Artikel
Bi-213 labelled anti-CD45 monoclonal antibody overcoming apoptosis-resistance by overcoming DNA-repair in HL-60 leukemia cells
Suche in Medline nach
Autoren
Veröffentlicht: | 20. März 2006 |
---|
Gliederung
Text
Aim: Targeted alpha-emitters such as Bi-213 are promising for treatment of a variety of cancer. However, the mechanisms by which alpha-emitters induce cell death are not well understood at the molecular level. Here, we examined induction of apoptosis, the role of activation of apoptosis pathways and induction of DNA-damage by Bi-213 labelled anti-CD45 monoclonal antibody ([Bi-213]anti-CD45) in apoptosis-sensitive/-resistant and radioresistant leukemia cells.
Methods: We examined induction of DNA damage and activation of apoptosis pathways of Bi-213 labelled anti-CD45 monoclonal antibody ([Bi-213]anti-CD45) in beta- and gamma-resistant HL-60 leukemia cells and in DNA-repair proficient and deficient cells. FACS analysis, Western blot analysis and Comet-assay were performed.
Results: We found a strong induction of apoptosis, activation of caspase-3, caspase-8 and caspase-9 and PARP cleavage in HL60 cells after treatment with 1, 3 or 10 Gy of [Bi-213]anti-CD45 after 24, 48 and 72 h. [Bi-213]anti-CD45-induced apoptosis was completely inhibited by z.VAD.fmk, a specific inhibitor of activation of caspases, indicating that caspases play an important role. Mitochondria were activated after [Bi-213]anti-CD45 treatment, resulting in caspase-9 activation. Bax, a death-promoting protein, was upregulated and Bcl-xL, a death-inhibiting protein, was downregulated after [Bi-213]anti-CD45 treatment. p21 were upregulated and cells showed a G2/M cell cycle arrest in a p53 deficient pathway after treatment with [Bi-213]anti-CD45. Upregulation of death inducing ligands such as CD95 ligand was not detected in HL-60 cells in contrast to the effect of beta- and gamma-irradiation, suggesting that [Bi-213]anti-CD45 induced apoptosis through the mitochondrial pathway. In addition, [Bi-213]anti-CD45 induced cell death in apoptosis-resistant HL-60 cells such as beta- and gamma-irradiation-resistant cells. [Bi-213]anti-CD45-triggered apoptosis was not inhibited by the DNA-repair mechanism of non homologous endjoining, predominant for repairing double strand breaks in mammalian cells.
Conclusions: Taken together we found that [Bi-213]anti-CD45 induces apoptosis and activates apoptosis pathways using exclusively the mitochondrial pathway. Induction of apoptosis is independent of CD95 receptor/CD95 ligand interaction and overrides DNA–repair. Bi-213-mediated alpha-radioimmunotherapy is a promising strategy for dose escalation of targeted cytotoxic therapy and single tumor cell kill.