RAS ONCOGENE P21 FARNESYL TRANSFERASE--NOVEL ANTICANCE

RAS癌基因P21法呢基转移酶--新型抗癌药物

基本信息

批准号：
3200379
负责人：
SAID M SEBTI
金额：
$ 16.8万
依托单位：
UNIVERSITY OF PITTSBURGH AT PITTSBURGH
依托单位国家：
美国
项目类别：
财政年份：
1992
资助国家：
美国
起止时间：
1992-05-01 至 1995-04-30
项目状态：
已结题

来源：
https://reporter.nih.gov/project-details/3200379
关键词：
antineoplastics athymic mouse bioassay farnesyl compound guanine nucleotide binding protein inhibitor /antagonist isozymes lipogenesis inhibitor lovastatin neoplastic growth nonhuman therapy evaluation oncogenes protein purification pyrophosphates synthetic protein tissue /cell culture transferase

项目摘要

Ras oncogenes are the most frequently identified oncogenes in human cancers. The ras oncogene product, p2lras, is post-translationary modified by the enzyme p2lras farnesyl:protein transferase which transfers farnesyl from farnesyl pyrophosphate, a cholesterol biosynthesis intermediate, to cysteine of the p21ras carboxy terminal CAAX (C=cysteine, A=aliphatic amino acid, X=any amino acid). This farnesylation leads to p2lras- membrane association which is an absolute requirement for p2lras- transforming activity in cultured cells. The overall goal of this proposal is to enhance our understanding of the importance of p2lras - membrane association in ras oncogene neoplasia in vivo and to design strategies to inhibit human tumor growth based on interference with the biochemical steps involved in p2lras membrane association. This will be accomplished by: 1) evaluating the antitumor activity of inhibitors of p2lras membrane association such as lovastatin and other cholesterol biosynthesis inhibitors in nude mice implanted with human tumors expressing various forms of ras oncogene, 2) purifying p2lras farnesyl:protein transferases from human tumors, 3) synthesizing novel analogues of farnesyl pyrophosphate, CAAX tetrapeptide derivatives as well as farnesyl pyrophosphate/CAAX transition state analogues and suicide inhibitors, and 4) determining the ability of these analogues to inhibit p2lras farnesyl:protein transferase, membrane association and human tumor growth in nude mice. The antitumor studies will be carried out in nude mice bearing human tumors expressing various forms of ras oncogenes. The p2lras farnesyl:protein transferases will be purified from human tumor cells by a recently published method using p2lras carboxy terminal CAAX peptide affinity chromatography. The synthetic analogues will first be screened based on their ability to inhibit the purified p2l- farnesyl:protein transferases. The selected inhibitors will then be tested for their ability to inhibit p2lras membrane association, human tumor cell growth in culture and in vivo in nude mice. The membrane association assay will be carried out by immunoprecipitation and western blotting of cytosolic and membrane-associated p2lras. This proposal seeks to broaden the spectrum of human tumors that can be treated chemotherapeutically by evaluating the antitumor activity of inhibitors of p2lras membrane association. The development of p2lras farnesyl:protein transferase assay has great potential as a powerful screening bioassay for the discovery of novel anticancer drugs that inhibit ras oncogene function.

RAS癌基因是人类中最常见的癌基因癌症。 RAS癌基因产品P2LRAS是翻译后修改的通过酶P2Lras farnesyl：蛋白质转移酶，转移Farnesyl 从胆固醇生物合成中间体的Farnesyl Pyrophophate到 p21ras羧基终端CAAX的半胱氨酸（C =半胱氨酸，A =脂肪族氨基酸，x =任何氨基酸）。这种法尼化导致P2lras-膜关联是P2LRAS转化的绝对要求培养细胞的活性。该提议的总体目标是增强我们对P2LRAS-膜重要性的理解 RAS癌基因肿瘤的关联，并设计策略基于干扰生化步骤抑制人类肿瘤的生长参与P2LRAS膜协会。这将通过：1）评估P2LRAS膜抑制剂的抗肿瘤活性洛伐他汀和其他胆固醇生物合成等关联植入人类肿瘤的裸小鼠的抑制剂，表达各种 RAS癌基因的形式，2）纯化P2Lras Farnesyl：蛋白质转移酶来自人类肿瘤，3）合成Farnesyl的新型类似物焦磷酸盐，CAAX四肽衍生物以及Farnesyl 焦磷酸/CAAX过渡状态类似物和自杀抑制剂，以及 4）确定这些类似物抑制P2LRA的能力 Farnesyl：蛋白质转移酶，膜关联和人类肿瘤生长在裸鼠。抗肿瘤研究将在裸鼠中进行表达各种形式的RAS癌基因的人类肿瘤。 p2lras Farnesyl：蛋白质转移酶将通过A从人类肿瘤细胞中纯化最近使用P2LRAS羧基终端CAAX肽发表的方法亲和色谱。合成类似物将首先筛选基于它们抑制纯化的P2L-法尼的能力：蛋白质转移酶。然后，将对选定的抑制剂进行测试能够抑制P2LRAS膜关联，人类肿瘤细胞生长的能力裸体小鼠的培养和体内。膜协会测定将是通过免疫沉淀和胞质和蛋白质印迹进行膜相关的P2LRA。该建议旨在扩大可以通过评估可以对化学治疗进行化学治疗的人类肿瘤 P2LRAS膜关联抑制剂的抗肿瘤活性。这 P2Lras Farnesyl的开发：蛋白质转移酶测定法潜在的筛选生物测定，以发现新颖抑制RAS癌基因功能的抗癌药物。