Selective inactivation of DNA-dependent protein kinase with antisense oligodeoxynucleotides: consequences for the rejoining of radiation-induced DNA double-strand breaks and radiosensitivity of human cancer cell lines.
The inhibition of DNA-dependent protein kinase activity with antisense-oligodeoxynucleotide (As-ODN) and its consequences for the rejoining of DNA-double-strand breaks (Dsbs) and radiation sensitivity was studied in human non-small cell lung cancer (NSCLC) cell lines. Cells were transfected with As-ODNs specific for the catalytic subunit of DNA-dependent protein kinase (DNA-PKcs). In comparison, cells were treated with Wortmannin, a potent but nonspecific inhibitor of DNA-PK activity. As-ODN efficiently reduced the kinase activity with an IC50 of about 100-200 IC50 of Wortmannin was at approximately 5-10 micro M. Treatment of cells with 300 nM As-ODN increased the fraction of residual Dsb at 4 h after irradiation by a factor of 4.4, 2.6, and 1.7 in A549, H460, and H661 cells, respectively. The respective values after treatment with 20 micro M Wortmannin were 5.3, 4.3, and 2.2. Inhibition of DNA-PK activity by As-ODN and Wortmannin also decreased the surviving fraction of the NSCLC cell lines. These data show that kinase activity of DNA-PKcs can be specifically inhibited with As-ODN as effective as Wortmannin and results in marked inhibition of DNA-Dsb rejoining and radiosensitization of NSCLC cell lines.
Sak
Ali
A
Department of Radiotherapy, University Essen, 45122 Essen, Germany. ali.sak@uni-essen.de
Stuschke
Martin
M
Wurm
Reinhard
R
Schroeder
Gisela
G
Sinn
Brigitte
B
Wolf
Gudrun
G
Budach
Volker
V
D000730:Androstadienes / Q000494:pharmacology; D002289:Carcinoma, Non-Small-Cell Lung / Q000201:enzymology* / Q000235:genetics / Q000532:radiotherapy; D004249:DNA Damage; D004260:DNA Repair / Q000187:drug effects* / Q000235:genetics; D004273:DNA, Neoplasm / Q000235:genetics / Q000528:radiation effects; D051747:DNA-Activated Protein Kinase; D004268:DNA-Binding Proteins; D004789:Enzyme Activation; D006801:Humans; D008175:Lung Neoplasms / Q000201:enzymology* / Q000235:genetics / Q000532:radiotherapy; D009687:Nuclear Proteins; D016376:Oligonucleotides, Antisense / Q000235:genetics / Q000494:pharmacology*; D017346:Protein Serine-Threonine Kinases / Q000037:antagonists & inhibitors* / Q000235:genetics / Q000378:metabolism; D012333:RNA, Messenger / Q000096:biosynthesis / Q000235:genetics / Q000378:metabolism; D011836:Radiation Tolerance / Q000187:drug effects* / Q000235:genetics; D014162:Transfection; D014407:Tumor Cells, Cultured; D000077191:Wortmannin
D000730:Androstadienes; D004273:DNA, Neoplasm; D004268:DNA-Binding Proteins; D009687:Nuclear Proteins; D016376:Oligonucleotides, Antisense; D012333:RNA, Messenger; D051747:DNA-Activated Protein Kinase; C497263:PRKDC protein, human; D017346:Protein Serine-Threonine Kinases; D000077191:Wortmannin