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In Vivo Diagnosis of Gene Localization and Expression by Nuclear Medicine Modality

通过核医学方式进行基因定位和表达的体内诊断

基本信息

批准号：
13670965
负责人：
NAKAMURA Kayoko
金额：
$ 2.56万
依托单位：
Keio University
依托单位国家：
日本
项目类别：
Grant-in-Aid for Scientific Research (C)
财政年份：
2001
资助国家：
日本
起止时间：
2001 至 2003
项目状态：
已结题

来源：
https://kaken.nii.ac.jp/grant/KAKENHI-PROJECT-13670965/
关键词：
Oligonucleotides Tc-99m Antisense mdr Gene imaging Multidrug resistance P-glycoprotein P-糖タンパク質

项目摘要

The aim of this investigation is to develop nuclear medicine techniques for non-invasive diagnosis of gene localization and expression. Gene here means the one injected for gene therapy or the other over-expressed due to the disease. Using human epidermoid tumor cell line ; KB-31 and its mdr-1 transfected cell line ; KB-G2 carried out the study. By co-working with Dr. Donald J Hnatowich, Professor of Massachusetts Medical School, the following results were obtained;1. Antisense (AS) and sense (S) oligonucleotides related to mdr-1 were synthesized and modified (all phosphothiolated) and then, labeled with Tc-99m via MAG3 or HYNIC ligands. Tc-99m-AS and Tc-99m-S were obtained with high specific activity without loosing their antisense-binding affinity.2. KB-31 or KB-G2 were treated with AS or S, followed by incubation with Tc-99m-MIBI, which is to evaluate P-glycoprotein (mdr-1 expression) function. The KB-G2 treated with AS suppressed the P-gp function, while it was not shown in KB-G2 treated by S. KB-31 did not show any influence by treating with AS or S. These results indicated the antisense effect of AS oligonucleiotied we used.3. Tc-99m-AS was accumulated in KB-G2 but not in KB-31, while Tc-99m-S was not taken up either in KB-G2 or KB-31. These results indicated that AS was accumulated into the cell by antisense effect.4. Tc-99m-AS or Tc-99m-S was injected into the mice bearing KB-G2 or KB-31. The uptake of Tc-99m-AS in the KB-G2 tumor was significantly higher than that in the KB-31 tumors. Tc-99m-AS uptake in KB-G2 tumor was significantly higher than Tc-99m-S. These results indicated that Tc-99m-AS was localized into the mdr-1-tumors specifically by antisense erect.All these results would lead the success for the antisense imaging.

本研究的目的是发展核医学技术，用于基因定位和表达的非侵入性诊断。这里的基因是指为基因治疗而注射的基因或由于疾病而过度表达的基因。采用人表皮样瘤细胞株KB-31及其转染mdr-1的细胞株KB-G2进行研究。通过与马萨诸塞州医学院教授Donald J Hnatowich博士的合作，获得了以下结果：1。合成并修饰与mdr-1相关的反义（AS）和正义（S）寡核苷酸（全部硫代磷酸化），然后通过MAG 3或HYNIC配体用Tc-99 m标记。获得了高比活性的Tc-99 m-AS和Tc-99 m-S，且不损失其反义结合亲和力. KB-31或KB-G2用AS或S处理，然后用Tc-99 m-MIBI孵育，这是为了评估P-糖蛋白（mdr-1表达）功能。AS对KB-G2细胞P-gp功能有抑制作用，而S对KB-G2细胞P-gp功能无抑制作用。KB-31不受AS或S处理的影响。这些结果表明我们所用的反义AS寡核苷酸具有一定的效果. Tc-99 m-AS在KB-G2中蓄积，而在KB-31中不蓄积，而Tc-99 m-S在KB-G2和KB-31中均不摄取。说明AS是通过反义作用进入细胞的.将Tc-99 m-AS或Tc-99 m-S分别注射于KB-G2或KB-31荷瘤小鼠。KB-G2肿瘤对~（99）Tc-AS的摄取明显高于KB-31肿瘤。KB-G2肿瘤对~（99）Tc-AS的摄取明显高于~（99）Tc-S。以上结果表明，Tc-99 m-AS在mdr-1肿瘤中具有特异性的反义显像定位，为反义显像的成功奠定了基础。