CAAX Protein Processing and Cancer Therapy

CAAX 蛋白质加工和癌症治疗

• 批准号: 6945847
• 负责人: Stephen G. Young
• 金额: $ 28.74万
• 依托单位: UNIVERSITY OF CALIFORNIA LOS ANGELES
• 依托单位国家: 美国
• 财政年份: 2003
• 资助国家: 美国
• 起止时间: 2003-08-01 至 2008-05-31
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/6945847
• 关键词: alkyltransferase; antineoplastics; carcinogenesis inhibitor; cell growth regulation; enzyme inhibitors; fibroblasts; gene targeting; genetically modified animals; guanine nucleotide binding protein; isoprenoid; laboratory mouse; lamins; neoplastic transformation; posttranslational modifications; protein isoforms; protein metabolism; tissue /cell culture

DESCRIPTION (provided by applicant): Proteins that terminate with a "CAAX" sequence motif (e.g., the Ras proteins, Rho proteins, and nuclear lamins) undergo three sequential posttranslational modifications. First, the cysteine (C) is isoprenylated by enzymes in the cytosol (farnesyltransferase or geranylgeranyltransferase I). Second, the last three amino acids of the protein (i.e., the -AAX) are released by Rce1, an integral membrane endoprotease of the endoplasmic reticulum. Third, the newly exposed isoprenylcysteine is methylated by isoprenylcysteine carboxyl methyltransferase (Icmt), also an integral membrane protein of the endoplasmic reticulum. Each of these modifications is important for the targeting of CAAX proteins to membrane surfaces and for their proper function. For example, inhibitors of farnesyltransferase block the membrane targeting and downstream effects of mutationally activated Ras proteins and have shown promise in the treatment of certain cancers. A potential therapeutic drawback of the farnesyltransferase inhibitors is that K-Ras, the Ras isoform most commonly involved in human cancers, is geranylgeranylated by geranylgeranyltransferase I in the setting of farnesyltransferase inhibitors. The existence of this alternate prenylation pathway has raised interest in defining the physiologic effects of blocking the enzymes involved in the "post-isoprenylation" processing of CAAX proteins (Rce 1 and Icmt), as those enzymes act on both farnesylated and geranylgeranylated proteins. The goal of this project is to gain insights into the potential therapeutic utility of interfering with protein processing by Icmt and Reel, using cell culture systems and genetically modified mice. The effects of a deficiency in Fntb (the gene encoding the beta-subunit of protein farnesyltransferase) will also be assessed. Many useful reagents have already been generated for this project, including mice with knockout and conditional alleles of lcmt, Reel, and Ftnb. Preliminary studies have uncovered many exciting findings, including the fact that Icmt deficiency strikingly reduces oncogenic transformation of fibroblasts by K-Ras. Specific Aim 1 of this project is to assess the physiologic impact of Icmt deficiency, further defining its effects on Ras transforming activity. Specific Aim 2 is to generate cell culture and animal models for Fntb deficiency, and to compare the impacts of Fntb deficiency, Icmt deficiency, and Rcel deficiency. Specific Aim 3 is to determine if deficiencies in Icmt and Reel render cells more sensitive to farnesyltransferase inhibition. These studies should define the therapeutic potential of blocking the three different steps of CAAX protein processing.

描述（由申请人提供）：用“ CAAX”序列基序终止的蛋白质（例如RAS蛋白，RHO蛋白和核层粘蛋白）进行了三种顺序翻译后修饰。首先，半胱氨酸（C）被胞质溶液中的酶（Farneylsylansferase或geranylylgeranylylansylansylansylansferase I）化。其次，蛋白质的最后三个氨基酸（即-AAX）由内质网的整体膜内肠酶RCE1释放。第三，新暴露的异on基半胱氨酸是通过异on-半胱氨酸羧基甲基转移酶（ICMT）甲基化的，这也是内质网的整体膜蛋白。这些修饰中的每一个对于将CAAX蛋白靶向膜表面及其适当功能都很重要。例如，Farnesylansfrass的抑制剂阻止了突变活化的Ras蛋白的膜靶向和下游效应，并在治疗某些癌症的治疗方面表现出了希望。 Farnesylansylansferase抑制剂的潜在治疗缺点是，在Farnesylansyransfers抑制剂的情况下，K-RAS是人类癌症中最常见的RAS同工型，是通过黄烷基凝血酶Transylansys I的黄烷基凝集烷基化的。这种替代婚前化途径的存在引起了人们对定义CAAX蛋白（RCE 1和ICMT）加工中涉及的酶的生理影响的兴趣，因为这些酶对法尼化和金甘烷基凝集素化蛋白作用。该项目的目的是洞悉使用细胞培养系统和转基因小鼠干扰ICMT和Reel蛋白质加工的潜在治疗效用。还将评估FNTB缺乏的影响（编码蛋白质Farneylansylansferase的β-亚基的基因）。该项目已经生成了许多有用的试剂，包括具有敲除和有条件等位基因的小鼠，LCMT，卷轴和FTNB。初步研究发现了许多令人兴奋的发现，包括ICMT缺乏症可显着降低K-RAS对成纤维细胞的致癌性转化。 该项目的具体目的1是评估ICMT缺乏症的生理影响，进一步定义其对RAS转化活动的影响。具体目标2是生成细胞培养和动物模型，以使FNTB缺乏症，并比较FNTB缺乏症，ICMT缺乏症和RCEL缺乏症的影响。具体目的3是确定ICMT和Reel中的缺陷是否使细胞对Farneylsylansferase抑制更敏感。这些研究应定义阻断CAAX蛋白质加工三个不同步骤的治疗潜力。