DNA-PK inhibition by NU7441 sensitizes breast cancer cells to ionizing radiation and doxorubicin
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DNA-dependent protein kinase (DNA-PK) plays a key role in the repair of DNA double-strand breaks (DSBs) that are probably the most deleterious form of DNA damage. Inhibition of DNA-PK has been considered as an attractive approach to decrease resistance to therapeutically induced DNA DSBs. Ionizing radiation (IR) and doxorubicin, which induce DSBs, are used in the treatment of breast cancer. We determined the cellular concentration of DNA-PK and other DSB-activated kinases: ATM and ATR and the effect of DNA-PK inhibition by NU7441 on DNA repair, cell cycle, and survival after IR or doxorubicin treatment in three human breast cancer cell lines (MCF-7, MDA-MB-231, and T47D) representing different breast cancer subtypes. T47D cells had the highest expression of DNA-PKcs, ATM, and ATR and the most rapid rate of DNA DSB repair. IR caused a 10- to 16-fold increase in DNA-PK activity and two to threefold induction of ATM in all 3 cell lines. NU7441 inhibited IR-induced DNA-PK activity in all cell lines with IC50s in the range 0.17–0.25 μM. NU7441 retarded the repair of DSB and significantly increased the sensitivity of all cell lines to IR (4- to 12-fold) and doxorubicin (3- to 13-fold). The greatest sensitiziation by NU7441 was observed in MDA-MB-231 cells. NU7441 affected the cell cycle distribution in all studied cell lines; increasing accumulation of cells in G2/M phase after DNA damage. Our data indicate that DNA-PK might be an effective target for chemo- and radio-potentiation in breast cancer and suggest that further development of DNA-PK inhibitors for clinical use is warranted.
KeywordsDNA-PK Ionizing radiation Doxorubicin Breast cancer
This work was partially funded by LLP/Erasmus Training Program and the Polish Ministry of Science and Higher Education (Grant No. 2 P04A 034 30).
Conflict of interest
The authors declare that they have no conflict of interest.
All experiments complied with the current laws of UK and Poland.
- 3.Andre F, Zielinski CC (2012) Optimal strategies for the treatment of metastatic triple-negative breast cancer with currently approved agents. Ann Oncol 23 Suppl 6:vi46–51Google Scholar
- 11.Taccioli GE, Amatucci AG, Beamish HJ, Gell D, Xiang XH, Torres Arzayus MI, Priestley A, Jackson SP, Marshak Rothstein A, Jeggo PA, Herrera VL (1998) Targeted disruption of the catalytic subunit of the DNA-PK gene in mice confers severe combined immunodeficiency and radiosensitivity. Immunity 9(3):355–366PubMedCrossRefGoogle Scholar
- 12.Rosenzweig KE, Youmell MB, Palayoor ST, Price BD (1997) Radiosensitization of human tumor cells by the phosphatidylinositol3-kinase inhibitors wortmannin and LY294002 correlates with inhibition of DNA-dependent protein kinase and prolonged G2-M delay. Clin Cancer Res 3(7):1149–1156PubMedGoogle Scholar
- 14.Peng Y, Zhang Q, Nagasawa H, Okayasu R, Liber HL, Bedford JS (2002) Silencing expression of the catalytic subunit of DNA-dependent protein kinase by small interfering RNA sensitizes human cells for radiation-induced chromosome damage, cell killing, and mutation. Cancer Res 62(22):6400–6404PubMedGoogle Scholar
- 19.Willmore E, Elliott SL, Mainou-Fowler T, Summerfield GP, Jackson GH, O’Neill F, Lowe C, Carter A, Harris R, Pettitt AR, Cano-Soumillac C, Griffin RJ, Cowell IG, Austin CA, Durkacz BW (2008) DNA-dependent protein kinase is a therapeutic target and an indicator of poor prognosis in B-cell chronic lymphocytic leukemia. Clin Cancer Res 14(12):3984–3992PubMedCrossRefGoogle Scholar
- 22.Song H, Hedayati M, Hobbs RF, Shao C, Bruchertseifer F, Morgenstern A, Deweese TL, Sgouros G (2013) Targeting aberrant DNA double strand break repair in triple negative breast cancer with alpha particle emitter radiolabeled anti-EGFR antibody. Mol Cancer TherGoogle Scholar
- 24.Douglas P, Cui X, Block WD, Yu Y, Gupta S, Ding Q, Ye R, Morrice N, Lees-Miller SP, Meek K (2007) The DNA-dependent protein kinase catalytic subunit is phosphorylated in vivo on threonine 3950, a highly conserved amino acid in the protein kinase domain. Mol Cell Biol 27(5):1581–1591PubMedCentralPubMedCrossRefGoogle Scholar
- 33.Kao J, Salari K, Bocanegra M, Choi YL, Girard L, Gandhi J, Kwei KA, Hernandez-Boussard T, Wang P, Gazdar AF, Minna JD, Pollack JR (2009) Molecular profiling of breast cancer cell lines defines relevant tumor models and provides a resource for cancer gene discovery. PLoS ONE 4(7):e6146PubMedCentralPubMedCrossRefGoogle Scholar
- 34.Neve RM, Chin K, Fridlyand J, Yeh J, Baehner FL, Fevr T, Clark L, Bayani N, Coppe JP, Tong F, Speed T, Spellman PT, DeVries S, Lapuk A, Wang NJ, Kuo WL, Stilwell JL, Pinkel D, Albertson DG, Waldman FM, McCormick F, Dickson RB, Johnson MD, Lippman M, Ethier S, Gazdar A, Gray JW (2006) A collection of breast cancer cell lines for the study of functionally distinct cancer subtypes. Cancer Cell 10(6):515–527PubMedCentralPubMedCrossRefGoogle Scholar