本發(fā)明涉及化學(xué)技術(shù)領(lǐng)域，尤其是一種含二氟甲基的螺[環(huán)丙烷-1,3'-吲哚啉]-2'-酮類化合物和藥物用途。

背景技術(shù)：

螺環(huán)氧化吲哚結(jié)構(gòu)單元廣泛存在于許多天然分子和臨床藥物分子中，在已發(fā)現(xiàn)的含有3,3'-螺環(huán)結(jié)構(gòu)的氧化吲哚類化合物中，很多都具有較高的生物活性，諸如抗腫瘤、抗病毒、抗hiv、抗氧化性等。其中，含有氧化吲哚螺環(huán)環(huán)丙烷結(jié)構(gòu)單元的分子更加引人注目，由于該類化合物可以作為黃體酮受體拮抗劑、rsv抑制劑、激酶抑制劑、抗腫瘤藥物等。

在另一方面，二氟甲基(-cf2h)，作為一個(gè)重要的含氟基團(tuán)，含有一個(gè)弱酸化的c-h鍵，是羥基、巰基及羥甲基的電子等排體，能夠調(diào)節(jié)分子的生物活性、代謝穩(wěn)定性以及親脂性等，在醫(yī)藥、農(nóng)藥等領(lǐng)域起著與其他氟烷基同樣重要的作用。因此，在分子中引入二氟甲基可賦予其諸多優(yōu)良性能，同時(shí)引入或保持了生物靶點(diǎn)的關(guān)鍵識(shí)別元素。這些決定了其在生物醫(yī)藥、生物成像等領(lǐng)域的巨大潛力和科研價(jià)值。

鑒于氧化吲哚螺環(huán)環(huán)丙烷化合物具有潛在的生物活性以及二氟甲基在藥物化學(xué)中的重要作用，因此，以二氟甲基重氮甲烷為砌塊，通過[3+2]反應(yīng)和縮環(huán)反應(yīng)策略與3-烯基氧化吲哚反應(yīng)，合成含有二氟甲基基團(tuán)的3,3'-氧化吲哚螺環(huán)環(huán)丙烷可能會(huì)產(chǎn)生一系列結(jié)構(gòu)和活性上有意義的新化合物，它們的合成可以為含二氟甲基的螺[環(huán)丙烷-1,3'-吲哚啉]-2'-酮類化合物在藥物發(fā)現(xiàn)和成藥性評(píng)價(jià)中的應(yīng)用建立物質(zhì)基礎(chǔ)。

技術(shù)實(shí)現(xiàn)要素：

本發(fā)明的目的在于提供一種含二氟甲基的螺[環(huán)丙烷-1,3'-吲哚啉]-2'-酮類化合物及其制備防治腫瘤疾病藥物的應(yīng)用，該類化合物是一類重要的藥物活性分子，對(duì)藥物篩選和制藥行業(yè)具有重要的應(yīng)用價(jià)值，并且該類分子的合成具有反應(yīng)條件溫和、操作簡(jiǎn)單、底物普適性好的優(yōu)點(diǎn)。

本發(fā)明是這樣實(shí)現(xiàn)的：原位生成的二氟甲基重氮甲烷與靛紅衍生的3-烯基氧化吲哚在無催化劑條件下，在有機(jī)溶劑中進(jìn)行[3+2]反應(yīng)和縮環(huán)反應(yīng)，獲得含二氟甲基的螺[環(huán)丙烷-1,3'-吲哚啉]-2'-酮類化合物。該類化合物具有如通式(ⅰ)所示的結(jié)構(gòu)：

式中，r¹為烷基、烷氧基或鹵素，r²為烷基，r³為烷基、芳基或?；?。

所述的有機(jī)溶劑為二氯甲烷、三氯甲烷、四氯化碳、乙腈、四氫呋喃、苯、甲苯、二甲苯、均三甲苯、1,4-二氧六環(huán)、乙二醇二甲醚、乙二醇二乙醚或甲基叔丁基醚。

原位生成的二氟甲基重氮甲烷與靛紅衍生的3-烯基氧化吲哚的反應(yīng)溫度為25-130℃，反應(yīng)時(shí)間為1-50小時(shí)。

所述的含二氟甲基的螺[環(huán)丙烷-1,3'-吲哚啉]-2'-酮類化合物在制備防治腫瘤疾病藥物的應(yīng)用。

本發(fā)明的反應(yīng)原理如下：

其中，r¹，r²，r³和有機(jī)溶劑如上所述。

通過采用上述技術(shù)方案，二氟乙胺(1)放置于反應(yīng)瓶中，依次加入有機(jī)溶劑、亞硝酸叔丁酯和乙酸，并在70℃條件下反應(yīng)10分鐘，停止加熱，即可原位得到二氟甲基重氮甲烷(2)。然后將靛紅衍生的3-烯基氧化吲哚(3)加入反應(yīng)瓶中，經(jīng)過[3+2]反應(yīng)和縮環(huán)反應(yīng)，合成了一系列含二氟甲基的螺[環(huán)丙烷-1,3'-吲哚啉]-2'-酮類化合物(4)。該類化合物含有潛在生物活性的二氟甲基基團(tuán)和3,3'-氧化吲哚螺環(huán)環(huán)丙烷骨架，可以為藥物發(fā)現(xiàn)和成藥性評(píng)價(jià)建立物質(zhì)基礎(chǔ)，具有重要的應(yīng)用價(jià)值。本發(fā)明原料易于獲得，可以在各種有機(jī)溶劑中進(jìn)行，具有反應(yīng)條件溫和、操作簡(jiǎn)單、底物普適性好的優(yōu)點(diǎn)。

附圖說明

附圖1為本發(fā)明的實(shí)施例化合物4a的單晶-x射線衍射結(jié)構(gòu)。

附圖2為本發(fā)明的實(shí)施例化合物4g的單晶-x射線衍射結(jié)構(gòu)。

附圖3為本發(fā)明的實(shí)施例化合物4r的單晶-x射線衍射結(jié)構(gòu)。

附圖4-6為本發(fā)明的實(shí)施例化合物4a的核磁共振譜圖。

具體實(shí)施方式

下面結(jié)合實(shí)施例及附圖進(jìn)一步介紹本發(fā)明，但本發(fā)明不僅限于下述實(shí)施例，可以預(yù)見本領(lǐng)域技術(shù)人員在結(jié)合現(xiàn)有技術(shù)的情況下，實(shí)施情況可能產(chǎn)生種種變化。

本發(fā)明的實(shí)施例1：含二氟甲基的螺[環(huán)丙烷-1,3'-吲哚啉]-2'-酮(4)的制備；化合物4a：在圓底燒瓶中，依次加入二氯甲烷(15ml)，二氟乙胺(1,97.3mg,1.2mmol)，亞硝酸叔丁酯(148.5mg,1.44mmol)，乙酸(14.5mg,0.24mmol)，在70℃條件下反應(yīng)10分鐘，停止加熱，即可原位制備獲得二氟甲基重氮甲烷(2)。再加入靛紅衍生的3-烯基氧化吲哚(3a，92.5mg,0.4mmol)，并在室溫下充分?jǐn)嚢?8小時(shí)，濃縮，再加入甲苯(15ml)，120℃條件下反應(yīng)2小時(shí)，薄層色譜檢測(cè)反應(yīng)完畢后，硅膠柱層析(300-400目)分離(石油醚：乙酸乙酯＝5：1)，得到淺黃色固體103.9mg，產(chǎn)率88％，熔點(diǎn)：88.9–90.0℃，核磁共振和高分辨質(zhì)譜測(cè)試分析結(jié)果如下：94:6trans/cis(determinedby¹⁹fnmranalysis)；¹hnmr(400mhz,cdcl3)δ(major)7.36-7.31(m,2h),7.06(t,j＝7.8hz,1h),6.91(d,j＝7.8hz,1h),6.40(td,j＝6.0,56.0hz,1h),4.19-4.11(m,2h),3.28(s,3h),2.99-2.95(m,2h),1.22(t,j＝7.2hz,3h)；¹³cnmr(100mhz,cdcl3)δ(major+minor)173.1,172.3,167.4,166.8,144.8,144.2,132.5,128.7,128.6,124.3,124.0,122.9,122.8,113.4(t,j＝236.3hz),108.9,108.5,108.2,62.0,61.9,35.8(d,j＝8.6hz),34.9(d,j＝9.7hz),34.6(t,j＝33.7hz),33.3(t,j＝33.9hz),32.8(d,j＝7.9hz),27.0,27.9,25.9,14.2,14.1；¹⁹fnmr(376mhz,cdcl3)δ(major+minor)-111.04(abd,j＝54.2,293.9hz,major),-112.94(abd,j＝55.4,297.4hz,minor),-114.43(abd,j＝54.1,297.6hz,minor),-115.88(abd,j＝54.1,293.4hz,major)；hrms(esi-tof)calcd.forc15h16f2no3[m+h]⁺:296.1093；found:296.1101.

附圖中單晶-x射線衍射結(jié)構(gòu)圖和核磁共振譜圖由現(xiàn)有技術(shù)軟件生成，本領(lǐng)域技術(shù)人員可以通過圖中的特點(diǎn)即可了解各化合物屬性，對(duì)于圖中的數(shù)字可能因提交原因不是很清晰，但不影響本發(fā)明的完全公開。

通過實(shí)施例制備的化合物4b～4s的制備方法同化合物4a，投料比與化合物4a相同，可得到化合物4b～4s，反應(yīng)產(chǎn)率見表1，但需強(qiáng)調(diào)的是本發(fā)明的化合物不限于表1所表示的內(nèi)容。

表1為含二氟甲基的螺[環(huán)丙烷-1,3'-吲哚啉]-2'-酮類化合物的化學(xué)結(jié)構(gòu)

表1：

本實(shí)施例制備化合物4b：紅色油狀物，產(chǎn)率91％；核磁共振和高分辨質(zhì)譜測(cè)試分析結(jié)果如下：93:7trans/cis(determinedby¹⁹fnmranalysis)；¹hnmr(400mhz,cdcl3)δ(major+minor)7.18(s,0.07h),7.16(s,0.93h),7.12(d,j＝7.9hz,0.93h),6.83(d,j＝7.9hz,0.07h),6.79(d,j＝7.9hz,0.93h),6.68(d,j＝7.9hz,0.07h),6.40(td,j＝7.2,55.8,hz,1h),4.36-4.30(m,0.14h),4.24-4.07(m,1.86h),3.25(d,j＝1.6hz,2.79h),3.20(d,j＝1.6hz,0.21h),2.99-2.90(m,1.86h),2.86-2.83(m,0.14h),2.34(s,0.21h),2.32(s,2.79h),1.39-1.35(m,0.21h),1.24-1.20(m,2.79h)；¹³cnmr(100mhz,cdcl3)δ(major+minor)172.2,167.6,166.8(d,j＝2.0hz),165.9,141.8,132.9,132.4,129.4,128.9,125.0,124.0,123.6,122.2,113.4(t,j＝236.3hz),108.2,61.9,61.2,35.9(d,j＝8.6hz),34.8(d,j＝9.7hz),34.5(t,j＝33.7hz),26.9,26.3,21.4,21.3,21.2,14.3,14.2；¹⁹fnmr(376mhz,cdcl3)δ(major+minor)-111.03(abdd,j＝5.9,54.7,294.3hz,major),-112.87(abdd,j＝8.6,55.9,297.2hz,minor)-114.51(abdd,j＝7.1,53.6,297.3hz,minor),-115.84(abdd,j＝7.0,56.1,294.6hz,major)；hrms(esi-tof)calcd.forc16h18f2no3[m+h]⁺:310.1249；found:310.1253.

本實(shí)施例制備化合物4c：橙色油狀物，產(chǎn)率81％；核磁共振和高分辨質(zhì)譜測(cè)試分析結(jié)果如下：>99:1trans/cis(determinedby¹⁹fnmranalysis)；¹hnmr(400mhz,cdcl3)δ7.16(dd,j＝2.6,8.6hz,1h),7.02(td,j＝2.6,8.6hz,1h),6.79(dd,j＝8.0,8.6hz,1h),6.38(td,j＝7.4,55.2hz,1h),4.25-4.09(m,2h),3.26(s,3h),3.02-3.01(m,1h),2.96-2.89(m,1h),1.23(t,j＝7.2hz,3h)；¹³cnmr(100mhz,cdcl3)δ172.0,166.7(d,j＝2.0hz,1c),159.2(d,j＝239.2hz,1c),140.2(d,j＝1.8hz,1c),125.6(d,j＝9.5hz,1c),114.9(d,j＝23.6hz,1c),113.2(t,j＝236.5hz,1c),111.4(d,j＝26.8hz,1c),108.9(d,j＝8.3hz,1c),62.2,35.9(dd,j＝2.0,8.6hz,1c),34.9(d,j＝4.5hz,1c),34.9(t,j＝28.9hz,1c),27.0,14.1；¹⁹fnmr(376mhz,cdcl3)δ-111.07(abddd,j＝1.6,5.9,54.7,295.3hz,1f),-116.01(abdd,j＝7.2,55.6,295.2hz,1f),-119.47(td,j＝4.1,8.8hz,1f)；hrms(esi-tof)calcd.forc15h15f3no3[m+h]⁺:314.0999；found:314.1010.

本實(shí)施例制備化合物4d：橙色油狀物，產(chǎn)率71％；核磁共振和高分辨質(zhì)譜測(cè)試分析結(jié)果如下：>99:1trans/cis(determinedby¹⁹fnmranalysis)；¹hnmr(400mhz,cdcl3)δ(major)7.37(d,j＝2.0hz,1h),7.02(dd,j＝2.0,8.4hz,1h),6.82(d,j＝8.4hz,1h),6.37(td,j＝7.2,55.2hz,1h),4.26-4.10(m,2h),3.26(s,3h),3.02-2.91(m,2h),1.24(t,j＝7.2hz,3h)；¹³cnmr(100mhz,cdcl3)δ(major+minor)172.3,166.7,145.3,134.7,125.3,124.0,112.7,122.3,113.2(t,j＝236.6hz),109.7,109.3,62.2,35.6(d,j＝8.6hz),34.9(d,j＝8.6hz),34.7(t,j＝34.0hz),27.2,27.0,14.2；¹⁹fnmr(376mhz,cdcl3)δ(major)-111.06(abdd,j＝5.6,54.5,295.4hz,major,1f),-116.01(abdd,j＝7.0,55.8,295.6hz,major,1f)；hrms(esi-tof)calcd.forc15h15clf2no3[m+h]⁺:330.0703；found:330.0708.

本實(shí)施例制備化合物4e：黃色油狀物，產(chǎn)率85％；核磁共振和高分辨質(zhì)譜測(cè)試分析結(jié)果如下：97:3trans/cis(determinedby¹⁹fnmranalysis)；¹hnmr(400mhz,cdcl3)δ(major)7.49(s,1h),7.44(dd,j＝1.9,8.4hz,1h),6.77(d,j＝8.4hz,1h),6.36(td,j＝7.2,55.3hz,1h),4.26-4.10(m,2h),3.25(s,3h),3.01-2.93(m,2h),1.24(t,j＝7.2hz,3h)；¹³cnmr(100mhz,cdcl3)δ(major)171.7,166.5,143.2,131.5,126.2,125.9,115.5,113.1(t,j＝236.7hz,1c),109.9,62.2,35.5(d,j＝8.6hz,1c),35.1(d,j＝9.5hz,1c),35.0(t,j＝34.0hz,1c),27.0,14.2；¹⁹fnmr(376mhz,cdcl3)δ(major+minor)-111.02(abdd,j＝3.7,54.6,295.3hz,major),-112.80(abdd,j＝8.2,55.9,298.9hz,minor),-114.42(abdd,j＝7.2,53.4,298.9hz,minor),-115.98(abdd,j＝6.7,55.8,295.7hz,major)；hrms(esi-tof)calcd.forc15h15brf2no3[m+h]⁺:374.0198；found:374.0204.

本實(shí)施例制備化合物4f：黃色油狀物，產(chǎn)率80％；核磁共振和高分辨質(zhì)譜測(cè)試分析結(jié)果如下：97:3trans/cis(determinedby¹⁹fnmranalysis)；¹hnmr(400mhz,cdcl3)δ(major)7.29(s,1h),7.20(d,j＝8.2hz,1h),6.89(d,j＝8.4hz,1h),6.38(td,j＝7.0,55.2hz,1h),4.20-4.11(m,2h),3.28(s,3h),3.03-2.94(m,2h),1.23-1.19(m,3h)；¹³cnmr(100mhz,cdcl3)δ(major)172.1,166.5,144.8,142.9,125.5,121.9,120.6(q,j＝255.2hz,1c),117.3,113.1(t,j＝236.2hz),108.9,62.3,35.7(d,j＝8.6hz),35.1(d,j＝9.5hz),34.9(t,j＝34.1hz),27.0,14.0；¹⁹fnmr(376mhz,cdcl3)δ(major+minor)-58.54(s,3f),-111.14(abdd,j＝5.6,54.6,295.2hz,major),-113.01(abdd,j＝8.3,55.9,299.7hz,minor),-114.46(abdd,j＝7.4,53.4,299.3hz,minor),-116.14(abdd,j＝7.1,55.6,295.6hz,major)；hrms(esi-tof)calcd.forc16h15f5no4[m+h]⁺:380.0916；found:380.0922.

本實(shí)施例制備化合物4g：黃色固體，產(chǎn)率92％；熔點(diǎn):102.0–103.8℃；核磁共振和高分辨質(zhì)譜測(cè)試分析結(jié)果如下：89:11trans/cis(determinedby¹⁹fnmranalysis)；¹hnmr(400mhz,cdcl3)δ(major+minor)7.39(d,j＝8.2hz,0.11h),7.28(d,j＝8.2hz,0.89h),7.05(d,j＝1.9hz,0.11h),7.02(dd,j＝1.8,8.2hz,0.89h),6.94(d,j＝1.8hz,0.11h),6.90(d,j＝1.8hz,0.89h),6.37(td,j＝7.1,55.2hz,1h),4.34-4.29(m,0.22h),4.23-4.07(m,1.78h),3.26(s,2.67h),3.22(s,0.33h)3.00-2.93(m,1.78h),2.65-2.57(m,0.22h),1.36(t,j＝7.2hz,0.33h),1.22(t,j＝7.2hz,2.67h)；¹³cnmr(100mhz,cdcl3)δ(major+minor)172.3,166.7,145.3,134.7,125.3,124.0,122.7,122.6,122.3,113.2(t,j＝236.6hz),109.7,109.3,62.2,62.1,35.6(d,j＝8.6hz),35.0(d,j＝8.6hz),34.7(t,j＝34.0hz),32.9(d,j＝7.6hz),27.2,27.0,14.2,14.1；¹⁹fnmr(376mhz,cdcl3)δ(major+minor)-111.03(abdd,j＝5.6,54.6,295.2hz,major),-113.05(abdd,j＝8.2,55.8,298.8hz,minor),-114.40(abdd,j＝7.4,53.6,298.7hz,minor),-115.96(abdd,j＝7.0,55.6,295.3hz,major)；hrms(esi-tof)calcd.forc15h15clf2no3[m+h]⁺:330.0703；found:330.0713.

本實(shí)施例制備化合物4h：黃色固體，產(chǎn)率83％；熔點(diǎn):103.7–105.2℃；核磁共振和高分辨質(zhì)譜測(cè)試分析結(jié)果如下：92:8trans/cis(determinedby¹⁹fnmranalysis)；¹hnmr(400mhz,cdcl3)δ(major+minor)7.32(d,j＝8.2hz,0.08h),7.22(d,j＝8.2hz,0.92h),7.18-7.16(m,1h),7.09(d,j＝1.6hz,0.08h),7.04(d,j＝1.6hz,0.92h),6.37(td,j＝7.2,55.1hz,1h),4.32-4.28(m,0.16h),4.23-4.06(m,1.84h),3.26(s,2.76h),3.21(s,0.24h),3.00-2.91(m,1.84h),2.65-2.57(m,0.16h),1.22(t,j＝7.2hz,2.76h),0.95(t,j＝7.3hz,0.24h)；¹³cnmr(100mhz,cdcl3)δ(major+minor)172.2,166.7,145.4,125.6,125.5,124.3,122.9,122.4,113.1(t,j＝236.6,hz),112.4,112.1,62.1,35.6(d,j＝8.7hz),35.0(d,j＝9.6hz),34.7(t,j＝33.9hz),27.1,27.0,14.2；¹⁹fnmr(376mhz,cdcl3)δ(major+minor)-111.03(abdd,j＝3.9,54.6,295.1hz,major),-113.05(abdd,j＝6.6,55.8,298.6hz,minor),-114.40(abdd,j＝7.1,55.5,298.8hz,minor),-115.96(abdd,j＝6.5,55.6,295.3hz,major)；hrms(esi-tof)calcd.forc15h15brf2no3[m+h]⁺:374.0198；found:374.0210.

本實(shí)施例制備化合物4i：黃色固體，產(chǎn)率96％；熔點(diǎn):83.3–85.0℃；核磁共振和高分辨質(zhì)譜測(cè)試分析結(jié)果如下：92:8trans/cis(determinedby¹⁹fnmranalysis)；¹hnmr(400mhz,cdcl3)δ(major+minor)7.05-7.03(m,0.16h),6.98-6.92(m,1.84h),6.90-6.87(m,0.92h),6.84-6.81(m,0.08h),6.40(td,j＝7.2hz,1h),4.34-4.28(m,0.16h),4.22-4.04(m,1.84h),3.86(s,3h),3.55(s,2.76h),3.50(s,0.24h),2.99-2.83(m,1.84h),2.61-2.53(m,0.16h),1.36(t,j＝7.1hz,0.24h),1.21(t,j＝7.2hz,2.76h)；¹³cnmr(100mhz,cdcl3)δ(major+minor)172.5,166.7,146.0,145.6,132.1,125.6,123.3,122.9,121.4,116.7,115.3,113.4(t,j＝236.3hz),112.7,112.5,62.0,61.9,61.3,56.1,36.0(d,j＝8.6hz),35.3(d,j＝8.6hz),34.8(t,j＝33.8hz),33.1(d,j＝8.7hz),30.6,30.4,14.3,14.2；¹⁹fnmr(376mhz,cdcl3)δ(major+minor)-111.12(abdd,j＝5.8,54.8,294.4hz,major),-112.85(abdd,j＝8.3,56.0,297.4hz,minor),-114.53(abdd,j＝7.2,53.5,297.3hz,minor),-116.05(abdd,j＝7.0,55.8,294.3hz,major)；hrms(esi-tof)calcd.forc16h18f2no4[m+h]⁺:326.1198；found:326.1202.

本實(shí)施例制備化合物4j：黃色固體，產(chǎn)率75％；熔點(diǎn):96.0–97.5℃；核磁共振和高分辨質(zhì)譜測(cè)試分析結(jié)果如下：90:10trans/cis(determinedby¹⁹fnmranalysis)；¹hnmr(400mhz,cdcl3)δ(major+minor)7.24(d,j＝7.8hz,0.10h),7.13(d,j＝7.4hz,0.90h),7.07-7.02(m,1h),6.99-6.95(m,0.90h),6.81-6.79(m,0.10h),6.38(td,j＝6.9,55.2hz,1h),4.23-4.07(m,2h),3.50(s,3h),3.01(d,j＝7.4hz,0.90h),2.97-2.92(m,0.90h),2.87(d,j＝9.3hz,0.10h),2.65-2.57(m,0.10h),1.22(t,j＝7.0hz,3h)；¹³cnmr(100mhz,cdcl3)δ(major+minor)172.0,170.9,167.2,166.5,147.9(d,j＝242.3hz),130.7(d,j＝23.9hz),126.8(d,j＝3.7hz),123.3(d,j＝6.6hz),120.1(d,j＝3.4hz),118.7(d,j＝3.3hz),116.9,116.7(d,j＝18.9hz),113.2(t,j＝236.6hz),62.1,62.0,36.0(d,j＝12.0hz),35.5(d,j＝9.6hz),35.0(t,j＝33.9hz),33.3(d,j＝7.9hz),29.7(d,j＝6.1hz),29.5(d,j＝6.0hz),14.2,14.1；¹⁹fnmr(376mhz,cdcl3)δ(major+minor)-111.09(abddd,j＝1.3,5.7,54.7,295.3hz,major),-112.99(abddd,j＝1.7,8.3,55.9,298.6hz,minor),-114.60(abdd,j＝7.3,53.5,298.5hz,minor),-116.13(abdd,j＝6.8,55.3,295.3hz,major),(-135.65)–(-135.69)(m,minor),(-136.40)–(-136.45)(m,major)；hrms(esi-tof)calcd.forc15h15f3no3[m+h]⁺:314.0999；found:314.1004.

本實(shí)施例制備化合物4k：白色固體，產(chǎn)率90％；熔點(diǎn):117.0–118.7℃；核磁共振和高分辨質(zhì)譜測(cè)試分析結(jié)果如下：93:7trans/cis(determinedby¹⁹fnmranalysis)；¹hnmr(400mhz,cdcl3)δ(major+minor)7.34(d,j＝7.8hz,0.07h),7.29(d,j＝1.2hz,0.07h),7.26-7.22(m,1.86h),6.95(t,j＝8.0hz,1h),6.38(td,j＝7.3hz,1h),4.21-4.08(m,2h),3.67(s,0.21h),3.66(s,2.79h),3.01(d,j＝7.8hz,0.93h),2.98-2.92(m,0.93h),2.88(d,j＝9.3hz,0.07h),2.64-2.60(m,0.07h),1.22(t,j＝7.2hz,3h)；¹³cnmr(100mhz,cdcl3)δ(major+minor)172.7,166.4,140.0,131.2,131.0,126.7,123.5,123.4,122.6,121.3,116.1,113.2(t,j＝236.7hz),62.2,62.1,35.8(d,j＝9.6hz),35.6(d,j＝8.6hz),35.1(t,j＝33.9hz),33.5(d,j＝7.6hz),30.8,30.5,14.2；¹⁹fnmr(376mhz,cdcl3)δ(major+minor)-111.07(abddd,j＝2.3,5.4,54.6,295.5hz,major),-112.87(abdd,j＝8.2,56.1,299.2hz,minor)-114.52(abddd,j＝3.0,7.6,53.4,299.1hz,minor),-116.08(abddd,j＝2.3,7.1,55.7,295.4hz,major)；hrms(esi-tof)calcd.forc15h14f2nnao3[m+na]⁺:352.0523；found:352.0532.

本實(shí)施例制備化合物4l：淺棕色固體，產(chǎn)率97％；熔點(diǎn):104.3–105.7℃；核磁共振和高分辨質(zhì)譜測(cè)試分析結(jié)果如下：89:11trans/cis(determinedby¹⁹fnmranalysis)；¹hnmr(400mhz,cdcl3)δ(major+minor)7.45(dd,j＝1.0,8.2hz,0.11h),7.42(dd,j＝1.0,8.2hz,0.89h),7.39(d,j＝8.8hz,0.11h),7.27(d,j＝7.6hz,0.89h),6.90(t,j＝7.9hz,1h),6.38(td,j＝7.2,55.2hz,1h),4.23-4.06(m,2h),3.68(s,0.33h),3.66(s,2.67h),3.02(d,j＝7.6hz,0.89h),2.98-2.91(m,0.89h),2.88(d,j＝9.4hz,0.11h),2.66-2.58(m,0.11h),1.21(t,j＝7.2hz,3h)；¹³cnmr(100mhz,cdcl3)δ(major+minor)173.8,172.8,167.1,166.4,141.4,134.5,134.3,127.0,125.2,123.8,123.7,123.1,121.8,113.2(t,j＝236.6hz),103.2,102.8,62.2,62.1,35.9(d,j＝9.6hz),35.5(d,j＝8.7hz),35.0(t,j＝34.0hz),33.6(d,j＝8.1hz),31.1,30.7,14.2；¹⁹fnmr(376mhz,cdcl3)δ(major+minor)-111.06(abdd,j＝4.3,54.6,295.3hz,major),-112.28(abdd,j＝6.8,56.4,299.2hz,minor),-114.49(abdd,j＝7.2,53.5,299.2hz,minor),-116.05(abdd,j＝6.6,54.6,295.3hz,major)；hrms(esi-tof)calcd.forc15h15brf2no3[m+h]⁺:374.0198；found:374.0202.

本實(shí)施例制備化合物4m：黃色固體，產(chǎn)率60％；熔點(diǎn):118.2–119.3℃；核磁共振和高分辨質(zhì)譜測(cè)試分析結(jié)果如下：>99:1trans/cis(determinedby¹⁹fnmranalysis)；¹hnmr(400mhz,cdcl3)δ7.35-7.31(m,2h),7.08-7.04(m,1h),6.91(d,j＝7.5hz,1h),6.40(td,j＝7.2,55.6hz,1h),3.70(s,3h),3.28(s,3h),3.02-2.94(m,2h)；¹³cnmr(100mhz,cdcl3)δ172.3,167.3,144.2,128.7,124.0,123.0(d,j＝2.7hz),122.0113.3(t,j＝236.4hz),108.6,52.8,35.9(d,j＝8.5hz),34.7(d,j＝9.5hz),34.6(t,j＝33.7hz),26.9；¹⁹fnmr(376mhz,cdcl3)δ-111.09(abdd,j＝5.3,54.9,295.1hz,1f),-115.91(abdd,j＝5.9,55.5,294.2hz,1f)；hrms(esi-tof)calcd.forc14h14f2no3[m+h]⁺:282.0936；found:282.0945.

本實(shí)施例制備化合物4n：淺黃色固體，產(chǎn)率87％；熔點(diǎn):63.2–64.0℃；核磁共振和高分辨質(zhì)譜測(cè)試分析結(jié)果如下：>99:1trans/cis(determinedby¹⁹fnmranalysis)；¹hnmr(400mhz,cdcl3)δ7.36-7.31(m,2h),7.06(t,j＝7.7hz,1h),6.91(d,j＝7.8hz,1h),6.41(td,j＝7.1,55.3hz,1h),4.12-3.98(m,2h),3.29(s,3h),3.02-2.95(m,2h)1.62-1.59(m,2h)0.86(t,j＝7.4hz,3h)；¹³cnmr(100mhz,cdcl3)δ172.4,166.9,144.2,128.7,124.1,122.9,122.8,113.4(t,j＝236.4hz,1c),108.6,67.6,35.9(d,j＝8.6hz,1c),34.9(d,j＝9.6hz,1c),34.7(t,j＝33.7hz,1c),26.9,21.9,10.3；¹⁹fnmr(376mhz,cdcl3)δ-111.04(abdd,j＝5.6,55.0,294.6hz,1f),-115.89(abdd,j＝7.0,55.6,294.4hz,1f)；hrms(esi-tof)calcd.forc16h18f2no3[m+h]⁺:310.1249；found:310.1255.

本實(shí)施例制備化合物4o：淺黃色固體，產(chǎn)率99％；熔點(diǎn):80.3–82.0℃；核磁共振和高分辨質(zhì)譜測(cè)試分析結(jié)果如下：87:13trans/cis(determinedby¹⁹fnmranalysis)；¹hnmr(400mhz,cdcl3)δ(major+minor)7.38-7.29(m,2h),7.08-7.02(m,1h),6.90(d,j＝8.2hz,0.87h),6.79(d,j＝7.8hz,0.13h),6.38(td,j＝7.2,55.7hz,1h),3.29(s,2.61h),3.23(s,0.39h),2.95-2.87(m,1.74h),2.81-2.78(m,0.13h),2.60-2.49(m,0.13h),1.41(s,1.17h),1.38(s,7.83h)；¹³cnmr(100mhz,cdcl3)δ(major+minor)172.6,165.7,144.1,132.2,128.7,128.6,128.5,125.0,124.2,124.1,122.9,122.7(d,j＝5.9hz),113.5(t,j＝236.2hz),108.9,108.5,108.2,82.9,35.9(d,j＝9.0hz),35.6(d,j＝9.0hz),34.3(t,j＝33.6hz),33.8(d,j＝8.0hz),28.2,28.1,26.9；¹⁹fnmr(376mhz,cdcl3)δ(major+minor)-110.97(abdd,j＝5.2,54.5,293.8hz,major),-112.66(abdd,j＝8.8,56.1,296.4hz,minor),-115.20(abdd,j＝6.9,53.5,296.3hz,minor),-115.90(abdd,j＝6.7,55.6,293.7hz,major)；hrms(esi-tof)calcd.forc17h20f2no3[m+h]⁺:324.1406；found:324.1415.

本實(shí)施例制備化合物4p：黃色固體，產(chǎn)率93％；熔點(diǎn):70.3–72.3℃；核磁共振和高分辨質(zhì)譜測(cè)試分析結(jié)果如下：85:15trans/cis(determinedby¹⁹fnmranalysis)；¹hnmr(400mhz,cdcl3)δ(major+minor)7.47(d,j＝7.8hz,0.15h),7.37(d,j＝7.8hz,0.85h),7.32(t,j＝7.8hz,1h),7.04(t,j＝6.0hz,0.85h),6.97(d,j＝8.0hz,0.15h),6.93(d,j＝7.8hz,0.85h),6.83(t,j＝8.2hz,0.15h),6.40(td,j＝8.0,56.0hz,1h),4.35-4.30(m,0.3h),4.25-4.07(m,1.7h),3.86-3.80(q,j＝7.2hz,2h),2.95(d,j＝7.5hz,0.85h),2.97-2.92(m,0.85h),2.85(d,j＝9.4hz,0.15h),2.65-2.56(m,0.15h),1.39-1.35(m,0.9h),1.32-1.21(m,5.1h)；¹³cnmr(100mhz,cdcl3)δ(major+minor)172.8,171.9,167.5,166.9,143.3,132.5,129.1,128.7,128.6,124.6,124.3,123.2,122.8,122.7,122.6,122.5,113.4(t,j＝236.5hz),109.0,108.7,108.4,62.0,61.3,35.9(d,j＝9.0hz),35.7,35.5,34.7(d,j＝9.6hz),34.6(t,j＝33.8hz),32.8(d,j＝7.8hz),14.3,14.2,12.9,12.8；¹⁹fnmr(376mhz,cdcl3)δ(major+minor)-111.06(abddd,j＝1.8,5.7,54.7,294.5hz,major),-113.01(abddd,j＝1.7,8.3,55.9,297.6hz,minor),-114.45(abdd,j＝7.3,53.7,297.6hz,minor),-113.01(abdd,j＝7.3,55.1,294.3hz,major)；hrms(esi-tof)calcd.forc16h18f2no3[m+h]⁺:310.1249；found:310.1252.

本實(shí)施例制備化合物4q：橙色油狀物，產(chǎn)率99％；核磁共振和高分辨質(zhì)譜測(cè)試分析結(jié)果如下：85:15trans/cis(determinedby¹⁹fnmranalysis)；¹hnmr(400mhz,cdcl3)δ(major+minor)7.50(d,j＝7.6hz,0.15h),7.39(d,j＝7.6hz,0.85h),7.36-7.20(m,6h),7.07-7.01(m,1h),6.87(d,j＝7.8hz,0.15h),6.81(d,j＝7.8hz,0.85h),6.47(td,j＝7.4,55.2hz,1h),5.08-4.90(m,2h),4.38-4.33(m,0.30h),4.27-4.09(m,1.70h),3.13-3.11(m,0.85h),3.07-3.00(m,0.85h),2.98-2.95(m,0.15h),2.74-2.66(m,0.15h),1.39(t,j＝7.2hz,0.45h),1.25(t,j＝7.2hz,2.55h)；¹³cnmr(100mhz,cdcl3)δ(major+minor)173.4,172.5,167.4,166.8,144.0,143.3,135.4,132.5,129.0,128.9,128.7,128.6,127.9,127.8,127.4,127.3,124.5,124.1,123.0,122.9,122.8,1227,113.5(t,j＝236.5hz),109.9,109.5,62.1,62.0,44.6,44.4,35.90(d,j＝9.0hz),35.0(d,j＝9.7hz),34.9(t,j＝33.8hz),33.5(t,j＝34.1hz),32.9(d,j＝7.7hz),14.1,14.2；¹⁹fnmr(376mhz,cdcl3)δ(major+minor)-110.88(abdd,j＝5.5,54.7,294.6hz,major),-112.87(abdd,j＝8.3,55.9,297.8hz,minor)-114.44(abdd,j＝7.3,53.7,297.6hz,minor),-112.87(abdd,j＝7.6,55.7,294.5hz,major)；hrms(esi-tof)calcd.forc21h20f2no3[m+h]⁺:372.1406；found:372.1418.

本實(shí)施例制備化合物4r：黃色油狀物，產(chǎn)率94％；核磁共振和高分辨質(zhì)譜測(cè)試分析結(jié)果如下：92:8trans/cis(determinedby¹⁹fnmranalysis)；¹hnmr(400mhz,cdcl3)δ(major+minor)7.57-7.54(m,2h),7.46-6.43(m,4h),7.26(t,j＝7.8hz,1h),7.10(t,j＝7.8hz,1h),6.89(d,j＝7.8hz,1h),6.44(td,j＝7.0,55.2hz,1h),4.30-4.12(m,2h),3.13-3.03(m,1.84h),2.97(d,j＝9.4hz,0.08h),2.76-2.68(m,0.08h),1.40(t,j＝7.2hz,0.24h),1.27(t,j＝7.2hz,2.76h)；¹³cnmr(100mhz,cdcl3)δ(major+minor)172.7,171.9,167.4,166.8,144.2,134.1,134.0,129.8,128.7,128.6,128.5,126.8,126.7,124.6,123.9,123.3,123.2,113.3(t,j＝236.5hz),110.2,109.9,62.1,62.0,36.1(d,j＝8.5hz),35.4(d,j＝9.6hz),35.2(t,j＝33.9hz),33.3(d,j＝7.9hz),14.2,14.1；¹⁹fnmr(376mhz,cdcl3)δ(major+minor)-110.83(abdd,j＝3.6,54.7,295.2hz,major),-115.18(abdd,j＝8.0,55.9,298.4hz,minor),-114.42(abdd,j＝7.0,53.6,298.2hz,minor),-115.94(abdd,j＝6.8,55.4,294.9hz,major)；hrms(esi-tof)calcd.forc20h18f2no3[m+h]⁺:358.1249；found:328.1253.

本實(shí)施例制備化合物4s：淺黃色固體，產(chǎn)率70％；熔點(diǎn):98.7–100.2℃；核磁共振和高分辨質(zhì)譜測(cè)試分析結(jié)果如下：87:13trans/cis(determinedby¹⁹fnmranalysis)；¹hnmr(400mhz,cdcl3)δ(major+minor)7.99(d,j＝8.2hz,0.13h),7.89(d,j＝8.2hz,0.87h),7.44(d,j＝7.8hz,0.13h),7.38-7.32(m,1.87h),7.19-7.13(m,1h),6.36(td,j＝7.0,55.3hz,1h),4.23-4.08(m,2h),3.04-3.00(m,1.74h),2.90(d,j＝9.4hz,0.13h),2.66-2.61(m,0.13h),1.65(s,7.83h),1.59(s,1.17h),1.27-1.19(m,3h)；¹³cnmr(100mhz,cdcl3)δ(major+minor)171.3,166.2,148.8,140.3,129.0,125.0,124.7,122.9,122.5,115.2,113.0(t,j＝236.8hz),85.3,62.2,36.8(d,j＝9.6hz),36.3(d,j＝9.6hz),35.6(t,j＝34.1hz),28.2,14.2；¹⁹fnmr(376mhz,cdcl3)δ(major+minor)-115.69(abd,j＝54.6,296.2hz,major),-112.80(abd,j＝52.2,303.1hz,minor),-114.78(abd,j＝53.3,297.6hz,minor),-115.99(abd,j＝55.7,296.2hz,major)；hrms(esi-tof)calcd.forc19h22f2no5[m+h]⁺:382.1461；found:382.1466.

本發(fā)明的式(1)化合物具有重要的生物活性，體外對(duì)人前列腺(pc-3)和人肺癌細(xì)胞(a549)共兩株腫瘤細(xì)胞的細(xì)胞毒性試驗(yàn)表明：此類式(1)所示的含二氟甲基的螺[環(huán)丙烷-1,3'-吲哚啉]-2'-酮類化合物對(duì)腫瘤細(xì)胞生長(zhǎng)具有抑制作用，有可能發(fā)展成為新的防治腫瘤藥物。

本發(fā)明的式(1)化合物或其可藥用鹽及其溶劑化物可以與藥學(xué)上常用的輔料或載體結(jié)合，制備得到具有腫瘤細(xì)胞生長(zhǎng)抑制活性從而可以用于防治腫瘤的藥物組合物。上述各類藥物組合物可以是固體形式，如片劑、膠囊劑、顆粒劑、粉劑；也可以是液體形式，如注射劑、懸浮劑和乳劑等。除此之外，還可以采用現(xiàn)代制藥界所公知的控釋劑或緩釋劑或納米劑。

本發(fā)明的式(1)化合物或其可藥用鹽及其溶劑化物可以與現(xiàn)已上市的抗腫瘤藥物如鉑類藥物順鉑(ddp)、脫氧胞昔類藥物吉西他濱(gemcitabine,gemzar,健擇)、紫杉醇(paclitaxel)、長(zhǎng)春花堿類藥物失碳長(zhǎng)春花堿(vinorebine,nvb諾維本)、喜樹堿類藥物伊立替康(irinatecan、cpt-11)、足葉乙甙(etoposide)等聯(lián)合使用，制備得到具有腫瘤生長(zhǎng)抑制活性的細(xì)胞毒性組合物，可用于治療腫瘤疾病。該類藥物組合物可以是固體形式，如片劑、膠囊劑、顆粒劑、粉劑；也可以是液體形式，如注射劑、懸浮劑和乳劑等。除此之外，還可以采用現(xiàn)代制藥界所公知的控釋劑或緩釋劑或納米劑。

藥理實(shí)施例1：化合物4f、4l、4o或4p對(duì)pc-3細(xì)胞的細(xì)胞毒性

pc-3(人前列腺癌)細(xì)胞用rpmi-1640培養(yǎng)基培養(yǎng)，培養(yǎng)基中含10％的胎牛血清，100u/ml青霉素及100u/ml的鏈霉素。細(xì)胞以每孔5000個(gè)細(xì)胞的濃度加入到96孔中，在37℃含5％co2潮濕空氣的培養(yǎng)箱中培養(yǎng)24小時(shí)。

細(xì)胞存活率的測(cè)定用改良mtt法。細(xì)胞經(jīng)過24小時(shí)的孵育后，分別將新配的化合物4f、4l、4o或4p的二甲基亞砜溶液以濃度梯度加入到各孔中，使孔中化合物最終濃度分別為6.25μmol/l,12.5μmol/l,25μmol/l,50μmol/l和100μmol/l。48小時(shí)后，每孔加入10μlmtt(5mg/ml)的磷酸鹽緩沖液，再繼續(xù)在37℃培養(yǎng)4小時(shí)后，離心5分鐘除去未轉(zhuǎn)化的mtt，每孔中加入150μl二甲基亞砜。以溶解還原的mtt晶體甲臜(formazan)，用酶標(biāo)儀在490nm波長(zhǎng)測(cè)定od值。其中化合物4f、4l、4o或4p對(duì)pc-3細(xì)胞半抑制濃度ic50由spss軟件(19版本)分析得到?；衔?f對(duì)pc-3腫瘤細(xì)胞的ic50為72.1μmol/l；化合物4l對(duì)pc-3腫瘤細(xì)胞的ic50為80.8μmol/l；化合物4o對(duì)pc-3腫瘤細(xì)胞的ic50為25.7μmol/l；化合物4p對(duì)pc-3腫瘤細(xì)胞的ic50為81.8μmol/l；而陽性對(duì)照順鉑對(duì)pc-3腫瘤細(xì)胞的ic50為23.7μmol/l。

實(shí)驗(yàn)結(jié)論：pc-3細(xì)胞是測(cè)試化合物對(duì)腫瘤細(xì)胞的細(xì)胞毒性的有效工具和評(píng)價(jià)指標(biāo)。本實(shí)驗(yàn)表明此類式(1)所示的含二氟甲基的螺[環(huán)丙烷-1,3'-吲哚啉]-2'-酮類化合物對(duì)pc-3細(xì)胞具有一定的細(xì)胞毒性，有可能發(fā)展成新的具有抗腫瘤作用的藥物。

藥理實(shí)施例2：化合物4n、4o、4q或4r對(duì)a549細(xì)胞的細(xì)胞毒性

a549(人肺癌細(xì)胞)用dmem培養(yǎng)基培養(yǎng)，培養(yǎng)基中含10％的胎牛血清，100u/ml的青霉素和100u/ml鏈霉素。細(xì)胞以每孔4000個(gè)細(xì)胞的濃度加入到96孔中，在37℃含5％co2潮濕空氣的培養(yǎng)箱中培養(yǎng)24小時(shí)。

細(xì)胞存活率的測(cè)定用改良mtt法。具體方法如藥理實(shí)施例1?；衔?n對(duì)a549腫瘤細(xì)胞的ic50為89.9μmol/l；化合物4o對(duì)a549腫瘤細(xì)胞的ic50為36.8μmol/l；化合物4q對(duì)a549腫瘤細(xì)胞的ic50為87.9μmol/l；化合物4r對(duì)a549腫瘤細(xì)胞的ic50為62.4μmol/l；而陽性對(duì)照順鉑對(duì)a540腫瘤細(xì)胞的ic50為23.0μmol/l。

實(shí)驗(yàn)結(jié)論：a549細(xì)胞是測(cè)試化合物對(duì)腫瘤細(xì)胞的細(xì)胞毒性的有效工具和評(píng)價(jià)指標(biāo)。本實(shí)驗(yàn)表明此類式(1)所示的含二氟甲基的螺[環(huán)丙烷-1,3'-吲哚啉]-2'-酮類化合物對(duì)a549細(xì)胞具有一定的細(xì)胞毒性，有可能發(fā)展成新的具有抗腫瘤作用的藥物。

從以上藥理實(shí)施例中我們可以看出該類化合物對(duì)這兩株腫瘤細(xì)胞都顯示有一定的細(xì)胞毒性?？梢娺@些化合物具有開發(fā)成為抗腫瘤藥物的潛力，值得繼續(xù)深入研究。