Fluorinated camptothecin drug derivatives, methods of making and using the same

Abstract

The present application relates to the technical field of pharmaceutical chemistry, and particularly relates to a fluorinated camptothecin drug derivative, a preparation method and application thereof. The present application provides a preparation method of a fluorinated camptothecin drug derivative, comprising the following steps: dispersing a camptothecin drug derivative as shown in formula (II) and a fluorination reagent in a solvent, carrying out a reaction at room temperature, and separating and purifying a reaction mixture obtained after the reaction, so as to obtain the fluorinated camptothecin drug derivative as shown in formula (I); the fluorinated camptothecin drug derivative in the present application is prepared by directly fluorinating the camptothecin drug derivative, the operation process is simple, the reaction condition is mild, the site selectivity is high, the reaction is efficient, and the fluorinated camptothecin drug derivative can be prepared by only one step reaction. In addition, the fluorinated camptothecin drug derivative in the present application has good antitumor activity, and provides a new scheme for antitumor drugs.

Description

TECHNICAL FIELD

[0001] The present application relates to the technical field of pharmaceutical chemistry, in particular to fluorinated camptothecin drug derivatives, and a preparation method and application thereof. BACKGROUND

[0002] Phenol is an important basic organic raw material, and many of its downstream products are related to many fields, mainly used for the production of phenolic resin, bisphenol A and caprolactam. The derivatives of phenol, such as halogenated phenol, nitrophenol and alkylphenol, can be used in the production of medicines, pesticides, paints, dyes, explosives and spices. Phenolic compounds exist widely in nature, and phenol structures are contained in spices and many natural products. The traditional method for modifying phenolic compounds is to first protect the hydroxyl group, then modify other positions of the hydroxyl group, and finally deprotect to achieve modification. This kind of method has harsh conditions, and the method has many steps and low reaction yield. The existence of these problems greatly reduces the progress of the modification research of phenolic compounds.

[0003] Camptothecin (CPT) and its derivatives are a class of natural alkaloids isolated from the bark of Camptotheca acuminata, which have anti-tumor activity. Some camptothecin drug derivatives have phenol functional groups (such as hydroxyl groups) in their structures, which have the characteristics of phenol and are classified as phenolic compounds. The modification of camptothecin drug derivatives is usually aimed at improving their anti-tumor activity and improving their pharmacokinetic properties (such as increasing water solubility and reducing drug resistance). Affected by the difficulty of modifying phenolic compounds, the modification process of camptothecin drug derivatives is also relatively complex. The problems and difficulties in the preparation of fluorinated camptothecin drug derivatives include: (1) complexity of the synthesis route: fluorination usually requires specific catalysts and conditions, which may not be compatible with other synthesis steps of camptothecin drug derivatives, resulting in a complex and difficult to control synthesis route; (2) selectivity control: in the fluorination process, high selectivity is often required to ensure that the fluorine atom is introduced only to the specific position, which may require fine condition control and post-processing steps to avoid the occurrence of side reactions; (3) strict reaction conditions: fluorination may require specific reaction conditions such as high temperature, high pressure or the use of toxic chemicals, which not only require high equipment requirements, but also pose a challenge to the safety of the operators; (4) control of side reactions and impurities: fluorination reactions may be accompanied by various side reactions, which may affect the activity and safety of the drug. Therefore, strict impurity control and purification steps are required; (5) catalyst design and selection: fluorination usually requires catalysts, and the selection and design of catalysts are crucial for improving reaction efficiency and selectivity; (6) biological activity evaluation: the biological activity of fluorinated camptothecin drug derivatives may be different from that of the original compound, so detailed biological activity evaluation is needed to determine whether fluorination improves the efficacy of the drug. SUMMARY

[0004] The present application aims at overcoming the deficiencies in the prior art, providing a fluorinated camptothecin drug derivative with a fluorination site at position 9 and a preparation method thereof, and applying it to the preparation of an antitumor drug, so as to overcome the problems in the prior art, such as harsh reaction conditions, multiple steps, and low reaction yield, in the preparation of fluorinated camptothecin drug derivatives by fluorination of camptothecin drug derivatives.

[0005] In order to achieve the above-mentioned purpose, the present application provides the following technical solutions.

[0006] In a first aspect, the present application provides a preparation method of a fluorinated camptothecin drug derivative, comprising the following steps:

[0007] dispersing a camptothecin drug derivative as shown in formula (II) and a fluorination reagent in a solvent, reacting at room temperature, separating and purifying the reaction mixture obtained after the reaction, so as to obtain the fluorinated camptothecin drug derivative as shown in formula (I); (II)

[0008] The present application directly fluorinates the camptothecin drug derivative by using a fluorination reagent to obtain a fluorinated camptothecin drug derivative with a fluorination site at position 9, and the process does not need to add a catalyst. Compared with general fluorination methods, this method has the advantages of not needing to add a catalyst, reducing reaction cost, toxicity, and operation difficulty. The reaction condition is mild, the site selectivity is high, the reaction is efficient, and the compound can be prepared by only one step. It is found by detection that the compound has better antitumor activity than camptothecin, and provides a new scheme for antitumor drugs.

[0009] The above-mentioned reaction process can be represented by the following reaction formula:

[0010]

[0011] As a preferred, the mass ratio of the amount of the camptothecin drug derivative and the amount of the fluorination reagent added is 1:1-3.

[0012] The excess of fluorinating agent helps to improve the selectivity of the reaction and reduce the generation of by-products. However, the excess of fluorinating agent can cause excessive fluorination, introduce unstable structures, and affect the stability and activity of the product. It is found through experiments that controlling the molar ratio of camptothecin drug derivative to fluorinating agent to be 1:1-3 can make the reaction efficiency and selectivity higher, and a relatively pure product can be obtained. If the molar ratio of camptothecin drug derivative to fluorinating agent is higher or lower than the above-mentioned range of 1:1 to 1:3, the following problems may occur, (1) too high ratio (less camptothecin drug derivative): insufficient amount of reactants, which can lead to incomplete reaction and reduced yield; excessive fluorinating agent can cause excessive fluorination of the product, introduce unwanted fluorine atoms, and affect the activity and stability of the product; the selectivity of the reaction can be reduced, and more side reactions can occur; the excess of fluorinating agent can increase the difficulty and cost of subsequent purification; (2) too low ratio (less fluorinating agent): too much reactant, which can reduce the reaction efficiency and slow down the reaction speed; excessive camptothecin drug derivative can increase the risk of side reactions and affect the purity and quality of the product; the fluorination of camptothecin drug derivative may not be fully achieved, leading to a product structure that does not meet expectations; the selectivity of the reaction can be reduced, and more impurities can be generated.

[0013] As a preferred, the molar ratio of the camptothecin drug derivative to the amount of fluorinating agent added is 1:1.5.

[0014] As a preferred, the fluorinating agent is one or more of N-fluorobenzenesulfonimide, 1-chloromethyl-4-fluoro-1,4-diazonium bicyclo2.2.2octane bis(tetrafluoroborate) salt, and 1-fluoropyridine tetrafluoroborate.

[0015] The selection of fluorinating reagents needs to consider the reaction conditions, the degree of fluorination of the product, the generation of by-products, the stability of the catalyst, and the cost, etc. The advantages of the selected N-fluorobenzenesulfonimide, 1-chloromethyl-4-fluoro-1,4-diazonium bicyclo[2.2.2]octane bis(tetrafluoroborate) salt, and 1-fluoropyridine tetrafluoroborate as fluorinating reagents are as follows: N-fluorobenzenesulfonimide: as a broad-spectrum fluorinating reagent, it can effectively introduce fluorine atoms. The reaction is usually carried out under mild conditions, and has less impact on the environment. The degree of fluorination can be controlled to obtain products with different fluorine contents. The reaction conditions are relatively simple and easy to operate; 1-chloromethyl-4-fluoro-1,4-diazonium bicyclo[2.2.2]octane bis(tetrafluoroborate) salt (also known as "diazonium compound"): the diazonium compound is usually used as a nitrogen carrier in fluorination reactions, and can effectively transfer fluorine atoms. It can react under a wide range of temperatures and reaction conditions. It usually provides high fluorination yield and good selectivity. The fluorination of some compounds that are difficult to fluorinate can be achieved; 1-fluoropyridine tetrafluoroborate: 1-fluoropyridine tetrafluoroborate is a relatively new fluorinating reagent, which has good selectivity and efficiency. It can carry out fluorination reaction under relatively mild conditions, and is suitable for the modification of sensitive compounds. Fluoropyridine compounds usually have good solubility and post-treatment properties.

[0016] As preferred, the solvent is one or more of acetonitrile, water, aqueous acetic acid, dichloromethane, dimethyl sulfoxide, 1,2-dichloroethane.

[0017] In selecting a solvent, the compatibility of the solvent with the reactants and products, the boiling point of the solvent, the temperature range of the reaction, the environmental impact of the solvent, and the cost are all factors to consider. In the synthesis of fluorinated camptothecin drug derivatives, selecting the appropriate solvent is crucial for the smooth progress of the reaction. Acetonitrile, water, aqueous acetic acid, dichloromethane, dimethyl sulfoxide, and 1,2-dichloroethane are used as solvents, each with some advantages: acetonitrile: acetonitrile is a polar organic solvent that can dissolve many organic compounds, including camptothecin drug derivatives and fluorination reagents. It can promote collisions between reactants, increasing reaction rates. Acetonitrile has a high boiling point, allowing it to be used over a wide temperature range, which helps control the reaction temperature; water: as the most common solvent, water is inexpensive, readily available, and environmentally friendly. It can form hydrogen bonds with fluorination reagents, increasing their solubility. Reactions carried out in water are generally safer because they usually do not require high temperatures or dangerous chemicals; aqueous acetic acid: aqueous acetic acid can provide an acidic environment, helping to control the pH of the reaction, which is necessary for some fluorination reactions. The acidity of acetic acid can promote the activity of some fluorination reagents, increasing reaction efficiency. Aqueous acetic acid has a relatively small environmental impact; dichloromethane: dichloromethane is a non-polar solvent that can dissolve many organic compounds. It is often used for reactants or products that are not easily dissolved in polar solvents; dimethyl sulfoxide (DMSO): DMSO is a polar organic solvent that can dissolve many organic compounds, including some that are difficult to dissolve in water. It can promote the polarization of electron clouds, helping to increase reaction rates; 1,2-dichloroethane: 1,2-dichloroethane is a non-polar solvent that can dissolve many organic compounds. It is often used for reactants or products that are not easily dissolved in polar solvents.

[0018] As a preferred, the fluorination reagent is 1-chloromethyl-4-fluoro-1,4-diazonium bicyclo 2.2.2 octane bis (tetrafluoroboric acid) salt, and the solvent is 8-12% aqueous acetic acid solution.

[0019] As a preferred, the separation and purification method comprises the following steps: adding saturated NaCl aqueous solution to the reaction mixture, extracting with dichloromethane, taking the organic layer, drying, filtering, and evaporating under reduced pressure to obtain the crude compound; the crude compound is subjected to silica gel column chromatography, the obtained eluent is dried and the solvent is removed under reduced pressure, thereby obtaining the fluorinated camptothecin drug derivative.

[0020] Further preferably, the separation and purification method comprises the following steps: adding saturated NaCl aqueous solution into the reaction mixture, extracting with dichloromethane, drying the organic layer with anhydrous sodium sulfate, filtering, and removing the solvent at room temperature by rotary evaporation to obtain the compound crude product; subjecting the compound crude product to silica gel column chromatography, taking a solution with a volume ratio of ethyl acetate to petroleum ether of 1:10 as the mobile phase, tracking TLC to collect the eluent with an Rf value of 0.3-0.5, removing the solvent from the collected eluent under reduced pressure, and drying to obtain the fluorinated camptothecin drug derivative as shown in formula (I).

[0021] In a second aspect, the present application provides a fluorinated camptothecin drug derivative prepared by the method.

[0022] In a third aspect, the present application provides the fluorinated camptothecin drug derivative prepared by the method or the use of the fluorinated camptothecin drug derivative in the preparation of an antitumor drug.

[0023] Preferably, the tumor includes cervical cancer, lung cancer, and liver cancer.

[0024] Compared with the prior art, the present application has the following beneficial effects:

[0025] The fluorinated camptothecin drug derivative in the present application is prepared by directly fluorinating the camptothecin drug derivative, and has the advantages of simple operation process, mild reaction condition, high site selectivity, and high reaction efficiency, and can be prepared by only one step reaction. In addition, the fluorinated camptothecin drug derivative in the present application has good antitumor activity, and provides a new scheme for antitumor drugs. BRIEF DESCRIPTION OF DRAWINGS

[0026] Figure 1 The figure is the nuclear magnetic hydrogen spectrum of the fluorinated camptothecin drug derivative in the present application.

[0027] Figure 2 The figure is the carbon spectrum of the fluorinated camptothecin drug derivative in the present application.

[0028] Figure 3 The figure is the fluorine spectrum of the fluorinated camptothecin drug derivative in the present application.

[0029] Figure 4 The figure is the comparison of the anti-cervical cancer activity of the fluorinated camptothecin drug derivative in the present application and the camptothecin drug derivative SN-38.

[0030] Figure 5 The figure is the comparison of the anti-lung cancer activity of the fluorinated camptothecin drug derivative in the present application and the camptothecin drug derivative SN-38.

[0031] Figure 6The anti-hepatoma activity of the fluorinated camptothecin drug derivative in the present application is compared with that of the anti-tumor drug camptothecin drug derivative SN-38. DETAILED DESCRIPTION

[0032] The present application will be further described below in conjunction with the accompanying drawings and specific examples. Those skilled in the art will be able to implement the present application based on these descriptions. In addition, the examples of the present application involved in the following descriptions are generally only examples of a part of the present application, rather than all examples. Therefore, all other examples obtained by those skilled in the art based on the examples in the present application without making creative efforts shall fall within the scope of protection of the present application.

[0033] Example 1 Preparation of fluorinated camptothecin drug derivative

[0034]

[0035] 1 mmol of camptothecin drug derivative SN-38 was added to 4 ml of 10% (w / w) acetic acid aqueous solution, 1.5 mmol of 1-chloromethyl-4-fluoro-1,4-diazonium bicyclo 2.2.2 octane bis (tetrafluoroboric acid) salt (Selectfluor) was added thereto, and the reaction was carried out at room temperature for 6 hours. After the reaction was completed, saturated NaCl aqueous solution was added to the reaction solution, and dichloromethane was used for extraction. The organic layer was dried over anhydrous sodium sulfate, filtered, and evaporated under reduced pressure to obtain the compound crude product. The compound crude product was subjected to silica gel column chromatography, and a solution of ethyl acetate and petroleum ether with a volume ratio of 1:9 was used as the mobile phase. TLC tracking was used to collect the eluate with an Rf value of 0.3-0.5. The collected eluate was dried by removing the solvent under reduced pressure to obtain 252 mg (yield 70%) of the fluorinated camptothecin drug derivative pure product shown as formula (I). The nuclear magnetic hydrogen spectrum thereof is shown as Figure 1 , the carbon spectrum is shown as Figure 2 , and the fluorine spectrum is shown as Figure 3 . 1 H NMR (500 MHz, DMSO-d6) δ 8.06 (dd, J = 17.8, 9.1 Hz, 1H), 7.61 (dd, J = 9.1, 5.2 Hz, 1H), 7.25 (d, J = 2.2 Hz, 2H), 6.54 (s, 1H), 5.42 (d, J = 2.1 Hz, 2H), 5.31 (d, J = 3.9 Hz, 2H), 3.60 (s, 2H), 1.86 (dq, J = 10.8, 6.9 Hz, 2H), 1.61 (s, 2H), 1.37 - 1.34 (m, 3H), 0.88 (d, J = 7.3 Hz, 3H).

[0036] Example 2 Preparation of fluorinated camptothecin drug derivative

[0037] The 1 mmol of the camptothecin drug derivative SN-38 was added into 4 ml of 10% (w / w) acetic acid aqueous solution, 1 mmol of Selectfluor was added into the solution, and the reaction was carried out at room temperature for 6 hours. After the reaction was completed, saturated NaCl aqueous solution was added into the reaction solution, dichloromethane was used to extract the solution, the organic layer was dried over anhydrous sodium sulfate, filtered, and evaporated under reduced pressure to obtain the compound crude product. The compound crude product was subjected to silica gel column chromatography, a solution of ethyl acetate and petroleum ether with a volume ratio of 1:9 was used as the mobile phase, and the eluate with an Rf value of 0.3-0.5 was collected by TLC tracking. The collected eluate was dried by removing the solvent under reduced pressure to obtain 601 mg of the fluorinated camptothecin drug derivative crude product shown in formula (I), with a purity of 93% and a yield of 95%.

[0038] Example 3 Preparation of a fluorinated camptothecin drug derivative

[0039] The 1 mmol of the camptothecin drug derivative SN-38 was added into 4 ml of 10% (w / w) acetic acid aqueous solution, 1 mmol of Selectfluor was added into the solution, and the reaction was carried out at room temperature for 6 hours. After the reaction was completed, saturated NaCl aqueous solution was added into the reaction solution, dichloromethane was used to extract the solution, the organic layer was dried over anhydrous sodium sulfate, filtered, and evaporated under reduced pressure to obtain the compound crude product. The compound crude product was subjected to silica gel column chromatography, a solution of ethyl acetate and petroleum ether with a volume ratio of 1:9 was used as the mobile phase, and the eluate with an Rf value of 0.3-0.5 was collected by TLC tracking. The collected eluate was dried by removing the solvent under reduced pressure to obtain 601 mg of the fluorinated camptothecin drug derivative crude product shown in formula (I), with a purity of 93% and a yield of 95%.

[0040] Example 4 Detection of the anti-tumor activity of a fluorinated camptothecin drug derivative

[0041] The tumor cells Hela (cervical cancer cells), A549 (lung cancer cells) and HepG2 (liver cancer cells) are selected, and the MTT method is used to detect the anti-tumor cell proliferation activity. Each kind of cell is inoculated into a 96-well plate containing 10% fetal bovine serum in 1640 culture solution at a concentration of 4000-5000 cells per well, and a note is added on the plate cover, and the plate is cultured at 5% CO2, 37℃ for 12 hours. After the cells are attached to the 96-well plate, the drug (fluorinated camptothecin drug derivative prepared in Example 1) is added to the sterile operation table with a pipette, so that the drug concentration of each well is 0.01 μM, 0.1 μM, 1 μM, 10 μM and 100 μM five concentration gradients, and five parallel groups are set for each concentration, and the anti-tumor drug SN-38 is used as a control, and the 96-well plate is again placed in a 5% CO2, 37℃ incubator for 24 hours. The 96-well plate is taken out, 10 μL of MTT reagent kit reagent (purchased from Promega Company) is added to each well, and incubated at 5% CO2, 37℃ for 4 hours in the dark. After the supernatant is aspirated, 150 μL of sterile DMSO is added to dissolve the formazan, and further dissolved in a 37℃ incubator for 5-10 min. Finally, the absorbance is measured by using an enzyme-labeled instrument. Thus, the inhibition rate and cytotoxicity of each cell are calculated, and the GraphPad Prism software is used for processing to calculate the IC 50 and IC 50 95% confidence interval, the anti-tumor activity detection results of Hela, A549 and HepG2 are shown in Figures 4 to 6 and Table 1.

[0042] Table 1

[0043] Compound A549 cell IC50 values (μm) HepG2 IC50 values (μm) Hela cell IC50 values (μm) Compound 3.34±0.4 7.28±0.5 1.935±0.4 SN-38 13.53±0.5 11.31±0.5 12.325±0.5

[0044] The experimental results show that the fluorinated camptothecin drug derivative in the present application has stronger anti-tumor activity than the original anti-tumor drug SN-38.

Claims

1. A method for preparing a fluorinated camptothecin drug derivative, characterized by, The method comprises the following steps: dispersing a camptothecin drug derivative as shown in formula (II) and a fluorination reagent in a solvent, reacting at room temperature, separating and purifying the reaction mixture obtained after the reaction, and thus obtaining the fluorinated camptothecin drug derivative as shown in formula (I); wherein the fluorination reagent is 1-chloromethyl-4-fluoro-1,4-diazonium bicyclo 2.2.2 octane bis (tetrafluoroboric acid) salt, the mass ratio of the camptothecin drug derivative to the added amount of the fluorination reagent is 1:1-3, and the solvent is an acetic acid aqueous solution.

2. The production method according to claim 1, wherein The mass ratio of the camptothecin drug derivative to the added amount of the fluorination reagent is 1:1.

5.

3. The production method according to claim 1, wherein The solvent is an 8-12% acetic acid aqueous solution.

4. The production method according to claim 1, wherein The separation and purification method comprises the following steps: adding a saturated NaCl aqueous solution to the reaction mixture, extracting with dichloromethane, drying the organic layer, filtering, and evaporating to dryness under reduced pressure to obtain a compound crude product; subjecting the compound crude product to silica gel column chromatography, collecting the obtained eluent, removing the solvent under reduced pressure, and drying, and thus obtaining the fluorinated camptothecin drug derivative.

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