Abstract

Cellular senescence is a fundamental biological process contributing to aging, often accompanied by extensive chromatin remodeling. Dynamic alterations of three‐dimensional (3D) genomic spatial structure, driven by chromatin reorganization, play a critical role in cell fate determination, but their relevance in therapy‐induced senescence (TIS) remains underexplored. Here, we perform an integrative multi‐omics analysis of Hi‐C, ATAC‐seq, CUT&RUN, and RNA‐seq in primary human fibroblasts undergoing TIS induced by ionizing radiation (RAD) or bleomycin (BLEO). We show that TIS leads to global chromatin decompaction, weakened compartmentalization, and destabilization of topologically associated domains (TADs), alongside widespread loss and rewiring of chromatin loops. Notably, RAD and BLEO elicit distinct changes in distance‐dependent compartment strength and enhancer–promoter (E‐P) loop patterns, reflecting divergent 3D regulatory programs. Importantly, TIS reshapes the chromatin environment around senescence‐associated secretory phenotype (SASP) genes, while their adjacent regions exhibit reduced chromatin interactions, allowing transcriptional activation. Our study reveals that 3D genome remodeling in TIS is highly plastic and context‐dependent and discloses spatial regulation of gene expression during therapy‐induced cellular senescence. In the course of TIS, cells undergo a profound epigenomic reorganization that underlies the development of a senescence‐associated phenotype and formation of an inflammatory microenvironment.