Key Laboratory of Structure-Based Drugs Design & Discovery of Ministry of Education, School of Pharmaceutical Engineering, Shenyang Pharmaceutical University, 103 Culture Road, Shenhe District, Shenyang 110016, China.
School of Pharmacy, China Medical University, 77 Puhe Road, North New Area, Shenyang, 110122, China.
Eur J Med Chem. 2022 Feb 5;229:114086. doi: 10.1016/j.ejmech.2021.114086. Epub 2021 Dec 29.
Many pyrimidine-based xanthine oxidase (XO) inhibitors with diverse chemotypes have been reported recently. Our previous study revealed that 2-(4-alkoxy-3-cyano)phenyl-6-imino-1,6-dihydropyrimidine-5-carboxylic acid derivatives exhibited remarkable XO inhibitory potency. Notably, an intramolecular hydrogen bond (IMHB) formed between amino and carboxylic groups could be observed. With the hope to expand the structure-activity relationships (SARs) and obtain potential pyrimidine-based XO inhibitors, IMHB interruption and scaffold hopping were carried out on these compounds to design 2-(4-alkoxy-3-cyanophenyl)pyrimidine-4/5-carboxylic acids (11a-11n and 15a-15j) and 6-(4-alkoxy-3-cyanophenyl)-1,2-dihydro-3H-pyrazolo[3,4-d]pyrimidin-3-ones (19a-19j). Among them, compound 19a (IC = 0.039 μM) was identified as the most promising compound with substantially higher in vitro inhibitory potency than allopurinol (IC = 7.590 μM) and comparable to febuxostat (IC = 0.028 μM). The SAR analysis revealed that interrupting the IMHB through the removal of the amino group could damage the XO inhibitory potency; pyrimidine-4-carboxylic acid moiety was more beneficial for the XO inhibitory potency than the pyrimidine-5-carboxylic acid moiety. Additionally, enzyme kinetics studies suggested that compounds 11a, 15a and 19a acted as mixed-type inhibitors for XO and the removal of 6-position amino group resulted in a weakened affinity to the free enzyme, but an enhanced binding to the enzyme-substrate complex. Molecular modeling provided a reasonable explanation for the SARs observed in this study. Furthermore, in vivo hypouricemic effects demonstrated that compounds 15a and 19a could effectively reduce serum uric acid levels at an oral dose of 10 mg/kg, with 19a demonstrating a stronger effect than 15a. Therefore, our study proved that 6-(4-alkoxy-3-cyanophenyl)-1,2-dihydro-3H-pyrazolo[3,4-d]pyrimidin-3-ones were potent pyrimidine-based XO inhibitors and compound 19a required further structural optimization as a potential and efficacious agents for the treatment of hyperuricemia and gout.
最近有许多嘧啶类黄嘌呤氧化酶（XO）抑制剂具有不同的化学结构。我们之前的研究表明，2-（4-烷氧基-3-氰基）苯基-6-亚氨基-1,6-二氢嘧啶-5-羧酸衍生物具有显著的 XO 抑制活性。值得注意的是，氨基和羧酸基团之间形成了分子内氢键（IMHB）。为了拓展构效关系（SARs）并获得潜在的嘧啶类 XO 抑制剂，我们对这些化合物进行了 IMHB 中断和骨架跃迁设计，得到了 2-（4-烷氧基-3-氰基苯基）嘧啶-4/5-羧酸（11a-11n 和 15a-15j）和 6-（4-烷氧基-3-氰基苯基）-1,2-二氢-3H-吡唑并[3,4-d]嘧啶-3-酮（19a-19j）。其中，化合物 19a（IC=0.039μM）被鉴定为最有前途的化合物，其体外抑制活性显著高于别嘌醇（IC=7.590μM），与非布司他（IC=0.028μM）相当。SAR 分析表明，通过去除氨基来中断 IMHB 可能会破坏 XO 的抑制活性；嘧啶-4-羧酸部分比嘧啶-5-羧酸部分更有利于 XO 的抑制活性。此外，酶动力学研究表明，化合物 11a、15a 和 19a 对 XO 表现为混合抑制作用，去除 6-位氨基会导致对游离酶的亲和力减弱，但与酶-底物复合物的结合增强。分子模拟为该研究中观察到的 SARs 提供了合理的解释。此外，体内降尿酸作用研究表明，化合物 15a 和 19a 可有效降低血清尿酸水平，口服剂量为 10mg/kg 时，19a 的作用强于 15a。因此，我们的研究证明 6-（4-烷氧基-3-氰基苯基）-1,2-二氢-3H-吡唑并[3,4-d]嘧啶-3-酮是有效的嘧啶类 XO 抑制剂，化合物 19a 需要进一步的结构优化，作为治疗高尿酸血症和痛风的潜在有效药物。
