Abstract 5578: Role of differentiation driver gene HOXD4 in anaplastic thyroid cancer

Anaplastic thyroid cancer (ATC) is a rare and extremely aggressive subset of thyroid cancer, noted for its refractory nature and poor prognosis. Conventional cancer treatments are ineffective in treating ATC while targeted therapies are of limited use due to a lack of targetable genetic lesions in this cancer type thus rendering a five-year survival rate of 4%. The low mutational burden of ATC prompted analysis of differential gene expression patterns using NCBI Gene Expression Omnibus publicly available transcriptomic datasets (GSE33630, GSE85457). Bioinformatic analysis of ATC compared to normal thyroid cancer tissue was carried out using GEO2R software. Strikingly, differentiation driver gene Homeobox D4 (HOXD4) was significantly upregulated in ATC patient tissue samples by 3-fold and 12-fold respectively. Kaplan-Meier analysis indicates a significant decrease in survival in patients with high HOXD4 levels (HR: 3.08, logrank: 0.019). Unlike most cancers, ATC lacks differentiation therefore losing its thyroid-like morphology and function. Thus, the marked upregulation of differentiation gene HOXD4 warranted further exploration in vitro. HOXD4 expression was upregulated in ATC cell line T238 by 23-fold compared to immortalized “normal” thyroid epithelial cell line Nthy-ori-3-1. CRISPRi knockdown of HOXD4 in T238 and SW1736, another ATC cell line, was needed for functional analysis rendering HOXD4 knockdown cell lines T238-HOX and SW1736-HOX as well as non-specific oligonucleotides control cell line. Cell proliferation was reduced in T238-HOX by 36%, 29%, 28%, and 38% and SW1736-HOX by 51%, 56%, 59%, and 52% at 24, 48, 72, and 96 hours respectively. Migratory capacity measured via scratch wound assay was significantly reduced in T238-HOX by 31% and 13% and SW1736-HOX at 45% and 22% at 16 and 22 hours respectively. Migration through Boyden chambers decreased by 72% in T238-HOX and 62% in SW1736-HOX and invasion was reduced by 40% and 51% respectively. We are currently evaluating the impact of HOXD4 knockdown on viability, clonogenicity, and senescence. The data reveal the novelty of HOXD4 as a putative carcinogenic driver of ATC and its function as both a diagnostic and prognostic biomarker, with the potential to be used as a therapeutic target. We are currently investigating various differentiating agents and their efficacy in a possible clinical application with or without other therapeutic agents. Citation Format: Kaci Kopec, Nicole R. DeSouza, David Garber, Steven A. Hemmerdinger, Augustine Moscatello, Humayun K. Islam, Jan Geliebter, Raj K. Tiwari. Role of differentiation driver gene HOXD4 in anaplastic thyroid cancer [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2025; Part 1 (Regular Abstracts); 2025 Apr 25-30; Chicago, IL. Philadelphia (PA): AACR; Cancer Res 2025;85(8_Suppl_1):Abstract nr 5578.