Development of a Novel Positron Emission Tomography Probe Deuterated [¹⁸F]FE-TMP ([¹⁸F]FE-TMP‑<i>d</i>₄), an Antagonist of Escherichia coli Dihydrofolate Reductase, for Reporter Gene Imaging of the Brain

[18F]Fluoroethoxy trimethoprim ([18F]FE-TMP, [18F]2), a unique antagonist of bacterial dihydrofolate reductase (ecDHFR), has been developed as a reporter gene imaging agent. Here, we developed new PET probes based on TMP. Simulations predicted that hydrophobic interactions around the benzene ring of 2 contributed to binding with ecDHFR. The more lipophilic fluoropropyl-TMP analog (3) showed a higher binding affinity for ecDHFR than 2. However, 18F-labeled 3 ([18F]3) underwent 18F-defluorination during PET imaging of ecDHFR-transfected mice. Subsequently, we evaluated FE-d4 analog (5) as a candidate exhibiting greater in vivo stability than 2. Metabolite analysis showed a lower contamination of radiolabeled metabolites in mouse brains with 18F-labeled 5 ([18F]5) than [18F]2. PET imaging with [18F]5 of a non-human primate brain was characterized by a distinct signal/noise ratio, which allowed in vivo visualization of brain circuits expressing the reporter gene, demonstrating the superior potential of [18F]5 as a PET probe for reporter gene imaging of the brain.[18F]氟乙氧基甲氧苄啶（[18F]Fluoroethoxy trimethoprim，[18F]FE-TMP、[18F]2）是一种独特的细菌二氢叶酸还原酶（ecDHFR）拮抗剂，已被开发为报告基因显像剂。本研究基于甲氧苄啶（trimethoprim, TMP）开发了新型正电子发射断层显像（PET）探针。分子模拟结果显示，化合物2的苯环周围的疏水相互作用可促进其与ecDHFR的结合。亲脂性更强的氟丙基-TMP类似物（3）对ecDHFR的结合亲和力高于化合物2。然而，经18F标记的类似物3（[18F]3）在ecDHFR转染小鼠的PET显像过程中发生了18F脱氟反应。随后，我们评估了FE-d4类似物（5）作为候选探针的性能，其体内稳定性优于化合物2。代谢物分析结果表明，与[18F]2相比，经18F标记的类似物5（[18F]5）在小鼠脑中的放射性标记代谢物污染程度更低。利用[18F]5对非人灵长类大脑进行的PET显像具有清晰的信噪比，可实现表达报告基因的脑环路的体内可视化，证实了[18F]5作为脑报告基因显像PET探针的优异应用潜力。