Structure-based design, synthesis, and biological evaluation of conformationally restricted novel 2-alkylthio-6-[1-(2,6-difluorophenyl)alkyl]-3,4-dihydro-5-alkylpyrimidin-4(3H)-ones as non-nucleoside inhibitors of HIV-1 reverse transcriptase
Structure-based design, synthesis, and biological evaluation of conformationally restricted novel 2-alkylthio-6-[1-(2,6-difluorophenyl)alkyl]-3,4-dihydro-5-alkylpyrimidin-4(3H)-ones as non-nucleoside inhibitors of HIV-1 reverse transcriptase
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DOI：
10.1021/jm010853h
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发表时间：
2001-08-02
影响因子：
7.3
通讯作者：
Loddo, R
中科院分区：
文献类型：
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作者：
Mai, A;Sbardella, G;Loddo, R
5-烷基-2-(烷硫基)-6-(2,6-二氟苄基)-3,4-二氢嘧啶-4(3H)-酮 (S-DABO, 2) 最近被描述为一类新型人类免疫缺陷病毒 1 型 (HIV-1) 非核苷逆转录酶 (RT) 抑制剂 (NNRTI)，在纳摩尔浓度下具有活性（Mai, A. 等人，J. Med.化学 1999, 42, 619-627)。在我们的先导化合物优化工作中，我们设计了新型构象限制性 S-DABO，3，其特征是在苄基碳 (Y = Me) 和嘧啶 5 位 (R = Me) 上有一个甲基。构象分析和对接模拟表明，两个甲基的存在将显着降低构象灵活性，而不会影响 R 对映体中装入 RT 非核苷结合袋的能力。为了开发结构-活性关系，我们制备了属于胸腺嘧啶（R = Me）和尿嘧啶（R = H）系列的几种3型同源物，具有各种2-烷硫基侧链（X = Me，i-Pr，n-Bu，i-Bu，s-Bu，c-戊基和c-己基）和不同于2,6-二氟苯基（Ar =苯基，2,6-二氯苯基， 1-萘基）。此外，除了α-甲基衍生物(Y=Me)之外，α-乙基衍生物(Y=Et)也包括在合成项目中。所有新化合物都在 MT-4 细胞中评估了细胞毒性和抗 HIV-1 活性，其中一些化合物使用均聚物模板引物针对高度纯化的重组野生型 HIV-1 RT 进行了检测。结果表示为CC50（细胞毒性）、EC50（抗HIV-1活性）、SI（选择性，由CC50/EC50比率给出）和IC50（RT抑制活性）值。在 2,6-二氟苄基胸腺嘧啶 (R = Me) 系列中，苄基碳的甲基化提高了抗 HIV-1 和 RT 抑制活性以及选择性。化合物 3w（Ar = 2,6-F-2-Ph，R = Y = Me，X = c-戊基）是迄今为止报道的 S-DABO 中最有效和最具选择性的（CC50 > 200 muM，EC50 = 6 nM，IC50 = 5 nM，SI > 33 333）。对纯对映体 (+)-3w（比 (-)-3w 活性高得多）进行的测定得出以下结果：CC50 > 200 muM，EC50 = 2 nM，IC50 = 8 nM，SI > 100 000，在 MKC-442 活性和选择性较低的条件下（CC50 > 200 muM，EC50 = 30 nM，IC50 = 40 nM，SI > 6666）。与尿嘧啶对应物 (R = H) 相比，2,6-二氟苯乙基胸腺嘧啶 (R = Me) 通常具有更高的效力。在 2,6-二氟苯基系列中，性能最好和最差的 2-烷硫基侧链分别是 2-c-戊硫基和 2-甲硫基。当苄基碳上的甲基被乙基取代时，活性保留或略有下降，因此表明包含该取代基的 RT 内空腔的尺寸与大于乙基的基团不相容。不同于 2,6-二氟苯基（苯基、1-萘基、2,6-二氯苯基）的芳基部分通常对活性有害，这与 2,6-氟对抑制剂的芳香部分和 Tyr188 之间假定的电荷转移相互作用所产生的有利电子效应一致。
5-Alkyl-2-(alkylthio)-6-(2,6-difluorobenzyl)-3,4-dihydropyrimidin-4(3H)-ones (S-DABOs, 2) have been recently described as a new class of human immunodeficiency virus type 1 (HIV-1) nonnucleoside reverse transcriptase (RT) inhibitors (NNRTIs) active at nanomolar concentrations (Mai, A. et al. J. Med. Chem. 1999, 42, 619-627). In pursuing our lead optimization efforts, we designed novel conformationally restricted S-DABOs, 3, featuring a methyl at the benzylic carbon (Y = Me) and at the pyrimidine 5-position (R = Me). Conformational analyses and docking simulations suggested that the presence of both methyls would significantly reduce conformational flexibility without compromising, in the R enantiomers, the capability of fitting into the RT non-nucleoside binding pocket. To develop structure-activity relationships, we prepared several congeners of type 3 belonging to the thymine (R = Me) and uracil (R = H) series, featuring various 2-alkylthio side chains (X = Me, i-Pr, n-Bu, i-Bu, s-Bu, c-pentyl, and c-hexyl) and aryl moieties different from the 2,6-difluorophenyl (Ar = phenyl, 2,6-dichlorophenyl, 1-naphthyl). Moreover, alpha -ethyl derivatives (Y = Et) were included in the synthetic project in addition to alpha -methyl derivatives (Y = Me). All of the new compounds were evaluated for their cytotoxicity and anti-HIV-1 activity in MT-4 cells, and some of them were assayed against highly purified recombinant wild-type HIV-1 RT using homopolymeric template primers. The results were expressed as CC50 (cytotoxicity), EC50 (anti-HIV-1 activity), SI (selectivity, given by the CC50/EC50 ratio), and IC50 (RT inhibitory activity) values. In the 2,6-difluorobenzylthymine (R = Me) series, methylation of the benzylic carbon improved anti-HIV-1 and RT inhibitory activities together with selectivity. Compound 3w (Ar = 2,6-F-2-Ph, R = Y = Me, X = c-pentyl) turned out the most potent and selective among the S-DABOs reported to date (CC50 > 200 muM, EC50 = 6 nM, IC50 = 5 nM, and SI > 33 333). Assays performed on the pure enantiomer (+)-3w, much more active than (-)-3w, yielded the following results: CC50 > 200 muM, EC50 = 2 nM, IC50 = 8 nM, and SI > 100 000, under conditions wherein MKC-442 was less active and selective (CC50 > 200 muM, EC50 = 30 nM, IC50 = 40 nM, SI > 6666). The 2,6-difluorophenylethylthymines (R = Me) were generally endowed with higher potency compared with the uracil counterparts (R = H). In the 2,6-difluorophenyl series the best and the least performant 2-alkylthio side chains were the 2-c-pentylthio and the 2-methylthio, respectively. When the methyl at the benzylic carbon was replaced by an ethyl, activity was retained or decreased slightly, thus suggesting that the dimensions of the cavity within the RT hosting this substituent would not be compatible with groups larger than ethyl. Aryl moieties different from the 2,6-difluorophenyl (phenyl, 1-naphthyl, 2,6-dichlorophenyl) were generally detrimental to activity, consistent with a favorable electronic effect exerted by the 2,6-fluorines on a putative charge-transfer interaction between the aromatic moieties of the inhibitor and Tyr188.