A pyrrolo[4,3,2-de]quinoline derivative

Abstract

The present invention relates to the technical field of anti-tumor drugs, and in particular to a pyrrolo[4,3,2-de]quinoline derivative, the structure of which is shown in the following formula I: #imgabs0#; wherein R 1 Selected from ‑CONH 2 ,‑CONHCH 3 , R 2 Selected from hydrogen, ‑COCH 3 ,‑COCF 3 , #imgabs1# or #imgabs2#; and the structural formula of the pyrrolo[4,3,2-de]quinoline derivative is not #imgabs3#. The compounds of the present invention can be obtained by a relatively short synthetic route using cheap and readily available 1,5-difluoro-2,4-dinitrobenzene as a starting material. The compounds of the present invention have inhibitory activity against at least one of the human tumor cell lines of human LoVo, RKO, HCT-116, HT-29, and DLD-1, and compounds 4b-4e exhibit nanomolar inhibitory activity against the four subtypes of intracellular phosphatidylinositol kinase PI3K and the key regulatory factor mTOR.

Description

Technical Field

[0001] The present invention relates to the technical field of anti-tumor drugs, and in particular to a pyrrolo[4,3,2-de]quinoline derivative. Background Art

[0002] In the current field of anti-tumor drug research and development, Lymphostin alkaloids, as a natural product with a unique pyrrolo[4,3,2-de]quinoline ring-fused tricyclic system, have shown potential value as PI3K / mTOR dual inhibitors and exhibited significant in vitro anticancer activity.

[0003] Lymphostin alkaloids include Lymphostin, Neolymphostin AD, etc., and the structural formula is shown below:

[0004] .

[0005] However, this type of alkaloids faces two core problems in practical application and further development.

[0006] First, the acquisition of Lymphostin alkaloids mainly relies on extraction and separation from microorganisms, which has significant limitations. The limited source of strains, low yield, high energy consumption and low efficiency of the production process make large-scale production extremely difficult. More importantly, it is difficult to obtain derivatives with diverse structures through this method, which limits the possibility of structure-activity relationship research and drug optimization. In addition, although there have been studies such as K. Tatsuta et al. of Waseda University who attempted to construct the pyrroloquinoline ring structure skeleton of Lymphostin through chemical synthesis, the method route is lengthy (up to 21 steps of reaction), and highly toxic reagents such as Tl(OCOCF3)3 are used in the process, which not only increases the safety risk of experimenters, but also limits its practical application value in industrial production and laboratory research.

[0007] Secondly, although Lymphostin alkaloids have been confirmed as dual inhibitors of PI3K / mTOR and have shown anti-tumor activity, their activity level still needs to be improved. In the current competition for the development of anti-tumor drugs, the high efficiency and selectivity of drugs are important criteria for evaluating their potential.

[0008] In summary, although Lymphostin alkaloids play an important role in the development of anti-tumor drugs, the limitations of their microbial preparation and the high difficulty of chemical synthesis, coupled with the fact that their current activity levels need to be improved, have jointly restricted the further development of this type of alkaloids. Therefore, exploring more efficient and safer synthesis methods, as well as developing new derivatives with higher activity and selectivity, will be an important direction for future research on Lymphostin alkaloids. Summary of the invention

[0009] In view of the technical problems that the current synthesis route of Lymphostin alkaloids is lengthy, inefficient, and uses highly toxic reagents, and there is still room for improvement in activity, the present invention provides a pyrrolo[4,3,2-de]quinoline derivative, which is obtained by a relatively short synthesis route using cheap and readily available 1,5-difluoro-2,4-dinitrobenzene as a starting material; the compound of the present invention has inhibitory activity against at least one human tumor cell line of human LoVo, RKO, HCT-116, HT-29, and DLD-1, and compounds 4b-4e all show nanomolar inhibitory activity against four subtypes of intracellular phosphatidylinositol kinase PI3K and the key regulatory factor mTOR.

[0010] The technical solution of the present invention is as follows:

[0011] A pyrrolo[4,3,2-de]quinoline derivative, the structure of which is shown in the following formula I:

[0012] ;

[0013] In the formula,

[0014] R1 is selected from -CONH2, -CONHCH3;

[0015] R2 is selected from hydrogen, -COCH3, -COCF3, or ;

[0016] and the structural formula of the pyrrolo[4,3,2-de]quinoline derivative is not .

[0017] That is, the pyrrolo[4,3,2-de]quinoline derivative is selected from the following compounds:

[0018] , , , , , , , , .

[0019] Preferably, the pyrrolo[4,3,2-de]quinoline derivative is selected from the following compounds:

[0020] , , , .

[0021] More preferably, the pyrrolo[4,3,2-de]quinoline derivative is selected from the following compounds:

[0022] , , .

[0023] Furthermore, the synthesis method of the pyrrolo[4,3,2-de]quinoline derivative comprises the following steps:

[0024] (1) Using 1,5-difluoro-2,4-dinitrobenzene as the starting material, compound VIII is obtained by ammoniation;

[0025] (2) Compound VIII and compound VII are reacted via Doebner-Miller reaction to synthesize quinoline ring compound VI;

[0026] (3) protecting the 5-amino group of compound VI with Boc to obtain an intermediate compound V with mono-Boc protection of the amino group;

[0027] (4) Reaction of compound V with bromination reagent phenyltrimethylammonium tribromide (PTAT) to obtain dihydropyrroloquinoline ring compound IV;

[0028] (5) The nitro groups at the 6- and 8-positions of compound IV are reduced by catalytic hydrogenation to obtain compound III;

[0029] (6) Compound II is prepared from Compound III by a method selected from the following:

[0030] ① Compound III is subjected to a Boc protecting group removal reaction to obtain compound II;

[0031] ② Compound III is subjected to substitution reaction and deprotection reaction (Boc protecting group, Ns protecting group) to obtain compound II;

[0032] (7) reacting compound II with aqueous ammonia or aqueous methylamine solution to obtain compound I;

[0033] The synthetic route is as follows:

[0034] .

[0035] Furthermore, step (1) is specifically as follows:

[0036] Aqueous ammonia was added to a tetrahydrofuran solution of 1,5-difluoro-2,4-dinitrobenzene, and the mixture was stirred at room temperature. The solvent was evaporated to dryness, and water was added, stirred, and filtered to obtain a yellow solid, which was compound VIII.

[0037] Furthermore, step (2) is specifically as follows:

[0038] Add compound VII to a trifluoroacetic acid suspension of compound VIII, heat to reflux, add dichloromethane to the reaction solution, wash with saturated sodium bicarbonate aqueous solution, wash with saturated brine, dry with anhydrous sodium sulfate, evaporate the solvent, and then perform silica gel column chromatography to obtain a yellow solid, which is compound VI.

[0039] Furthermore, step (3) is specifically as follows:

[0040] To a tetrahydrofuran suspension of compound VI, add 4-dimethylaminopyridine (DMA) and Boc2O, stir, react at 30-60°C, add water to the reaction solution, extract with ethyl acetate, wash with saturated brine, dry with anhydrous sodium sulfate, evaporate the solvent, and then separate by silica gel column chromatography to obtain a yellow-red oil and a light yellow solid. The obtained light yellow solid is the intermediate compound V.

[0041] Further, the yellow-red oil obtained in step (3) is dissolved in anhydrous acetonitrile solution, magnesium perchlorate is added, stirred and reacted at room temperature, phosphoric acid aqueous solution is added to the reaction solution, extracted with ethyl acetate, washed with saturated sodium bicarbonate solution, washed with saturated brine, dried over anhydrous sodium sulfate, the solvent is evaporated, and then silica gel column chromatography is performed to obtain a light yellow solid, which is the intermediate compound V.

[0042] Furthermore, step (4) is specifically as follows:

[0043] Add phenyltrimethylammonium tribromide to a tetrahydrofuran solution of compound V, react at 30-60°C, add water to the reaction solution, extract with ethyl acetate, wash with saturated sodium bicarbonate solution, wash with saturated brine, dry with anhydrous sodium sulfate, evaporate the solvent, and then perform silica gel column chromatography to obtain a red solid which is compound IV.

[0044] Furthermore, step (5) is specifically as follows:

[0045] Dissolve compound IV in a mixed solvent of dichloromethane and methanol, add catalyst Pd / C, replace hydrogen, stir and react at room temperature, filter, wash with methanol, evaporate the solvent, and then perform silica gel column chromatography to obtain a red solid, which is compound III.

[0046] Furthermore, in step (6), method ① is specifically:

[0047] Dissolve compound III in dichloromethane, add anhydrous zinc bromide, stir and react at room temperature, add saturated sodium bicarbonate aqueous solution to the reaction solution, extract with dichloromethane, wash with saturated brine, dry with anhydrous sodium sulfate, evaporate the solvent, and then perform silica gel column chromatography to obtain compound II.

[0048] Furthermore, method ② in step (6) is specifically:

[0049] Compound III is first reacted with o-nitrobenzenesulfonyl chloride (NsCl) to obtain compound XI; then, compound IX is added to an anhydrous dichloromethane solution of compound XI under a triethylamine environment to react to obtain compound X; anhydrous zinc bromide is added to an anhydrous dichloromethane solution of compound X, a saturated sodium bicarbonate aqueous solution is added to the reaction solution, dichloromethane extraction is performed, saturated saline water is washed, anhydrous sodium sulfate is dried, the solvent is evaporated, and silica gel column chromatography is performed; the obtained product is dissolved in N,N-dimethylformamide, anhydrous potassium carbonate and thiophenol are added, stirred at room temperature and reacted, a sodium bicarbonate aqueous solution is added to the reaction solution, dichloromethane extraction is performed, water is washed, saturated saline water is washed, anhydrous sodium sulfate is dried, the solvent is evaporated, and silica gel column chromatography is performed to obtain compound II;

[0050] The reaction route is as follows:

[0051] ;

[0052] Wherein, R2 is selected from -COCH3, -COCF3, or .

[0053] Furthermore, step (7) is specifically as follows:

[0054] Dissolve compound II in methanol, add ammonia water or methylamine aqueous solution, reflux under nitrogen protection, then cool to room temperature, evaporate the reaction liquid, and perform silica gel column chromatography to obtain the product.

[0055] The beneficial effects of the present invention are:

[0056] The pyrrolo[4,3,2-de]quinoline derivative provided by the present invention has a novel structure, exhibits superior anti-tumor activity or kinase inhibitory activity in in vitro experiments, and can more effectively inhibit at least one human tumor cell line or specific kinase compared to Lymphostin, and has great potential in the development of future anti-cancer drugs. At the same time, the pyrrolo[4,3,2-de]quinoline derivative provided by the present invention can be obtained by a simpler synthesis method, which overcomes many difficulties in the chemical synthesis of Lymphostin alkaloids and provides the possibility of large-scale production and drug optimization. DETAILED DESCRIPTION

[0057] In order to enable those skilled in the art to better understand the technical solutions in the present invention, the technical solutions in the embodiments of the present invention are described clearly and completely below. Obviously, the described embodiments are only part of the embodiments of the present invention, not all of the embodiments. Based on the embodiments of the present invention, all other embodiments obtained by ordinary technicians in this field without creative work should fall within the scope of protection of the present invention.

[0058] Specific implementation method The general formula of the pyrrolo[4,3,2-de]quinoline derivative synthesized in each embodiment is:

[0059] ;

[0060] In the formula, the substituents of R1 and R2 are shown in Table 1 below.

[0061] Table 1 Pyrrolo[4,3,2-de]quinoline derivatives

[0062]

[0063] Example 1 1-tert-Butyloxycarbonylamino-6,8-diaminopyrrolo[4,3,2-de]quinoline-1,4(2 H Preparation of ethyl 4-formate (Compound III)

[0064] (1) Preparation of 4,6-dinitro-1,3-phenylenediamine (Compound VIII):

[0065] ;

[0066] To a 15 mL THF solution of 2.0 g (9.8 mmol) 1,5-difluoro-2,4-dinitrobenzene, add 2.0 mL of ammonia water dropwise. After the addition is complete, stir at room temperature and a yellow solid gradually precipitates. Allow to react overnight. TLC shows that the raw material has reacted completely. Evaporate the solvent, add a small amount of water, stir and filter to obtain 1.75 g of a yellow solid, with a yield of 90%, a purity of 99.3%, and a melting point of 298-300°C.

[0067] 1 H NMR (600 MHz, DMSO- d 6): δ 8.84 (s, 1H), 7.63 (brs, 4H), 6.14 (s, 1H).

[0068] 13 C NMR (150 MHz, DMSO- d 6): δ 149.1, 127.8, 123.9, 98.1.

[0069] HRMS (ESI) m / z: [M+H] + calcd for C6H7N4O4199.0462; found 199.0460.

[0070] (2) Preparation of ethyl 4-methyl-5-amino-6,8-dinitroquinoline-2-carboxylate (Compound VI):

[0071] ;

[0072] To a suspension of 1.5 g (7.58 mmol) of compound VIII in 20 mL of trifluoroacetic acid, add 1.59 g (11.20 mmol) of (E)-4-oxo-2-pentenoic acid ethyl ester (Compound VII), and after the addition, heat to reflux. After reflux for 14 hours, spot the plate, and TLC shows that the raw material reaction is complete. Add dichloromethane to the reaction solution, wash with saturated sodium bicarbonate aqueous solution, wash with saturated brine, dry with anhydrous sodium sulfate, evaporate the solvent, and then perform silica gel column chromatography (petroleum ether: ethyl acetate = 3:1) to obtain 1.28 g of yellow solid, yield: 53%, purity: 95.7%, melting point: 203-205°C.

[0073] 1 H NMR (400 MHz, DMSO- d 6): δ 8.85 (brs, 2H), 8.81 (s, 1H), 8.00 (s,1H), 4.40 (q, J = 7.2 Hz, 2H), 3.03 (s, 3H), 1.34 (t, J = 7.2 Hz, 3H).

[0074] 13 C NMR (150 MHz, DMSO- d 6): δ 163.6, 150.9, 149.7, 149.1, 143.5,136.4, 124.8, 123.6, 123.2, 121.7, 61.9, 23.3, 14.0.

[0075] HRMS (ESI) m / z: [M+H] + Calculate for C 13 H 13 N4O6321.0829; found: 321.0826.

[0076] (3) Preparation of ethyl 4-methyl-5-tert-butoxycarbonylamino-6,8-dinitroquinoline-2-carboxylate (Compound V):

[0077] ;

[0078] To a suspension of 900 mg (2.81 mmol) of compound VI in 30 mL of tetrahydrofuran, 180 mg (1.47 mmol) of DMAP and 1.50 g (6.87 mmol) of Boc2O were added successively, and the mixture was stirred after addition. After reacting at 30°C for 14 hours, TLC showed that the reaction of the raw materials was complete. Water was added to the reaction solution, extracted with ethyl acetate, washed with saturated brine, dried over anhydrous sodium sulfate, and the solvent was evaporated. Then, silica gel column chromatography (petroleum ether: ethyl acetate = 5:1) was performed to obtain 1.14 g of a yellow-red oil, with a yield of 78%. In addition, the amino mono-Boc-protected product, compound V, was also isolated, with a light yellow solid of 189 mg, with a yield of 16%.

[0079] Add 90 mg (0.4 mmol) of magnesium perchlorate to 30 mL of anhydrous acetonitrile solution of 520 mg (1.0 mmol) of yellow-red oil, stir at room temperature after addition. After 8 hours of reaction at room temperature, spot the plate, TLC shows that the reaction of the raw material is complete. Add 1 M phosphoric acid aqueous solution to the reaction solution, extract with ethyl acetate, wash with saturated sodium bicarbonate solution, wash with saturated brine, dry with anhydrous sodium sulfate, evaporate the solvent, and then perform silica gel column chromatography (petroleum ether: ethyl acetate = 4:1) to obtain 378 mg of light yellow solid, yield: 90%, purity: 96.3%, melting point: 195-198°C.

[0080] 1 H NMR (600 MHz, DMSO- d 6): δ 9.94 (brs, 1H), 9.00 (s, 1H), 8.20 (s,1H), 4.44 (q, J = 7.2 Hz, 2H), 3.00 (s, 3H), 1.49 (s, 9H), 1.37 (t, J = 7.2 Hz,3H).

[0081] 13 C NMR (150 MHz, DMSO- d 6): δ 163.8, 153.8, 151.2, 151.0, 147.6,144.8, 140.8, 132.2, 128.5, 127.2, 119.5, 81.4, 62.6, 28.5, 23.4, 14.5.

[0082] HRMS (ESI) m / z: [M+Na] +Calculate for C 18 H 20 N4O8Na 443.1173; found:443.1168.

[0083] (4) 1-tert-Butyloxycarbonylamino-6,8-dinitropyrrolo[4,3,2-de]quinoline-1,4(5 H Preparation of ethyl 4-formate (Compound IV):

[0084] ;

[0085] Add 448 mg (1.19 mmol) of phenyltrimethylammonium tribromide to a solution of 500 mg (1.19 mmol) of compound V in 35 mL of anhydrous tetrahydrofuran, and react at 30°C. After stirring for a few minutes, the reaction solution gradually turned into a red turbid liquid. After reacting at 30°C for 5 hours, spot the plate, and TLC shows that the raw material has reacted. Add water to the reaction solution, extract with ethyl acetate, wash with saturated sodium bicarbonate solution, wash with saturated brine, dry with anhydrous sodium sulfate, evaporate the solvent, and then chromatograph on a silica gel column (petroleum ether: ethyl acetate = 9:1) to obtain 360 mg of a red solid, with a yield of 72%, a purity of 96.8%, and a melting point of >135°C (decomp.).

[0086] 1 H NMR (300 MHz, DMSO- d 6): δ 10.19 (brs, 1H), 8.39 (s, 1H), 7.63 (s,1H), 7.31 (s, 1H), 4.40 (q, J = 7.2 Hz, 2H), 1.56 (s, 9H), 1.35 (t, J = 7.2 Hz, 3H).

[0087] 13 C NMR (150 MHz, CDCl3): δ 161.2, 147.7, 137.3, 130.9, 147.0, 128.8,125.1, 122.9, 122.5, 116.7, 116.1, 110.0, 87.9, 62.8, 27.7, 14.1.

[0088] HRMS (ESI) m / z: [M+H] + Calculate for C 18 H 19 N4O8419.1197; found: 419.1182.

[0089] (5) 1-tert-Butyloxycarbonylamino-6,8-diaminopyrrolo[4,3,2-de]quinoline-1,4(2 H Preparation of ethyl 4-formate (Compound III):

[0090] ;

[0091] 340 mg (0.81 mmol) of compound IV was dissolved in a mixed solvent of 10 mL of dichloromethane and 5 mL of methanol, and a catalytic amount of 10% Pd / C was added. After the addition was completed, the hydrogen was replaced and stirred at room temperature for 8 hours. TLC showed that the reaction of the raw material was complete. Filter, wash with a large amount of methanol, evaporate the solvent, and then perform silica gel column chromatography (dichloromethane: methanol = 100: 1) to obtain 226 mg of a red solid, with a yield of 77%, a purity of 97.9%, and a melting point of >145°C (decomp.).

[0092] 1 H NMR (400 MHz, DMSO- d 6): δ 7.69 (s, 1H), 6.38 (s, 1H), 6.23 (brs,2H), 5.49 (brs, 2H), 5.16 (s, 2H), 4.36 (q, J = 7.2 Hz, 2H), 1.54 (s, 9H), 1.35(t, J = 7.2 Hz, 3H).

[0093] 13 C NMR (150 MHz, CDCl3): δ 165.7, 152.6, 143.6, 143.5, 140.3, 133.3,130.5, 129.6, 113.9, 112.9, 106.0, 81.4, 61.6, 55.5, 28.3, 14.2.

[0094] HRMS (ESI) m / z: [M+H] + Calculate for C 18 H 23 N4O4359.1714; found: 359.1722.

[0095] Example 2 Preparation of Compounds 2a-2d

[0096] ;

[0097] (1) 1-tert-Butyloxycarbonylamino-8-o-nitrobenzenesulfonylamino-6-aminopyrrolo[4,3,2-de]quinoline-1,4(2 H Preparation of ethyl 4-formate (Compound XI):

[0098] First, compound III was prepared according to the method of Example 1, and then 7 mg (0.056 mmol) of DMAP, 0.12 mL (0.838 mmol) of triethylamine and 68 mg (0.31 mmol) of o-nitrobenzenesulfonyl chloride were added to a solution of 100 mg (0.28 mmol) of compound III in 10 mL of anhydrous dichloromethane. After addition, the mixture was stirred at room temperature. After reacting at room temperature for 6 hours, water was added to the reaction solution, extracted with dichloromethane, washed with saturated brine, dried with anhydrous sodium sulfate, and the solvent was evaporated. Then, silica gel column chromatography (dichloromethane: methanol = 200: 1) was performed to obtain 119 mg of a brown-yellow solid, with a yield of 79%, a purity of 97.6%, and a melting point of >168°C (decomp.).

[0099] 1 H NMR (400 MHz, DMSO- d 6): δ 11.38 (brs, 1H), 8.02 (d, J=7.6 Hz, 1H), 7.95 (d, J=7.6 Hz, 1H), 7.83 (dd, J=7.6, 7.6 Hz, 1H), 7.76 (dd, J=7.6, 7.6Hz, 1H), 7.15 (s, 1H), 5.93 (brs, 2H), 5.19 (s, 2H), 4.39 (q, J = 7.2 Hz, 2H), 1.56 (s, 9H), 1.35 (t, d= 7.2 Hz, 3 H).

[0100] 13 C NMR (150 MHz, DMSO- d 6): δ 165.2, 153.5, 148.1, 147.2, 145.4,140.0, 133.5, 133.3, 133.0, 131.7, 131.3, 128.5, 124.5, 121.2, 121.0, 113.4,106.7, 83.8, 62.0, 55.9, 28.3, 14.2.

[0101] HRMS (ESIMS): calcd for C 24 H 26 N5O8S [M+H] +: 544.1497. Found: 544.1503.

[0102] (2) Preparation of compounds a'-d':

[0103] To a solution of 1.0 mmol of compound XI in 20 mL of anhydrous dichloromethane, 2.0 mmol of triethylamine was added, and then 1.05 mmol of acyl chloride (R2Cl, R2 is selected from -COCH3, -COCF3, or After the reaction of the raw materials is completed, water is added to the reaction solution, extracted with dichloromethane, washed with saturated brine, dried over anhydrous sodium sulfate, the solvent is evaporated, and then silica gel column chromatography is performed to obtain compounds a'-d'.

[0104] ①1-tert-Butyloxycarbonylamino-6-acetylamino-8-o-nitrobenzenesulfonylaminopyrrolo[4,3,2-de]quinoline-1,4(2 H )-4-carboxylic acid ethyl ester (Compound a'):

[0105] ;

[0106] Brown solid, yield: 63%, purity: 95.9%, melting point: >149℃ (decomp.).

[0107] 1 H NMR (400 MHz, DMSO- d 6): δ 11.29 (brs, 1H), 9.82 (brs, 1H), 8.87(s, 1H), 8.15 (d, J=7.6 Hz, 1H), 7.97 (m, 2H), 7.83 (dd, J=7.6, 7.6 Hz, 1H), 7.73 (dd, J=7.6, 7.6 Hz, 1H), 5.27(s, 2H), 4.41 (q, J = 7.2 Hz, 2H), 2.27 (s,3H), 1.58 (s, 9H), 1.37 (t, d= 7.2 Hz, 3 H).

[0108] 13 C NMR (150 MHz, DMSO- d6): δ 170.6, 165.3, 154.1, 149.5, 146.3,144.9, 141.4, 134.5, 133.1, 132.6, 131.2, 130.8, 129.1, 124.5, 121.3, 121.1,113.5, 106.7, 83.6, 61.9, 55.8, 28.1, 24.5, 14.3.

[0109] HRMS (ESIMS): calcd for C 26 H 28 N5O9S [M+H] + : 586.1602. Found: 586.1593.

[0110] ②1-tert-Butyloxycarbonylamino-6-trifluoroacetylamino-8-o-nitrobenzenesulfonylaminopyrrolo[4,3,2-de]quinoline-1,4(2 H )-4-carboxylic acid ethyl ester (compound b'):

[0111] ;

[0112] Brown solid, yield: 45%, purity: 95.3%, melting point: >168℃ (decomp.).

[0113] 1 H NMR (400 MHz, DMSO- d 6): δ 11.37 (brs, 1H), 9.99 (brs, 1H), 8.91(s, 1H), 8.17 (d, J=7.6 Hz, 1H), 8.08 (m, 2H), 7.86 (dd, J=7.6, 7.6 Hz, 1H), 7.77 (dd, J=7.6, 7.6 Hz, 1H), 5.28 (s, 2H), 4.41 (q, J = 7.2 Hz, 2H), 1.58(s, 9H), 1.37 (t, d= 7.2 Hz, 3 H).

[0114] 13 C NMR (150 MHz, DMSO- d6): δ 171.8, 166.5, 155.3, 149.9, 146.8,145.4, 141.7, 134.8, 133.7, 132.9, 131.3, 130.9, 129.4, 124.7, 121.6, 121.3,113.8, 106.8, 83.7, 62.3, 55.9, 28.3, 24.6, 14.4.

[0115] HRMS (ESIMS): calcd for C 26 H 25 N5O9F3S [M+H] + : 640.1320. Found: 640.1312.

[0116] ③1-tert-Butyloxycarbonylamino-6-propionylamino-8-o-nitrobenzenesulfonylaminopyrrolo[4,3,2-de]quinoline-1,4(2 H )-4-carboxylic acid ethyl ester (Compound c'):

[0117] ;

[0118] Brown solid, yield: 59%, purity: 95.5%, melting point: >153°C (decomp.).

[0119] 1 H NMR (400 MHz, DMSO- d 6): δ 11.28 (brs, 1H), 9.81 (brs, 1H), 8.86 (s, 1H), 8.13 (d, J=7.6 Hz, 1H), 7.96 (m, 2H), 7.81 (dd, J=7.6, 7.6 Hz, 1H), 7.72 (dd, J=7.6, 7.6 Hz, 1H), 5.28 (s, 2H), 4.41 (q, J = 7.2 Hz, 2H), 2.53(q, J = 7.2 Hz, 2H), 1.58 (s, 9H), 1.37 (t, d= 7.2 Hz, 3 H), 1.18 (t, J = 7.2Hz, 3H).

[0120] 13 C NMR (150 MHz, DMSO- d6): δ 170.6, 165.3, 154.1, 149.5, 146.3,144.9, 141.4, 134.5, 133.1, 132.6, 131.2, 130.8, 129.1, 124.5, 121.3, 121.1,113.5, 106.7, 83.6, 61.8, 56.5, 31.2, 28.6, 14.6, 10.7.

[0121] HRMS (ESIMS): calcd for C 27 H 30 N5O9S [M+H] + : 600.1759. Found: 600.1751.

[0122] ④1-tert-Butyloxycarbonylamino-6-isobutyrylamino-8-nitrobenzenesulfonylaminopyrrolo[4,3,2-de]quinoline-1,4(2 H )-4-carboxylic acid ethyl ester (Compound d'):

[0123] ;

[0124] Brown solid, yield: 66%, purity: 95.7%, melting point: >164℃ (decomp.).

[0125] 1 H NMR (400 MHz, DMSO- d 6): δ 11.26 (brs, 1H), 9.80 (brs, 1H), 8.87(s, 1H), 8.16 (d, J=7.6 Hz, 1H), 7.95 (m, 2H), 7.83 (dd, J=7.6, 7.6 Hz, 1H), 7.73 (dd, J=7.6, 7.6 Hz, 1H), 5.26 (s, 2H), 4.41 (q, J = 7.2 Hz, 2H), 2.81(m, 1H), 1.58 (s, 9H), 1.37 (t, d= 7.2 Hz, 3 H), 1.23 (d, J = 6.8 Hz, 6H).

[0126] 13 C NMR (150 MHz, DMSO- d6): δ 170.3, 165.1, 154.3, 149.4, 146.5,144.7, 141.3, 134.5, 133.1, 132.8, 131.1, 130.8, 129.2, 124.5, 121.1, 121.0,113.6, 106.8, 83.8, 61.9, 56.3, 35.9, 28.8, 19.7, 14.5.

[0127] HRMS (ESIMS): calcd for C 28 H 32 N5O9S [M+H] + : 614.1915. Found: 614.1901.

[0128] (3) Preparation of compounds a"-d":

[0129] Add 3.0 mmol of anhydrous zinc bromide to a 20 mL anhydrous dichloromethane solution of 0.5 mmol of compound a'-d', stir at room temperature after the addition is complete. After the reaction of the raw materials is complete, add saturated sodium bicarbonate aqueous solution to the reaction solution, extract with dichloromethane, wash with saturated brine, dry with anhydrous sodium sulfate, evaporate the solvent, and then perform silica gel column chromatography to obtain compound a"-d".

[0130] ①6-Acetylamino-8-o-nitrobenzenesulfonylaminopyrrolo[4,3,2-de]quinoline-4-carboxylic acid ethyl ester (Compound a”):

[0131] ;

[0132] Brown solid, yield: 78%, purity: 97.3%, melting point: >229℃ (decomp.).

[0133] 1 H NMR (400 MHz, CD3OD): d 8.72 (s, 1H), 8.33 (s, 1H), 8.21 (1H, s),8.18 (ddd, J = 8.0, 7.5, 1.0 Hz, 1H), 8.13 (dd, J = 8.0, 1.0 Hz, 1H), 7.90(dd, J = 7.5, 1.5 Hz, 1H), 7.85 (ddd, J = 8.0, 8.0, 1.5 Hz, 1H), 4.41 (q, J =7.2 Hz, 2H), 2.26 (s, 3H), 1.38 (t, d= 7.2 Hz, 3 H).

[0134] 13 C NMR (150 MHz, CD3OD): d 170.8, 165.6, 154.3, 149.6, 146.7, 144.9,141.7, 134.6, 133.5, 132.3, 131.5, 130.9, 129.8, 124.6, 121.3, 121.1, 113.6,106.8, 61.9, 24.6, 14.1.

[0135] HRMS (ESIMS): calcd for C 21 H 18 N5O7S [M+H] + : 484.0921. Found: 484.0910.

[0136] ②6-Trifluoroacetylamino-8-o-nitrobenzenesulfonylaminopyrrolo[4,3,2-de]quinoline-4-carboxylic acid ethyl ester (Compound b”):

[0137] ;

[0138] Brown solid, yield: 49%, purity: 96.8%, melting point: >263℃ (decomp.).

[0139] 1 H NMR (400 MHz, CD3OD): d 8.81 (s, 1H), 8.37 (s, 1H), 8.23 ​​(1H, s),8.19 (ddd, J = 8.0, 7.5, 1.0 Hz, 1H), 8.16 (dd, J = 8.0, 1.0 Hz, 1H), 7.93(dd, J = 7.5, 1.5 Hz, 1H), 7.87 (ddd, J = 8.0, 8.0, 1.5 Hz, 1H), 4.41 (q, J =7.2 Hz, 2H), 1.38 (t, d= 7.2 Hz, 3 H).

[0140] 13 C NMR (150 MHz, CD3OD): d170.9, 165.7, 154.5, 149.8, 146.8, 145.1,141.8, 134.9, 133.6, 132.4, 131.6, 130.9, 129.9, 124.7, 121.5, 121.2, 116.7,113.7, 106.9, 61.8, 14.1.

[0141] HRMS (ESIMS): calcd for C 21 H 15 F3N5O7S [M+H] + : 538.0639. Found: 538.0631.

[0142] ③6-Propanylamino-8-o-nitrobenzenesulfonylaminopyrrolo[4,3,2-de]quinoline-4-carboxylic acid ethyl ester (Compound c"):

[0143] ;

[0144] Brown solid, yield: 71%, purity: 97.7%, melting point: >236℃ (decomp.).

[0145] 1 H NMR (400 MHz, CD3OD): d 8.71 (s, 1H), 8.32 (s, 1H), 8.22 (1H, s),8.18 (ddd, J = 8.0, 7.5, 1.0 Hz, 1H), 8.13 (dd, J = 8.0, 1.0 Hz, 1H), 7.90(dd, J = 7.5, 1.5 Hz, 1H), 7.85 (ddd, J = 8.0, 8.0, 1.5 Hz, 1H), 4.41 (q, J =7.2 Hz, 2H), 2.66 (q, J = 7.2 Hz, 2H), 1.38 (t, d= 7.2 Hz, 3 H), 1.18 (t, J =7.2 Hz, 3H).

[0146] 13 C NMR (150 MHz, CD3OD): d170.6, 165.3, 154.1, 149.3, 146.5, 144.7,141.3, 134.5, 133.4, 132.1, 131.3, 130.8, 129.6, 124.5, 121.6, 121.3, 113.5,106.9, 61.8, 32.7, 14.1, 11.2.

[0147] HRMS (ESIMS): calcd for C 22 H 20 N5O7S [M+H] + : 498.1078. Found: 498.1063.

[0148] ④6-isobutyrylamino-8-o-nitrobenzenesulfonylaminopyrrolo[4,3,2-de]quinoline-4-carboxylic acid ethyl ester (Compound d"):

[0149] ;

[0150] Brown solid, yield: 76%, purity: 98.1%, melting point: >251℃ (decomp.).

[0151] 1 H NMR (400 MHz, CD3OD): d 8.73 (s, 1H), 8.33 (s, 1H), 8.21 (1H, s),8.18 (ddd, J = 8.0, 7.5, 1.0 Hz, 1H), 8.13 (dd, J = 8.0, 1.0 Hz, 1H), 7.90(dd, J = 7.5, 1.5 Hz, 1H), 7.85 (ddd, J = 8.0, 8.0, 1.5 Hz, 1H), 4.41 (q, J =7.2 Hz, 2H), 3.08 (m, 1H), 1.38 (t, d= 7.2 Hz, 3 H), 1.23 (d, J = 6.8 Hz,6H).

[0152] 13 C NMR (150 MHz, CD3OD): d170.7, 165.7, 154.4, 149.5, 146.8, 144.7,141.8, 134.7, 133.6, 132.2, 131.6, 130.9, 129.7, 124.6, 121.3, 121.1, 113.7,106.8, 61.9, 32.1, 21.3, 14.2.

[0153] HRMS (ESIMS): calcd for C 23 H 22 N5O7S [M+H] + : 512.1234. Found: 512.1226.

[0154] (4) Preparation of compounds 2a-2d:

[0155] To a 10 mL DMF solution of 0.5 mmol of compound a”-d”, add 1.5 mmol of anhydrous potassium carbonate and 0.7 mmol of thiophenol, stir at room temperature after the addition. After the reaction of the raw materials is complete, add sodium bicarbonate aqueous solution to the reaction solution, extract with dichloromethane, wash with water, wash with saturated brine, dry with anhydrous sodium sulfate, evaporate the solvent, and then chromatograph on a silica gel column to obtain compounds 2a-2d.

[0156] ①6-Acetylamino-8-aminopyrrolo[4,3,2-de]quinoline-4-carboxylic acid ethyl ester (Compound 2a):

[0157] ;

[0158] Red solid, yield: 67%, purity: 99.3%, melting point: >228℃ (decomp.).

[0159] 1 H NMR (400 MHz, DMSO- d 6): d 9.94 (brs, 1H), 8.64 (s, 1H), 8.39 (brs,2H), 8.38 (s, 1H), 8.11 (s, 1H), 4.44 (q, J = 7.2 Hz, 2H), 2.32 (s, 3H), 1.40(t, d= 7.2 Hz, 3 H).

[0160] 13 C NMR (150 MHz, DMSO- d 6): d169.4, 164.5, 153.8, 145.1, 141.9, 141.7,138.6, 134.5, 126.7, 123.1, 121.7, 101.3, 61.6, 24.7, 14.1.

[0161] HRMS (ESIMS): calcd for C 15 H 15 N4O3[M+H] + : 299.1139. Found: 299.1137.

[0162] ②6-Trifluoroamino-8-acetylaminopyrrolo[4,3,2-de]quinoline-4-carboxylic acid ethyl ester (Compound 2b):

[0163] ;

[0164] Red solid, yield: 57%, purity: 97.8%, melting point: >256℃ (decomp.).

[0165] 1 H NMR (400 MHz, DMSO- d 6): d 10.34 (brs, 1H), 8.76 (s, 1H), 8.51 (brs,2H), 8.45 (s, 1H), 8.28 (s, 1H), 4.44 (q, J = 7.2 Hz, 2H), 1.40 (t, d= 7.2Hz, 3 H).

[0166] 13 C NMR (150 MHz, DMSO- d 6): d 171.6, 166.4, 154.5, 145.7, 142.9, 142.5,139.9, 134.6, 126.3, 123.1, 121.4, 117.1, 99.9, 62.9, 14.3.

[0167] HRMS (ESIMS): calcd for C 15 H 12 F3N4O3[M+H] + : 353.0856. Found: 353.0861.

[0168] ③6-Propanylamino-8-aminopyrrolo[4,3,2-de]quinoline-4-carboxylic acid ethyl ester (Compound 2c):

[0169] ;

[0170] Red solid, yield: 63%, purity: 98.1%, melting point: >237℃ (decomp.).

[0171] 1 H NMR (400 MHz, DMSO- d 6): d 9.93 (brs, 1H), 8.65 (s, 1H), 8.39 (brs,2H), 8.37 (s, 1H), 8.12 (s, 1H), 4.44 (q, J = 7.2 Hz, 2H), 2.66 (q, J = 7.2Hz, 2H), 1.40 (t, d= 7.2 Hz, 3H), 1.18 (t, J = 7.2 Hz, 3H).

[0172] 13 C NMR (150 MHz, DMSO- d 6): d 169.3, 164.6, 153.7, 145.2, 141.7, 141.3,138.7, 134.6, 126.3, 123.2, 121.8, 101.5, 61.6, 32.2, 14.1, 11.7.

[0173] HRMS (ESIMS): calcd for C 16 H 17 N4O3[M+H] + : 313.1295. Found: 313.1288.

[0174] ④6-isobutylamino-8-acylaminopyrrolo[4,3,2-de]quinoline-4-carboxylic acid ethyl ester (Compound 2d):

[0175] ;

[0176] Red solid, yield: 76%, purity: 98.9%, melting point: >241℃ (decomp.).

[0177] 1 H NMR (400 MHz, DMSO- d 6): d9.96 (brs, 1H), 8.67 (s, 1H), 8.39 (brs,2H), 8.38 (s, 1H), 8.13 (s, 1H), 4.44 (q, J = 7.2 Hz, 2H), 3.08 (m, 1H), 1.40(t, d= 7.2 Hz, 3 H), 1.23 (d, J = 6.8 Hz, 6H).

[0178] 13 C NMR (150 MHz, DMSO- d 6): d 169.7, 164.3, 153.9, 145.4, 141.8, 141.6,138.8, 134.7, 126.6, 123.3, 121.9, 101.2, 61.9, 32.1, 19.8, 14.1.

[0179] HRMS (ESIMS): calcd for C 17 H 19 N4O3[M+H] + : 327.1452. Found: 327.1441.

[0180] Example 3 Preparation of 6,8-diaminopyrrolo[4,3,2-de]quinoline-4-carboxamide (Compound 3a)

[0181] ;

[0182] First, compound III was prepared according to the method of Example 1, and then 400 mg (1.78 mmol) of anhydrous zinc bromide was added to a solution of 100 mg (0.28 mmol) of compound III in 10 mL of anhydrous dichloromethane. After the addition, the mixture was reacted at room temperature. After the reaction of the raw materials was completed, a saturated sodium bicarbonate aqueous solution was added to the reaction solution, extracted with dichloromethane, washed with saturated brine, dried over anhydrous sodium sulfate, and the solvent was evaporated. Then, the mixture was dissolved in 10 mL of methanol, 5 mL of ammonia water was added, and the mixture was refluxed for 24 h under nitrogen protection, cooled to room temperature, and the reaction solution was evaporated to dryness, and then subjected to silica gel column chromatography (dichloromethane: methanol = 10: 1) to obtain 25 mg of a dark red solid, with a yield of 39%, a purity of 96.1%, and a melting point of >178°C (decomp.).

[0183] 1 H NMR (400 MHz, CD3OD): d 8.57 (s, 1H), 8.01 (s, 1H), 5.87 (s, 1H).

[0184] 13 C NMR (150 MHz, CD3OD): d 166.7, 154.1, 153.9, 138.8, 138.7, 137.1,134.9, 124.3, 121.9, 121.3, 95.6.

[0185] HRMS (ESIMS): calcd for C 11 H 10 N5O [M+H] + : 228.0880. Found: 228.0872.

[0186] Example 4 6,8-diamino-N-methylpyrrolo[4,3,2-de]quinoline-4-carboxamide (Compound 4a):

[0187] ;

[0188] First, compound III was prepared according to the method of Example 1, and then 400 mg (1.78 mmol) of anhydrous zinc bromide was added to a solution of 100 mg (0.28 mmol) of compound III in 10 mL of anhydrous dichloromethane. After the addition, the mixture was reacted at room temperature. After the reaction of the raw materials was completed, a saturated aqueous sodium bicarbonate solution was added to the reaction solution, extracted with dichloromethane, washed with saturated brine, dried over anhydrous sodium sulfate, and the solvent was evaporated. Then, the mixture was dissolved in 10 mL of methanol, 5 mL of methylamine aqueous solution was added, and the mixture was refluxed for 24 h under nitrogen protection, cooled to room temperature, and the reaction solution was evaporated to dryness, and then subjected to silica gel column chromatography (dichloromethane: methanol = 10: 1) to obtain 34 mg of a dark red solid, with a yield of 43%, a purity of 96.6%, and a melting point of >183°C (decomp.).

[0189] 1 H NMR (400 MHz, CD3OD): d 8.56 (s, 1H), 7.99 (s, 1H), 5.85 (s, 1H), 5.85 (s, 1H), 3.01 (d, J = 4.8 Hz, 3H).

[0190] 13 C NMR (150 MHz, CD3OD): d 166.7, 154.1, 153.9, 138.8, 138.7, 137.1,134.9, 124.3, 121.9, 121.3, 95.6, 26.9.

[0191] HRMS (ESIMS): calcd for C 12 H 12 N5O [M+H] + : 242.1036. Found: 242.1032.

[0192] Example 5 Preparation of Compounds 3b-3e:

[0193] ;

[0194] First, compound 2a-2d was prepared according to the method of Example 3, then 0.2 mmol of compound 2a-2d was dissolved in 10 mL of methanol, 5 mL of aqueous ammonia was added, and the mixture was refluxed for 24 h under nitrogen protection. The mixture was cooled to room temperature, and the reaction solution was evaporated to dryness and subjected to silica gel column chromatography to obtain compound 3b-3e.

[0195] ①6-Acetylamino-8-aminopyrrolo[4,3,2-de]quinoline-4-carboxamide (Compound 3b):

[0196] ;

[0197] Dark red solid, yield: 78%, purity: 96.9%, melting point: >211℃ (decomp.).

[0198] 1 H NMR (400 MHz, CD3OD): d 8.63 (s, 1H), 8.09 (s, 1H), 5.97 (s, 1H), 2.33 (s, 3H).

[0199] 13 C NMR (150 MHz, CD3OD): d 170.1, 169.5, 156.8, 151.7, 146.3, 143.5,143.1, 137.9, 135.3, 128.6, 123.7, 110.6, 24.5.

[0200] HRMS (ESIMS): calcd for C 13 H 12 N5O2[M+H] + : 270.0986. Found: 270.0972.

[0201] ②6-Trifluoroamido-8-aminopyrrolo[4,3,2-de]quinoline-4-carboxamide (Compound 3c):

[0202] ;

[0203] Dark red solid, yield: 71%, purity: 96.5%, melting point: >243℃ (decomp.).

[0204] 1 H NMR (400 MHz, CD3OD): d 8.71 (s, 1H), 8.13 (s, 1H), 6.08 (s, 1H).

[0205] 13 C NMR (150 MHz, CD3OD): d 171.6, 169.8, 157.3, 152.6, 147.5, 145.1,144.3, 138.8, 136.7, 129.6, 124.5, 116.8, 111.2.

[0206] HRMS (ESIMS): calcd for C 13 H9F3N5O2[M+H] + : 324.0703. Found: 324.0698.

[0207] ③6-Propanylamino-8-aminopyrrolo[4,3,2-de]quinoline-4-carboxamide (Compound 3d):

[0208] ;

[0209] Dark red solid, yield: 76%, purity: 97.1%, melting point: >223℃ (decomp.).

[0210] 1 H NMR (400 MHz, CD3OD): d 8.62 (s, 1H), 8.08 (s, 1H), 5.96 (s, 1H), 2.66 (q, J = 7.2 Hz, 2H), 1.18 (t, J = 7.2 Hz, 3H).

[0211] 13 C NMR (150 MHz, CD3OD): d 169.9, 169.1, 156.6, 151.5, 146.1, 143.3,143.0, 137.7, 135.1, 128.5, 123.6, 110.3, 32.3, 11.6.

[0212] HRMS (ESIMS): calcd for C 14 H 14 N5O2[M+H] + : 284.1142. Found: 284.1133.

[0213] ④6-isobutyrylamino-8-aminopyrrolo[4,3,2-de]quinoline-4-carboxamide (Compound 3e):

[0214] ;

[0215] Dark red solid, yield: 80%, purity: 97.8%, melting point: >230℃ (decomp.).

[0216] 1 H NMR (400 MHz, CD3OD): d 8.65 (s, 1H), 8.11 (s, 1H), 5.99 (s, 1H), 3.08 (m, 1H), 1.23 (d, J = 6.8 Hz, 6H).

[0217] 13 C NMR (150 MHz, CD3OD): d 170.2, 169.3, 156.7, 151.5, 146.6, 143.3,143.1, 137.7, 135.1, 128.8, 123.6, 110.8, 32.2, 19.8.

[0218] HRMS (ESIMS): calcd for C 15 H 16 N5O2[M+H] + : 298.1299. Found: 298.1281.

[0219] Example 6 Preparation of Compounds 4b-4e:

[0220] ;

[0221] First, compound 2a-2d was prepared according to the method of Example 3, then 0.2 mmol of compound 2a-2d was dissolved in 10 mL of methanol, 5 mL of methylamine aqueous solution was added, and the mixture was refluxed for 24 h under nitrogen protection, cooled to room temperature, and the reaction solution was evaporated to dryness and subjected to silica gel column chromatography to obtain compound 4b-4e.

[0222] ①6-Acetylamino-8-amino-N-methylpyrrolo[4,3,2-de]quinoline-4-carboxamide (Compound 4b):

[0223] ;

[0224] Dark red solid, yield: 63%, purity: 97.3%, melting point: >219℃ (decomp.).

[0225] 1 H NMR (400 MHz, CD3OD): d 8.62 (s, 1H), 8.07 (s, 1H), 5.95 (s, 1H), 3.00 (d, J = 4.8 Hz, 3H), 2.32 (s, 3H).

[0226] 13 C NMR (150 MHz, CD3OD): d 170.2, 169.4, 156.5, 151.3, 146.1, 143.2,143.0, 137.7, 135.1, 128.3, 123.1, 110.3, 26.9, 24.6.

[0227] HRMS (ESIMS): calcd for C 14 H 14 N5O2[M+H] + : 284.1142. Found: 284.1137.

[0228] ②6-Trifluoroamido-8-amino-N-methylpyrrolo[4,3,2-de]quinoline-4-carboxamide (Compound 4c):

[0229] ;

[0230] Dark red solid, yield: 51%, purity: 97.8%, melting point: >256℃ (decomp.).

[0231] 1 H NMR (400 MHz, CD3OD): d 8.70 (s, 1H), 8.12 (s, 1H), 6.06 (s, 1H), 3.01 (d, J = 4.8 Hz, 3H).

[0232] 13 C NMR (150 MHz, CD3OD): d171.5, 169.7, 157.1, 152.3, 147.4, 145.3,144.1, 138.7, 136.5, 129.3, 124.1, 116.7, 111.3, 27.2.

[0233] HRMS (ESIMS): calcd for C 14 H 11 F3N5O2[M+H] + : 338.0859. Found: 338.0851.

[0234] ③6-Propanylamino-8-amino-N-methylpyrrolo[4,3,2-de]quinoline-4-carboxamide (Compound 4d):

[0235] ;

[0236] Dark red solid, yield: 67%, purity: 97.6%, melting point: >229℃ (decomp.).

[0237] 1 H NMR (400 MHz, CD3OD): d 8.63 (s, 1H), 8.07 (s, 1H), 5.98 (s, 1H), 2.98 (d, J = 4.8 Hz, 3H), 2.65 (q, J = 7.2 Hz, 2H), 1.18 (t, J = 7.2 Hz, 3H).

[0238] 13 C NMR (150 MHz, CD3OD): d 169.8, 169.3, 156.5, 151.7, 146.3, 143.4,143.2, 137.6, 135.3, 128.7, 123.5, 110.4, 32.3, 26.9, 11.6.

[0239] HRMS (ESIMS): calcd for C 15 H 16 N5O2[M+H] + : 298.1299. Found: 298.1296.

[0240] ④6-isobutyrylamino-8-amino-N-methylpyrrolo[4,3,2-de]quinoline-4-carboxamide (Compound 4e):

[0241] ;

[0242] Dark red solid, yield: 69%, purity: 98.1%, melting point: >238℃ (decomp.).

[0243] 1 H NMR (400 MHz, CD3OD): d 8.66 (s, 1H), 8.11 (s, 1H), 5.98 (s, 1H), 3.08 (m, 1H), 2.99 (d, J = 4.8 Hz, 3H), 1.23 (d, J = 6.8 Hz, 6H).

[0244] 13 C NMR (150 MHz, CD3OD): d 169.7, 169.1, 156.6, 151.4, 146.3, 143.1,143.0, 137.6, 135.3, 128.9, 123.8, 110.6, 32.3, 26.9, 19.8.

[0245] HRMS (ESIMS): calcd for C 16 H 18 N5O2[M+H] + : 312.1455. Found: 312.1449.

[0246] Preparation of Comparative Example Compounds 1a-1d

[0247] (1) Preparation of compound ad:

[0248] ;

[0249] First, compound III was prepared according to the method of Example 1, and then 2.0 mmol of triethylamine was added to a 20 mL anhydrous dichloromethane solution of 1.0 mmol of compound III, and then 1.05 mmol of acyl chloride (R3Cl, R3 is selected from -COCH3, -COCF3, or After the reaction of the raw materials is completed, water is added to the reaction solution, extracted with dichloromethane, washed with saturated brine, dried over anhydrous sodium sulfate, the solvent is evaporated, and then silica gel column chromatography is performed to obtain compound ad.

[0250] ①1-tert-Butyloxycarbonylamino-8-acetylamino-6-aminopyrrolo[4,3,2-de]quinoline-1,4(2 H)-4-Ethyl carboxylate (Compound a):

[0251] ;

[0252] Orange solid, yield: 89%, purity: 98.1%, melting point: >233℃ (decomp.).

[0253] 1 H NMR (400 MHz, DMSO- d 6): d 11.18 (brs, 1H), 7.86 (s, 1H), 7.80 (s,1H), 5.75 (brs, 2H), 5.26 (s, 2H), 4.40 (q, J = 7.2 Hz, 2H), 2.07 (s, 3H),1.55 (s, 9H), 1.35 (t, d= 7.2 Hz, 3 H).

[0254] 13 C NMR (150 MHz, DMSO- d 6): d 165.4, 158.5, 146.5, 146.4, 142.4, 140.9,131.8, 128.3, 124.4, 113.6, 82.6, 61.7, 56.5, 28.4, 14.6.

[0255] HRMS (ESI) m / z: [M+H] + Calculate for C 20 H 25 N4O5401.1819; found: 401.1818.

[0256] ②1-tert-Butyloxycarbonylamino-8-trifluoroacetylamino-6-aminopyrrolo[4,3,2-de]quinoline-1,4(2 H )-4-carboxylic acid ethyl ester (compound b):

[0257] ;

[0258] Orange-yellow solid, yield: 21%, purity: 95.5%, melting point: >256℃ (decomp.).

[0259] 1 H NMR (400 MHz, DMSO- d 6): d12.62 (brs, 1H), 9.06 (s, 1H), 8.01 (s,1H), 5.93 (brs, 2H), 5.38 (s, 2H), 4.41 (q, J = 7.2 Hz, 2H), 1.56 (s, 9H),1.35 (t, d= 7.2 Hz, 3 H).

[0260] 13 C NMR (150 MHz, DMSO- d 6): d 166.3, 158.8, 146.7, 146.5, 143.1, 141.2,131.6, 128.2, 124.3, 116.7, 113.8, 86.7, 62.8, 55.1, 28.6, 14.6.

[0261] HRMS (ESI) m / z: [M+H] + Calculate for C 20 H 22 N4O5F3455.1537; found: 455.1533.

[0262] ③1-tert-Butyloxycarbonylamino-8-propionylamino-6-aminopyrrolo[4,3,2-de]quinoline-1,4(2 H )-4-carboxylic acid ethyl ester (Compound c):

[0263] ;

[0264] Orange solid, yield: 78%, purity: 98.3%, melting point: >238℃ (decomp.).

[0265] 1 H NMR (400 MHz, DMSO- d 6): d 11.22 (brs, 1H), 7.87 (s, 1H), 7.81 (s,1H), 5.76 (brs, 2H), 5.25 (s, 2H), 4.41 (q, J = 7.2 Hz, 2H), 2.51 (q, J = 7.2Hz, 2H), 1.54 (s, 9H), 1.35 (t, d= 7.2 Hz, 3H), 1.16 (t, J = 7.2 Hz, 3H).

[0266] 13C NMR (150 MHz, DMSO- d 6): d 165.6, 158.1, 146.3, 146.3, 142.5, 140.7,131.6, 128.7, 124.3, 113.8, 82.9, 61.8, 56.3, 31.1, 28.5, 14.6, 10.7.

[0267] HRMS (ESI) m / z: [M+H] + Calculate for C 21 H 27 N4O5415.1976; found: 415.1971.

[0268] ④1-tert-Butyloxycarbonylamino-8-isobutyrylamino-6-aminopyrrolo[4,3,2-de]quinoline-1,4(2 H )-4-carboxylic acid ethyl ester (Compound d):

[0269] ;

[0270] Orange solid, yield: 83%, purity: 98.6%, melting point: >249℃ (decomp.).

[0271] 1 H NMR (400 MHz, DMSO- d 6): d 11.16 (brs, 1H), 7.88 (s, 1H), 7.80 (s,1H), 5.73 (brs, 2H), 5.27 (s, 2H), 4.41 (q, J = 7.2 Hz, 2H), 2.81 (m, 1H),1.56 (s, 9H), 1.35 (t, d= 7.2 Hz, 3 H), 1.21 (d, J = 6.8 Hz, 6H).

[0272] 13 C NMR (150 MHz, DMSO- d 6): d 165.6, 158.1, 146.3, 146.3, 142.5, 140.7,131.6, 128.7, 124.3, 113.8, 82.9, 61.8, 56.3, 35.7, 28.5, 19.8, 14.6.

[0273] HRMS (ESI) m / z: [M+H] + Calculate for C 22 H 29 N4O5429.2132; found: 429.2128.

[0274] (2) Preparation of compounds 1a-1d:

[0275] ;

[0276] To a 10 mL anhydrous dichloromethane solution of 0.5 mmol of compound ad, add 3.0 mmol of anhydrous zinc bromide, stir at room temperature after the addition is complete. After the reaction of the raw materials is complete, add saturated sodium bicarbonate aqueous solution to the reaction solution, extract with dichloromethane, wash with saturated brine, dry with anhydrous sodium sulfate, evaporate the solvent, and then perform silica gel column chromatography to obtain compounds 1a-1d.

[0277] ①6-Amino-8-acetylaminopyrrolo[4,3,2-de]quinoline-4-carboxylic acid ethyl ester (Compound 1a):

[0278] ;

[0279] Purple solid, yield: 89%, purity: 99.1%, melting point: >268℃ (decomp.).

[0280] 1 H NMR (400 MHz, DMSO- d 6): d 10.74 (brs, 1H), 8.66 (s, 1H), 8.34 (s,1H), 7.72 (s, 1H), 7.58 (brs, 2H), 4.44 (q, J = 7.2 Hz, 2H), 2.25 (s, 3H),1.40 (t, d= 7.2 Hz, 3 H).

[0281] 13 C NMR (150 MHz, DMSO- d 6): d 170.4, 165.3, 154.1, 145.2, 142.7, 142.3,139.2, 134.2, 126.2, 123.2, 121.1, 99.7, 61.2, 24.5, 14.

[0282] HRMS (ESIMS): calcd for C 15 H 15N4O3[M+H] + : 299.1139. Found: 299.1123.

[0283] ②6-Amino-8-trifluoroacetylaminopyrrolo[4,3,2-de]quinoline-4-carboxylic acid ethyl ester (Compound 1b):

[0284] ;

[0285] Purple solid, yield: 63%, purity: 97.3%, melting point: >276℃ (decomp.).

[0286] 1 H NMR (400 MHz, DMSO- d 6): d 10.91 (brs, 1H), 8.79 (s, 1H), 8.45 (s,1H), 7.88 (s, 1H), 7.63 (brs, 2H), 4.44 (q, J = 7.2 Hz, 2H), 1.41 (t, d= 7.2Hz, 3 H).

[0287] 13 C NMR (150 MHz, DMSO- d 6): d 171.6, 166.4, 154.5, 145.7, 142.9, 142.5,139.9, 134.6, 126.3, 123.1, 121.4, 117.1, 99.9, 62.9, 14.3.

[0288] HRMS (ESI) m / z: [M+H] + Calculate for C 15 H 12 N4O3F3353.0856; found: 353.0853.

[0289] ③6-amino-8-propionylaminopyrrolo[4,3,2-de]quinoline-4-carboxylic acid ethyl ester (Compound 1c):

[0290] ;

[0291] Purple solid, yield: 81%, purity: 98.7%, melting point: >263℃ (decomp.).

[0292] 1 H NMR (400 MHz, DMSO-d 6): d 10.76 (brs, 1H), 8.65 (s, 1H), 8.33 (s,1H), 7.72 (s, 1H), 7.57 (brs, 2H), 4.44 (q, J = 7.2 Hz, 2H), 2.65 (q, J = 7.2Hz, 2H), 1.40 (t, d= 7.2 Hz, 3H) , 1.18 (t, J = 7.2 Hz, 3H).

[0293] 13 C NMR (150 MHz, DMSO- d 6): d 170.3, 165.1, 154.0, 145.3, 142.6, 142.3,139.1, 134.3, 126.3, 123.1, 121.4, 99.6, 61.6, 32.1, 14.2, 11.8.

[0294] HRMS (ESIMS): calcd for C 16 H 17 N4O3[M+H] + : 313.1295. Found: 313.1287.

[0295] ④6-Amino-8-isobutyrylaminopyrrolo[4,3,2-de]quinoline-4-carboxylic acid ethyl ester (Compound 1d):

[0296] ;

[0297] Purple solid, yield: 76%, purity: 98.1%, melting point: >245℃ (decomp.).

[0298] 1 H NMR (400 MHz, DMSO- d 6): d 10.73 (brs, 1H), 8.62 (s, 1H), 8.31 (s,1H), 7.71 (s, 1H), 7.58 (brs, 2H), 4.44 (q, J = 7.2 Hz, 2H), 3.08 (m, 1H),1.40 (t, d= 7.2 Hz, 3 H), 1.23 (d, J = 6.8 Hz, 6H).

[0299] 13C NMR (150 MHz, DMSO- d 6): d 170.4, 165.3, 154.1, 145.4, 142.3, 142.1,139.3, 134.5, 126.6, 123.2, 121.3, 99.1, 61.9, 32.1, 21.3, 14.2.

[0300] HRMS (ESIMS): calcd for C 17 H 19 N4O3[M+H] + : 327.1452. Found: 327.1449.

[0301] Test Case

[0302] Using the CCK-8 method, the anti-tumor activity of 2a-2d, 3a-3e and 4a-4e synthesized in the examples and 1a-1d synthesized in the comparative examples was tested at the in vitro cell level. The results are shown in Table 2. It was found that the anti-cancer activity of the compound 1a-1d whose amino group at position 8 was acylated was greatly reduced. The anti-cancer activity of the compound 2a-2d and the compound 3c whose amino group at position 8 was not acylated and R1 was -CONH2 was better than that of the clinical first-line anti-tumor drug oxaliplatin. Compounds 3a, 3b, 3d, 3e and 4a-4e showed anti-tumor activity that was significantly better than oxaliplatin, and the inhibitory activity of at least one of the human tumor cell lines LoVo, RKO, HCT-116, HT-29 and DLD-1 was better than that of Lymphostin. Among them, the anti-tumor activity of compounds 4b-4e was the best, reaching the nanomolar level.

[0303] Table 2 In vitro antitumor activity results of pyrrolo[4,3,2-de]quinoline derivatives and positive controls (IC 50 value)

[0304]

[0305] The anti-kinase activity of compounds 4b-4e with the best anti-tumor activity was tested using the method reported in the literature (Kashem, MA et al. Three mechanistically distinct kinase assays compared: Measurement of intrinsic ATPase activity identified the most comprehensive set of ITK inhibitors. J. Biomol. Screen. 2007, 12(1), 70-83.).

[0306] The results are shown in Table 3. These four compounds showed nanomolar inhibitory activity against the four PI3K isoforms (PI3Kα, PI3Kβ, PI3Kγ and PI3Kδ) and the key regulatory factor mTOR. They are typical PI3K / mTOR dual inhibitors, and their inhibitory activity is comparable to that of the positive control drug, the classic PI3K / mTOR dual inhibitor BEZ235 (which entered Phase II clinical trials).

[0307] It is worth mentioning that the inhibitory activity of compounds 4b, 4c and 4d against at least one kinase surpassed that of Lymphostin, among which compound 4b showed very low IC on all tested targets. 50 The value of 5.3447 μmol / L was 0.177 μmol / L, which showed extremely strong inhibitory activity and was the best among the four compounds. This result marks an important progress in the field of optimizing kinase inhibitors and provides a strong candidate molecule for the development of new strategies for cancer treatment.

[0308] Table 3 Kinase inhibitory activity results (IC 50 value)

[0309]

[0310] Although the present invention has been described in detail by way of preferred embodiments, the present invention is not limited thereto. Without departing from the spirit and essence of the present invention, a person of ordinary skill in the art may make various equivalent modifications or substitutions to the embodiments of the present invention, and such modifications or substitutions shall be within the scope of the present invention. Any person of ordinary skill in the art may easily conceive of changes or substitutions within the technical scope disclosed by the present invention, and such changes or substitutions shall be within the scope of protection of the present invention.

Claims

1. A pyrrolo[4,3,2-de]quinoline derivative, characterized in that: The structure is shown in Formula I below: ； In the formula, R1 is -CONH2, R2 is selected from hydrogen or ; or, R1 is -CONHCH3, R2 is selected from hydrogen, -COCH3, -COCF3, or .

2. The pyrrolo[4,3,2-de]quinoline derivative according to claim 1, characterized in that Selected from the following compounds: 、 、 、 。 3. The pyrrolo[4,3,2-de]quinoline derivative according to claim 1, characterized in that Selected from the following compounds: 、 、 。 4. The pyrrolo[4,3,2-de]quinoline derivative according to claim 1, characterized in that The synthesis method comprises the following steps: (1) Using 1,5-difluoro-2,4-dinitrobenzene as the starting material, compound VIII is obtained by ammoniation; (2) Compound VIII and compound VII are reacted via Doebner-Miller reaction to synthesize quinoline ring compound VI; (3) protecting the 5-amino group of compound VI with Boc to obtain an intermediate compound V with mono-Boc protection of the amino group; (4) Reaction of compound V with bromination reagent phenyltrimethylammonium tribromide to obtain dihydropyrroloquinoline ring compound IV; (5) The nitro groups at the 6- and 8-positions of compound IV are reduced by catalytic hydrogenation to obtain compound III; (6) When R2 is hydrogen, compound III is subjected to a Boc protecting group removal reaction to obtain compound II; When R2 is -COCH3, -COCF3, or When the compound III undergoes substitution reaction and deprotection reaction, the compound II is obtained; (7) When R1 is -CONH2, compound II is reacted with aqueous ammonia to obtain compound I; When R1 is -CONHCH3, compound II is reacted with aqueous methylamine solution to obtain compound I; The synthetic route is as follows: 。 5. The pyrrolo[4,3,2-de]quinoline derivative according to claim 4, characterized in that Step (4) is as follows: Add phenyltrimethylammonium tribromide to a tetrahydrofuran solution of compound V, react at 30-60°C, add water to the reaction solution, extract with ethyl acetate, wash with saturated sodium bicarbonate solution, wash with saturated brine, dry with anhydrous sodium sulfate, evaporate the solvent, and then perform silica gel column chromatography to obtain a red solid which is compound IV.

6. The pyrrolo[4,3,2-de]quinoline derivative according to claim 4, characterized in that Step (5) is as follows: Dissolve compound IV in a mixed solvent of dichloromethane and methanol, add catalyst Pd / C, replace hydrogen, stir and react at room temperature, filter, wash with methanol, evaporate the solvent, and then perform silica gel column chromatography to obtain a red solid, which is compound III.

7. The pyrrolo[4,3,2-de]quinoline derivative according to claim 4, characterized in that When R2 is hydrogen, step (6) is specifically: Dissolve compound III in dichloromethane, add anhydrous zinc bromide, stir and react at room temperature, add saturated sodium bicarbonate aqueous solution to the reaction solution, extract with dichloromethane, wash with saturated brine, dry with anhydrous sodium sulfate, evaporate the solvent, and then perform silica gel column chromatography to obtain compound II.

8. The pyrrolo[4,3,2-de]quinoline derivative according to claim 4, characterized in that When R2 is -COCH3, -COCF3, or When , step (6) is specifically as follows: Compound III is first reacted with o-nitrobenzenesulfonyl chloride to obtain compound XI; then, R2Cl is added to an anhydrous dichloromethane solution of compound XI under a triethylamine environment to react to obtain compound X; anhydrous zinc bromide is added to an anhydrous dichloromethane solution of compound X, a saturated sodium bicarbonate aqueous solution is added to the reaction solution, dichloromethane extraction is performed, saturated brine is washed, anhydrous sodium sulfate is dried, the solvent is evaporated, and silica gel column chromatography is performed; the obtained product is dissolved in N,N-dimethylformamide, anhydrous potassium carbonate and thiophenol are added, stirred at room temperature and reacted, sodium bicarbonate aqueous solution is added to the reaction solution, dichloromethane extraction is performed, water is washed, saturated brine is washed, anhydrous sodium sulfate is dried, the solvent is evaporated, and then silica gel column chromatography is performed to obtain compound II; The reaction route is as follows: 。

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