[Inhibition of 8-chloroadenosine on proteasome activity of HL-60 cells].

Abstract

BACKGROUND & OBJECTIVE:

The proteasome is a multicatalytic proteinase complex in eukaryotic cells, and it can be used as a new target for screening anti-tumor drugs according to certain previous researches. The inhibition activity of 8-CA (8-chloroadenosine, a new anti-tumor drug synthesized in our lab) in sarcoma 180, hepatocarcinoma 22, and human gastric carcinoma xenograft tumor has been confirmed. However,the relationship between the mechanism of the inhibition and proteasome is still unclear. The present study was designed to explore the effect of 8-CA on 3 kinds of enzyme activities (chymotrypsin-like activity, trypsin-like activity, and peptidyl-glutamyl- peptide hydrolyzing activity) of 20S proteasome in human progranulocyte leukemia HL-60 cells.

METHODS:

HL-60 cells were treated with 8-CA at different concentrations for 24, 48, and 72 hours. Then total protein of HL-60 cells was extracted, and the three enzyme activities of 20S proteasomes in the total protein was examined. And the chymotrypsin-like activity of the purified proteasome from the total protein was also examined by immunoprecipitation. The enzyme activity was indicated by fluorescence absorbance (A) produced by degradation of specific substrates by proteasomes.

RESULTS:

In HL-60 cells, three enzyme activities of the proteasomes were significantly inhibited after exposure to 0.1 micromol/L 8-CA for 48 hours; and the inhibition effect was concentration dependent. When the concentration of 8-CA reached 5 micromol/L, the inhibition rates of three proteasome activities (chymotrypsin-like activity,trypsin-like activity,and peptidyl- glutamyl-peptide hydrolyzing activity) were 44.96%, 54.52%, 48.36% for 48 hours and 67.53%, 70.48%, 64.08% for 72 hours, respectively. The inhibition rate of 1 micromol/L and 5 micromol/L 8-CA on chymotrypsin-like proteasome activity of HL-60 cells for 48 hours were 68.14% (P< 0.01) and 92.75% (P< 0.001), respectively.

CONCLUSION:

8-CA inhibits proteasome activity of HL-60 cells with a time- and concentration-dependent manner.

• Authors: Yi Zhang, Jun Wu, Bo Xu, Li-He Zhang, Jing-Rong Cui
• Journal: Ai zheng = Aizheng = Chinese journal of cancer (Ai Zheng) Vol. 23 Issue 2 Pg. 155-9 (Feb 2004) China
• PMID: 14960234 (Publication Type: Journal Article, Research Support, Non-U.S. Gov't)

Chemical References

• Multienzyme Complexes
• NF-kappa B
• 2-Chloroadenosine
• Cysteine Endopeptidases
• Proteasome Endopeptidase Complex
• 8-chloroadenosine

Topics

• 2-Chloroadenosine (analogs & derivatives, pharmacology)
• Apoptosis (drug effects)
• Cysteine Endopeptidases
• HL-60 Cells (drug effects)
• Humans
• Multienzyme Complexes (antagonists & inhibitors)
• NF-kappa B (metabolism)
• Precipitin Tests
• Proteasome Endopeptidase Complex