The Effect Of Nimotuzumab On The Radiochemotherapy Sensitivity Of Esophageal Squamous Cancer Cells

Background and objective:Esophageal cancer which derived from the mucous membrane of esophagus includes squamous cell carcinoma and adenocarcinoma.The main histological typle of esophageal cancer is squamous cell carcimoma in our country. Esophageal cancer is the eighth most common cancer worldwide. Most patients are in III/IV stage when diagnosed, especially in developing countries, which also results in a poor prognosis. Chemoradiotherapy are still reserved as a significant approach to patients suffering from advanced carcinoma, however, the effectiveness was limited, only20to40%of patients were observed to be responsive to it. Thus, how to improve the therapeutic benefit remains to be a research hotspot. Nimotuzumab is a humanized monoclonal antibody which can bind to domain III of the extracellular region of the EGFR with a moderate affinity and inhibits its downstream signal pathway. Compared to other annti-EGFR monoclonal antibodies, it carries a slight side effect. Nimotuzumab revealed a remarkable anti-proliferative, pro-apoptotic and anti-angiogenic effect in vitro as well as in vivo studies and enhanced the radiotherapy sensitivity of malignant tumors, such as glioma and nasopharyngeal carcinoma. Due to its modulation effect on radiochemotherapy sensitivity, it has been combined with chemotherapeutic drugs and radiotherapy. Moreover, it has been approved in the treatment of different tumors in different countries. In our contry, it has been recommended for the treatment of head and neck cancer patients. In this study, we observed the effect of nimotuzumab on the radiochemotherapy sensitivity of esophageal squamous cancer cells and explored the mechanism underlying the combinational scheme.Methods:1.The mean fluorescence intensity of EGFR expression on TE-1and Eca109esophageal cancer cells were analyzed by Flow cytomy, while, the degree of EGFR expression was detected by immunohistochemistry.2. MTT assay was applied to evaluate the effect of the combination of nimotuzumab and paclitaxel(PTX)/cis-platinum(DDP)/5-florouracil(5-Fu)on the proliferation of Eca109and TE-1esophageal cancer cells.3. The effect of h-R3on radiotherapy sensitivity of Eca109and TE-1esophageal cancer cells was observed by Cell cloning experiments.4. Flow cytometry was used to analyse the changes of apoptosis rate of Eca109and TE-1cells induced by the the combination scheme.5. The changes of cell cycle distribution underlying the Eca109and TE-1cells were also detected by flow cytometry.6. Transmission electron microscope reserved as a significant method to observe the effect of nimotuzumab on the autophagy of Eca109and TE-1cells.7. The mRNA expression level of Becline-1expressed on Eca109and TE-1cells were analyzed by RT-PCR and the protein expression level of becline-1was measured by western blotting.Results:1. Eca109cells highly expressed EGFR, but TE-1cells carried a low expression level of EGFR.2. Nimotuzumab could enhanced the response of PTX/DDP/5-Fu to Eca109cells, but no enhancement was observed on TE-1cells.3. Nimotuzumab could enhanced radiotherapy sensitivity of Eca109cells, however, it have no obvious effect on that of TE-1cells.4. The combination of nimotuzumab and PTX/DDP induced a higher apoptosis rate underlying the Eca109cells, but had no obvious effect on apoptosis rate of TE-1cells.5. The retardant of G0/G1phase of the cell cycle and the decrease of S phase of the cell cycle induced by nimotuzumab were observed in Eca109cell line but not in TE-1cell line.6. Nimotuzumab induced an enhancement of autophagy existing in Eca109cells, but had no apparent effect on that of TE-1cells.7. Nimotuzumab could up-regulate Becline-1expression on Eca109cells but exerted no effect on expression level of Becline-1in TE-1cells Conclusion:1. Nimotuzumab could enhance the chemosensitivity and radiosensitivity of Eca109esophageal cancer cells which possess of a high level of EGFR expression, but the TE-1cells with a low level of EGFR expression revealed no obvious response to nimotuzumab.2. The superiority of the combinational project was achieved by a series of possible mechanisms, such as the the retardant of G0/G1phase of the cell cycle and the enhancement of apoptosis and autophagy.