| BackgroundOn December 15,2020,the International Agency for Research on Cancer(IARC)of the World Health Organization released the latest data of global cancer burden in 2020.In 2020,there will be 19.29 million new cancer cases and 9.96 million deaths.Lung cancer still accounts for 1.8 million cancer deaths,far more than other types of cancer,ranking first in cancer deaths.In China,there will be 4.57 million new cancer cases and 3 million deaths in 2020,of which lung cancer accounts for 17.9%and 23.8%respectively,and is still the largest cancer in China.In the past 40 years,the death rate of lung cancer in China has increased fourfold,and it has been the top cause of cancer death,accounting for 27.3%of all cancer deaths in China.Therefore,lung cancer is still the most important cancer in China.The concept of maintenance therapy appeared at the end of last century,and formally entered the NCCN guidelines in 2009.After that,researchers have carried out a variety of clinical trials on various drugs,from the third generation chemotherapy drugs to antiangiogenic drugs,and then to small molecule targeted drugs,including the hot spots in the field of cancer in recent years,immunotherapy drugs and so on.However,most drugs have certain limitations in the process of maintenance therapy For example,pemetrexed benefits more in patients with non-squamous cell carcinoma;the effect of gemcitabine maintenance therapy is closely related to the general state of patients;the patients with gefitinib sensitivity are Asian,female,adenocarcinoma,and non-smoking history;bevacizumab benefits more significantly in patients with EGFR mutation;cetuximab has ethnic differences in survival benefits,and the immune response to PD-1/PD-L1 pathway Inhibitors in the Chinese market has not yet been fully affirmed and so on.At the same time,Chinese medicine has accumulated rich clinical experience and broad mass base in China for thousands of years,and began to play a unique advantage in the field of maintenance treatment.Clinical practice has proved that traditional Chinese medicine has the characteristics of effective,low toxicity,low price and long-term use,which is a strong evidence that it can be used in the maintenance treatment stage.According to the theory of traditional Chinese medicine,the fundamental cause of disease is the deficiency of vital energy.In traditional Chinese medicine,"Zhengqi"refers to the normal function of Qi and blood in human organs,meridians and collaterals.When Zhengqi is full and in normal operation,the body has certain disease resistance ability and self-healing ability.Immunity in modern medicine refers to the ability of the body to recognize foreign bodies and eliminate antigenic foreign bodies,which is basically consistent with the function of "healthy qi" in traditional Chinese medicine.Shenqi Fuzheng injection is composed of Codonopsis pilosula and Astragalus membranaceus.It can nourish Qi,replenish qi and generate blood,promote the recovery of hematopoiesis and immune function,and inhibit tumor cell metastasis.As a typical Chinese patent medicine,the effect on immune system has always been one of the focuses of anti-tumor research in traditional Chinese medicine.Jinxu Wang separated 23 components from Shenqi Fuzheng Injection by high speed counter current chromatography(HSCCC),and proved that most of them had immunopotentiation activity.Previous clinical trials at home and abroad were mostly conducted to verify the immune enhancement function of Shenqi Fuzheng injection.The only basic exploration of several immune mechanisms focused on macrophages and Jurka T cells,but NK cells as the key link between tumor immunosuppressive state and tumor progression were less studied.Previous clinical studies have shown that Shenqi Fuzheng injection can reduce chemotherapy-related fatigue of NSCLC,especially in emotional,physical and cognitive dimensions.In terms of maintenance treatment,previous small sample clinical studies have shown that there may be a certain degree of comparability in progression free survival.At the same time,the basic experimental results suggest that the main way of action of Shenqi Fuzheng injection may be related to the enhancement of body immunity.Based on the above foundation,this study will be divided into the following two parts:first,through a prospective observational study to explore the effect of Shenqi Fuzheng Injection on disease-free progression,disease control,safety and immunity of patients with advanced NSCLC.Secondly,NK-92 cells and lung cancer A549 cells were co cultured to simulate the tumor immune microenvironment,and the effects of Shenqi Fuzheng Injection on the biological behaviors of the two cells in the co culture system were observed.The possible targets and pathways of Shenqi Fuzheng Injection in tumor inhibition at the gene level were further explored by gene chip technology.ObjectiveObjective to explore the clinical efficacy and safety of Shenqi Fuzheng Injection assisted NCCN maintenance therapy through prospective clinical study.2 to establish the co culture model of NK-92 cells and lung cancer A549 cells,to observe the effect of Shenqi Fuzheng Injection on the recognition ability,killing effect and cytokine secretion of NK-92 cells,and to explore the possible target and pathway of Shenqi Fuzheng Injection on the gene level of NK-92 cells by gene chip technology.MethodClinical study:Patients with stage ?-? NSCLC who received chemotherapy in six research institutions including Guang'anmen Hospital of Chinese Academy of traditional Chinese medicine from December 2018 to December 2020 were collected.According to the actual medication situation,the subjects were divided into two groups:Shenqi Fuzheng Injection maintenance treatment group,referred to as Shenqi group(Shenqi Fuzheng injection was added on the basis of Western medicine group),western medicine NCCN guideline maintenance treatment group,referred to as western medicine group.The duration,course of treatment,progress and other information were collected and recorded.The efficacy and safety evaluation indexes of the two groups were analyzed and compared.Basic experiment:The effect cells(NK-92 cells)and target cells(A549 cells)were used as the research objects to establish the co culture model in vitro.The best effect target ratio and the best intervention concentration of the drug were selected by CCK-8 method.Then the cytotoxicity of Shenqi Fuzheng Injection on NK-92 cells was determined by LDH method.The release of IFN-? and TNF-? from NK-92 cells was detected by ELISA,and NK-92 cells were detected by flow cytometry Objective to evaluate the cytotoxicity of Shenqi Fuzheng Injection on NK-92 cells from multiple perspectives,and to explore whether the killing ability of NK-92 cells can be enhanced by enhancing the recognition ability by detecting the expression of activated mic A/B and inhibitory HLA-ABC receptor on the surface of A549 cells.Then,gene chip technology was used to screen the differential genes with up or down expression after drug intervention,and Perform Go function and KEGG enrichment analysis for the differential genes,so as to find the possible target of Shenqi Fuzheng intervention on NK-92 cell function.Research resultsClinical study:As of December 31,2020,125 cases were enrolled in the group,including 62 in western medicine group and 63 in Shenqi group.There were three cases of incomplete study in the two groups,the completion rate of both groups was above 95%,and the baseline data of the two groups were relatively balanced.In terms of gender,age,smoking history,clinical stage and complications,the two groups were more balanced,and the difference was not statistically significant(P>0.05),which was comparable.The difference between the two groups was statistically significant in the pathological type,metastasis site,ECOG score and the number of previous first-line chemotherapy courses.2.Progression free survival and survival rate:PFS in western medicine group and Shenqi group were 4.44 months and 4.11 months respectively.There was no statistical significance between the two groups(p=0.4593,95%CI 3.25-5.65).The survival rate of the two groups was higher than the western medicine group,but the difference was not statistically significant.3.Hierarchical analysis of progression free survival period:Age stratification:PFS of the western medicine group and Shenqi group was 4.60 months and 4.63 months respectively,while the PFS of the two groups over 65 years old was 3.71 months and 3.84 months respectively,and the difference was not statistically significant(P>0.05);Gender stratification:the PFS of Western medicine group and Shenqi group was 3.98 months and 3.94 months in male subjects,and that of female group was 5.95 months and 4.50 months respectively,and the difference was not statistically significant(P>0.05);BMI stratification:the PFS of the subjects with BMI<20 was 2.78 months and 3.37 months in the TCM and the Shenqi group,respectively.The PFS of BMI>20 group was 4.53 months and 4.01 months respectively,and the difference between BMI<20 group and BMI group was not statistically significant(P>0.05),and the difference between BMI>20 groups was statistically significant(p=0.0475);Pathological classification:the PFS of Western medicine group and Shenqi group were 2.99 months and 4.01 months respectively in squamous cell carcinoma subjects,4.53 months and 4.50 months in adenocarcinoma subjects,respectively,and there was no significant difference between the two groups(P>0.05);Clinical staging:the PFS of the western medicine group and Shenqi group in phase III was 6.47 months and 4.02 months,while the PFS of the two groups in phase ? was 3.94 months and 4.27 months respectively.There was no statistical significance between the two groups in phase ?(P>0.05),and the difference between the two groups in phase ? was statistically significant(p=0.0379).ECOG score was divided into two groups:PFS of subjects with ECOG=1 was 4.60 months and 3.84 months,while western medicine group showed some advantages(p=0.0157),and subjects with ECOG=2 had 1.20 months and 2.92 months respectively,and the reference Qi group showed some advantages(p=0.034 7);The PFS of the two groups were 4.40 months and 3.94 months respectively,and the PFS of the 5-6 patients was 4.01 months and 4.01 months respectively,and the difference between the two groups was not statistically significant(P>0.05);4.Cox regression analysis:gender,age,BMI and ECOG scores were independent risk factors affecting disease progression:>65 years old disease progression risk was 1.3992 times of age<=65 years old,female disease progression risk was 0.7011 times of male,BMI<20 disease progression risk was 1.6670 times of BMI>=20,0.9413 and 0.5321 times of 2 points in ECOG=0 and ECOG=1 point respectively.5.Disease control rate:after the sixth course,the disease control rate of Shenqi group was better than that of Western medicine group(76.00%vs 41.67%,P=0.038),and there was no statistical significance after the second and fourth courses(P>0.05).6 clinical symptoms:The score of fact-4.0 scale:the baseline of the two groups was balanced,and the observation value of Shenqi group was lower than that of Western medicine group(p=0.0126),but the observation value after the sixth course was better than that of Western medicine group(p=0.0287),and the relative baseline change of Shenqi group was more significant than that of Western medicine group,and the difference was statistically significant(P<0.0001);Score of symptom scale of traditional Chinese medicine:the data of the two groups were unbalanced at baseline.Compared with the observation values after the second,fourth and sixth courses,the scores of Shenqi group were higher than those in the western medicine group,indicating that the subjects included in the Shenqi group had more severe symptoms of TCM and the general situation was poor.But the observation values of Shenqi group were more significant than that of Western medicine group after the second,fourth and sixth courses(P<0.0001)Score of Piper Fatigue Scale:the data baseline of the two groups was comparable,and the observation value difference was not statistically significant after the second,fourth and sixth courses,but after the second and sixth courses,the relative baseline changes of Shenqi group were more significant than that of the western medicine group,and the difference was statistically significant(P<0.05)7.Immune function:the number of cd3+,cd4+,cd4+/cd8+and NK cells in Shenqi group showed an upward trend,especially the increase trend of cd3+and NK cells was more significant,while the control group showed a downward trend,and the difference between the two groups was significant(P<0.05),8 adverse reactions:the incidence of appetite decline,nausea,fatigue and other symptoms in Shenqi group was lower than that of Western medicine group,and the difference was statistically significant(P<0.001),There was no significant difference between the two groups in the incidence of pain and hair loss(P>0.05);in the aspect of the decrease of hemoglobin,leukocyte and neutrophil,Shenqi group was better than that of Western medicine group(P<0.05);the serum glutamic transaminase and ?-glutamyl-tripeptide enzyme were higher than that of the western medicine group(P<0.01).Basic experiment:The results of CCK-8 showed that the best target ratio of NK-92 and A549 cells was 20:1,and the best intervention concentration of Shenqi fuzheng injection was 60 ?L/ml.two In the co culture system of NK-92 cells and A549,the cell lysis increased significantly,which was significantly different from that of NK-92 and A549 cells(P<0.01);compared with the control group,the cell lysis increased slightly,but the difference was not statistically significant(P>0.05).The addition of Shenqi Fuzheng injection was not statistically significant(P>0.05)Compared with A549 and NK-92 cells,the co culture system of the solution was significantly increased,and the difference was statistically significant(P<0.01).The intervention of Shenqi Fuzheng Injection and the stimulation of A549 cells can promote the secretion of IFN-y,TNF-? and expression of granule enzyme CD107a(P<0.05).When the injection of Shenqi Fuzheng is added into the co culture system of NK-92 and A549,the secretory cytokines and degranulation of NK-92 and A549 are significantly higher than that of the single culture group(P<0.01).The positive regulation not only exists in the single layer culture of A549,but also exists in the co culture system of A549 and NK-92,and the difference between the two conditions is not statistically significant(P>0.05),but it has no regulation effect on HLA ABC.After the dry prognosis of Shenqifuzheng injection,395 genes were expressed in NK-92 cells,106 genes were up regulated and 289 genes were down regulated,but the most of the 20 genes with the largest FC value gap were up regulated genes.The results of the enrichment analysis of go showed that the correlation between the difference genes and BP was the highest,mainly focused on the functions of lipid metabolism,protein folding and cell membrane depolarization.The results of KEGG pathway analysis showed that the effects of Shenqi Fuzheng Injection on cell genes were mainly in cell metabolism,disease stimulation,cell function and genetic material regulation.research conclusionConclusion:Shenqi Fuzheng injection cannot prolong PFS and increase survival benefit of patients on the basis of Western medicine maintenance treatment.Shenqi Fuzheng Injection Combined with western medicine maintenance treatment shows certain advantages in disease control,improvement of symptoms,enhancement of immunity and reduction of adverse reactions.And in improving the symptoms of the subjects,there is a positive correlation between time and efficacy,that is,the closer to the later stage of the research process,the more advantages of traditional Chinese medicine treatment can be displayed.Shenqi Fuzheng injection can cooperate with NK-92 to exert cytotoxicity on A549 cells by regulating the receptor on the surface of target cells,activating NK-92 cells and promoting the secretion of granzyme CD 107a,cytokines TNF-? and IFN-?.The results of gene chip,go and KEGG enrichment analysis further confirmed that the intervention of Shenqi Fuzheng injection had a greater correlation with the biological process of NK cells killing target cells through direct contact with perforin granzyme pathway and secreting various cytokines through exocytosis. |
PDF Full Text Request