CN101602707A - Naphthaline lactam derivatives and application thereof in tumor cell proliferation inhibition - Google Patents

Abstract

A naphthalene lactam derivative and application thereof in tumor cell proliferation inhibition belong to the technical field of pharmaceutical chemistry. The naphthalene lactam derivative is a derivative with high biological active groups such as acetylene bonds, cyano groups, amino groups or amide structures and the like by a synthesis method of firstly carrying out dehydrocondensation on 2-bit carbonyl and a compound with active methylene by using different halogenated alkanes, introducing different electrophilic substituent groups into 6 bits, and then substituting active hydrogen of 1-bit imine. The experiment of the naphthaline lactam derivative on the proliferation inhibition of tumor cells is carried out by selecting a tetrazolium salt reduction method and aiming at 7721 human liver cancer cells MCF-7 human breast cancer cells Hela human cervical cancer cells or BGC-823 human gastric cancer cells, and the result shows that the naphthaline lactam derivative has good activity and selectivity on the proliferation inhibition of the tumor cells.

Description

A class of naphthalenolactam derivatives and their application in tumor cell proliferation inhibition

technical field

The invention relates to a class of naphthalene lactam derivatives and their application in inhibiting tumor cell proliferation, belonging to the technical field of medicinal chemistry.

Background technique

Naphtholactam has special biological activity due to its special lactam structure, which has always attracted the interest and attention of bioorganic chemists. Mari'a L.Lo'pez-Rodri'guez et al. interacted with the N-substituted naphthalene lactam with optimized structure and the 5-HT7R receptor, and the pKi values were all less than 7.3 (mari'a L.Lo'pez-Rodri'guez , et al. J. Med. Chem. 2003, 46, 5638-5650). Yansong Gu et al. introduced alkyl-substituted 1,4-cycloheptanediamine at the 6-position of naphthalene lactam, and synthesized a series of naphthalene lactam derivatives for regulating the disorder or deficiency of 5-HT ₂ C receptors (Gu et al .US Patent 6,667,303B1).

However, based on the inherent structure of naphthalene lactam, it is not easy to modify. Although it has a wide range of and good biological activities, the number of naphthalene lactam derivatives that have been reported so far is still limited.

Contents of the invention

In order to overcome the deficiencies in the prior art, the present invention provides a class of naphthyl lactam derivatives and their application in inhibiting tumor cell proliferation. Naphtholactam was used as the parent to make further modifications in order to screen out new derivatives with stronger tumor cell proliferation inhibitory activity.

The technical scheme of the present invention is: a class of naphthalene lactam derivatives has a general chemical structure formula:

in:

R ₁ is =C(CN) ₂ , =C(COOCH ₃ ) ₂ ,

R ₂ is -H, -Cl, -Br, -CN or -NO ₂ ;

R ₃ is -CH ₂ CH ₂ C≡CH, -CH ₂ CH ₂ Br, -CH ₂ CH ₂ NH ₂ ,

-CH ₂ CH ₂ NHCH ₂ CH ₂ N(CH ₃ ) ₂ , -CH ₂ CH ₂ NHCH ₂ CH ₂ N(CH ₂ CH ₃ ) ₂ ,

-CH ₂ CH ₂ NCH ₂ CH ₂ CH ₂ CH ₃ or

The naphthalenolactam derivatives are formulated into liquid medicines with different concentrations, and the tetrazolium salt reduction method is used to treat 7721 human liver cancer cells, MCF-7 human breast cancer cells, Hela human cervical cancer cells or BGC-823 human gastric cancer cells. The test method is to inoculate a certain number of tumor cells in the logarithmic growth phase in a 96-well microculture plate at 90 μl/well according to different tumor growth rates, and add 10 μl/well of the prepared drug solution after culturing for 24 hours. For cell lines, each concentration was replicated in triplicate. Set up a cell-free zero well. If the prepared medicinal solution has color, do a cell-free zero-adjustment hole for the corresponding medicinal solution concentration. After the tumor cells were cultured at 37°C and 5% CO ₂ for 48 hours, 20 μl/well of 5 mg/ml tetrazolium salt solution prepared with normal saline was added; Sodium bicarbonate-5% isobutanol-0.01mol/l HCl prepared 50 μl/well of the triple solution, and placed in a CO ₂ incubator overnight. Then measure the OD570 value with a microplate reader. The inhibitory rate of described naphthalene lactam derivative to cancer cell growth is calculated according to the following formula:

Tumor inhibition rate=(OD value of control group-OD value of treatment group)/OD value of control group×100%.

The derivatives are as follows: 2-[(1-propargyl-benzo[c,d]indole)-2(1H)-ylidene]malononitrile (derivative 1), 2-[(1- Bromoethyl-benzo[c,d]indole)-2(1H)-ylidene]malononitrile (derivative 2), 2-{[(2-aminoethyl)-benzo[c,d] Indole]-2(1H)-ylidene}malononitrile (derivative 3), (E)-N-{[1-(2-aminoethyl)-benzo[c,d]indole]-2 (1H)-ylidene}-4-nitroaniline (derivative 4), (E)-N-[(1-propargyl-benzo[c,d]indole)-2(1H)-ylidene] -4-nitroaniline (derivative 5), (E)-N-{[1-(2-aminoethyl)-benzo[c,d]indole]-2(1H)-ylidene}-4 -Carboxyaniline (derivative 6), (E)-N-[(1-propargyl-benzo[c,d]indole)-2(1H)-ylidene]-4-carboxyaniline (derivative 7 ), 2-[(1-propargyl-benzo[c,d]indole)-2(1H)-ylidene]malonate (derivative 8), 2-{[2-(N , N'-dimethyl-ethylenediamine)-ethyl-benzo[c,d]indole]-2(1H)-ylidene}malononitrile (derivative 9), 2-{[2-( N,N'-diethyl-ethylenediamine)-ethyl-benzo[c,d]indole]-2(1H)-ylidene}malononitrile (derivative 10), 2-[(2- Butylamine-ethyl-benzo[c,d]indole)-2(1H)-ylidene]malononitrile (derivative 11), 2-[(2-piperazine-ethyl-benzo[c, d]indole)-2(1H)-ylidene]malononitrile (derivative 12), 2-[benzo[c,d]indole-2(1H)-ylidene]malononitrile (derivative 13) , 2-[6-chloro-benzo[c,d]indole-2(1H)-ylidene]malononitrile (derivative 14), 2-[6-bromo-benzo[c,d]indole -2(1H)-ylidene]malononitrile (derivative 15), 2-[6-cyano-benzo[c,d]indole-2(1H)-ylidene]malononitrile (derivative 16) , 2-[6-nitro-benzo[c,d]indole-2(1H)-ylidene]malononitrile (derivative 17), (E)-N-[benzo[c,d]indole Indole-2(1H)-ylidene]-4-nitroaniline (derivative 18), (E)-N-[6-nitro-benzo[c,d]indole-2(1H)-ylidene] -4-nitroaniline (derivative 19), (E)-N-[6-cyano-benzo[c,d]indole-2(1H)-ylidene]-4-nitroaniline (derivative 20).

The synthetic method of described naphthalene lactam derivative is as follows:

The synthesis of derivatives 1-8 is carried out in two steps:

That is to use naphthalene lactam as the raw material, benzene, toluene or chlorobenzene as the solvent, add 1mL _POCl3 as the dehydrating agent, and the reaction temperature is between 100-120°C, respectively choose malononitrile or methyl malonate with active methylene Esters, or primary aromatic amines with electron-withdrawing groups such as p-nitroaniline or p-carboxyaniline undergo dehydration condensation to obtain corresponding intermediates. Among them, the reaction with malononitrile gave derivative 13, and the reaction with p-nitroaniline gave derivative 18.

After the above intermediates are purified, react with halogenated alkanes such as bromoethylamine, bromoacetylene or dibromoethane to obtain the corresponding de-HBr products. The reaction environment is selected from acetonitrile/t-BuOK system or DMF/NaOCH according to the degree of difficulty of the reaction. ₃ systems.

Derivatives 9-12 are derived from derivative 2, catalyzed by basic conditions, respectively refluxed with corresponding amines in acetonitrile, and HBr is removed to obtain corresponding products.

The synthetic route of derivative 14-20 is:

Still using naphthalene lactam as raw material, react with SO ₂ Cl ₂ in acetic acid at 50°C for 0-5h, with liquid Br ₂ in 1,2-dichloroethane at 50°C for 0.5h or with concentrated HNO ₃ Reaction in concentrated H ₂ SO ₄ under ice bath for 3h to obtain an intermediate in which R ₂ is -Cl, -Br, -NO ₂ .

The above intermediates were reacted with malononitrile/POCl ₃ at 100°C to obtain derivatives 14, 15, and 17.

The 6-nitro-1,8-naphthalenolactam in the above intermediate was reacted with p-nitroaniline/POCl ₃ at 120°C to obtain derivative 19.

The 6-bromo-1,8-naphtholactam in the above intermediate was reacted with CuCN in N-methylpyrrolidone (NMP) at 130° C. for 4 h to obtain 6-cyano-1,8-naphtholactam.

6-cyano-1,8-naphthalenolactam reacted with malononitrile/POCl ₃ at 100°C to obtain derivative 16.

6-cyano-1,8-naphthalenolactam reacted with p-nitroaniline/POCl ₃ at 120°C to obtain derivative 20.

The growth inhibitory activity of derivatives on tumor cells in vitro was tested by the reduction method of microculture tetrozolium (MTT) on 7721 human liver cancer cells, MCF-7 human breast cancer cells, Hela human cervical cancer cells and BGC-823 human gastric cancer cells. In vitro inhibition test.

The beneficial effect of the present invention is: this kind of derivative is based on naphthalene lactam, respectively dehydrogenates the 2-position carbonyl and the compound with active methylene by using different halogenated alkanes, and introduces different electrophilic Substituting the group, and then substituting the active hydrogen of the 1-position imine to obtain a class of derivatives with high biological activity groups such as acetylenic bond, cyano group, amine group or amide structure. The experiment on the inhibition of tumor cell proliferation by such derivatives was carried out by using the tetrazolium salt reduction method for 7721 human liver cancer cells, MCF-7 human breast cancer cells, Hela human cervical cancer or BGC-823 human gastric cancer cells. The results showed that , the naphthyl lactam derivatives have good activity and selectivity for inhibiting tumor cell proliferation.

Detailed ways

The present invention will be further described below by way of examples.

Example 1

Synthesis of 2-[(1-propargyl-benzo[c,d]indole)-2(1H)-ylidene]malononitrile (derivative 1):

(1) Add 1.69g of naphthalene lactam (0.01mol), 0.6g of malononitrile (0.01mol), and _1.1mL of POCl3 into a 25mL two-necked flask, add 15mL of toluene, heat to 100°C under magnetic stirring, and react at constant temperature for 4 hours. The reaction solution was poured into methanol, and after cooling, it was directly suction-filtered to obtain a brown solid crude product. After vacuum drying, the crude product was separated by column chromatography to obtain 1.82 g of yellow spongy crystals with a yield of 83%. Melting point: >300°C.

ESI-HRMS (m/z): Calculated: 217.064, Found: 217.0639.

¹ HNMR (d6-DMSO, 400MHz): δ(ppm): 5.30(s, 1H), 7.21(d, J=7.2Hz, 1H), 7.58(t, J ₁ =7.6Hz, J ₂ =8.0Hz, 1H), 7.68(d, J=8.0Hz, 1H), 7.83(t, _J1 =8.0Hz, _J2 =7.2Hz, 1H), 8.17(d, J=8.0Hz, 1H), 8.55(d, J=8.0Hz, 1H).

(2) Dissolve 0.217g of derivative 13 (1mmol) in 10mL of DMF, add 0.067g of NaOCH ₃ (1.2mmol), heat up to 60°C under magnetic stirring, add 1mL of bromoacetylene dropwise, and follow the reaction by thin-plate chromatography. After about 45min, The response is complete. The cooled reaction solution was poured into 150 mL of ice water, and the aqueous phase was extracted with dichloromethane. After the organic phase was dried with anhydrous MgSO ₄ , the crude product was obtained by rotary evaporation. Purified by column chromatography to obtain 0.18 g of golden yellow crystals with a yield of 71%. Melting point: >300°C.

ESI-HRMS (m/z): Calculated: 255.0796, Found: 255.0797.

¹ H NMR (d6-DMSO, 400MHz): δ (ppm): 2.50 (s, 1H), 5.23 (s, 2H), 7.23 (d, J = 7.2Hz, 1H), 7.62 (t, J ₁ = 8.0Hz , J ₂ =7.2Hz, 1H), 7.70(d, J=8.4Hz, 1H), 7.81(t, J ₁ =8.0Hz, J ₂ =8.4Hz, 1H), 8.14(d, J=8.0Hz, 2H), 8.69 (d, J = 7.6 Hz, 1H).

Example 2

Synthesis of 2-[(1-bromoethyl-benzo[c,d]indole)-2(1H)-ylidene]malononitrile (derivative 2):

Dissolve 0.217g of derivative 13 (1mmol) in 10mL of anhydrous acetonitrile, add 0.2g of K ₂ CO ₃ , 1mL of dibromoethane, and heat to reflux under magnetic stirring. After 5h, the reaction is complete, and the crude solid is obtained by direct vacuum evaporation, washed with water After drying in vacuo, a yellow solid was obtained. Purified by column chromatography to obtain 0.175 g of yellow crystals with a yield of 80% and a melting point of 232.3°C.

ESI-HRMS (m/z): Calculated: 323.0058, Found: 323.0067.

¹ HNMR (CDCl3, 400MHz): δ (ppm): 3.83 (t, J ₁ =6.4Hz, J ₂ =6.8Hz, 2H), 4.85 (t, J ₁ =6.4Hz, J ₂ =6.8Hz, 2H) , 7.25(d, J=7.6Hz, 1H), 7.61(t, _J1 =8.0Hz, _J2 =7.6Hz, 1H), 7.70(d, J=8.0Hz, 1H), 7.82(t, _J1 =8.0Hz, J ₂ =7.6Hz, 1H), 8.15(d, J=8.4Hz, 2H), 8.71(d, J=7.6Hz, 1H).

Example 3

Synthesis of 2-{[(2-aminoethyl)-benzo[c,d]indole]-2(1H)-ylidene}malononitrile (derivative 3):

Dissolve 0.217g of derivative 13 (1mmol) in 10mL of DMF, add 0.134g of NaOCH ₃ (2.4mmol), heat up to 100°C, react for half an hour, add 0.243g of bromoethylamine hydrobromide (1.2mmol), reflux Reaction 4h. The cooled reaction solution was poured into 150 mL of ice water, and the aqueous phase was extracted with dichloromethane. After the organic phase was dried with anhydrous MgSO ₄ , the crude product was obtained by rotary evaporation. Purified by column chromatography to obtain 0.176 g of yellow crystals with a yield of 68% and a melting point of 213.2°C.

¹ HNMR (d6-DMSO, 400MHz): δ (ppm): 1.84 (t, NH ₂ , 2H), 2.54 (m, NCH ₂ CH ₂ N, 2H), 2.87 (m, NCH ₂ CH ₂ N, 2H) , 6.32(d, J=7.6Hz, 1H), 6.54(t, J ₁ =7.6Hz, J ₂ =8.8Hz, 1H), 7.54(d, J=8.8Hz, 1H), 7.34(d, J= 7.6Hz, 1H), 7.35 (t, _J1 = 7.6Hz, _J2 = 7.2Hz, 1H), 7.82 (d, J = 7.2Hz, 1H).

ESI-HRMS (m/z): Calculated: 260.1062, Found: 260.1045.

Example 4

Synthesis of (E)-N-{[1-(2-aminoethyl)-benzo[c,d]indole]-2(1H)-ylidene}-4-nitroaniline (derivative 4):

(1) Add 1.69g of naphthalene lactam (0.01mol) and 1.65g of p-nitroaniline (0.012mol) into a 25mL two-necked flask, add 10mL of chlorobenzene, and heat to 120°C under magnetic stirring. Add 1.1mL POCl ₃ dropwise to the uniformly stirred reaction system. After constant temperature reaction for 2 hours, the solution turns brown. After cooling to room temperature, add 2mL methanol dropwise to the reaction solution. Washed with water to obtain a red solid crude product. After drying in vacuum, recrystallize with acetic acid to obtain 2.62 g of red crystals with a yield of 90%.

(2) Dissolve 0.289g derivative 18 (1mmol) into 10mL acetonitrile, add 0.269gt-BuOK (2.4mmol), heat up to reflux, react for half an hour, add 0.243g hydrobromide of bromoethylamine (1.2 mmol), reflux reaction 5h. The cooled reaction solution was poured into 150 mL of ice water, filtered with suction, and dried in vacuo to obtain a crude product. Purified by column chromatography to obtain 200 mg of reddish-brown crystals with a yield of 60% and a melting point of 152.3-153.6°C.

¹ HNMR (d6-DMSO, 400MHz): δ (ppm): 2.05 (t, NH ₂ , 2H), 2.94 (m, NCH ₂ CH ₂ N, 2H), 3.07 (m, NCH ₂ CH ₂ N, 2H) , 6.85(d, J=7.6Hz, 1H), 7.10(d, J=7.6Hz, 1H), 7.24(d, J=8.4Hz, 2H), 7.428(t, J ₁ =8.0Hz, J ₂ = 7.6Hz, 1H), 7.46(d, J=7.6Hz, 1H), 7.51(t, _J1 =7.6Hz, _J2 =8.4Hz, 1H), 7.89(d, J=8.0Hz, 1H), 8.31 (d, J=8.4Hz, 2H).

ESI-HRMS (m/z): Calculated: 332.1273, Found: 332.1258.

Example 5

E) Synthesis of -N-[(1-propargyl-benzo[c,d]indole)-2(1H)-ylidene]-4-nitroaniline (derivative 5):

Dissolve 0.289g of derivative 18 (1mmol) in 10mL of DMF, add 0.067g of NaOCH ₃ (1.2mmol), raise the temperature to 80°C under magnetic stirring, add 1mL of bromoacetylene dropwise, and track the reaction by thin-plate chromatography. After 2h, the reaction is complete. The cooled reaction solution was poured into 150 mL of ice water, and the aqueous phase was extracted with dichloromethane. After the organic phase was dried with anhydrous MgSO ₄ , the crude product was obtained by rotary evaporation. Purified by column chromatography to obtain 0.27 g of brown granular crystals with a yield of 70% and a melting point of 170.5-171.8°C.

ESI-HRMS (m/z): Calculated: 327.1008, Found: 327.0988.

¹ HNMR (CDCl3, 400MHz): δ (ppm): 2.31(s, 2H), 4.90(s, 2H), 6.96(d, J=7.2Hz, 1H), 7.07(d, J=7.2Hz, 1H) , 7.22(d, J=8.8Hz, 2H), 7.38(t, _J1 =7.6Hz, _J2 =8.0Hz, 1H), 7.46(d, J=8.4Hz, 1H), 7.51(t, _J1 =8.8Hz, J ₂ =7.2Hz, 1H), 7.89 (d, J=8.0Hz, 1H), 8.31 (d, J=8.8Hz, 2H).

Example 6

Synthesis of (E)-N-{[1-(2-aminoethyl)-benzo[c,d]indole]-2(1H)-ylidene}-4-carboxyaniline (derivative 6):

(1) Add 1.69g naphthalene lactam (0.01mol) and 1.63g p-aminobenzoic acid (0.012mol) into a 25mL two-neck flask, add 10mL chlorobenzene, add 1.1mLPOCl ₃ dropwise to the uniformly stirred reaction system, and magnetically stir reflux for 4 hours. After cooling to room temperature, add 2 mL of methanol dropwise to the reaction liquid, mix evenly and cool, then directly suction filter, wash with a large amount of methanol, and wash with water to obtain a brown solid crude product. After vacuum drying, it was recrystallized with acetic acid to obtain 1.7 g of a brown solid with a yield of 59%.

(2) Dissolve 0.288g of the intermediate (1mmol) obtained in step (1) into 10mL of acetonitrile, add 0.269g of t-BuOK (2.4mmol), heat up to reflux, and react for half an hour, then add 0.243g of bromoethylamine Bromate (1.2mmol), reflux for 5h. The cooled reaction solution was poured into 150 mL of ice water, extracted with dichloromethane, and rotary evaporated to obtain a brown crude product. Purified by column chromatography, 150mg of brown crystals were obtained, the yield was 45%, and the melting point was 176.8-178.1°C.

ESI-HRMS (m/z): Calculated: 331.1231, Found: 331.1274.

¹ HNMR (d6-DMSO, 400MHz): δ (ppm): 2.14 (t, NH ₂ , 2H), 2.84 (m, NCH ₂ CH ₂ N, 2H), 2.98 (t, NCH ₂ CH ₂ N, 2H) , 6.86(d, J=7.2Hz, 1H), 7.08(d, J=7.2Hz, 1H), 7.12(d, J=8.8Hz, 2H), 7.28(t, J ₁ =8.0Hz, J ₂ = 7.2Hz, 1H), 7.42(d, J=8.4Hz, 1H), 7.53(t, _J1 =7.2Hz, _J2 =8.4Hz, 1H), 7.89(d, J=8.0Hz, 1H), 8.31 (d, J = 8.8 Hz, 2H), 8.44 (s, OH, 1H).

Example 7

Synthesis of (E)-N-[(1-propargyl-benzo[c,d]indole)-2(1H)-ylidene]-4-carboxyaniline (derivative 7):

Dissolve 0.288g (1mmol) of the intermediate obtained in step (1) of Derivative 6 into 10mL of acetonitrile, add 0.269gt-BuOK (2.4mmol), heat up to reflux, and react for half an hour, drop 1mL of bromoacetylene, and react 3h. The cooled reaction liquid was poured into 150 mL of ice water, extracted with dichloromethane, and rotary evaporated to obtain a brown crude product. Purified by column chromatography to obtain 170 mg of brown crystals with a yield of 52%.

ESI-HRMS (m/z): Calculated: 326.1055, Found: 326.1089.

¹ HNMR (d6-DMSO, 400MHz): δ (ppm): 2.31 (s, 1H), 4.89 (s, NCH ₂ , 2H), 6.96 (d, J = 7.6Hz, 1H), 7.07 (d, J = 7.2Hz, 1H), 7.22(d, J=8.8Hz, 2H), 7.38(t, _J1 =8.0Hz, _J2 =7.6Hz, 1H), 7.46(d, J=8.4Hz, 1H), 7.51 (t, _J1 = 7.2Hz, J2 = 8.4Hz, 1H), 7.89 (d _, J = 8.0Hz, 1H), 8.31 (d, J = 8.4Hz, 2H), 8.5 (s, OH, 1H) .

Example 8

Synthesis of dimethyl 2-[(1-propargyl-benzo[c,d]indole)-2(1H)-ylidene]malonate (derivative 8):

(1) Add 1.69g of naphthalenolactam (0.01mol) and 1.32g of dimethyl malonate (0.01mol) into a 25mL two-neck flask, add 15mL of toluene, and heat to 120°C under magnetic stirring. Add 1.1mL POCl ₃ dropwise to the reaction system. After constant temperature reaction for 4 hours, add 2mL methanol to the reaction system. After cooling, directly suction filter, wash with a large amount of methanol, and wash with water to obtain a brown solid crude product. After vacuum drying, the crude product was separated by column chromatography to obtain 1.55 g of yellow crystals with a yield of 54%.

(2) 0.283 g (1 mmol) of the intermediate obtained in step (1) was dissolved in 10 mL of DMF, 0.067 g of NaOCH ₃ (1.2 mmol) was added, and the temperature was raised to 100° C. under magnetic stirring, then 1 mL of bromoacetylene was added dropwise and reacted for 2 h. The cooled reaction solution was poured into 150 mL of ice water, filtered with suction, and dried in vacuo to obtain a crude product. Purified by column chromatography to obtain 0.2 g of yellow crystals with a yield of 63% and a melting point of 257.6°C.

¹ HNMR (d6-DMSO, 400MHz): δ (ppm): 1.88 (s, 1H), 3.75 (s, NCH ₂ , 2H), 6.58 (d, J=7.8Hz, 1H), 7.15 (t, J ₁ =6.8Hz, J ₂ =6.3Hz, 1H), 7.17(d, J=6.3Hz, 1H), 7.52(d, J=7.2Hz, 1H), 7.45(t, J ₁ =7.2Hz, J ₂ = 7.6Hz, 1H), 7.63 (d, J = 7.6Hz, 1H), 4.76 (s, _OCH3 , 6H).

ESI-HRMS (m/z): Calculated: 321.1001, Found: 321.1045.

Example 9

2-{[2-(N,N'-dimethyl-ethylenediamine)-ethyl-benzo[c,d]indole]-2(1H)-ylidene}malononitrile (derivative 9) , 2-{[2-(N, N'-diethyl-ethylenediamine)-ethyl-benzo[c,d]indole]-2(1H)-ylidene}malononitrile (derivative 10 ), 2-[(2-butylamine-ethyl-benzo[c,d]indole)-2(1H)-ylidene]malononitrile (derivative 11), 2-[(2-piperazine- Synthesis of ethyl-benzo[c,d]indole)-2(1H)-ylidene]malononitrile (derivative 12):

Dissolve 0.324g of derivative 2 (1mmol) in 10mL of acetonitrile, add 2mmol of the corresponding amine and 2mL of triethylamine, reflux at 60°C, track the reaction to completion by TLC, generally takes 20-30h. After the reaction, the cooled reaction solution was poured into 150 mL of ice water, extracted with dichloromethane, and the organic phase was dried with anhydrous MgSO ₄ and then rotary evaporated to obtain a crude product. After vacuum drying, the crude product was purified by column chromatography to obtain the corresponding pure target derivative.

Derivative 9: ¹ HNMR (d6-DMSO, 400MHz): δ (ppm): 2.31 (s, NCH ₃ , 6H), 2.52 (t, NCH ₂ CH ₂ N, 2H), 2.70 (m, NCH ₂ CH ₂ N, 2H), 4.0(m, NH, 1H), 2.82(m, NCH2CH2N _, 2H), _3.22 (t, _{NCH2CH2N ,} 2H), 7.32(d, J=7.6Hz, 1H ), 7.10(t, J ₁ =7.6Hz, J ₂ =7.6Hz, 1H), 7.18(d, J=8.0Hz, 1H), 7.34(d, J=8.8Hz, 1H), 7.17(t, J ₁ = 7.6 Hz, J ₂ = 8.0 Hz, 1H), 7.75 (d, J = 8.4 Hz, 1H).

ESI-HRMS (m/z): Calculated: 331.1797, Found: 331.1763.

Derivative 10: ESI-HRMS (359.2145 m/z), Derivative 11: ESI-HRMS (316.1662 m/z), Derivative 12: ESI-HRMS (329.168 m/z).

Example 10

Synthesis of 2-[6-chloro-benzo[c,d]indole-2(1H)-ylidene]malononitrile (derivative 14):

(1) Dissolve 1.69g naphthalene lactam (0.01mol) completely in 25mL acetic acid, raise the temperature to 50°C, slowly add 0.8mLSO ₂ Cl ₂ (0.01mol) to the reaction solution dropwise, the temperature of the reaction system rises and suddenly turns yellow A large amount of solids were generated, the stirring was continued for 0.5 h, and the direct suction filtration yielded 1.90 g of dry bright yellow solids, with a yield of 94%.

(2) The experimental method is the same as the step (1) of derivative 1, and the yield is 78%.

Derivative 12: ESI-HRMS (251.027 m/z), melting point > 300°C.

Example 11

Synthesis of 2-[6-bromo-benzo[c,d]indole-2(1H)-ylidene]malononitrile (derivative 15):

(1) 1.69 g of naphthalene lactam (0.01 mol) was dissolved in 20 mL of 1,2-dichloroethane. Take 0.51mL of liquid bromine (0.01mol) and dissolve it in 10mL of 1,2-dichloroethane. The Br ₂ -ClCH ₂ CH ₂ Cl system was added dropwise to the reaction system, and the temperature was slowly raised to 50° C., and reacted for 3 hours. Cool, stand still, and directly suction filter to obtain 2.05 g of a yellow solid with a yield of 83% and a melting point of 254.9°C.

(2) The experimental method is the same as the step (1) of derivative 1, and the yield is 82%.

¹ HNMR (d6-DMSO, 400MHz): δ (ppm): 7.77 (d, J = 7.6Hz, 1H), 7.92 (t, J ₁ = 7.6Hz, J ₂ = 8.4Hz, 1H), 8.60 (d, J=8.0Hz, 1H), 8.29(d, J=8.4Hz, 1H), 7.03(d, J=8.0Hz, 1H), 9.88(s, NH, 1H).

ESI-HRMS (m/z): Calculated: 297.9745, Found: 297.9711.

Example 12

Synthesis of 2-[6-cyano-benzo[c,d]indole-2(1H)-ylidene]malononitrile (derivative 16):

(1) Dissolve 1.24 g (5 mmol) of the intermediate 6-bromo-1,8-naphtholactam obtained in the synthesis step (1) of derivative 15 in 15 mL of N-methylpyrrolidone (NMP), and add 0.627 g of cyanide under stirring Copper (7mmol) was reacted at a constant temperature of 130°C for 4h. After cooling, the reaction solution was poured into 200 mL of ice water, filtered with suction, washed with ammonia water to remove the light green color, and dried to obtain 0.91 g of a yellow solid with a yield of 94%.

(2) The experimental method is the same as step (1) of derivative 1, and the yield is 75%.

¹ HNMR (d6-DMSO, 400MHz): δ (ppm): 7.82 (d, J = 7.6Hz, 1H), 7.99 (t, J ₁ = 7.6Hz, J ₂ = 8.4Hz, 1H), 8.63 (d, J=7.6Hz, 1H), 8.34(d, J=8.4Hz, 1H), 7.12(d, J=7.6Hz, 1H), 9.95(s, NH, 1H).

ESI-HRMS (m/z): Calculated: 242.0592, Found: 242.0574.

Example 13

Synthesis of 2-[6-nitro-benzo[c,d]indole-2(1H)-ylidene]malononitrile (derivative 17):

(1) Dissolve 1.69g of naphthalenolactam (0.01mol) in 10mL of concentrated sulfuric acid, and control the temperature below 10°C in an ice-water bath. After stirring evenly, add 2.4mL of concentrated nitric acid dropwise to it, and the color of the solution becomes lighter. After 3 hours of reaction , the reaction solution was poured into ice water, a yellow solid was precipitated, filtered by suction, and dried in vacuum to obtain 19.0 g of a solid, with a yield of 89%.

(2) The experimental method is the same as the step (1) of derivative 1, and the yield is 78%.

ESI-HRMS (m/z): Calculated: 262.0491, Found: 262.0434.

Example 14

Synthesis of (E)-N-[6-nitro-benzo[c,d]indole-2(1H)-ylidene]-4-nitroaniline (derivative 19):

Taking the intermediate 6-nitro-1,8-naphtholactam obtained in the synthesis step (1) of derivative 17, the experimental operation was as in the synthesis step (1) of derivative 4, and the yield was 72%.

ESI-HRMS (m/z): Calculated: 334.0702, Found: 334.0775.

Example 15

Synthesis of (E)-N-[6-cyano-benzo[c,d]indole-2(1H)-ylidene]-4-nitroaniline (derivative 20):

Taking the intermediate 6-cyano-1,8-naphtholactam obtained in the synthesis step (1) of derivative 16, the experimental operation was as in the synthesis step (1) of derivative 4, and the yield was 68%.

ESI-HRMS (m/z): Calculated: 314.0804, Found: 314.0844.

¹ HNMR (d6-DMSO, 400MHz): δ(ppm): 4.31(s, 1H), 6.56(d, J=7.2Hz, 1H), 7.67(d, J=7.2Hz, 1H), 7.28(d, J=8.8Hz, 2H), 7.50(d, J=8.4Hz, 1H), 7.57(t, _J1 =7.6Hz, _J2 =8.4Hz, 1H), 7.85(d, J=8.0Hz, 1H) , 8.41 (d, J=8.8Hz, 2H).

Example 16

Determination of in vitro tumor cell growth inhibitory activity of derivatives:

7721 human liver cancer cells, MCF-7 human breast cancer cells, Hela human cervical cancer cells and BGC-823 human gastric cancer cells were tested in vitro by reduction method of tetrazolium salt (microculture tetrozolium, MTT).

The specific operation of tetrazolium salt (MTT) reduction method is: according to different tumor growth rates, a certain number of tumor cells in logarithmic growth phase 90 μl/well are inoculated in a 96-well micro-culture plate, and 10 μl of drug solution is added after culturing for 24 hours. /well, for each cell line, each concentration was three replicate wells. Set up a cell-free zero well. If the drug has a color, do a cell-free zero well for the corresponding drug concentration. After the tumor cells were cultured at 37°C and 5% CO for 48 hours, add MTT (Sigma) solution 5 mg/ml and prepare 20 μl/well with physiological _saline ; after continuing to culture for 4 hours, add triple solution (10% SDS-5% Isobutanol-0.01mol/l HCl) 50 μl/well, overnight in a CO ₂ incubator. Then measure the OD570 value with a microplate reader. Calculate the inhibitory rate of the test substance to the growth of cancer cells according to the following formula:

Tumor inhibition rate=(OD value of control group-OD value of treatment group)/OD value of control group×100%

It can be seen from the tabular data that derivatives 3, 4, 9, and 19 exhibit good tumor cell proliferation inhibitory activity, derivative 7 has selectivity for 7721 human liver cancer cells, and derivative 11 for MCF-7 human breast cancer cells Proliferation inhibitory effect.

Table 1. Growth inhibition rate of derivatives on 7721 human liver cancer cells

Table 2. Growth inhibition rate of derivatives to MCF-7 human breast cancer cells

Table 3. The growth inhibition rate of derivatives to Hela human cervical cancer

Table 4. Growth inhibition rate of derivatives on BGC-823 human gastric cancer cells

Claims (2)

1. A class of naphthalene lactam derivatives, characterized in that: the derivatives have a general chemical structure of:

in: R ₁ is =C(CN) ₂ , =C(COOCH ₃ ) ₂ ,

or

R ₂ is -H, -Cl, -Br, -CN or -NO ₂ ; R ₃ is -CH ₂ CH ₂ C≡CH, -CH ₂ CH ₂ Br, -CH ₂ CH ₂ NH ₂ , -CH ₂ CH 2 NHCH _{2 CH 2 N} (CH ₃ ) ₂ , -CH ₂ CH ₂ NHCH ₂ CH ₂ N(CH ₂ CH ₃ ) ₂ , -CH ₂ CH ₂ NCH ₂ CH ₂ CH ₂ CH ₃ or

2. The application of a class of naphthalene lactam derivatives according to claim 1 in the inhibition of tumor cell proliferation, characterized in that: said naphthalene lactam derivatives are formulated into medicinal solutions with different concentrations, and tetrazole Salt reduction method was used to test 7721 human liver cancer cells, MCF-7 human breast cancer cells, Hela human cervical cancer cells or BGC-823 human gastric cancer cells. Inoculate 90 μl/well in a 96-well micro-culture plate, add 10 μl/well of the prepared drug solution after 24 hours of culture, for each cell line, each concentration is three replicate wells; if the prepared drug solution has color The corresponding concentration of the drug solution should be adjusted to zero wells without cells; after the tumor cells were cultured at 37°C and 5% CO ₂ for 48 hours, 20 μl/well of 5 mg/ml tetrazolium salt solution prepared with normal saline was added; continue to culture for 4 After 1 hour, add 50 μl/well of the triple solution prepared by 10% sodium dodecylsulfonate-5% isobutanol-0.01mol/1 HCl, and put it in a _CO2 incubator overnight; then use a microplate reader to measure the OD570 value ; Calculate the inhibitory rate of described naphthalene lactam derivatives to the growth of cancer cells according to the following formula: Tumor inhibition rate=(OD value of control group-OD value of treatment group)/OD value of control group×100%.