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p53-independent G1 cell cycle arrest of human colon carcinoma cells HT-29 by sulforaphane is associated with induction of p21CIP1 and inhibition of expression of cyclin D1

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Abstract

Isothiocyanate sulforaphane (SFN) is a potent cancer chemopreventive agent. We investigated the mechanisms underlying the anti-proliferative effects of SFN in the human colon carcinoma cell line, HT-29. We demonstrate that SFN inhibits the growth of HT-29 cells in a dose- and time-dependent manner. Treatment of serum-stimulated HT-29 cells with SFN suppressed the re-initiation of cell cycle by inducing a G1 phase cell cycle arrest. At high doses (>25 μM), SFN dramatically induces the expression of p21CIP1 while significantly inhibits the expression of the G1 phase cell cycle regulatory genes such as cyclin D1, cyclin A, and c-myc. This regulation can be detected at both the mRNA and protein levels as early as 4 h post-treatment of SFN at 50 μM. Additionally, SFN activates MAPKs pathways, including ERK, JNK and p38. Exposure of HT-29 cells with both SFN and an antioxidant, either NAC or GSH, completely blocked the SFN-mediated activation of these MAPK signaling cascades, regulation of cyclin D1and p21CIP1 gene expression, and G1phase cell cycle arrest. This finding suggests that SFN-induced oxidative stress plays a role in these observed effects. Furthermore, the activation of the ERK and p38 pathways by SFN is involved in the upregulation of p21CIP1 and cyclin D1, whereas the activation of the JNK pathway plays a contradictory role and may be partially involved in the downregulation of cyclin D1. Because cyclin D1 and p21CIP1 play opposing roles in G1 phase cell cycle progression regulation, blocking the activation of each MAPK pathway with specific MAPK inhibitors, is unable to rescue the SFN-induced G1 phase cell cycle arrest in HT-29 cells.

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Abbreviations

SFN:

Sulforaphane

MAPK:

Mitogen-activated protein kinase

NF-κB:

Nuclear factor-kappa B

GSH:

Glutathione

NAC:

N-acetyl-L-cysteine

MTS:

3-(4,5-dimethylthiazol-2-yl)-5-(3-carboxyphenyl)-2-(4-sulfophenyl)-2H-tetrazolium, inner salt

ERK:

Extracellular signal-regulated protein kinase JNK, c-Jun NH2-terminal kinase

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Acknowledgements

This work was supported in part by NIH Grant R01 CA-073674-07. The authors thank Ms. Patra Pharm in the Flow Cytometry Core Facility for her excellent assistance.

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Authors and Affiliations

  1. Department of Pharmaceutics, Ernest Mario School of Pharmacy, Rutgers, The State University of New Jersey, 160 Frelinghuysen Road, Piscataway, NJ, 08854, USA

    Guoxiang Shen, Changjiang Xu, Chi Chen, Vidya Hebbar & Ah-Ng Tony Kong

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  1. Guoxiang Shen
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  2. Changjiang Xu
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  3. Chi Chen
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Correspondence to Ah-Ng Tony Kong.

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Shen, G., Xu, C., Chen, C. et al. p53-independent G1 cell cycle arrest of human colon carcinoma cells HT-29 by sulforaphane is associated with induction of p21CIP1 and inhibition of expression of cyclin D1. Cancer Chemother Pharmacol 57, 317–327 (2006). https://doi.org/10.1007/s00280-005-0050-3

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