Ataxia Telangiectasia Mutated and p21CIP1 Modulate Cell Survival of Drug-Induced Senescent Tumor Cells: Implications for Chemotherapy

Abstract

Purpose: Premature or stress-induced senescence is a major cellular response to chemotherapy in solid tumors and contributes to successful treatment. However, senescent tumor cells are resistant to apoptosis and may also reenter the cell cycle.We set out to find a means to specifically induce senescent tumor cells to undergo cell death and not to reenter the cell cycle that may have general application in cancer therapy. Experimental Design: We investigated the mechanisms regulating cell survival in drug-induced senescent tumor cells. Using immunofluorescence and flow cytometry^based techniques, we established the status of the ataxia telangiectasia mutated (ATM) signaling pathway in these cells.We assayed the requirement of ATM signaling and p21 CIP1 expression for survival in premature senescent tumor cells using pharmacologic inhibitors and antisense oligonucleotides. Results: The ATM/ATR (ATM-and Rad3-related) signaling pathway was found to be constitutively active in drug-induced senescent tumor cells. We found that blocking ATM/ATR signaling with pharmacologic inhibitors, including the novel ATM inhibitors KU55933 and CGK733, induced senescent breast, lung, and colon carcinoma cells to undergo cell death. We show that the mechanism of action of this effect is directly via p21 CIP1

Down-regulation of Wild-type p53-induced Phosphatase 1 (Wip1) Plays a Critical Role in Regulating Several p53-dependent Functions in Premature Senescent Tumor Cells

“…Induction of Premature Senescence, Senescence-associated ␤-Galactosidase Activity, and Micronuclei Detection-Unless otherwise stated, senescence was induced by treating cells with the DNA-damaging agents doxorubicin (200 nM for MCF-7 cells and 600 nM for A549 cells) for 72 h as previously described (20). Because forced Wip1 expression leads to death of premature senescent cells, all the experiments were conducted from 1 to 7 days after drug release.…”

“…Immunofluorescence Microscopy-Detection of ␥-H2AX, phosphorylated ATM (Ser 1981 ) and phosphorylated Chk2 (Thr 68 ) was performed as previously described (20). The anti-␥-H2AX antibodies (JBW301) were purchased from Upstate Biotechnology (Milton Keynes, UK), and anti-phospho-ATM (Ser 1981 ) and anti-phospho-Chk2 (Thr 68 ) were from Cell Signaling (Beverly, MA).…”

“…To investigate the role of Wip1 phosphatase in modulating premature senescence in tumor cells, we treated the lung adenocarcinoma cell line A549 with doxorubicin for 72 h as previously described (20). We analyzed Wip1 protein level during drug treatment and thereafter during the development of the senescent phenotype over several days after the initial DNA damage.…”

NF-κB-dependent cytokine secretion controls Fas expression on chemotherapy-induced premature senescent tumor cells

“…Induction of premature senescence and senescence-associated b-galactosidase (SA-b-gal) activity Unless otherwise stated, senescence was induced by treating cells with the DNA-damaging agents doxorubicin (200 nM for MCF-7 cells and 600 nM for A549 cells) for 72 h, as previously described (Crescenzi et al, 2008). To allow the development of a fully senescent phenotype, cells were analyzed from 4 to 21 days after replating.…”

Long-Term In vivo Effects of Cisplatin on γ-H2AX Foci Signaling in Peripheral Lymphocytes of Tumor Patients After Irradiation

“…An increased inhibition of radiationinduced g-H2AX foci by 60% was seen using ATM and DNAdependent protein kinase inhibitors. The small molecule compounds NU7027, Ku55933, and CGK733 used in the present study are potent and specific kinase inhibitors for DNAdependent protein kinase (23), ATM (24), and ATM/ataxia telangiecasia and Rad3 related (25), respectively. The results indicate that ATM and DNA-dependent protein kinase are involved in radiation-induced g-H2AX phosphorylation in resting human lymphocytes in a partly overlapping and competing manner.…”