Identification of GPX3 and JUN as Tumor Suppressors in Thyroid Cancer through Integrated WGCNA and Mendelian Randomization

<b>Background:</b> Thyroid cancer (TC) ranks among the most common malignancies globally, with an increasing incidence among younger populations. While papillary thyroid carcinoma (PTC) generally has a favorable prognosis, other forms of TC, such as anaplastic thyroid carcinoma (ATC), are associated with poor outcomes. Although specific mutations, such as BRAF^V600E, have been identified in certain types of TC, the underlying mechanisms remain largely unclear. Therefore, there is a critical need to further explore therapeutic targets associated with malignant tumors to improve treatment outcomes. <b>Method:</b> We integrated eQTL data from European populations with RNA-Seq data from TC patients obtained from TCGA and multiple GEO databases. Through differential expression analysis, WGCNA, and Mendelian randomization (MR) analysis, we sought to identify potential gene therapy targets in TC. Additionally, we explored the biological behaviors of these targets using various cellular biology assays, such as MTT, colony formation, wound healing, and Transwell assays. Molecular biology techniques, including Western blot, were employed to investigate the underlying mechanisms. <b>Result:</b> Differential expression analysis across six GEO datasets identified 649 genes associated with TC. Subsequent WGCNA analysis of the GSE6339 dataset revealed 2,739 genes, and MR analysis further identified 189 genes. The intersection of these datasets highlighted four key genes: TIAM1, RAP1GAP, GPX3, and JUN. GO analysis linked these genes to "response to oxidative stress" and "regulation of GTPase activity". KEGG pathway analysis demonstrated significant enrichment in pathways including "Glutathione metabolism", "cAMP signaling pathway", "Rap1 signaling pathway", "Tight junction", and "Thyroid hormone synthesis". Further, single-gene GSEA analyses suggested distinct pathways through which each gene may influence TC progression. Immune profiling revealed marked differences in immune cell populations, notably CD8+ T cells, monocytic lineage cells, neutrophils, NK cells, and T cells, between normal and cancerous thyroid tissues. Notably, RAP1GAP, GPX3, and JUN were implicated in the regulation of Treg and follicular helper T cell functions. The differential expression of these genes was rigorously validated using TCGA dataset and six additional GEO datasets. While the tumor-suppressive roles of TIAM1 and RAP1GAP have been previously established, our findings reveal that the overexpression of GPX3 and JUN significantly impairs the proliferative and migratory capacities of TC cells, underscoring their potential as therapeutic targets. <b>Conclusion:</b> This study identifies GPX3 and JUN as critical tumor suppressor genes in TC, with their function closely linked to T regulatory cells and follicular helper T cells. The overexpression of GPX3 and JUN demonstrates significant tumor-suppressive activity, highlighting their potential as effective therapeutic targets in combating TC.