^(125)I偶联反义核酸对裸鼠肾癌移植瘤的治疗作用

Effects of ^(125)I labeled antisense oligonucleotides against Ki67 mRNA on the growth and apoptosis of implanted human renal carcinoma cells in nude mice

目的观察125I偶联K i67基因反义核酸(ASODNs)对裸鼠肾癌移植瘤的治疗作用。方法合成K i67基因ASODNs,氯胺-T法125I偶联ASODNs(125I-ASODNs)。BALB/c种系裸鼠接种人肾癌786-0细胞。125I-ASODNs治疗组12只瘤体注射125I-ASODNs 0.1 m l(7.4 MBq/L、10 nmol),ASODNs治疗组12只瘤体注射ASODNs 0.1 m l(10 nmol),连续4 d。治疗后第3、6、12 d每组分别处死小鼠4只,取瘤组织检测肿瘤体积,免疫组化、W estern b lot技术检测K i67表达,免疫组化TUNEL法检测细胞凋亡率。结果125I-ASODNs组治疗后3、6、12 d小鼠肿瘤体积分别为(51.8±13.7)、(76.1±22.9)、(636.9±73.8)mm3,与ASODNs处理组(70.3±18.9)、(185.7±37.7)、(868.9±128.2)mm3比较,差异有统计学意义(P<0.01);K i67抗原表达率分别为(16.5±2.1)、(20.8±1.0)、(28.6±2.4)%,ASODNs组为(26.8±2.3)、(29.2±1)、(33.6±2.6)%,K i67蛋白量比分别为(54.8±2.6)、(58.6±2.9)、(63.9±3.5)%,与ASODNs组(72.6±5.1)、(79.7±3.4)、(85.7±4.4)%比较差异有统计学意义(P<0.01);肿瘤细胞凋亡率分别为(22.6±2.0)、(24.9±2.0)、(27.7±2.2)%,与ASODNs组(15.1±1.6)、(17.8±1.9)、(18.3±2.3)%比较差异有统计学意义(P<0.01)。结论125I-ASODNs比ASODNs具有更强的抑制裸鼠肾癌移植瘤生长及促进凋亡作用。

Objective To investigate the effects of ^125I labeled antisense oligonucleotides against Ki67 gene （^125I-ASODNs） on the growth and apoptosis of implanted human renal carcinoma （HRC） cells in nude mice. Methods ASODNs targeting Ki67 gene was synthesized and labeled with ^125I by Chlorasep- tine-T methods. Human renal carcinoma 786-0 cells were implanted subcutaneously in BALB/c nude mice to develop the implanted tumor model of HRC. In ^125I-ASODNs treated group （n = 12） ,0. 1 ml ^125I-ASODNs （7.4 MBq/L, 10 nmol）was directly injected into tumors in nude mice every day for 4 days. In ASODNs treated group （n = 12） ,nude mice were treated with injection of 0.1ml ASODNs （10 nmol） in the same way. The 12 nude mice in each group were killed at 3,6 and 12 days ,respectively,after treatment. The tumor sizes were measured and tumor volumes were calculated. The Ki67 expression of the tumor was detected by immunohistochemical technique and Western blot method, respectively. The apoptosis of tumor was detected by TUNEL assay. Results At 3,6 and 12 days after treatment, the tumor volumes in ^125I-ASODNs group （51.8 ± 13.7,76. 1 ± 22.9,636.9 ±73.8 mms） were significantly less than those in ASODNs group （70.3 ±18.9,185.7±37.7,868.9 ±128.2 mm^3,P〈0.01）. The Ki67 antigen expression rates in ^125I. ASODNs group （ 16.5 ± 2. 1,20.8 ±1.0,28.6 ± 2.4 % ） were significantly lower than those in ASODNsgroup （26.8 ±2.3,29.2 ± 1,33.6 ± 2.6 % ,P 〈 0.01 ）. The Ki67 protein ratios in ^125I-ASODNs group （54.8 ± 2.6,58.6 ± 2.9,63.9 ± 3.5 % ） were significantly lower than those in ASODNs group （72.6 ± 5.1,79.7 ±3.4,85.7 ±4.4 %,P 〈0.01）. The apoptosis rates of tumor cells in ^125I-ASODNs group （22.6 ± 2.0,24.9 ± 2.0,27.7 ± 2.2 % ） were significantly higher than those in ASODNs group （ 15.1 ± 1. 6,17.8 ±1.9,18.3±2.3 %,P〈0.01）. Conclusions Effects of ASODNs targeting Ki67 gene on the growth and apoptosis of implanted HRC cells in nude mice were augmented significantly when ASODNs were labeled with ^125I.