背景:通过N-琥珀酰亚胺基-4-[18F]氟苯甲酸酯([18F]SFB)的单结构域抗体(sdAb)的放射性氟化已被证明是开发基于sdAb的PET示踪剂的有前景的策略。在假体组(PG)[18F]SFB生产自动化的同时,已成功报道,没有报道大规模sdAb标记的实用方法。因此,我们优化并自动化了PG生产,使随后有效的手动缀合反应与抗成纤维细胞活化蛋白(FAP)-αsdAb(4AH29)和抗叶酸受体(FR)-αsdAb(2BD42)。FAP的α同种型和FR都是确定的肿瘤标志物。已知FAP-α主要由乳腺癌相关成纤维细胞过度表达,卵巢,和其他癌症,而其在正常组织中的表达较低或检测不到。FR-α在上皮癌中表达升高,如卵巢,脑癌和肺癌.非侵入性成像技术,例如PET成像,使用针对特定肿瘤标志物的示踪剂可以提供肿瘤及其环境的分子信息,诊断的助手,癌症治疗的治疗选择和评估。
结果:[18F]使用全自动三步合成SFB,一锅反应。总操作时间为54分钟,得到[18F]SFB,RCP>90%,RCYd.c.为44±4%(n=13)。纯化后的手动缀合反应产生RCP>95%的[18F]FB-sdAb,合成结束活性>600MBq和表观摩尔活性>10GBq/μmol。总体RCYd.c.,校正为QMA上的[18F]F-陷阱,[18F]FB-2BD42和[18F]FB-4AH29分别为9%(n=1)和5±2%(n=3)。
结论:[18F]SFB合成在TrasisAllInOne模块上成功实现了自动化和升级。抗hFAP-α和抗hFR-αsdAb是放射性氟化的,产生相似的RCYsd.c.和RCPs,显示了这种方法作为sdAb通用放射性氟化策略的潜力。放射性氟化sdAb显示出有利的生物分布模式,并且对于进一步表征FAP-α和FR-α成像的新PET示踪剂具有吸引力。
BACKGROUND: Radiofluorination of single domain antibodies (sdAbs) via N-succinimidyl-4-[18F]fluorobenzoate ([18F]SFB) has shown to be a promising strategy in the development of sdAb-based PET tracers. While automation of the prosthetic group (PG) [18F]SFB production, has been successfully reported, no practical method for large scale sdAb labelling has been reported. Therefore, we optimized and automated the PG production, enabling a subsequently efficient manual conjugation reaction to an anti-fibroblast activation protein (FAP)-α sdAb (4AH29) and an anti-folate receptor (FR)-α sdAb (2BD42). Both the alpha isoform of FAP and the FR are established tumour markers. FAP-α is known to be overexpressed mainly by cancer-associated fibroblasts in breast, ovarian, and other cancers, while its expression in normal tissues is low or undetectable. FR-α has an elevated expression in epithelial cancers, such as ovarian, brain and lung cancers. Non-invasive imaging techniques, such as PET-imaging, using tracers targeting specific tumour markers can provide molecular information over both the tumour and its environment, which aides in the diagnosis, therapy selection and assessment of the cancer treatment.
RESULTS: [18F]SFB was synthesized using a fully automated three-step, one-pot reaction. The total procedure time was 54 min and results in [18F]SFB with a RCP > 90% and a RCY d.c. of 44 ± 4% (n = 13). The manual conjugation reaction after purification produced [18F]FB-sdAbs with a RCP > 95%, an end of synthesis activity > 600 MBq and an apparent molar activity > 10 GBq/µmol. Overall RCY d.c., corrected to the trapping of [18F]F- on the QMA, were 9% (n = 1) and 5 ± 2% (n = 3) for [18F]FB-2BD42 and [18F]FB-4AH29, respectively.
CONCLUSIONS: [18F]SFB synthesis was successfully automated and upscaled on a Trasis AllInOne module. The anti-hFAP-α and anti-hFR-α sdAbs were radiofluorinated, yielding similar RCYs d.c. and RCPs, showing the potential of this method as a generic radiofluorination strategy for sdAbs. The radiofluorinated sdAbs showed a favourable biodistribution pattern and are attractive for further characterization as new PET tracers for FAP-α and FR-α imaging.