多肽UNAM的生物合成及其在DC-CIK细胞培养中的应用  

Biosynthesis and application of polypeptide UNAM in DC-CIK cell culture

作　　者：史北辰 冯茜 韩强 陈浩东 孙越 周国栋 金媛媛[4] 张志斐 杨兆勇[4] SHI Bei-chen;FENG Qian;HAN Qiang;CHEN Hao-dong;SUN Yue;ZHOU Guo-dong;JIN Yuan-yuan;ZHANG Zhi-fei;YANG Zhao-yong(School of Pharmacy,North China University of Science and Technology,Tangshan 063210,China;Department of Internal Medicine,The Fourth People's Hospital of Zibo,Shandong 255067,China;Department of Oncology,The Fourth People's Hospital of Zibo,Shandong 255067,China;Institute of Medicinal Biotechnology,Chinese Academy of Medical Sciences&Peking Union Medical College,Beijing 100050,China)

机构地区：[1]华北理工大学药学院,唐山063210 [2]淄博市第四人民医院内科,山东255067 [3]淄博市第四人民医院肿瘤科,山东255067 [4]中国医学科学院北京协和医学院医药生物技术研究所代谢工程室,北京100050

出　　处：《中国医药生物技术》2021年第4期316-321,共6页Chinese Medicinal Biotechnology

基　　金：河北省自然科学基金(B2020209001);国家自然科学基金面上项目(81872782);国家自然科学基金国际(地区)合作与交流项目(81761128016)。

摘　　要：目的制备尿苷二磷酸-N-乙酰胞壁酸(UNAM),并研究其提升DC-CIK细胞识别及杀伤肿瘤细胞的作用。方法通过设计体外酶促反应合成多肽UNAM的路线,构建体外酶促反应体系,按照UNAM的路线图合成多肽产物,并采用制备液相纯化和收集最终产物。使用常规培养方法和添加UNAM的方法分别对DC-CIK细胞进行诱导培养,观察培养后的DC-CIK细胞对荷瘤小鼠的治疗效果。结果利用大肠杆菌异源表达系统,获得了UNAM生物合成途径中的NahK、GlmU、MurA和MurB 4个酶,并成功合成了分枝杆菌细胞壁肽聚糖生物合成中的中间产物UNAM。在动物实验中,UNAM-DC-CIK细胞对荷瘤小鼠的治疗效果比常规方法的DC-CIK细胞更好,其带瘤生存期更长(43 d),优于DC-CIK治疗组(38 d)和对照组(25 d)。结论UNAM可以提升DC-CIK细胞的增殖能力和对肿瘤的杀伤能力。Objective Study on the preparation of UDP-N-acetylmuramic acid(UNAM)and culture of DC-CIK cells using UNAM as an inducer,in order to enhance the activity and improve the recognition and killing effect of DC-CIK on tumor cells.Methods Through the design of in vitro enzymatic reaction route to synthesize peptide UNAM,the in vitro enzymatic reaction system was constructed,and the final product was purified and collected by preparative liquid chromatography.DC-CIK cells were induced by conventional culture method in the presence or absence of UNAM,and the therapeutic effect of the DC-CIK cells on tumor bearing mice was observed.Results Four enzymes,NahK、GlmU、MurA and MurB,involved in the biosynthetic pathway of UNAM were obtained by Escherichia coli heterologous expression system.UNAM,an intermediate in the biosynthesis of peptidoglycan from mycobacterial cell wall,was successfully synthesized by the enzymatic reaction in vitro.In animal experiments,the therapeutic effect of UNAM-DC-CIK cells on tumor bearing mice was better than that of DC-CIK cells cultured by conventional methods,and the survival time is longer.The longest survival time in the UNAM-DC-CIK treatment group,DC-CIK treatment group and control group was 43 days,38 days and 25 days,respectively.Conclusion The proliferation and killing ability of UNAM-DC-CIK cells cultured with UNAM were higher than that of DC-CIK cells cultured by conventional methods.

关 键 词：多肽UNAM 生物合成 DC-CIK细胞 抗肿瘤 

分 类 号：Q813.11[生物学—生物工程] R73-3[医药卫生—肿瘤]