McConnell Brain Imaging Centre, McGill University, Montreal, Canada.
Mol Imaging Biol. 2011 Apr;13(2):332-41. doi: 10.1007/s11307-010-0329-6.
The transferrin receptor (TfR) is one of the most attractive targets to overcome the blood-brain barrier (BBB). It has recently been shown that THRPPMWSPVWP binds to the TfR and is subsequently internalized into TfR-expressing cells. Here, we evaluated the ability of THRPPMWSPVWP to become internalized into human TfR-expressing cells via endocytosis to determine its potential to act as a carrier system for the transport of small molecules across the BBB.
To validate the underlying concept of a conjugate consisting of a small brain imaging tracer and a large peptidic carrier molecule, a conjugate of the high affinity D2 receptor ligand fallypride and the TfR targeting peptide THRPPMWSPVWP has been synthesized. Furthermore, two derivatives of THRPPMWSPVWP were labeled with (68)Ga in high radiochemical yields (>96%) and a radiochemical purity of 96-98% and evaluated in vitro and in vivo.
The fallypride-THRPPMWSPVWP conjugate still displayed a K(i) of 27 nM. The uptake of the (68)Ga-labeled peptides into TfR-bearing cells was investigated using U87MG and HT-29 cells to assess the capability of the peptide to act as a carrier molecule targeting the TfR. The in vitro binding studies revealed negligible uptake of the tested (68)Ga-labeled conjugates ranging from 0.08% to 0.66% after 60 min incubation at 37°C. Initial in vivo experiments with (68)Ga-DOTA-S-maleimido-THRPPMWSPVWP in two healthy rats showed a mean brain uptake of 0.037% injected dose per gram, confirming the results obtained in vitro.
These results suggest that the accumulation of the (68)Ga-radiolabeled conjugates of the TfR-binding peptide THRPPMWSPVWP into TFR expressing human cell lines is nonspecific and too low to render this peptide suitable as a possible carrier molecule for a receptor-mediated transport of compounds across the BBB.
转铁蛋白受体（TfR）是克服血脑屏障（BBB）的最有吸引力的靶标之一。最近已经表明，THRPPMWSPVWP 与 TfR 结合，并随后被内化到表达 TfR 的细胞中。在这里，我们评估了 THRPPMWSPVWP 通过内吞作用进入人 TfR 表达细胞的能力，以确定其作为小分子跨 BBB 转运载体系统的潜力。
为了验证由小分子脑成像示踪剂和大肽载体分子组成的缀合物的基本概念，已经合成了高亲和力 D2 受体配体 fallypride 和 TfR 靶向肽 THRPPMWSPVWP 的缀合物。此外，两种 THRPPMWSPVWP 的衍生物用（68）Ga 标记，放射化学产率（>96%）和放射化学纯度（96-98%）高，并在体外和体内进行了评估。
fallypride-THRPPMWSPVWP 缀合物仍显示出 K(i)为 27 nM。使用 U87MG 和 HT-29 细胞研究（68）Ga 标记肽进入 TfR 携带细胞的摄取，以评估该肽作为靶向 TfR 的载体分子的能力。体外结合研究表明，在 37°C 孵育 60 分钟后，测试的（68）Ga 标记缀合物的摄取率低至 0.08%至 0.66%。在两名健康大鼠中进行的初步体内实验显示，（68）Ga-DOTA-S-马来酰亚胺-THRPPMWSPVWP 的脑摄取平均值为每克 0.037%注射剂量，证实了体外获得的结果。
这些结果表明，TfR 结合肽 THRPPMWSPVWP 的（68）Ga 放射性标记缀合物在表达 TfR 的人细胞系中的积累是非特异性的，太低而不能使该肽适合作为跨 BBB 进行化合物受体介导转运的可能载体分子。
