چکیده (انگلیسی):

Apoptosis is associated with cascades of
biochemical changes, including caspase activation,
cleavage of poly-ADP-ribose polymerase (PARP),
and fragmentation of genomic DNA. Knowledge of
the kinetics of these changes in drug-induced
apoptosis is important for designing
pharmacodynamic studies. We have shown that the
slow manifestation of apoptosis contributes to the
delayed pharmacological effects of paclitaxel (Cancer
Res. 58:2141-2148, 1998). The present study
examined the timing of the biochemical changes in
paclitaxel-induced apoptosis in human prostate PC3
cancer cells. After treatment with 20 nM paclitaxel,
the fraction of cells that detached from the culture
flask increased with time to reach 68% at the end of
the 96-hour experiment. In contrast, the control
samples showed <1% detachment. The attached and
detached paclitaxel-treated cells showed different
biochemical properties. The detached cells exhibited
the full spectrum of apoptotic changes, whereas the
attached cells only showed activation of caspase-3-
like proteases but not PARP cleavage, DNA
fragmentation, nor release of DNA fragments to the
cytoplasm. Activation of caspases in the attached cells
was several-fold lower and occurred at a later time (ie,
24 vs 12 hours) compared to the detached cells. In the
detached cells, caspase activation was first detected at
12 hours and peaked at 36 hours, whereas PARP
cleavage was first detected at 24 hours and was
completed prior to 72 hours. In contrast, the extent of
internucleosomal DNA fragmentation and the release
of DNA-histone complex to the cytoplasm (both were
first detected at 24 hours) were cumulative over time
up to the last time point of 96 hours. In summary, in
paclitaxel-induced apoptosis, caspase activation was
followed with a 12-hour lag time by PARP cleavage,
internucleosomal DNA fragmentation, and release of
DNA-histone complex to the cytoplasm. There was no
detectable lag time between PARP cleavage and DNA
fragmentation. The observation that only the detached
cells but not the attached cells showed the full
spectrum of apoptotic changes suggests that
detachment is either a part of the initiation/execution
phases of apoptosis and/or is required for their
completion