Overexpression of E2F1 in human gastric carcinoma is involved in anti-cancer drug resistance

BackgroundRoutine chemotherapy often cannot achieve good therapeutic effects because of multidrug resistance (MDR). MDR is frequently caused by the elevated expression of the MDR1 gene encoding P-glycoprotein (P-gp). E2F1 is a frequently overexpressed protein in human tumor cells that increases the activity of the MDR1 promoter, resulting in higher P-gp levels. The upregulation of P-gp might contribute to the survival of tumor cells during chemotherapy. E2F1 confers anticancer drug resistance; however, we speculate whether E2F1 affects MDR through other pathways. This study investigated the possible involvement of E2F1 in anticancer drug resistance of gastric carcinoma in vitro and in vivo.MethodsA cisplatin-resistant SGC7901/DDP gastric cancer cell line with stable overexpression of E2F1 was established. Protein expression levels of E2F1, MDR1, MRP, TAp73, GAX, ZEB1, and ZEB2 were detected by western blotting. The influence of overexpression of E2F1 on anticancer drug resistance was assessed by measuring IC50 of SGC7901/DDP cells to cisplatin, doxorubicin, and 5-fluorouracil, as well as the rate of doxorubicin efflux, apoptosis, and cell cycle progression detected by flow cytometry. We determined the in vivo effects of E2F1-overexpression on tumor size in nude mice, and apoptotic cells in tumor tissues were detected by deoxynucleotidyl transferase-mediated dUTP-biotin nick end labeling and hematoxylin and eosin staining.ResultsThe SGC7901/DDP gastric cancer cell line stably overexpressing E2F1 exhibited significantly inhibited sensitivity to cisplatin, doxorubicin, and 5-fluorouracil. Flow cytometry confirmed that the percentage of apoptotic cells decreased after E2F1 upregulation, and that upregulation of E2F1 potentiated S phase arrest of the cell cycle. Furthermore, upregulation of E2F1 significantly decreased intracellular accumulation of doxorubicin. Western blot revealed that E2F1 upregulation suppressed expression of GAX, and increased the expression of MDR1, MRP, ZEB1, TAp73, and ZEB2.ConclusionsOverexpression of E2F1 promotes the development of MDR in gastric carcinoma, suggesting that E2F1 may represent an efficacious target for gastric cancer therapy.

[1]  W. Sadee,et al.  Growth factor signaling and resistance to cancer chemotherapy. , 2004, Current topics in medicinal chemistry.

[2]  B. Kowtharapu,et al.  E2F1 confers anticancer drug resistance by targeting ABC transporter family members and Bcl-2 via the p73/DNp73-miR-205 circuitry , 2012, Cell cycle.

[3]  B. Pützer,et al.  Alterations of ΔTA‐p 73 splice transcripts during melanoma development and progression , 2004, International journal of cancer.

[4]  G. Blandino,et al.  The Transcriptional Repressor ZEB Regulates p73 Expression at the Crossroad between Proliferation and Differentiation , 2001, Molecular and Cellular Biology.

[5]  C. Speyer,et al.  Regulation of the Expression and Activity of the Antiangiogenic Homeobox Gene GAX/MEOX2 by ZEB2 and MicroRNA-221 , 2010, Molecular and Cellular Biology.

[6]  Zhiming Wang,et al.  miR-301a Is a Candidate Oncogene that Targets the Homeobox Gene Gax in Human Hepatocellular Carcinoma , 2012, Digestive Diseases and Sciences.

[7]  Leila Mohammadi,et al.  BMC Cancer , 2001 .

[8]  P. Jiang,et al.  Suppression of p53 activity by Siva1 , 2009, Cell Death and Differentiation.

[9]  Wen He,et al.  Down Regulation of CIAPIN1 Reverses Multidrug Resistance in Human Breast Cancer Cells by Inhibiting MDR1 , 2012, Molecules.

[10]  T. Aikou,et al.  Expression of the multidrug‐resistance‐associated protein (MRP) gene in human colorectal, gastric and non‐small‐cell lung carcinomas , 1996, International journal of cancer.

[11]  T. Mak,et al.  TAp73 enhances the pentose phosphate pathway and supports cell proliferation , 2013, Nature Cell Biology.

[12]  M. Saceda,et al.  Tumour cells resistance in cancer therapy , 2007, Clinical & translational oncology : official publication of the Federation of Spanish Oncology Societies and of the National Cancer Institute of Mexico.

[13]  B. Pützer,et al.  E2F1 in melanoma progression and metastasis. , 2010, Journal of the National Cancer Institute.

[14]  Jean-Pierre Gillet,et al.  Mechanisms of multidrug resistance in cancer. , 2010, Methods in molecular biology.

[15]  Young-Duck Kim,et al.  E2F1 Expression is Related with the Poor Survival of Lymph Node-positive Breast Cancer Patients Treated with Fluorouracil, Doxorubicin and Cyclophosphamide , 2003, Breast Cancer Research and Treatment.

[16]  S. Cole.  Targeting multidrug resistance protein 1 (MRP1, ABCC1): past, present, and future. , 2014, Annual review of pharmacology and toxicology.

[17]  Ying Han,et al.  Octarepeat peptides of prion are essential for multidrug resistance in gastric cancer cells , 2012, Journal of digestive diseases.

[18]  D. Gorski,et al.  Inhibition of endothelial cell activation by the homeobox gene Gax. , 2003, The Journal of surgical research.

[19]  I. Tannock,et al.  Drug penetration in solid tumours , 2006, Nature Reviews Cancer.

[20]  Zhong-Zhen Zhao,et al.  Apoptosis Sensitization by Euphorbia Factor L1 in ABCB1-Mediated Multidrug Resistant K562/ADR Cells , 2013, Molecules.

[21]  L. Nelles,et al.  Interaction between Smad-interacting Protein-1 and the Corepressor C-terminal Binding Protein Is Dispensable for Transcriptional Repression of E-cadherin* , 2003, Journal of Biological Chemistry.

[22]  H. Rotheneder,et al.  Regulation of the MDR1 promoter by E2F1 and EAPP , 2013, FEBS letters.

[23]  P. Duerksen-Hughes,et al.  Methyl methanesulfonate induces apoptosis in p53-deficient H1299 and Hep3B cells through a caspase 2- and mitochondria-associated pathway. , 2012, Environmental toxicology and pharmacology.

[24]  Y. Iwamoto,et al.  Involvement of P‐glycoprotein and MRP1 in resistance to cyclic tetrapeptide subfamily of histone deacetylase inhibitors in the drug‐resistant osteosarcoma and Ewing's sarcoma cells , 2006, International journal of cancer.

[25]  R. Halaban,et al.  Deregulated E2f Transcriptional Activity in Autonomously Growing Melanoma Cells , 2000, The Journal of experimental medicine.

[26]  Jingqing Yang,et al.  Reversal of Multidrug Resistance in Gastric Cancer Cells by E2F‐1 Downregulation In Vitro and In Vivo , 2014, Journal of cellular biochemistry.