Probing for the effect of thyroid hormone receptors in different cancer phenotypes.

摘要

Thyroid hormone receptors (TRs) are ligand-dependent transcription factors that mediate many of the actions of the thyroid hormone in growth, development, and differentiation. Over the past decades significant advances have been made in the understanding of TR actions in maintaining normal cellular functions. However, the role/s of TRs in human cancer remain unclear to this day. TR alterations in the form of somatic mutations have been associated with several types of cancer, implying that TRs function as tumor suppressors. These TR mutants are usually impaired in T3-driven gene transcription and in many cases can function as dominant-negative inhibitors of wild-type TR activity. The Koh lab has generated tools to complement mutations with impaired transactivation. The hormone analog QH2 behaves as a "functionally orthogonal" ligand that rescues the transcriptional activity of the synthetic mutant TRbeta(H435A) that otherwise is not responsive to T3. QH2 can activate the TRbeta(H435A) mutant without stimulating wild-type TRs, including TRalpha;, which has been associated with cardiotoxic side effects. TRbeta; mutations at His435 have been found in genetic diseases such as resistance to thyroid hormone (RTH) and pituitary cancer (TSHoma), making QH2 and ideal candidate to potentially restore some of the cancer-associated phenotypes in in vitro assays. This orthogonal ligand-receptor pair may provide means to assess the role of mutant TRs in cancer progression. A second novel hormone analog developed in the Koh laboratory, JZ07, is a potent antagonist of wild-type TRs. If wild-type TRs function as tumor inducing factors due to their dual role in cancer development and progression, we envisioned JZ07 could attenuate this effect and inhibit metastasis-associated phenotypes. Many in vitro studies use supraphysiological concentrations of T3 to induce changes in cancer- associated phenotypes and to infer the role of TR. JZ07 will allow us to assess the role of a TR antagonist on TR function under normal or "euthyroid" conditions. This dissertation evaluates the use of these TR modulators as tools to assess the contribution of TRs in cancer-associated phenotypes. Expression of TRs proved to inhibit the invasive and migratory activities of cells, suggesting TRs could function as tumor suppressors. Evaluation of the hormone analogs QH2 and JZ07 induced additional effects in the different metastatic phenotypes. QH2 did not affect the increased invasive and proliferative activities of the mutant TRbeta(H435A), however TRbeta(H435A) did not repress invasion as would be expected if TRbeta(H435A) behaved similar to unliganded TRbeta(wt). The orthogonal ligand enhanced migration in the mutant TRbeta(H435A) but clear non-genomic effects are also observed in wild-type expressing cell lines. The novel hormone analog JZ07 successfully recapitulated its antagonistic behavior in the invasive activity of wild-type TRs in an isotype-dependent manner and further inhibited invasion in TRbeta(wt) cells. Nevertheless, the inhibitory effect in the migration activity of the mutant TRbeta(H435A) and Neo control cells cannot be explained by classical transactivation action alone. Together, this suggests that the role of TRs in cancer-associated phenotypes of migration, invasion and proliferation likely involve non-genomic actions of TRs and the affects of T3 on these cell phenotypes likely involves T3 acting on cellular targets other than TRs.