Background: Fibroblast activation protein (FAP)-targeted theranostic radiopharmaceuticals have shown desired tumor-to-background organ selectivity due to the ubiquitous presence of FAP within the tumor microenvironment. However, suboptimal tumor retention and fast clearance have hindered their use to deliver effective cancer therapies. With well-documented FAP-targeting moieties and linkers appending them to optimal chelators, the development of copper radiopharmaceuticals has attracted considerable interest, given the fact that an ideal theranostic pair of copper radionuclides (64Cu: t1/2= 12.7 h; 17.4% β+; Eβ+max= 653 keV and67Cu: t1/2= 2.58 d; 100% β−; Eβ−max= 562 keV) are available. Herein, we report our design, synthesis, and comparative evaluation of monovalent and divalent FAP-targeted theranostic conjugates constructed from our previously reported bifunctional chelator scaffold (BFS) based on 1,4,8,11-tetraaza-bicyclo [6.6.2]hexadecane-4,11-diacetic acid (CB-TE2A), which forms the most stable complex with Cu(II).Methods: After synthesis and characterization, the monovalent and divalent conjugates were radiolabeled with64Cu for in vitro cell assays, followed by in vivo positron emission tomography (PET) imaging evaluation in relevant mouse models.Results: Both64Cu-labeled conjugates showed high in vitro stability and anticipated FAP-mediated cell binding and internalization. The divalent one showed significantly higher FAP-specific tumor uptake than its monovalent counterpart.Conclusions: Our results demonstrate that the BFS-based multivalent approach can be practically used to generate FAP-targeted radiotheranostic agents for effective cancer diagnosis and treatment.
背景：成纤维细胞激活蛋白（FAP）靶向诊疗一体化放射性药物因其在肿瘤微环境中的广泛存在而展现出理想的肿瘤与背景器官选择性。然而，次优的肿瘤滞留和快速清除限制了其在有效癌症治疗中的应用。基于已有充分文献记载的FAP靶向基团及将其连接至最佳螯合剂的连接子，铜放射性药物的开发引起了广泛关注，这得益于铜放射性核素（64Cu：半衰期12.7小时；β+衰变率17.4%；最大β+能量653 keV；67Cu：半衰期2.58天；β-衰变率100%；最大β-能量562 keV）构成了理想的诊疗配对。本文报道了我们基于先前报道的双功能螯合剂支架（BFS）——该支架采用与Cu(II)形成最稳定配合物的1,4,8,11-四氮杂-双环[6.6.2]十六烷-4,11-二乙酸（CB-TE2A）——设计、合成并对比评估了单价与二价FAP靶向诊疗偶联物。 方法：在完成合成与表征后，将单价与二价偶联物用64Cu进行放射性标记，用于体外细胞实验，随后在相关小鼠模型中进行体内正电子发射断层扫描（PET）成像评估。 结果：两种64Cu标记的偶联物均表现出高体外稳定性及预期的FAP介导的细胞结合与内化作用。其中二价偶联物显示出显著高于单价偶联物的FAP特异性肿瘤摄取。 结论：我们的研究结果表明，基于BFS的多价策略可实际应用于开发FAP靶向放射性诊疗药物，以实现有效的癌症诊断与治疗。
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