Indole pyrazole c-n axis chiral compound and synthesis method thereof

Abstract

The application relates to the technical field of chemical synthesis, and particularly discloses an indole pyrazole C-N axis chiral compound and a synthesis method thereof. The structural formula of the indole pyrazole C-N axis chiral compound is shown in formula (I), the compound is a kind of axis chiral compound with high resistance to selection, R1 is a substituted or unsubstituted phenyl, naphthyl or alkyl, R2 is a phenyl, R3 is a substituted or unsubstituted phenyl, and R4 is an ethyl. The compound shown in formula (I) can inhibit the growth of cancer cells, thereby interfering with the malignant behavior of the cancer cells.

Description

Technical Field

[0001] This invention belongs to the field of chemical synthesis technology, specifically relating to an indolepyrazole compound with a chiral CN axis and its synthesis method. Background Technology

[0002] Discovering a scaffold structure with pharmacological activity, a broader range of effects, and more significant therapeutic efficacy has always been a fascinating yet highly challenging research field. Indolepyrazole derivatives are fused heterocyclic systems formed by the fusion of indole and pyrazole rings through shared edges, creating a relatively rigid and well-planar fused heterocyclic system. Due to their unique structural characteristics, they represent scaffolds of immense medical value. Specifically, indolepyrazole possesses a variety of biological activities, including anti-inflammatory, anticancer, anticonvulsant, and other activities. For example, indolepyrazole is present as a core component in various antiemetics, hypnotics, and drugs such as Alectinib and Granise. In recent years, scientists' research on fused heterocyclic systems has moved beyond traditional broad-spectrum activity screening and entered a stage of precise design and functionalization. Developing drugs with superior activity and fewer side effects, starting from stereochemical dimensions such as axial chirality, has become a current hot topic.

[0003] Axial chirality is a form of stereoisomerism that arises from the pairing of four substituents in a non-planar arrangement around a stereogenetic axis. Key examples include trans-restricted isomers, spirolones, allenes, and spirochiral structures. Among all these subclasses, trans-restricted isomers have attracted considerable attention due to their wide applications in medicinal chemistry, chiral materials, ligands, and organocatalysts. Research on the construction of axially chiral diaryl compounds has matured in recent decades. The construction strategies for axially chiral diaryl compounds are mainly divided into four types: first, conventional resolution, catalytic kinetic resolution, and dynamic kinetic resolution; second, direct aryl-aryl coupling; third, construction of a new unit and direct coupling with an aryl group; and fourth, the transformation from a chiral center to axial chirality. Compared to the construction of axially chiral diaryl compounds, the development of axially chiral diaryl compounds with five-membered heterocyclic rings has been slow due to inherent limitations, and related reports are scarce.

[0004] In view of this, the present invention is proposed. Summary of the Invention

[0005] To address the shortcomings of existing technologies, the present invention aims to provide a synthetic method for a class of indolepyrazole CN-axis chiral compounds. This synthetic method features mild and simple reaction conditions and good substrate versatility. The structural formula of these compounds is shown in formula (I): ; R1 is a substituted or unsubstituted phenyl, naphthyl, or alkyl group; R2 is a phenyl group; R3 is a substituted or unsubstituted phenyl group; and R4 is an ethyl group. The compound is an axially chiral compound with high resistance selectivity.

[0006] Furthermore, R1 is selected from one of phenyl, p-fluorophenyl, p-iodophenyl, p-methoxyphenyl, p-cyanophenyl, tert-butyl, and naphthyl; R3 is selected from one of phenyl, p-chlorophenyl, and p-bromophenyl.

[0007] The synthesis method of compound (I) is as follows: using the first substrate and the second substrate as raw materials, chiral phosphoric acid as catalyst, under the protection of inert gas, in reaction solvent A, at 70~90℃ for 10~55 hours, to obtain the target product (I); The structural formula of the first substrate is: ; The structural formula of the second substrate is: ; The reaction solvent A is 1,1,2,2-tetrachloroethane or carbon tetrachloride; The chiral phosphate is selected from one of (s)-3,3'-bis-9-phenanthroline-1,1'-binaphthol phosphate, (s)-3,3'-bis(2-naphthyl)-5,5',6,6',7,7',8,8'-octahydro-1,1'-binaphthol phosphate, (s)-binaphthol phosphate, and (s)-3,3'-bis[3,5-bis(trifluoromethyl)phenyl]-1,1'-binaphthol phosphate.

[0008] The preferred method for synthesizing compound (I) is as follows: In an inert atmosphere, the first substrate 2,3-diketone ester and the enamine compound derived from the second substrate 1,3-diketone are condensed at 70°C for 10–55 hours in the presence of (S)-3,3'-bis-9-phenanthroline-1,1'-binaphthol phosphate, using carbon tetrachloride as a solvent, to generate an imine intermediate. This intermediate undergoes isomerization to an enamine intermediate. Subsequently, activated by the formation of two hydrogen bonds with chiral phosphoric acid, the enamine intermediate undergoes chiral selective intramolecular cyclization to generate a centrally chiral intermediate. Finally, with the aid of chiral phosphoric acid, the dehydration reaction of the centrally chiral intermediate produces a pyrrole ring, thereby yielding the target product. The reaction formula for the synthesis is as follows:

[0009] Furthermore, the molar ratio of the first substrate, the second substrate, and chiral phosphoric acid is 1~3:1~2:0.05~0.2; Furthermore, the molar ratio of the first substrate, the second substrate, and chiral phosphoric acid is 1.2:1:0.1.

[0010] Furthermore, the first substrate, 2,3-diketone, was prepared according to the method described in the literature (Organic Chemistry Frontiers, 2016, 12(3), 1686-1690).

[0011] Furthermore, the preparation method of the enamine compound derived from the second substrate 1,3-diketone includes the following steps: adding 1,3-cyclohexanedione, 1,2-dichloroethane, and trifluoroacetic acid to a dry container under inert gas protection, then adding a pyrazole derivative, and heating the reaction mixture at 80°C overnight; after the reactants have reacted completely, concentrating to dryness, and purifying the crude product by silica gel column chromatography to obtain the enamine compound derived from 1,3-diketone.

[0012] The structural formula of the pyrazole derivative is: .

[0013] Furthermore, the molar ratio of 1,3-cyclohexanedione, trifluoroacetic acid, and pyrazole derivative is 1.35:0.2:1.

[0014] Compared with the prior art, the advantages and beneficial effects of the present invention are as follows: This invention successfully developed a novel chiral acid-catalyzed, transselective cyclization reaction. Through chiral phosphoric acid catalysis, a condensation reaction of a 2,3-diketone ester with a 1,3-diketone-derived enamine compound generates an imine intermediate, which is subsequently isomerized to an enamine intermediate. Activated by the formation of two hydrogen bonds with chiral phosphoric acid, the enamine intermediate undergoes chiral-selective intramolecular cyclization to generate a centrally chiral intermediate. Finally, with the aid of chiral phosphoric acid, the target product is generated. This invention provides a highly efficient and enantioselective method for the synthesis of chiral CN-axis indolepyrazole compounds. The discovery of the chiral phosphoric acid catalytic system is key to achieving high efficiency and enantioselectivity. Attached Figure Description

[0015] Figure 1 The 1H NMR spectrum of compound 3a provided in the specific implementation embodiment; Figure 2 The carbon NMR spectrum of compound 3a provided in the specific implementation embodiment; Figure 3 The 1H NMR spectrum of compound 3b provided in the specific implementation embodiment; Figure 4 The carbon NMR spectrum of compound 3b provided in the specific implementation embodiment; Figure 5 The 1H NMR spectrum of compound 3c provided in the specific embodiment; Figure 6 The carbon NMR spectrum of compound 3c provided in the specific implementation embodiment; Figure 7 The 3d nuclear magnetic resonance hydrogen spectrum of the compound provided in the specific embodiment; Figure 8 The image shows the carbon NMR spectrum of the compound at 3d as provided in the specific implementation embodiment. Figure 9 The nuclear magnetic resonance fluorine spectrum of compound 3d provided in the specific embodiment; Figure 10 The 1H NMR spectrum of compound 3e provided in the specific implementation embodiment; Figure 11 The carbon NMR spectrum of compound 3e provided in the specific implementation embodiment; Figure 12 The 1H NMR spectrum of compound 3f provided in the specific implementation embodiment;

[0016] Figure 13 The carbon NMR spectrum of compound 3f provided in the specific implementation embodiment; Figure 14 The hydrogen nuclear magnetic resonance spectrum of 3g of the compound provided in the specific embodiment; Figure 15 The carbon NMR spectrum of compound 3g provided in the specific embodiment; Figure 16 The 1H NMR spectrum of compound 3h provided in the specific implementation embodiment; Figure 17 The carbon NMR spectrum of compound 3h provided in the specific implementation embodiment; Figure 18 The 1H NMR spectrum of compound 3i provided in the specific implementation scheme; Figure 19 The carbon NMR spectrum of compound 3i provided in the specific implementation scheme; Figure 20 The X-ray diffraction pattern of compound 3i provided in the specific implementation scheme is shown. Detailed Implementation

[0017] The following examples are only used to illustrate specific embodiments of the present invention and do not limit the scope of protection of the present invention. Unless otherwise specified, the experimental methods used in the following examples are conventional methods; the materials and reagents used are commercially available unless otherwise specified.

[0018] In the following embodiments: EA stands for Ethyl Acetate. DCE stands for Dichloroethane (1,2-dichloroethane). TFA stands for Trifluoroacetic acid. CPA refers to Chiral Phosphoric Acid.

[0019] Example 1: Compound Preparation (1) The first substrate, 2,3-diketone ester, was synthesized according to the method in reference [1]:

[0020] In the above reaction formula, R3 is selected from one of phenyl, p-chlorophenyl, and p-bromophenyl, and R4 is ethyl.

[0021] [1] Jian Cui, Ya-Nan Duan, Jun Yu, Chi Zhang. Iodosobenzene-mediateddirect and efficient oxidation of β-dicarbonyls to vicinal tricarbonylscatalyzed by iron(iii) salts. Organic Chemistry Frontiers, 2016, 12(3), 1686-1690. (2) Preparation of enamine compounds derived from the second substrate 1,3-diketone:

[0022] In the above reaction formula, R1 is selected from one of phenyl, p-fluorophenyl, p-iodophenyl, p-methoxyphenyl, p-cyanophenyl, tert-butyl, and naphthyl, and R2 is phenyl.

[0023] In a dry container, 0.00405 mol of 1,3-cyclohexanedione, 20 mL of 1,2-dichloroethane, and 0.0006 mol of trifluoroacetic acid were added under nitrogen protection. Then, 0.003 mol of 4-methyl-1,3-diphenyl-1H-pyrazole-5-amine (a pyrazole derivative) was added. The reaction mixture was heated at 80 °C overnight and analyzed by TLC. After complete reaction, the eluent was evaporated to dryness under reduced pressure. Isocratic elution was performed using 200-300 mesh silica gel with a 1:1 volume ratio of petroleum ether and ethyl acetate as eluents. The eluent with an Rf value of 0.3 was collected and evaporated to dryness under reduced pressure to obtain the enamine derivative of 1,3-dione.

[0024] When R1 is p-fluorophenyl, p-iodophenyl, p-methoxyphenyl, p-cyanophenyl, tert-butyl, or naphthyl, the preparation method of the compound is the same as above, and the feed ratio is also the same.

[0025] (3) Preparation of compound 3a:

[0026] In a dry container, add the first substrate 2,3-diketone ester (0.24 mmol), the second substrate 1,3-diketone-derived enamine compound (0.2 mmol), and (s)-3,3'-di-9-phenanthyl-1,1'-binaphthol phosphate (0.02 mmol). Under nitrogen protection, add carbon tetrachloride solvent (2 mL) and heat to 70 °C. Analyze by TLC. After the starting material disappears, concentrate under reduced pressure and evaporate to dryness. Elute isocratically using 200-300 mesh silica gel with petroleum ether and ethyl acetate in a 50:1 v / v ratio. Develop with petroleum ether and ethyl acetate in a 10:1 v / v ratio. Collect the eluent with an Rf value of 0.6 and evaporate to dryness under reduced pressure to give compound 3a. Yield 70%, ee value (enantiomer excess) > 99%.

[0027] Enantioselectivity was determined using an Agilent HPLC system with a CHIRALPAK column. The chiral stationary phase was a Daicel Chiralpak AD-H column manufactured by Daicel Chiral Technologies (Shanghai) Co., Ltd., with a UV absorption detection wavelength of 254 nm.

[0028] Compound 3a: white solid; 1 H NMR (500 MHz, CDCl3) δ 11.31 (s, 1H), 7.81 - 7.75 (m, 2H), 7.49 -7.43 (m, 2H), 7.41 - 7.36 (m, 1H), 7.30 - 7.25 (m, 2H), 7.25 - 7.16 (m, 4H),7.06 (s, 2H), 6.87 (d, J = 7.8 Hz, 1H), 6.76 (t, J = 8.3 Hz, 3H), 4.11 - 3.99(m, 2H), 2.15 (s, 3H), 0.82 (t, J = 7.1 Hz, 3H). 13C NMR (126 MHz, CDCl3) δ168.13, 151.90, 150.07, 146.03, 139.26, 138.21, 133.23, 132.56, 130.10,129.14, 128.82, 128.62, 128.09, 127.30, 127.21, 126.71, 122.35, 115.17,114.24, 109.26, 107.29, 102.02, 61.26, 13.30, 9.33. HRMS (ESI) m / z: [M+H] + Calculated for C 33 H 27 N3O3H + 514.2125, found 514.2125. The preparation methods of compounds 3b to 3h are the same as those of 3a, and the raw material feeding ratios are also the same as those of 3a.

[0029] Compound 3b: white solid; 1 H NMR (600 MHz, CDCl3) δ 11.26 (s, 1H), 7.80 (dd, J = 8.3, 1.4 Hz,2H), 7.47 (t, J = 7.5 Hz, 2H), 7.39 (t, J = 7.4 Hz, 1H), 7.30 (t, J = 8.0 Hz,1H), 7.26 (s, 2H), 7.24 - 7.19 (m, 3H), 7.06 (s, 2H), 6.88 (d, J = 7.9 Hz,1H), 6.77 (t, J = 8.3 Hz, 3H), 4.08 (qq, J = 10.8, 7.2 Hz, 2H), 2.16 (s, 3H), 0.88 (t, J = 7.1 Hz, 3H). 13C NMR (126 MHz, CDCl3) δ 167.86, 151.92, 150.21,144.38, 139.25, 138.11, 135.08, 133.05, 132.28, 129.25, 128.66, 128.61,128.20, 127.52, 127.43, 127.30, 126.99, 122.26, 115.11, 114.25, 109.43,107.64, 102.03, 61.46, 29.73, 13.44, 9.34. HRMS (ESI) m / z calcd forC 33 H 26 ClN3O3[M+H] + =548.1735, found = 548.1753. Compound 3c: Yellow solid; 1 H NMR (600 MHz, CDCl3) δ 11.25 (s, 1H), 7.47 (t, J = 7.7 Hz, 2H), 7.39 (s, 1H), 7.30 (t, J = 8.0 Hz, 1H), 7.27 - 7.18 (m, 7H), 6.88 (d, J = 7.9Hz, 1H), 6.77 (t, J = 7.5 Hz, 3H), 4.08 (qq, J = 10.8, 7.2 Hz, 2H), 2.15 (s,3H), 0.88 (t, J = 7.2 Hz, 3H). 13 C NMR (126 MHz, CDCl3) δ 167.84, 151.93,150.21, 144.36, 139.26, 138.11, 133.04, 132.27, 130.47, 129.26, 129.10,128.67, HRMS (ESI) m / z: [M+H] + calculated for C 33 H 26 BrN3O3H + 592.1230, found 592.1230. Compound 3d: Yellow solid; 1 H NMR (600 MHz, CDCl3) δ 11.31 (s, 1H), 7.79 - 7.74 (m, 2H), 7.46 (t,J = 7.5 Hz, 2H), 7.38 (t, J = 7.4 Hz, 1H), 7.31 - 7.22 (m, 4H), 7.10 (s, 2H), 6.92 - 6.85 (m, 3H), 6.76 (d, J = 8.1 Hz, 1H), 6.73 - 6.68 (m, 2H), 4.15 -4.01 (m, 2H), 2.16 (s, 3H), 0.84 (t, J = 7.2 Hz, 3H). 13 C NMR (151 MHz, CDCl3)δ 168.05, 162.36, 160.72, 151.98, 150.21, 145.77, 139.18, 134.37, 133.07,132.60, 130.04, 128.94, 128.65, 128.18, 127.28, 126.83, 124.26, 124.20,116.08, 115.93, 115.16, 114.30, 109.38, 107.38, 101.84, 61.36, 13.32, 9.34. 19 FNMR (471 MHz, CDCl3) δ -113.91 (dt, J = 8.7, 4.3 Hz). HRMS (ESI) m / z calcdfor C 33 H 26 FN3O3[M+H] + =532.2031, found = 532.2047. Compound 3e: white solid; 1H NMR (600 MHz, CDCl3) δ 11.31 (s, 1H), 7.77 (d, J = 8.4 Hz, 2H), 7.44 (t, J = 7.7 Hz, 2H), 7.36 (t, J = 7.4 Hz, 1H), 7.27 (d, J = 8.6 Hz, 4H),7.10 (s, 2H), 6.86 (d, J = 7.9 Hz, 1H), 6.76 (d, J = 8.1 Hz, 1H), 6.72 - 6.64(m, 5H), 4.11 - 4.00 (m, 2H), 3.78 (s, 3H), 2.14 (s, 3H), 0.82 (t, J = 7.2Hz, 3H). 13 C NMR (151 MHz, CDCl3) δ 168.16, 158.74, 151.88, 149.62, 146.13,139.41, 133.34, 132.46, 131.50, 130.19, 128.85, 128.59, HRMS (ESI) m / z calcd for C 34 H 29 N3O4[M+H] + =544.2231, found =544.2250. Compound 3f: white solid; 1 H NMR (600 MHz, CDCl3) δ 11.30 (s, 1H), 7.74 (d, J = 9.7 Hz, 2H), 7.42 (t, J = 7.7 Hz, 2H), 7.32 (h, J = 7.3 Hz, 7H), 7.26 (s, 1H), 7.19 (t, J= 8.0 Hz, 1H), 6.79 (d, J = 7.9 Hz, 1H), 6.44 (d, J = 9.0 Hz, 1H), 4.22 -4.08 (m, 2H), 1.97 (s, 3H), 1.21 (s, 9H), 0.95 (d, J = 7.2 Hz, 3H). 13C NMR(151 MHz, CDCl3) δ 168.40, 151.73, 147.05, 146.19, 140.38, 134.12, 131.49,130.67, 129.20, 128.51, 127.55, 127.46, 127.00, 126.42, 114.93, 113.44,108.81, 107.23, 102.18, 61.56, 61.34, 29.33, 14.16, 13.45, 9.46. HRMS (ESI)m / z calcd for C 31 H 31 N3O3[M+H] + =494.2438, found =494.2438. Compound 3g: Pink solid; 1 H NMR (600 MHz, CDCl3) δ 11.31 (s, 1H), 7.77 (d, J = 7.3 Hz, 2H), 7.46 (t, J = 7.7 Hz, 2H), 7.39 (t, J = 7.4 Hz, 1H), 7.31 - 7.22 (m, 4H), 0.84 (t, J = 7.2 Hz, 3H). 13 CNMR (151 MHz, CDCl3) δ 168.05, 162.36, 160.72, 151.98, 150.21, 145.77,139.19, 134.40, 134.38, 133.08, 132.60, 130.04, 128.94, 128.66, 128.18,127.28, 126.83, 124.26, 124.20, 116.08, 115.93, 115.17, 114.30, 109.38,107.39, 101.84, 61.36, 29.73, 13.32, 9.34. HRMS (ESI) m / z calcd for C 34 H 26 N4O3[M+H] +=539.2078, found =539.2076. Compound 3h: white solid; 1 H NMR (600 MHz, CDCl3) δ 11.32 (s, 1H), 7.81 (ddd, J = 11.6, 7.8, 1.5Hz, 3H), 7.66 (s, 1H), 7.53 (d, J = 5.5 Hz, 1H), 7.49 - 7.44 (m, 4H), 7.41 -7.38 (m, 1H), 7.32 (t, J = 8.0 Hz, 1H), 7.26 (s, 3H), 7.21 - 7.18 (m, 2H), 6.94 - 6.89 (m, 3H), 6.85 (d, J = 8.1 Hz, 1H), 3.99 (q, J = 7.2 Hz, 2H), 2.19(s, 3H), 0.74 (t, J = 7.2 Hz, 3H). 13 C NMR (151 MHz, CDCl3) δ 168.07, 151.94,150.29, 146.10, 139.35, 135.63, 133.22, 133.17, 132.82, 132.02, 130.06,129.09, 128.82, 128.66, 128.27, 128.14, 127.64, 127.35, 127.13, 126.79,126.76, 126.39, 120.73, 120.71, 115.18, 114.43, 109.33, 107.28, 102.06,61.22, 13.21, 9.36. HRMS (ESI) m / z calcd for C 37 H 29 N3O3[M+H] + =564.2282, found =564.2282. Compound 3i: white solid; 1H NMR (600 MHz, CDCl3) δ 11.31 (s, 1H), 7.76 (d, J = 9.7 Hz, 2H), 7.52 (d, J = 8.8 Hz, 2H), 7.45 (d, J = 7.9 Hz, 2H), 7.39 (d, J = 7.3 Hz, 1H),7.30 - 7.22 (m, 4H), 7.09 (s, 2H), 6.87 (d, J = 10.8 Hz, 1H), 6.73 (d, J =8.1 Hz, 1H), 6.52 (s, 2H), 4.16 - 3.99 (m, 2H), 2.14 (s, 3H), 0.84 (t, J =7.2 Hz, 3H). 13 C NMR (151 MHz, CDCl3) δ 168.03, 152.00, 150.52, 145.74, 139.05,138.20, 137.97, 132.96, 132.52, 130.18, 130.02, 128.95, HRMS (ESI) m / z: [M+H] + Calculated for C 33 H 26 IN3O3H + 640.1092, found 640.1092. X-ray crystallography data: Crystallization process of compound 3i: Single crystals of 3i were obtained by slowly evaporating a mixed solution of petroleum ether and dichloromethane containing 3i at room temperature. The crystal data and structural refinement results of compound 3i, selected from suitable single crystals, are listed in Appendix Table 2.

[0030] Crystallographic data of compound 3i: C 33 H 26IN3O3 (M = 639.47 g / mol): Triclinic system, space group P-1 (No. 2), a = 9.0702(2) Å, b = 11.5298(4) Å, c = 14.9469(4) Å, α = 80.862(3)°, β = 74.630(2)°, γ = 68.161(3)°, V = 1395.88(8) Å 3 , Z = 2, T = 150.00(10) K, μ(Cu Kα)= 9.330 mm -1 Calculated density = 1.521 g / cm³ 3 14,383 diffraction points were measured (6.146° ≤ 2θ ≤ 149.58°), of which 5,459 independent diffraction points (Rint = 0.0315, Rsigma = 0.0228) were used for all calculations. The final R1 = 0.0364 (I>2σ(I)), wR2 = 0.0993 (all data).

[0031] The structural formula of compound 3b-3i is as follows: , ,

[0032] , ,

[0033] ,

[0034] Table 1: Yields of compounds 3a-3i

[0035] Table 2: Crystal data and structural refinement of compound 3i

[0036] Example 2: Cytotoxicity Experiment 1. Experimental Materials The normal human liver L02 cells, human gastric cancer AGS cells, and human liver cancer HepG2 cells used in this experiment were purchased from European certified cell culture collections; RPMI 1640 medium was purchased from Hyclone, USA; fetal bovine serum (FBS) was purchased from Biontech, Germany; penicillin-streptomycin solution (100×) was purchased from Beyotime Biotechnology Co., Ltd.; and tetramethylthiazolyl blue (MTT) and dimethyl sulfoxide were purchased from Sigma-Aldrich, USA.

[0037] 2. Experimental Methods 2.1 Cell Culture L02 cells, AGS cells, and HepG2 cells were cultured in RPMI 1640 complete medium (hereinafter referred to as RPMI 1640 medium) containing 10% fetal bovine serum (FBS) and 1% penicillin-streptomycin in an incubator at 37°C, 45-65% humidity, and 5% CO2. When the cells reached the logarithmic growth phase, they were passaged, experimented on, and cryopreserved.

[0038] 2.2 MTT assay for cell viability Cell plating and drug administration: L02 cells, AGS cells, and HepG2 cells in logarithmic growth phase and in good condition were collected and plated at a concentration of 1×10⁻⁶ cells / cells. 4 Cells were seeded at a density of [number] cells / well in 96-well plates and incubated at 37°C, 5% CO2 for 24 h. The old culture medium was then aspirated from the wells. The test compound, dissolved in DMSO, was diluted with RPMI 1640 medium to the corresponding concentrations (0.1, 1, 10, 100 μM) and added to each well of the 96-well plate at 100 μL. Three replicates were performed for each concentration. Each plate also included a positive control group (cisplatin, administered via the same route as the test compound group), a normal cell group (containing only cells and RPMI 1640 medium), and a blank control group (containing only RPMI 1640 medium and no cells). After administration, the 96-well plates were incubated at 37°C, 5% CO2 for 72 h. Cell viability was assessed by adding 20 μL of MTT solution (5 mg / mL) to each well after 72 h of incubation and continuing incubation for another 1.5 h. Subsequently, the culture medium in the well plate was removed, 150 μL of DMSO was added to each well, and the plate was placed on a horizontal shaker and shaken at medium speed for 5 min. The absorbance at 562 nm wavelength was then measured using a microplate reader.

[0039] Data Processing: The relative inhibition rate of cell growth by the drug was calculated using the following formula: Cell Inhibition Rate = [1 - (X - C0) / (C - C0)] × 100%, where C, C0, and X represent the average absorbance values ​​of the three wells in the normal cell group, blank control group, and drug-treated group, respectively. Finally, the cell inhibition rate curve was fitted using Graphpad Prism 5.0 software, and the IC50 of the cell growth inhibition rate of the test compound was calculated. 50 Value; and according to the formula SI-1 = IC50 for L02 cells 50 / IC of AGS cells 50 SI-2 = IC50 of L02 cells 50 / IC on HepG2 cells 50 Calculate the selectivity index of the compound.

[0040] 2.3 Experimental Results Table 3: IC50 of the cell inhibition rate of the compound50 value

[0041] The experimental results above show that compounds 3a-3i have an effect on the IC50 of AGS cells and HepG2 cells. 50 The values ​​were significantly lower than those of the positive control drug cisplatin, indicating that the compounds of this invention have a significant inhibitory effect on tumor cell growth and have the potential to be developed into novel anti-tumor drugs. The selectivity indices of compounds 3a-3i are all greater than 1, indicating that the compounds of this invention may have a wide therapeutic window, providing important preliminary safety evidence for further in vivo anti-tumor research and development.

Claims

1. An indolepyrazole CN-axis chiral compound, characterized in that, The structural formula of the compound is shown in formula (Ⅰ): ； Wherein, R1 is a substituted or unsubstituted phenyl, naphthyl or alkyl group, R2 is a phenyl group, R3 is a substituted or unsubstituted phenyl group, and R4 is an ethyl group.

2. The compound according to claim 1, characterized in that, R1 is selected from one of phenyl, p-fluorophenyl, p-iodophenyl, p-methoxyphenyl, p-cyanophenyl, tert-butyl, and naphthyl; R3 is selected from one of phenyl, p-chlorophenyl, and p-bromophenyl.

3. A method for preparing the compound according to claim 1 or 2, characterized in that, The process includes the following steps: using the first and second substrates as raw materials, chiral phosphoric acid as a catalyst, and under inert gas protection, reacting in a reaction solvent at 70-90°C for 10-55 hours to obtain the target compound; The structural formula of the first substrate is The structural formula of the second substrate is: .

4. The preparation method according to claim 3, characterized in that, The chiral phosphoric acid is selected from (s)-3,3'-bis-9-phenanthroline-1,1'-binaphthol phosphate, (s)-3,3'-bis(2-naphthyl)-5,5',6,6',7,7',8,8'-octahydro-1,1'-binaphthol phosphate, (s)-binaphthol phosphate, and (s)-3,3'-bis[3,5-bis(trifluoromethyl)phenyl]-1,1'-binaphthol phosphate; the reaction solvent is 1,1,2,2-tetrachloroethane or carbon tetrachloride.

5. The preparation method according to claim 4, characterized in that, The chiral phosphoric acid is (S)-3,3'-bis-9-phenanthroline-1,1'-binaphthol phosphate, the reaction temperature is 70℃, and the reaction solvent is carbon tetrachloride.

6. The preparation method according to claim 3, characterized in that, The molar ratio of the first substrate, the second substrate, and chiral phosphoric acid is 1~3:1~2:0.05~0.

2.

7. The preparation method according to claim 6, characterized in that, The molar ratio of the first substrate, the second substrate, and chiral phosphoric acid is 1.2:1:0.

1.

8. The preparation method according to claim 3, characterized in that, The second substrate was prepared by the following method: 1,3-cyclohexanedione, 1,2-dichloroethane, and trifluoroacetic acid were added to a dry container under inert gas protection, followed by the addition of a pyrazole derivative. The reaction mixture was heated at 80°C overnight. After the reactants had reacted completely, the mixture was concentrated to dryness, and the crude product was purified by silica gel column chromatography to obtain the second substrate. The structural formula of the pyrazole derivative is: .

9. The preparation method according to claim 8, characterized in that, The molar ratio of 1,3-cyclohexanedione, trifluoroacetic acid, and pyrazole derivative is 1.35:0.2:

1.

10. The use of the compound of claim 1 or 2 in the preparation of an antitumor drug; preferably, the use is in the preparation of a drug for treating gastric adenocarcinoma and / or liver cancer.

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