Phthalein derivatives as a new tool for selectivity in thymidylate synthase inhibition
Phthalein derivatives as a new tool for selectivity in thymidylate synthase inhibition
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DOI：
10.1021/jm9900016
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发表时间：
1999-06-17
影响因子：
7.3
通讯作者：
La Colla, P
中科院分区：
文献类型：
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作者：
Costi, PM;Rinaldi, M;La Colla, P
从前导化合物酚酞衍生的一组新的酞衍生物被设计用于特异性地补充细菌形式的胸苷酸合成酶（Lactobacillus casei, LcTS）与人类TS （hTS）酶的结构特征。筛选新化合物对不同物种的TS酶，即干酪乳杆菌（LcTS）、卡氏肺囊虫（PcTS）、新型隐球菌（CnTS）和人胸腺苷酸合成酶（hTS）的活性和特异性。测定了所有化合物对LcTS的表观抑制常数（Ki），测定了对四种TS的抑制因子（IF，每种抑制剂存在和不存在时酶促反应初始速率之比）。我们发现IF和Ki这两个活性参数之间存在很强的相关性，因此我们将更简单的IF作为筛选因子，以加速生物学评价。化合物5b、5c、5ba和6bc对LcTS表现出明显的抑制作用，而对hTS基本没有抑制作用，首次证明了TSs具有显著的物种特异性。由于酶之间的序列同源性，一些化合物也表现出对CnTS的高活性和特异性。特别是，3 -羟基-3-（3-氯-4-羟基苯基）-6-硝基- 1h， 3h -萘[1,8-c，d]吡喃-1- 1，（6bc）对CnTS的IF < 0.04 (K-i = 0.45 μ M)，而在化合物的最大溶解度浓度（200 μ M）下，在hTS实验中保持无活性。在细胞培养试验中发现，大多数化合物对人类细胞系无细胞毒性，但对几种革兰氏阳性细菌有细胞毒性。这些结果与酶的测定结果一致。有趣的是，在细胞培养实验中，几种化合物也对新生Cr.具有选择性活性。一般来说，抗革兰氏阳性细菌的活性和选择性最高的化合物是那些在酶分析中设计和发现的针对LcTS和hTS的特异性化合物。最初的先导化合物对大多数测试细胞系的选择性最低。据我们所知，这些化合物是第一个对细菌TS有选择性的TS抑制剂。
A new set of phthalein derivatives stemming from the lead compound, phenolphthalein, were designed to specifically complement structural features of a bacterial form of thymidylate synthase (Lactobacillus casei, LcTS) versus the human TS (hTS) enzyme. The new compounds were screened for their activity and their specificity against TS enzymes from different species, namely, L. casei (LcTS), Pneumocystis carinii (PcTS), Cryptococcus neoformans (CnTS), and human thymidylate synthase (hTS). Apparent inhibition constants (Ki) for all the compounds against LcTS were determined, and inhibition factors (IF,ratio between the initial rates of the enzymatic reaction in the presence and absence of each inhibitor) against each of the four TS species were measured. A strong correlation was found between the two activity parameters, IF and Ki, and therefore the simpler IF was used as a screening factor in order to accelerate biological evaluation. Compounds 5b, 5c, 5ba, and 6bc showed substantial inhibition of LcTS while remaining largely inactive against hTS, illustrating for the first time remarkable species specificity among TSs. Due to sequence homology between the enzymes, several compounds also showed high activity and specificity for CnTS. In particular, 3-hydroxy-3-(3-chloro-4-hydroxyphenyl)-6-nitro-1H,3H-naphtho[1,8-c,d]pyran-1-one, (6bc) showed an IF < 0.04 for CnTS (K-i = 0.45 mu M) while remaining inactive in the hTS assay at the maximum solubility concentration of the compound (200 mu M). In cell culture assays most Of the compounds were found to be noncytotoxic to human cell lines but were cytotoxic against several species of Grampositive bacteria. These results are consistent with the enzymatic assays. Intriguingly, several compounds also had selective activity against Cr. neoformans in cell culture assay. In general, the most active and selective compounds against the Gram-positive bacteria were those designed and found in the enzyme assay to be specific for LcTS versus hTS. The original lead compound was least selective against most of the cell lines tested. To our knowledge these compounds are the first TS inhibitors selective for bacterial TS with respect to hTS.