CN110156796B

CN110156796B - 异恶唑拼接3,3’-硫代吡咯啉酮双螺环氧化吲哚化合物及其制备方法及应用

Info

Publication number: CN110156796B
Application number: CN201910499289.9A
Authority: CN
Inventors: 田民义; 刘雄利; 王关炼; 常顺琴; 徐圣文; 汪军鑫; 左雄; 周英
Original assignee: Guizhou University
Current assignee: Guizhou University
Priority date: 2019-06-11
Filing date: 2019-06-11
Publication date: 2022-03-15
Anticipated expiration: 2039-06-11
Also published as: CN110156796A

Abstract

本发明公开了一种异恶唑拼接3,3'‑硫代吡咯啉酮双螺环氧化吲哚化合物，本发明以将各种取代的3‑NCS氧化吲哚、靛红衍生的硝基异恶唑烯烃化合物，按摩尔比为1:1.3的比例在有机溶剂中,在手性有机碱性小分子催化下，进行Michael加成环化反应，获得异恶唑拼接3,3'‑硫代吡咯啉酮双螺环氧化吲哚化合物。该类骨架化合物包含潜在的生物活性异恶唑基团，可以为生物活性筛选提供化合物源，对药物的筛选和制药行业具有重要的应用价值。本发明操作简单易行，原料合成便宜易得，可以在各种有机溶剂中进行，也具有较好的空气稳定性，适用性广，对于各种取代基都有很好的兼容性。且该骨架化合物对人白血病细胞生长具有抑制活性。

Description

异恶唑拼接3,3’-硫代吡咯啉酮双螺环氧化吲哚化合物及其制备方法及应用

技术领域

本发明涉及化学技术领域，尤其是一种异恶唑拼接3,3’-硫代吡咯啉酮双螺环氧化吲哚化合物及其制备方法及应用。

背景技术

根据药物设计中药效团和骨架迁越原理,把具有生物活性基团拼接到具有活性分子骨架中在有机化学和医药化学中是极其重要的研究领域。(1)多官能团氧化吲哚广泛存在天然产物和合成药物分子中，其中，尤其3,3’-吡咯双螺环氧化吲哚因为具有广泛的生物活性，吸引了许多化学工作者及医药化学团队的广泛关注，例如，化合物I是具有抗细菌活性；化合物II具有抗真菌活性，化合物III具有抗肿瘤活性。(2)异恶唑基团也普遍存在天然产物和药物分子中。例如:异噁唑基团也普遍存在天然产物和药物分子中。如:许多天然产物和药物(Cloxacillin,muscimol,Isoxicam,leflunomide,等)共享一个异恶唑分子单元，这些化合物在解除病痛、经济发展中起着重大作用。因此,根据药物设计中药效团和骨架迁越原理,鉴于3,3’-吡咯双螺环氧化吲哚骨架化合物具有潜在的生物活性，异恶唑基团属于潜在的生物活性官能团。因此，把异恶唑基团拼接到3,3’-吡咯双螺环氧化吲哚骨架，合成一系列新的潜在多活性官能团的氧化吲哚衍生物，可以为生物活性筛选提供化合物源，对药物的筛选和制药行业具有重要的应用价值(如附图6所示)。

发明内容

本发明的目的是：提供一种异恶唑拼接3,3’-硫代吡咯啉酮双螺环氧化吲哚化合物及其制备方法与应用，它是一类重要的医药中间体类似物和药物分子类似物，对药物筛选和制药行业具有重要的应用价值，且其合成方法非常经济简便。

本发明还发现该类化合物在制备防治肿瘤疾病药物中的应用。

本发明是这样实现的：异恶唑拼接3,3’-硫代吡咯啉酮双螺环氧化吲哚化合物，该化合物具有如下通式(I)的结构：

式中，R¹为苯基、甲基、乙基或苄基；R²为甲基、H或氟；R³为苯基、甲基、乙基或苄基；R⁴为H或氯或溴或甲基。

异恶唑拼接3,3’-硫代吡咯啉酮双螺环氧化吲哚化合物的制备方法，将各种取代的3-NCS氧化吲哚、靛红衍生的硝基异恶唑烯烃化合物，按摩尔比为1:1.3的比例在有机溶剂中,在手性有机碱性小分子催化下，进行Michael加成环化反应，获得异恶唑拼接3,3’-硫代吡咯啉酮双螺环氧化吲哚化合物。

合成路线举例如下：

反应机理举例如下：

其中3-NCS氧化吲哚1以及靛红衍生的硝基异恶唑烯烃化合物2的结构式，其取代基满足式中，R¹为苯基、甲基、乙基或苄基；R²为甲基、H或氟；R³为苯基、甲基、乙基或苄基；R⁴为H或氯或溴或甲基。

所述的有机小分子碱性催化剂为手性双功能金鸡纳碱或其衍生的硫脲或芳酰胺、环己基二胺衍生的硫脲或芳酰胺、1,2-二苯基二胺衍生的的硫脲或芳酰胺、等。

有机小分子碱性催化剂举例如下(但需强调的是本发明的有机小分子碱性催化剂不限于如下表示的内容)：

所述的有机溶剂为乙腈、甲苯、二氯甲烷、或氯仿。

各种取代的3-NCS氧化吲哚1、靛红衍生的硝基异恶唑烯烃化合物2，在有机溶剂中反应温度为-35-25℃，反应时间为0.5-1.0小时。

异恶唑拼接3,3’-硫代吡咯啉酮双螺环氧化吲哚化合物在制备防治肿瘤疾病药物中的应用。

通过采用上述技术方案，将各种取代的3-NCS氧化吲哚、靛红衍生的硝基异恶唑烯烃化合物，按摩尔比为1:1.3的比例在有机溶剂中,在手性有机碱性小分子催化下，进行Michael加成环化反应，获得异恶唑拼接3,3’-硫代吡咯啉酮双螺环氧化吲哚化合物，该类骨架包含潜在的生物活性异恶唑基团，可以为生物活性筛选提供化合物源，对药物的筛选和制药行业具有重要的应用价值。且该化合物对人白血病细胞(K562)具有抑制活性的作用。本发明操作简单易行，原料合成便宜易得，可以在各种有机溶剂中进行，也具有较好的空气稳定性，适用性广，对于各种取代基都有很好的兼容性。

附图说明

附图1及附图2为本发明的实施例1的化合物3aa谱图数据；

附图3及附图4为本发明的实施例1的化合物3aa液相数据；

附图5为本发明的实施例化合物3ab和3ad单晶图；

附图6位本发明的化合物设计合成路线示意。

具体实施方式

本发明的实施例：

在反应管中依次加入20.4mg N-甲基-3-NCS氧化吲哚1a(0.1mmol)，37.1mg靛红衍生的硝基异恶唑烯烃化合物2a(0.13mmol,1.3equiv.)，180mg

分子筛,3.24mg催化剂奎宁(10mmol％,0.01mmol)和25.0mL二氯甲烷溶液，-35℃反应0.5h，TLC检测基本反应完全，直接上样经柱层析(洗脱剂：V(石油醚)：V(乙酸乙酯)＝4:1)纯化得44.0mg化合物3aa，白色固体，熔点：215.2-217.0℃，dr：>20:1；产率93％,90％ee。[α]_D ²⁰＝129.64(c 0.37,CH₂Cl₂)；The ee was determined by HPLC analysis using a Chiralpak IF column(80/20hexane/i-PrOH；flow rate:1.0mL/min；λ＝254nm；τ_major＝32.94min；τ_minor＝25.33min)；核磁共振和高分辨质谱测试等结果如下：¹H NMR(DMSO-d₆,400MHz)δ:2.07(s,3H),3.10(s,3H),3.11(s,3H),5.88(s,1H),6.97-7.02(m,2H),7.08-7.12(m,1H),7.15-7.18(m,1H),7.29-7.33(m,1H),7.36-7.40(m,1H),7.64(d,J＝7.6Hz,1H),7.75(d,J＝7.2Hz,1H),11.32(brs,1H)；¹³C NMR(DMSO-d₆,100MHz)δ:11.3,27.4,27.5,49.6,70.5,72.6,109.5,110.0,123.0,123.3,125.1,126.9,127.8,130.3,131.6,144.0,145.1,155.4,166.0,173.5,173.8,200.2；HRMS(ESI-TOF)m/z:Calcd.for C₂₄H₁₉N₅NaO₅S[M+Na]⁺:512.0999；Found:512.1004.

化合物3ab至3fe的制备方法同化合物3aa，投料比与化合物3aa相同，可得到化合物3ab至3fe，反应产率和立体选择性见表1和表2，但需强调的是本发明的化合物不限于表1和表2所表示的内容。

表1为一种异恶唑拼接3,3’-硫代吡咯啉酮双螺环氧化吲哚化合物的化学结构

表2为一种异恶唑拼接3,3’-硫代吡咯啉酮双螺环氧化吲哚化合物的化学结构

本实施例制备化合物3ab:白色固体,熔点：174.5-176.1℃；产率96％；88％ee,>20:1dr,[α]_D ²⁰＝137.00(c 0.46,CH₂Cl₂)；The ee was determined by HPLC analysisusing a Chiralpak IF column(80/20hexane/i-PrOH；flow rate:1.0mL/min；λ＝254nm；τ_major＝83.70min；τ_minor＝51.02min)；核磁共振和高分辨质谱测试等结果如下：¹H NMR(DMSO-d₆,400MHz)δ:2.09(s,3H),3.12(s,3H),4.75(d,J＝16.0Hz,1H),5.00(d,J＝16.0Hz,1H),6.00(s,1H),6.77(d,J＝8.0Hz,1H),7.03(d,J＝8.0Hz,1H),7.07-7.10(m,1H),7.17-7.26(m,5H),7.33(d,J＝6.8Hz,2H),7.37-7.41(m,1H),7.70(d,J＝7.6Hz,1H),7.81(d,J＝7.6Hz,1H),11.42(br s,1H)；¹³C NMR(DMSO-d₆,100MHz)δ:11.4,27.5,43.9,49.7,70.6,72.7,110.0,110.2,124.9,127.6,127.8,127.9,129.0,131.8,136.2,144.0,144.1,155.4,165.8,173.8,200.0；HRMS(ESI-TOF)m/z:Calcd.for C₃₀H₂₃N₅NaO₅S[M+Na]⁺:588.1312；Found:588.1315.

本实施例制备化合物3ac:白色固体,熔点：139.9-142.0℃；产率92％；82％ee,>20:1dr,[α]_D ²⁰＝140.61(c 0.27,CH₂Cl₂)；The ee was determined by HPLC analysisusing a Chiralpak IF column(90/10hexane/i-PrOH；flow rate:1.0mL/min；λ＝254nm；τ_major＝49.71min；τ_minor＝18.79min)；核磁共振和高分辨质谱测试等结果如下：¹H NMR(DMSO-d₆,400MHz)δ:1.06-1.09(m,3H),2.07(s,3H),3.11(s,3H),3.66-3.68(m,2H),5.90(s,1H),7.01-7.09(m,3H),7.16-7.20(m,1H),7.26-7.31(m,1H),7.37-7.41(m,1H),7.61(d,J＝7.2Hz,1H),7.78(d,J＝7.2Hz,1H),11.32(br s,1H)；¹³C NMR(DMSO-d₆,100MHz)δ:11.3,12.6,27.4,49.4,70.4,72.5,109.6,109.9,122.8,123.3,125.3,127.1,127.8,130.4,131.6,144.1,155.4,166.0,173.1,173.8,200.0；HRMS(ESI-TOF)m/z:Calcd.forC₂₅H₂₁N₅NaO₅S[M+Na]⁺:526.1156；Found:526.1159.

本实施例制备化合物3ad:白色固体,熔点：242.7-243.6℃；产率90％；94％ee,>20:1dr,[α]_D ²⁰＝219.68(c 0.21,CH₂Cl₂)；The ee was determined by HPLC analysisusing a Chiralpak IF column(90/10hexane/i-PrOH；flow rate:1.0mL/min；λ＝254nm；τ_major＝51.34min；τ_minor＝22.04min)；核磁共振和高分辨质谱测试等结果如下：¹H NMR(DMSO-d₆,400MHz)δ:2.11(s,3H),3.12(s,3H),5.95(s,3H),6.67(d,J＝7.2Hz,1H),7.04(d,J＝7.6Hz,1H),7.10-7.14(m,1H),7.19-7.26(m,2H),7.36-7.47(m,4H),7.55-7.59(m,2H),7.66(d,J＝7.6Hz,1H),7.81(d,J＝7.2Hz,1H),11.43(br s,1H)；¹³C NMR(DMSO-d₆,100MHz)δ:11.3,27.4,49.6,70.7,72.7,109.9,125.4,126.5,127.1,128.9,130.3,131.6,134.4,144.1,144.9,155.4,166.4,173.7,200.0；HRMS(ESI-TOF)m/z:Calcd.forC₂₉H₂₁N₅NaO₅S[M+Na]⁺:574.1156；Found:574.1155.

本实施例制备化合物3ae:白色固体,熔点：158.9-160.7℃；产率90％；83％ee,>20:1dr,[α]_D ²⁰＝216.00(c 0.33,CH₂Cl₂)；The ee was determined by HPLC analysisusing a Chiralpak IF column(93/7hexane/i-PrOH；flow rate:1.0mL/min；λ＝254nm；τ_major＝23.77min；τ_minor＝31.83min)；核磁共振和高分辨质谱测试等结果如下：¹H NMR(DMSO-d₆,400MHz)δ:2.10(s,3H),3.10(s,3H),3.11(s,3H),5.86(s,1H),7.00-7.06(m,2H),7.16-7.20(m,1H),7.36-7.42(m,2H),7.56(s,1H),7.65(d,J＝7.6Hz,1H),11.43(brs,1H)；¹³C NMR(DMSO-d₆,100MHz)δ:11.3,27.4,27.7,49.4,70.5,72.7,110.0,111.1,123.3,124.9,126.2,126.5,126.7,129.6,144.0,144.3,155.5,165.9,173.2,173.7,199.4；HRMS(ESI-TOF)m/z:Calcd.for C₂₄H₁₈ClN₅NaO₅S[M+Na]⁺:546.0609；Found:546.0607.

本实施例制备化合物3af:白色固体,熔点：119.6-121.3℃；产率90％；80％ee,>20:1dr,[α]_D ²⁰＝137.34(c 0.37,CH₂Cl₂)；The ee was determined by HPLC analysisusing a Chiralpak IF column(80/20hexane/i-PrOH；flow rate:1.0mL/min；λ＝254nm；τ_major＝23.75min；τ_minor＝29.04min)；核磁共振和高分辨质谱测试等结果如下：¹H NMR(DMSO-d₆,400MHz)δ:2.11(s,3H),4.73(d,J＝16.4Hz,1H),5.04(d,J＝16.0Hz,1H),5.98(s,1H),6.82(d,J＝8.8Hz,1H),7.03(d,J＝7.6Hz,1H),7.17-7.28(m,4H),7.30-7.41(m,4H),7.62(d,J＝2.0Hz,1H),7.69(d,J＝7.6Hz,1H)；¹³C NMR(DMSO-d₆,100MHz)δ:11.4,27.4,44.1,49.6,70.6,72.8,110.0,111.8,123.4,124.7,126.9,127.6,129.0,129.7,131.9,135.8,143.2,144.1,155.5,165.7,173.5,173.6,199.2；HRMS(ESI-TOF)m/z:Calcd.for C₃₀H₂₂ClN₅NaO₅S[M+Na]⁺:622.0922；Found:622.0927.

本实施例制备化合物3ag:白色固体,熔点：252.2-254.0℃；产率93％；75％ee,>20:1dr,[α]_D ²⁰＝237.05(c 0.24,CH₂Cl₂)；The ee was determined by HPLC analysisusing a Chiralpak IC column(85/15hexane/i-PrOH；flow rate:1.0mL/min；λ＝254nm；τ_major＝18.57min；τ_minor＝37.38min)；核磁共振和高分辨质谱测试等结果如下：¹H NMR(DMSO-d₆,400MHz)δ:2.10(s,3H),3.10(s,3H),3.11(s,3H),5.86(s,1H),6.99-7.03(m,2H),7.16-7.20(m,1H),7.36-7.40(m,1H),7.52-7.55(m,1H),7.60(d,J＝7.2Hz,1H),7.66(s,1H),11.43(br s,1H)；¹³C NMR(DMSO-d₆,100MHz)δ:11.3,27.4,27.6,49.4,70.5,72.7,110.0,111.7,114.2,123.3,124.9,126.4,128.8,129.9,133.2,144.1,144.7,155.5,165.9,173.1,173.6,199.4；HRMS(ESI-TOF)m/z:Calcd.for C₂₄H₁₈BrN₅NaO₅S[M+Na]⁺:590.0104；Found:590.0109.

本实施例制备化合物3ba:白色固体,熔点：213.6-215.2℃；产率90％；87％ee,>20:1dr,[α]_D ²⁰＝98.63(c 0.39,CH₂Cl₂)；The ee was determined by HPLC analysisusing a Chiralpak IE column(85/15hexane/i-PrOH；flow rate:1.0mL/min；λ＝254nm；τ_major＝33.76min；τ_minor＝29.77min)；核磁共振和高分辨质谱测试等结果如下：¹H NMR(DMSO-d₆,500MHz)δ:1.09-1.12(m,3H),2.11(s,3H),3.14(s,3H),3.64-3.74(m,2H),5.96(s,1H),7.02(d,J＝8.0Hz,1H),7.07(d,J＝8.0Hz,1H),7.13-7.19(m,2H),7.34-7.41(m,2H),7.71(d,J＝8.0Hz,1H),7.77(d,J＝7.5Hz,1H),11.39(br s,1H)；¹³C NMR(DMSO-d₆,125MHz)δ:10.8,12.0,27.0,34.9,54.9,70.0,72.1,109.0,109.4,122.5,122.7,124.6,126.0,127.4,129.8,131.2,142.5,144.6,154.9,165.4,173.0,199.8；HRMS(ESI-TOF)m/z:Calcd.for C₂₅H₂₁N₅NaO₅S[M+Na]⁺:526.1156；Found:526.1158.

本实施例制备化合物3bd:白色固体,熔点：223.3-225.4℃；产率91％；94％ee,>20:1dr,[α]_D ²⁰＝142.01(c 0.20,CH₂Cl₂)；The ee was determined by HPLC analysisusing a Chiralpak IF column(90/10hexane/i-PrOH；flow rate:1.0mL/min；λ＝254nm；τ_major＝23.40min；τ_minor＝14.71min)；核磁共振和高分辨质谱测试等结果如下：¹H NMR(DMSO-d₆,500MHz)δ:1.15-1.18(m,3H),2.19(s,3H),3.73-3.78(m,2H),6.07(s,1H),6.76(d,J＝8.0Hz,1H),7.14(d,J＝8.0Hz,1H),7.20-7.27(m,2H),7.31-7.34(m,1H),7.44-7.48(m,3H),7.51-7.54(m,1H),7.62-7.66(m,2H),7.77(d,J＝8.0Hz,1H),7.87(d,J＝7.5Hz,1H),11.54(br s,1H)；¹³C NMR(DMSO-d₆,125MHz)δ:10.9,12.1,35.0,49.0,70.2,72.2,109.4,109.5,122.7,125.0,126.6,127.0,128.5,129.7,134.0,142.5,144.5,155.0,165.8,172.7,172.9,199.5；HRMS(ESI-TOF)m/z:Calcd.for C₃₀H₂₃N₅NaO₅S[M+Na]⁺:588.1312；Found:588.1315.

本实施例制备化合物3be:白色固体,熔点：164.8-166.2℃；产率94％；79％ee,>20:1dr,[α]_D ²⁰＝206.38(c 0.26,CH₂Cl₂)；The ee was determined by HPLC analysisusing a Chiralpak IF column(55/45hexane/i-PrOH；flow rate:1.0mL/min；λ＝254nm；τ_major＝33.77min；τ_minor＝60.19min)；核磁共振和高分辨质谱测试等结果如下：¹H NMR(DMSO-d₆,500MHz)δ:1.08-1.12(m,3H),2.14(s,3H),3.15(s,3H),3.64-3.74(m,2H),5.94(s,1H),7.06-7.10(m,2H),7.15-7.20(m,1H),7.37-7.41(m,1H),7.44-7.46(m,1H),7.63-7.66(m,2H),11.48(br s,1H)；¹³CNMR(DMSO-d₆,125MHz)δ:11.3,12.4,27.6,35.4,49.3,70.5,72.7,110.0,111.1,124.9,126.3,126.6,129.6,130.3,131.7,142.9,144.3,155.4,165.8,173.1,173.3,199.5,218.0；HRMS(ESI-TOF)m/z:Calcd.for C₂₅H₂₀ClN₅NaO₅S[M+Na]⁺:560.0766；Found:560.0769.

本实施例制备化合物3ca:白色固体,熔点：126.5-128.2℃；产率93％；77％ee,15:1dr,[α]_D ²⁰＝145.20(c 0.27,CH₂Cl₂)；The ee was determined by HPLC analysis usinga Chiralpak IC column(90/10hexane/i-PrOH；flow rate:1.0mL/min；λ＝254nm；τ_major＝17.68min；τ_minor＝34.33min)；核磁共振和高分辨质谱测试等结果如下：¹H NMR(CDCl₃,400MHz)δ:2.10(s,3H),3.17(s,3H),4.79(d,J＝15.6Hz,1H),4.89(d,J＝15.6Hz,1H),6.17(s,1H),6.68(d,J＝8.0Hz,1H),6.76(d,J＝7.6Hz,1H),7.02-7.11(m,2H),7.15-7.20(m,6H),7.24-7.28(m,1H),7.52-7.54(m,1H),7.64-7.66(m,1H),8.73(br s,1H)；¹³C NMR(CDCl₃,100MHz)δ:10.3,26.3,43.8,49.0,68.9,70.9,107.9,109.2,121.9,122.3,126.5,126.9,127.9,129.3,134.0,141.6,143.8,154.0,164.4,172.3,172.8,200.1；HRMS(ESI-TOF)m/z:Calcd.for C₃₀H₂₃N₅NaO₅S[M+Na]⁺:588.1312；Found:588.1312.

本实施例制备化合物3ce:白色固体,熔点：186.7-188.1℃；产率92％；91％ee,12:1dr,[α]_D ²⁰＝220.92(c 0.29,CH₂Cl₂)；The ee was determined by HPLC analysis usinga Chiralpak IC column(75/25hexane/i-PrOH；flow rate:1.0mL/min；λ＝254nm；τ_major＝18.71min；τ_minor＝46.66min)；核磁共振和高分辨质谱测试等结果如下：¹H NMR(CDCl₃,400MHz)δ:2.13(s,3H),3.13(s,3H),4.78(d,J＝15.6Hz,1H),4.87(d,J＝15.6Hz,1H),6.15(s,1H),6.67-6.71(m,1H),7.05-7.09(m,1H),7.14-7.25(m,7H),7.50-7.52(m,1H),7.57(d,J＝7.6Hz,1H),8.83(br s,1H)；¹³C NMR(CDCl₃,100MHz)δ:10.3,26.5,43.9,48.7,68.8,71.0,108.8,109.3,125.4,126.5,126.9,127.1,127.9,129.3,141.7,142.4,154.1,164.1,171.9,172.7,199.2；HRMS(ESI-TOF)m/z:Calcd.for C₃₀H₂₂ClN₅NaO₅S[M+Na]⁺:622.0922；Found:622.0925.

本实施例制备化合物3cf:白色固体,熔点：200.1-201.7℃；产率94％；76％ee,>20:1dr,[α]_D ²⁰＝293.56(c 0.15,CH₂Cl₂)；The ee was determined by HPLC analysisusing a Chiralpak IC column(65/35hexane/i-PrOH；flow rate:1.0mL/min；λ＝254nm；τ_major＝38.46min；τ_minor＝25.56min)；核磁共振和高分辨质谱测试等结果如下：¹H NMR(DMSO-d₆,400MHz)δ:2.13(s,3H),4.73-4.83(m,2H),4.94-5.08(m,2H),6.10(s,1H),6.83-6.88(m,2H),7.18-7.36(m,13H),7.66(s,1H),7.75(d,J＝7.2Hz,1H),11.79(br s,1H)；¹³CNMR(DMSO-d₆,100MHz)δ:11.4,44.0,49.9,70.7,73.0,110.7,111.8,124.9,127.0,127.6,127.7,129.0,129.1,135.8,135.9,142.9,143.3,155.6,165.4,173.5,174.0,199.5；HRMS(ESI-TOF)m/z:Calcd.for C₃₆H₂₆ClN₅NaO₅S[M+Na]⁺:698.1235；Found:698.1242.

本实施例制备化合物3ch:白色固体,熔点：126.5-127.8℃；产率96％；70％ee,>20:1dr,[α]_D ²⁰＝54.69(c 0.55,CH₂Cl₂)；The ee was determined by HPLC analysisusing a Chiralpak IC column(65/35hexane/i-PrOH；flow rate:1.0mL/min；λ＝254nm；τ_major＝77.23min；τ_minor＝41.94min)；核磁共振和高分辨质谱测试等结果如下：¹H NMR(DMSO-d₆,400MHz)δ:2.10(s,3H),2.29(s,3H),4.72-4.84(m,2H),4.93-5.01(m,2H),6.12(s,1H),6.67(d,J＝8.0Hz,1H),6.87(d,J＝8.0Hz,1H),7.01(d,J＝7.6Hz,1H),7.18-7.33(m,12H),7.49(s,1H),7.85(d,J＝6.4Hz,1H),11.69(br s,1H)；¹³C NMR(DMSO-d₆,100MHz)δ:11.4,21.3,43.9,50.1,70.8,72.8,110.1,110.7,127.5,127.6,127.7,129.0,129.1,136.0,136.2,141.8,142.9,155.4,165.5,173.8,174.2,200.4；HRMS(ESI-TOF)m/z:Calcd.for C₃₇H₂₉N₅NaO₅S[M+Na]⁺:678.1782；Found:678.1780.

本实施例制备化合物3de:白色固体,熔点：140.8-142.1℃；产率94％；77％ee,>20:1dr,[α]_D ²⁰＝296.52(c 0.12,CH₂Cl₂)；The ee was determined by HPLC analysisusing a Chiralpak IC column(82/18hexane/i-PrOH；flow rate:1.0mL/min；λ＝254nm；τ_major＝36.02min；τ_minor＝14.39min)；核磁共振和高分辨质谱测试等结果如下：¹H NMR(DMSO-d₆,400MHz)δ:2.14(s,3H),3.13(s,3H),5.97(s,1H),6.68(d,J＝8.0Hz,1H),7.07-7.09(m,1H),7.20-7.23(m,1H),7.29-7.33(m,1H),7.36(d,J＝6.8Hz,2H),7.43-7.50(m,2H),7.58-7.61(m,2H),7.67(s,1H),7.72(d,J＝7.6Hz,1H),11.53(br s,1H)；¹³C NMR(DMSO-d₆,100MHz)δ:11.3,27.7,49.6,70.6,72.9,110.3,111.2,124.5,126.7,127.0,129.9,130.4,134.1,143.9,144.3,155.6,166.2,173.2,173.3,199.5；HRMS(ESI-TOF)m/z:Calcd.for C₂₉H₂₀ClN₅NaO₅S[M+Na]⁺:608.0766；Found:608.0769.

本实施例制备化合物3ea:白色固体,熔点：249.8-251.6℃；产率95％；96％ee,>20:1dr,[α]_D ²⁰＝93.77(c 0.41,CH₂Cl₂)；The ee was determined by HPLC analysisusing a Chiralpak IF column(98/2hexane/i-PrOH；flow rate:1.0mL/min；λ＝254nm；τ_major＝24.56min；τ_minor＝16.83min)；核磁共振和高分辨质谱测试等结果如下：¹H NMR(DMSO-d₆,500MHz)δ:2.11(s,3H),3.11(s,3H),3.14(s,3H),5.91(s,1H),6.93(d,J＝8.0Hz,1H),7.01(d,J＝7.5Hz,1H),7.13-7.17(m,1H),7.21-7.23(m,1H),7.32-7.36(m,1H),7.53(s,1H),7.65(d,J＝7.5Hz,1H),11.34(br s,1H)；¹³C NMR(DMSO-d₆,125MHz)δ:10.8,20.7,26.9,27.0,49.1,70.0,72.2,82.3,109.0,109.2,122.5,124.8,126.7,127.3,129.8,131.8,141.2,144.6,154.9,165.5,173.1,173.2,187.3,199.7,217.3；HRMS(ESI-TOF)m/z:Calcd.for C₂₅H₂₁N₅NaO₅S[M+Na]⁺:526.1156；Found:526.1162.

本实施例制备化合物3ed:白色固体,熔点：125.5-127.2℃；产率94％；92％ee,>20:1dr,[α]_D ²⁰＝170.00(c 0.19,CH₂Cl₂)；The ee was determined by HPLC analysisusing a Chiralpak IF column(98/2hexane/i-PrOH；flow rate:1.0mL/min；λ＝254nm；τ_major＝42.45min；τ_minor＝15.71min)；核磁共振和高分辨质谱测试等结果如下：¹H NMR(DMSO-d₆,500MHz)δ:2.19(s,3H),2.46(s,3H),3.16(s,3H),6.02(s,1H),6.75(d,J＝7.5Hz,1H),7.00(d,J＝8.0Hz,1H),7.21-7.24(m,1H),7.28-7.33(m,2H),7.44(d,J＝7.5Hz,2H),7.52-7.55(m,1H),7.63-7.66(m,3H),7.71(d,J＝7.5Hz,1H),11.47(br s,1H)；¹³C NMR(DMSO-d₆,125MHz)δ:11.3,21.2,27.4,49.6,70.6,72.7,109.7,109.8,125.5,127.0,127.3,130.2,130.3,131.8,132.3,134.4,141.7,144.9,155.4,166.3,173.1,173.6,199.7,218.0；HRMS(ESI-TOF)m/z:Calcd.forC₃₀H₂₃N₅NaO₅S[M+Na]⁺:588.1312；Found:588.1317.

本实施例制备化合物3fa:白色固体,熔点：125.2-127.0℃；产率92％；83％ee,>20:1dr,[α]_D ²⁰＝151.30(c 0.19,CH₂Cl₂)；The ee was determined by HPLC analysisusing a Chiralpak IE column(80/20hexane/i-PrOH；flow rate:1.0mL/min；λ＝254nm；τ_major＝19.21min；τ_minor＝13.82min)；核磁共振和高分辨质谱测试等结果如下：¹H NMR(DMSO-d₆,500MHz)δ:2.12(s,3H),3.13(s,3H),3.14(s,3H),5.91(s,1H),7.00(d,J＝8.0Hz,1H),7.07-7.09(m,1H),7.12-7.15(m,1H),7.29-7.35(m,2H),7.61-7.65(m,2H),11.37(br s,1H)；¹³C NMR(DMSO-d₆,125MHz)δ:10.8,27.0,27.1,49.2,70.0,72.2,109.0,110.5,114.3(d,J_CF＝25.0Hz),117.6(d,J_CF＝22.5Hz),122.6,125.8,126.2,126.3,127.1,129.9,131.3,140.0,144.6,155.0,158.3(d,J_CF＝237.5Hz),165.4,172.9,173.3,199.9；HRMS(ESI-TOF)m/z:Calcd.for C₂₄H₁₈FN₅NaO₅S[M+Na]⁺:530.0905；Found:530.0909.

本实施例制备化合物3fd:白色固体,熔点：187.8-188.6℃；产率93％；94％ee,>20:1dr,[α]_D ²⁰＝187.10(c 0.13,CH₂Cl₂)；The ee was determined by HPLC analysisusing a Chiralpak IC column(85/15hexane/i-PrOH；flow rate:1.0mL/min；λ＝254nm；τ_major＝29.02min；τ_minor＝20.26min)；核磁共振和高分辨质谱测试等结果如下：¹H NMR(DMSO-d₆,500MHz)δ:2.21(s,3H),3.19(s,3H),6.01(s,1H),6.73(d,J＝8.0Hz,1H),7.13-7.16(m,1H),7.19-7.22(m,1H),7.29-7.32(m,1H),7.36-7.40(m,1H),7.43(d,J＝7.5Hz,2H),7.51-7.54(m,1H),7.63-7.66(m,2H),7.69-7.74(m,2H),11.50(br s,1H)；¹³C NMR(DMSO-d₆,125MHz)δ:11.2,27.6,49.6,70.6,72.7,109.7,110.9,114.9(d,J_CF＝25.0Hz),118.1(d,J_CF＝23.8Hz),123.6,126.5,126.9,127.0,127.2,128.9,130.2,130.4,131.8,134.4,140.5,144.8,155.5,158.8(d,J_CF＝238.8Hz),166.2,173.0,173.6,200.0；HRMS(ESI-TOF)m/z:Calcd.for C₂₉H₂₀FN₅NaO₅S[M+Na]⁺:592.1061；Found:592.1068.

本实施例制备化合物3fe:白色固体,熔点：167.7-168.2℃；产率91％；83％ee,12:1dr,[α]_D ²⁰＝66.02(c 0.27,CH₂Cl₂)；The ee was determined by HPLC analysis using aChiralpak IF column(60/40hexane/i-PrOH；flow rate:1.0mL/min；λ＝254nm；τ_major＝29.62min；τ_minor＝25.50min)；核磁共振和高分辨质谱测试等结果如下：¹H NMR(DMSO-d₆,500MHz)δ:2.15(s,3H),3.13(s,3H),3.14(s,3H),5.88(s,1H),7.05-7.08(m,2H),7.28-7.32(m,1H),7.42-7.44(m,1H),7.60-7.62(m,1H),7.64(d,J＝2.0Hz,1H),11.44(br s,1H)；¹³C NMR(DMSO-d₆,125MHz)δ:10.8,27.1,27.2,49.1,70.0,72.3,110.5,114.2(d,J_CF＝26.3Hz),117.6(d,J_CF＝22.5Hz),125.9,126.2,128.9,129.8,139.9,143.8,155.1,158.2(d,J_CF＝237.5Hz),165.2,172.6,173.1,199.0；HRMS(ESI-TOF)m/z:Calcd.forC₂₄H₁₇ClFN₅NaO₅S[M+Na]⁺:564.0515；Found:564.0519.

本发明的式(1)化合物具有重要的生物活性，体外对人白血病细胞(K562)的细胞毒性试验表明：此类式(1)所示的结构的异恶唑拼接3,3’-硫代吡咯啉酮双螺环氧化吲哚化合物对肿瘤细胞生长具有抑制作用，有可能发展成为新的防治肿瘤药物。但需强调的是本发明的化合物不限于人白血病细胞(K562)表示的细胞毒性。

药理实施例：化合物3af,3ce,3cf和3fe对K562细胞的细胞毒性

K562(人慢性髓系白血病细胞)用RPMI-1640培养基培养，培养基中含10％的胎牛血清，100U/mL的青霉素和100U/mL链霉素。细胞以每孔5000个细胞的浓度加入到96孔中，在37℃含5％CO₂潮湿空气的培养箱中培养24小时。

细胞存活率的测定用改良MTT法。细胞经过24小时的孵育后，分别将新配的化合物3af,3ce,3cf和3fe的二甲基亚砜溶液以浓度梯度加入到各孔中，使孔中化合物最终浓度分别为5μmol/L,10μmol/L,20μmol/L,40μmol/L和80μmol/L。48小时后，每孔加入10μL MTT(5mg/mL)的磷酸盐缓冲液，再继续在37℃培养4小时后，离心5分钟除去未转化的MTT，每孔中加入150μL二甲基亚砜。以溶解还原的MTT晶体甲臜(formazan)，用酶标仪在490nm波长测定OD值。化合物3af对K562肿瘤细胞的IC₅₀为47.1μmol/L；化合物3ce对K562肿瘤细胞的IC₅₀为32.4μmol/L；化合物3cf对K562肿瘤细胞的IC₅₀为25.3μmol/L；化合物3fe对K562肿瘤细胞的IC₅₀为23.6μmol/L；而阳性对照顺铂对K562肿瘤细胞的IC₅₀为21.0μmol/L。

实验结论：K562细胞是测试化合物对肿瘤细胞的细胞毒性的有效工具和评价指标。本实验表明此类式(1)所示的异恶唑拼接3,3’-硫代吡咯啉酮双螺环氧化吲哚化合物对K562细胞具有较强的细胞毒性，和肿瘤治疗一线用药顺铂同一数量级，有可能发展成新的具有抗肿瘤作用的药物。可见这些化合物具有开发成为抗肿瘤药物的潜力，值得继续深入研究下去。

Claims

1.异恶唑拼接3,3'-硫代吡咯啉酮双螺环氧化吲哚化合物，其特征在于：该化合物具有如通式(Ⅰ)所示的结构：

2.一种如权利要求1所述的异恶唑拼接3,3'-硫代吡咯啉酮双螺环氧化吲哚化合物的制备方法，其特征在于：其合成路线如下：

3.一种如权利要求1所述的异恶唑拼接3,3'-硫代吡咯啉酮双螺环氧化吲哚化合物在制备防治白血病药物中的应用。