Investigating the role of prostaglandin system dysregulation in radiation resistance of triple negative breast cancer cells

Abstract

The overall goal of this research program was to elucidate the molecular mechanisms underlying radiation resistance in breast cancer. Prostaglandin (PG) signaling dysregulation has been implicated in various aspects of cancer progression, including tumor growth, metastasis, and inflammation, but its potential role in promoting radiation resistance remains underexplored. Among the key components of the PG system, prostaglandin-endoperoxide synthase 2 (PTGS2) emerged as a critical mediator potentially driving this resistance mechanism. Therefore, the primary aim of this study was to investigate the role of PTGS2 dysregulation in the development of radiation resistance in triple-negative breast cancer cells. To address this gap, radiation resistant variants of MDA-MB-231 cells were generated by repeated exposures to x-ray radiation (57 Gy total dose) followed by clonal cell selection. RT-qPCR and Western blot analysis revealed robust upregulation of PTGS2 mRNA and protein expression in radiation resistant cells compared to the parental MDA-MB-231Control cells. Whole-transcriptome analysis further corroborated these results, whereby radiation resistant variants demonstrated a 16.7-fold (p<0.05) increase in PTGS2 mRNA expression. In addition, 6 Gy radiation challenge further increased PTGS2 expression 48 hours post radiation exposure (76% increase; p<0.05) in radiation resistant variants, whereas the MDA-MB-231Control cells showed lack of PTGS2 expression changes. The next goal was to determine whether the upregulation of PTGS2 expression in MDA-MB-231 cells promoted the development of radiation resistance. Here, PTGS2 was overexpressed using the CRISPR/dCas9 system, and expression was verified using RT-qPCR and Western blotting. Surprisingly, PTGS2 overexpression led to decreased survival following radiation exposure. This suggests that although PTGS2 is upregulated in radiation-resistant cells, its overexpression alone makes the cells more susceptible to radiation-induced damage. In addition, PTGS2 overexpression reduced adhesion to fibronectin and increased anchorage independent growth. These changes suggest that PTGS2 promotes a more aggressive cancer phenotype by enhancing cell detachment and survival in non-anchorage conditions. Taken together, this study highlights the role of PTGS2 in the development of radioresistance, suggesting that PTGS2 and its downstream effectors within the prostaglandin signaling pathway may serve as promising therapeutic targets for therapy resistant cancers.