van der Wal Steffen, de Korne Clarize M, Sand Laurens G L, van Willigen Danny M, Hogendoorn Pancras C W, Szuhai Karoly, van Leeuwen Fijs W B, Buckle Tessa
Interventional Molecular Imaging Laboratory, Department of Radiology, Leiden University Medical Center, Albinusdreef 2, 2333 ZA, Leiden, The Netherlands.
Department of Pathology, Leiden University Medical Center, Albinusdreef 2, 2333 ZA, Leiden, The Netherlands.
Chembiochem. 2018 Jun 4;19(16):1758-65. doi: 10.1002/cbic.201800229.
The availability of a receptor for theranostic pretargeting approaches was assessed by use of a new click-chemistry-based deactivatable fluorescence-quenching concept. The efficacy was evaluated in a cell-based model system featuring both membranous (available) and internalized (unavailable) receptor fractions of the clinically relevant receptor chemokine receptor 4 (CXCR4). Proof of concept was achieved with a deactivatable tracer consisting of a CXCR4-specific peptide functionalized with a Cy5 dye bearing a chemoselective azide handle (N -Cy5-AcTZ14011). Treatment with a Cy7 quencher dye (Cy7-DBCO) resulted in optically silent Cy7-[click]-Cy5-AcTZ14011. In situ, a >90 % FRET-based reduction of the signal intensity of N -Cy5-AcTZ14011 [K =(222.4±25.2) nm] was seen within minutes after quencher addition. In cells, discrimination between the membranous and the internalized receptor fraction could be achieved through quantitative assessment of quenching/internalization kinetics. Similar evaluation of an activatable tracer variant based on the same targeting moiety (Cy5-S-S-Cy3-AcTZ14011) was unsuccessful in vitro. As such, using the described deactivatable approach to screen membrane receptors and their applicability in receptor-(pre-)targeted theranostics can become straightforward.
通过使用一种基于新型点击化学的可失活荧光猝灭概念，评估了用于诊疗预靶向方法的受体的可用性。在一个基于细胞的模型系统中评估了疗效，该系统具有临床相关受体趋化因子受体4（CXCR4）的膜性（可用）和内化（不可用）受体部分。通过一种可失活的示踪剂实现了概念验证，该示踪剂由用带有化学选择性叠氮化物手柄（N -Cy5-AcTZ14011）的Cy5染料功能化的CXCR4特异性肽组成。用Cy7猝灭染料（Cy7-DBCO）处理产生了光学上沉默的Cy7-[点击]-Cy5-AcTZ14011。在原位，加入猝灭剂后几分钟内，基于荧光共振能量转移（FRET）的N -Cy5-AcTZ14011信号强度降低了>90%[K =(222.4±25.2) nm]。在细胞中，通过对猝灭/内化动力学的定量评估，可以区分膜性和内化受体部分。基于相同靶向部分（Cy5-S-S-Cy3-AcTZ14011）的可激活示踪剂变体的类似评估在体外未成功。因此，使用所描述的可失活方法来筛选膜受体及其在受体（预）靶向诊疗中的适用性可以变得很直接。
