Folate receptor β (FR-β) is one of the markers expressed on macrophages and a promising target for imaging of inflammation. Here, we report the radiosynthesis and preclinical evaluation of [⁶⁸Ga]Ga-NOTA-folate (⁶⁸Ga-FOL). First, we determined the affinity of ⁶⁸Ga-FOL using human FR-β expressing cells. Then, we studied atherosclerotic mice with ⁶⁸Ga-FOL and ¹⁸F-FDG PET/CT. After sacrifice, the tissues excised were measured with a γ-counter for ex vivo biodistribution. Further, the tracer distribution and co-localization with macrophages in aorta cryosections were studied using autoradiography, hematoxylin-eosin staining and immunostaining with anti-Mac-3 antibody. Specificity of ⁶⁸Ga-FOL was assessed in a blocking study with excess of folate glucosamine. As a last step, human radiation doses were extrapolated from rat PET data. We were able to produce ⁶⁸Ga-FOL at high radioactivity concentration, with high molar activity and radiochemical purity. The cell binding studies showed high (5.1 ± 1.1 nM) affinity of ⁶⁸Ga-FOL to FR-β. The myocardial uptake of ⁶⁸Ga-FOL (SUV 0.43 ± 0.06) was 20-folds lower compared to ¹⁸F-FDG (SUV 10.6 ± 1.8, P = 0.001). The autoradiography and immunohistochemistry of aorta revealed that ⁶⁸Ga-FOL radioactivity co-localized with Mac-3-positive macrophage-rich atherosclerotic plaques. The plaque-to-healthy vessel wall ratio of ⁶⁸Ga-FOL (2.44 ± 0.15) was significantly higher than that of ¹⁸F-FDG (1.93 ± 0.22, P = 0.005). Blocking studies verified ⁶⁸Ga-FOL specificity to FR. As estimated from rat data the human effective dose was 0.0105 mSv/MBq. The organ with highest absorbed dose was kidney (0.1420 mSv/MBq). In conclusion, ⁶⁸Ga-FOL is a promising new FR-β-targeted tracer for imaging macrophage-associated inflammation.

中文翻译：

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叶酸受体β阳性巨噬细胞PET显像的[68Ga] Ga-NOTA-叶酸盐的放射合成和临床前评价

叶酸受体β（FR-β）是在巨噬细胞上表达的标志物之一，是炎症成像的有希望的靶标。在这里，我们报道[ ⁶⁸ Ga] Ga-NOTA-叶酸（⁶⁸ Ga-FOL）的放射合成和临床前评价。首先，我们使用表达人FR-β的细胞确定了⁶⁸ Ga-FOL的亲和力。然后，我们研究了⁶⁸ Ga-FOL和¹⁸ F-FDG PET / CT的动脉粥样硬化小鼠。处死后，用γ计数器测量切除的组织的离体生物分布。此外，使用放射自显影，苏木精-伊红染色和用抗Mac-3抗体免疫染色研究了主动脉冰冻切片中示踪剂的分布和与巨噬细胞的共定位。特异性⁶⁸Ga-FOL在一项封闭研究中评估了过量的叶酸葡萄糖胺。最后一步，从大鼠PET数据推断出人体辐射剂量。我们能够以高放射性浓度，高摩尔活性和放射化学纯度生产⁶⁸ Ga-FOL。细胞结合研究显示⁶⁸ Ga-FOL对FR-β具有高亲和力（5.1±1.1 nM）。与¹⁸ F-FDG（SUV 10.6±1.8，P = 0.001）相比，⁶⁸ Ga-FOL（SUV 0.43±0.06）的心肌摄取低20倍。主动脉的放射自显影和免疫组织化学分析显示⁶⁸ Ga-FOL放射性与富含Mac-3的巨噬细胞的动脉粥样硬化斑块共定位。斑块与健康血管壁之比为⁶⁸Ga-FOL（2.44±0.15）明显高于¹⁸ F-FDG（1.93±0.22，P = 0.005）。阻断研究证实了⁶⁸ Ga-FOL对FR的特异性。根据大鼠数据估计，人的有效剂量为0.0105 mSv / MBq。吸收剂量最高的器官是肾脏（0.1420 mSv / MBq）。总之，⁶⁸ Ga-FOL是一种有前途的新型FR-β靶向示踪剂，可用于对巨噬细胞相关的炎症进行成像。