Targeting Novel Sodium Iodide Symporter Interactors ADP-Ribosylation Factor 4 and Valosin-Containing Protein Enhances Radioiodine Uptake

Abstract

The sodium iodide symporter (NIS) is required for iodide uptake, which facilitates thyroid hormone biosynthesis. NIS has been exploited for over 75 years in ablative radioiodine (RAI) treatment of thyroid cancer, where its ability to transport radioisotopes depends on its localization to the plasma membrane. The advent of NIS-based in vivo imaging and theranostic strategies in other malignancies and disease modalities has recently increased the clinical importance of NIS. However, NIS trafficking remains illdefined. Here, we used tandem mass spectrometry followed by coimmunoprecipitation and proximity ligation assays to identify and validate two key nodes-ADP-ribosylation factor 4 (ARF4) and valosin-containing protein (VCP)-controlling NIS trafficking. Using cell-surface biotinylation assays and highly inclined and laminated optical sheet microscopy, we demonstrated that ARF4 enhanced NIS vesicular trafficking from the Golgi to the plasma membrane, whereas VCP-a principal component of endoplasmic reticulum (ER)-associated degradation-governed NIS proteolysis. Gene expression analysis indicated VCP expression was particularly induced in aggressive thyroid cancers and in patients who had poorer outcomes following RAI treatment. Two repurposed FDA-approved VCP inhibitors abrogated VCP-mediated repression of NIS function, resulting in significantly increased NIS at the cell-surface and markedly increased RAI uptake in mouse and human thyroid models. Collectively, these discoveries delineate NIS trafficking and highlight the new possibility of systemically enhancing RAI therapy in patients using FDAapproved drugs.Significance: These findings show that ARF4 and VCP are involved in NIS trafficking to the plasma membrane and highlight the possible therapeutic role of VCP inhibitors in enhancing radioiodine effectiveness in radioiodine-refractory thyroid cancer.

New mechanisms of radioiodide uptake revealed via a novel high throughput drug screening approach in thyroid cancer

“…S8). We previously identified ebastine as enhancing the localisation of NIS to the PM (35). Larger fold changes in radioiodide uptake were apparent in cells treated with ebastine (EBT) and SAHA than with SAHA alone ( Fig.…”

“…Human thyroid carcinoma cell lines TPC-1 and 8505C, and human breast lines MDA-MB-231 and MCF7, were cultured as before (35), at low passage, authenticated by short tandem repeat analysis (NorthGene) and tested for Mycoplasma contamination (EZ-PCR; Geneflow).…”

“…Full-length human NIS cDNA (36) Table S1). RAI uptake, Western blotting and NIS mRNA expression assays were used to monitor NIS activity as previously (35,36). The expression of specific mRNAs was determined with a 7500 Real-time PCR system (Applied Biosystems).…”

“…The collection of normal human thyroid tissue was approved by the Local Research Ethics Committee, and subjects gave informed written consent. Human primary thyrocytes were isolated as previously (35,37). Only cultures with positive TSH dose responses were progressed to cell experiments after 6 days.…”

“…As our most prominent drugs in terms of fold induction of NIS function were ebastine, carebastine and the histone deacetylase SAHA ( Supplementary Table S4), we investigated their potential for mechanistic interaction. Thyroid cancers frequently show repression of NIS mRNA expression (11), and VCP is critical to the post-translational processing of NIS (35).…”