTA) precedes follicular thyroid carcinoma (FTC) by definition with a favorable prognosis compared to FTC. However, the genetic mechanism of FTA to FTC progression remains unknown. For this, it is required to disclose FTA and FTC genomes in mutational and evolutionary perspectives. We performed whole-exome sequencing and copy number profiling of 14 FTAs and 13 FTCs, which exhibited previously-known gene mutations (NRAS, HRAS, BRAF, TSHR and EIF1AX) and copy number alterations (CNAs) (22q loss and 1q gain) in follicular tumors. In addition, we found eleven potential cancer-related genes with mutations (EZH1, SPOP, NF1, TCF12, IGF2BP3, KMT2C, CNOT1, BRIP1, KDM5C, STAG2 and MAP4K3) that have not been reported in thyroid follicular tumors. Of note, FTA genomes showed comparable levels of mutations to FTC in terms of the number, sequence composition and functional consequences (potential driver mutations) of mutations. Analyses of evolutionary ages using somatic mutations as molecular clocks further identified that FTA genomes were as old as FTC genomes. Whole-transcriptome sequencing did not find any gene fusions with potential significance. Our data indicate that FTA genomes may be as old as FTC genomes, thus suggesting that follicular thyroid tumor genomes during the transition from FTA to FTC may stand stable at genomic levels in contrast to the discernable changes at pathologic and clinical levels. Also, the data suggest a possibility that the mutational profiles obtained from early biopsies may be useful for the molecular diagnosis and therapeutics of follicular tumor patients.

Calebiro D, Grassi ES, Eszlinger M, et al.
Recurrent EZH1 mutations are a second hit in autonomous thyroid adenomas.
J Clin Invest. 2016; 126(9):3383-8 [PubMed] Free Access to Full Article Related Publications
Autonomous thyroid adenomas (ATAs) are a frequent cause of hyperthyroidism. Mutations in the genes encoding the TSH receptor (TSHR) or the Gs protein α subunit (GNAS) are found in approximately 70% of ATAs. The involvement of other genes and the pathogenesis of the remaining cases are presently unknown. Here, we performed whole-exome sequencing in 19 ATAs that were paired with normal DNA samples and identified a recurrent hot-spot mutation (c.1712A>G; p.Gln571Arg) in the enhancer of zeste homolog 1 (EZH1) gene, which codes for a catalytic subunit of the polycomb complex. Targeted screening in an independent cohort confirmed that this mutation occurs with high frequency (27%) in ATAs. EZH1 mutations were strongly associated with known (TSHR, GNAS) or presumed (adenylate cyclase 9 [ADCY9]) alterations in cAMP pathway genes. Furthermore, functional studies revealed that the p.Gln571Arg EZH1 mutation caused increased histone H3 trimethylation and increased proliferation of thyroid cells. In summary, this study revealed that a hot-spot mutation in EZH1 is the second most frequent genetic alteration in ATAs. The association between EZH1 and TSHR mutations suggests a 2-hit model for the pathogenesis of these tumors, whereby constitutive activation of the cAMP pathway and EZH1 mutations cooperate to induce the hyperproliferation of thyroid cells.

Hsu YC, Liu CL, Yang PS, et al.
Interaction of Age at Diagnosis with Transcriptional Profiling in Papillary Thyroid Cancer.
World J Surg. 2016; 40(12):2922-2929 [PubMed] Related Publications
BACKGROUND: Age is an important prognostic factor for papillary thyroid cancer (PTC). However, little is known about why advanced age is associated with poor prognosis. The study investigated the changes in transcriptional profiling related to age.
METHODS: RNA sequencing data of PTC samples were retrieved from The Cancer Genome Atlas data portal. Spearman's correlation was used to test the association between age and gene expression. Correlation in the same direction to disease severity was considered functionally relevant. Functional enrichment analysis and pathway annotations were performed.
RESULTS: There was no correlation between age and thyroid-specific genes, except for a weak, negative association between age and TSHR expression. Among 272 genes with a positive association between gene expression and age, the most prominent alteration was metabolic pathways, particularly glycolysis. Among 482 genes with a negative association between gene expression and age, the most enriched biological process was immune-related functions, particularly natural killer cell-mediated cytotoxicity.
CONCLUSIONS: Our analysis characterized the age-associated molecular landscape in PTC. Metabolic alterations and immune dysregulation are probable mechanisms involving in worse prognosis in older patients with PTC.

Hart SN, Ellingson MS, Schahl K, et al.
Determining the frequency of pathogenic germline variants from exome sequencing in patients with castrate-resistant prostate cancer.
BMJ Open. 2016; 6(4):e010332 [PubMed] Free Access to Full Article Related Publications
OBJECTIVES: To determine the frequency of pathogenic inherited mutations in 157 select genes from patients with metastatic castrate-resistant prostate cancer (mCRPC).
DESIGN: Observational.
SETTING: Multisite US-based cohort.
PARTICIPANTS: Seventy-one adult male patients with histological confirmation of prostate cancer, and had progressive disease while on androgen deprivation therapy.
RESULTS: Twelve patients (17.4%) showed evidence of carrying pathogenic or likely pathogenic germline variants in the ATM, ATR, BRCA2, FANCL, MSR1, MUTYH, RB1, TSHR and WRN genes. All but one patient opted in to receive clinically actionable results at the time of study initiation. We also found that pathogenic germline BRCA2 variants appear to be enriched in mCRPC compared to familial prostate cancers.
CONCLUSIONS: Pathogenic variants in cancer-susceptibility genes are frequently observed in patients with mCRPC. A substantial proportion of patients with mCRPC or their family members would derive clinical utility from mutation screening.
TRIAL REGISTRATION NUMBER: NCT01953640; Results.

Seagle BL, Eng KH, Yeh JY, et al.
Discovery of candidate tumor biomarkers for treatment with intraperitoneal chemotherapy for ovarian cancer.
Sci Rep. 2016; 6:21591 [PubMed] Free Access to Full Article Related Publications
Tumor mRNA expression was used to discover genes associated with worse survival or no survival benefit after intraperitoneal (IP) chemotherapy. Data for high grade serous ovarian cancer patients treated with IP (n = 90) or IV-only (n =  398) chemotherapy was obtained from The Cancer Genome Atlas. Progression free survival (PFS) and overall survival (OS) were compared between IP and IV groups using Kaplan-Meier analysis and Cox regression. Validations were performed by analyses of microarray and RNA-Seq mRNA expression data. PFS and OS were compared between IP and IV groups by permutation testing stratified by gene expression. P-values are two-tailed. IP chemotherapy increased PFS (26.7 vs 16.0 months, HR 0.43 (0.28-0.66), p = 0.0001) and OS (49.6 vs 38.2 months, HR 0.46 (0.25-0.83), p = 0.01). Increased expression of NCAM2 and TSHR and decreased expression of GCNT1 was associated with decreased PFS and OS after IV chemotherapy (p < 0.05). High tumor expression of LMAN2, FZD4, FZD5, or STT3A was associated with no significant PFS increase after IP compared to IV chemotherapy. Low expression of APC2 and high expression of FUT9 was associated with 5.5 and 7.2 months, respectively, decreased OS after IP compared to IV chemotherapy (p ≤ 0.007).

Pagan M, Kloos RT, Lin CF, et al.
The diagnostic application of RNA sequencing in patients with thyroid cancer: an analysis of 851 variants and 133 fusions in 524 genes.
BMC Bioinformatics. 2016; 17 Suppl 1:6 [PubMed] Free Access to Full Article Related Publications
BACKGROUND: Thyroid carcinomas are known to harbor oncogenic driver mutations and advances in sequencing technology now allow the detection of these in fine needle aspiration biopsies (FNA). Recent work by The Cancer Genome Atlas (TCGA) Research Network has expanded the number of genetic alterations detected in papillary thyroid carcinomas (PTC). We sought to investigate the prevalence of these and other genetic alterations in diverse subtypes of thyroid nodules beyond PTC, including a variety of samples with benign histopathology. This is the first clinical evaluation of a large panel of TCGA-reported genomic alterations in thyroid FNAs.
RESULTS: In FNAs, genetic alterations were detected in 19/44 malignant samples (43% sensitivity) and in 7/44 histopathology benign samples (84% specificity). Overall, after adding a cohort of tissue samples, 38/76 (50%) of histopathology malignant samples were found to harbor a genetic alteration, while 15/75 (20%) of benign samples were also mutated. The most frequently mutated malignant subtypes were medullary thyroid carcinoma (9/12, 75%) and PTC (14/30, 47%). Additionally, follicular adenoma, a benign subtype of thyroid neoplasm, was also found to harbor mutations (12/29, 41%). Frequently mutated genes in malignant samples included BRAF (20/76, 26%) and RAS (9/76, 12%). Of the TSHR variants detected, (6/7, 86%) were in benign nodules. In a direct comparison of the same FNA also tested by an RNA-based gene expression classifier (GEC), the sensitivity of genetic alterations alone was 42%, compared to the 91% sensitivity achieved by the GEC. The specificity based only on genetic alterations was 84%, compared to 77% specificity with the GEC.
CONCLUSIONS: While the genomic landscape of all thyroid neoplasm subtypes will inevitably be elucidated, caution should be used in the early adoption of published mutations as the sole predictor of malignancy in thyroid. The largest set of such mutations known to date detects only a portion of thyroid carcinomas in preoperative FNAs in our cohort and thus is not sufficient to rule out cancer. Due to the finding that variants are also found in benign nodules, testing only GEC suspicious nodules may be helpful in avoiding false positives and altering the extent of treatment when selected mutations are found.

Kartal K, Onder S, Kosemehmetoglu K, et al.
Methylation status of TSHr in well-differentiated thyroid cancer by using cytologic material.
BMC Cancer. 2015; 15:824 [PubMed] Free Access to Full Article Related Publications
BACKGROUND: The role of methylation status of the thyroid stimulating hormone receptor gene (TSHr) in the discrimination of benign and malignant thyroid nodules has already been studied using paraffin blocks and cell lines. As cytological sampling plays an important role in assessment of thyroidal nodules, we have investigated the potential clinical use of TSHr methylation status of fine needle aspiration specimens reported according to Bethesda System.
METHOD: Sixty nine patients who had both cytological and pathological diagnosis of the same nodule were selected. Four groups were composed according to cytological and pathological diagnoses: Benign (B), papillary thyroid carcinoma (PTC), atypia of unknown significance (AUS) and follicular neoplasia (FN). The latter 2 groups were further sub-classified into 2 as benign (AUS-B and FN-B) and malignant (AUS-M and FN-M) according to final pathological diagnosis. DNAs were isolated from the fine needle aspiration cytology specimens and the methylation status of TSHr promotor region was investigated by using methylation specific polymerase chain reaction.
RESULTS: Overall, TSHr methylation was present in 58% of cases; 71% of malignant and 46% of benign nodules. PTC group showed the highest TSHr methylation rate (87%), followed by 61% in AUS, 44% in B, and 30% in FN (p = 0.016). TSHr methylation rate was significantly higher in PTC group when compared to B (p = 0.013) and FN-B (p = 0.004) groups; but not in FN-M (p = 0.115) or AUS (p = 0.096) groups. All 9 cases of papillary thyroid carcinoma with lymph node metastasis showed TSHr methylation. Positive predictive value, negative predictive value, sensitivity and specificity of TSHr methylation in determination of malignancy were calculated as 60, 66, 71 and 54%, respectively.
CONCLUSION: The eminent ratio of TSHr methylation in well-differentiated thyroid carcinoma against benign thyroidal nodules adduced that TSHr methylation status can be utilized as a tumor marker for well-differentiated thyroid cancer; however, it has a limited value. The determination of methylation status of TSHr gene had no efficiency on decision of the malignant potential for the nodules which are cytologically classified as atypia of undetermined significance.

Liu M, Watson LT, Zhang L
HMMvar-func: a new method for predicting the functional outcome of genetic variants.
BMC Bioinformatics. 2015; 16:351 [PubMed] Free Access to Full Article Related Publications
BACKGROUND: Numerous tools have been developed to predict the fitness effects (i.e., neutral, deleterious, or beneficial) of genetic variants on corresponding proteins. However, prediction in terms of whether a variant causes the variant bearing protein to lose the original function or gain new function is also needed for better understanding of how the variant contributes to disease/cancer. To address this problem, the present work introduces and computationally defines four types of functional outcome of a variant: gain, loss, switch, and conservation of function. The deployment of multiple hidden Markov models is proposed to computationally classify mutations by the four functional impact types.
RESULTS: The functional outcome is predicted for over a hundred thyroid stimulating hormone receptor (TSHR) mutations, as well as cancer related mutations in oncogenes or tumor suppressor genes. The results show that the proposed computational method is effective in fine grained prediction of the functional outcome of a mutation, and can be used to help elucidate the molecular mechanism of disease/cancer causing mutations. The program is freely available at http://bioinformatics.cs.vt.edu/zhanglab/HMMvar/download.php.
CONCLUSION: This work is the first to computationally define and predict functional impact of mutations, loss, switch, gain, or conservation of function. These fine grained predictions can be especially useful for identifying mutations that cause or are linked to cancer.

Bastos AU, Oler G, Nozima BH, et al.
BRAF V600E and decreased NIS and TPO expression are associated with aggressiveness of a subgroup of papillary thyroid microcarcinoma.
Eur J Endocrinol. 2015; 173(4):525-40 [PubMed] Related Publications
BACKGROUND: Thyroid cancer incidence has dramatically increased worldwide over the last two decades. The rise is mostly due to an increased detection of small papillary thyroid carcinomas (PTCs) (≤20  mm), predominantly microPTC (≤10  mm). Although small tumors generally have an excellent outcome, a considerable percentage may have a more aggressive disease and worse prognosis. The clinical challenge is to preoperatively identify those tumors that are more likely to recur.
AIM: To improve risk stratification and patient management, we sought to determine the prognostic value of BRAF V600E, NRAS or RET/PTC mutations in patients with PTC measuring <20  mm, mainly microPTC.
METHODS: The prevalence of RET/PTC fusion genes was examined by quantitative RT-PCR. BRAF V600E and NRAS Q61 mutations were determined by PCR sequencing. To further elucidate why some small PTC are less responsive to radioactive iodine treatment therapy, we explored if these genetic alterations may modulate the expression of iodine metabolism genes (NIS, TPO, TG, TSHR and PDS) and correlated with clinico-pathological findings that are predictors of recurrence.
RESULTS: This study shows that tumors measuring ≤20  mm exhibited higher prevalence of BRAF V600E mutation, which correlated with aggressive histopathological parameters, higher risk of recurrence, and lower expression of NIS and TPO. Although this correlation was not found when microPTC were evaluated, we show that tumors measuring 7-10  mm, which were positive for BRAF mutation, presented more aggressive features and lower expression of NIS and TPO.
CONCLUSION: We believe that our findings will help to decide the realistic usefulness of BRAF V600E mutation as a preoperative marker of poor prognosis in small PTC, primarily in microPTC.

Zheng H, Wang M, Jiang L, et al.
BRAF-Activated Long Noncoding RNA Modulates Papillary Thyroid Carcinoma Cell Proliferation through Regulating Thyroid Stimulating Hormone Receptor.
Cancer Res Treat. 2016; 48(2):698-707 [PubMed] Free Access to Full Article Related Publications
PURPOSE: The importance of long noncoding RNAs (lncRNAs) in tumorigenesis has recently been demonstrated. However, the role of lncRNAs in development of thyroid cancer remains largely unknown.
MATERIALS AND METHODS: Using quantitative reverse transcription polymerase chain reaction, expression of three lncRNAs, including BRAF-activated long noncoding RNA (BANCR), papillary thyroid cancer susceptibility candidate 3 (PTCSC3), and noncoding RNA associated with mitogen-activated protein kinase pathway and growth arrest (NAMA), was investigated in the current study.
RESULTS: Of the three lncRNAs (BANCR, PTCSC3, and NAMA), expression of BANCR was significantly up-regulated while PTCSC3 and NAMA were significantly down-regulated in papillary thyroid carcinoma (PTC) compared to that in normal tissue. BANCR-knockdown in a PTC-derived cell line (IHH-4) resulted in significant suppression of thyroid stimulating hormone receptor (TSHR). BANCR-knockdown also led to inhibition of cell growth and cell cycle arrest at G0/G1 phase through down-regulation of cyclin D1. In addition, BANCR was enriched by polycomb enhancer of zeste homolog 2 (EZH2), and silencing BANCR led to decreased chromatin recruitment of EZH2, which resulted significantly reduced expression of TSHR.
CONCLUSION: These findings indicate that BANCR may contribute to the tumorigenesis of PTC through regulation of cyclin D1 and TSHR.

Tong GX, Mody K, Wang Z, et al.
Mutations of TSHR and TP53 Genes in an Aggressive Clear Cell Follicular Carcinoma of the Thyroid.
Endocr Pathol. 2015; 26(4):315-9 [PubMed] Related Publications
Clear cell follicular carcinoma is a rare type of thyroid cancer and some with aggressive biological behavior. The cytoplasmic clearing of the neoplastic cells has been attributed to the accumulation of various substances, such as glycogen, lipid, mucin, and thyroglobulin, or distension of mitochondria or endoplasmic reticulum. However, the molecular mechanisms responsible for the characteristic appearance of the cell cytoplasm and the biological behavior remain unknown. We report here a case of aggressive clear cell follicular carcinoma of the thyroid with molecular profile using targeted next generation sequencing (NGS) that presented as a metastatic tumor in a woman with a history of breast carcinoma. The NGS data revealed the coexisting of a well-characterized loss-of-function TP53 R248Q mutation and a putative gain-of-function mutation of TSHR L272V, which was suggested by the overexpression of thyroglobulin and SLC5A5 (NIS) genes in this tumor. TP53 mutations are usually related with dedifferentiation, progression, and metastasis of thyroid carcinomas. Identification of TP53 R248Q in this tumor correlated with its aggressive clinical behavior. Gain-of-function mutation of TSHR can overstimulate the thyroid follicular cells as the elevated level of TSH does and might have contributed to the development of clear cell morphology in this tumor. This report represents the first case of clear cell follicular carcinoma of the thyroid with NGS analysis and more molecular characterization is needed to elucidate the pathogenesis and provide more prognosis-relevant information for this uncommon variant of thyroid carcinomas.

Campennì A, Giovinazzo S, Curtò L, et al.
Thyroid hemiagenesis, Graves' disease and differentiated thyroid cancer: a very rare association: case report and review of literature.
Hormones (Athens). 2015 Jul-Sep; 14(3):451-8 [PubMed] Related Publications
OBJECTIVE: Thyroid hemiagenesis is a rare congenital disorder characterized by the absence of a lobe and/or of isthmus. Studies on the association between thyroid hemiagenesis, Graves' disease and differentiated thyroid cancer are rare.
CASE PRESENTATION: We describe the medical and surgical history of a patient in whom a molecular evaluation was performed. A 36-year-old man presented with symptoms and signs of hyperthyroidism of a few months' duration. Hyperthyroidism was confirmed biochemically and anti-TSH-receptor antibodies were positive. Thyroid ultrasonography showed no left lobe and demonstrated a diffused enlargement of the right lobe; an ipoechoic, non-homogenous nodule 15 millimeters in size was identified in the middle part of the lobe. A 99mTc-pertechnetate thyroid scintigraphy (111 MBq) confirmed thyroid hemiagenesis due to the absence of the left lobe. Treatment with methimazole (30 mg/day) was started. As the patient's hyperthyroidism improved, he underwent fine-needle needle aspiration cytology (FNAC) of the right nodule. Cytology was suspicious for malignancy (THY4) and the patient was referred for surgery. Histopathological findings revealed a papillary thyroid carcinoma. The molecular analysis did not show PAX8 or TSHR mutations in the thyroid tissue nor mutations of BRAF, H-RAS, N-RAS or K-RAS genes in the tumor.
CONCLUSION: Though thus far studies on the association of thyroid hemiagenesis, Graves' disease and differentiated thyroid cancer are extremely rare, the possibility of the development of thyroid cancer must be taken into account in patients affected by thyroid hemiagenesis and the nodular variant of Graves' disease.

Broecker-Preuss M, Baten J, Sheu-Grabellus SY, et al.
Expression of the cAMP binding protein EPAC1 in thyroid tumors and growth regulation of thyroid cells and thyroid carcinoma cells by EPAC proteins.
Horm Metab Res. 2015; 47(3):200-8 [PubMed] Related Publications
The thyrotropin receptor-cAMP pathway is central in growth regulation of thyroid cells and thyroid tumorigenesis, and it regulates expression of thyroid specific genes. Recently, 2 new protein kinase A-independent cAMP effectors named EPAC1 and 2 were described that activate additional intracellular pathways. The aim of our study was to investigate the role of EPAC proteins in growth regulation of thyroid cells and thyroid carcinomas. EPAC1 expression was investigated immunohistochemically in tissues of various thyroid tumors. Utilizing MTT assay, the effect of EPAC stimulation on proliferation in thyroid carcinoma cells and in non-transformed rat FRTL5 cells was investigated. The activation of intracellular signaling pathways was examined by RAP pull-down assay and Western blots. EPAC1 expression was strong in non-oxyphilic follicular thyroid adenomas and carcinomas and in follicular papillary thyroid carcinomas. It was moderate in oxyphilic follicular tumors and classical and tall cell papillary carcinomas. In contrast, EPAC1 expression was low in poorly differentiated carcinomas and very low in anaplastic carcinomas. Thyroid carcinoma cell lines showed no or very weak EPAC1 expression and exhibited no growth-promoting effect after EPAC stimulation. Non-transformed rat FRTL5 cells were growth-stimulated by an EPAC-specific cAMP-analogue and showed EPAC-dependent activation of RAP, ERK, and p70S6 kinase. EPAC1 expression and cellular response to EPAC activation in rat FRTL5 cells reflect cellular responses to cAMP and TSH stimulation in non-transformed thyroid cells. In undifferentiated thyroid carcinomas, loss of EPAC1 expression may be in accordance with the loss of thyroid-specific functions and the loss of responsiveness of the TSHR-cAMP pathway.

He H, Li W, Liyanarachchi S, et al.
Genetic predisposition to papillary thyroid carcinoma: involvement of FOXE1, TSHR, and a novel lincRNA gene, PTCSC2.
J Clin Endocrinol Metab. 2015; 100(1):E164-72 [PubMed] Free Access to Full Article Related Publications
CONTEXT: By genome-wide association studies, the risk allele [A] of SNP rs965513 predisposes strongly to papillary thyroid carcinoma (PTC). It is located in a gene-poor region of 9q22, some 60 kb from the FOXE1 gene. The underlying mechanisms remain to be discovered.
OBJECTIVE: Our objective was to identify novel transcripts in the 9q22 locus and correlate gene expression levels with the genotypes of rs965513.
DESIGN: We performed 3' and 5' rapid amplification of cDNA ends and RT-PCR to detect novel transcripts. One novel transcript was forcibly expressed in a cell line followed by gene expression array analysis. We genotyped rs965513 from PTC patients and measured gene expression levels by real-time RT-PCR in unaffected thyroid tissue and matched tumor.
SETTING: This was a laboratory-based study using cells from clinical tissue samples and a cancer cell line.
MAIN OUTCOME MEASURES: We detected previously uncharacterized transcripts and evaluated the gene expression levels and the correlation with the risk allele of rs965513, age, gender, chronic lymphocyte thyroiditis (CLT), and TSH levels.
RESULTS: We found a novel long intergenic noncoding RNA gene and named it papillary thyroid cancer susceptibility candidate 2 (PTCSC2). Transcripts of PTCSC2 are down-regulated in PTC tumors. The risk allele [A] of rs965513 was significantly associated with low expression of unspliced PTCSC2, FOXE1, and TSHR in unaffected thyroid tissue. We also observed a significant association of age and CLT with PTCSC2 unspliced transcript levels. The correlation between the rs965513 genotype and the PTCSC2 unspliced transcript levels remained significant after adjusting for age, gender, and CLT. Forced expression of PTCSC2 in the BCPAP cell line affected the expression of a subset of noncoding and coding transcripts with enrichment of genes functionally involved in cell cycle and cancer.
CONCLUSIONS: Our data suggest a role for PTCSC2, FOXE1, and TSHR in the predisposition to PTC.

Nikiforov YE, Carty SE, Chiosea SI, et al.
Highly accurate diagnosis of cancer in thyroid nodules with follicular neoplasm/suspicious for a follicular neoplasm cytology by ThyroSeq v2 next-generation sequencing assay.
Cancer. 2014; 120(23):3627-34 [PubMed] Related Publications
BACKGROUND: Fine-needle aspiration (FNA) cytology is a common approach to evaluating thyroid nodules, although 20% to 30% of FNAs have indeterminate cytology, which hampers the appropriate management of these patients. Follicular (or oncocytic) neoplasm/suspicious for a follicular (or oncocytic) neoplasm (FN/SFN) is a common indeterminate diagnosis with a cancer risk of approximately 15% to 30%. In this study, the authors tested whether the most complete next-generation sequencing (NGS) panel of genetic markers could significantly improve cancer diagnosis in these nodules.
METHODS: The evaluation of 143 consecutive FNA samples with a cytologic diagnosis of FN/SFN from patients with known surgical outcomes included 91 retrospective samples and 52 prospective samples. Analyses were performed on a proprietary sequencer using the targeted ThyroSeq v2 NGS panel, which simultaneously tests for point mutations in 13 genes and for 42 types of gene fusions that occur in thyroid cancer. The expression of 8 genes was used to assess the cellular composition of FNA samples.
RESULTS: In the entire cohort, histologic analysis revealed 104 benign nodules and 39 malignant nodules. The most common point mutations involved the neuroblastoma RAS viral oncogene homolog (NRAS), followed by the Kirsten rat sarcoma viral oncogene homolog (KRAS), the telomerase reverse transcriptase (TERT) gene, and the thyroid-stimulating hormone receptor (TSHR) gene. The identified fusions involved the thyroid adenoma associated (THADA) gene; the peroxisome proliferator-activated receptor γ (PPARG) gene; and the neurotrophic tyrosine kinase, receptor, type 3 (NTRK3) gene. Performance characteristics were similar in the retrospective and prospective groups. Among all FN/SFN nodules, preoperative ThyroSeq v2 performed with 90% sensitivity (95% confidence interval [CI], 80%-99%), 93% specificity (95% CI, 88%-98%), a positive predictive value of 83% (95% CI, 72%-95%), a negative predictive value of 96% (95% CI, 92%-100%), and 92% accuracy (95% CI, 88%-97%).
CONCLUSIONS: The current results indicate that comprehensive genotyping of thyroid nodules using a broad NGS panel provides a highly accurate diagnosis for nodules with FN/SFN cytology and should facilitate the optimal management of these patients.

Khan MS, Pandith AA, Masoodi SR, et al.
Epigenetic silencing of TSHR gene in thyroid cancer patients in relation to their BRAF V600E mutation status.
Endocrine. 2014; 47(2):449-55 [PubMed] Related Publications
Promoter hypermethylation of multiple genes have been identified to play a role in thyroid cancers and most prominent among them is TSHR gene promoter hypermethylation in particular showing a close association with BRAF gene-altered status. Thus, the aim of this study was to analyze the TSHR gene promoter hypermethylation in a series of thyroid tumor tissues in the backdrop of their BRAF gene mutational status. Methylation-specific PCR (MS-PCR) was used for detection of promoter methylation while BRAF gene mutational status was analyzed by PCR followed by DNA sequencing in the same series of 60 thyroid tumor tissues. The promoter region of TSHR gene was found to be methylated in 25 % (15 of 60) of the thyroid cancer patients. Patients having elevated TSH levels showed strong association with methylation (OR = 4.0, P = 0.02). BRAF V600E mutation was found in 25 % (15 of 60) patients and among them TSHR promoter was methylated in 73.3 % (11 of 15) patients and only 26.7 % (4 of 15) patients with mutated BRAF showed the absence of TSHR promoter methylation. We found a significant association between the presence of methylation in TSHR with the BRAF V600E mutation-positive cases (P < 0.05). In conclusion, our study showed a high implication of TSHR gene methylation and its significant association with BRAF V600E mutation in thyroid tumors, depicting a positive connection between TSHR pathway and MAP Kinase pathway.

Vicchio TM, Giovinazzo S, Certo R, et al.
Lack of association between autonomously functioning thyroid nodules and germline polymorphisms of the thyrotropin receptor and Gαs genes in a mild to moderate iodine-deficient Caucasian population.
J Endocrinol Invest. 2014; 37(7):625-30 [PubMed] Related Publications
BACKGROUND: Mutations of the thyrotropin receptor (TSHR) and/or Gαs gene have been found in a number of, but not all, autonomously functioning thyroid nodules (AFTNs). Recently, in a 15-year-old girl with a hyperfunctioning papillary thyroid carcinoma, we found two somatic and germline single nucleotide polymorphisms (SNPs): a SNP of the TSHR gene (exon 7, codon 187) and a SNP of Gαs gene (exon 8, codon 185). The same silent SNP of the TSHR gene had been reported in patients with AFTN or familial non-autoimmune hyperthyroidism. No further data about the prevalence of the two SNPs in AFTNs as well as in the general population are available in the literature.
AIM: To clarify the possible role of these SNPs in predisposing to AFTN.
METHODS: Germline DNA was extracted from blood leukocytes of 115 patients with AFTNs (43 males and 72 females, aged 31-85 years, mean ± SD = 64 ± 13) and 100 sex-matched healthy individuals from the same geographic area, which is marginally iodine deficient. The genotype distribution of the two SNPs was investigated by restriction fragment length polymorphism-polymerase chain reaction.
RESULTS: The prevalence of the two SNPs in our study population was low and not different to that found in healthy individuals: 8 % of patients vs. 9 % of controls were heterozygous for the TSHR SNP and 4 % patients vs. 6 % controls were heterozygous for the Gαs SNP. One patient harbored both SNPs.
CONCLUSIONS: These results suggest that these two SNPs do not confer susceptibility for the development of AFTN.

Boutin A, Eliseeva E, Gershengorn MC, Neumann S
β-Arrestin-1 mediates thyrotropin-enhanced osteoblast differentiation.
FASEB J. 2014; 28(8):3446-55 [PubMed] Free Access to Full Article Related Publications
Thyrotropin (TSH) activation of the TSH receptor (TSHR), a 7-transmembrane-spanning receptor (7TMR), may have osteoprotective properties by direct effects on bone. TSHR activation by TSH phosphorylates protein kinases AKT1, p38α, and ERK1/2 in some cells. We found TSH-induced phosphorylation of these kinases in 2 cell lines engineered to express TSHRs, human embryonic kidney HEK-TSHR cells and human osteoblastic U2OS-TSHR cells. In U2OS-TSHR cells, TSH up-regulated pAKT1 (7.1±0.5-fold), p38α (2.9±0.4-fold), and pERK1/2 (3.1±0.2-fold), whereas small molecule TSHR agonist C2 had no or little effect on pAKT1 (1.8±0.08-fold), p38α (1.2±0.09-fold), and pERK1/2 (1.6±0.19-fold). Furthermore, TSH increased expression of osteoblast marker genes ALPL (8.2±4.6-fold), RANKL (21±5.9-fold), and osteopontin (OPN; 17±5.3-fold), whereas C2 had little effect (ALPL, 1.7±0.5-fold; RANKL, 1.3±0.6-fold; and OPN, 2.2±0.7-fold). β-Arrestin-1 and -2 can mediate activatory signals by 7TMRs. TSH stimulated translocation of β-arrestin-1 and -2 to TSHR, whereas C2 failed to translocate either β-arrestin. Down-regulation of β-arrestin-1 by siRNA inhibited TSH-stimulated phosphorylation of ERK1/2, p38α, and AKT1, whereas down-regulation of β-arrestin-2 increased phosphorylation of AKT1 in both cell types and of ERK1/2 in HEK-TSHR cells. Knockdown of β-arrestin-1 inhibited TSH-stimulated up-regulation of mRNAs for OPN by 87 ± 1.7% and RANKL by 73 ± 2.4%, and OPN secretion by 74 ± 10%. We conclude that TSH enhances osteoblast differentiation in U2OS cells that is, in part, caused by activatory signals mediated by β-arrestin-1.

D'Agostino M, Sponziello M, Puppin C, et al.
Different expression of TSH receptor and NIS genes in thyroid cancer: role of epigenetics.
J Mol Endocrinol. 2014; 52(2):121-31 [PubMed] Related Publications
The TSH receptor (TSHR) and sodium/iodide symporter (NIS) are key players in radioiodine-based treatment of differentiated thyroid cancers. While NIS (SLC5AS) expression is diminished/lost in most thyroid tumors, TSHR is usually preserved. To examine the mechanisms that regulate the expression of NIS and TSHR genes in thyroid tumor cells, we analyzed their expression after inhibition of ras-BRAF-MAPK and PI3K-Akt-mTOR pathways and the epigenetic control occurring at the gene promoter level in four human thyroid cancer cell lines. Quantitative real-time PCR was used to measure NIS and TSHR mRNA in thyroid cancer cell lines (TPC-1, BCPAP, WRO, and FTC-133). Western blotting was used to assess the levels of total and phosphorylated ERK and Akt. Chromatin immunoprecipitation was performed for investigating histone post-translational modifications of the TSHR and NIS genes. ERK and Akt inhibitors elicited different responses of the cells in terms of TSHR and NIS mRNA levels. Akt inhibition increased NIS transcript levels and reduced those of TSHR in FTC-133 cells but had no significant effects in BCPAP. ERK inhibition increased the expression of both genes in BCPAP cells but had no effects in FTC-133. Histone post-translational modifications observed in the basal state of the four cell lines as well as in BCPAP treated with ERK inhibitor and FTC-133 treated with Akt inhibitor show cell- and gene-specific differences. In conclusion, our data indicate that in thyroid cancer cells the expression of TSHR and NIS genes is differently controlled by multiple mechanisms, including epigenetic events elicited by major signaling pathways involved in thyroid tumorigenesis.

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