RNA-seq analysis of transcriptional co-regulators of NIS in a murine model of triple negative breast cancer

Introduction: The sodium iodide symporter (NIS; <i>SLC5A5</i>) is the sole transporter of iodide, exploited in radioiodide treatment of differentiated thyroid cancer. NIS is expressed in breast cancer (BrCa), but its intracellular retention prevents sufficient radioiodide uptake for tumoural ablation. Pre-clinical studies suggest that upregulation of <i>SLC5A5</i> is promoted by retinoic acid receptor alpha (RARα), and that this mechanism is epigenetically corrupted in BrCa via upregulation of the RARα co-repressor PRAME. We recently reported that the histone deacetylase (HDAC) inhibitor SAHA - combined with the disulfiram metabolite Cu(DDC)₂ - significantly increased NIS function in MDA-MB-231 orthotopic tumours. Here, we employed RNA-seq analysis of these tumours to determine whether transcriptional co-regulators impact NIS expression in BrCa.<br/><br/>Methods: Global changes in gene expression in response to SAHA and Cu(DDC)₂ were appraised by RNA-seq analysis. Differentially expressed genes (DEGs) were identified and analysed using DSeq2 and DAVID.<br/><br/>Results: PRAME-overexpressing MDA-MB-231 cells demonstrated significantly lower anti-proliferative responses to ATRA (pan RAR ligand) and AM580 (RARa ligand). PRAME overexpression led to trans-repression of RARα canonical target genes and <i>SLC5A5</i> in response to ATRA. We therefore established a panel of BrCa cell lines (MDA-MB-231, SUM52, AU565, SKBR3) stably transfected with either CRISPR-VP64 (activator) or CRISPR-KRAB (repressor), measuring the expression of <i>SLC5A5</i> and the ability to transport radioiodide. Gene ontology enrichment analysis and pathway mapping of <i>in vivo</i> BrCa tumours treated with SAHA and Cu(DDC)₂ highlighted transcriptional regulation as the most significantly altered biological process (474/1230; q=9.05x10^-15) and chromatin remodelling linked to HDAC4 (q=2.5x10^-7). Interestingly, 1081 genes regulated by RARα were significantly dysregulated due to drug treatment (q=3.21x10^-84).<br/><br/>Discussion: These studies reveal that our drug strategies to enhance NIS function in orthotopic breast tumours are accompanied by unexpectedly potent transcriptional activation, yielding further insight into the mechanisms that control SLC5A5 expression in breast cancer.