靶向CD93分子探针制备及其生物学实验研究

CD93分子作为一种新型新生血管生成激活剂,有可能成为肿瘤监测的重要分子靶点,因此制备125I标记抗CD93单抗(125I-anti-CD93 mAb),研究其在荷瘤鼠体内生物学分布及磷屏显像特点,探讨其对A549肺癌靶向性及CD93阳性肿瘤监测的可行性。本文制备125I-anti-CD93 mAb及125I-IgG并进行鉴定;建立A549肺癌荷瘤小鼠模型,经尾静脉注射125I标记抗CD93单抗,并设125I-IgG为对照组,注药后不同时间进行生物学分布及磷屏放射自显影;肿瘤组织行HE染色及CD93免疫组织化学染色;采用统计软件对定量数据(x+s)进行统计学处理,组间数据用t检验,P<0.05为差异有统计学意义。结果表明,125I-anti-CD93 mAb及125I-IgG标记率、放射性比活度、放化纯度分别为91.37%和90.24%,1096.44 MBq/mg和1082.88 MBq/mg,96.49%和94.82%,放置72 h后两标记物的稳定性仍>90%,两者间无统计学差异(生理盐水组P=0.93,t=0.09,血清组P=0.13,t=1.90)。荷瘤鼠生物学分布结果显示125I-anti-CD93 mAb主要经肝肾代谢,注射后48 h时肿瘤组织放射性摄取率为(6.42±0.71)%ID/g,T/NT(瘤/对侧肌肉)摄取比值为4.45±0.86,显著高于对照组125I-IgG组(2.45±0.33/1.71±0.24),P<0.05,t=3.07和P<0.01,t=5.10。动态全身磷屏自显影结果可见,24 h两组小鼠轮廓清楚,48 h时实验组肿瘤显像最清晰,肿瘤放射性摄取比(肿瘤/对侧肌肉)为3.34±0.18,明显高于对照组(1.62±0.19),P<0.01,t=6.52。免疫组化染色结果证实肿瘤高表达CD93。125I-anti-CD93 mAb制备简便,对A549肺癌组织靶向性好,有望成为CD93阳性肿瘤监测的新型分子探针。

Preparation of CD93 Targeting Probe and its Biological Experiments

As a novel activator in the process of angiogenesis,CD93 may be an important molecular target for tumor surveillance,therefore,125I-anti-CD93 mAb was prepared to study the biological distribution and the characteristics of phosphor screen imaging of 125I-anti-CD93 mAb in tumor-bearing mice,to explore its targeting to A549 lung cancer and the feasibility of CD93 positive tumor monitoring.Radionuclide markers 125I-anti-CD93 mAb and 125I-IgG were prepared and identified;A549 lung cancer bearing mice model was established.125I-labeled anti-CD93 monoclonal antibody was injected into tail vein,and 125I-IgG was used as control.Biological distribution and phosphorus screen autoradiography were performed at different time after injection;HE staining and CD93 immunohistochemical staining were performed on tumor tissues.Quantitative data(x+s)were statistically processed by statistical software,and t-test was used for data between groups,P<0.05 was considered statistically significant.The results showed that the labeling rate,specific activity and radiochemical purity of 125I-anti-CD93 mAb and 125I-IgG were 91.37%and 90.24%,1096.44 MBq/mg and 1082.88 MBq/mg,96.49%and 94.82%respectively,and the stability of the two markers was still above 90%after 72 hours,there was no statistical difference between the two(normal saline group P=0.93,t=0.09,serum group P=0.13,t=1.90).Biological distribution of the tumor-bearing mice showed that 125I-anti-CD93 mAb were mainly metabolized through the liver and kidney,the tumor tissue(6.42±0.71)%ID/g and the T/NT(tumor/contralateral muscle)uptake ratio was 4.45±0.86 at 48 hours after injection,significantly higher than the control group 125I-IgG group(2.45±0.33/1.71±0.24),P<0.05,t=3.07 and P<0.01,t=5.10.The results of dynamic whole body phosphorus screen autoradiography showed that the contours of two groups was clear at 24 hours,and the most clear tumor imaging of the experimental group was obtained at 48 hours after the injection,the tumor radioactivity uptake ratio(tumor area/opposite-area)was 3.34±0.18,significantly higher than that of the control group(1.62±0.19),P<0.01,t=6.52.Immunohistochemical staining showed that the tumor was highly expressed CD93.125I-anti-CD93 mAb is easy to prepare and has good targeting effect on A549 lung cancer tissue.There CD93 may be a new molecular probe for CD93 positive tumor monitoring.