Distantly metastatic differentiated thyroid carcinoma is kinase-driven and enriched for DNA repair and DNA methylation gene alterations.

Although distant metastasis is uncommon in differentiated thyroid carcinoma (DTC), it remains the leading cause of thyroid cancer-related mortality. The genetic landscape of distantly metastatic DTC (DMDTC) has not been well characterized in large cohorts. This study aimed to identify functional genetic alterations in DMDTC and validate their biological significance. We included 78 patients with DMDTC and performed DNA-based next-generation sequencing (NGS) in all cases, followed by RNA-based NGS for fusion gene detection, along with a review of previously reported isolated cases. Plasmids harbouring novel variants, including SPON1::ALK and RFTN1::BRAF fusions, and mutations in PTEN (c.322_345del, c.740del, c.968dup), STK11 (c.842C>T, c.1225C>T), and DNMT3A (c.891G>A, c.2312G>A, c.2595A>T, c.2606G>A) were constructed and transfected into TPC-1 and HEK293T cells to investigate downstream signalling. The methylation status of differentially methylated genes (DMGs) associated with DNMT3A mutations was analysed using the Infinium MethylationEPIC v2.0 BeadChip, with several DMGs validated by real-time quantitative PCR. The cohort consisted of 25 males and 53 females, with a mean age of 60.3 years at the diagnosis of metastasis. Histological types included papillary carcinoma (31 cases), follicular carcinoma (44 cases), and oncocytic carcinoma (3 cases). The lung and bone were the most common metastatic sites. Multiple metastases and older age were associated with metastasis-free and overall survival. Genetic alterations involving phosphorylation signalling pathways were identified in 61 cases, among which pathological alterations of DNA damage repair (DDR)-related genes were detected in ten cases. Novel RFTN1::BRAF and SPON1::ALK fusions, along with PTEN (c.740del, c.968dup) and STK11 (c.842C>T) mutations, could enhance downstream phosphorylation levels. DNMT3A mutations (c.891G>A, c.2312G>A, c.2595A>T, c.2606G>A) induced genome-wide methylation dysregulation, with altered expression of SLC12A7, FLNC, HMGB2, BNC2, and DAPK1. This study shows that DMDTCs are characterized by dysregulated phosphorylation signalling, accompanied by chromosomal instability and aberrant methylation, thus underscoring DDR gene-targeted therapy as a promising strategy. © 2026 The Pathological Society of Great Britain and Ireland.