p53-deficient cells display increased sensitivity to anthracyclines after loss of the catalytic subunit of the DNA-dependent protein kinase.
DNA-dependent protein kinase (DNA-PK) is involved in non-homologous end joining which repairs DNA double-strand breaks introduced by irradiation and radiomimetic agents. DNA-PK interacts with p53 but may also have p53-independent functions. The present study investigated whether disruption of the gene for the catalytic subunit DNA-PKcs affects chemosensitivity in p53-deficient cells. Drug sensitivity of DNA-PKcs(+/+)/p53(+/+), DNA-PKcs(+/+)/p53(-/-), DNA-PKcs(-/-)/p53(+/+), and DNA-PKcs(-/-)/p53(-/-) mouse lung-fibroblasts was determined by the MTT assay, the clonogenic assay, and trypan blue exclusion. Susceptibility to apoptosis was determined by DNA fragmentation (TUNEL) and by caspase-3 cleavage. We show that p53-deficient cells were 2 to 3-fold resistant to treatment with doxorubicin, epirubicin, cisplatin, and docetaxel as compared to wild-type cells. We further demonstrate that the additional loss of DNA-PKcs function in p53-deficient cells resulted in a 2-fold increase in sensitivity to doxorubicin and epirubicin as documented by the MTT assay, clonogenic assay, and trypan blue exclusion. Doxorubicin-induced hypersensitivity in these cells correlated with a transient G2/M checkpoint activation but did not seem to correlate with apoptosis. The data indicate that additional loss of DNA-PKcs in p53-deficient cells reverses anthracycline-resistance imposed by p53-deficiency, and that DNA-PKcs modulates p53-independent pathways responding to DNA damage induced by anthracyclines. They also indicate that processes other than apoptosis may contribute to the increased cytotoxicity to anthracyclines. DNA-PKcs may thus be a potential target for functional inhibition, which might increase the efficacy of some anti-tumour agents in the treatment of cancers mutated in the p53 gene.
Fedier
André
A
Division of Gynaecology, Department of Obstetrics and Gynaecology, University Hospital of Zurich, Switzerland.
Moawad
Ahmed
A
Haller
Urs
U
Fink
Daniel
D
D000818:Animals; D018943:Anthracyclines / Q000494:pharmacology*; D000970:Antineoplastic Agents / Q000494:pharmacology; D017209:Apoptosis; D053148:Caspase 3; D020169:Caspases / Q000378:metabolism; D020134:Catalytic Domain; D002460:Cell Line; D004249:DNA Damage; D053938:DNA Fragmentation; D004260:DNA Repair; D051747:DNA-Activated Protein Kinase; D004268:DNA-Binding Proteins; D004305:Dose-Response Relationship, Drug; D005347:Fibroblasts / Q000378:metabolism; D016195:G2 Phase; D020287:In Situ Nick-End Labeling; D020128:Inhibitory Concentration 50; D051379:Mice; D008822:Mice, Transgenic; D008938:Mitosis; D009154:Mutation; D017346:Protein Serine-Threonine Kinases / Q000737:chemistry / Q000502:physiology*; D013778:Tetrazolium Salts / Q000494:pharmacology; D013844:Thiazoles / Q000494:pharmacology; D013997:Time Factors; D016159:Tumor Suppressor Protein p53 / Q000378:metabolism / Q000502:physiology*
D018943:Anthracyclines; D000970:Antineoplastic Agents; D004268:DNA-Binding Proteins; D013778:Tetrazolium Salts; D013844:Thiazoles; D016159:Tumor Suppressor Protein p53; D051747:DNA-Activated Protein Kinase; D017346:Protein Serine-Threonine Kinases; C505409:Casp3 protein, mouse; D053148:Caspase 3; D020169:Caspases; C022616:thiazolyl blue