1). Growth Activation Alone is not Sufficient to Cause Metastatic Thyroid Cancer in a Mouse Model of Follicular Thyroid Carcinoma. Thyroid stimulating hormone (TSH) is the major stimulator of thyrocyte proliferation, but its role in thyroid carcinogenesis remains unclear. To address this question, we used TRbetaPV/PV mice. These mice, harboring a dominantly negative mutation (PV) of the thyroid hormone beta receptor (TRbeta), exhibit increased serum thyroid hormone and elevated TSH. To eliminate TSH-growth stimulating effect, TRbetaPV/PV mice were crossed with TSH receptor gene knockout (TSHR-/-) mice. Wild-type siblings of TRbetaPV/PV mice were treated with anti-thyroid agent, propylthiouracil (PTU) to elevate serum TSH for evaluating long-term TSH effect (WT-PTU mice). Thyroids from TRbetaPV/PVTSHR-/- showed impaired growth with no occurrence of FTC. Both WT-PTU and TRbetaPV/PV mice displayed enlarged thyroids, but only TRbetaPV/PV mice developed metastatic FTC. Molecular analyses indicate that PV acted, via multiple mechanisms, to activate the integrins-Src-focal adhesion kinase-p38 MAPK pathway and to affect cytoskeletal re-structuring to increase tumor cell migration, and invasion. Thus, growth stimulated by TSH is a pre-requisite, but not sufficient for metastatic cancer to occur. Additional genetic alterations (such as PV), destined to alter focal adhesion and migration capacities, are required to empower hyperplastic follicular cells to invade and to metastasize. These in vivo findings provide new insights in understanding carcinogenesis of the human thyroid. 2). Inhibition of mTORC1 Signaling Reduces Tumor Growth but does not Prevent Cancer Progression in a Mouse Model of Thyroid Cancer. Selective drugs targeting dysregulated oncogenic pathways are promising cancer therapies. Because the mTORC1 pathway is hyperactivated in human follicular thyroid cancer (FTC), we hypothesized that its inhibition could block cancer development and progression. We, therefore, analyzed the effect of a treatment with a specific mTORC1 inhibitor (RAD001) in a faithful mouse model of FTC with constitutive mTORC1 activation (TRbeta PV/PVPten +/- mice). The treatment did not prevent capsular and vascular invasion of the thyroid and the occurrence of lung metastasis. However, it substantially decelerated thyroid tumor growth, thereby prolonging TRbeta PV/PVPten +/- mouse lifespan. RAD001 efficiently inhibited mTORC1 activity, as shown by the reduced phosphorylation of its downstream targets involved in the activity of the translation machinery, such as p70S6K, eukaryotic translation initiation factor 4E binding protein 4E-BP1, and the eukaryotic initiation factors eIF-4B and eIF-4G. Whereas mTORC1signaling inhibition did not alter cell apoptosis, it induced a significant decrease in cell proliferation that was associated with the reduced abundance and altered activity of key regulators of cell cycle progression. Altogether, our data indicate that mTORC1 signaling plays a major role in the integration of the mitogenic signal in FTC. Therefore, our preclinical study with a relevant mouse model of FTC demonstrates for the first time that RAD001 efficaciously stabilizes cancer growth although it does not prevent its fatal outcome. In conclusion, our work underscores that in the treatment of FTC patients, RAD001 can only be used in combination with drugs and therapies inducing tumor shrinkage and blocking metastasis. 3). Thyroid Hormone Receptors are Tumor Suppressors in a Mouse Model of Metastatic Follicular Thyroid Carcinoma. Aberrant expression and mutations of TRs are closely associated with several types of human cancers. To test the hypothesis that TRs could function as tumor suppressors, we took advantage of mice with deletion of all functional TRs (TRalpha1 -/-TRbeta -/- mice). As these mice aged, they spontaneously developed follicular thyroid carcinoma with pathological progression from hyperplasia to capsular invasion, vascular invasion, anaplasia, and metastasis to the lung, similar to human thyroid cancer. Detailed molecular analysis revealed that known tumor promoters such as pituitary tumor transforming gene were activated and tumor suppressors such as peroxisome proliferator-activated receptor gamma and p53 were suppressed during carcinogenesis. In addition, consistent with the human cancer, AKT-mTOR-p70S6K signaling and vascular growth factor and its receptor were activated to facilitate tumor progression. This report presents in vivo evidence that functional loss of both TRalpha1 and TRbeta genes promotes tumor development and metastasis. Thus, TRs were identified as novel tumor suppressors in a mouse model of metastatic follicular thyroid cancer. 4). Pituitary Homeobox 2 (PITX2) Promotes Thyroid Carcinogenesis by Activation of Cyclin D2. Pituitary homeobox 2 (PITX2), a Paired -like homeodomain transcription factor and a downstream effector of beta-catenin signaling, plays substantial roles in normal embryonic development but its possible involvement in tumorigenesis was unknown. In this study, we extend its function in human cancer. Remarkably, we found that PITX2 was frequently expressed in human follicular cell-derived (papillary, follicular, and anaplastic) thyroid cancer tissues but not in normal thyroids, indicating for the first time that overactivated PITX2 may contribute to thyroid cancer. Cell-based and biochemical studies were performed to uncover the molecular mechanism of PITX2 action in thyroid cancer. Knockdown of PITX2 gene expression in human thyroid cancer cells significantly reduced cell proliferation and soft-agar colony formation. Biochemical analysis of cell cycle regulators upon PITX2 knockdown revealed down-regulation of Cyclin D1, Cyclin D2, and dephosphorylation of Rb. Chromatin immunoprecipitation and promoter reporter assay indicated that Cyclin D2 was a direct target gene of PITX2. Consistently, we observed that high expression levels of Cyclin D2 were frequently associated with PITX2 expression in follicular cell-derived thyroid cancer tissues. To confirm our results in vivo, we took advantage of a mouse model of thyroid cancer (TRbeta PV/PV mouse). Consistently, the aberrant elevation of Pitx2 levels in the thyroid cancer of TRbetaPV/PV mice was accompanied by up-regulation of Cyclin D1, Cyclin D2, and increased phosphorylation of Rb. Collectively, our findings demonstrate that the overactivated PITX2-Cyclin D2 pathway promotes thyroid tumorigenesis, and they provide the first evidence implicating an oncogenic role of PITX2 in human cancer.

Agency
National Institute of Health (NIH)
Institute
National Cancer Institute (NCI)
Type
Investigator-Initiated Intramural Research Projects (ZIA)
Project #
1ZIABC011191-02
Application #
8157686
Study Section
Project Start
Project End
Budget Start
Budget End
Support Year
2
Fiscal Year
2010
Total Cost
$762,355
Indirect Cost
Name
National Cancer Institute Division of Basic Sciences
Department
Type
DUNS #
City
State
Country
Zip Code
Kim, Won Gu; Cheng, Sheue-Yann (2018) Mechanisms Linking Obesity and Thyroid Cancer Development and Progression in Mouse Models. Horm Cancer 9:108-116
Park, Sunmi; Willingham, Mark C; Qi, Jun et al. (2018) Metformin and JQ1 synergistically inhibit obesity-activated thyroid cancer. Endocr Relat Cancer 25:865-877
Zhu, Xuguang; Cheng, Sheue-Yann (2018) Analysis of Thyroid Tumorigenesis in Xenograft Mouse Model. Methods Mol Biol 1801:207-223
Zhu, Xuguang; Cheng, Sheue Yann (2017) Epigenetic Modifications: Novel Therapeutic Approach for Thyroid Cancer. Endocrinol Metab (Seoul) 32:326-331
Park, Jeong Won; Zhao, Li; Willingham, Mark C et al. (2017) Inhibition of STAT3 signaling blocks obesity-induced mammary hyperplasia in a mouse model. Am J Cancer Res 7:727-739
Enomoto, Keisuke; Zhu, Xuguang; Park, Sunmi et al. (2017) Targeting MYC as a Therapeutic Intervention for Anaplastic Thyroid Cancer. J Clin Endocrinol Metab 102:2268-2280
Zhu, Xuguang; Enomoto, Keisuke; Zhao, Li et al. (2017) Bromodomain and Extraterminal Protein Inhibitor JQ1 Suppresses Thyroid Tumor Growth in a Mouse Model. Clin Cancer Res 23:430-440
Zhu, Xuguang; Kim, Dong Wook; Zhao, Li et al. (2016) SAHA-induced loss of tumor suppressor Pten gene promotes thyroid carcinogenesis in a mouse model. Endocr Relat Cancer 23:521-33
Park, Jeong Won; Zhao, Li; Willingham, Mark C et al. (2016) Loss of tyrosine phosphorylation at Y406 abrogates the tumor suppressor functions of the thyroid hormone receptor ?. Mol Carcinog :
Park, Jeong Won; Han, Cho Rong; Zhao, Li et al. (2016) Inhibition of STAT3 activity delays obesity-induced thyroid carcinogenesis in a mouse model. Endocr Relat Cancer 23:53-63

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