<p>Aptamers are short oligonucleotides capable of binding to various molecular targets with high affinity and specificity. These short sequences are conventionally selected through the systematic evolution of ligands by exponential enrichment (SELEX) process. In this study, the non-SELEX in silico strategy was used to simulate the process of aptamer synthesis and subsequent affinity evaluation. We hypothesized that a candidate RNA aptamer could function as an antagonist to nuclear thyroid hormone receptors (TRs), thereby inhibiting their interaction with thyroid hormone response elements (TREs). Using knowledge-based approaches, TRE sequences were retrieved from the literature, and representative loci across the human genome were modeled. Through RNA structure prediction, molecular docking, and molecular dynamics simulations, several single-stranded RNA aptamers with strong binding affinity toward TRs were identified. Among them, one candidate demonstrated the most favorable interaction with thyroid hormone receptor alpha. Pending experimental validation, this aptamer holds potential as a novel therapeutic agent for hyperthyroidism by acting as a TR-blocking molecule.</p>