Selective Therapeutic Potential of a H2O2-Inducible DNA Interstrand Cross-linker in Anaplastic Thyroid Carcinoma

Abstract

We aimed to investigate hydrogen peroxide-inducible DNA interstrand cross-link (HP-ICL) as a targeted therapy for anaplastic thyroid cancer (ATC) due to its higher H2O2 content than normal cells. In vitro analysis included fluorescence microscopy for H2O2 levels and exposure of ATC cells to various HP-ICL concentrations followed by assessment of cell viability, apoptosis, cell cycle, and DNA damage using methyl thiazolyl tetrazolium (MTT), flow cytometry, and a γH2AX assay. Protein levels related to apoptosis and the phosphatidylinositol-3-kinase/protein kinase B/mammalian target of rapamycin (PI3K/AKT/mTOR) pathway were measured by Western blotting. An ATC xenograft mouse model was used to evaluate the HP-ICL's in vivo effects. ATC cells had higher H2O2 levels than normal thyroid cells. HP-ICL treatment caused a dose-dependent decrease in cell viability and an increase in apoptosis, with a slight G2/M phase arrest. A 30 µM HP-ICL treatment doubled γH2AX foci. Bcl-2 levels decreased, while Bax, cleaved-Caspase 3, and PARP increased in a dose-dependent manner. It also inhibited p-PI3K, p-AKT, and p-mTOR. In vivo, the HP-ICL significantly inhibited tumor growth while maintaining body weight and without causing organ damage or altering thyroid hormone levels. Additionally, tumor sections exhibited increased TUNEL staining, decreased Ki67 expression, and reduced levels of p-PI3K, p-AKT, and p-mTOR. The HP-ICL significantly inhibited ATC both in vitro and in vivo, suggesting its potential as an effective therapy for ATC.

Keywords: DNA cross-linking agents; apoptosis; hydrogen peroxide; thyroid cancer.