Related Experiment Videos
[The mechanism of apoptosis induced by anticancer agents in human leukemia HL-60 cells]
A Yoshida1, T Ueda, T Nakamura
1First Department of Internal Medicine, Fukui Medical School.
Abstract:
The present study was designed to investigate the contribution of proteases in anticancer agents-induced apoptosis of human leukemia HL-60 cells. A serine protease inhibitor TPCK and an ICE-like protease inhibitor VAD-FMK prevented etoposide, camptothecin and ara-C-induced internucleosomal DNA cleavage. Using a cell-free system, we have examined the inhibitory mechanism of these inhibitors on etoposide-induced internucleosomal DNA cleavage. We found that cell lysates prepared from etoposide-treated HL-60 cells undergoing apoptosis contain the significant activity to induce internucleosomal DNA fragmentation in isolated nuclei. On the other hand, we could not detect such activity in the cell lysates from untreated HL-60 cells. Treatment of the cell lysates with a serine protease inhibitor TPCK abrogated the DNA fragmenting activity. An ICE-like protease inhibitor VAD-FMK had no effect on this DNA fragmenting activity in vitro. However, the formation of TPCK-sensitive DNA fragmenting activity in etoposide-treated cells was blocked by the VAD-FMK. These data indicate that serine and ICE-like proteases may be involved in etoposide-induced apoptosis at the different stages, and especially a serine protease may be closely associated with the final step for induction of DNA fragmentation during apoptosis in HL-60 cells.
Insights
Anticancer drugs trigger apoptosis in leukemia cells by activating proteases. A serine protease inhibitor blocked DNA fragmentation, suggesting its key role in the final stages of apoptosis.
Area of Science:
- Molecular Biology
- Cell Biology
- Biochemistry
Context:
- Apoptosis, or programmed cell death, is crucial for development and disease.
- Proteases play significant roles in apoptotic pathways.
- Understanding protease involvement in anticancer drug action is vital for therapy.
Purpose:
- Investigate the role of proteases in anticancer agent-induced apoptosis in human leukemia HL-60 cells.
- Elucidate the specific mechanisms of protease inhibitors TPCK and VAD-FMK in etoposide-induced DNA cleavage.
Summary:
- Anticancer agents like etoposide, camptothecin, and ara-C induce internucleosomal DNA cleavage in HL-60 cells.
- Serine protease inhibitor TPCK and ICE-like protease inhibitor VAD-FMK prevented this DNA cleavage.
- Cell lysates from etoposide-treated cells showed DNA fragmentation activity, inhibited by TPCK but not VAD-FMK in vitro.
- VAD-FMK blocked the formation of TPCK-sensitive activity in treated cells, indicating sequential protease involvement.
Impact:
- Serine and ICE-like proteases are involved in etoposide-induced apoptosis at different stages.
- A serine protease appears critical for the final DNA fragmentation step in HL-60 cell apoptosis.
- Findings may inform the development of targeted therapies for leukemia.