Vorobyeva Anzhelika, Din Moeen-Ud, Schulga Alexey, Konovalova Elena, Abouzayed Ayman, Bragina Olga, Li Ruonan, Gräslund Torbjörn, Deyev Sergey M, Oroujeni Maryam
Department of Immunology, Genetics and Pathology, Uppsala University, 751 85, Uppsala, Sweden.
Molecular Immunology Laboratory, Shemyakin and Ovchinnikov Institute of Bioorganic Chemistry, Russian Academy of Sciences, 117997, Moscow, Russia.
EJNMMI Radiopharm Chem. 2025 May 30;10(1):26. doi: 10.1186/s41181-025-00347-6.
Epithelial cell adhesion molecule (EpCAM) is a transmembrane glycoprotein, which is overexpressed in several types of malignancies. Designed ankyrin repeat protein (DARPin) Ec1 is a 19 kDa engineered scaffold protein that binds with high affinity to EpCAM. Radiolabelled Ec1 might be used as a companion diagnostic for the selection of patients for personalized therapy. This study aimed to investigate the influence of different radiometal-chelator complexes on the biodistribution and imaging contrast of Ga-labelled Ec1. To investigate this, two macrocyclic chelators, 1,4,7-triazacyclononane-N,N,N-triacetic acid (NOTA) and 1-(1,3-carboxypropyl)-1,4,7-triazacyclononane-4,7-diacetic acid (NODAGA) were conjugated to the C-terminus of the Ec1. The previously developed DARPin Ec1 conjugated to 1,4,7,10-tetraazacylododecane-1,4,7,10-tetraacetic acid (DOTA) was used as a comparator.
All Ec1 variants were successfully labelled with Ga. The use of NOTA and NODAGA provided twice higher radiochemical yield and improved label stability compared to DOTA. All labelled Ec1 variants bound to the EpCAM-expressing cells with nanomolar affinity and preserved targeting specificity in vitro and in vivo. Biodistribution studies in mice bearing EpCAM-expressing SKOV-3 xenografts showed that [Ga]Ga-Ec1-NOTA had lower uptake in most normal organs while maintaining tumor uptake. Among all variants, [Ga]Ga-Ec1-NOTA showed the lowest liver uptake, with no significant differences in tumor uptake. Additionally, [Ga]Ga-Ec1-NOTA provided the highest tumor-to-blood ratio compared to [Ga]Ga-Ec1-DOTA and [Ga]Ga-Ec1-NODAGA.
[Ga]Ga-Ec1-NOTA is the preferred radioconjugate for PET imaging of EpCAM expression.
上皮细胞粘附分子（EpCAM）是一种跨膜糖蛋白，在多种恶性肿瘤中过度表达。设计锚蛋白重复蛋白（DARPin）Ec1是一种19 kDa的工程化支架蛋白，能与EpCAM高亲和力结合。放射性标记的Ec1可作为选择患者进行个性化治疗的伴随诊断剂。本研究旨在探讨不同放射性金属螯合剂复合物对镓标记的Ec1生物分布和成像对比度的影响。为研究此问题，将两种大环螯合剂，1,4,7-三氮杂环壬烷-N,N,N-三乙酸（NOTA）和1-(1,3-羧丙基)-1,4,7-三氮杂环壬烷-4,7-二乙酸（NODAGA）与Ec1的C末端偶联。将先前开发的与1,4,7,10-四氮杂环十二烷-1,4,7,10-四乙酸（DOTA）偶联的DARPin Ec1用作对照。
所有Ec1变体均成功用镓标记。与DOTA相比，使用NOTA和NODAGA可提供两倍更高的放射化学产率并提高标记稳定性。所有标记的Ec1变体均以纳摩尔亲和力与表达EpCAM的细胞结合，并在体外和体内保持靶向特异性。对携带表达EpCAM的SKOV-3异种移植瘤的小鼠进行的生物分布研究表明，[镓]镓-Ec1-NOTA在大多数正常器官中的摄取较低，同时保持肿瘤摄取。在所有变体中, [镓]镓-Ec1-NOTA的肝脏摄取最低，肿瘤摄取无显著差异。此外，与[镓]镓-Ec1-DOTA和[镓]镓-Ec1-NODAGA相比，[镓]镓-Ec1-NOTA提供了最高的肿瘤与血液比值。
[镓]镓-Ec1-NOTA是用于EpCAM表达PET成像的首选放射性偶联物。
