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目的 制备131I标记的抗神经纤毛蛋白-1(NRP-1)单克隆抗体A6 (131I-A6),探讨其作为NRP-1靶向显像新型分子探针的可行性.方法 (1)采用Iodogen法对A6进行131I标记,检测其标记率、放化纯和体外稳定性.(2)以人胶质瘤细胞株U87MG为实验细胞进行体外实验,测定131I-A6生物学活性、结合率及其与受体的亲和力.(3)将荷U87MG胶质瘤模型裸鼠采用随机抽样法分为5组,每组5只,分别于注射1.2 MBq 131I-A6后24、48、72、96和120 h处死,计算各脏器放射性摄取(％ID/g)、肿瘤/血液(T/B)和肿瘤/肌肉(T/M)比值.(4)取6只荷瘤裸鼠,以随机抽样法分为未阻断组和竞争阻断组,前组注射3.7 MBq 131I-A6;后组注射3.7 MBq 131I-A6和未标记的700 μg A6,均分别于注射后24、48、72、96和120 h行SPECT/CT显像.采用两样本t检验对实验数据进行统计学分析.结果 (1) 131I-A6标记率为(95.46±3.34)％,放化纯＞95％;131I-A6在室温下PBS溶液中放置至96 h,其放化纯仍＞85％.(2)131I-A6与U87MG胶质瘤细胞特异性结合率1h达到最高值,为(15.80±1.30)％,在加入未标记的抗体A6时,U87MG细胞对131 I-A6明显受抑制(t=2.862,P＜0.05);与细胞表面抗原的亲和力(Kd)为(1.67±0.14) nmol/L.(3)24h时131I-A6在荷瘤裸鼠血液放射性最高,为(8.00±1.42) ％ID/g;其次是肝脏和肿瘤组织,分别为(7.68±1.56)和(6.00±1.24) ％ID/g;脑、骨、肌肉组织放射性计数较低.给药后24h,T/B和T/M分别为0.78±0.10和3.20±0.30,随时间延长比值逐渐增高,在120 h达到最高,分别为1.87±0.50和7.13±0.24.(4)体内显像示,注射131I-A6后24 h肿瘤略显影,随时间延长变清晰,120 h显影为最清晰,阻断后未见肿瘤显影.结论 131I-A6的标记方法简单易行,标记率高,产物稳定性好,具有良好的NRP-1靶向性和特异性,有望成为一种新型肿瘤诊断和治疗的药物.

Objective To synthesize 131I labeled anti-neuropilin-1 monoclonal antibody A6 (131IA6) and evaluate its biodistribution and imaging in malignant glioma xenografts.Methods (1) A6 was labeled with 131I by Iodogen method under the optimum labeling conditions,then the labeling efficiency,radiochemical purity and stability were measured in vitro.(2) In vitro bioactivity,cellular uptake and receptor affinity of 131I-A6 with U87MG cells were measured.(3) The nude mice bearing human U87MG cells were randomly divided into 5 groups with 5 in each group.The nude mice were sacrificed by cervical dislocation and dissected at 24,48,72,96,and 120 h,respectively,after intravenous injection of 1.2 MBq 131I-A6.The biodistribution of the agent was measured as ％ID/g,and the ratios of tumor/blood (T/B) and tumor/muscle (T/M) were calculated.(4) SPECT/CT imaging was performed in 6 mice including 3 in the competitive inhibition control group at 24,48,72,96,and 120 h post injection.Two-sample t test was used for data analysis.Results (1) The labeling yield of 131I-A6 was (95.46±3.34)％,and the radiochemical purity was more than 95％.At 96 h of incubation in PBS,the radiochemical purity was more than 85％.(2)131I-A6 had rapid accumulation in U87MG cells and reached the peak of (15.80±1.30)％ at 1 h.When the probe was incubated with large excesses of non-radioactive A6,the uptake level of 131I-A6 in U87MG cells was significantly inhibited (t=2.862,P＜0.05).Kd of 131I-A6 binding to NRP-1 was (1.67±0.14) nmol/L in U87MG cells.(3) Biodistribution study showed that the uptake in blood,liver and tumor was (8.00±1.42),(7.68±1.56) and (6.00±1.24) ％ID/g at 24 h,respectively.The uptake in muscle,brain and bone was lower.The T/B and T/M were 0.78±0.10 and 3.20±0.30 at 24 h,and they reached the highest level of 1.87±0.50 and 7.13±0.24 at 120 h.(4) The SPECT imaging showed that the tumors could be visualized at 24 h and delineated more clearly at 120 h post injection of 131I-A6.Conclusions 131I-A6 can be easily synthesized by Iodogen method with high radiochemical purity.The specific tumor uptake of 131I-A6,which correlates with NRP-1 expression in gliomas,suggests that it may be a new promising tumor targeting radiotracer.