Animal Study Of Anti-EGGR Therapy To Improve The Effect Of Radiotherapy For Recurrent Esophageal Carcinoma

BackgroundEsophageal cancer is of high incidence in our country, which annual morbidity and mortality rates ranked third and fourth among all cancers. As most patients are diagnosed as locally adbanced disease and had no opportunity to get cutative surgery, definitive chemo-radiotherapy or radiotherapy was the main treatment modality. However, local tumor recurrence rate might be high as 30-40% after radiotherapy, which is one of the main reasons for treatment failure. At this moment, the re-irradiation combined with systemic chemotherapy is often the only choice, but the re-irradiation is very difficuly because that:(1) in previous radical radiotherapy local normal tissue had received high radiation doses, so a radical irradiation dose could not be given to in the recurred tumor, (2) in the previous irradiation, the influence on microenvironment of the tumor bed (mainly tumor microvascules damage) may lead to increasing of hypoxia fraction in recurred tumors which leads to radiotherapy resistance.Tumor xenograft model for recurrent tumor after irradiation were developed to simulate the clinical situation. The widely accepted approach is giving a certain dose of radiation to the transplantation site before transplanting. It was found in variety of animal experiments using different recurrent tumor models that the hypoxia level in the recurrent tumor was significantly higher than the normally transplanted. In addtion, epidermal growth factor receptor (EGFR) overexpression was found in hypoxia area. Therefore, EGFR targeted therapy combined with radiotherapy is expected to become an effective means to control the tumor recurrence. Nimotuzumab, an important EGFR-targeted antibody, is currently used clinically combined with radiotherapy and approved for treating esophageal.ObjectiveTo establish animal models of recurrent esophageal cancer. To investigate the relationship hypoxic state and EGFR expression in the different models of esophageal cancer recurrence by immunohistochemistry, Western Blot and other technologies. To observe the effects of treatments for recurrent esophageal cancer using nimotuzumab plus radiotherapy.MethodsIn Western Blot test; EGFR, p-EGFR and HIF-? expression was test in EC 109 esophageal cancer cells under normal and hypoxic condition with/without nimotuzumab treatments. Cell cycle and apoptosis were determined by flow cytometry analysis. Three kinds of tumor models were established:(1) recurrent tumor model 1:, the right hind leg was irradiation with lOGy X-ray 24 hours before tumor implantion (2) recurrent tumor model 1:tumors regrowing after 20Gy irradiation. (3) common xenograft model. Tumor samples were prepared and the expression of HIF-1? and EGFR were evaluated by immunohistochemistry (IHC) staining. When the tumors grew up to the inclusion criteria, the mice were randomly to recieve:non-treatment controlled, the irradiation alone, nimotuzumab alone group and irradiation+nimotuzumab. Paired T-test was uesd to compare the tumor volume differences of the two groups.P<0.05 was considered statistically significant.ResultsWestern blot resulted that HIF-1? and EGFR expression increased in hypoxic EC 109 cells. Nimotuzumab treatments lead to down-regulation of pEGFR. In flowcytometry analyisis, nimotuzumab combined with radiotherapy arrest the cells in the cell cycle G2. Meanwhile, higher apoptosis rate was found in nimotuzumab plus radiotherapy group. IHC staining showed increased expression of EGFR on the cell membrane in the recurrent tumor model. In different tumor models, tumor growth delay assay demonstrated that nimotuzumab combined with radiotherapy significantly delayed the tumor growth than radiotherapy alone or nimotuzumab alone, P<0.05.ConclusionsIn EC109 cells, EGFR expression were correlated with HIF-1 a expression in both cell culture and tumor tissue. Nimotuzumab could down-regulared the pEGFR expression. In recurrence tumor models, nimotuzumab could significantly delay the tumor growth as combined with irradiation. Our study provided theoretical and experimental information for new treatment for the recurrent esophageal cancer.