Thyroid cancers are among the most common endocrine malignancies and encompass a broad spectrum of biological behaviors ranging from indolent to highly aggressive forms. Accurate prognostic stratification at the time of diagnosis is crucial for optimal patient management and treatment for patients with thyroid cancer. The telomerase reverse transcriptase (TERT) promoter mutation has emerged as an essential molecular marker, predicting the aggressive clinical course in thyroid carcinomas. The TERT promoter hotspot mutation (C228T and C250T) serves as a major indicator of unfavorable prognosis in differentiated thyroid carcinomas, and their frequency escalates from well-differentiated to poorly differentiated and to anaplastic thyroid carcinomas (ATCs) [1,2]. The presence of TERT promoter mutation is associated with resistance to radioactive iodine treatment, distant metastasis, tumor recurrence, and the dedifferentiation process in thyroid cancers [1,[3], [4], [5]].

To detect the presence of TERT promoter mutations, molecular methods including Sanger sequencing and next-generation sequencing have been usually employed in the clinical setting. However, these techniques can be time-consuming, technically demanding, and not always cost-effective for routine screening. Immunohistochemistry (IHC) can stand out as a potential alternative method because of its well-established protocols, affordability, and simplicity [6]. While TERT IHC might not be suitable for predicting the presence of TERT promoter mutations in follicular thyroid carcinoma (FTC) [7], its efficacy in other types of thyroid carcinomas remains less elucidated.

Beyond the genetic mutations, epigenetic modifications have emerged as essential prognostic determinants in thyroid tumors. DNA methylation, a crucial epigenetic mechanism, plays an essential role in regulating gene expression. Abnormalities in DNA methylation, such as hypermethylation of tumor suppressors and hypomethylation of oncogenes, have been related to possible tumorigenesis in thyroid cancer [8]. 5-Hydroxymethylcytosine (5hmC) is an intermediate in the DNA demethylation pathway, and recent studies have suggested a marked decrease in 5hmC levels in various cancers [9]. A recent study reported the relevance of 5hmC loss to TERT promoter mutations in papillary thyroid carcinoma (PTC), proposing that 5hmC IHC may offer an alternative method to predict the presence of TERT promoter mutation [10]. Furthermore, the potential role of 5hmC as a valuable biomarker for predicting the presence of lymph node (LN) metastasis in PTCs has also been suggested [11].

In this study, we investigated the immunohistochemical expression of TERT and 5hmC according to the status of TERT promoter mutations in various thyroid carcinomas. This study aimed to elucidate the diagnostic utility of these markers in predicting the presence of TERT promoter mutations and the aggressiveness of various thyroid carcinomas.