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Gene therapy for metastatic colon cancer in the liver by using CEA promoter

利用 CEA 启动子对肝脏转移性结肠癌进行基因治疗

基本信息

批准号：
11671140
负责人：
KONDO Satoshi
金额：
$ 2.24万
依托单位：
KOKKAIDO UNIVERSITY
依托单位国家：
日本
项目类别：
Grant-in-Aid for Scientific Research (C)
财政年份：
1999
资助国家：
日本
起止时间：
1999 至 2000
项目状态：
已结题

来源：
https://kaken.nii.ac.jp/grant/KAKENHI-PROJECT-11671140/
关键词：
Cancer gene therapy Colon cancer Liver metastasis CEA promoter N116Y Adenovirus vector Pancreatic cancer N116Y ベクター

项目摘要

As a first step, we established a liver metastasis model of human cancer cell line in nude mice. 1 x 10E6 of cancer cells were intrasplenically inoculated in nude mice. As we could not establish a liver metastasis model of colon cancer cell line, pancreatic cancer cell line was used as a substitute for the model.Subsequently, we constructed replication-deficient recombinant N116Y adenoviruses driven by the cytomegalovirus (CMV) promoter (Ad CMV-N116Y) and the carcinoembryonic antigen (CEA) promoter (Ad CEA-N116Y). A dominant negative H-ras mutant, N116Y, was derived from the v-H-ras oncogene by substituting asparagine with tyrosine at codon 116, which is in the GTP-binding domain. As a control vector, we constructed a beta-galactosidase (LacZ) expressing adenovirus vector driven by the CEA promoter (Ad CEA-LacZ).We examined the effect of Ad CEA-N116Y on the metastatic growth of cancer colonies in liver. Ad CEA-N116Y effectively reduced the number of metastatic colonies without any complication by injecting intrasplenically 5 days after tumor cell inoculation. The result of Ad CEA-LacZ injected mice showed that only cancer cells in liver were positively stained while the liver cells were negative. Thus, N116Y can selectively suppress the metastatic growth of human tumor cell by using the CEA promoter-driven adenovirus vector indicating that N116Y gene therapy may be potentially useful for the treatment of cancer patients with liver metastasis.

第一步，我们建立了人肝癌细胞株裸鼠肝转移模型。将I X IOE 6个癌细胞脾内接种于裸鼠中。由于无法建立结肠癌细胞系肝转移模型，我们以胰腺癌细胞系作为模型的替代物，构建了由巨细胞病毒（CMV）启动子和癌胚抗原（CEA）启动子驱动的复制缺陷型重组N116 Y腺病毒（Ad CMV-N116 Y）。N116 Y是一种显性失活的H-ras突变体，它是由v-H-ras癌基因的第116位密码子（位于GTP结合区）的天冬酰胺被酪氨酸取代而得。作为对照载体，我们构建了CEA启动子驱动的β-半乳糖苷酶（LacZ）表达腺病毒载体（Ad CEA-LacZ），观察了Ad CEA-N116 Y对肝癌转移生长的影响。在肿瘤细胞接种后5天，通过脾内注射Ad CEA-N116 Y有效地减少了转移集落的数量，而没有任何并发症。AdCEA-LacZ注射小鼠的结果显示，只有肝脏中的癌细胞呈阳性染色，而肝细胞呈阴性。因此，N116 Y可以通过使用CEA启动子驱动的腺病毒载体选择性地抑制人肿瘤细胞的转移生长，表明N116 Y基因治疗可能用于治疗具有肝转移的癌症患者。