CN106749089A - The preparation of new fluoro thiazole hydrazone compounds and its application in antineoplastic - Google Patents

Abstract

The present invention relates to the preparation of a class of fluorothiazole hydrazone compounds I-IV and the combination of such compounds, as well as their application in anti-tumor and/or anti-cancer drugs. The present invention also relates to a preparation method of such compounds. The general chemical structure of the compound is as follows: R ₁ , R ₂ and R ₃ in the general structural formula are respectively one of H, Br, F and CF ₃ , R ₁ in compound I is Br, R ₂ and R ₃ are H; in compound II, R ₁ is H, R ₂ and R ₃ are F; R ₂ is CF ₃ in compound III, R ₁ and R ₃ are H; R ₂ is H in compound IV, R ₁ and R ₃ are CF ₃ ; the present invention also relates to containing at least A compound or a salt thereof with a physiologically acceptable inorganic or organic acid, if appropriate, a pharmaceutically acceptable excipient and/or diluent or excipient; the invention also relates to a compound containing at least one of the formula The dosage form of the above-mentioned compound of the compound or its salt is tablet, capsule, solution for infusion, suppository, patch, powder, suspension and the like.

Description

Preparation of novel fluorothiazole hydrazone compounds and their application in antitumor drugs

technical field

The invention relates to the technical field of medicines, in particular to a class of fluorothiazole hydrazone compounds I-IV and their pharmacological activity and pharmaceutical application. The compound and its derivatives can be used to prevent and/or treat diseases such as tumors or cancers caused by various factors.

Background technique

Malignant tumors seriously affect people's health and have become the second leading cause of human death, second only to cardiovascular and cerebrovascular diseases. According to the statistics of the World Health Organization, more than 10 million cancer patients are newly diagnosed every year in the world, and the total number of cancer deaths worldwide reaches 7 million people every year. With the aggravation of environmental pollution, the incidence of cancer is increasing year by year, showing a trend of multiple occurrences. The WHO 2014 report predicts that global cancer cases will show a rapid growth trend, from 14 million in 2012 to 19 million in 2025. , will reach 24 million by 2035, and the death toll will also increase from 6 million to 10 million per year. In 2012, China had 3.07 million new cancer patients and caused about 2.2 million deaths, accounting for 21.9% and 26.8% of the global total respectively. Cancer has become the leading cause of death among urban and rural residents in my country. Tumors not only seriously threaten the health of the people, bring economic losses to patients and their families, but also cause heavy burdens to the country and society. Finding new anti-tumor drugs with high efficiency, low toxicity, and new mechanisms of action that can selectively kill or inhibit tumor cells has become an important direction for the development of anti-tumor drugs.

Thiazole compounds are a very important class of heterocyclic compounds, with antibacterial, anti-tuberculosis, anti-cancer, anti-virus, lowering blood pressure, anti-inflammatory and analgesic, lowering blood sugar, anti-epileptic, anti-parasitic and anti-oxidation fields. There are many thiazole drugs used in the clinical treatment of various diseases. Hydrazone compounds refer to a class of compounds containing -NHN=CH-groups in their molecular structures, which are the products of condensation reactions between hydrazine and carbonyl compounds such as aldehydes or ketones, which have a wide range of biological activities (such as antibacterial, antitumor, etc.) And hypoglycemic, etc.), it has a good application prospect in the field of medicine. The present invention couples active groups of thiazole and hydrazone compounds to design and synthesize fluorothiazole hydrazone compounds. In the prior art, there is no report on the fluorothiazole hydrazone compounds provided by the present invention and the pharmaceutical composition thereof as active ingredients. There is no report on the application of such derivatives or their pharmaceutical compositions in the preparation or treatment of drugs for diseases such as tumors caused by various factors.

Contents of the invention

The invention relates to the technical field of medicines, in particular to a class of fluorothiazole hydrazone compounds I-IV, a preparation method thereof, pharmacological activity and pharmaceutical use. The compounds and their derivatives can be used to prevent and/or treat diseases such as tumors and cancers caused by various factors.

To achieve the above object, the technical scheme adopted in the present invention is as follows:

The present invention designs and synthesizes a class of fluorothiazole hydrazone compounds I-IV, and its chemical structure is as follows:

R ₁ , R ₂ and R ₃ in the general structural formula are respectively one of H, Br, F and CF ₃ , R ₁ in compound I is Br, R ₂ and R ₃ are H; in compound II, R ₁ is H, R ₂ and R ₃ are F; in compound III, R ₂ is CF ₃ , R ₁ and R ₃ are H; in compound IV, R ₂ is H, and R ₁ and R ₃ are CF ₃ .

One or two or more mixtures of the compounds I-IV have antitumor activity, and the compound I-IV referred to here can be the pharmaceutically acceptable compound I-IV itself and the compound I-IV One of chemical equivalents such as salt, but not limited to the above chemical equivalents.

The present invention also provides the application of compounds I-IV of the present invention in the preparation for preventing and/or treating tumors and/or cancers and other related diseases caused by various factors.

When the compound of the present invention is used as a medicine, it can be used directly or in the form of a pharmaceutical composition. The pharmaceutical composition contains 0.1-99%, preferably 0.5-90%, of the compound of the present invention, and the rest are pharmaceutically acceptable carriers and/or excipients.

The pharmaceutical carrier or excipient is one or more solid, semi-solid and liquid diluents, fillers and pharmaceutical preparation adjuvants. The pharmaceutical composition of the present invention is used in a dose per body weight. The medicine of the present invention can be administered in three forms of injection (intravenous injection, intramuscular injection), oral administration and external application.

The purpose of the present invention is to provide a new class of fluorothiazole hydrazone compounds and their preparation methods, which can prevent and/or treat diseases such as tumors and/or cancers caused by various factors, such as lung cancer and myeloma , neuroblastoma, liver cancer, gastric cancer, breast cancer, pancreatic cancer, bladder cancer, bowel cancer, prostate cancer and other tumors and/or cancer diseases, or one or more of them.

1. Compound synthesis and structure identification

The preparation method steps of fluorothiazole hydrazone compound I-IV of the present invention are as follows:

Step 1: Preparation of intermediate 3,4-difluorobenzylidene thiosemicarbazide (compound a):

Weigh 10mmol 3,4-difluorobenzaldehyde (3.07g) and 11mmol thiosemicarbazide (2.165g) and place in a 500mL reaction flask, then add 30mL ethanol (95%) and stir evenly, then add 1mL glacial acetic acid dropwise, and then 65 Reflux and stir at -70°C for about 10 hours, evaporate most of the ethanol under reduced pressure, add 20 ml of ice water, filter to obtain a precipitate, wash the precipitate with ice water (30 mL for three times), and recrystallize from ethanol to obtain the intermediate 3,4-difluorobenzylidene thiosemicarbazide (compound a).

Step 2: Preparation of fluorothiazole hydrazone compounds I-IV:

Weigh 1mmol of the intermediate 3,4-difluorobenzylidene thiosemicarbazide (215mg) and 2-5mmol of anhydrous sodium acetate into a reaction flask, add 10-20mL of absolute ethanol, and then add 0.9-1.1mmol of the corresponding Substitute 2-bromoacetophenone, then heat and reflux at 60-90°C for 10-14 hours, cool the reaction solution, filter to obtain a solid, and then recrystallize from methanol to obtain thiazole hydrazone compound I-IV.

2. Determination of the toxicity of fluorothiazole hydrazone compounds I-IV of the present invention to various tumor cells

The cytotoxicity of synthetic derivatives to human tumor cells cultured in vitro was detected by the commonly used tetrazolium salt (MTT) method. Cell lines selected for in vitro anti-tumor experiments: human lung cancer cell A549, human cloned colon adenocarcinoma cell CaCo-2, human liver cancer cell HepG2, human glioma cell U87, human umbilical vein endothelial cell HUVEC, etc. Determination method: Take the cells in the logarithmic growth phase, inoculate the cell suspension into a 96-well plate, pre-culture for 24 hours in an incubator at 37°C, 100% relative humidity, 5% CO ₂ , and 95% air, and then add drugs . In addition, a negative control (equal concentration of DMSO) and a blank background (without adding cells) were set for each concentration, and three replicate wells were set for each group. Continue culturing for another 24 hours, then add MTT solution to each well, continue culturing for 4 hours, and carefully suck off the supernatant (suspended cells need to be centrifuged first, and then suck off the supernatant). Add DMSO to each well, set a micro-oscillator to vibrate for 5 minutes to completely dissolve the crystals, and measure the OD value by using a microplate reader at 492 nm single-wavelength colorimetry. Inhibition rate (%)=(mean OD value of experimental group−mean OD value of blank group)/(mean OD value of control group−mean OD value of blank group)×100%, and calculate IC ₅₀ value.

The present invention has the following advantages:

The compound provided by the present invention has strong inhibitory activity on various tumor cell lines tested by MTT method, and has low toxicity to normal cell human umbilical vein endothelial cells HUVEC, which proves that the compound of the present invention has good anti-tumor activity , has a good application prospect in the development of antitumor drugs.

detailed description

In order to better understand the essence of the present invention, the following examples of the present invention will be used to illustrate the preparation methods and pharmacological effects of the compounds of the present invention, fluorothiazole hydrazone compounds I-IV, but the present invention is not limited by these examples .

Example 1: 4-(3-bromophenyl)-2-(2-(3,4-difluorobenzylidene)hydrazino)thiazole (4-(3-bromophenyl)-2-(2-( 3,4-difluorobenzylidene) hydrazinyl) thiazole, compound I) preparation

(1) Weigh 10mmol 3,4-difluorobenzaldehyde (3.07g) and 11mmol thiosemicarbazide (2.165g) into a 500mL reaction flask, add 30mL ethanol (95%) and stir evenly, then add 1mL glacial acetic acid dropwise , then reflux and stir at 65-70°C for about 10 hours, evaporate most of the ethanol under reduced pressure, add 20 ml of ice water, filter to obtain a precipitate, wash the precipitate with ice water (30 mL for three times), and recrystallize from ethanol to obtain the intermediate 3,4-difluoro Benzylidenethiosemicarbazide (compound a).

(2) Weigh 1mmol of intermediate 3,4-difluorobenzylidene thiosemicarbazide (215mg) and 4mmol of anhydrous sodium acetate into a 250mL reaction bottle, add 20mL of absolute ethanol, and then add 1mmol of 2,3' -Dibromoacetophenone (278mg), then heated to reflux at 60-90°C for 12 hours, cooled the reaction solution, filtered and dried to obtain gray solid 4-(3-bromophenyl)-2-(2-(3,4- Difluorobenzylidene)hydrazino)thiazole (77%); ¹ H-NMR (500MHz, DMSO- _d6 )δ:8.03(1H,s),7.99(1H,s),7.84(1H,d,J =7.5Hz), 7.66(1H,m),7.47(4H,overlap),7.35(1H,dd,J=8.0,7.5Hz); ¹³ C-NMR(125MHz,DMSO- _d6 )δ:168.1,150.8, 148.81, 148.83, 139.0, 136.8, 132.2, 130.7, 130.1, 128.1, 124.3, 123.2, 122.1, 118.0, 114.5, 105.5; ESIMS: m/z393[M ⁺ H] ⁺ HRESIMS: calc forC16H10N3F2SBr]9303H.9303H. , found 393.9777.

Example 2: 2-(2-(3,4-difluorobenzylidene)hydrazino)-4-(3,4-difluorophenyl)thiazole (2-(2-(3,4-difluorobenzylidene )hydrazinyl)-4-(3,4-difluorophenyl)thiazole, compound II)

The preparation method of Compound II is similar to that of Compound I, except that the raw material 2,3'-dibromoacetophenone is replaced by 3',4'-difluoro-2-bromoacetophenone from Example 1 Ketone, compound II was prepared, gray solid, yield 82%, ¹ H-NMR (500MHz, DMSO- _d6 ) δ: 12.26 (1H, s), 7.95 (1H, s), 7.78 (1H, m), 7.58 -7.65 (2H, overlap), 7.38-7.45 (4H, overlap); ¹³ C-NMR (125MHz, DMSO- _d6 ) δ: 168.6, 151.1, 150.2, 149.2, 148.8, 148.3, 139.5, 138.7, 132.7, 132.6, 123.7, 122.6, 118.4, 118.1, 114.8, 114.6, 105.4; ESIMS: m/z352[M+H] ⁺ HRESIMS: calc for C16H9N3F4S[M+H] ⁺ 352.0526, found 352.0452.

Example 3: 2-(2-(3,4-difluorobenzylidene)hydrazino)-4-(4-(trifluoromethyl)phenyl)thiazole (2-(2-(3,4 -difluorobenzylidene)hydrazinyl)-4-(4-(trifluoromethyl)phenyl)thiazole, compound III)

The preparation method of compound III is similar to the preparation method of compound I, and its difference from Example 1 is that the raw material 2,3'-dibromoacetophenone is replaced by 4'-trifluoromethyl-2-bromoacetophenone , to prepare compound III, gray solid, yield 78%, ¹ H-NMR (500MHz, DMSO- _d6 )δ: 12.31 (1H, s), 8.01 (2H, d, J=8.0Hz), 7.96 (1H, s), 7.69 (2H, d, J = 8.0Hz), 7.61 (1H, dd, J = 9.5, 9.5Hz), 7.50 (1H, s), 7.38-7.45 (2H, overlap); ¹³ C-NMR ( 125MHz, DMSO- _d6 )δ: 168.8, 151.2, 149.5, 149.3, 139.5, 138.7, 132.6, 128.0, 127.9, 126.4, 125.9, 123.7, 118.3, 114.9, 107.0; ESIMS: m/z 384[M+H] ⁺ HRESIMS: calc forC17H10N3F5S[M+H] ⁺ 384.0588, found 384.0535.

Example 4: 2-(2-(3,4-difluorobenzylidene)hydrazino)-4-(3,5-bis(trifluoromethyl)phenyl)thiazole (4-(3,5 -bis(trifluoromethyl)phenyl)-2-(2-(3,4-difluorobenzylidene)hydrazinyl)thiazole, compound IV)

The preparation method of compound IV is similar to the preparation method of compound I, except that the raw material 2,3'-dibromoacetophenone is replaced by 3',5'-bis(trifluoromethyl)- 2-Bromoacetophenone, compound IV was prepared, gray solid, yield 90%, ¹ H-NMR (500MHz, DMSO- _d6 ) δ: 10.92(1H,s), 8.16(2H,s), 7.76(1H ¹³ C-NMR (125MHz, DMSO- _d6 ) δ: 169.5, 151.8, 149.8, 148.2, 139.8, 136.7, 131.8(2C), 131.6, 125.7(2C), 123.3(2C), 123.2, 120.8, 117.4, 114.5, 106.4; ESIMS: m/z 452[M+H] ⁺ HRESIMS: calc forC18H9N3F8S[M+H] ⁺ 452.0462,found452.0446.

Example 5: Determination of the toxicity of fluorothiazole hydrazone compounds I-IV to various tumor cells

The cytotoxicity of synthetic compounds on human tumor cells cultured in vitro was detected by the commonly used tetrazolium salt (MTT) method. Cell lines selected for in vitro anti-tumor experiments: human lung cancer cell A549, human cloned colon adenocarcinoma cell CaCo-2, human liver cancer cell HepG2, human glioma cell U87, human umbilical vein endothelial cell HUVEC, etc. Determination method: take the cells in the logarithmic growth phase, inoculate the cell suspension into a 96-well plate, so that the number of cells per well is 3×10 ³ , at 37°C, 100% relative humidity, containing 5% CO2 by volume, After 24 hours of pre-cultivation in an incubator with 95% air, the drugs were added. In addition, the concentrations of 1.25, 2.5, 5.0, 10.0, 20.0 μg/mL of fluorothiazole hydrazone compounds I-IV, each concentration set a negative control (isoconcentration DMSO) and blank background (no cells), each Each group had 3 replicate wells. Continuously culture for another 24 hours, then add 20 microliters of 5 mg/ml MTT solution to each well, continue to cultivate for 4 hours, and carefully suck off the supernatant (suspended cells need to be centrifuged first, and then suck off the supernatant). Add 150 microliters of DMSO to each well, set the micro-oscillator to vibrate for 5 minutes to completely dissolve the crystals, and measure the OD value by using a microplate reader at 492 nm single-wavelength colorimetry. Inhibition rate (%)=(mean OD value of experimental group-mean OD value of blank group)/(mean OD value of control group-mean OD value of blank group)x100%, and SPSS17.0 software was used to calculate _IC50 value (Table 1).

Table 1: Data table of compounds I-IV's inhibitory activity on tumor cells

The experimental results show that the compounds provided by the present invention all show good anti-tumor activity, have strong inhibitory activity in vitro to various tumor cell lines tested, and have low toxicity to normal human umbilical vein endothelial cells HUVEC, The compounds of the present invention can be used to prevent and/or treat tumor or cancer-related diseases and symptoms caused by various factors.

Embodiment 6: the preparation of compound injection

After dissolving fluorothiazole hydrazone compounds I-IV with a small amount of DMSO (weight ratio: 1:0.1-1:0.5, here is 1:0.4), add water for injection as usual (weight ratio: 1:20 -1:200, here is 1:200), finely filtered, potted and sterilized to make injection.

Embodiment 7: Preparation of compound powder injection

Dissolve fluorothiazole hydrazone compounds I-IV with a small amount of DMSO (weight ratio: 1:0.1-1:0.5, here is 1:0.5), and dissolve them in sterile water for injection (weight ratio: : 1:20-1:60, here is 1:60), stir to dissolve, filter with a sterile suction filter funnel, then aseptic fine filter, pack in ampoules, freeze-dry at low temperature and then aseptically melt seal Get powder injection.

Embodiment 8: Preparation of compound powder

The fluorothiazole hydrazone compounds I-IV were added to the excipient at a weight ratio of 9:1 to the excipient (Tween 80: propylene glycol: cyclodextrin: lactose=1:2:4:12) , made into powder.

Embodiment 9: Preparation of compound tablet

The fluorothiazole hydrazone compounds I-IV are respectively mixed with excipients (hydroxypropylmethylcellulose E5: microcrystalline cellulose MCC102: magnesium stearate: (8% povidone K30) = 15:15: 2:0.1) The weight ratio is 5:1 by adding excipients, granulating and compressing into tablets.

Embodiment 10: Preparation of compound oral liquid

Fluorothiazole hydrazone compounds I-IV were added to distilled water containing 20% simple syrup and 0.1% sodium benzoate, respectively, and the concentration of 15 μg/mL oral liquid was prepared according to the conventional oral liquid preparation method.

Embodiment 11: the preparation of compound capsule

Fluorothiazole hydrazone compounds I-IV are mixed with excipients (medicinal starch: glucose: hydrolyzed gelatin: glycine = 30:10:1:1) in a weight ratio of 5:1 to make capsules .

Embodiment 12: the preparation of compound capsule

Fluorothiazole hydrazone compounds I-IV are mixed respectively with excipients (medicinal starch: glucose: hydrolyzed gelatin: glycine = 30:10:1:1) in a ratio of 3:1 by weight to make capsules .

Claims (7)

1. A class of fluorothiazole hydrazone compounds, characterized in that they have the following general structural formula: R ₁ , R ₂ and R ₃ in the general structural formula are respectively one of H, Br, F and CF ₃ . 2. according to the described fluorothiazole hydrazone compound of claim 1, it is characterized in that: The fluorothiazole hydrazone compound is one of the fluorothiazole hydrazone compounds I-IV, In compound I, R ₁ is Br, R ₂ and R ₃ are H; In compound II, R ₁ is H, R ₂ and R ₃ are F; In compound III, R ₂ is CF ₃ , R ₁ and R ₃ are H; In compound IV, R ₂ is H, R ₁ and R ₃ are CF ₃ . 3. the application of a fluorothiazole hydrazone compound described in claim 1 or 2, characterized in that: One or more of the fluorothiazole hydrazone compounds, pharmaceutically acceptable salts of fluorothiazole hydrazone compounds, and chemical equivalents of fluorothiazole hydrazone compounds have antitumor activity, and the fluorothiazole hydrazone compounds have antitumor activity. One or two or more of thiazole hydrazone compounds, pharmaceutically acceptable salts of fluorothiazole hydrazone compounds and chemical equivalents of fluorothiazole hydrazone compounds are used as active ingredients in the preparation of prevention and/or treatment of tumors, The application of one or two or more drugs in cancer and other related diseases and symptoms. 4. according to the described application of claim 3, it is characterized in that: One or two or more of the fluorothiazole hydrazone compounds, pharmaceutically acceptable salts of fluorothiazole hydrazone compounds and chemical equivalents of fluorothiazole hydrazone compounds can be mixed with pharmaceutically acceptable drugs The mixture of carriers can be made into tablets, capsules, oral liquids, granules, pills or injections for treating tumors, cancer-related diseases and symptoms, but not limited to the above dosage forms. 5. According to the application according to claim 3 or 4, it is characterized in that: said prevention and/or treatment of one or two or more of related diseases and symptoms such as tumors and cancers, especially those related to lung cancer, liver cancer and gastric cancer One or two or more of tumors and/or cancer diseases such as breast cancer, pancreatic cancer, colon cancer, and prostate cancer. 6. One or two or more drugs for preventing and/or treating tumors, cancers and other related diseases and symptoms, characterized in that: the pharmacological properties of the fluorothiazole hydrazone compounds and fluorothiazole hydrazone compounds One or two or more of acceptable salts and chemical equivalents of fluorothiazole hydrazone compounds can be used directly or in the form of a pharmaceutical composition; the pharmaceutical composition contains 0.1-99%, preferably 0.5-90 % of one or two or more of the fluorothiazole hydrazone compounds, pharmaceutically acceptable salts of fluorothiazole hydrazone compounds, and chemical equivalents of fluorothiazole hydrazone compounds, and the rest are pharmaceutically acceptable acceptable pharmaceutical carriers and/or excipients. 7. a preparation method of the bromophenol thiosemicarbazide compound described in claim 1 or 2, is characterized in that: Step 1: Preparation of intermediate 3,4-difluorobenzylidene thiosemicarbazide (compound a): Weigh 10mmol 3,4-difluorobenzaldehyde (3.07g) and 10-15mmol thiosemicarbazide into a reaction flask, then add 20-30mL ethanol (volume concentration 90-95%) and stir evenly, then add 0.5-1.5 mL of glacial acetic acid, then reflux and stir at 65-70°C for about 8-12 hours, evaporate most of the ethanol under reduced pressure, add 15-30 ml of water, filter, and wash the precipitate with 20-40 ml of water (2-4 times). Crystallization to obtain intermediate 3,4-difluorobenzylidene thiosemicarbazide (compound a); Step 2: Preparation of fluorothiazole hydrazone compounds I-IV: Weigh 1mmol of the intermediate 3,4-difluorobenzylidene thiosemicarbazide (215mg) and 2-5mmol of anhydrous sodium acetate into a reaction flask, add 10-20mL of absolute ethanol, and then add 0.9-1.1mmol of the corresponding Substitute 2-bromoacetophenone, then heat and reflux at 60-90°C for 10-14 hours, cool the reaction solution, filter to obtain a solid, and then recrystallize from methanol to obtain thiazole hydrazone compound I-IV.