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Tumor-specific RNAi targeting eIF4E suppresses tumor growth, induces apoptosis and enhances cisplatin cytotoxicity in human breast carcinoma cells.
Breast Cancer Res Treat. 2009 Feb; 113(3):443-56.BC

Abstract

BACKGROUND

Eukaryotic initiation factor eIF4E, an important regulator of translation, plays a crucial role in the malignant transformation, progression and chemoresistance of many human solid tumors. The overexpression of this gene has been found in a variety of human malignancies including breast carcinoma. In the present study, we attempted to explore the possibility of eIF4E as a therapeutic target for the treatment of human breast carcinoma using breast carcinoma cell line (MCF-7).

MATERIALS AND METHODS

The survivin promoter-driven eIF4E-shRNA vector was constructed on the basis of pSUPER.retro vector. Then, we established stably transfected MCF-7 and MCF210 (normal human mammary epithelial cell) cells expressing eIF4E-shRNAs or control-shRNAs. Firstly, the changes of eIF4E expression were detected by RT-PCR and Western blot assays. Next, the optimal shRNA vector (eIF4E-shRNA2) was selected to knock down eIF4E expression and investigate the effect of eIF4E-shRNA on eIF4E-regulated gene expression and cell proliferation both in vitro and in vivo. Followingly, the changes of cell cycle and apoptosis in the stably transfectants (MCF-7) were detected by flow cytometry and TUNEL methods, while we also explored possible apoptosis pathways. Finally, we investigated the effect of shRNA targeting eIF4E on the chemosensitivity of breast carcinoma cells to cisplatin in vitro and in vivo.

RESULTS

Two survivin promoter-driven eIF4E-shRNA vectors were successfully constructed. eIF4E-shRNA2 but not eIF4E-shRNA1 efficiently downregulated the levels of eIF4E expression in the stably transfected MCF-7-s2 cells but not in the stably transfected MCF210-s2 cells, while MCF-7-s2 showed obvious proliferation suppression but MCF210-s2 did not. The downregulation of eIF4E expression significantly reduced the levels of VEGF, FGF-2 and cyclinD1 expression, suppressed cell growth, induced cell cycle arrest in G(0)/G(1) phase and subsequent apoptosis by activating caspase 3 in MCF-7 cells. The results of FCM and TUNEL staining assays indicated that the classic apoptosis characters of the MCF-7 cells stably expressing eIF4E-shRNA2 manifested an apoptosis rate of 18.3%, significantly higher than those in the control groups (P < 0.05). Moreover, we found that downregulation of c-IAP1, c-IAP2 and c-Myc but not Bcl-2 family proteins were involved in the apoptosis induced by eIF4E-shRNA2. In tumorigenicity assay, xenograft tumors developed from MCF-7-s2 cells in mice showed a significant slowdown in the growth speed and formation rate compared with control groups. Furthermore, we also testified that eIF4E-shRNA could synergistically enhance the cytotoxicity effects of cisplatin to MCF-7 cells both in vitro and in vivo.

CONCLUSION

Survivin promoter-driven RNA interference system could efficiently and specifically downregulate eIF4E expression in human breast carcinoma cells but not in normal human mammary epithelial cell cells. Thus, eIF4E might play an important role in chemosensitivity to cisplatin, and survivin promoter-driven RNAi targeting eIF4E can be used as adjuvant therapy for human breast carcinomas with tumor specificity and high efficacy.

Authors+Show Affiliations

Department of Clinical Diagnosis, Tangdu Hospital, Fourth Military Medical University, Xi'an, Shaanxi Province, PR China.No affiliation info availableNo affiliation info availableNo affiliation info availableNo affiliation info availableNo affiliation info availableNo affiliation info available

Pub Type(s)

Journal Article

Language

eng

PubMed ID

18327707

Citation

Dong, Ke, et al. "Tumor-specific RNAi Targeting eIF4E Suppresses Tumor Growth, Induces Apoptosis and Enhances Cisplatin Cytotoxicity in Human Breast Carcinoma Cells." Breast Cancer Research and Treatment, vol. 113, no. 3, 2009, pp. 443-56.
Dong K, Wang R, Wang X, et al. Tumor-specific RNAi targeting eIF4E suppresses tumor growth, induces apoptosis and enhances cisplatin cytotoxicity in human breast carcinoma cells. Breast Cancer Res Treat. 2009;113(3):443-56.
Dong, K., Wang, R., Wang, X., Lin, F., Shen, J. J., Gao, P., & Zhang, H. Z. (2009). Tumor-specific RNAi targeting eIF4E suppresses tumor growth, induces apoptosis and enhances cisplatin cytotoxicity in human breast carcinoma cells. Breast Cancer Research and Treatment, 113(3), 443-56. https://doi.org/10.1007/s10549-008-9956-x
Dong K, et al. Tumor-specific RNAi Targeting eIF4E Suppresses Tumor Growth, Induces Apoptosis and Enhances Cisplatin Cytotoxicity in Human Breast Carcinoma Cells. Breast Cancer Res Treat. 2009;113(3):443-56. PubMed PMID: 18327707.
* Article titles in AMA citation format should be in sentence-case
TY - JOUR T1 - Tumor-specific RNAi targeting eIF4E suppresses tumor growth, induces apoptosis and enhances cisplatin cytotoxicity in human breast carcinoma cells. AU - Dong,Ke, AU - Wang,Rui, AU - Wang,Xi, AU - Lin,Fang, AU - Shen,Jian-Jun, AU - Gao,Ping, AU - Zhang,Hui-Zhong, Y1 - 2008/03/10/ PY - 2007/12/18/received PY - 2008/02/26/accepted PY - 2008/3/11/pubmed PY - 2009/2/12/medline PY - 2008/3/11/entrez SP - 443 EP - 56 JF - Breast cancer research and treatment JO - Breast Cancer Res Treat VL - 113 IS - 3 N2 - BACKGROUND: Eukaryotic initiation factor eIF4E, an important regulator of translation, plays a crucial role in the malignant transformation, progression and chemoresistance of many human solid tumors. The overexpression of this gene has been found in a variety of human malignancies including breast carcinoma. In the present study, we attempted to explore the possibility of eIF4E as a therapeutic target for the treatment of human breast carcinoma using breast carcinoma cell line (MCF-7). MATERIALS AND METHODS: The survivin promoter-driven eIF4E-shRNA vector was constructed on the basis of pSUPER.retro vector. Then, we established stably transfected MCF-7 and MCF210 (normal human mammary epithelial cell) cells expressing eIF4E-shRNAs or control-shRNAs. Firstly, the changes of eIF4E expression were detected by RT-PCR and Western blot assays. Next, the optimal shRNA vector (eIF4E-shRNA2) was selected to knock down eIF4E expression and investigate the effect of eIF4E-shRNA on eIF4E-regulated gene expression and cell proliferation both in vitro and in vivo. Followingly, the changes of cell cycle and apoptosis in the stably transfectants (MCF-7) were detected by flow cytometry and TUNEL methods, while we also explored possible apoptosis pathways. Finally, we investigated the effect of shRNA targeting eIF4E on the chemosensitivity of breast carcinoma cells to cisplatin in vitro and in vivo. RESULTS: Two survivin promoter-driven eIF4E-shRNA vectors were successfully constructed. eIF4E-shRNA2 but not eIF4E-shRNA1 efficiently downregulated the levels of eIF4E expression in the stably transfected MCF-7-s2 cells but not in the stably transfected MCF210-s2 cells, while MCF-7-s2 showed obvious proliferation suppression but MCF210-s2 did not. The downregulation of eIF4E expression significantly reduced the levels of VEGF, FGF-2 and cyclinD1 expression, suppressed cell growth, induced cell cycle arrest in G(0)/G(1) phase and subsequent apoptosis by activating caspase 3 in MCF-7 cells. The results of FCM and TUNEL staining assays indicated that the classic apoptosis characters of the MCF-7 cells stably expressing eIF4E-shRNA2 manifested an apoptosis rate of 18.3%, significantly higher than those in the control groups (P < 0.05). Moreover, we found that downregulation of c-IAP1, c-IAP2 and c-Myc but not Bcl-2 family proteins were involved in the apoptosis induced by eIF4E-shRNA2. In tumorigenicity assay, xenograft tumors developed from MCF-7-s2 cells in mice showed a significant slowdown in the growth speed and formation rate compared with control groups. Furthermore, we also testified that eIF4E-shRNA could synergistically enhance the cytotoxicity effects of cisplatin to MCF-7 cells both in vitro and in vivo. CONCLUSION: Survivin promoter-driven RNA interference system could efficiently and specifically downregulate eIF4E expression in human breast carcinoma cells but not in normal human mammary epithelial cell cells. Thus, eIF4E might play an important role in chemosensitivity to cisplatin, and survivin promoter-driven RNAi targeting eIF4E can be used as adjuvant therapy for human breast carcinomas with tumor specificity and high efficacy. SN - 1573-7217 UR - https://www.unboundmedicine.com/medline/citation/18327707/Tumor_specific_RNAi_targeting_eIF4E_suppresses_tumor_growth_induces_apoptosis_and_enhances_cisplatin_cytotoxicity_in_human_breast_carcinoma_cells_ L2 - https://doi.org/10.1007/s10549-008-9956-x DB - PRIME DP - Unbound Medicine ER -