CN104744484A - 一种3-呋喃[2,3-b]并吡啶-4-吲哚马来酰亚胺衍生物及其制备和应用 - Google Patents
一种3-呋喃[2,3-b]并吡啶-4-吲哚马来酰亚胺衍生物及其制备和应用 Download PDFInfo
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- C07D491/04—Ortho-condensed systems
- C07D491/044—Ortho-condensed systems with only one oxygen atom as ring hetero atom in the oxygen-containing ring
- C07D491/048—Ortho-condensed systems with only one oxygen atom as ring hetero atom in the oxygen-containing ring the oxygen-containing ring being five-membered
Landscapes
- Chemical & Material Sciences (AREA)
- Organic Chemistry (AREA)
- Plural Heterocyclic Compounds (AREA)
Abstract
本发明提供一类具有新型结构的3-呋喃[2,3-b]并吡啶-4-吲哚马来酰亚胺衍生物及其制备和应用。该类化合物以取代吲哚为起始原料，依次经酰化、醇解、N-烷基化得到中间体，中间体与取代的2-(呋喃[2,3-b]并吡啶-3-基)乙酰胺在叔丁醇钾的作用下缩合得到目标产物3-呋喃[2,3-b]并吡啶-4-吲哚马来酰亚胺衍生物，该类化合物具有高效选择性的GSK-3β抑制活性，可用于治疗GSK-3β介导疾病如糖尿病、双向精神障碍、阿尔茨海默病、癌症、炎性疾病、抑郁症、神经保护和精神分裂症。本发明设计合理，制备方法简单实用。
Description
技术领域
本发涉及一种3-呋喃[2,3-b]并吡啶-4-吲哚马来酰亚胺衍生物及其制备和应用。
背景技术
糖原合成酶激酶-3 (glycogen synthas kinase-3，GSK-3)是一种多功能的丝氨酸/苏氨酸类蛋白激酶，在所有真核生物中都有分布。在哺乳动物中主要有2种亚型：GSK-3α和GSK-3β。其中GSK-3β通过参与胰岛素、Wnt／β-连环蛋白、Hedgehog以及 Notch等信号传导通路，在调控细胞的分化、代谢、凋亡及基因表达等方面都起着重要作用。这些信号传导途径的失调与2型糖尿病、阿尔茨海默病、癌症等疾病密切相关。例如在2型糖尿病中，过度表达的GSK-3β可以通过磷酸化糖原合成酶（GS）而抑制其活性，降低细胞外葡萄糖向糖原转化的能力；另外过度表达的GSK-3β也会使胰岛素受体底物-1（IRS-1）中多个丝氨酸残基磷酸化，消弱胰岛素信号的传导，导致靶组织对胰岛素的反应性降低，主要表现为脂肪和肌肉组织对葡萄糖的摄取减少。上述原因均会导致血糖浓度的上升。在阿尔茨海默尔氏病中，过表达的GSK-3β会导致Tau蛋白过度磷酸化，引起神经元纤维缠结，并会导致神经元死亡。因此，GSK-3β抑制剂可用于预防和治疗糖尿病和阿尔茨海默尔氏病等疾病。开发具有高活性和选择性的GSK-3β抑制剂已成为当前新药开发的热点。
迄今为止，已发现了多种类型的GSK-3β抑制剂，以小分子抑制剂居多，如 paullones类化合物、靛玉红类化合物、氨基嘧啶类化合物、Li离子和TDZD等。其中Li离子、TDZD为非ATP竞争型抑制剂，其余为ATP竞争型抑制剂。这些小分子抑制剂与ATP竞争活性口袋的作用方式大多为与催化区的 Asp133和Val135形成2至3个氢键，另外结构水也可作为水桥连接蛋白残基和小分子形成氢键。然而目前已开发的GSK-3β的抑制剂在抑制活性和选择性方面有待于进一步提高，开发具有新型结构的高效特异性GSK-3β抑制剂已成为当务之急。
发明内容
针对现有技术中存在的上述问题，本发明的目的是提供一种3-呋喃[2,3-b]并吡啶-4-吲哚马来酰亚胺衍生物及其制备和应用，可用于治疗GSK-3β介导的疾病如糖尿病、双向精神障碍、阿尔茨海默病、癌症、炎性疾病、抑郁症、神经保护和精神分裂症等。
所述的一种3-呋喃[2,3-b]并吡啶-4-吲哚马来酰亚胺衍生物，其特征在于该衍生物的结构通式如下：
其中：
R1，R3为一取代或二取代基，取代基独立的选自：氢；碳原子数为1-3的直链烷基、支链烷基和环烷基或碳原子数为1-3的直链烷氧基、支链烷氧基和环烷氧基或卤素；
R2为氢； 碳原子数为1-5的直链烷基、支链烷基和环烷基或(CH2)nR4，其中n=1-5, R4为吡咯、咪唑、三氮唑、吗啉、四氢吡咯、哌啶或二甲氨基。
所述的3-呋喃[2,3-b]并吡啶-4-吲哚马来酰亚胺衍生物，其特征在于R1为H、5-F、5-Br或5-Cl，n为3，R3为H，R4为咪唑或吗啉。
所述的3-呋喃[2,3-b]并吡啶-4-吲哚马来酰亚胺衍生物的制备方法，其特征在于，以如式（1）所示的取代吲哚为起始原料，依次经酰化、醇解得到如式（2）所示的中间体2、中间体2经N-烷基化得到如式（3）所示的中间体3，中间体3与如式（4）所示的取代的2-(呋喃[2,3-b]并吡啶-3-基)乙酰胺在叔丁醇钾的作用下缩合得到如式（5）所示的目标产物3-呋喃[2,3-b]并吡啶-4-吲哚马来酰亚胺衍生物，反应式如下：
所述的一种3-呋喃[2,3-b]并吡啶-4-吲哚马来酰亚胺衍生物的制备方法，其特征在于包括如下步骤：
1）中间体2的制备
将取代吲哚加入溶剂无水乙醚中搅拌溶解，控制温度0-5℃，缓慢滴加草酰氯的乙醚溶液，滴完后保温反应1h，然后降温至-30 ~ -20℃，滴加甲醇钠的甲醇溶液，滴完后搅拌30min，然后将反应液倒入冰水中，过滤，水洗，二氯甲烷洗，干燥后得到淡黄色中间体2；
2）中间体3的制备
将步骤1）得到的中间体2加入无水DMF中搅拌溶解，将反应液降温至0~5 ℃，分批加入70%的NaH，加完后升至室温反应30 min，然后在0℃加入卤代烃，加完后将反应液升至室温反应1h，冷却后将反应液倒入水中，后处理得到淡黄色中间体3，所述的后处理方法如下：依次经乙酸乙酯萃取，有机相用饱和食盐水洗，无水硫酸钠干燥，减压浓缩，残余物用硅胶柱层析（乙酸乙酯：石油醚的体积比为1：3）得到淡黄色中间体3c；
3）目标产物5的制备
将中间体4和步骤 2）得到的中间体3、加入无水THF中搅拌溶解，控制温度-5 ~ 0℃，缓慢滴入叔丁醇钾的叔丁醇溶液，滴完后升至室温反应2h，然后加入浓盐酸搅拌30 min，再将反应液倒入10% 碳酸氢钠溶液中，乙酸乙酯萃取后有机相用饱和食盐水洗涤，无水硫酸钠干燥，过滤，减压浓缩，残余物用硅胶柱层析（二氯甲烷与甲醇的体积比为 50:1）提纯，得到目标产物5。
所述的一种3-呋喃[2,3-b]并吡啶-4-吲哚马来酰亚胺衍生物及其药学上能接受的盐在制备GSK-3β介导疾病治疗药物中的应用。
本发明通过采用上述技术，提供一类具有新型结构的3-呋喃[2,3-b]并吡啶-4-吲哚马来酰亚胺衍生物及其制备方法，其原料简单易得，合成路线设计合理、制备方法简单实用，后处理方便，得到的该类化合物具有高效选择性的GSK-3β抑制活性，可用于制备GSK-3β介导疾病治疗药物，能用于GSK-3β介导疾病如糖尿病、双向精神障碍、阿尔茨海默病、癌症、炎性疾病、抑郁症、神经保护和精神分裂症的治疗。
附图说明
图1为本发明的GSK-3β抑制剂对tau蛋白396位丝氨酸磷酸化的影响图。
具体实施方式
本发明结合实施例作进一步的说明。以下的实施例是说明本发明的，而不是以任何方式限制本发明。
中间体及目标化合物的制备：
实施例1: 中间体2的制备
在三口烧瓶中加入0.026 mol 取代吲哚，30 mL无水乙醚，搅拌溶解，控制温度0-5℃，缓慢滴加3.4 g（0.026 mol）草酰氯的乙醚（5 mL）溶液，滴完后保温反应1h, 然后降温至-30 ~ -20℃左右，滴加16.3 g甲醇钠的甲醇溶液（17.5%，0.052 mol），滴完后搅拌30 min，然后将反应液倒入100 mL冰水中，过滤，水洗（3 × 10 mL），二氯甲烷洗（2 × 10 mL），干燥后得到淡黄色中间体2，
2-(7-甲基-1H-吲哚-3-基)-2-氧代乙酸甲酯（2b）
收率：73.2%，熔点: 183-185 oC. 1H NMR (500 MHz, DMSO-d 6 ) δ 12.48 (brs, 1H), 8.41 (d, J = 3.5 Hz, 1H), 8.00 (d, J = 7.9 Hz, 1H), 7.18 (t, J = 7.5 Hz, 1H), 7.33 (d, J = 7.1Hz, 1H), 3.90 (s, 3H), 2.53 (s, 3H)，
2-(5-氟-1H-吲哚-3-基)-2-氧代乙酸甲酯（2c）
收率：57.3%，熔点: > 250 oC. 1H NMR (500 MHz, DMSO-d 6 ) δ 12.52 (brs, 1H), 8.53 (s, 1H), 7.84 (dd, J = 9.6, 2.6 Hz, 1H), 7.59-7.56 (m, 1H), 7.17 (d, J = 7.1Hz, 1H), 3.90 (s, 3H)，
2-(5-氯-1H-吲哚-3-基)-2-氧代乙酸甲酯（2e）
收率：58.9%，熔点：> 250 oC. 1H NMR (500 MHz, DMSO-d 6 ) δ 12.57 (brs, 1H), 8.54 (s, 1H), 8.14 (d, J = 2.0 Hz, 1H), 7.58 (d, J = 8.6 Hz, 1H), 7.33 (dd, J = 8.6, 2.0 Hz, 1H), 3.90 (s, 3H)。
实施例2：中间体3c-k的制备
在三口烧瓶中加入（4.9 mmol）2，10 ml无水DMF，搅拌溶解，将反应液降温至0~5℃，分批加入0.17 g (4.9 mmol) 70%的NaH，加完后升至室温反应30 min，然后在0℃加入5.9 mmol卤代烃，加完后将反应液升至室温反应1h，冷却后将反应液倒入100 mL中水中，乙酸乙酯萃取（3 × 50 mL），合并有机相，饱和食盐水洗（3 × 150 mL），无水硫酸钠干燥，减压浓缩，残余物用硅胶柱层析（乙酸乙酯：石油醚 = 1：3）提纯得到淡黄色中间体3c-k，
二甲基-1H-吲哚-3-基)-2-氧代乙酸甲酯（3c）
收率63.5%, 熔点: 97-98℃. 1H NMR (500 MHz, CDCl3) δ 8.32 (d, J = 8.0 Hz, 1H), 8.20 (s, 1H), 7.20 (t, J = 7.6 Hz, 1H), 7.04 (d, J = 9.0 Hz, 1H), 4.08 (s, 3H), 3.95 (s, 3H), 2.74 (s, 3H)，
2-(5-氟-1-甲基-1H-吲哚-3-基)-2-氧代乙酸甲酯 (3d)
收率67.4%，熔点: 128-129℃. 1H NMR (500 MHz, CDCl3) δ 8.36 (s, 1H), 8.10 (dd, J = 9.3, 2.5 Hz , 1H), 7.30-7.27 (m, 1H), 7.11-7.06 (m, 1H), 3.96 (s, 3H), 3.87 (s, 3H)，
2-(6-氟-1-甲基-1H-吲哚-3-基)-2-氧代乙酸甲酯 (3e)
收率70.1%, 熔点: 121-122℃. 1H NMR (500 MHz, CDCl3) δ 8.39-8.35 (m, 1H), 8.34 (s, 1H), 7.11 (td, J = 9.5, 2.0 Hz, 1H), 7.05 (dd, J = 9.0, 2.0 Hz, 1H), 3.96 (s, 3H), 3.84 (s, 3H)，
2-(5-氯-1-甲基-1H-吲哚-3-基)-2-氧代乙酸甲酯 (3f)
收率 61.7%, 熔点: 117-118℃. 1H NMR (500 MHz, CDCl3) δ 8.44 (d, J = 2.0 Hz, 1H), 8.38 (s, 1H), 7.34-7.28 (m, 2H), 3.97 (s, 3H), 3.89 (s, 3H)，
2-(6-氯-1-甲基-1H-吲哚-3-基)-2-氧代乙酸甲酯 (3g)
收率62.7% ，熔点: 135-137℃. 1H NMR (500 MHz, CDCl3) δ 8.36-8.34 (m, 2H), 7.38 (d, J = 2.0 Hz, 1H), 7.33 (dd, J = 8.0, 2.0 Hz, 1H), 3.97 (s, 3H), 3.86 (s, 3H)，
2-(5-溴-1-甲基-1H-吲哚-3-基)-2-氧代乙酸甲酯 (3h)
收率 65.3%, 熔点: 126-127℃. 1H NMR (500 MHz, CDCl3) δ 8.59 (d, J = 2.0 Hz, 1H), 8.36 (s, 1H), 7.46 (dd, J = 8.5, 2.0 Hz, 1H), 7.25 (d, J = 8.5 Hz, 1H), 3.96 (s, 3H), 3.88 (s, 3H)，
2-(6-溴-1-甲基-1H-吲哚-3-基)-2-氧代乙酸甲酯 (3i)
收率 67.9%, 熔点: 138-139℃. 1H NMR (500 MHz, CDCl3) δ 8.34 (s, 1H), 8.30 (d, J = 8.5 Hz, 1H), 7.54 (d, J = 1.5 Hz, 1H), 7.47 (dd, J = 8.5, 1.5 Hz, 1H), 3.96 (s, 3H), 3.86 (s, 3H)，
2-(5-甲氧基-1-甲基-1H-吲哚-3-基)-2-氧代乙酸甲酯 (3j)
收率78.5%, 熔点: 131-132℃. 1H NMR (500 MHz, CDCl3) δ 8.30 (s, 1H), 7.95 (d, J = 2.5 Hz, 1H), 7.26 (s, 1H), 6.99 (dd, J =9.0, 2.5 Hz, 1H), 3.96 (s, 3H), 3.92 (s, 3H), 3.86 (s, 3H)，
2-(1-烯丙基-1H-吲哚-3-基)-2-氧代乙酸甲酯 ( 3k)
收率 54.8% , 熔点: 67-68℃. 1H NMR (500 MHz, CDCl3) δ 8.48-8.42 (m, 1H), 8.39 (s, 1H), 7.41-7.32 (m, 3H), 6.07-5.99 (m, 1H), 5.34 (d, J = 10.9 Hz, 1H), 5.22 (d, J = 17.0, 1H), 4.80 (d, J = 5.5 Hz, 2H), 3.96 (s, 3H)。
实施例3：目标产物5a-r 的制备
在三口瓶中加入0.28 mmol 2-(呋喃[2,3-b]并吡啶-3-基)乙酰胺4、 0.37 mmol 3a-r和10 mL无水THF，搅拌溶解，控制温度-5 ~ 0℃左右, 缓慢滴入（0.84ml, 0.84 mmol）叔丁醇钾的叔丁醇溶液，滴完后升至室温反应2h, 然后加入5 mL浓盐酸, 搅拌30 min，再将反应液倒入100 mL 10% 碳酸氢钠溶液中，乙酸乙酯萃取（3 × 50 mL），合并有机相，有几层用饱和食盐水洗涤，无水硫酸钠干燥，过滤，减压浓缩，残余物用硅胶柱层析（二氯甲烷：甲醇 = 50:1）提纯，得到目标产物5a-r，
3-(呋喃[2,3-b]并吡啶-3-基)-4-(1H-吲哚-3-基) -1H-吡咯-2,5-酮 (5a)
收率10.1%, 熔点: 210-212℃.1H NMR (500 MHz, DMSO-d 6 ) δ 12.14 (brs, 1H), 11.44 (brs, 1H), 8.23 (s, 1H), 7.83 (d, J = 8.5 Hz, 1H), 7.69-7.58 (m, 2H), 7.44 (d, J = 8.0 Hz, 1H), 7.31 (t, J = 7.5 Hz, 1H), 7.08-7.04 (m, 1H), 6.72 (t, J = 8.0 Hz, 1H), 6.51 (d, J = 8.5 Hz, 1H). ESI-MS: m/z [M+H]+ 330. Anal. Calcd for C19H11N3O3: C, 69.30; H, 3.37; N, 12.76. Found: C, 69.38; H, 3.49; N, 12.48，
3-(呋喃[2,3-b]并吡啶-3-基)-4-(1-甲基-1H-吲哚-3-基) -1H-吡咯-2,5-酮 (5b)
收率 17.5%, 熔点: > 250℃. 1H NMR (500 MHz, DMSO-d 6 ) δ 11.25 (brs, 1H), 8.36 (s, 1H), 8.22 (dd, J = 4.5, 1.5 Hz, 1H), 8.02 (s, 1H), 7.49 (d, J = 8.0 Hz, 1H), 7.30 (dd, J = 8.0, 1.5 Hz, 1H), 7.10 (t, J = 8.0 Hz, 1H), 7.06-7.01 (m, 1H), 6.81-6.73 (m, 2H), 3.91 (s, 3H). ESI-MS: m/z [M+H]+ 344. Anal. Calcd for C20H13N3O3: C, 69.96; H, 3.82; N, 12.24. Found: C, 69.77; H, 3.68; N, 12.11，
3- (1,7-甲基-1H-吲哚-3-基) -4- (呋喃[2,3-b]并吡啶-3-基)- 1H-吡咯-2,5-酮 (5c)
收率 19.8%， 熔点: 243-245℃. 1H NMR (500 MHz, DMSO-d 6 ) δ 11.23 (brs, 1H), 8.32 (s, 1H), 8.22 (dd, J = 4.5, 1.5 Hz, 1H), 7.90 (s, 1H), 7.34 (dd, J = 8.0, 1.5 Hz, 1H), 7.08-7.04 (m, 1H), 6.77 (d, J = 8.0 Hz, 1H), 6.61-6.55 (m, 2H), 4.16 (s, 3H), 2.71 (s, 3H). ESI-MS: m/z [M+H]+ 358. Anal. Calcd for C21H15N3O3: C, 70.58; H, 4.23; N, 11.76. Found: C, 70.46; H, 4.51; N, 11.99，
3- (5-氟-1-甲基-1H-吲哚-3-基) -4- (呋喃[2,3-b]并吡啶-3-基)- 1H-吡咯-2,5-酮(5d)
收率 28.4% ，熔点: 232-234℃. 1H NMR (500 MHz, DMSO-d 6 ) δ 11.27 (brs, 1H), 8.39 (s, 1H), 8.24 (dd, J = 4.5, 1.5 Hz, 1H), 8.03 (s, 1H), 7.53-7.49 (m, 1H), 7.29 (dd, J = 8.0, 1.5 Hz, 1H), 7.08-7.04 (m, 1H), 6.96 (td, J = 9.5, 2.5 Hz, 1H). 6.51 (dd, J = 10.0, 2.5 Hz, 1H), 3.89 (s, 3H). ESI-MS: m/z [M+H]+ 362. Anal. Calcd for C20H12FN3O3: C, 66.48; H, 3.35; N, 11.63. Found: C, 66.75; H, 3.57; N, 11.91，
3- (6-氟-1-甲基-1H-吲哚-3-基) -4- (呋喃[2,3-b]并吡啶-3-基)- 1H-吡咯-2,5-酮(5e)
收率25.7%，熔点: 216-217℃. 1H NMR (500 MHz, DMSO-d 6 ) δ 11.27 (brs, 1H), 8.40 (s, 1H), 8.23 (dd, J = 4.5, 1.5 Hz, 1H), 7.98 (s, 1H), 7.40 (dd, J = 10.0, 2.0 Hz, 1H), 7.25 (dd, J = 8.0, 1.5 Hz, 1H), 7.07-7.03 (m, 1H), 6.84-6.79 (m, 1H), 6.67-6.63 (td, J = 9.5, 2.5 Hz, 1H), 3.87 (s, 3H). ESI-MS: m/z [M+H]+ 362. Anal. Calcd for C20H12FN3O3: C, 66.48; H, 3.35; N, 11.63. Found: C, 66.25; H, 3.28; N, 11.44，
3- (5-氯-1-甲基-1H-吲哚-3-基) -4- (呋喃[2,3-b]并吡啶-3-基)- 1H-吡咯-2,5-酮 (5f)
收率 26.2%，熔点: 111-113℃.1H NMR (500 MHz, DMSO-d 6 ) δ 11.28 (brs, 1H), 8.41 (s, 1H), 8.25 (dd, J = 4.5, 1.5 Hz, 1H), 8.02 (s, 1H), 7.52 (d, J = 8.5 Hz, 1H), 7.27 (dd, J = 8.0, 1.5 Hz, 1H), 7.11 (dd, J = 8.5, 1.5 Hz, 1H), 7.08-7.05 (m, 1H), 6.84 (d, J = 1.5 Hz, 1H), 3.88 (s, 3H). ESI-MS: m/z [M+H]+ 378. Anal. Calcd for C20H12ClN3O3: C, 63.59; H, 3.20; N, 11.12. Found: C, 63.31; H, 3.31; N, 1139，
3- (6-氯-1-甲基-1H-吲哚-3-基) -4- (呋喃[2,3-b]并吡啶-3-基)- 1H-吡咯-2,5-酮 (5g)
收率 21.7% ， 熔点: 209-211℃.1H NMR (500 MHz, DMSO-d 6 ) δ 11.29 (brs, 1H), 8.41 (s, 1H), 8.24 (dd, J = 4.5, 1.5 Hz, 1H), 8.00 (s, 1H), 7.65 (s, 1H), 7.25 (d, J = 8.0 Hz, 1H), 7.08-7.04 (m, 1H), 6.84-6.79 (m, 2H), 3.90 (s, 3H). ESI-MS: m/z [M+H]+ 378. Anal. Calcd for C20H12ClN3O3: C, 63.59; H, 3.20; N, 11.12. Found: C, 63.44; H, 3.36; N, 11.28，
3- (5-溴-1-甲基-1H-吲哚-3-基) -4- (呋喃[2,3-b]并吡啶-3-基)- 1H-吡咯-2,5-酮(5h)
收率 14.7% ，熔点: 146-148℃. 1H NMR (500 MHz, DMSO-d 6) δ 11.21 (brs, 1H), 8.41 (s, 1H), 8.26 (dd, J = 4.5, 1.5 Hz, 1H), 8.01 (s, 1H), 7.48 (d, J = 8.5 Hz, 1H), 7.28 (dd, J = 8.0, 1.5 Hz, 1H), 7.22 (dd, J = 8.0, 2.0 Hz, 1H), 7.09-7.05 (m, 1H), 6.99 （d, J = 1.5 Hz, 1H), 3.88 (s, 3H). ESI-MS: m/z [M+H]+ 422. Anal. Calcd for C20H12BrN3O3: C, 56.89; H, 2.86; N, 9.95. Found: C, 57.01; H, 2.66; N, 10.06，
3- (6-溴-1-甲基-1H-吲哚-3-基) -4- (呋喃[2,3-b]并吡啶-3-基)- 1H-吡咯-2,5-酮 (5i)
收率 11.5% ，熔点: > 250℃.1H NMR (500 MHz, DMSO-d 6 ) δ 11.29 (brs, 1H), 8.40 (s, 1H), 8.24 (dd, J = 4.5, 1.5 Hz, 1H), 7.99 (s, 1H), 7.79 (d, J = 1.5Hz, 1H), 7.26 (dd, J = 8.0, 1.5 Hz, 1H), 7.08-7.04 (m, 1H), 6.92 (dd, J = 8.5, 1.5 Hz, 1H), 6.79 (d, J = 8.5 Hz, 1H), 3.89 (s, 3H). ESI-MS: m/z [M+H]+ 422. Anal. Calcd for C20H12BrN3O3: C, 56.89; H, 2.86; N, 9.95. Found: C, 56.76; H, 2.73; N, 9.85，
3-(呋喃[2,3-b]并吡啶-3-基)-4-(5-甲氧基-1-甲基-1H-吲哚-3-基)-1H-吡咯-2,5-酮 (5j)
收率 22.3% ，熔点: > 250℃， 1H NMR (500 MHz, DMSO-d 6 ) δ 11.22 (brs, 1H), 8.33 (s, 1H), 8.24 (dd, J = 4.5, 1.5 Hz, 1H), 8.04 (s, 1H), 7.44 (dd, J = 8.0, 1.5 Hz, 1H), 7.37 (d, J = 8.5 Hz, 1H), 7.10-7.07 (m, 1H), 6.68 (dd, J = 8.5, 2.5 Hz, 1H), 6.12 (d, J = 2.5 Hz, 1H). 3.88 (s, 3H), 3.05 (s, 3H). ESI-MS: m/z [M+H]+ 374. Anal. Calcd for C21H15N3O4: C, 67.56; H, 4.05; N, 11.25. Found: C, 67.73; H, 4.21; N, 11.43，
3- (1-烯丙基-1H-吲哚-3-基) -4- (呋喃[2,3-b]并吡啶-3-基)- 1H-吡咯-2,5-酮(5k)
收率 20.8%，熔点: 221-223℃， 1H NMR (500 MHz, DMSO-d 6 ) δ 11.26 (brs, 1H), 8.40 (s, 1H), 8.21 (dd, J = 4.5, 1.5 Hz, 1H), 7.96 (s, 1H), 7.47 (d, J = 8.5Hz, 1H), 7.23 (dd, J = 8.0, 1.5 Hz, 1H), 7.08 (td, J = 8.5, 1.5Hz , 1H), 7.03-6.90 (m, 1H), 6.89 (d, J = 8.5 Hz, 1H). 6.77 (t, J = 8.0 Hz 1H), 6.05-5.97 (m, 1H), 5.20 (dd, J = 10.5, 1.5 Hz, 1H), 5.04 (dd, J = 17.0, 1.5 Hz, 1H), 4.95 (d, J = 5.5 Hz, 2H). ESI-MS: m/z [M+H]+ 370. Anal. Calcd for C22H15N3O3: C, 71.54; H, 4.09; N, 11.38. Found: C, 71.69; H, 4.18; N, 11.55，
3- (1-丁基-1H-吲哚-3-基) -4- (呋喃[2,3-b]并吡啶-3-基)- 1H-吡咯-2,5-酮(5l)
收率 16.5% ，熔点: 205-207℃.1H NMR (500 MHz, DMSO-d 6 ) δ 11.24 (brs, 1H), 8.40 (s, 1H), 8.21 (dd, J = 4.5, 1.5 Hz, 1H), 7.93 (s, 1H), 7.54 (d, J = 8.0Hz, 1H), 7.18 (dd, J = 8.0, 1.5 Hz, 1H), 7.09 (t, J = 8.0 Hz, 1H), 6.98-6.93 (m, 2H), 6.78 (t, J = 8.0Hz, 1H), 4.27 (t, J = 7.0, 2H), 1.75-1.67 (m, 2H), 1.26-1.19 (m, 2H), 0.88 (t, J = 7.0 Hz, 3H). ESI-MS: m/z [M+H]+ 386. Anal. Calcd for C23H19N3O3: C, 71.67; H, 4.97; N, 10.90. Found: C, 71.85; H, 4.79; N, 11.02，
3-(呋喃[2,3-b]并吡啶-3-基)-4-(1-异丙基-1H-吲哚-3-基)-1H-吡咯-2,5-酮(5m)
收率18.4% ，熔点: > 250℃. 1H NMR (500 MHz, DMSO-d 6 ) δ 11.25 (brs, 1H), 8.41 (s, 1H), 8.21 (dd, J = 4.5, 1.5 Hz, 1H), 7.90 (s, 1H), 7.59 (d, J = 8.0 Hz, 1H), 7.13-7.07 (m, 2H), 7.04-6.99 (m, 2H), 6.82 (td, J = 8.0, 1.5 Hz, 1H), 5.86-5.80 (m, 1H), 1.43 (d, J = 7.0 Hz, 6H). ESI-MS: m/z [M+H]+ 372. Anal. Calcd for C22H17N3O3: C, 71.15; H, 4.61; N, 11.31. Found: C, 71.42; H, 4.80; N, 11.17，
3-(1-(3-(1H-咪唑-1-基)丙基)-1H-吲哚-3-基)-4-(呋喃[2,3-b]并吡啶-3-基)-1H-吡咯-2,5-酮(5n)
收率13.4% ，熔点: 184-186℃。1H NMR (500 MHz, DMSO-d 6 ) δ 11.31 (brs, 1H), 8.40 (s, 1H), 8.19 (dd, J = 4.5, 1.5 Hz, 1H), 7.95 (s, 1H), 7.64 (s, 1H), 7.46 (d, J = 8.5 Hz, 1H), 7.29-7.19 (m, 2H), 7.14-7.05 (m, 1H),7.05-6.98 (m, 1H), 6.95-6.88 (m, 2H), 6.79 (t, J = 7.5 Hz, 1H), 4.26 (t, J = 7.0 Hz, 2H), 3.98 (t, J = 7.0 Hz, 2H), 2.29-2.18 (m, 2H). ESI-MS: m/z [M+H]+ 438. Anal. Calcd for C25H19N5O3: C, 68.64; H, 4.38; N, 16.01. Found: C, 68.43; H, 4.42; N, 16.27，
3-(1-(3-(1H-咪唑-1-基)丙基)-5-溴-1H-吲哚-3-基)-4-(呋喃[2,3-b]并吡啶-3-基)-1H-吡咯-2,5-酮 (5o)
收率19.5% ，熔点: 96-98℃. 1H NMR (500 MHz, DMSO-d 6 ) δ 11.28 (brs, 1H), 8.42 (s, 1H), 8.22 (dd, J = 4.5, 1.5 Hz, 1H), 7.93 (s, 1H), 7.84 (s, 1H), 7.47 (d, J = 8.5 Hz, 1H), 7.29-7.19 ( m, 3H), 7.14 (d, J = 1.5 Hz, 1H), 7.06-6.99 (m, 2H), 4.24 (t, J = 7.0 Hz, 2H), 3.97 (t, J = 7.0 Hz, 2H), 2.27-2.08 (m, 2H). ESI-MS: m/z [M+H]+ 516. Anal. Calcd for C25H18BrN5O3: C, 58.15; H, 3.51; N, 13.56. Found: C, 58.33; H, 3.68; N, 13.77，
3- (1-(3-(1H-咪唑-1-基)丙基)-5-氯-1H-吲哚-3-基)-4-(呋喃[2,3-b]并吡啶-3-基)- 1H-吡咯-2,5-酮 (5p)
收率 16.3% ，熔点: 107-109℃. 1H NMR (500 MHz, DMSO-d 6 ) δ 11.32 (brs, 1H), 8.44 (s, 1H), 8.29（s, 1H）, 8.22 (dd, J = 4.5, 1.5 Hz, 1H), 7.96 (s, 1H), 7.55 (d, J = 8.5 Hz, 1H), 7.44 (s, 1H), 7.27-7.22 ( m, 2H), 7.13 (dd, J = 8.5, 1.5 Hz, 1H), 7.09-7.05 (m, 1H), 6.98 (d, J = 1.5 Hz, 1H). 4.28 (t, J = 7.0 Hz, 2H), 4.06 (t, J = 7.0 Hz, 2H), 2.29-2.17 (m, 2H). ESI-MS: m/z [M+H]+ 472. Anal. Calcd for C25H18ClN5O3: C, 63.63; H, 3.84; N, 14.84. Found: C, 63.46; H, 3.97; N,14.65，
3-(呋喃[2,3-b]并吡啶-3-基)-4-(1-(3-羟丙基)-1H-吲哚-3-基)-1H-吡咯-2,5-酮(5q)
收率 12.8%， 熔点: 193-195℃. 1H NMR (500 MHz, DMSO-d 6 ) δ 11.25 (brs, 1H), 8.40 (s, 1H), 8.20 (dd, J = 4.5, 1.5 Hz, 1H), 7.95 (s, 1H), 7.53 (d, J = 8.0 Hz), 7.19 (dd, J = 8.0, 1.5 Hz, 1H), 7.09 (t, J = 8.0 Hz, 1H), 7.00-6.98 (m, 1H), 6.89 (d, J = 8.0 Hz, 1H), 6.79 (t, J = 8.0 Hz, 1H), 4.89 (t, J = 5.0 Hz, 1H), 4.33 (t, J = 7.0 Hz, 2H), 3.40-3.38 (m, 2H), 1.91-1.86 (m, 2H). ESI-MS: m/z [M+H]+ 388. Anal. Calcd for C22H17N3O4: C, 68.21; H, 4.42; N, 10.85. Found: C, 68.47; H,4.22; N, 10.74，
3-(呋喃[2,3-b]并吡啶-3-基)-4-(1-(3-吗啉丙基)-1H-吲哚-3-基)-1H-吡咯-2,5-酮
(5r)
收率 10.6%，熔点: 200-201℃. 1H NMR (500 MHz, CDCl3) δ 8.38 (brs, 1H), 8.23 (dd, J = 4.5, 1.5 Hz, 1H), 8.17 (s, 1H), 7.92 (s, 1H), 7.39 (d, J = 8.5 Hz, 1H), 7.28 (d, J = 1.5 Hz, 1H), 7.15-7.09 (m, 1H), 6.90-6.85 (m, 2H), 6.80 (t, J = 8.0Hz, 1H), 4.33 (t, J = 6.5 Hz, 2H), 3.74-3.68 (m 4H), 2.51-2.45 (m, 4H), 2.32 (t, J = 7.0 Hz, 2H), 2.01-2.04 (m, 2H). ESI-MS: m/z [M+H]+ 457. Anal. Calcd for C26H24N4O4: C, 68.41; H, 5.30; N, 12.27. Found: C, 68.59; H, 5.21; N, 12.53。
2．目标化合物对GSK-3β活性及选择性测试:
（1）酶水平的GSK-3β抑制活性测定：
通过大肠杆菌表达系统表达C端6×His融合GSK-3β蛋白，并经过Ni2+亲和纯化方法纯化，激酶活性检测采用10L反应体系的Invitrogen的Z-LYTE激酶试剂盒，每个样品3复孔。利用酶标仪Envision多标记微孔板检测仪 （PerkinElmer公司产品）检测400 nm激发下 445 nm以及 520 nm处的荧光强度，利用试剂盒提供的公式计算得到样品孔的底物磷酸化率，以此反应激酶活性高低。抑制率大于50%的用GraphPad Prism 软件拟合得出IC50值, 实验中采用的阳性化合物为星形孢菌素，部分测试结果见下表1：
表1目标化合物对GSK-3β的抑制活性
(2) 目标化合物对GSK-3β的选择性
我们选择性了部分代表性的化合物，在0.8 ug的浓度下检测了它们对PKCE，JAK2, BRAF, IKK2等激酶的抑制率，对于抑制率大于50%的化合物进一步测定了其IC50，实验中采用的阳性化合物为星形孢菌素，测试结果见下表2：
表2 目标化合物对GSK-3β的选择性
结果表明，星形孢菌素虽然对GSK-3β具有很强的抑制活性（IC50=38 nM），但其对受试的其它酶如PKCE, JAK2, BRAF, IKK2 和DraK2同样具有很强的抑制活性, 缺少选择性。本发明设计合成的化合物5n, 5o和5p对GSK-3β具有很强的抑制活性，对受试的其它酶几乎没有抑制活性，显示了很好的选择性。
细胞水平的GSK-3β抑制活性的测定
老年痴呆症患者的神经元细胞，能够分泌较多的A-beta淀粉样蛋白。该蛋白诱发神经元tau蛋白的过度磷酸化，从而引起神经元纤维缠结现象，神经元纤维缠结是老年痴呆症的主要症状。GSK-3β是磷酸化tau蛋白的主要激酶，该激酶对tau蛋白396位丝氨酸等一系列残基进行磷酸化。因此GSK-3β激酶的特异性抑制剂对减弱tau蛋白磷酸化，减轻老年痴呆症症状，具有一定的药效作用。本模型针对人神经肿瘤细胞SH-SY5Y，用A-beta淀粉样蛋白诱导tau蛋白过度磷酸化，检测GSK-3β抑制剂对tau蛋白396位丝氨酸磷酸化的影响，从而评价这些抑制剂在细胞水平的活性, 我们选择了部分化合物进行了细胞水平的GSK-3β抑制活性的测定，具体方法为: SH-SY5Y细胞用含有10%胎牛血清,1%青霉素和1%链霉素的DMEM/F12(1:1)培养基在5% CO2环境的37℃培养箱中培养。培养基每两天换一次。在12孔板中接每孔25万的SH-SY5Y细胞，培养过夜，饥饿12h后，加入浓度为1M化合物，1h后再加入20M Aβ25-35，继续处理6小时。6小时后，用冰冷的PBS洗两次后，用1×SDS buffer收样。样品10%SDS聚丙烯酰胺凝胶跑电泳，将蛋白转于PVDF膜上。膜用5%牛奶封闭1h，一抗（EPIT MICS公司的Tau Phospho(ser396)）孵育过夜，二抗为兔抗，孵育1h，用ECL曝光显影, 部分测试结果见图1。
如图1所示，化合物5n，5o，5p能够显著减少A-beta淀粉样蛋白诱发的神经元tau蛋白的过度磷酸化，在细胞水平显示了GSK-3β的抑制活性。
综上所述，3-呋喃[2,3-b]并吡啶-4-吲哚马来酰亚胺衍生物是一类高效特异性GSK-3β抑制剂，在GSK-3β介导疾病如糖尿病、双向精神障碍、阿尔茨海默病、癌症、炎性疾病、抑郁症、神经保护和精神分裂症中具有很好的应用前景。
Claims (5)
1.一种3-呋喃[2,3-b]并吡啶-4-吲哚马来酰亚胺衍生物，其特征在于该衍生物的结构通式如下：
其中：
R1，R3为一取代或二取代基，取代基独立的选自：氢；碳原子数为1-3的直链烷基、支链烷基和环烷基或碳原子数为1-3的直链烷氧基、支链烷氧基和环烷氧基或卤素；
R2为氢； 碳原子数为1-5的直链烷基、支链烷基和环烷基或(CH2)nR4，其中n=1-5, R4为吡咯、咪唑、三氮唑、吗啉、四氢吡咯、哌啶或二甲氨基。
2.根据权利要求1所述的3-呋喃[2,3-b]并吡啶-4-吲哚马来酰亚胺衍生物，其特征在于R1为H、5-F、5-Br或5-Cl，n为3，R3为H，R4为咪唑或吗啉。
3.一种根据权利要求1所述的3-呋喃[2,3-b]并吡啶-4-吲哚马来酰亚胺衍生物的制备方法，其特征在于，以如式（1）所示的取代吲哚为起始原料，依次经酰化、醇解得到如式（2）所示的中间体2、中间体2经N-烷基化得到如式（3）所示的中间体3，中间体3与如式（4）所示的取代的2-(呋喃[2,3-b]并吡啶-3-基)乙酰胺在叔丁醇钾的作用下缩合得到如式（5）所示的目标产物3-呋喃[2,3-b]并吡啶-4-吲哚马来酰亚胺衍生物，反应式如下：
。
4.根据权利要求2所述的一种3-呋喃[2,3-b]并吡啶-4-吲哚马来酰亚胺衍生物的制备方法，其特征在于包括如下步骤：
1）中间体2的制备
将取代吲哚加入溶剂无水乙醚中搅拌溶解，控制温度0-5℃，缓慢滴加草酰氯的乙醚溶液，滴完后保温反应1h，然后降温至-30 ~ -200C，滴加甲醇钠的甲醇溶液，滴完后搅拌30min，然后将反应液倒入冰水中，过滤，水洗，二氯甲烷洗，干燥后得到淡黄色中间体2；
2）中间体3的制备
将步骤1）得到的中间体2加入无水DMF中搅拌溶解，将反应液降温至0~5 ℃，分批加入70%的NaH，加完后升至室温反应30 min，然后在0℃加入卤代烃，加完后将反应液升至室温反应1h，冷却后将反应液倒入水中，后处理得到淡黄色中间体3；
3）目标产物5的制备
中间体4和步骤 2）得到的中间体3在无水THF中搅拌溶解，控制温度-5 ~ 0℃，缓慢滴入叔丁醇钾的叔丁醇溶液，滴完后升至室温反应2h，然后加入浓盐酸搅拌30 min，再将反应液倒入10% 碳酸氢钠溶液中，乙酸乙酯萃取后有机相用饱和食盐水洗涤，无水硫酸钠干燥，过滤，减压浓缩，残余物用硅胶柱层析提纯，得到目标产物5。
5.一种根据权利要求1所述的3-呋喃[2,3-b]并吡啶-4-吲哚马来酰亚胺衍生物及其药学上能接受的盐在制备GSK-3β介导疾病治疗药物中的应用。
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| CN108178764A (zh) * | 2018-01-09 | 2018-06-19 | 天津科技大学 | 呋喃并[2,3-b]吡啶类化合物及无金属催化的合成方法 |
| CN112375086A (zh) * | 2020-12-11 | 2021-02-19 | 浙江工业大学 | 一种手性螺[呋喃-3,3′-假吲哚]-2-酮类化合物的制备方法 |
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|---|---|---|---|---|
| CN105777751A (zh) * | 2016-04-26 | 2016-07-20 | 浙江工业大学 | 一种双芳基马来酰亚胺类化合物及其药学上可接受的盐及其制备方法和应用 |
| CN109535158A (zh) * | 2016-04-26 | 2019-03-29 | 浙江工业大学 | 一种双芳基马来酰亚胺类化合物及其药学上可接受的盐及其制备方法和应用 |
| CN109535158B (zh) * | 2016-04-26 | 2021-04-23 | 浙江工业大学 | 一种双芳基马来酰亚胺类化合物及其药学上可接受的盐及其制备方法和应用 |
| CN108178764A (zh) * | 2018-01-09 | 2018-06-19 | 天津科技大学 | 呋喃并[2,3-b]吡啶类化合物及无金属催化的合成方法 |
| CN112375086A (zh) * | 2020-12-11 | 2021-02-19 | 浙江工业大学 | 一种手性螺[呋喃-3,3′-假吲哚]-2-酮类化合物的制备方法 |
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