^(99m)Tc标记的CXCR4小干扰RNA对人乳腺癌目的基因表达的影响  

Effect of CXCR4-siRNA labeled with ^(99m)Tc on the target gene expression of human breast cancer

作　　者：姜廷军[1] 李丽 曹学良[1] 栾厦[1] 付鹏[3] 赵长久[1] JIANG Ting-jun;LI Li;CAO Xue-liang;LUAN Sha;FU Peng;ZHAO Chang-jiu(Department of Nuclear Medicine,The Fourth Affiliated Hospital of Harbin Medical University,Harbin 150001,China;Department of PET-CT,Heilongjiang Province Land Reclamation Headquarters General Hospital,Harbin 150088,China;Department of Nuclear Medicine,The First Affiliated Hospital of Harbin Medical University,Harbin 150001,China)

机构地区：[1]哈尔滨医科大学附属第四医院核医学科,黑龙江哈尔滨150001 [2]黑龙江省农垦总局总医院PET-CT中心,黑龙江哈尔滨150088 [3]哈尔滨医科大学附属第一医院核医学科,黑龙江哈尔滨150001

出　　处：《哈尔滨医科大学学报》2018年第3期206-211,共6页Journal of Harbin Medical University

基　　金：黑龙江省自然科学基金面上项目(H2015066)

摘　　要：目的探讨以靶向趋化因子受体4(CXCR4)为靶点的^(99m)Tc标记小干扰RNA(small interference RNA,siRNA)对人乳腺癌MDA-MB-231细胞目的基因表达的影响,为后续的乳腺癌CXCR4-siRNA基因显像奠定基础。方法人工设计合成三段针对CXCR4 mRNA的siRNA,同时设计出阴性对照序列,质体分别包裹三段干扰序列,转染MDA-MB-231细胞,24 h后进行PCR实验,检测CXCR4 mRNA的表达情况,在48 h后进行Western blot实验,检测CXCR4蛋白的表达,综合分析,选出最佳干扰序列。使用螯合剂HYNIC对干扰效果最好siRNA序列及siRNA(阴性对照)进行放射性^(99m)Tc标记,分别检测其标记率、放射性化学纯度、稳定性及干扰活性。多组间计量资料比较用方差分析进行比较,两组间计量资料比较用LSD-t检验进行比较。结果三段不同序列siRNA作用于MDA-MB-231细胞后,CXCR4 siRNA1作用后的CXCR4 mRNA及蛋白表达量减少的情况与其它两条干扰序列差异有统计学意义。^(99m)Tc-HYNICCXCR4 siRNA1的标记率为(61. 26±2. 47)%(n=5),阴性对照^(99m)Tc-HYNIC-CXCR4 siRNA的标记率为(60. 85±2. 76)%(n=5)。^(99m)Tc-HYNIC-CXCR4 siRNA1的稳定性好,放射性化学纯度均能达到90%。^(99m)Tc标记前后CXCR4 siRNA干扰活性无差别。结论 CXCR4 siRNA探针能与CXCR4 mRNA上的目的序列结合,抑制目的基因的表达。合成的探针分子稳定性好,具备活体乳腺癌显像的基础条件,有望为乳腺癌精准诊疗提供一条新路径。Objective To investigate the effect of human chemokine receptor 4 （CXCR4） small interference RNA （siRNA） labeled with ^99mTc on the target gene expression of human breast cancer MDA-MB-231 cells, and to lay the foundation for the subsequent development of CXCR4-siRNA gene in breast cancer. Methods Three siRNAs targeted to CXCR4 mRNA were designed and synthesized, and negative control sequences were designed. The plastics were wrapped in three sections of the interference line MDA-MB-231. The mRNA and protein h transfection was investigated and Western sequence, its carrier transfection breast cancer cell expression of the target gene with RT-PCR after 24 blot was used after 48 h to filter out the target sequence which interference effects is best. The HYNIC as its chelating, radiolabeled the CXCR4 siRNA sequence that interference effects was best and siRNA （negative control） radioactive with ^99mTc. The labeling efficiency, stability, radiochemical purity, the activity of molecular hybridi-zation and the interference activity after radiolabeled were analysed. Multivariate metering data were compared using the variance analysis, and the comparison between the two groups was compared with the LSD-t test. Results Three different sequences of siRNA were administered to MDA-MB-231 cells, CXCR4 siRNA1 after the role of CXCR4 mRNA and protein expression decreased with the other two interference sequence difference was statistically significant. The labeling rate of ^99mTc-HYNIC-CXCR4 siRNA1 was （61.26 ±2.47 ） % （ n = 5 ） , and the labeling rate of the negative control ^99mTc-HYNIC-CXCR4 siRNA was （60. 85 ± 2. 76） % （ n = 5 ）. ^99mTc-HYNIC-CXCR4 siRNA1 had good stability. Radioactive chemical purity reached 90%. There was no difference in CXCR4 siRNA interference activity before and after 99roTe labeling. Conclusion The CXCR4 siRNA probe binds to the target sequence on CXCR4 mRNA, inhibiting the expression of the gene of interest. Synthetic probe molecular stability, with the basic condition

关 键 词：靶向趋化因子受体4 SIRNA MDA-MB-231细胞 脂质体2000 ^99MTC 

分 类 号：R737.9[医药卫生—肿瘤]