Camptothecin-20-site ester derivative as well as preparation method, pharmaceutical composition and application thereof

Abstract

The invention relates to novel camptothecin-20-site ester derivatives (formulas I and II) or salts based on nitrogen-containing heterocyclic-1 '(N)-acetyl, a preparation method of the derivatives, a pharmaceutical composition containing the derivatives and application of the derivatives or salts as drugs, especially as antitumor drugs. The compounds have good commercial application prospects in preparation of the antitumor drugs.

Description

Technical Field

[0001] The present invention relates to a new class of camptothecin-20-position ester derivatives (Formula I, II) or salts based on nitrogen-containing heterocyclic-1'(N)-acetyl groups, their preparation methods, pharmaceutical compositions containing them, and their use as drugs, especially as anti-tumor drugs.

[0002] Background Art

[0003] As we all know, cancer remains the leading cause of death in humans. Small molecule drugs are widely used in the clinical treatment of cancer. However, many anti-tumor chemical agents cannot fully exert their potential therapeutic effects and even often cause serious side effects. This is mainly due to some of their own defects, such as poor solubility in aqueous solution, non-selective delivery to the target, and poor stability of their pharmacophores, which cannot ensure that they can fully reach the target site in the body.

[0004] Camptothecins (CPT) and their related analogs represent an important class of small molecule drugs that can be used to treat cancer but have some of the above defects. Camptothecin is an alkaloid isolated from Camptotheca acuminata, a plant of the Aquilegiaceae family. Studies have shown that it has anticancer activity against many solid tumors. Camptothecin mainly acts on DNA topoisomerase I (Topo I, an enzyme highly expressed in many cancer cells) and is the most classic specific inhibitor of Topo I. Studies have shown that the anti-tumor activity of camptothecin is attributed to its ability to stabilize the drug-DNA topoisomerase I (Topo I)-DNA ternary complex, interfere with the catalytic cycle of Topo I, and thus prevent the recombination of Topo I-induced single-stranded DNA breaks. Currently, Topo I has become one of the hottest targets for designing new anticancer drugs. The National Cancer Institute (NCI) drug mechanism analysis computer network system has listed Topo I inhibitors as one of the six major categories of anti-tumor drugs that are being studied. In addition to the United States and Japan, France, Germany, Italy and South Korea are also conducting research on camptothecin derivatives.

[0005] In view of the serious toxicity of camptothecin, such as bone marrow suppression, hemorrhagic cystitis and diarrhea, its application in cancer treatment is limited. Natural camptothecin is poorly soluble in water and has poor lipid solubility. In order to increase the water solubility of camptothecin, clinical trials in the 1970s evaluated the antitumor activity of camptothecin sodium salt. However, its sodium salt form is severely toxic to the human body and has low anticancer activity, so the Phase II clinical trial was interrupted. The discovery of topoisomerase I in the 1980s once again aroused the interest of oncologists in camptothecin. In 1989, studies found that semi-synthetic derivatives of camptothecin have extraordinary antitumor activity against nude mouse xenograft tumors. However, the antitumor activity of camptothecin in humans is very low, because there is a balance between the closed lactone form and the open carboxylate form in the body of camptothecin, and the open form depends on the pH value and the animal species. In particular, human serum albumin (HSA) preferentially binds to the open form of camptothecin to form a stable complex, which shifts the balance to the open form, making the content of the lactone form with antitumor activity in the body too low.

[0006] The camptothecin molecule contains four six-membered rings and one five-membered ring. The E ring is an α-hydroxy lactone with a chiral center (20S configuration). There is also an intramolecular hydrogen bond in the camptothecin molecule, which makes the carbonyl group of the E ring very active and easily open under the attack of nucleophilic reagents. Structure-activity relationship (SAR) studies have shown that the hydroxy lactone ring E of camptothecin is the most critical structural feature for anti-tumor activity. Experiments have shown that in nude mice, the activity of the closed lactone form of camptothecin is far superior to that of the open-ring form of camptothecin sodium salt. Therefore, the stability of the camptothecin lactone ring in vivo is an important factor affecting anti-tumor activity. The severe toxicity of camptothecin (MTD of mice is 12 mg / kg) is mainly because the open-ring form of camptothecin is tightly bound to HSA after the drug enters the human body, causing the equilibrium of camptothecin in the body to shift from the closed lactone form to the open-ring form, and camptothecin quickly opens the ring (half-life is less than half an hour).

[0007] Further SAR studies have shown that the natural 20S configuration of ring E lactone and camptothecin is crucial for its antitumor activity, while substitutions at positions 7, 9, and 10 of camptothecin can interfere with the binding of the open-ring form to HSA and improve the stability of the lactone ring in vivo, and the solubility of camptothecin derivatives can also be improved after substitution at position 7. Therefore, most structural modifications involve the A and B rings of CPT, among which there are a wide range of possibilities for variation, especially at positions 7, 9, 10, and 11. At the same time, according to literature reports and our previous studies, some camptothecin derivatives esterified with the 20-hydroxyl group can improve the lactone ring stability and water solubility of CPT, thereby improving the antitumor activity of camptothecin and reducing its toxicity. Some esterified derivatives not only have higher anticancer activity than irinotecan in vitro and in vivo, but also have much lower toxicity than CPT in vivo. The SAR also shows that the formation principle of hydrogen bonds and the fact that heterocyclic atoms, especially nitrogen atoms, easily form positive charge centers are conducive to the formation of CPTs-DNA-TOP I ternary complexes.

[0008] At present, the two water-soluble camptothecin derivatives irinotecan and topotecan abroad have definite efficacy and a wide anti-tumor spectrum compared with the parent compound camptothecin, and have been widely used in clinical practice. There are also several derivatives such as 9-nitrocamptothecin, DX-8951f, GG-211, Bay38-3441, BN80915 and CKD-602 undergoing II / III clinical studies. In China, only camptothecin, 10-hydroxycamptothecin, irinotecan and topotecan are on the market. Since camptothecin and 10-hydroxycamptothecin are unmodified natural products, they are highly toxic and have low anti-tumor activity, which limits their clinical application; moreover, patients are prone to drug resistance after long-term use of these drugs, and there is an urgent need to find highly effective and low-toxic anti-tumor drugs.

[0009] The compound of the present invention has significant cytotoxic activity against cancer cells and shows excellent anti-tumor activity. Summary of the invention

[0010] In order to overcome the deficiencies of the prior art, the object of the present invention is to provide a novel camptothecin derivative based on nitrogen-containing heterocyclic-1′(N)-acetyl esterification at the 20-position;

[0011] Another object of the present invention is to provide a novel method for preparing a camptothecin derivative based on esterification of the 20-position of a nitrogen-containing heterocyclic-1′(N)-acetyl group;

[0012] Another object of the present invention is to provide a novel camptothecin derivative based on nitrogen-containing heterocyclic-1'(N)-acetyl esterification at the 20-position and its composition for use as an anti-tumor drug.

[0013] One aspect of the present invention relates to a pharmaceutical composition, which comprises a compound of general formula (I, II) and its isomers as active ingredients and a carrier commonly used in the pharmaceutical field.

[0014] Another aspect of the present invention relates to the anti-tumor use of the compound of general formula (I, II) or a pharmaceutical composition containing the same.

[0015] Specifically, the present invention relates to compounds represented by general formula (I, II)

[0016] Among them, according to the compound represented by general formula (I) of the present invention:

[0017]

[0018] R 1 is hydrogen or C 1-8 Alkyl, C 1-8 Acyl, C 1-7 Hydroxyalkyl, C 3-6 Cycloalkyl, chloroacetyl, benzoyl, CH 2 NO 2 , CH 2 NO,CH 2 NR 5 R 6 , where R 5 and R 6 H, C 1-6 Alkyl, aryl substituted alkyl, hydroxy substituted C 1-6 Alkyl, amino substituted C 1-6 Alkyl, mono- or di-substituted amino C 1-6 Alkyl or cyclic amino group;

[0019] R 2 Selected from hydrogen, halogen, C 1-6 Alkyl, C 1-6 Alkoxy, hydroxy, RCOO, cyano, aldehyde, nitro, amino, halo 1-6 Alkyl, carboxyl, C 1-6 Alkoxycarbonyl, C 1-6 Alkoxycarbonylamino or CH 2 NR 7 R 8 , where R 7 and R 8 They are hydrogen, C 1-6 Alkyl, aryl substituted alkyl, hydroxy substituted C 1-6 Alkyl, amino substituted C 1-6 Alkyl, mono- or di-substituted amino C 1-6 Alkyl or cyclic amino, or R 2 Selected from CH 2 R 9 , where R 9C 1-6 Alkoxy or cyano, or R 2 Selected from NR 10 R 11 , where R 10 , R 11 C 1-6 Alkyl, aryl, dialkylaminoalkyl, amino C 1-6 Alkyl or hydroxyl C 1-6 alkyl;

[0020] R 3 For hydrogen, halogen, C 1-6 Alkyl, C 1-6 Alkoxy, amino substituted C 1-6 Alkoxy, hydroxy, RCOO, cyano, aldehyde, nitro, amino, halo 1-6 Alkyl, carboxyl, C 1-6 Alkoxycarbonyl, C 1-6 Alkoxycarbonylamino, amino C 1-6 Alkyl, hydroxyl C 1-6 Alkyl or [4-(1-piperidinyl)-1-1-piperidinyl]carbonyloxy;

[0021] R 2 Can and R 3 Ringing;

[0022] R 4 Selected from hydrogen, halogen, C 1-6 Alkyl, C 1-6 Alkoxy, RCOO, cyano, aldehyde, nitro, amino, halo C 1-6 Alkyl, carboxyl, C 1-6 Alkoxycarbonyl or C 1-6 Alkoxycarbonylamino;

[0023] R 3 Can and R 4 Ringing;

[0024] It is characterized in that: R is selected from hydrogen, halogen, nitro, amino, C 1-6 Alkyl, C 1-6 Alkoxy, cyano or aldehyde groups.

[0025] X is hydrogen or fluorine, chlorine, bromine, iodine, carboxyl, nitro, nitroso, amino, hydroxyl, C 1-8 Alkyl, C 1-8 Acyl, C 1-7 Hydroxyalkyl, C 3-6 Cycloalkyl, chloroacetyl, benzoyl, CH 2 NO 2 , CH 2 NO,CH 2 NR 5 R6 , where R 5 and R 6 H, C 1-6 Alkyl, aryl substituted alkyl, hydroxy substituted C 1-6 Alkyl, amino substituted C 1-6 Alkyl, mono- or di-substituted amino C 1-6 Alkyl or cyclic amino groups.

[0026] The above C 1-8 The alkyl group is selected from C 1-8 Straight or branched alkyl, C 1-8 The acyl group is selected from C 1-8 Straight-chain or branched alkanoyl, C 1-8 Alkenyl or C 1-8 Alkynoyl, C 1-7 The hydroxyalkyl group is selected from C 1-7 Straight or branched chain hydroxyalkyl;

[0027] More specifically, the C 1-8 The alkyl group is selected from methyl, ethyl, propyl, isopropyl, butyl, isobutyl, pentyl, isopentyl, hexyl, heptyl, etc.; C 1-8 The acyl group is selected from acetyl, propionyl, butyryl, isobutyryl, valeryl, isovaleryl, hexanoyl, heptanoyl, etc.; C 1-7 The hydroxyalkyl group is selected from hydroxymethyl, hydroxyethyl, hydroxypropyl, hydroxyisopropyl, hydroxybutyl, hydroxyisobutyl, hydroxypentyl, hydroxyisopentyl, hydroxyhexyl, hydroxyheptyl, etc.; C 3-6 Cycloalkyl is selected from cyclopentyl, cyclohexyl;

[0028] The above-mentioned camptothecin-20-ester derivative based on nitrogen-containing heterocyclic-1′(N)-acetyl group and its pharmaceutically acceptable salt are characterized in that the C20-hydroxyl group is in (S)-configuration.

[0029] According to the present invention, the compound represented by general formula (II):

[0030]

[0031] R 1 is hydrogen or C 1-8 Alkyl, C 1-8 Acyl, C 1-7 Hydroxyalkyl, C 3-6 Cycloalkyl, chloroacetyl, benzoyl, CH 2 NO 2 , CH 2 NO,CH 2 NR 5 R 6 , where R 5 and R 6 H, C 1-6Alkyl, aryl substituted alkyl, hydroxy substituted C 1-6 Alkyl, amino substituted C 1-6 Alkyl, mono- or di-substituted amino C 1-6 Alkyl or cyclic amino group;

[0032] R 2 Selected from hydrogen, halogen, C 1-6 Alkyl, C 1-6 Alkoxy, hydroxy, RCOO, cyano, aldehyde, nitro, amino, halo 1-6 Alkyl, carboxyl, C 1-6 Alkoxycarbonyl, C 1-6 Alkoxycarbonylamino or CH 2 NR 7 R 8 , where R 7 and R 8 They are hydrogen, C 1-6 Alkyl, aryl substituted alkyl, hydroxy substituted C 1-6 Alkyl, amino substituted C 1-6 Alkyl, mono- or di-substituted amino C 1-6 Alkyl or cyclic amino, or R 2 Selected from CH 2 R 9 , where R 9 C 1-6 Alkoxy or cyano, or R 2 Selected from NR 10 R 11 , where R 10 , R 11 C 1-6 Alkyl, aryl, dialkylaminoalkyl, amino C 1-6 Alkyl or hydroxyl C 1-6 alkyl;

[0033] R 3 For hydrogen, halogen, C 1-6 Alkyl, C 1-6 Alkoxy, amino substituted C 1-6 Alkoxy, hydroxy, RCOO, cyano, aldehyde, nitro, amino, halo 1-6 Alkyl, carboxyl, C 1-6 Alkoxycarbonyl, C 1-6 Alkoxycarbonylamino, amino C 1-6 Alkyl, hydroxyl C 1-6 Alkyl or [4-(1-piperidinyl)-1-1-piperidinyl]carbonyloxy;

[0034] R 2 Can and R 3 Ringing;

[0035] R 4 Selected from hydrogen, halogen, C1-6 Alkyl, C 1-6 Alkoxy, RCOO, cyano, aldehyde, nitro, amino, halo C 1-6 Alkyl, carboxyl, C 1-6 Alkoxycarbonyl or C 1-6 Alkoxycarbonylamino;

[0036] R 3 Can and R 4 Ringing;

[0037] It is characterized in that: R is selected from hydrogen, halogen, nitro, amino, C 1-6 Alkyl, C 1-6 Alkoxy, cyano or aldehyde groups.

[0038] X is hydrogen or fluorine, chlorine, bromine, iodine, carboxyl, nitro, nitroso, amino, hydroxyl, C 1-8 Alkyl, C 1-8 Acyl, C 1-7 Hydroxyalkyl, C 3-6 Cycloalkyl, chloroacetyl, benzoyl, CH 2 NO 2 , CH 2 NO,CH 2 NR 5 R 6 , where R 5 and R 6 H, C 1-6 Alkyl, aryl substituted alkyl, hydroxy substituted C 1-6 Alkyl, amino substituted C 1-6 Alkyl, mono- or di-substituted amino C 1-6 Alkyl or cyclic amino groups.

[0039] n is 1-4.

[0040] The above C 1-8 The alkyl group is selected from C 1-8 Straight or branched alkyl, C 1-8 The acyl group is selected from C 1-8 Straight-chain or branched alkanoyl, C 1-8 Alkenyl or C 1-8 Alkynoyl, C 1-7 The hydroxyalkyl group is selected from C 1-7 Straight or branched chain hydroxyalkyl;

[0041] More specifically, the C 1-8 The alkyl group is selected from methyl, ethyl, propyl, isopropyl, butyl, isobutyl, pentyl, isopentyl, hexyl, heptyl, etc.; C 1-8 The acyl group is selected from acetyl, propionyl, butyryl, isobutyryl, valeryl, isovaleryl, hexanoyl, heptanoyl, etc.; C 1-7The hydroxyalkyl group is selected from hydroxymethyl, hydroxyethyl, hydroxypropyl, hydroxyisopropyl, hydroxybutyl, hydroxyisobutyl, hydroxypentyl, hydroxyisopentyl, hydroxyhexyl, hydroxyheptyl, etc.; C 3-6 Cycloalkyl is selected from cyclopentyl, cyclohexyl;

[0042] The above-mentioned camptothecin-20-ester derivative based on nitrogen-containing heterocyclic-1′(N)-acetyl group and its pharmaceutically acceptable salt are characterized in that the C20-hydroxyl group is in (S)-configuration.

[0043] According to the present invention, the compound of the present invention may exist in the form of isomers, and generally the "compound of the present invention" includes the isomers of the compound.

[0044] The compounds of the present invention may exist as cis-trans isomers of the double bond, and the asymmetric center may have an S configuration or an R configuration. The present invention includes all possible stereoisomers and mixtures of two or more isomers. If cis / trans isomers exist, the present invention relates to the cis form and the trans form and mixtures of these forms. If a single isomer is required, it can be separated according to conventional methods or prepared by stereoselective synthesis.

[0045] According to an embodiment of the present invention, the compound of the present invention also includes its pharmacologically acceptable salt, salt hydrate, ester or prodrug.

[0046] The present invention also relates to a method for preparing the compounds of the present invention, wherein camptothecin having a hydroxyl group at position 20 or its analogue having a substituent at position 7 is subjected to a condensation reaction with a corresponding nitrogen-containing heterocyclic-1′(N)-acetic acid or an acyl halide to prepare the corresponding compound.

[0047] The condensation reaction occurs in the presence of a suitable condensation agent and catalyst.

[0048] The preferred condensing agent is selected from 1,3-dicyclohexylcarbodiimide (DCC), dipyridyl carbonate (2-DPC), 1-(3-dimethylaminopropyl)-3-ethylcarbodiimide hydrochloride (EDCI), 1,3-diisopropylcarbodiimide (DIPC), 4-pyrrolidinylpyridine. The more preferred condensing agent is 1-(3-dimethylaminopropyl)-3-ethylcarbodiimide hydrochloride (EDCI);

[0049] The preferred catalyst is selected from tertiary amines. The preferred tertiary amine is selected from 4-dimethylaminopyridine (DMAP), 4-pyrrolidinopyridine. The more preferred tertiary amine is selected from 4-dimethylaminopyridine (DMAP);

[0050] The reaction temperature is between 0°C and 60°C, preferably between 20°C and 30°C, and more preferably at room temperature;

[0051] The reaction is carried out in a suitable solvent, preferably an anhydrous aprotic solvent, more preferably alkyl halide, DMF, tetrahydrofuran (THF), N,N-dimethylformamide (DMF), chloroform, pyridine, dichloroethane, etc. The most preferred solvent is dichloromethane.

[0052] The present invention therefore also relates to a pharmaceutical composition containing the compound of the present invention as an active ingredient and conventional pharmaceutical excipients or adjuvants or other pharmacologically acceptable carriers. Usually the pharmaceutical composition of the present invention contains 0.1-95% by weight of the compound of the present invention.

[0053] The pharmaceutical composition of the compounds of this invention can be prepared according to methods well known in the art. When used for this purpose, if necessary, the compounds of this invention can be combined with one or more solid or liquid pharmaceutical excipients and / or adjuvants to prepare suitable application forms or dosage forms that can be used as human or veterinary medicines.

[0054] The compound of the present invention or the pharmaceutical composition containing the same can be administered in a unit dosage form, and the administration route can be enteral or parenteral, such as oral, intramuscular, subcutaneous, nasal, oral mucosa, skin, peritoneum or rectum.

[0055] The administration route of the compound of the present invention or the pharmaceutical composition containing the compound can be injection, which includes intravenous injection, intramuscular injection, subcutaneous injection, intradermal injection and acupoint injection.

[0056] The dosage form for administration can be a liquid dosage form or a solid dosage form. For example, the liquid dosage form can be a true solution, a colloid, a microparticle dosage form, an emulsion dosage form, or a suspension dosage form. Other dosage forms include tablets, capsules, dripping pills, aerosols, pills, powders, solutions, suspensions, emulsions, granules, suppositories, freeze-dried powder injections, and the like.

[0057] The compound of the present invention can be prepared into common preparations, sustained-release preparations, controlled-release preparations, targeted preparations and various microparticle drug delivery systems.

[0058] For example, in order to formulate a unit dosage form into tablets, various carriers well known in the art can be widely used. Examples of carriers include diluents and absorbents, such as starch, dextrin, calcium sulfate, lactose, mannitol, sucrose, sodium chloride, glucose, urea, calcium carbonate, kaolin, microcrystalline cellulose, aluminum silicate, etc.; wetting agents and binders, such as water, glycerol, polyethylene glycol, ethanol, propanol, starch slurry, dextrin, syrup, honey, glucose solution, acacia slurry, gelatin slurry, sodium carboxymethyl cellulose, shellac, methylcellulose, potassium phosphate, polyvinyl pyrrolidone, etc.; disintegrants, such as dry starch, alginate, agar powder, brown seaweed starch, sodium bicarbonate and citric acid, calcium carbonate, polyoxyethylene sorbitan fatty acid esters, sodium lauryl sulfate, methylcellulose, ethylcellulose, etc.; disintegration inhibitors, such as sucrose, tristearin, cocoa butter, hydrogenated oil, etc.; absorption promoters, such as quaternary ammonium salts, sodium lauryl sulfate, etc.; lubricants, such as talc, silicon dioxide, corn starch, stearate, boric acid, liquid paraffin, polyethylene glycol, etc. The tablets can be further made into coated tablets, such as sugar-coated tablets, film-coated tablets, enteric-coated tablets, or double-layer tablets and multi-layer tablets.

[0059] For example, in order to make the dosing unit into a pill, various carriers known in the art can be widely used. Examples of carriers include diluents and absorbents, such as glucose, lactose, starch, cocoa butter, hydrogenated vegetable oil, polyvinyl pyrrolidone, Gelucire, kaolin, talc, etc.; binders, such as gum arabic, tragacanth gum, gelatin, ethanol, honey, liquid sugar, rice paste or flour paste, etc.; disintegrants, such as agar powder, dry starch, alginate, sodium dodecyl sulfate, methyl cellulose, ethyl cellulose, etc.

[0060] For example, in order to prepare the dosing unit into a capsule, the active ingredient compound of the present invention is mixed with the above-mentioned various carriers, and the resulting mixture is placed in a hard gelatin capsule or a soft capsule. The active ingredient compound of the present invention can also be prepared into a microcapsule, suspended in an aqueous medium to form a suspension, or loaded into a hard capsule or prepared as an injection for use.

[0061] For example, the compound of the present invention is made into an injection preparation, such as a solution, a suspension solution, an emulsion, or a freeze-dried powder injection. This preparation can be aqueous or non-aqueous, and can contain one and / or more pharmacodynamically acceptable carriers, diluents, adhesives, lubricants, preservatives, surfactants, or dispersants. For example, the diluent can be selected from water, ethanol, polyethylene glycol, 1,3-propylene glycol, ethoxylated isostearyl alcohol, polyoxyethylene sorbitol fatty acid esters, and the like. In addition, in order to prepare an isotonic injection, an appropriate amount of sodium chloride, glucose, or glycerol can be added to the injection preparation. In addition, conventional cosolvents, buffers, pH regulators, and the like can also be added. These adjuvants are commonly used in the art.

[0062] In addition, if necessary, colorants, preservatives, perfumes, flavoring agents, sweeteners or other materials may be added to the pharmaceutical preparations.

[0063] The dosage of the pharmaceutical composition of the compound of the present invention depends on many factors, such as the nature and severity of the disease to be prevented or treated, the sex, age, weight, personality and individual response of the patient or animal, the route of administration, the number of administrations, etc. Therefore, the therapeutic dosage of the present invention can vary widely.

[0064] Generally speaking, the dosage of the pharmaceutical ingredients of the present invention is well known to those skilled in the art. It can be appropriately adjusted according to the actual amount of the drug contained in the final preparation of the compound composition of the present invention to achieve the requirement of the therapeutically effective amount and complete the prevention or treatment purpose of the present invention.

[0065] Suitable daily doses of the compounds of the present invention range from 1 to 1000 mg / m 2 , especially 10-100 mg / m 2 The above dosage can be administered in a single dosage form or divided into several dosage forms, such as two, three or four dosage forms, which is limited by the clinical experience of the administering physician and the administration regimen including the use of chemotherapy and radiotherapy.

[0066] In the in vitro anti-tumor activity experiment, the compounds of the present invention showed good anti-tumor activity. The half inhibitory concentration (IC 50 ) are all lower than topotecan, and the IC 50 The cytotoxic activity of most of the compounds of the present invention is better than that of topotecan, especially the IC values ​​of the compounds of the present invention in human non-small cell lung cancer (A549) and colon cancer (HCT-8) cells. 50 The lowest can reach 4×10 -9 mol / L and 7×10 -9 mol / L, and their activities are 15 and 40 times that of topotecan, respectively. The IC values ​​of some preferred compounds of the present invention for A549, Bel7402, HCT-8, A2780 50 Comparable to camptothecin.

[0067] The compound of the present invention has a good commercial application prospect in the preparation of anti-tumor drugs. The tumors include but are not limited to non-small cell lung cancer, colorectal cancer, gastric cancer, liver cancer, and ovarian cancer. BRIEF DESCRIPTION OF THE DRAWINGS

[0068] Figure 1It is a camptothecin-20-position esterified derivative based on nitrogen-containing heterocyclic-1'(N)-acetyl group as shown in formula I and II of the present invention.

[0069] Figure 2 The present invention shows a synthetic route for the camptothecin-20-position esterified derivatives based on nitrogen-containing heterocyclic-1′(N)-acetyl groups as shown in formulas I and II of the present invention.

[0070] Figure 3 The synthetic route of the nitrogen-containing heterocyclic-1′(N)-acetic acid (Compound 1-4) and its camptothecin-20-esterified derivative (Compound 5-11) described in Example 1-7 of the present invention. DETAILED DESCRIPTION

[0071] The following examples are used to further illustrate the present invention, but they do not mean any limitation of the present invention. The methods are conventional methods unless otherwise specified. The raw materials can be obtained from public commercial channels unless otherwise specified.

[0072] The reactants camptothecin and 7-ethylcamptothecin used in the following examples are available from commercial sources, and the other reagents, solvents and catalysts used are available from commercial sources.

[0073] Example 1, 5-carboxyluracil-1-acetic acid (Compound 1)

[0074] In a 25mL round-bottom flask, 2.56g (45.7mmol) potassium hydroxide and 8ml water were added. After stirring and dissolving, 1.56g (10.0mmol) 5-carboxyuracil was added, followed by 1.7g (12.2mmol) bromoacetic acid in 5ml aqueous solution under steady stirring at 40°C. The reaction mixture was stirred at 60°C for 5 hours, then cooled in an ice bath, and the pH was adjusted to 5.5 with hydrochloric acid. After filtering through a membrane, the crude product was recrystallized in water to obtain 1.13g of a white solid, which was 5-carboxyuracil-1-acetic acid (Compound 1). Yield: 52.8%, mp 260-262°C.

[0075] The structural characterization data are as follows:

[0076] 1 H NMR (400 MHz, DMSO-d 6 , δppm): 12.95 (br s, 2H, COOH), 12.25 (s, 1H, NH), 8.72 (s, 1H, 6-H), 4.62 (s, 2H, OOCCH 2 -N); 13 C-NMR (100 MHz, DMSO-d 6, δppm): δ169.0, 163.7, 163.3, 153.7, 149.8, 102.0, 49.4; MS (ESI - ): m / z 213[MH] - , 427[2M-H] - ;HRESIMS: m / z 215.0302[M+H] + (calcd for C 7 H 7 N 2 O 6 , 215.0299).

[0077] Example 2, 5-methyluracil-1-acetic acid (Compound 2)

[0078] In a 25mL round-bottom flask, 2-56g (45.7mmol) potassium hydroxide and 8ml water were added. After stirring and dissolving, 1.26g (10.0mmol) 5-methyluracil was added, followed by 1.7g (12.2mmol) bromoacetic acid in 5ml aqueous solution under steady stirring at 40°C. The reaction mixture was stirred at 60°C for 5 hours, then cooled in an ice bath, and the pH was adjusted to 5.5 with hydrochloric acid. After filtering through a membrane, the crude product was recrystallized in water to obtain 1.26g of a white solid, which was 5-methyluracil-1-acetic acid (Compound 2). Yield: 68.4%, mp 292-294°C.

[0079] The structural characterization data are as follows:

[0080] 1 H NMR (400 MHz, DMSO-d 6 , δppm): 13.13 (br s, 1H, COOH), 11.35 (s, 1H, NH), 7.48 (s, 1H, 6-H), 4.37 (s, 2H, OOCCH 2 -N), 1.75 (s, 3H, 5-CH 3 ); 13 C-NMR (100 MHz, DMSO-d 6 , δppm): 169.7, 164.3, 150.9, 141.7, 108.3, 48.4, 11.8; MS (ESI - ): m / z 183[MH] - , 367[2M-H] - ;HRESIMS: m / z 185.0556[M+H] + (calcd for C 7 H 9 N2 O 4 , 185.0557).

[0081] Example 3, 5-ethyluracil-1-acetic acid (Compound 3)

[0082] In a 25mL round-bottom flask, add 2.56g (45.7mmol) potassium hydroxide and 8ml water, stir and dissolve, then add 1.40g (10.0mmol) 5-ethyluracil, then add 1.7g (12.2mmol) bromoacetic acid in 5ml aqueous solution under steady stirring at 40°C. The reaction mixture is stirred at 60°C for 5 hours, then cooled by ice bath, and the pH is adjusted to 5.5 with hydrochloric acid. After filtering through a membrane, the crude product is recrystallized in water to obtain 1.26g white solid, which is 5-ethyluracil-1-acetic acid (Compound 3). Yield: 63.7%, mp>300°C.

[0083] The structural characterization data are as follows:

[0084] 1 H NMR (400 MHz, DMSO-d 6 , δppm): 13.12 (br s, 1H, COOH), 11.345 (s, 1H, NH), 7.46 (s, 1H, 6-H), 4.40 (s, 2H, OOCCH 2 -N), 2.19 (m, 2H, 5-CH 2 Me), 1.03(t, J=7.2Hz, 3H, 5-Et-CH 3 ); 13 C-NMR (100 MHz, DMSO-d 6 , δppm): 169.6, 163.9, 150.8, 141.2, 114.1, 48.5, 19.4, 12.8; MS (ESI - ): m / z 197[MH] - , 395[2M-H] - ;HRESIMS: m / z 199.0717[M+H] + (calcd for C 8 H 11 N 2 O 4 , 199.0713).

[0085] Example 4, 3,5-dichloro-4-pyridone-1-acetic acid (Compound 4)

[0086] In a 25mL round-bottom flask, 2.56g (45.7mmol) potassium hydroxide and 8ml water were added. After stirring and dissolving, 1.63g (10.0mmol) 3,5-dichloro-4-pyridone was added, followed by 5ml aqueous solution of 1.7g (12.2mmol) bromoacetic acid at 40°C with steady stirring. The reaction mixture was stirred at 60°C for 5 hours, then cooled by ice bath, and pH was adjusted to 5.5 with hydrochloric acid. After filtering through a membrane, the crude product was recrystallized in water to obtain 2.01g of white solid, which was 3,5-dichloro-4-pyridone-1-acetic acid (Compound 4). Yield: 91.0%, mp 236-238°C.

[0087] The structural characterization data are as follows:

[0088] 1 H NMR (400 MHz, DMSO-d 6 , δppm): 12.79 (br s, 1H, COOH), 8.30 (s, 2H, 2-H&6-H), 4.82 (s, 2H, OOCCH 2 -N); 13 C-NMR (100 MHz, DMSO-d 6 , δppm): 169.1, 166.4, 140.0, 140.0, 121.7, 121.7, 56.2; HRESIMS: m / z 221.9722[M+H] + (calcd for C 7 H 6 Cl 2 NO 3 , 221.9719).

[0089] Example 5, 20S-camptothecin ester of 5-carboxyuracil-1-acetic acid (Compound 5)

[0090] In a 10 mL round-bottom flask, 14.1 mg (0.041 mmol) of camptothecin, 26.5 mg (0.124 mmol) of 5-carboxyuracil-1-acetic acid (Compound 1), 30.0 mg (0.157 mmol) of 1-[3-(dimethylamino)propyl]-3-ethylcarbodiimide hydrochloride (EDCI), 12.0 mg (0.098 mmol) of DMAP and 5 mL of dichloromethane were added. The reaction mixture was stirred at room temperature for 8 to 12 hours, and the reaction was monitored by TLC until the reaction was completed (clear or nearly clear). 40 ml of chloroform was added to dilute the reaction solution, and the organic phase was washed with water (35 ml) and saturated sodium bicarbonate aqueous solution (35 ml) in turn, and dried with anhydrous sodium sulfate or anhydrous magnesium sulfate. After filtering to remove the sodium sulfate or magnesium sulfate, the solvent was removed by concentration under reduced pressure. The residue was dissolved in chloroform and chromatographed on a silica gel column (eluent: chloroform-methanol 98:2 to 95:5 for gradient elution) to obtain 11.2 mg of a light yellow solid, which was 20S-camptothecin ester of 5-carboxyuracil-1-acetic acid, i.e., compound 5, with a yield of 50.2% and mp of 252-254°C.

[0091] The structural characterization data are as follows:

[0092] 1 H NMR (400 MHz, DMSO-d 6 , δppm): 11.97 (br s, 1H, NH), 8.80 (m, 1H, Ar-H), 8.66 (s, 1H, 6'-H), 8.18 (t, 1H, J=9.2Hz, Ar-H), 8.09 (d, 1H, J=7.6Hz, Ar-H), 7.87 (m , 1H, Ar-H), 7.71 (t, 1H, J=6.8Hz, Ar-H), 7.24 (s, 1H, Ar-H), 5.51 (s, 2H, H-17), 5.26 (s, 2H, H-5), 5.13 (m, 2H, OOCCH 2 -N), 2.19 (m, 2H, 19-CH 2 ), 0.93 (t, 3H, J=7.6Hz, 18-CH 3 ); 13 C-NMR (100 MHz, DMSO-d 6, δppm): 166.7, 161.8, 156.4, 152.3, 150.0, 149.9, 147.9, 146.0, 144.7, 131.5, 130.4, 129.7, 129.7, 128.9, 128.5, 127.9, 127.7, 126.0, 118.9, 95.3, 77.0, 66.4, 50.2, 42.0, 42.0, 30.4, 7.5; HRESIMS: m / z 545.1312[M+H] + (calcd for C 27 H 22 N 4 O 9 , 545.1303).

[0093] Example 6, 20S-camptothecin ester of 5-methyluracil-1-acetic acid (Compound 6)

[0094] In a 10 mL round-bottom flask, 14.1 mg (0.041 mmol) of camptothecin, 22.8 mg (0.124 mmol) of 5-methyluracil-1-acetic acid (Compound 2), 30.0 mg (0.157 mmol) of 1-[3-(dimethylamino)propyl]-3-ethylcarbodiimide hydrochloride (EDCI), 12.0 mg (0.098 mmol) of DMAP and 5 mL of dichloromethane were added. The reaction mixture was stirred at room temperature for 8 to 12 hours, and the reaction was monitored by TLC until the reaction was completed (clear or nearly clear). 40 ml of chloroform was added to dilute the reaction solution, and the organic phase was washed with water (35 ml) and saturated sodium bicarbonate aqueous solution (35 ml) in turn, and dried with anhydrous sodium sulfate or anhydrous magnesium sulfate. After filtering to remove the sodium sulfate or magnesium sulfate, the solvent was removed by concentrating under reduced pressure. The residue was dissolved in chloroform and chromatographed on a silica gel column (eluent: chloroform-methanol 98:2 to 95:5 for gradient elution) to obtain 12.3 mg of a light yellow solid, which is 20S-camptothecin ester of 5-methyluracil-1-acetic acid, i.e., compound 6, with a yield of 58.3% and mp of 263-264°C.

[0095] The structural characterization data are as follows:

[0096] 1 H NMR (400 MHz, DMSO-d 6, δppm): 11.39 (s, 1H, NH), 8.63 (s, 1H, Ar-H), 8.15 (d, J=8.4Hz, 1H, Ar-H), 8.07 (d, J=8.0Hz, 1H, Ar-H), 7.86 (t, 1H, J=7.2Hz, Ar -H), 7.69 (t, 1H, J=7.6Hz, Ar-H), 7.55 (s, 1H, 6′-H), 7.20 (s, 1H, Ar-H), 5.51 (s, 2H, H-17), 5.23 (s, 2H, H-5), 4.78 (m, 2H, OOCCH 2 -N), 2.17(dd, 2H, J=14.4Hz, 7.2Hz, 19-CH 2 ), 1.72(s, 3H, 5′-CH 3 ), 0.93 (t, 3H, J=7.2Hz, 18-CH 3 ); 13 C-NMR (100 MHz, DMSO-d 6 , δppm): 167.5, 166.8, 164.2, 156.4, 152.3, 150.8, 147.8, 146.0, 144.6, 141.1, 131.5, 130.4, 129.7, 128.9, 128.4, 127.9, 127.6, 118.9, 108.8, 95.2, 76.9, 66.3, 50.1, 48.1, 30.4, 11.8, 7.5; HRESIMS: m / z 515.1565[M+H] + (calcd for C 27 H 23 N 4 O 7 , 515.1561).

[0097] Example 7, 20S-camptothecin ester of 5-ethyluracil-1-acetic acid (Compound 7)

[0098] In a 10 mL round-bottom flask, 14.1 mg (0.041 mmol) of camptothecin, 24.6 mg (0.124 mmol) of 5-ethyluracil-1-acetic acid (Compound 3), 30.0 mg (0.157 mmol) of 1-[3-(dimethylamino)propyl]-3-ethylcarbodiimide hydrochloride (EDCI), 12.0 mg (0.098 mmol) of DMAP and 5 mL of dichloromethane were added. The reaction mixture was stirred at room temperature for 8 to 12 hours, and the reaction was monitored by TLC until the reaction was completed (clear or nearly clear). 40 ml of chloroform was added to dilute the reaction solution, and the organic phase was washed with water (35 ml) and saturated sodium bicarbonate aqueous solution (35 ml) in turn, and dried with anhydrous sodium sulfate or anhydrous magnesium sulfate. After filtering to remove the sodium sulfate or magnesium sulfate, the solvent was removed by concentration under reduced pressure. The residue was dissolved in chloroform and chromatographed on a silica gel column (eluent: chloroform-methanol 98:2 to 95:5 for gradient elution) to obtain 13.1 mg of a light yellow solid, which is 20S-camptothecin ester of 5-ethyluracil-1-acetic acid, i.e., compound 7, with a yield of 60.3% and mp of 285-287°C.

[0099] The structural characterization data are as follows:

[0100] 1 H NMR (400 MHz, DMSO-d 6 , δppm): 11.37 (s, 1H, NH), 8.68 (d, J=5.2Hz, 1H, Ar-H), 8.17 (m, 2H, Ar-H), 7.88 (t, 1H, J=7.6Hz, Ar-H), 7.72 (m, 1H, Ar-H), 7.52 (s, 1H, 6′-H), 7.21 (s, 1H, Ar-H), 5.51 (s, 2H, H-17), 5.28 (s, 2H, H-5), 4.81 (m, 2H, OOCCH 2 -N), 2.17(dd, 4H, J=14.8, 7.2Hz, 5′-CH 2 &19-CH 2 ), 0.97 (m, 6H, 5′α-CH 3 &18-CH 3 ); 13 C-NMR (100 MHz, DMSO-d 6, δppm): 167.9, 167.3, 164.2, 156.9, 152.8, 151.2, 148.4, 146.5, 145.2, 141.0, 132.0, 130.9, 130. 2, 129.4, 129.0, 128.4, 128.2, 119.4, 115.1, 95.7, 66.8, 50.7, 48.7, 30.8, 19.8, 13.2, 7.9; MS (ESI + ): m / z 551[M+Na] + , 1079[2M+Na] + ;HRESIMS: m / z 529.1715[M+H] + (calcd for C 28 H 25 N 4 O 7 , 529.1718).

[0101] Example 8, 7-ethyl-20S-camptothecin ester of 5-methyluracil-1-acetic acid (Compound 8)

[0102] In a 10 mL round-bottom flask, 15.4 mg (0.041 mmol) of 7-ethyl-camptothecin, 22.8 mg (0.124 mmol) of 5-methyluracil-1-acetic acid (Compound 2), 30.0 mg (0.157 mmol) of 1-[3-(dimethylamino)propyl]-3-ethylcarbodiimide hydrochloride (EDCI), 12.0 mg (0.098 mmol) of DMAP and 5 mL of dichloromethane were added. The reaction mixture was stirred at room temperature for 8 to 12 hours, and the reaction was monitored by TLC until the reaction was completed (clear or nearly clear). 40 ml of chloroform was added to dilute the reaction solution, and the organic phase was washed with water (35 ml) and saturated sodium bicarbonate aqueous solution (35 ml) in turn, and dried with anhydrous sodium sulfate or anhydrous magnesium sulfate. After filtering to remove the sodium sulfate or magnesium sulfate, the solvent was removed by concentration under reduced pressure. The residue was dissolved in chloroform and chromatographed on a silica gel column (eluent: chloroform-methanol 98:2 to 95:5 for gradient elution) to obtain 11.2 mg of an orange solid, which is 7-ethyl-20S-camptothecin ester of 5-methyluracil-1-acetic acid, i.e., compound 8, with a yield of 50.3% and mp of 290-292°C.

[0103] The structural characterization data are as follows:

[0104] 1 H NMR (400 MHz, DMSO-d 6, δppm): 11.40 (s, 1H, NH), 8.28 (d, 1H, J=8.4Hz, Ar-H), 8.17 (d, 1H, J=8.4Hz, Ar-H), 7.86 (t, 1H, J=7.2Hz, Ar-H), 7.73 (t, 1H, J=7.6Hz, Ar-H), 7.54 (s, 1H, 6'-H), 7.19 (s, 1H, Ar-H), 5.52 (s, 2H, H-17), 5.31 (m, 2H, H-5), 4.77 (m, 2H, OOCCH 2 -N), 3.21 (dd, 2H, J=13.6, 7.2Hz, 7-CH 2 ), 2.17 (dd, 2H, J=14.4, 7.6Hz, 19-CH 2 ), 1.72(s, 3H, 5′-CH 3 ), 1.30 (t, 3H, J=7.2Hz, 7α-CH 3 ), 0.93 (t, 3H, J=7.2Hz, 18-CH 3 ); 13 C-NMR (100 MHz, DMSO-d 6 , δppm): 167.5, 166.8, 164.2, 156.4, 151.7, 150.8, 148.4, 146.6, 145.5, 144.6, 141.1, 130.0, 129.8, 127 .9, 127.6, 126.5, 124.0, 118.8, 108.8, 95.1, 76.8, 66.4, 49.5, 48.1, 30.3, 22.2, 13.9, 11.8, 7.4; MS (ESI + ): m / z 543[M+H] + , 1085[2M+H] + , 1107[2M+Na] + ;MS(ESI - ): m / z 541[MH] - ;HRESIMS: m / z 543.1880[M+H] + (calcd for C 29 H 28 N 4 O 7 , 543.1874).

[0105] Example 9, 7-ethyl-20S-camptothecin ester of 5-ethyluracil-1-acetic acid (Compound 9)

[0106] In a 10 mL round-bottom flask, 15.4 mg (0.041 mmol) of 7-ethyl-camptothecin, 24.6 mg (0.124 mmol) of 5-ethyluracil-1-acetic acid (Compound 3), 30.0 mg (0.157 mmol) of 1-[3-(dimethylamino)propyl]-3-ethylcarbodiimide hydrochloride (EDCI), 12.0 mg (0.098 mmol) of DMAP and 5 mL of dichloromethane were added. The reaction mixture was stirred at room temperature for 8 to 12 hours, and the reaction was monitored by TLC until the reaction was completed (clear or nearly clear). 40 ml of chloroform was added to dilute the reaction solution, and the organic phase was washed with water (35 ml) and saturated sodium bicarbonate aqueous solution (35 ml) in turn, and dried with anhydrous sodium sulfate or anhydrous magnesium sulfate. After filtering to remove the sodium sulfate or magnesium sulfate, the solvent was removed by concentration under reduced pressure. The residue was dissolved in chloroform and chromatographed on a silica gel column (eluent: chloroform-methanol 98:2 to 95:5 for gradient elution) to obtain 14.8 mg of a light yellow solid, which was 7-ethyl-20S-camptothecin ester of 5-ethyluracil-1-acetic acid, i.e., compound 9, with a yield of 64.8% and mp>290°C (dec.).

[0107] The structural characterization data are as follows:

[0108] 1 H NMR (400 MHz, DMSO-d 6 , δppm): 11.38 (s, 1H, NH), 8.28 (d, 1H, J=8.4Hz, Ar-H), 8.17 (d, 1H, J=8.4Hz, Ar-H), 7.87 (t, 1H, J=7.6Hz, Ar-H), 7.74 (t, 1 H, J=7.6Hz, Ar-H), 7.52 (s, 1H, 6'-H), 7.20 (s, 1H, Ar-H), 5.52 (s, 2H, H-17), 5.34 (d, 2H, J=6.8Hz, H-5), 4.81 (m, 2H, OOCCH 2 -N), 3.21 (dd, J=15.2, 7.2Hz, 2H, 7-CH 2 ), 2.17 (dd, 4H, J=14.0, 7.2Hz, 19-CH 2 &5′-CH 2 ), 1.30 (t, 3H, J=7.6Hz, 7α-CH 3 ), 0.97 (t, 3H, J=7.2Hz, 5′α-CH 3 ), 0.92 (t, 3H, J=7.6Hz, 18-CH 3 ); 13 C-NMR (100 MHz, DMSO-d6 , δppm): 167.4, 166.8, 163.8, 156.5, 151.7, 150.7, 148.4, 146.6, 145.6, 144.7, 140.5, 130.0, 129.8, 128.0 , 127.6, 126.5, 124.0, 118.8, 114.6, 95.1, 76.9, 66.3, 49.5, 48.1, 30.3, 22.2, 19.3, 13.9, 12.7, 7.4; MS (ESI + ): m / z 557[M+H] + , 579[M+Na] + , 1113[2M+H] + , 1135[2M+Na] + ;MS(ESI - ): m / z 555[MH] - ;HRESIMS: m / z 557.2034[M+H] + (calcd for C 30 H 29 N 4 O 7 , 557.2031).

[0109] Example 10, 20S-camptothecin ester of 3,5-dichloro-4-pyridone-1-acetic acid (Compound 10)

[0110] In a 10 mL round-bottom flask, 14.1 mg (0.041 mmol) of camptothecin, 27.5 mg (0.124 mmol) of 3,5-dichloro-4-pyridone-1-acetic acid (Compound 4), 30.0 mg (0.157 mmol) of 1-[3-(dimethylamino)propyl]-3-ethylcarbodiimide hydrochloride (EDCI), 12.0 mg (0.098 mmol) of DMAP and 5 mL of dichloromethane were added. The reaction mixture was stirred at room temperature for 8 to 12 hours, and the reaction was monitored by TLC until the reaction was completed (clear or nearly clear). 40 ml of chloroform was added to dilute the reaction solution, and the organic phase was washed with water (35 ml) and saturated sodium bicarbonate aqueous solution (35 ml) in turn, and dried with anhydrous sodium sulfate or anhydrous magnesium sulfate. After filtering to remove the sodium sulfate or magnesium sulfate, the solvent was removed by concentration under reduced pressure. The residue was dissolved in chloroform and chromatographed on a silica gel column (eluent: chloroform-methanol 98:2 to 95:5 for gradient elution) to obtain 11.2 mg of orange-yellow powder, which is 20S-camptothecin ester of 3,5-dichloro-4-pyridone-1-acetic acid, i.e., compound 10, yield: 49.5%, mp 256-258°C.

[0111] The structural characterization data are as follows:

[0112] 1 H NMR (300 MHz, DMSO-d 6 , δppm): 8.69 (d, 1H, J=5.2Hz, Ar-H), 8.37 (s, 2H, 2′-H&6′-H), 8.19 (m, 2H, Ar-H), 7.89 (t, J=7.8Hz, 1H, Ar- H), 7.73 (t, 1H, J=7.8Hz, Ar-H), 7.31 (s, 1H, Ar-H), 5.53 (s, 2H, H-17), 5.30 (s, 2H, H-5), 5.12 (s, 2H, OOCCH 2 -N), 2.17 (dd, 2H, J=14.4, 6.9Hz, 19-CH 2 ), 0.91 (t, 3H, J=7.2Hz, 18-CH 3 ); 13 C-NMR (100 MHz, DMSO-d 6 , δppm): 167.1, 166.8, 166.4, 156.4, 152.4, 147.9, 146.1, 144.5, 139.6, 139.6, 131.5, 130.4, 129.8, 1 28.8, 128.5, 127.9, 127.7, 122.0, 122.0, 118.8, 95.4, 77.7, 66.3, 55.6, 50.2, 30.2, 7.4; HRESIMS: m / z 552.0723[M+H] + (calcd for C 27 H 21 Cl 2 N 3 O 6 , 552.0724).

[0113] Example 11, 7-ethyl-20S-camptothecin ester of 3,5-dichloro-4-pyridone-1-acetic acid (Compound 11)

[0114] In a 10 mL round-bottom flask, 15.4 mg (0.041 mmol) of 7-ethyl-camptothecin, 27.5 mg (0.124 mmol) of 3,5-dichloro-4-pyridone-1-acetic acid (Compound 4), 30.0 mg (0.157 mmol) of 1-[3-(dimethylamino)propyl]-3-ethylcarbodiimide hydrochloride (EDCI), 12.0 mg (0.098 mmol) of DMAP and 5 mL of dichloromethane were added. The reaction mixture was stirred at room temperature for 8 to 12 hours, and the reaction was monitored by TLC until the reaction was completed (clear or nearly clear). 40 ml of chloroform was added to dilute the reaction solution, and the organic phase was washed with water (35 ml) and saturated sodium bicarbonate aqueous solution (35 ml) in turn, and dried with anhydrous sodium sulfate or anhydrous magnesium sulfate. After filtering to remove the sodium sulfate or magnesium sulfate, the solvent was removed by concentration under reduced pressure. The residue was dissolved in chloroform and chromatographed on a silica gel column (eluent: chloroform-methanol 98:2 to 95:5 for gradient elution) to obtain 10.1 mg of a light yellow solid, which was 7-ethyl-20S-camptothecin ester of 3,5-dichloro-4-pyridone-1-acetic acid, i.e., compound 11, yield: 42.6%, mp>300°C (dec.).

[0115] The structural characterization data are as follows:

[0116] 1 H NMR (400 MHz, DMSO-d 6 , δppm): 8.36 (s, 2H, 2′-H&6′-H), 8.28 (d, 1H, J=8.4Hz, Ar-H), 8.19 (d, 1H, J=8.4Hz, Ar-H), 7.87 (t, 1H, J=7.2Hz , Ar-H), 7.74 (t, 1H, J=7.2Hz, Ar-H), 7.28 (s, 1H, Ar-H), 5.54 (s, 2H, H-17), 5.34 (m, 2H, H-5), 5.32 (s, 2H, OOCCH 2 -N), 3.23(dd, 2H, J=14.8, 7.2Hz, 7-CH 2 ), 2.16 (dd, 2H, J=15.2, 7.2Hz, 19-CH 2 ), 1.30 (t, 3H, J=7.6Hz, 7α-CH 3 ), 0.91 (t, 3H, J=7.2Hz, 18-CH 3 ); 13 C-NMR (100 MHz, DMSO-d 6, δppm): 167.1, 166.8, 166.4, 156.4, 151.8, 148.4, 146.7, 145.6, 144.5, 139.6, 139.6, 130.0, 129.7, 128 .0, 127.6, 126.6, 124.1, 122.0, 122.0, 118.7, 95.3, 77.7, 66.4, 55.6, 49.5, 30.1, 22.2, 13.9, 7.4; MS (ESI + ): m / z 580[M+H] + , 1181[2M+Na] + ;MS(ESI - ): m / z 1257[2M+HSO 4 ] - ;HRESIMS: m / z 580.1042[M+H] + (calcd for C 29 H 24 Cl 2 N 3 O 6 , 580.1037).

[0117] Pharmacological experiments

[0118] Experimental example: MTT human cancer cell killing test

[0119] Drugs and reagents: DMSO was of analytical grade; RPMI1640 was a product of GIBCO; calf serum was a product of the Beijing Military Region Veterinary Prevention and Treatment Center.

[0120] Instrument: BIORAD 550 microplate reader.

[0121] MTT assay: Tumor cells with good growth were collected and prepared into 1×10 -4 / mL cell suspension was inoculated into a 96-well culture plate, with 100 μL (containing 1000-1500 tumor cells) per well, and placed at 37°C with 5% CO 2 After 24 hours of incubation in the incubator, the drug was added. The experiment set up blank control and solvent control. The test sample was set to 4 concentrations, with 3 parallel wells for each concentration. The incubator was placed at 37°C and 5% CO 2 Culture in an incubator for 4 days. Discard the culture medium, add 100 μL of MTT solution (0.4 mg / mL, prepared in RPMI1640) to each well, and incubate at 37°C for 4 hours. Discard the supernatant, add 150 μL of MTT solution to each well to dissolve the particles, and after slight shaking, use a BIORAD 550 microplate reader to measure the absorbance OD value at a detection wavelength of 540 nm and a reference wavelength of 450 nm.

[0122] Result calculation: The dose-response curve can be obtained by plotting the different drug concentrations and the inhibition rate on cells, from which the half inhibition concentration (IC 50 )(μmol / L). 50 Compounds with concentrations ≤20 μM were recorded as effective compounds.

[0123] The above experiments were repeated three times for each compound sample.

[0124] The anti-tumor activity results of the derivatives of the present invention against human tumor cell lines are shown in Table 1.

[0125] Table 1. Antitumor activity results of the derivatives of the present invention against human tumor cell lines

[0126]

[0127] A549: human lung adenocarcinoma cells; Bel 7402: human liver cancer cells; BGC-823: human gastric adenocarcinoma cells; HCT-8: human colorectal adenocarcinoma cells; A2780: human ovarian cancer cells.

[0128] From the in vitro anti-tumor activity screening, most of the compounds of the present invention have a half inhibitory concentration (IC 50 ) are all lower than topotecan, and the IC 50 The cytotoxic activity of most of the compounds of the present invention is better than that of topotecan, especially compound 8 in human non-small cell lung cancer (A549) and colorectal adenocarcinoma (HCT-8) cells. 50 The lowest can reach 4×10 -9 mol / L and 7×10 -9 mol / L, and their activities are 15 and 40 times that of topotecan, respectively. The IC values ​​of some preferred compounds of the present invention, such as 5, 6, 8, 10, and 11, against A549, HCT-8, and A2780 are 50 Equivalent to or slightly better than camptothecin, IC5, 8, 11 for Bel7402 50 Comparable to camptothecin.

Claims

1. A camptothecin-20-ester derivative based on a nitrogen-containing heterocyclic-1′(N)-acetyl group and a pharmaceutically acceptable salt thereof having the following general formula (I): In the formula (I): R1 is hydrogen or C 1-8 Alkyl, C 1-8 Acyl, C 1-7 Hydroxyalkyl, C 3-6 Cycloalkyl, chloroacetyl, benzoyl, CH2NO2, CH2NO, CH2NR5R6, where R5 and R6 are H, C 1-6 Alkyl, aryl substituted alkyl, hydroxy substituted C 1-6 Alkyl, amino substituted C 1-6 Alkyl, mono- or di-substituted amino C 1-6 Alkyl or cyclic amino group; R2 is selected from hydrogen, halogen, C 1-6 Alkyl, C 1-6 Alkoxy, hydroxy, RCOO, cyano, aldehyde, nitro, amino, halo 1-6 Alkyl, carboxyl, C 1-6 Alkoxycarbonyl, C 1-6 Alkoxycarbonylamino or CH2NR7R8, wherein R7 and R8 are hydrogen, C 1-6 Alkyl, aryl substituted alkyl, hydroxy substituted C 1-6 Alkyl, amino substituted C 1-6 Alkyl, mono- or di-substituted amino C 1-6 Alkyl or cyclic amino, or R2 is selected from CH2R9, wherein R9 is C 1-6 Alkoxy or cyano, or R2 is selected from NR 10 R 11 , where R 10 , R 11 C 1-6 Alkyl, aryl, dialkylaminoalkyl, amino C 1-6 Alkyl or hydroxyl C 1-6 alkyl; R3 is hydrogen, halogen, C 1-6 Alkyl, C 1-6 Alkoxy, amino substituted C 1-6 Alkoxy, hydroxy, RCOO, cyano, aldehyde, nitro, amino, halo 1-6 Alkyl, carboxyl, C 1-6 Alkoxycarbonyl, C 1-6 Alkoxycarbonylamino, amino C 1-6 Alkyl, hydroxyl C 1-6 Alkyl or [4-(1-piperidinyl)-1-1-piperidinyl]carbonyloxy; R2 can form a ring with R3; R4 is selected from hydrogen, halogen, C 1-6 Alkyl, C 1-6 Alkoxy, RCOO, cyano, aldehyde, nitro, amino, halo C 1-6 Alkyl, carboxyl, C 1-6 Alkoxycarbonyl or C 1-6 Alkoxycarbonylamino; R3 can form a ring with R4; Features: R is selected from hydrogen, halogen, nitro, amino, C 1-6 Alkyl, C 1-6 Alkoxy, cyano or aldehyde groups. X is hydrogen or fluorine, chlorine, bromine, iodine, carboxyl, nitro, nitroso, amino, hydroxyl, C 1-8 Alkyl, C 1-8 Acyl, C 1-7 Hydroxyalkyl, C 3-6 Cycloalkyl, chloroacetyl, benzoyl, CH2NO2, CH2NO, CH2NR5R6, where R5 and R6 are H, C 1-6 Alkyl, aryl substituted alkyl, hydroxy substituted C 1-6 Alkyl, amino substituted C 1-6 Alkyl, mono- or di-substituted amino C 1-6 Alkyl or cyclic amino groups.

2. The camptothecin-20-ester derivative based on nitrogen-containing heterocyclic-1′(N)-acetyl group and its pharmaceutically acceptable salt according to claim 1, characterized in that: C 1-8 The alkyl group is selected from C 1-8 Straight or branched alkyl, C 1-8 The acyl group is selected from C 1-8 Straight chain or branched chain alkanoyl, C 1-8 Alkenyl or C 1-8 Alkynoyl, C 1-7 The hydroxyalkyl group is selected from C 1-7 Straight chain or branched chain hydroxyalkyl.

3. The camptothecin-20-ester derivative based on nitrogen-containing heterocyclic-1′(N)-acetyl group and its pharmaceutically acceptable salt according to claim 1, characterized in that: The hydroxyl group at C20 is in (S)-configuration.

4. A camptothecin-20-ester derivative based on a nitrogen-containing heterocyclic-1′(N)-acetyl group and a pharmaceutically acceptable salt thereof having the following general formula (II): In the formula (II): R1 is hydrogen or C 1-8 Alkyl, C 1-8 Acyl, C 1-7 Hydroxyalkyl, C 3-6 Cycloalkyl, chloroacetyl, benzoyl, CH2NO2, CH2NO, CH2NR5R6, where R5 and R6 are H, C 1-6 Alkyl, aryl substituted alkyl, hydroxy substituted C 1-6 Alkyl, amino substituted C 1-6 Alkyl, mono- or di-substituted amino C 1-6 Alkyl or cyclic amino group; R2 is selected from hydrogen, halogen, C 1-6 Alkyl, C 1-6 Alkoxy, hydroxy, RCOO, cyano, aldehyde, nitro, amino, halo 1-6 Alkyl, carboxyl, C 1-6 Alkoxycarbonyl, C 1-6 Alkoxycarbonylamino or CH2NR7R8, wherein R7 and R8 are hydrogen, C 1-6 Alkyl, aryl substituted alkyl, hydroxy substituted C 1-6 Alkyl, amino substituted C 1-6 Alkyl, mono- or di-substituted amino C 1-6 Alkyl or cyclic amino, or R2 is selected from CH2R9, wherein R9 is C 1-6 Alkoxy or cyano, or R2 is selected from NR 10 R 11 , where R 10 , R 11 C 1-6 Alkyl, aryl, dialkylaminoalkyl, amino C 1-6 Alkyl or hydroxyl C 1-6 alkyl; R3 is hydrogen, halogen, C 1-6 Alkyl, C 1-6 Alkoxy, amino substituted C 1-6 Alkoxy, hydroxy, RCOO, cyano, aldehyde, nitro, amino, halo 1-6 Alkyl, carboxyl, C 1-6 Alkoxycarbonyl, C 1-6 Alkoxycarbonylamino, amino C 1-6 Alkyl, hydroxyl C 1-6 Alkyl or [4-(1-piperidinyl)-1-1-piperidinyl]carbonyloxy; R2 can form a ring with R3; R4 is selected from hydrogen, halogen, C 1-6 Alkyl, C 1-6 Alkoxy, RCOO, cyano, aldehyde, nitro, amino, halo C 1-6 Alkyl, carboxyl, C 1-6 Alkoxycarbonyl or C 1-6 Alkoxycarbonylamino; R3 can form a ring with R4; Features: R is selected from hydrogen, halogen, nitro, amino, C 1-6 Alkyl, C 1-6 Alkoxy, cyano or aldehyde groups. X is hydrogen or fluorine, chlorine, bromine, iodine, carboxyl, nitro, nitroso, amino, hydroxyl, C 1-8 Alkyl, C 1-8 Acyl, C 1-7 Hydroxyalkyl, C 3-6 Cycloalkyl, chloroacetyl, benzoyl, CH2NO2, CH2NO, CH2NR5R6, where R5 and R6 are H, C 1-6 Alkyl, aryl substituted alkyl, hydroxy substituted C 1-6 Alkyl, amino substituted C 1-6 Alkyl, mono- or di-substituted amino C 1-6 Alkyl or cyclic amino groups. n is 1-4.

5. The camptothecin-20-ester derivative based on nitrogen-containing heterocyclic-1'(N)-acetyl group and its pharmaceutically acceptable salt according to any one of claims 1 or 4, characterized in that: R1C 1-8 The alkyl group is selected from methyl, ethyl, propyl, isopropyl, butyl, isobutyl, pentyl, isopentyl, hexyl, heptyl, etc.; C 1-8 The acyl group is selected from acetyl, propionyl, butyryl, isobutyryl, valeryl, isovaleryl, hexanoyl, heptanoyl, etc.; C 1-7 The hydroxyalkyl group is selected from hydroxymethyl, hydroxyethyl, hydroxypropyl, hydroxyisopropyl, hydroxybutyl, hydroxyisobutyl, hydroxypentyl, hydroxyisopentyl, hydroxyhexyl, hydroxyheptyl, etc.; C 3-6 The cycloalkyl group is selected from cyclopentyl and cyclohexyl.

6. The method for preparing a camptothecin-20-position ester derivative based on a nitrogen-containing heterocyclic-1′(N)-acetyl group according to any one of claims 1 or 4, characterized in that: The corresponding compound is prepared by condensing camptothecin in which the 20-position is a hydroxyl group or its analogue having a substituent at the 7-position with the corresponding nitrogen-containing heterocyclic-1'(N)-acetic acid or acyl halide.

7. The preparation method according to claim 6, characterized in that The condensation reagent used in the condensation reaction is selected from 1,3-dicyclohexylcarbodiimide (DCC), dipyridine carbonate (2-DPC), 1-(3-dimethylaminopropyl)-3-ethylcarbodiimide hydrochloride (EDCI), 1,3-diisopropylcarbodiimide (DIPC), and 4-pyrrolidinopyridine.

8. The preparation method according to claim 6, characterized in that The catalyst used in the condensation reaction is selected from tertiary amines.

9. The preparation method according to claim 6, characterized in that The solvent used in the condensation reaction is selected from dichloromethane or N,N-dimethylformamide, chloroform, pyridine, and dichloroethane.

10. A pharmaceutical composition, characterized in that Contains an effective dose of any one of the compounds according to claims 1 to 5, and a pharmacologically acceptable carrier.

11. The pharmaceutical composition according to claim 10, characterized in that The pharmaceutical composition can be in the form of tablets, capsules, pills, injections, sustained-release preparations, controlled-release preparations and various microparticle delivery systems.

12. Use of any compound according to claims 1 to 5 in the preparation of anti-tumor drugs.

13. Use according to claim 12, characterized in that The tumors include non-small cell lung cancer, colorectal cancer, gastric cancer, liver cancer, and ovarian cancer.

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