靶向胃癌干细胞单克隆抗体2C12的鉴定及功能研究  

Identification and functional study of 2C12 monoclonal antibody against gastric cancer stem cells

作　　者：张弢 李建周 马小军 郑蕊 舒雄[2] 王淼舟[3] ZHANG Tao;LI Jianzhou;MA Xiaojun;ZHENG Rui;SHU Xiong;WANG Miaozhou(The Second People's Hospital of Xining,Qinghai Xining 810003,China;Beijing Jishuitan Hospital/Beijing Institute of Traumatology and Orthopedics,Beijing 100035,China;Qinghai University Affiliated Hospital,Qinghai Xining 810012,China.)

机构地区：[1]西宁市第二人民医院,青海西宁810003 [2]北京积水潭医院/北京市创伤骨科研究所,北京100035 [3]青海大学附属医院,青海西宁810012

出　　处：《现代肿瘤医学》2023年第8期1386-1390,共5页Journal of Modern Oncology

基　　金：青海省科技厅应用基础研究项目(编号:2020-ZJ-779)。

摘　　要：目的:研究胃癌干细胞的功能性单克隆抗体2C12体外功能实验,为胃癌干细胞的靶向治疗提供候选单抗药物。方法:以胃癌细胞系SNU-5为细胞模型,流式细胞术检测其亲本细胞和通过无血清悬浮培养的球体细胞中2C12+细胞的表达水平,双色细胞免疫荧光检测单抗2C12和CD90识别的抗原在SNU-5细胞中的表达情况。流式细胞术分选SNU-5细胞中2C12+和2C12-细胞,采用无血清悬浮培养成球实验和Transwell小室法分别检测其自我更新能力和侵袭能力。用单抗2C12处理SNU-5的球体细胞后,检测其对SNU-5的球体细胞自我更新能力和侵袭能力的影响。结果:2C12+细胞在SNU-5的球体细胞中的表达水平明显高于亲本细胞。免疫荧光结果显示单抗2C12识别的抗原分子能与CD90在SNU-5细胞上共定位。流式细胞术分选结果表明SNU-5细胞中2C12+细胞成球数明显高于2C12-细胞(103.3±9.07 vs 50.67±5.86)(P<0.01),侵袭能力也明显较高(209.3±12.9 vs 99.0±3.61)(P<0.01)。对2C12单抗的体外功能研究发现,不同浓度的单抗2C12(0μg/mL、20μg/mL、40μg/mL)能显著抑制SNU-5的球体细胞的成球能力(122.0±5.66、83.0±5.66、59.0±4.24),抑制率达到31.97%和51.64%,同时侵袭能力也显著降低(178.0±8.49、106.5±5.0、78.0±2.83),抑制率达到40.17%和56.18%。结论:单抗2C12是一株抗胃癌干细胞的功能性单抗,能够显著抑制胃癌干细胞的干性相关特征,为胃癌干细胞靶向治疗的候选抗体药物。Objective:To identify functional monoclonal antibody(2C12)that can target gastric cancer stem cells(CSCs)and provide antibody candidate drugs for targeted therapy of gastric CSCs.Methods:Taking gastric cancer cell line SNU-5 as a cell model,the expression of 2C12+cells from parent cells and sphere cells was detected by flow cytometry.The expression of antigen protein,which was identified by both monoclonal antibody 2C12 and CD90 in SNU-5 cells,was detected by two-color immunofluorescence.Flow cytometry was used to separate 2C12+cells and 2C12-cells.The spheroidization experiment in serum-free suspension culture and Transwell method were used to detect their self-renewal ability and invasion ability.After SNU-5 sphere cells were treated with 2C12,SNU-5 sphere cells'ability of self-renewal and invasion was detected.Results:The expression of 2C12+cells from sphere cells was significantly higher than that of parent cells from SNU-5 cells.The cell immunofluorescence analysis showed that the antigen identified by 2C12 and CD90 could be co-localized in SNU-5 cells.Sphere formation abilities in 2C12+and 2C12-SNU-5 cells were identified(103.3±9.07 vs 50.67±5.86)(P<0.01).The invasion ability of 2C12+cells was significantly higher than that of 2C12-cells(209.3±12.9 vs 99.0±3.61)(P<0.01).The study on functional properties of antibody in vitro illustrated that different concentrations of 2C12(0μg/mL,20μg/mL,40μg/mL)could significantly inhibit sphere formation of sphere cells from SNU-5 cells(122.0±5.66,83.0±5.66,59.0±4.24),and the inhibition rate reached 31.97%and 51.6%,meanwhile,2C12 could inhibit the invasion ability of sphere cells from SNU-5 cells(178.0±8.49,106.5±5.0,78.0±2.83),and the inhibition rate was 40.17%and 56.18%.Conclusion:2C12 monoclonal antibody is one of functional monoclonal antibodies targeting gastric CSCs,which can significantly inhibit the stem-related characteristics of gastric CSCs,and could be a candidate drug for the targeted therapy of gastric CSCs.

关 键 词：胃癌干细胞 单克隆抗体 成球能力 侵袭能力 

分 类 号：R735.2[医药卫生—肿瘤]