| Objective This study intends to use antinib to inhibit tumor angiogenesis,improve the hypoxic microenvironment of NSCLC,combine Carbogen gas with high concentration oxygen inhalation,and combine with modern radiotherapy technology to study its effect on the radiotherapy sensitivity of NSCLC.Methods After treating nude mouse model with anrotini combined with Carbogen,hypoxia probe of PET-CT was used to detect the change of oxygenation status in the tumor area.Immunohistochemical method was used to detect the expression levels of hypoxic factor and apoptotic factor,and the proliferation and apoptosis levels of lung cancer cell line A549 were detected at the cell level after hypoxic and oxygen-enriched combination with antinib and radiotherapy.In clinical trials,the efficacy of amlotinib and Carbogen combined with radiotherapy in NSCLC patients was evaluated,and it was finally determined whether amlotinib combined with Carbogen could increase the apoptosis of NSCLC cells and thus improve radiotherapy sensitivity in NSCLC.Conclusion The combination of antinib with Carbogen improved the tumor microenvironment,and caused hi F-1 expression to decrease by acting on tumor cells,which led to the decreased expression of apoptotic factor Bcl-2 and increased expression of caspase-3,inhibited the proliferation ability of tumor cells,and improved the radiotherapy sensitivity.The combination of the three factors can improve the radiotherapy sensitivity of NSCLC.The treatment-related adverse reactions can be tolerated by clinical patients and can be safely used in clinical practice,which is worthy of further promotion and research.Part Ⅰ Study on the effect of anrotidine combined with Carbogen inhalation on the sensitivity of NSCLC to radiotherapy in nude mice and its mechanismObjective This study was intended to investigate the effects of anrotinib combined with Carbogen on tumor microenvironment and the synergistic effects of combined radiotherapy on tumorigenesis in nude mice with NSCLC.Methods Using A549 cell line is constructed non-small cell lung cancer in nude mice tumor models,joint ROM and Carbogen for Ann,the nude mouse model of tumor tissue by micro PET detection lack of oxygen to improve the situation,and the lack of oxygen in the tumor tissue area immunofluorescence and radiation from the imaging detection,clear markers pp mo indications and lack of oxygen GULT-1 the expression of change,at the same time as a tumor in nude mice model of Ann ROM for and after Carbogen combined radiotherapy effect,preparation of lung cancer pathological specimens,Immunohistochemistry was used to detect the expression levels of hif-1a,ki-67,apoptotic factors Caspase3 and bcl-2 in the samples.Results 1.The success of this research is constructed non-small cell lung cancer in nude mice tumor models,after 18 f-FDG PET the micro examination,Ann,for he combined effects Carbogen group of lung cancer in nude mice tissues for 18 f-FDG uptake was significantly lower than the air inhaled group(P = 0.02),than pure Carbogen inhalation lung cancer in nude mice group of 18 f-FDG uptake was also significantly lower than the air inhaled group(P = 0.03),and normal tissue in nude mice in the experimental group and control group of 18 f-FDG uptake have little impact;2.Immunofluorescence tests revealed pure air suction,nude mouse lung tumor tissue area lack oxygen probe pp indications and GULT-1 higher expression,organization region shows clear,lack of oxygen in the experimental group for ernesto,joint Carbogen inhalation and pure Carbogen inhalation group,lack of oxygen of pp mo indications express decline and fluid in regions littered GLUT-1 higher expression,show that before the lack of oxygen area after the combination and pure Carbogen role,have been in the oxygenation state;3.In autolithography studies,the uptake of 18f-fdg was detected in tumor tissues of nude mice after inhalation of anrotidine in combination with Carbogen,and the intake amount(4.19± 1.32%)was significantly lower than that of the air inhalation group(9.67± 4.35,n=5 mice,P<0.01).In addition,in the anrotidine and Carbogen group,the expression of piperonidazole was decreased in the area with high expression of tumor tissue glut-1.4.Immunohistochemical assay results showed that the expression levels of hif-1a,ki-67 and bcl-2 in the group treated with anrotidine and Carbogen were significantly higher than those in the group treated with radiotherapy alone,while the expression levels of apoptotic factor Caspase3 were significantly lower.Conclusion 1.In the tumor-forming model of nude mice with non-small cell lung cancer,the combination of anrotidine and Carbogen can improve the oxygenation state in the hypoxic area of the tumor and improve the hypoxic microenvironment.2.In the tumorigenic model of NSCLC in nude mice,the expression of hif-1 in tumor tissues decreased,the expression of ki-67 decreased,the expression of apoptotic factor bcl-2 decreased and the expression of caspase-3 increased after the combination of amrotenib and Carbogen and radiotherapy,the proliferation level of tumor was significantly reduced and the radiosensitivity was enhanced.Part Ⅱ The study of the effect and mechanism of anrotidine combined with hyperoxia combined radiotherapy on A549 lung cancer cellsObjective the purpose of this study was to investigate the mechanism of radiosensitivity changes in A549 cell lines cultured under different oxygen concentrations after the improvement of anrotinib in hypoxic microenvironment.Methods After A549 cell lines were treated with hypoxic culture combined with radiotherapy and normoxic culture combined with radiotherapy,the proliferation level of cells was detected by MTS and scour method,and the apoptosis level was detected by flow cytometry.The expression levels of HIF-1a,Ki-67,apoptotic factors Caspase3 and Bcl-2 were detected by western blot.Results 1.The MTS test results showed that the cell proliferation level was the lowest in the normoxic 24h+ radiotherapy 4Gy+ amputinib group,while the cell proliferation level was the highest in the simple hypoxic group(P=0.01).2.In the scratch experiment,it was found that compared with the normox24 h + radiotherapy 4Gy+ anrotinib group,the cell scratch growth rate of the control group with low oxygen for 24 h was the fastest,and the cell scratch growth rate of the normox24 h + radiotherapy 4Gy+ anrotinib group was also faster than that of the normox24 h + radiotherapy 4Gy+ anrotinib group.3.Flow cytometry analysis showed that compared with the apoptosis level of cells in the hypoxic 24h+ radiotherapy 4Gy+ anrotinib group,the apoptosis rate of cells in the anrotinib group at the later stage was significantly increased in the normoxic 24h+ radiotherapy 4Gy+ anrotinib group(P<0.05).4.Western blot results showed that Caspase3 expression decreased and increased in the iso-24 h+ radiotherapy 4Gy+ amrotinib group,and the expression levels of hif-1a,ki-67 and bcl-2 all decreased.Conclusion One of the mechanisms by which the combination of amputinib and hyperoxia enhanced the sensitivity of A549 cells to radiotherapy may be that after the interaction between the two to improve the tumor microenvironment,the expression of hif-1 in A549 cells of non-small cell lung cancer was decreased,and the expression of apoptotic factor bcl-2 was decreased and the expression of Caspase3 was increased,leading to the decrease of proliferation ability.Part Ⅲ Clinical observation of anrotidine combined with Carbogen in the treatment of locally advanced non-small cell lung cancerObjective In this clinical study,we intended to observe whether anrotidine combined with Carbogen and three-dimensional conformal radiotherapy could improve the local control rate and short-term efficacy of patients with NSCLC.Methods 50 pathologically confirmed elderly patients with stage III non-small cell lung cancer were randomly divided into experimental group(errotinib combined with Carbogen radiotherapy)and control group(Carbogen combined radiotherapy).Patients in both groups received prescription doses of 60-66 gy /30-33 times,the same standard three-dimensional conformal radiotherapy,and Carbogen gas inhalation was given 5min before each radiotherapy,and the inhalation time was 15±6min until the end of radiotherapy,and the gas flow was set to 10L/mi.Usage of amrotinib: patients in the experimental group were given the recommended dose of 12mg/ time,once a day,taken orally before breakfast,for 2 consecutive weeks,and stopped for 1 week.Results All enrolled patients completed the scheduled treatment and follow-up.At the end of 3 and 6 months,CR was 52% and 24%,60% and 32%,respectively,in the experimental group and the control group(p<0.05),and the total effective rate was 84% and 48%,84% and 56%,respectively(p<0.05).In this study,the PR sensitization ratio of Carbogen combined with radiotherapy was 1.35 and CR sensitization ratio was 1.2.The experimental group was between 30-40 gy,and the regression rate of the two groups was not significantly different.However,the regression rate of the two groups was statistically significant at 50-60 gy,indicating that the local control rate of the experimental group was better.Adverse reactions of observation,the results did not see obvious liver,kidney and nerve toxicity,heart toxicity level III and above reaction,not observed clinical grade Ⅳ radiation reaction was observed.The elevated blood pressure in the experimental group was higher than that in the control group.Hand-foot syndrome did not appear in the control group.Gastrointestinal reactions were similar in both groups.Conclusion 1.The combination of anrotinib and Carbogen in the radiotherapy of NSCLC can improve the local control rate,PR rate and CR rate,suggesting that the combination of the two can improve the radiotherapy sensitivity of NSCLC.2.When anrotidine combined with Carbogen combined radiotherapy is clinically applied to elderly patients with non-small cell lung cancer,the side effects of anrotidine including hypertension and hand-foot syndrome can be tolerated after active treatment,but close attention should be paid to the treatment process. |
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