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NSC126188, a piperazine alkyl derivative, induces apoptosis via upregulation of RhoB in HeLa cells

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Summary

We describe here a piperazine alkyl derivative, NSC126188, which induced apoptosis of HeLa cells by upregulating RhoB expression. NSC126188 caused multi-septation of fission yeast and hypersensitized a ∆rho3 mutant, which implicates the involvement of functional human homolog RhoB. The treatment of cells with NSC126188 induced apoptosis and a dramatic increase in RhoB expression. In addition, RhoB knockdown using siRNA rescued cells from apoptosis, indicating a crucial role of RhoB in NSC126188-induced apoptosis. In a reporter assay using luciferase and EGFP under control of the RhoB promoter, NSC126188 increased both luciferase activity and the expression of EGFP, implicating transcriptional activation of RhoB by NSC126188. Furthermore, NSC126188 demonstrated in vivo anti-tumor activity, inhibiting tumor growth by 66.8% in a nude mouse xenograft using PC-3 human prostate cancer cells. These results suggest that NSC126188 is a potential lead compound and that upregulation of RhoB is associated with NSC126188-induced apoptosis.

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Acknowledgments

We are grateful to Young-Joo Jang and Hyunji Lee for their technical assistances. This work was supported in part by grants from the 21st Century Frontier for Functional Analysis of the Human Genome and the KRIBB Initiative from the Korea Research Council of Fundamental Science and Technology.

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Correspondence to Hwan Mook Kim or Misun Won.

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Bo-Kyung Kim, Dong-Myung Kim and Kyung-Sook Chung equally contributed to this work

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Table S1

Determination of GI50 values of NSC126188 in various human cancer cell lines. Cancer cells were treated with NSC126188 at 40 µM followed by 2-fold serial dilutions of the compound for 48 h. Growth inhibition was examined compared to that of non-treated cells. The GI50, the concentration of NSC126188 that caused 50% cell death, was determined. (PPT 118 kb)

Figure S1

The dose-dependent growth inhibition of HeLa cells by NSC126188. HeLa cells were treated with 40 µM followed by 2-fold serial dilutions of NSC126188 for 48 h. Figure. S2. a. Microscopic observation of cells treated with 10 µM NSC126188. B. Annexin V staining of HeLa cells treated with 10 µM NSC126188 for 12 h. Cells cultivated in the presence of NSC126188 showed intense Annexin V-FITC staining. Figure S3. Microscopic observation of cells treated with siRNA. The HeLa cells were treated with siRhoB (10 nM) for 36 h in the absence and presence of NSC126188 (3 µM). The scrambled siRNA (siScrambled) was used as a negative control. (PPT 2752 kb)

Figure S4

Diagram of the RhoB promoter. The RhoB promoter (594 bp) was cloned into the pGL2-basic plasmid vector for reporter assays. The sites for CCAAT, NF-Y, SP1, and AP2 were included in the construct. Reporter genes, either luciferase or EGFP, were cloned under control of the RhoB promoter and were used to quantitate promoter activity. Fgure S5. In vivo xenograft assay of NSC126188 using PC-3 cells. A. Changes in body weights. B. Tumor volumes during treatment of cancer (PPT 123 kb)

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Kim, BK., Kim, DM., Chung, KS. et al. NSC126188, a piperazine alkyl derivative, induces apoptosis via upregulation of RhoB in HeLa cells. Invest New Drugs 29, 853–860 (2011). https://doi.org/10.1007/s10637-010-9433-3

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  • DOI: https://doi.org/10.1007/s10637-010-9433-3

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