**4. Radioiodine-refractory thyroid cancer**

Approximately 80 years after the first clinical use of radioiodine therapy for the diagnosis and treatment of differentiated thyroid cancer [41], radioiodine therapy is still the first choice of treatment after thyroidectomy for primary and metastatic differentiated thyroid carcinomas. However, 30% of metastatic differentiated thyroid tumors show dedifferentiation and lose their ability to accumulate radioiodine, thus making adjuvant treatment with radioiodine ineffective (radioiodine-refractory) [42]. Current therapeutic strategies for symptomatic radioiodine-refractory thyroid cancers include the implementation of local therapy whenever possible. However, in the case of diffuse significant progression of distant metastatic disease, systemic therapy is currently based on anti-angiogenic multi-targeted tyrosine kinase inhibitors [43]. The two multi-targeted

#### *The Molecular Basis for Radioiodine Therapy DOI: http://dx.doi.org/10.5772/intechopen.108073*

tyrosine kinase inhibitors sorafenib and lenvatinib have been approved by regulatory authorities for use in radioiodine-refractory differentiated thyroid carcinomas [44, 45]. These agents have shown promising results with a significant improvement of median progression-free survival over placebo, but generally with similar overall survival. More recently, novel highly selective inhibitors targeting oncogenic chromosomal rearrangements involving the proto-oncogenes RET and NTRK have been approved for clinical use in radioiodine-refractory differentiated thyroid carcinomas [46, 47]. Therefore, the presence of druggable oncogenes should be screened in patients with metastatic disease, and whenever present, a selective inhibitor might be considered.

The underlying molecular basis for the loss of radioiodine accumulation in radioiodine-refractory metastatic thyroid carcinomas is thyroid dedifferentiation, which results in a decreased expression of the genes involved in the iodide metabolism. Radioiodine therapy effectivity is ultimately dependent on functional NIS expression at the plasma membrane of the thyroid tumor cells, as deficient radioiodide accumulation is the major cause of treatment failure [5]. However, NIS gene expression is frequently downregulated in differentiated thyroid cancer compared with normal thyroid tissue or even totally silenced as evidenced in poorly differentiated carcinomas. Multiple transcriptional and posttranscriptional mechanisms have been postulated to explain NIS gene repression in thyroid tumors, including transcriptional repression of the transcription factor Pax8 that regulates NIS gene transcriptional expression, and by TGFB1-induced activation of SMAD signaling leading to NOX4-dependent ROS production, which in turn impairs Pax8-dependent NIS gene expression [48, 49]. Immunohistochemical studies have revealed that NIS is frequently expressed (or even overexpressed) at different levels in differentiated thyroid carcinomas compared with adjacent normal tissue [50]. However, NIS expression is mainly located in intracellular compartments, indicating that a posttranslational mechanism is involved in radioiodide resistance due to defective NIS expression at the plasma membrane [51, 52]. Significantly, loss-of-function NIS variants have not been identified in either benign cold thyroid nodules or thyroid tumors [53, 54], demonstrating that the intracellular retention of NIS is not caused by structural defects, as reported in patients with dyshormonogenic congenital hypothyroidism. Therefore, the paradoxical observation of reduced radioiodine uptake and intracellularly retained NIS protein expression highlights the importance of elucidating the posttranslational mechanisms regulating NIS plasma membrane expression.

The pituitary tumor-transforming gene (PTTG) binding factor (PBF) has been reported as being an NIS-interacting protein involved in NIS intracellular retention in thyroid cancer, as ectopic PBF overexpression results in reduced iodide accumulation caused by NIS endocytosis from the plasma membrane [55]. The phosphorylation of the PBF residue Tyr-174 is required for PBF-mediated NIS endocytosis, as PP1 inhibited Src kinase activity restores iodide accumulation in thyroid cancer cell lines [56]. Moreover, chemical inhibition of the NIS-interacting protein valosin-containing protein (VCP), a principal component of the endoplasmic reticulum-associated degradation protein quality control process involved in NIS proteolysis, increases NIS expression at the plasma membrane and radioiodide accumulation in thyroid cancer models [57]. Recently, high-throughput drug screening has revealed multiple cellular processes that are central to NIS regulation, including proteasomal degradation and autophagy, which can be drugged to enhance radioiodide uptake [58]. Moreover, functional defects in the glycosylphosphatidylinositol (GPI) transamidase complex due to BRAFV600E-triggered PIGU repression cause partially glycosylated NIS molecules to be retained in the endoplasmic reticulum, probably due to a deficiency in

an unidentified GPI-anchored protein that is necessary for proper NIS anterograde plasma membrane transport [59].

Constitutive activation of mitogen-activated protein kinase (MAPK) signaling induces a partial to complete loss of differentiation in thyroid cancers. In agreement, *in vitro* studies have revealed that BRAFV600E impairs NIS expression, thereby reducing iodide uptake [60]. In patients, BRAFV600E expression in papillary thyroid carcinoma was correlated with radioiodine-refractory recurrences and defective NIS expression or intracellular retention [61]. In line with this, transgenic mice expressing the oncogene BRAFV600E in thyroid follicular cells developed papillary thyroid tumors, with these tumors neither concentrating radioiodine nor responding to radioiodine therapy [62]. Interestingly, the blockage of BRAFV600E kinase activity with either the BRAFV600E-selective ATP competitive inhibitor PLX4720, a vemurafenib progenitor, or further downstream with the allosteric MEK 1/2 kinase inhibitor selumetinib, restored thyroid-specific gene expression (including NIS) and radioiodine incorporation into these tumors, which rendered them susceptible to therapeutic doses of radioiodine [62].

When used as single agents, vemurafenib and selumetinib are comparatively ineffective inhibitors of BRAFV600E-driven thyroid cancer. Although they are potent inhibitors of MAPK-ERK signaling in thyroid cancer, this inhibition is followed by a strong rebound effect that reactivates ERK signaling. Blockage of ERK-dependent negative feedback mechanisms increases the expression of the tyrosine kinase HER3, with its activation after dimerization with HER2 by the autocrine-secreted ligand neuregulin leading to ERK activation involving CRAF signaling [63]. Significantly, the allosteric MEK 1/2 inhibitor CH5126766, when bound to the protein, impairs its phosphorylation by upstream (A, B, or C) RAF kinases and reduces reactivation of ERK signaling, which overcomes the adaptive resistance of BRAFV600E-promoted thyroid cancer to MAPK inhibitors and markedly enhances the effectiveness of radioiodine therapy [64].
