Shan Liang
National Center for Biotechnology Information, NLM, NIH
Antibody fragments such as diabodies and minibodies are ideal for molecular imaging because of their compact size and efficient antigen binding (1-4). However, most radiometal-labeled antibody fragments exhibit high kidney retention, which leads to decreased absolute tumor uptake (5-7). In the case of radiometals such as Cu, which have greater tumor retention than radioiodine labels, the advantage is offset by prolonged kidney retention. Use of I results in reduced kidney uptake due to metabolism and redistribution, but at the expense of lower tumor uptake (1, 8, 9). An approach to avoid the kidney retention issue is to increase the apparent molecular weight of antibody fragments through attachment of polyethylene glycol (PEG) (5). This approach has been successfully applied to the engineered antibody fragments against carcinoembryonic antigen (CEA), CD3, and others (5, 10). Increased tumor retention, longer serum half-life (t), and decreased kidney uptake have been observed for PEGylated fragments compared with non-PEGylated forms (5, 11). Li et al. reported that attachment of 1,4,7,10-tetraazacyclododecane-,','','''-tetraacetic acid (DOTA) and PEG3400 to anti-CEA diabody can significantly reduce kidney uptake of the diabody compared with kidney uptake of the parental diabody while preserving high tumor uptake (5, 12). Li et al. further generated a PEGylated (PEG3400) anti–tumor-associated glycoprotein 72 (TAG-72) diabody (AVP04-07), which reduces kidney retention to levels usually seen with intact immunoglobulin G. Kidney uptake appears to be a function of apparent molecular size (as modified by PEG) and other less defined factors intrinsic to a diabody (1). However, PEG3400 is less than ideal for clinical use because of its inherent polydispersity and the consequent inability to manufacture products that are easy to characterize chemically. Oversubstitution may also inhibit the immunoreactivity of antibody fragments. In the case of anti-CEA diabody, Li et al. showed that it is necessary to limit the number of PEG3400 to approximately one per monomer (5). Li et al. therefore evaluated the effect of monodispersed PEG on the kidney retention and tumor uptake of anti-TAG-72 diabody (1). The investigators synthesized monodispersed PEG (DOTA-PEG27-Cys-VS, where Cys-VS is vinyl sulfone conjugated to the thiol of cysteine) and conjugated this to the amino groups on the anti-TAG-72 diabody (named as PEG27 AVP04-07). Biodistribution studies with In-labeled PEG27 AVP04-07 demonstrated low kidney uptake without reduction of the tumor uptake and tumor/blood ratio. Positron emission tomography (PET) imaging with Cu-labeled PEG27 AVP04-07 (Cu-PEG27 AVP04-07) showed high-contrast tumor images (tumor/blood ratio: 8) within 24 h after injection (1).
双特异性抗体和微型抗体等抗体片段由于其紧凑的尺寸和高效的抗原结合能力，是分子成像的理想选择(1 - 4)。然而，大多数放射性金属标记的抗体片段在肾脏中的滞留率很高，这导致肿瘤的绝对摄取量降低(5 - 7)。对于像铜这样的放射性金属，其在肿瘤中的滞留时间比放射性碘标记更长，但肾脏滞留时间延长抵消了这一优势。使用碘会因代谢和重新分布而降低肾脏摄取，但代价是肿瘤摄取较低(1, 8, 9)。避免肾脏滞留问题的一种方法是通过连接聚乙二醇(PEG)来增加抗体片段的表观分子量(5)。这种方法已成功应用于针对癌胚抗原(CEA)、CD3等的工程化抗体片段(5, 10)。与未聚乙二醇化的形式相比，聚乙二醇化片段的肿瘤滞留增加、血清半衰期(t)延长且肾脏摄取减少(5, 11)。Li等人报道，与亲本双特异性抗体的肾脏摄取相比，将1,4,7,10 - 四氮杂环十二烷 - N,N',N'',N''' - 四乙酸(DOTA)和PEG3400连接到抗CEA双特异性抗体上可显著降低双特异性抗体的肾脏摄取，同时保持高肿瘤摄取(5, 12)。Li等人进一步制备了聚乙二醇化(PEG3400)抗肿瘤相关糖蛋白72(TAG - 72)双特异性抗体(AVP04 - 07)，其将肾脏滞留降低到通常在完整免疫球蛋白G中看到的水平。肾脏摄取似乎是表观分子大小(由PEG修饰)和双特异性抗体固有的其他不太明确的因素的函数(1)。然而，PEG3400由于其固有的多分散性以及因此无法制造易于化学表征的产品，不太适合临床使用。过度取代也可能抑制抗体片段的免疫反应性。在抗CEA双特异性抗体的情况下，Li等人表明有必要将PEG3400的数量限制在每个单体约一个(5)。因此，Li等人评估了单分散PEG对抗TAG - 72双特异性抗体的肾脏滞留和肿瘤摄取的影响(1)。研究人员合成了单分散PEG(DOTA - PEG27 - Cys - VS，其中Cys - VS是与半胱氨酸的硫醇共轭的乙烯砜)，并将其与抗TAG - 72双特异性抗体上的氨基共轭(命名为PEG27 AVP04 - 07)。用铟标记的PEG27 AVP04 - 07进行的生物分布研究表明肾脏摄取低，同时肿瘤摄取和肿瘤/血液比值没有降低。用铜标记的PEG27 AVP04 - 07(Cu - PEG27 AVP04 - 07)进行的正电子发射断层扫描(PET)成像显示，注射后24小时内肿瘤图像对比度高(肿瘤/血液比值: 8)(1)。
