Radiosensitivity Enhancement By Combined Treatment Of Nimotuzumab And Celecoxib On Nasopharyngeal Carcinoma

Objective: In this study,the treatment compliance,toxicities,and preliminary therapeutic efficacy were observed to evaluate the feasibility of the combinational treatment modality of concurrent chemoradiation plus nimotuzumab and celecoxib for locoregionally advanced nasopharyngeal carcinoma.The cytotoxicity and radiosensitivity of nimotuzumab and celecoxib on nasopharyngeal carcinoma cells and the potential mechanisms were investigated in vitro.Detection of tumor EGFR levels by immunohistochemistry was conducted to demonstrate the association between nuclear EGFR and the clinical/pathologic data of the nasopharyngeal carcinoma patients.Methods and Materials: In the phase II clinical trials,patients with locoregionally advanced nasopharyngeal carcinoma were treated with the regimen of induction chemotherapy,followed by concurrent chemoradiation plus nimotuzumab or the combination of celecoxib,nimotuzumab,and radiation treatments.The treatment compliance and toxicities were assessed from the start of protocol therapy.Survival curve was estimated with the Kaplan–Meier method.Human nasopharyngeal carcinoma cells,CNE1 and CNE2,were treated with nimotuzumab,celecoxib,or their combination.MTT and clonogenic survival assays were administrated to evaluate the radiosensitivity of nimotuzumab or celecoxib or the combination of both drugs on nasopharyngeal carcinoma cells.Immunoblotting were performed to identify the role of celecoxib and/or nimotuzumab with or without radiation on the expression of cytoplasm and nuclear EGFR.Futhermore,immunohistochemistry was conducted to determine the expression of EGFR.Kaplan–Meier method and Cox regression analysis were employed to determine the association between nuclear EGFR expression and the clinical and pathologic variables such as gender,age,clinical T category,Clinical N category,histologic grade and progression-free survival.Results:(1)Twenty-three patients were enrolled in the clinical trial of induction chemotherapy,followed by concurrent chemoradiation plus nimotuzumab.Results showed that all patients received a full course of induction chemotherapy and radiotherapy,19(82.6%)patients completed the scheduled concurrent chemotherapy,and 22(95.7%)patients received ?6 weeks of nimotuzumab.During the period of concurrent chemoradiation and nimotuzumab,grade 3-4 toxicities occurred in 14(60.9%)patients: 8(34.8%)had grade 3-4 oral mucositis,6(26.1%)had grade 3 neutropenia.With a median follow-up of 24.1 months,the 2-year progression-free survival and overall survival were 83.5% and 95.0%,respectively.(2)Thirty-one patients were enrolled in the clinical trial of the combinational treatment of celecoxib,nimotuzumab and radiation.Results showed that all patients received a full course of induction chemotherapy,radiotherapy and celecoxib.All patients received ?6 weeks of nimotuzumab.During the period of radiotherapy,grade 3-4 toxicities occurred in 12(38.7%)patients: 8(25.8%)had grade 3-4 oral mucositis,4(12.9%)had grade 3 neutropenia.With a median follow-up of 30.5 months,the 2-year progression-free survival and overall survival were 89.2% and 96.8%,respectively.(3)MTT assays showed that celecoxib(0~200?mol/L)showed dramatically dose and time-dependent toxic effects in two cell lines.Nimotuzumab was not toxic to either cell line,even when they were treated with 200?g/ml for 72 h.(4)Colony formation assays revealed that combined nimotuzumab and celecoxib increases radiosensitivity compared to either drug alone in CNE-2 cells.Combined nimotuzumab and celecoxib cooperatively blocks radiation-induced activation of cytoplasm and nuclear EGFR signaling pathways in CNE2 cells.However,celecoxib alone or in combination with nimotuzumab failed to inhibit cytoplasm and nuclear EGFR signaling pathways,and demonstrated no radiosensitivity enhancement in CNE1 cells.(5)High nuclear EGFR expression occurred in 45.2% of patients.There was no association between nuclear EGFR expression and any of the clinical and pathologic variables.Conclusion:(1)The combinational treatment modality of radiotherapy plus celecoxib and nimotuzumab was feasible,with good compliance,acceptable toxicity,and encouraging preliminary clinical outcome.(2)Radiation activates cytoplasm and nuclear EGFR signaling pathways in nasopharyngeal carcinoma cells CNE1 and CNE2.(3)Nimotuzumab,celecoxib,or their combination did not increase the radiosensitivity of CNE1 cells.(4)Radiosensitivity enhancement was observed when nimotuzumab and celecoxib were administrated concurrently,which might be relevant to the inhibition of cytoplasm and nuclear EGFR signaling pathways.