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Shogaols at proapoptotic concentrations induce G2/M arrest and aberrant mitotic cell death associated with tubulin aggregation

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Abstract

Shogaols have been previously reported to induce cancer cell death via multiple mechanisms, among which one analog 6-shogaol has been reported to cause microtubule damage through specific reaction with sulfhydryl groups in tubulin. In this study, a series of shogaols with different side chain lengths (4-, 6-, 8- and 10-shogaol) was synthesized and evaluated for antiproliferative activity in HCT 116 colon carcinoma and SH-SY5Y neuroblastoma cells. 4- and 6-shogaol were identified as lead compounds possessing the strongest antiproliferative activity. In the soft agar assay, the lead shogaols displayed dose-dependent inhibition on cancer cell colony formation under anchorage-independent conditions. Using HCT 116 as the selected cancer cell line, the molecular events linking shogaols-induced G2/M cell cycle arrest to apoptosis characterized by caspase 3 and PARP cleavage were investigated. At sublethal concentrations, the halt at G2/M phase was alleviated along time and cells survived. Conversely, proapoptotic concentrations of 4- and 6-shogaol induced irreversible G2/M arrest that was at least in part associated with down-regulation of cell cycle checkpoint proteins cdk1, cyclin B and cdc25C, as well as spindle assembly checkpoint proteins mad2, cdc20 and survivin. A dose- and time-dependent accumulation of insoluble tubulin in the insoluble fractions of cell lysates provided evidence that G2 checkpoint failure led to disruption of microtubule turnover. In summary, our results conclude that shogaols cause apoptosis by inducing aberrant mitosis at least through the attenuation of cell cycle and spindle assembly checkpoint proteins.

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Abbreviations

CAK:

Cdk activating kinase

Cdk:

Cyclin-dependent kinase

mad2:

Mitotic-arrest deficient homologue-2

p-cdk1:

Phospho cyclin-dependent kinase 1

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Acknowledgments

This work was supported by National University of Singapore Academic Research Fund Tier 1 R-148-000-116-112 and National Medical Research Council Grant No. NMRC/NIG/0050/2009 (E–H. Chew).

Conflict of interest

The authors declare that they have no conflicts of interest.

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Authors and Affiliations

  1. Department of Pharmacy, Faculty of Science, National University of Singapore, 18 Science Drive 4, Singapore, 117543, Republic of Singapore

    Fei-Fei Gan, Amrita A. Nagle, Xiaohui Ang, Olivia Huixian Ho, Sock-Hoon Tan, Hong Yang, Wai-Keung Chui & Eng-Hui Chew

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  1. Fei-Fei Gan
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  3. Xiaohui Ang
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Correspondence to Eng-Hui Chew.

Additional information

Xiaohui Ang, Olivia Huixian Ho and Sock-Hoon Tan contributed equally to this work.

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Gan, FF., Nagle, A.A., Ang, X. et al. Shogaols at proapoptotic concentrations induce G2/M arrest and aberrant mitotic cell death associated with tubulin aggregation. Apoptosis 16, 856–867 (2011). https://doi.org/10.1007/s10495-011-0611-3

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