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Cell Death Associated with Abnormal Mitosis Observed by Confocal Imaging in Live Cancer Cells
Published on: August 21, 2013
Cell death associated with abnormal mitosis observed by confocal imaging in live cancer cells
Asher Castiel1, Leonid Visochek, Leonid Mittelman
1Cancer Research Center, Sheba Medical Center.
Abstract:
Phenanthrene derivatives acting as potent PARP1 inhibitors prevented the bi-focal clustering of supernumerary centrosomes in multi-centrosomal human cancer cells in mitosis. The phenanthridine PJ-34 was the most potent molecule. Declustering of extra-centrosomes causes mitotic failure and cell death in multi-centrosomal cells. Most solid human cancers have high occurrence of extra-centrosomes. The activity of PJ-34 was documented in real-time by confocal imaging of live human breast cancer MDA-MB-231 cells transfected with vectors encoding for fluorescent γ-tubulin, which is highly abundant in the centrosomes and for fluorescent histone H2b present in the chromosomes. Aberrant chromosomes arrangements and de-clustered γ-tubulin foci representing declustered centrosomes were detected in the transfected MDA-MB-231 cells after treatment with PJ-34. Un-clustered extra-centrosomes in the two spindle poles preceded their cell death. These results linked for the first time the recently detected exclusive cytotoxic activity of PJ-34 in human cancer cells with extra-centrosomes de-clustering in mitosis, and mitotic failure leading to cell death. According to previous findings observed by confocal imaging of fixed cells, PJ-34 exclusively eradicated cancer cells with multi-centrosomes without impairing normal cells undergoing mitosis with two centrosomes and bi-focal spindles. This cytotoxic activity of PJ-34 was not shared by other potent PARP1 inhibitors, and was observed in PARP1 deficient MEF harboring extracentrosomes, suggesting its independency of PARP1 inhibition. Live confocal imaging offered a useful tool for identifying new molecules eradicating cells during mitosis.
Insights
Phenanthrene derivative PJ-34 disrupts supernumerary centrosome clustering in cancer cells during mitosis, causing cell death. This targeted approach offers a new strategy for cancer treatment by inducing mitotic failure in multi-centrosomal cells.
Area of Science:
- Cell Biology
- Cancer Research
- Molecular Pharmacology
Background:
- Most solid human cancers exhibit supernumerary centrosomes, leading to genomic instability.
- Centrosome clustering is crucial for maintaining bipolar spindle formation during mitosis.
- Dysregulation of centrosome number and function is a hallmark of cancer cells.
Purpose of the Study:
- To investigate the effect of phenanthrene derivatives, specifically PJ-34, on centrosome behavior in multi-centrosomal cancer cells.
- To elucidate the mechanism by which PJ-34 induces cell death in cancer cells.
- To evaluate the potential of PJ-34 as a targeted anti-cancer therapeutic.
Main Methods:
- Live-cell confocal imaging of human breast cancer cells (MDA-MB-231) expressing fluorescent markers for centrosomes (γ-tubulin) and chromosomes (histone H2b).
- Treatment with phenanthrene derivatives, including PJ-34, to observe real-time cellular responses during mitosis.
- Analysis of chromosome arrangement and centrosome distribution following drug treatment.
Main Results:
- PJ-34 effectively prevented the clustering of supernumerary centrosomes in multi-centrosomal cancer cells.
- De-clustered centrosomes migrated to spindle poles, leading to aberrant chromosome segregation and mitotic failure.
- PJ-34 demonstrated selective cytotoxicity towards multi-centrosomal cancer cells, sparing normal cells with two centrosomes.
- The cytotoxic effect was observed even in PARP1-deficient cells, suggesting a mechanism independent of PARP1 inhibition.
Conclusions:
- PJ-34 induces cancer cell death by disrupting centrosome clustering and causing mitotic failure.
- This mechanism represents a novel therapeutic strategy targeting the high prevalence of supernumerary centrosomes in human cancers.
- Live confocal imaging is a valuable tool for identifying novel anti-mitotic agents with selective cancer cell-killing properties.
