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A multistep model for paclitaxel-induced apoptosis in human breast cancer cell lines
A L Blajeski1, T J Kottke, S H Kaufmann
1Department of Molecular Pharmacology and Experimental Therapeutics, Mayo Clinic, Rochester, Minnesota 55905, USA.
Abstract:
Despite extensive previous investigation, the events occurring between paclitaxel-induced mitotic arrest and the subsequent onset of apoptosis remain incompletely understood. In the present study, the sequential morphological and biochemical changes that occur after paclitaxel treatment were examined in MDA-MB-468 (p53 mutant) and MCF-7 (p53 wild-type) breast cancer cells. Flow cytometry indicated that paclitaxel induces tetraploidy that persists until the onset of apoptosis in both cell lines. Light and electron microscopy indicated that the cells transiently arrest in mitosis and then enter a multinucleated interphase state characterized by the absence of punctate staining for CENP-F, a G(2) marker, but the presence of cyclin E, a G(1) cyclin, and p21(waf1/cip1), a cyclin-dependent kinase inhibitor. Despite high p21(waf1/cip1) levels, paclitaxel-treated cells incorporated thymidine into DNA. Aphidicolin inhibited this DNA synthesis but not the subsequent onset of apoptosis. Conversely, the broad-spectrum caspase inhibitor benzyloxycarbonyl-val-ala-asp(OMe)-fluoromethylketone inhibited apoptosis and enhanced the number of multinucleated cells but did not facilitate generation of octaploid cells. These results are consistent with a multistep model in which breast cancer cells exposed to paclitaxel undergo an aberrant mitotic exit; proceed through a tetraploid, multinucleated G(1) state; initiate an aphidicolin-suppressible process of DNA repair; and subsequently undergo apoptosis.
Insights
Paclitaxel treatment causes breast cancer cells to arrest in mitosis, become multinucleated, and initiate DNA repair before apoptosis. This study clarifies the cell death pathway following paclitaxel exposure.
Area of Science:
- Cell Biology
- Cancer Research
- Pharmacology
Background:
- Paclitaxel is a key chemotherapy agent, but the precise mechanisms linking its induction of mitotic arrest to apoptosis are not fully understood.
- Investigating these intermediate events is crucial for understanding drug resistance and developing more effective cancer therapies.
Purpose of the Study:
- To elucidate the sequential morphological and biochemical events in breast cancer cells following paclitaxel treatment.
- To differentiate the roles of DNA synthesis and caspase activity in paclitaxel-induced cell death.
Main Methods:
- Utilized MDA-MB-468 (p53 mutant) and MCF-7 (p53 wild-type) breast cancer cell lines.
- Employed flow cytometry, light and electron microscopy, and biochemical assays.
- Investigated the effects of aphidicolin (DNA synthesis inhibitor) and a caspase inhibitor.
Main Results:
- Paclitaxel induced persistent tetraploidy and a multinucleated interphase state in both cell lines.
- Cells exhibited DNA synthesis despite high p21(waf1/cip1) levels, which was inhibited by aphidicolin.
- Caspase inhibition blocked apoptosis but increased multinucleated cells, without promoting octaploidy.
Conclusions:
- Paclitaxel-treated breast cancer cells undergo aberrant mitotic exit and a tetraploid, multinucleated G1 phase.
- An aphidicolin-suppressible DNA repair process precedes apoptosis.
- This multistep model provides new insights into paclitaxel's mechanism of action in breast cancer.