| Chemotherapy is an important therapeutic strategy for cancer treatment and remains the mainstay for the management of human malignancies; however, chemotherapy fails to eliminate all tumor cells because of intrinsic or acquired drug resistance, which is the most common cause of tumor recurrence. Human cancers are generally initially responsive to standard chemotherapies; however, response is almost inevitably followed by the development of drug-resistant phenotype, which leads to tumor recurrence and metastasis. The mechanisms responsible for drug resistance are complex and still poorly understood. It may be due to either the specific nature and genetic background of the cancer cell itself or the genetic changes that follow toxic chemotherapy. Drug resistance to therapy is classified by two categories:intrinsic and acquired. Intrinsic resistance would make the therapy ineffective because prior to receiving the therapy, the cancer cells have already resistant to anti-cancer drugs due to multiple mechanisms. Acquired resistance develops during the treatment, although the tumor cells were not initially resistant to anti-cancer drugs. The most common reasons for the acquisition of resistance to anti-cancer drugs are due to expression of one or more energy-dependent transporters that detect and eject anti-cancer drugs from cells, insensitivity to drug-induced apoptosis and the induction of drug-detoxification mechanisms.The Notch signaling pathway plays an important role in the proliferation and differentiation of cells. It is an evolutionarily conserved pathway that regulates critical cell fate decisions. Emerging evidence suggest that the Notch signaling network is frequently deregulated in human malignancies with up-regulated expression of Notch receptors and their ligands were found in breast, lung, colon, lymphomas and pancreatic cancer. Recently, Notch pathway has been reported to be involved in drug resistance. More importantly, the studies have demonstrated that Notch regulates the formation of cancer stem cells (CSCs) and contributes to the acquisition of the epithelial-mesenchymal transition (EMT) phenotype, which are critically associated with drug resistance.Experimental evidence also revealed that Notch was involved in anti-cancer drug resistance, indicating that targeting Notch could be a novel therapeutic approach for the treatment of cancer by overcoming drug resistance of cancer cells, which may lead to the elimination of CSCs or EMT type cells which are typically drug-resistant, and are believed to be the "root cause" of tumor recurrence. Notch receptor activation is irreversible as it involves proteolysis-mediated release of the Notch intracellular domain (NICD) which can be interrupted by y-secretase inhibitor (GSI), translocation to the nucleus and transactivation of gene targets which in turn affect numerous pathways involving cell-fate determination. It has been reported that blocking the Notch signaling pathway by GSIs could enhance the drug sensitivity of some cancer cells to chemotherapy. Up to date, there is no published evidence that the Notch signaling involves in drug resistance of nasopharyngeal carcinoma. Nasopharyngeal carcinoma was highly incident malignancy in China In the treatment of nasopharyngeal carcinoma, there have been significant research advances including the introduction of chemoradiation and adjuvant chemotherapy, though the overall survival rate remains only in50%-60%and many nasopharyngeal carcinomas eventually develop resistance to radiotherapy and chemotherapy.In this work, we discussed the correlation and mechanisms of Notch signaling pathway in drug resistance of NPC, which will supply new specific molecular-targeted drug for treatment of advanced stage especially recurrent nasopharyngeal carcinoma patients.Objective:Detect the Notch receptor expression of nasopharyngeal carcinomas tissue and the apoptosis of NPC cells after suppression of the Notch signaling pathway, and further discuss the mechanism of Notch signaling in drug resistant of nasopharyngeal carcinoma, which will supply new specific molecular drug for nasopharynggeal carcinoma treatment.Methods:1. Detected the Notch receptor expression of the inflammatory mucous membrane of nasopharynx, primary and recurrent nasopharyngeal carcinomas tissue with QDs-immunofluorescence; Detected the Notch receptor expression of CNE-1and CNE-2cells with Western-blot.2. The CNE-1and CNE-2cells was treated with different concentration of Cisplatin and CNE-2treated with different concentration of DAPT combined with10uM Cisplatin. After48h, the apoptosis rate of the above cells was detected by flow cytometry; Created and cultured the Cisplatin resistant CNE-2cells named CNE-2/CDDP which was treated with different concentration of DAPT combined with10?M Cisplatin. After48h, the apoptosis rate of the CNE-2/CDDPcells was detected by flow cytometry.3. The CNE-2cells were only treated with different concentration of DAPT, and then the proliferatin and cell cycle were detected by CCK-8and flow cytometry. Meanwhile, the proteins regulating cell cycle and cell differentiation were detected by wetern-blot; The CNE-2/CDDP cells were only treated with different concentration of DAPT, and then the proteins associated with EMT and regulating the drug resistant were also detected by wetern-blot.Results:1. There was almost no expression or weakly positive expression of Notch receptors in inflammatory mucous membrane of nasopharynx. The Notch-1and Notch-4receptors were positive expression in the primary nasopharyngeal carcinomas tissue, and Notch-2,3were weakly positive expression. In the recurrent nasopharyngeal carcinomas tissue, the Notch-1and Notch-4recepotors were stronger positive expression than the primary nasopharyngeal carcinomas tissue, while there was no difference of Notch-2,3expression. Notch-1and Notch-4recepotors were highly expression in CNE-2cells but not in CNE-1. There was no difference of Notch-2,3expresssion between CNE-1and CNE-2, but it was lower than the expression of Notch-1,4. The results in cells were accordant with tissues. All the differences show statistical significance (P<0.05).2. There was obviously different sensitivity to Cisplatin between CNE-1and CNE-2cells. The apoptosis rate of CNE-1was obviously higher than CNE-2when treated with same concentration of Cisplatin. CNE-2cells were treated with different concentration of DAPT combined with10?M Cisplatin, and then the apoptosis rate was increased in a dose-dependent relationship with DAPT. When the CNE-2cells were only treated with different concentration of DAPT, there was no obvious apoptosis, but the cell proliferation was inhibited and cycle was blocked in G1phase. Meanwhile, the expression of CyclinE/CDK2, c-myc and p-ERK were obviously declined after suppression of Notch by DAPT. CNE-2/CDDP cells were treated with different concentration of DAPT combined with10?M Cisplatin and then the apoptosis rate was increased in a dose-dependent relationship with DAPT. When the CNE-2/CDDP cells were only treated with different concentration of DAPT, there was no obvious apoptosis, the proteins expression involved in EMT and drug resistant were obviously declined after suppression of Notch by DAPT. All the differences show statistical significance (P <0.05).Conclusion:The Notch signaling was involved in the development of nasopharyngeal carcinoma; especially the Notch-1and Notch-4receptors were confirmed that it participated in the cisplatin resistant of nasopharyngeal carcinoma. The cisplatin sensitivity of CNE-2/CDDP which was resistant to the cisplatin toxic effect was improved when suppressed the Notch signaling by DAPT. It was supposed tha the Notch signaling regulated the acquired drug resistant of NPC by regulating the EMT and then regulating the expression of P-gp and ERCC1. The Notch signaling also regulated intrinsic durg resistant of NPC through regulating the cell cycle and cell differentiation... |
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