CAAX Processing Enzymes as Anticancer Targets

描述（由申请人提供）：许多细胞内信号传导蛋白（例如，Ras和Rho蛋白）和几个核纤层蛋白终止于羧基末端CAAX基序。CAAX蛋白经历三个连续的翻译后修饰。首先，半胱氨酸（即，CAAX基序的C）被一对胞质酶法尼基转移酶（FTase）和香叶基香叶基转移酶I（GGT酶I）法尼基化或香叶基香叶基化。第二，最后三个氨基酸（即，-AAX）被Ras和α-因子转化酶（Rcel）（内质网（ER）的整合膜蛋白酶）切割掉。第三，新暴露的羧基末端异戊二烯半胱氨酸被另一种ER蛋白异戊二烯半胱氨酸羧基甲基转移酶（Icmt）甲基化。这些翻译后修饰使CAAX蛋白的C-末端更加疏水，增强蛋白质与膜表面的附着并促进某些蛋白质-蛋白质相互作用。 激活Ras突变已在30%的人类癌症中检测到，并且在白血病和骨髓增生性疾病中很常见。FTase抑制剂已用于治疗携带突变激活的Ras蛋白的癌症。不幸的是，K-Ras和N-Ras--最常与人类癌症有关的Ras亚型--在FTase抑制的情况下容易被GGT酶I香叶基香叶基化。这种替代的异戊二烯化途径将注意力集中在该途径中的其他酶上，如GGT酶I、Rcel和Icmt。令人惊讶的是，没有关于抑制这些其他酶对小鼠癌症发展的影响的数据。在这个项目中，这个空白将得到解决。 在初步研究中，已经产生了同时携带Cre诱导型潜在致癌Kras 2等位基因（KrasLsL）和诱导型Mx 1-Cre转基因的小鼠。在这些小鼠中诱导Cre激活了潜在的Ras等位基因，并导致完全成熟的、致命的骨髓增生性疾病，这让人联想到人类的慢性骨髓性白血病或青少年骨髓单核细胞白血病。最近，已经产生了翻译后加工酶（FTase、GGT酶I、Rcel和Icmt）的条件性“floxed”等位基因。因此，现在有可能培育Cre表达可用于同时激活潜在致癌K-Ras等位基因和抑制CAAX加工酶的小鼠。使用这些小鼠，我们将确定有缺陷的CAAX处理的影响，Ras诱导的骨髓增生性疾病的发展，进展和致死率。

DESCRIPTION (provided by applicant): Many intracellular signaling proteins (e.g., the Ras and Rho proteins) and several nuclear lamins terminate with a carboxyl-terminal CAAX motif. CAAX proteins undergo three sequential posttranslational modifications. First, the cysteine (i.e., the C of the CAAX motif) is farnesylated or geranylgeranylated by a pair of cytosolic enzymes--farnesyltransferase (FTase) and geranylgeranyltransferase I (GGTase I). Second, the last three amino acids (i.e., the -AAX) are cleaved off by Ras and a-factor converting enzyme (Rcel), an integral membrane protease of the endoplasmic reticulum (ER). Third, the newly exposed carboxyl-terminal isoprenylcysteine is methylated by another ER protein, isoprenylcysteine carboxyl methyltransferase (Icmt). These posttranslational modifications render the C-terminus of CAAX proteins more hydrophobic, enhancing the attachment of the proteins to membrane surfaces and facilitating certain protein-protein interactions. Activating Ras mutations have been detected in 30% of all human cancers, and are common in leukemia and myeloproliferative diseases. Inhibitors of FTase have been used to treat cancers that harbor mutationally activated Ras proteins. Unfortunately, K-Ras and N-Ras--the Ras isoforms most often implicated in human cancers--are readily geranylgeranylated by GGTase I in the setting FTase inhibition. This alternate isoprenylation pathway has focused attention on other enzymes in the pathway, such as GGTase I, Rcel, and Icmt. Surprisingly, there are no data on the impact of inhibiting these other enzymes on the development of cancer in mice. In this project, this void will be addressed. In preliminary studies, mice harboring both a Cre-inducible latent oncogenic Kras2 allele (KrasLsL) and the inducible Mx1-Cre transgene have been generated. Induction of Cre in those mice activates the latent Ras allele and results in a full-fledged, lethal, myeloproliferative disease that is reminiscent of chronic myelogenous leukemia or juvenile myelomonocytic leukemia in humans. Recently, conditional "floxed" alleles for the posttranslational processing enzymes (FTase, GGTase I, Rcel, and Icmt) have been generated. Thus, it is now possible to breed mice in which Cre expression can be used to simultaneously activate the latent oncogenic K-Ras allele and inactivate the CAAX processing enzymes. Using these mice, we will define the impact of defective CAAX processing on the development, progression, and lethality of Ras-induced myeloproliferative disease.