Endocrine Abstracts

Purpose: Treatment with tyrosine kinase inhibitors (TKI) has been associated with alterations in circulating thyroid hormone levels, possibly related to perturbations in peripheral thyroid hormone metabolism. In this study, we evaluated the effect of the multi-kinase inhibitor vandetanib on the expression of the three deiodinase selenoenzymes.

Materials and Methods: Cell models that endogenously express either D1 (HepG2 and HEK-293), D2 (HeLa and KMH-2) or D3 (K1 and Cal-62), respectively, were treated with vandetanib 2 uM in time-course experiments. Deiodinase expression levels were assessed by RT-PCR. PDGFRA-EGFP mice were administered with 50 mg/kg vandetanib daily and sacrificed after 14 or 28 days. Fibro/ adipogenic progenitors (FAPs) were isolated by FACS sorting after muscle digestion. D2 deiodination assay was performed using liquid chromatography-tandem mass spectrometry (LC-MS/MS).

Results: In vitro, vandetanib determined a strong cell-specific downregulation of Dio2 expression in KMH-2 cells, and no changes in deiodinase expression were found in other cell lines. The effect of vandetanib was then analyzed in mice in two tissues wherein D2 action is highly relevant, i.e. brown adipose tissue (BAT) and pituitary. 28 days after daily treatment with vandetanib, Dio2 mRNA levels in both BAT and pituitary were unchanged compared to controls. Therefore, we hypothesized that specific cell types dispersed in selected organs would play an important role in the vandetanib-mediated effects on TH metabolism. We found that FAPs (i.e. interstitial mesenchymal progenitors crucial for muscle regeneration process) from heart and skeletal muscle in mice express elevated levels of Dio2 and other genes relevant for TH metabolism. Strikingly, Dio2 expression in FAPs from both heart and skeletal muscle dramatically decreased after treatment for 14 days with vandetanib, and Dio2 inhibition persisted after 28 days of treatment. Importantly, liver expression of Dio1 and Dio3 in vandetanib-treated mice was unchanged compared to untreated controls, as well as markers of TH action such as Malic Enzyme and Spot14. We also measured D2 activity in skeletal muscle FAPs from vandetanib-treated mice compared to controls and found that the D2 activity in mice treated with vandetanib for 14 days was reduced of about 60% compared to controls.

Conclusion: Vandetanib determined a dramatic downregulation of D2 expression and activity in mesenchymal precursors from skeletal and heart muscle. Given the widespread diffusion of mesenchymal cells within the body, our results may explain at least partially the alterations in thyroid hormone levels that occur in TKI-treated patients.