Much like replicative senescence, the irreversible cell-cycle arrest induced by eroded telomeres, accelerated senescence occurs when replicative cells suffer irreparable
DNA double-strand breaks (DSBs). Along with apoptosis and
necrosis, senescence is a desirable outcome in
cancer treatment with ionizing radiation (IR) or
chemotherapy. In both normal and
cancer cells, DSBs promote the assembly of IR-induced foci (IRIF), domains of modified
chromatin that serve a key role in DNA damage signaling. IRIF persistence is a critical determinant of accelerated senescence, making drugs that promote persistent IRIF an attractive strategy to sensitize
cancer to genotoxic
therapy. As an IRIF reporter, we have expressed an inducible
green fluorescent protein (GFP) fusion to the IRIF-binding domain (IBD) of 53BP1 (GFP-IBD) in the
breast cancer cell line MCF7. Within minutes of exposure to IR, the GFP-IBD relocalizes to form fluorescent nuclear foci, which disperse within several hours. A pair of high-content screening assays for IRIF formation and persistence were established in multiwell plates based on imaging and quantifying GFP-IBD foci per Hoechst-stained MCF7 nucleus at 2 hours and 24 hours. Using the
ataxia telangiectasia-mutated inhibitor
CGK733 to block IRIF formation and the
topoisomerase II inhibitor etoposide to prevent IRIF resolution, we obtained a Z' >0.8 both for IRIF formation at 2 hours and IRIF persistence at 24 hours. Screening the diverse drugs and natural products in the National Cancer Institute Developmental
Therapeutics Program Approved Oncology Drugs Set, the National Institutes of Health Clinical Collection, and the MicroSource Spectrum Collection yielded multiple hits that significantly delayed IRIF resolution. Secondary screening suggested some of these otherwise nontoxic drugs also enhance accelerated senescence, indicating strong potential for their repurposing as
radiation sensitizers to improve the efficacy of
cancer therapy.