Vogt A, Qian Y, Blaskovich M A, Fossum R D, Hamilton A D, Sebti S M
Department of Pharmacology, School of Medicine, University of Pittsburgh, Pennsylvania 15261.
J Biol Chem. 1995 Jan 13;270(2):660-4. doi: 10.1074/jbc.270.2.660.
Cysteine farnesylation of the carboxyl-terminal tetrapeptide CAAX (C = Cys, A = Leu, Ile, or Val, X = Met or Ser) of the oncogene product Ras is required for its malignant transformation activity. As a consequence farnesyltransferase (FTase), the enzyme responsible for this lipid modification, has become one of the most sought-after targets for anticancer drug development. We have recently designed peptide mimics of the COOH-terminal Cys-Val-Ile-Met of KB-Ras where the dipeptide Val-Ile was replaced by aminobenzoic acid derivatives. Although these peptidomimetics are potent inhibitors of FTase in vitro, they retain several undesirable peptide features that hamper their use in vivo. We report here the design, synthesis, and biological activity of the first non-peptide mimetics of CAAX where the tripeptide AAX was replaced by biphenyl derivatives. (R)-4-[N-(3-mercapto-2-aminopropyl)]amino-3'- carboxybiphenyl, where the cysteine is linked to the biphenyl derivative through a secondary amine, contains no amino acids, lacks peptidic features, and has no hydrolyzable bonds. This peptidomimetic is a potent inhibitor of FTase in vitro (IC50 = 50-150 nM) and disrupts Ras processing in whole cells. Furthermore, this non-peptide mimetic of CAAX is highly selective for FTase (666-fold) relative to the closely related geranylgeranyltransferase I. This selectivity is also respected in vivo since the processing of Ras but not the geranylgeranylated Rap1A was disrupted in whole cells. Structure activity relationship studies revealed that FTase recognition and inhibitory potency of CAAX peptidomimetics require free thiol and carboxylate groups separated by a hydrophobic moiety, and that precise positioning of these functional groups must correspond to that of the parent CAAX. The true CAAX peptidomimetic described in this manuscript has several desirable features for further development as a potential anticancer agent. It is not metabolically inactivated by FTase, does not require a pro-drug strategy for inhibition in vivo, and is selective for farnesylation relative to geranylgeranylation.
癌基因产物Ras的羧基末端四肽CAAX（C = 半胱氨酸，A = 亮氨酸、异亮氨酸或缬氨酸，X = 甲硫氨酸或丝氨酸）的半胱氨酸法尼基化是其恶性转化活性所必需的。因此，负责这种脂质修饰的酶法尼基转移酶（FTase）已成为抗癌药物开发中最受追捧的靶点之一。我们最近设计了KB-Ras羧基末端半胱氨酸-缬氨酸-异亮氨酸-甲硫氨酸的肽模拟物，其中二肽缬氨酸-异亮氨酸被氨基苯甲酸衍生物取代。尽管这些拟肽在体外是有效的FTase抑制剂，但它们保留了几个不利于其体内应用的肽特性。我们在此报告了首批CAAX非肽模拟物的设计、合成及生物活性，其中三肽AAX被联苯衍生物取代。（R）-4-[N-（3-巯基-2-氨基丙基）]氨基-3'-羧基联苯，其中半胱氨酸通过仲胺与联苯衍生物相连，不含氨基酸，没有肽的特性，也没有可水解的键。这种拟肽在体外是有效的FTase抑制剂（IC50 = 50 - 150 nM），并能破坏全细胞中的Ras加工过程。此外，这种CAAX非肽模拟物相对于密切相关的香叶基香叶基转移酶I对FTase具有高度选择性（666倍）。由于在全细胞中Ras的加工过程被破坏而香叶基香叶基化的Rap1A没有被破坏，这种选择性在体内也得到了体现。构效关系研究表明，CAAX拟肽对FTase的识别和抑制效力需要被疏水部分隔开的游离巯基和羧基，并且这些官能团的精确定位必须与母体CAAX的定位一致。本手稿中描述的真正的CAAX拟肽具有作为潜在抗癌药物进一步开发的几个理想特性。它不会被FTase代谢失活，在体内抑制不需要前药策略，并且相对于香叶基香叶基化对法尼基化具有选择性。
