RADIOACTIVE LABELING OF BLOOD CELLS AND THEIR PRODUCTS

该项目的总体目标是改进收获方法 和放射性标记血细胞成分，以研究动力学和 标记细胞的组织分布，并探讨其诊断 回注后的效用。迄今为止，最大的临床影响， 放射性标记的白细胞成像已经被用于检测 标记的中性粒细胞的感染，以及标记的中性粒细胞的肿瘤定位。 淋巴细胞然而，目前的方法存在严重的技术问题。 困难包括标记细胞的放射性泄漏，以及 细胞核放射自分解造成的细胞损伤。 我们将继续开发高比活度放射性标记的 作为试剂的（氨基苯乙烯基）吡啶鎓（ASP）膜渗透染料， 为了标记混合的白细胞用于炎症部位的成像（131 I，129 I， 111 In或99 mTc）和用于标记体内放射敏感性淋巴细胞 追踪研究（131 I或111 In）。我们准备并使用了 放射性碘标记的膜渗透染料，以标记淋巴细胞和混合的 白细胞产量高。对后者的初步研究已经 在犬的诱导炎症部位表现出高度定位。 由于放射性核素仅限于细胞膜，标记染料 对淋巴细胞的放射性毒性比使用的放射性药物小 目前。通过测量大鼠T细胞迁移模式的变化， 淋巴细胞标记的放射性标记染料的剂量增加，我们将 确定与111 In-8-羟基喹啉相比放射毒性的降低。的 外周血淋巴细胞（PBL）和肿瘤浸润的能力 用新染料标记的淋巴细胞（TIL）定位于肿瘤中， 在Balb/c小鼠的EMT 6乳腺癌肉瘤肿瘤中进行测试。的 标记染料的放射毒性降低应允许更高剂量的 管理的活动，从而增加信息内容， 过继免疫治疗前的图像。 一种经典的荧光钙结合配体装载方法 将适于用99 mTc复合物标记白细胞。 我们将合成几种配体， （N2 S2）部分和可在细胞内水解的基团。的 这些新的中性99 mTcvO（N2 S2）配合物的总体亲脂性将 使它们扩散到白细胞中。 细胞内酯酶将 将所述络合物转化为新的带电99 mTcV（N2 S2）络合物，从而 将极性复合物捕获在细胞内。白细胞标记 新的药物将在麻醉的动物中进行测试， 诱发痛风和大肠杆菌通过诊断成像技术进行诊断。 用这些试剂标记的白细胞应该提供改善的图像， 从99 mTc-HMPAO标记的99 mTc泄漏的身体区域 白细胞使图像模糊。 我们将测量粒细胞集落刺激因子（G- CSF）对诱导部位放射性标记白细胞摄取的影响 炎症如果G-CSF增加中性粒细胞摄取， 炎症，该程序最终可用于检测 白细胞减少症患者感染。

The overall goal of this project is to improve the methods of harvesting and radiolabeling blood cellular components, to study the kinetics and tissue distribution of the labeled cells, and to explore their diagnostic utility after reinjection. To date, the greatest clinical impact of imaging radiolabeled leukocytes has been the detection of foci of infection by labeled neutrophils, and in tumor localization by labeled lymphocytes. However, current methods suffer from serious technical difficulties including radioactivity leaking out of labeled cells, and cell damage from autoradiolysis of cell nuclei. We will continue the development of high specific activity radiolabeled (aminostyryl)pyridinium (ASP) membrane permeant dyes as agents with which to label mixed leukocytes for imaging sites of inflammation (131I, 129I, 111In, or 99mTc) and for labeling radiosensitive lymphocytes for in vivo tracking studies (131I or 111In). We have prepared and used radioiodinated membrane permeant dyes to label both lymphocytes and mixed leukocytes in high yield. Preliminary studies with the latter have demonstrated high localization in induced sites of inflammation in dogs. Since radionuclides are restricted to the cell membrane, labeled dyes will be less radiotoxic to lymphocytes than the radiopharmaceuticals used currently. By measuring changes in the migration pattern of rat T- lymphocytes labeled with increasing doses of radiolabeled dyes, we will determine the reduction in radiotoxicity compared to 111In-oxine. The ability of peripheral blood lymphocytes (PBLs) and tumor infiltrating lymphocytes (TILs) labeled with the new dyes to localize in tumors will be tested in EMT6 mammary carcinosarcoma tumors in Balb/c mice. The decreased radiotoxicity of the labeled dyes should allow higher doses of administered activity and thereby increase the information content of the images prior to adoptive immunotherapy. A now classic approach for loading fluorescent calcium-binding ligands into cells will be adapted for labeling leukocytes with 99mTc complexes. We will synthesize several ligands containing both a diaminedisulfide (N2S2) moiety and groups which can be hydrolyzed intracellularly. The overall lipophilicity of these new neutral 99mTcvO(N2S2) complexes will allow them to diffuse into leukocytes. Intracellular esterases will then convert the complex to a new charged 99mTcV(N2S2) complex thereby trapping the polar complex within the cell. Leukocytes labeled with the new agents will be tested in anesthetized animals bearing sodium urate induced gout and E coli. abscesses by diagnostic imaging techniques. Leukocytes labeled with these agents should provide improved images in regions of the body where leakage of 99mTc from 99mTc-HMPAO labeled leukocytes obscures the image. We will measure the effect of granulocyte colony-stimulating factor (G- CSF) on the uptake of radiolabeled leukocytes in induced sites of inflammation. If G-CSF increases neutrophil uptake in sites of inflammation, the procedure could ultimately be used to detect sites of infection in patients with leukopenia.