Purpose: This study was motivated by the need for better positron emission tomography (PET)-compatible tools to image bacterial infection. Our previous efforts have targeted bacteria-specific metabolism via assimilation of carbon-11 labeled d-amino acids into the bacterial cell wall. Since the chemical determinants of this incorporation are not fully understood, we sought a high-throughput method to label d-amino acid derived structures with fluorine-18. Our strategy employed a chemical biology approach, whereby an azide (-N₃) bearing d-amino acid is incorporated into peptidoglycan muropeptides, with subsequent “click” cycloaddition with an ¹⁸F-labeled strained cyclooctyne partner. Procedures: A water-soluble, ¹⁸F-labeled and dibenzocyclooctyne (DBCO)-derived radiotracer ([¹⁸F]FB-sulfo-DBCO) was synthesized. This tracer was incubated with pathogenic bacteria treated with azide-bearing d-amino acids, and incorporated ¹⁸F was determined via gamma counting. In vitro uptake in bacteria previously treated with azide-modified d-amino acids was compared to that in cultures treated with amino acid controls. The biodistribution of [¹⁸F]FB-sulfo-DBCO was studied in a cohort of healthy mice with implications for future in vivo imaging. Results: The new strain-promoted azide–alkyne cycloaddition (SPAAC) radiotracer [¹⁸F]FB-sulfo-DBCO was synthesized with high radiochemical yield and purity via N-succinimidyl 4-[¹⁸F]fluorobenzoate ([¹⁸F]SFB). Accumulation of [¹⁸F]FB-sulfo-DBCO was significantly higher in several bacteria treated with azide-modified d-amino acids than in controls; for example, we observed 7 times greater [¹⁸F]FB-sulfo-DBCO ligation in Staphylococcus aureus cultures incubated with 3-azido-d-alanine versus those incubated with d-alanine. Conclusions: The SPAAC radiotracer [¹⁸F]FB-sulfo-DBCO was validated in vitro via metabolic labeling of azide-bearing peptidoglycan muropeptides. d-Amino acid-derived PET radiotracers may be more efficiently screened via [¹⁸F]FB-sulfo-DBCO modification.

中文翻译：

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使用 [18F]FB-sulfo-DBCO 对叠氮化物修饰细菌进行生物正交放射性标记

目的：这项研究的动机是需要更好的正电子发射断层扫描 (PET) 兼容工具来成像细菌感染。我们之前的努力通过将碳 11 标记的d-氨基酸同化到细菌细胞壁中来针对细菌特异性代谢。由于这种掺入的化学决定因素尚未完全了解，我们寻求一种高通量方法来用氟 18标记d-氨基酸衍生结构。我们的策略采用了化学生物学方法，其中将带有d-氨基酸的叠氮化物 (-N ₃ )掺入肽聚糖胞肽中，随后与¹⁸ F 标记的应变环辛炔伴侣进行“点击”环加成。程序：合成了水溶性、¹⁸ F 标记的二苯并环辛炔 (DBCO) 衍生放射性示踪剂 ([ ¹⁸ F]FB-磺基-DBCO)。将该示踪剂与用含叠氮化物的d-氨基酸处理的病原菌一起孵育，并通过伽马计数确定掺入的¹⁸ F。将先前用叠氮化物修饰的d-氨基酸处理的细菌的体外摄取与用氨基酸对照处理的培养物中的体外摄取进行比较。在一组健康小鼠中研究了[ ¹⁸ F]FB-sulfo-DBCO的生物分布，这对未来的体内成像具有重要意义。结果：通过N-琥珀酰亚胺基4-[ ¹⁸ F]氟苯甲酸酯（[ ¹⁸ F]SFB）以高放射化学产率和纯度合成了新型菌株促进的叠氮-炔环加成（SPAAC）放射性示踪剂[ ¹⁸ F]FB-磺基-DBCO 。在用叠氮化物修饰的d-氨基酸处理的几种细菌中，[ ¹⁸ F]FB-sulfo-DBCO的积累显着高于对照组；例如，我们观察到，与 3-叠氮基-d-丙氨酸孵育的金黄色葡萄球菌培养物相比，与 d-丙氨酸孵育的金黄色葡萄球菌培养物中，[ 18 ^F ]FB-磺基-DBCO 连接增加了 7 倍。结论： SPAAC 放射性示踪剂 [ ¹⁸ F]FB-sulfo-DBCO通过叠氮化物肽聚糖胞肽的代谢标记在体外得到验证。通过[ ¹⁸ F]FB-磺基-DBCO 修饰可以更有效地筛选d-氨基酸衍生的 PET 放射性示踪剂。