Knox R J, Jenkins T C, Hobbs S M, Chen S, Melton R G, Burke P J
Enact Pharma Plc, Salisbury, United Kingdom.
Cancer Res. 2000 Aug 1;60(15):4179-86.
A novel prodrug activation system, endogenous in human tumor cells, is described. A latent enzyme-prodrug system is switched on by a simple synthetic, small molecule co-substrate. This ternary system is inactive if any one of the components is absent. CB 1954 [5-(aziridin-1-yl)-2,4-dinitrobenzamide] is an antitumor prodrug that is activated in certain rat tumors via its 4-hydroxylamine derivative to a potent bifunctional alkylating agent. However, human tumor cells are resistant to CB 1954 because they are unable to catalyze this bioactivation efficiently. A human enzyme has been discovered that can activate CB 1954, and it has been shown to be commonly present in human tumor cells. The enzyme is NQO2 [NAD(P)H quinone oxidoreductase 2], but its activity is normally latent, and a nonbiogenic co-substrate such as NRH [nicotinamide riboside (reduced)] is required for enzymatic activity. There is a very large (100-3000-fold) increase in CB 1954 cytotoxicity toward either NQO2-transfected rodent or nontransfected human tumor cell lines in the presence of NRH. Other reduced pyridinium compounds can also act as co-substrates for NQO2. Thus, the simplest quaternary salt of nicotinamide, 1-methyl-3-carboxamidopyridinium iodide, was a co-substrate for NQO2 when reduced to the corresponding 1,4-dihydropyridine derivative. Increased chain length and/or alkyl load at the 1-position of the dihydropyridine ring improved specific activity, and compounds more active than NRH were found. However, little activity was seen with either the 1-benzyl or 1-(2-phenylethyl) derivatives. A negatively charged substituent at the 3-position of the reduced pyridine ring also negated the ability of these compounds to act as cosubstrates for NQO2. In particular, 1-carbamoylmethyl-3-carbamoyl-1,4dihydropyridine was shown to be a co-substrate for NQO2 with greater stability than NRH, with the ability to enter cells and potentiate the cytotoxicity of CB 1954. Furthermore, this agent is synthetically accessible and suitable for further pharmaceutical development. NQO2 activity appears to be related to expression of NQO1 (DT-diaphorase), an enzyme that is known to have a favorable distribution toward certain human cancers. NQO2 is a novel target for prodrug therapy and has a unique activation mechanism that relies on a synthetic co-substrate to activate an apparently latent enzyme. Our findings may reopen the use of CB 1954 for the direct therapy of human malignant disease.
本文描述了一种存在于人类肿瘤细胞内的新型前药激活系统。一种潜在的酶-前药系统可通过一种简单的合成小分子共底物激活。如果该三元系统中任何一个组分缺失，则其无活性。CB 1954 [5-(氮丙啶-1-基)-2,4-二硝基苯甲酰胺] 是一种抗肿瘤前药，在某些大鼠肿瘤中可通过其4-羟胺衍生物激活为一种强效双功能烷化剂。然而，人类肿瘤细胞对CB 1954具有抗性，因为它们无法有效地催化这种生物激活过程。现已发现一种人类酶可激活CB 1954，并且已证明其在人类肿瘤细胞中普遍存在。该酶为NQO2 [NAD(P)H醌氧化还原酶2]，但其活性通常处于潜伏状态，酶活性需要非生物源共底物如NRH [烟酰胺核糖（还原型）]。在存在NRH的情况下，CB 1954对NQO2转染的啮齿动物或未转染的人类肿瘤细胞系的细胞毒性会有非常大（100 - 3000倍）的增加。其他还原型吡啶化合物也可作为NQO2的共底物。因此烟酰胺的最简单季铵盐，即1-甲基-3-甲酰胺基吡啶碘化物，在还原为相应的1,4-二氢吡啶衍生物时是NQO2的共底物。二氢吡啶环1位链长度增加和/或烷基负载增加可提高比活性，并且发现了比NRH活性更高的化合物。然而，1-苄基或1-(2-苯乙基)衍生物几乎没有活性。还原吡啶环3位的带负电荷取代基也会使这些化合物失去作为NQO2共底物的能力。特别是，1-氨甲酰甲基-3-氨甲酰基-1,4-二氢吡啶被证明是NQO2的共底物，其稳定性高于NRH，能够进入细胞并增强CB 1954的细胞毒性。此外，该试剂可通过合成获得，适合进一步的药物开发。NQO2活性似乎与NQO1（DT-黄递酶）的表达有关，NQO1是一种已知在某些人类癌症中分布良好的酶。NQO2是前药治疗的一个新靶点，具有独特的激活机制，依赖于合成共底物来激活一种明显潜伏的酶。我们的发现可能会重新开启将CB 1954用于人类恶性疾病直接治疗的应用。
