The Growth Stimulatory Effects of Thyrotropin and Thyroid Hormones on Thyroid Cancer Depend on Expression of Thyrotropin Receptor and Integrins
Authors: Thakur Shilpa , Cardenas Stephanie , Klubo-Gwiezdzinska Joanna 
Published: May 2021

Abstract
Background: The long-term management of metastatic thyroid cancer (TC) consists of thyrotropin (TSH) suppression with supraphysiologic doses of thyroid hormones (TH) via a negative feedback loop. The goal of TSH suppression is to prevent TSH stimulation of the TSH receptor (TSHR), as it has been shown to promote proliferation of cancer cells. However, TH (T3 and T4) have also been shown to stimulate cancer cell proliferation via αvβ3 integrin signaling. Since both TSH and TH have mitogenic potential, we aimed to investigate which one is a more potent growth stimulus-TSH or TH by analyzing its growth stimulatory effects in TC models. Methods: We analyzed the mRNA expression of TSHR and ITGAV (αv), ITGB3 (β3) integrins in 496 human TC tissue samples, including 65 paired samples of normal tissue (NT) and the corresponding tumor included in The Cancer Genome Atlas (TCGA). We used 13 TC cell lines and analyzed the mRNA expression of 24 genes (4 thyroid-specific genes, 2 TH receptor genes, and 18 integrin genes) with an emphasis on the expression of cell surface receptors αv, β3 integrins, and TSHR. The protein expression of αv, β3, and TSHR was analyzed by immunoblotting. To test the effects of TH and TSH on cell proliferation and expression of αv, β3, and TSHR, cells were treated with varying concentrations of TSH (0.01, 0.1, 1, 10 mIU/mL), T3 (0.1, 1, 10 100 nM) and T4 (1, 10, 100, 1000 nM) for 72 h. Results: Analysis of the RNA seq data from TCGA revealed a significantly higher expression of TSHR in NT compared with TC (log fold change 0.59, p&lt;0.001), lower expression of αv integrin in NT compared with TC (log fold change -0.3, p=0.001), and comparable expression of β3 integrin (log fold change 0.20, p=0.3). Based on the mRNA expression data of 13 TC cell lines, we selected 6 cell lines (FTC133, TPC1, XTC1, OCUT2, C643, THJ16T) characterized by variable αv, β3, and TSHR expression. The TPC1 and OCUT2 cells with high to moderate αVβ3 expression responded to T4 (1000nM; p&lt;0.001) and T3 treatment (100nM; p&lt;0.001) respectively, with increased proliferation, while the cell lines characterized by low to no β3 and/or low αV expression (FTC133, XTC1, C643, and THJ16T) did not change their growth rate in response to TH. The C643 and XTC1 cells characterized by a preserved low-to-moderate TSHR expression responded to TSH treatment (10mIU/mL) with increased proliferation (p&lt;0.05), while the growth curve of cell lines with very low to no TSHR expression (FTC133, TPC1, OCUT1, THJ16T) was not affected. Analysis of the effects of TH and TSH on the mRNA expression of αV, β3, and TSHR was observed to be cell-line specific. Conclusion: The growth stimulatory effects of TSH and TH on TC cells depend on its concentration and expression of TSHR and αVβ3, respectively. Since TC is characterized by relatively lower TSHR and higher αV integrin expression than NT, treatment with supraphysiologic doses of TH in patients with metastatic TC needs to be individualized.