Decristoforo Clemens, Faintuch-Linkowski Bluma, Rey Ana, von Guggenberg Elisabeth, Rupprich Marco, Hernandez-Gonzales Ignacio, Rodrigo Teodoro, Haubner Roland
Department of Nuclear Medicine, Medical University of Innsbruck, A-6020 Innsbruck, Austria.
Nucl Med Biol. 2006 Nov;33(8):945-52. doi: 10.1016/j.nucmedbio.2006.09.001.
There has been increasing interest in peptides containing the Arg-Gly-Asp (RGD) sequence for targeting of alpha(v)beta(3) integrins to image angiogenesis. [(18)F]Galacto-RGD has been successfully used for positron emission tomography applications in patients. Here we report on the preclinical characterization of a (99m)Tc-labeled derivative for single-photon emission computed tomography. c(RGDyK) was derivatized with HYNIC at the amino group of the lysine [c(RGDyK(HYNIC)) or HYNIC-RGD]. (99m)Tc labeling was performed using coligands (tricine and EDDA), as well as (99m)Tc(CO)(3)(H(2)O)(3). Radiolabeled peptides were characterized with regard to lipophilicity, protein binding and stability in buffer, serum and tissue homogenates. Integrin receptor activity was determined in internalization assays using alpha(v)beta(3)-receptor-positive M21 and alpha(v)beta(3)-receptor-negative M21L melanoma cells. Biodistribution was evaluated in normal and nude mice bearing M21, M21L and small cell lung tumors. HYNIC-RGD could be labeled at high specific activities using tricine, tricine-trisodium triphenylphosphine 3,3',3''-trisulfonate (TPPTS), tricine-nicotinic acid (NA) or EDDA as coligands. [(99m)Tc]EDDA/HYNIC-RGD, [(99m)Tc]tricine-TPPTS/HYNIC-RGD and [(99m)Tc]tricine-NA/HYNIC-RGD showed protein binding (<5%) considerably lower than (99m)Tc(3)/HYNIC-RGD and [(99m)Tc]tricine/HYNIC-RGD. [(99m)Tc]EDDA/HYNIC-RGD revealed high in vitro stability accompanied by low lipophilicity with a log P value of -3.56, comparable to that of [(18)F]Galacto-RGD. In M21 cells for this compound, the highest level of specific and rapid cell uptake (1.25% mg protein(-1)) was determined. In vivo, rapid renal excretion, low blood retention, low liver and muscle uptakes and low intestinal excretion 4 h postinjection were observed. Tumor uptake values were 2.73% ID/g in M21 alpha(v)beta(3)-receptor-positive tumors versus 0.85% ID/g in receptor-negative tumors 1 h postinjection. Small cell lung tumors could be visualized using gamma camera imaging. [(99m)Tc]EDDA/HYNIC-RGD shows encouraging properties to target alpha(v)beta(3) receptors in vivo with high stability and favorable pharmacokinetics. Tumor uptake studies showed specific targeting of alpha(v)beta(3)-receptor-positive tumors with tumor-to-organ ratios comparable to those of [(18)F]Galacto-RGD.
含有精氨酸 - 甘氨酸 - 天冬氨酸（RGD）序列的肽用于靶向α(v)β(3)整合素以成像血管生成，这方面的研究兴趣日益增加。[(18)F]半乳糖 - RGD已成功用于患者的正电子发射断层扫描应用。在此，我们报告一种用于单光子发射计算机断层扫描的(99m)Tc标记衍生物的临床前特性。c(RGDyK)在赖氨酸的氨基处用HYNIC进行衍生化[c(RGDyK(HYNIC))或HYNIC - RGD]。使用共配体（三羟甲基氨基甲烷和乙二胺二乙酸）以及(99m)Tc(CO)(3)(H(2)O)(3)进行(99m)Tc标记。对放射性标记的肽进行了亲脂性、蛋白质结合以及在缓冲液、血清和组织匀浆中的稳定性表征。在使用α(v)β(3)受体阳性的M21和α(v)β(3)受体阴性的M21L黑色素瘤细胞的内化试验中测定整合素受体活性。在正常小鼠以及荷有M21、M21L和小细胞肺癌肿瘤的裸鼠中评估生物分布。使用三羟甲基氨基甲烷、三羟甲基氨基甲烷 - 三苯基膦三磺酸钠（TPPTS）、三羟甲基氨基甲烷 - 烟酸（NA）或乙二胺二乙酸作为共配体，可高比活度标记HYNIC - RGD。[(99m)Tc]乙二胺二乙酸/HYNIC - RGD、[(99m)Tc]三羟甲基氨基甲烷 - TPPTS/HYNIC - RGD和[(99m)Tc]三羟甲基氨基甲烷 - NA/HYNIC - RGD显示出的蛋白质结合（<5%）明显低于(99m)Tc(3)/HYNIC - RGD和[(99m)Tc]三羟甲基氨基甲烷/HYNIC - RGD。[(99m)Tc]乙二胺二乙酸/HYNIC - RGD显示出高体外稳定性，亲脂性低，log P值为 - 3.56，与[(18)F]半乳糖 - RGD相当。对于该化合物在M21细胞中，测定出最高水平的特异性和快速细胞摄取（1.25% mg蛋白(-1)）。在体内，观察到注射后4小时快速经肾排泄、低血滞留、低肝和肌肉摄取以及低肠道排泄。注射后1小时，在M21α(v)β(3)受体阳性肿瘤中的肿瘤摄取值为2.73% ID/g，而在受体阴性肿瘤中为0.85% ID/g。使用γ相机成像可观察到小细胞肺癌肿瘤。[(99m)Tc]乙二胺二乙酸/HYNIC - RGD显示出令人鼓舞的特性，能够在体内靶向α(v)β(3)受体，具有高稳定性和良好的药代动力学。肿瘤摄取研究显示，α(v)β(3)受体阳性肿瘤的特异性靶向，肿瘤与器官的比值与[(18)F]半乳糖 - RGD相当。
