Optimization Of EGFRmAb Modified Gold Nanorods Mediated Photothermal Therapy For Head And Neck Squamous Cell Carcinoma In Mice

Objectives:Our research and numerous studies at home and abroad have demonstrated from the protein and gene level that EGFR is highly expressed in head and neck squamous cell carcinoma[1-4].Based on this,our preliminary work indicates that EGFR monoclonal antibody and gold nanorod conjugate can enter the cancer cells in vitro through endocytosis of EGFR.This project will further study the parameters related to the induction of apoptosis of head and neck squamous carcinoma cells by nano-gold rod photothermal effect,and To establish a nude mouse xenograft model of laryngeal squamous cell carcinoma and nasopharyngeal squamous cell carcinoma,and to verify that the conjugate of EGFR monoclonal antibody and nanogold rod is selectively enriched in the xenografted tumor of the head and neck squamous cell carcinoma and enters the cancer cell.By controlling the NIR laser irradiation parameters(time and power),the authors explored the conditions and procedures for optimizing the EGFR monoclonal antibody and nano-gold rod photothermal therapy for the treatment of head and neck squamous cell carcinoma xenografts in nude mice to obtain the best effect.At the same time,the systematic safety evaluation of this combination therapy method provides a basis and basis for exploring a new method for the targeted treatment of human head and neck squamous cell carcinoma with EGFR monoclonal antibody combined with nanogold photothermal effect.Method:1.MTT assay was used to detect the effect of nano-gold rods on the proliferation of normal cells and head and neck squamous carcinoma cells at different concentrations.Electron transmission microscopy was used to observe the distribution of gold nanorods into the head and neck squamous cell carcinoma cells,and the microscopic findings of apoptosis and necrosis induced by photothermal therapy.3.Establish a nude mouse xenograft model of laryngeal squamous cell carcinoma and nasopharyngeal squamous cell carcinoma4.To study the conjugate of EGFR monoclonal antibody and nano-gold rod in the nude mice xenografts of head and neck squamous cell carcinoma and enter the cancer cells.The tail vein injection was performed with PEG2000-AuNRs and EGFR mAb-AuNRs conjugates respectively.The direct injection of AuNRs into the tumor was used as the positive control group,and the ablation of the tumor in nude mice after NIR laser irradiation was compared.Draw tumor growth curve and survival curve.5.EGFR monoclonal antibody combined with nanogold photothermal effect targeted therapy for the treatment of head and neck squamous cell carcinoma transplanted tumor optimization and efficacy evaluation.The dose was optimized from three doses of EGFR mAb-AuNRs conjugate,near-infrared(NIR)irradiation mode and treatment procedure.Results:1.Hep-2 cells,CNE-2 cells,NP-69 cells and BEAS-2B cells are typical epithelial cells with adherent growth and similar characteristics.2.The effective killing concentration(IC50)of EGFR mAb-AuNRs on two tumor cells is almost unaffected by normal human epithelial cells.The cell inhibition rate increases as the concentration of gold nanorods is increased.3.Tumor cells+EGFR/AuNRs+NIR irradiation group showed obvious cytostatic effect;AuNRs inhibited the proliferation of CNE-2 cells,but had no effect on Hep-2 and two normal human epithelial cells.4.Observed by transmission electron microscopy:In vitro experiments,gold nanorods selectively enter the head and neck squamous cell carcinoma,mainly in the mitochondria and lysosomes,and cause cervical squamous cell cancer death after NIR laser irradiation.5.Direct injection of AuNRs into the tumors of the two tumor-bearing nude mice is effective,has a significant ablation effect on the transplanted tumor,and significantly prolongs the survival of the tumor-bearing nude mice,and the Hep-2 tumor-bearing The prognosis of nude mice is better.In the tail vein injection of EGFR mAb-AuNRs,the growth inhibition of transplanted tumors in nude mice was not obvious.6.The optimal single-time NIR laser irradiation duration of PPTT in nude mice with CNE-2 tumor was 6 minutes;the optimal irradiation power was:3 W/cm2(high necrotic mode)2.5 W/cm2(high apoptotic mode);Excellent tumor injection dose:280ug/kg.The optimal single-time NIR laser irradiation duration of PPTT in Hep-2 tumor-bearing nude mice was 6 minutes;the optimal irradiation power was 3 W/cm2;the optimal tumor injection dose was 280 ug/kg.Conclusions:1.EGFR/AuNR photothermal therapy can prevent damage to normal cells while killing head and neck squamous cancer cells.2.The photothermal treatment of tumor-injected AuNRs has a significant effect on Hep-2 and CNE-2,and the inhibition rate of Hep-2 xenografts can be 100%.The effect of tail vein injection of EGFR/AuNRs is not clear.3.The optimum single-time NIR laser irradiation duration of PPTT in nude mice with CNE-2 tumor was 6 minutes;the optimal irradiation power was:3 W/cm2(high necrotic mode)2.5 W/cm2(high apoptotic mode);Excellent tumor injection dose:280ug/kg.4.The optimal single-time NIR laser irradiation duration of PPTT in Hep-2 tumor-bearing nude mice was 6 minutes;the optimal irradiation power was 3 W/cm2;the optimal tumor injection dose was 280 ug/kg.