CN105503863A - Novel anti-tumor compound - Google Patents

Abstract

The invention provides a pyrazolopyridine derivative anti-tumor compound with excellent anti-tumor activity. The anti-tumor compound is shown as a formula I, wherein X represents -NH2, -NHOH, -NHX<1>, amino acid residue and the like (shown as the accompanying diagram); Y represents hydrogen atoms, alkyl groups, naphthenic bases, benzyl groups, aryl groups, heteroaryl, alkyl amino groups, -COX2 or -CONHX3, wherein the aryl group, the heteroaryl and the alkyl group are respectively, independently and randomly substituted by one or a plurality of materials from halogen, trifluoromethyl, amino groups, alkyl amino groups, hydroxyl, hydroxyalkyl, alkoxy, cyanogroup, nitryl, aryl groups and heteroaryl; the definitions of R1, R2, X1 and X2 are identical to the definitions in the specifications. The invention also provides a preparation method of an anti-tumor agent and application of the anti-tumor agent to lung cancer, colon cancer and ovarian cancer anti-tumor medicine.

Description

Novel antitumor compounds

Technical Field

The field belongs to the field of antitumor drugs, and particularly relates to a novel antitumor compound or a medicinal salt thereof, and a preparation method and application thereof.

Background

Tumor is still the most common and serious disease directly endangering human life in the world today, and the incidence rate is second only to cardiovascular diseases. At present, tumor chemotherapy achieves certain progress, obviously prolongs the survival time of patients, but still does not achieve satisfactory curative effect. In recent years, the research on molecular level of oncology and tumor focus parts and the discovery of a plurality of new treatment targets provide possibility for the development of novel antitumor drugs. With the continuous and deep research on the signal transduction pathway of tumor cells, the design and research of novel antitumor drugs are receiving more and more attention. The existing antitumor drugs still far fail to meet the requirements of the growing cancer patients, and the antitumor drugs are still important directions for research and development.

The invention patent CN103534254A discloses a tricyclic and tetracyclic pyrazolo [3,4-b ] pyridine compound as an antitumor agent and its pharmaceutically acceptable salts, tautomers, stereoisomers or mixtures of stereoisomers in any proportion, such as mixtures of enantiomers, especially racemic mixtures, and its preparation method and use, especially as an antitumor agent. The structure is as follows:

the invention patent CN102131807A discloses a compound useful as a protein kinase inhibitor of the formula, also provides pharmaceutically acceptable compositions comprising said compound and methods of using said compositions in the treatment of various diseases, conditions or disorders, the structure of which is shown below:

the invention patent CN101544634A discloses a drug compound 2-phenyl-3-substituted pyrazolo [1,5-A ] for treating diseases such as tumor, virus and the like]Pyridine derivatives, and a preparation method and application thereof. Wherein R is₁、R₂、R₃And R₄The definitions of (1) and (b) in the specification are the same. Also discloses a pharmaceutical composition consisting of the compound as an anti-tumor and anti-virus active ingredient I or pharmaceutically acceptable salt thereof and one or more pharmaceutically acceptable carriers, excipients or diluents. The structure is as follows:

a series of pyridopyrazole compounds with biological activity are yet to be researched, and the development of a novel pyridylpyrazole compound with excellent antitumor activity has great significance in the field of antitumor medicines.

Disclosure of Invention

The invention aims to provide a pyrazolopyridine antitumor compound or a pharmaceutically acceptable salt thereof and a preparation method thereof.

The invention also provides a pharmaceutical composition containing the antitumor compound shown in the formula I and application thereof.

The object of the invention can be achieved by the following measures:

a compound shown in formula I, or pharmaceutically acceptable salt, hydrate and isomer thereof,

wherein,

x represents-NH₂、-NHOH、-NHX¹Amino acid residues, a,

Y represents a hydrogen atom, an alkyl group, a cycloalkyl group, a benzyl group, an aryl group, a heteroaryl group, an alkylamino group or-COX²or-CONHX³Wherein aryl, heteroaryl, alkyl are each independently optionally substituted with one or more groups selected from halogen, trifluoromethyl, amino, alkylamino, hydroxy, hydroxyalkyl, alkoxy, cyano, nitro, aryl and heteroaryl;

X¹the aryl-amino-substituted aryl-substituted heteroaryl is selected from substituted or unsubstituted aryl, heteroaryl, alkyl, cycloalkyl, amino and alkylamino, and the substituent is selected from one or more of halogen, trifluoromethyl, amino, alkylamino, hydroxyl, hydroxyalkyl, alkoxy, cyano, nitro, aryl, ester group and heteroaryl;

the amino acid residue is a substitution residue formed after one hydrogen is absent in the amino group of the amino acid;

q is a substituted or unsubstituted 4-to 8-membered heterocycloalkyl group containing at least one N atom, wherein the 5-to 8-membered heterocycle contains one or more N, O, S atoms and the 4-to 8-membered heterocycle is further substituted with one or more groups selected from alkyl, aryl, heteroaryl, or cyano;

R¹、R²each independently selected from alkyl;

X²selected from the group consisting of cycloalkyl, aryl, heteroaryl, alkylamino, wherein aryl, heteroaryl, alkyl are each independently optionally substituted with one or more groups selected from the group consisting of halogen, trifluoromethyl, amino, alkylamino, hydroxy, hydroxyalkyl, alkoxy, cyano, nitro, aryl, and heteroaryl;

X³the aryl is selected from substituted or unsubstituted alkyl, cycloalkyl, aryl or heteroaryl, and the substituent is one of halogen, nitro or alkyl.

Further, Y and X in formula I¹In the groups, the aryl is independently selected from 5-6 membered monocyclic aryl, and the heteroaryl is independently selected from 5-6 membered monocyclic heteroaryl containing nitrogen or sulfur.

Further, the alkyl groups in formula I are independently selected from C_1-6An alkyl group; cycloalkyl is independently selected from C_3-8A cycloalkyl group.

Further, the aryl groups described in formula I are independently selected from substituted or unsubstituted phenyl groups.

Further, the heteroaryl groups in formula I are independently selected from substituted or unsubstituted pyridyl or thienyl.

In one scheme, Q is substituted or unsubstituted piperazinyl, and the substituent is C_1-6Alkyl, phenyl or pyridyl.

In one embodiment, the amino acid residues are: an amino acid residue formed by the amino group of the amino acid group of the carboxy-substituted or non-substituted alanine, phenylalanine, tyrosine, isoleucine, aspartic acid or valine which lacks one hydrogen, wherein the substituent is C_1-4An alkyl group.

Illustrative, non-limiting specific examples of compounds of the present invention are given below:

further, the invention also provides a preparation method of the compound in the general formula I, but not limited to the method described below. All starting materials are prepared or purchased directly according to the general rules of the target molecule and by protocols in these routes, methods well known to those of ordinary skill in the art of organic chemistry. A process for preparing a compound of formula I or a pharmaceutically acceptable salt, hydrate, or isomer thereof as shown in process one and process two:

the first method,

The method comprises the following steps:

a person skilled in the art can select the compound a as a starting material according to the structural characteristics of the target molecule of the formula I, dissolve the compound a in an organic solvent, and reduce the compound a to the compound b under the action of a reducing agent.

Step two:

dissolving the compound b in an organic solvent, and reacting with methanesulfonyl chloride under the catalysis of an organic base to generate a compound c.

Step three:

and (3) dissolving the compounds c and d in an organic solvent, completely reacting at 90-120 ℃ under the catalytic action of cesium carbonate, adding water for washing, extracting by using the organic solvent, drying by spinning, and separating and purifying by using a column to obtain a compound e.

Step four:

and (3) carrying out saponification reaction on the compound e to obtain a compound f.

Step five:

according to the structural characteristics of the target molecule in the formula I, an amine compound X-H containing amino or imino is selected to perform amidation reaction with a compound f in a solvent, and a compound g is obtained.

Step six:

and removing the BOC protecting group from the compound g to obtain a compound h.

Step seven:

according to the structural characteristics of the target molecule shown in the formula I, carboxyl, acyl chloride, halogenated hydrocarbon, halogenated aromatic hydrocarbon and methyl sulfonate compound Z (such as 4-picolinic acid, benzoyl chloride, bromoethane, 4-bromonitrobenzene, cyclopentyl methanesulfonate and the like) containing the structural characteristics corresponding to the Y group are selected as reaction raw materials to react with the compound h in a solvent to obtain the compound I.

When Y represents-CONHX³The synthetic route of the target compound is shown as follows:

reacting the compound H with triphosgene to obtain a compound H-1, and then reacting the compound H with the compound H₂N-X₃Reaction to obtain a compound I'

Method two

The method comprises the following steps:

and removing Boc protection from the compound e to obtain a compound j.

Step two:

according to the structural characteristics of the target molecule shown in the formula I, carboxyl, acyl chloride, halogenated hydrocarbon, halogenated aromatic hydrocarbon and methyl sulfonate compound Z (such as 4-picolinic acid, benzoyl chloride, bromoethane, 4-bromonitrobenzene, cyclopentyl methanesulfonate and the like) containing the structural characteristics corresponding to the Y group are selected as reaction raw materials to react with the compound e in a solvent to obtain a compound k.

Step three:

and (3) carrying out saponification reaction on the compound k to obtain a compound i.

Step four:

according to the structural characteristics of the target molecule of the formula I, an amine compound X-H containing amino or imino is selected to perform amidation reaction with a compound I in a solvent to obtain the target compound I.

In the first and second methods, the group X, the group Y and the group X³The definition of (A) is the same as that in the general formula I in the specification.

The related amidation reaction is the conventional amidation reaction in the field, and can be carried out under the action of a catalyst, wherein the catalyst is selected from 1, 3-Dicyclohexylcarbodiimide (DCC), N, N ' -Diisopropylcarbodiimide (DIC), 1- (3-dimethylaminopropyl) -3-Ethylcarbodiimide (EDC) and hydrochloride thereof, 1- (3-dimethylaminopropyl) -3-ethylcarbonyldiamine methyl iodide, N, N-Diisopropylethylamine (DIEA), 1-hydroxybenzotriazole (HoBt), 2- (7-azobenzotriazol) -N, N, N ', N ' -tetramethyluronium Hexafluorophosphate (HATU), benzotriazol-N, N, N ', N ' -tetramethyluronium Hexafluorophosphate (HBTU), 6-chlorobenzotriazole-1, 1,3, 3-tetramethyluronium Hexafluorophosphate (HCTU), 2- (1H-benzotriazol L-1-yl) -1, 1,3, 3-tetramethyluronium tetrafluoroborate (TBTU), 2-succinimidyl-1, 1,3, 3-tetramethyluronium tetrafluoroborate (TSTU), 5-norbornene-2, 3-dicarbonyl-N, N, N ', N' -tetramethyluronium tetrafluoroborate (TNTU). The ratio of the condensing agent is about 1 to 3 times. In addition, the amidation reaction may be carried out in the presence of a condensation catalyst such as diethyl azodicarboxylate/triphenylphosphine.

The invention further provides a pharmaceutical composition of the anti-tumor compound shown in the formula I, wherein the pharmaceutical composition contains a therapeutically effective amount of the compound or the pharmaceutical salt thereof as an active ingredient, and one or more pharmaceutically acceptable carriers.

The pharmaceutical composition preferably contains 0.1 to 99.5 weight percent of the anti-tumor compound or the pharmaceutically acceptable salt thereof of the invention as an active ingredient, and more preferably contains 0.5 to 99.5 weight percent of the active ingredient.

Pharmaceutical compositions containing the active ingredient of the compounds of formula I may be in a form suitable for oral administration, for example as tablets, dragees, lozenges, aqueous or oily suspensions, dispersible powders or granules, emulsions, hard or soft capsules, or syrups. Oral compositions may be prepared according to any method known in the art for preparing pharmaceutical compositions, and such compositions may contain one or more ingredients selected from the group consisting of: sweetening agents, flavoring agents, coloring agents and preserving agents in order to provide a pleasant to the eye and palatable pharmaceutical preparation. Tablets contain the active ingredient in admixture with non-toxic pharmaceutically acceptable excipients which are suitable for the manufacture of tablets. These excipients may be inert excipients, such as calcium carbonate, sodium carbonate, lactose, calcium phosphate or sodium phosphate; granulating and disintegrating agents, such as microcrystalline cellulose, croscarmellose sodium, corn starch or alginic acid; binding agents, for example starch, gelatin, polyvinylpyrrolidone or acacia, and lubricating agents, for example magnesium stearate, stearic acid or talc. These tablets may be uncoated or they may be coated by known techniques which mask the taste of the drug or delay disintegration and absorption in the gastrointestinal tract and thereby provide a sustained action over a longer period. For example, water soluble taste masking substances such as hydroxypropylmethyl cellulose or hydroxypropyl cellulose, or time extending substances such as ethyl cellulose, cellulose acetate butyrate may be used.

Oral formulations may also be presented as hard gelatin capsules wherein the active ingredient is mixed with an inert solid diluent, for example, calcium carbonate, calcium phosphate or kaolin, or as soft gelatin capsules wherein the active ingredient is mixed with a water soluble carrier, for example polyethylene glycol, or an oil vehicle, for example peanut oil, liquid paraffin or olive oil.

Aqueous suspensions contain the active materials in admixture with excipients suitable for the manufacture of aqueous suspensions. Such excipients are suspending agents, for example sodium carboxymethylcellulose, methylcellulose, hydroxypropylmethylcellulose, sodium alginate, polyvinylpyrrolidone and acacia; dispersing or wetting agents may be a naturally occurring phosphatide, for example lecithin, or condensation products of an alkylene oxide with fatty acids, for example polyoxyethylene stearate, or condensation products of ethylene oxide with long chain aliphatic alcohols, for example heptadecaethyleneoxycetanol (heptadecaethyleneoxycetanol), or condensation products of ethylene oxide with partial esters derived from fatty acids and a hexitol such as polyethylene oxide sorbitol monooleate, or condensation products of ethylene oxide with partial esters derived from fatty acids and hexitol anhydrides, for example polyethylene oxide sorbitan monooleate. The aqueous suspensions may also contain one or more preservatives, for example ethyl or n-propyl paraben, one or more colouring agents, one or more flavouring agents and one or more sweetening agents, such as sucrose, saccharin or aspartame.

Oil suspensions may be formulated by suspending the active ingredient in a vegetable oil, for example arachis oil, olive oil, sesame oil or coconut oil, or in a mineral oil such as liquid paraffin. The oil suspension may contain a thickening agent, for example beeswax, hard paraffin or cetyl alcohol. Sweetening agents such as those set forth above, and flavoring agents may be added to provide a palatable preparation. These compositions can be preserved by the addition of antioxidants such as butylated hydroxyanisole or alpha-tocopherol.

Dispersible powders and granules suitable for preparation of an aqueous suspension by the addition of water provide the active ingredient in admixture with a dispersing or wetting agent, suspending agent or one or more preservatives. Suitable dispersing or wetting agents and suspending agents are illustrative of the examples given above. Other excipients, for example sweetening, flavoring and coloring agents, may also be present. These compositions are preserved by the addition of an antioxidant such as ascorbic acid.

The pharmaceutical compositions of the invention may also be in the form of oil-in-water emulsions. The oily phase may be a vegetable oil, for example olive oil or arachis oil, or a mineral oil, for example liquid paraffin or mixtures thereof. Suitable emulsifying agents may be naturally-occurring phosphatides, for example soy bean lecithin, and esters or partial esters derived from fatty acids and hexitol anhydrides, for example sorbitan monooleate, and condensation products of the said partial esters with ethylene oxide, for example polyethylene oxide sorbitol monooleate. The emulsions may also contain sweetening agents, flavouring agents, preservatives and antioxidants. Syrups and elixirs may be formulated with sweetening agents, for example glycerol, propylene glycol, sorbitol or sucrose. Such formulations may also contain a demulcent, a preservative, a colorant and an antioxidant.

The pharmaceutical compositions may be in the form of a sterile injectable aqueous solution. Among the acceptable vehicles and solvents that may be employed are water, ringer's solution and isotonic sodium chloride solution. The sterile injectable preparation may be a sterile injectable oil-in-water microemulsion in which the active ingredient is dissolved in the oil phase. For example, the active ingredient is dissolved in a mixture of soybean oil and lecithin. The oil solution is then treated to form a microemulsion by adding to a mixture of water and glycerol. The injection solution or microemulsion may be injected into the bloodstream of a patient by local bulk injection. Alternatively, it may be desirable to administer the solutions and microemulsions in a manner that maintains a constant circulating concentration of the compounds of the present invention. To maintain such a constant concentration, a continuous intravenous delivery device may be used. An example of such a device is an intravenous pump of the model deltec add-plus.tm.5400.

The pharmaceutical compositions may be in the form of sterile injectable aqueous or oleaginous suspensions for intramuscular and subcutaneous administration. The sterile injectable preparation may also be a sterile injectable solution or suspension prepared in a non-toxic parenterally-acceptable diluent or solvent, for example as a solution in 1, 3-butanediol. In addition, sterile, fixed oils are conventionally employed as a solvent or suspending medium. For this purpose, any blend fixed oil may be used, including synthetic mono-or diglycerides. In addition, fatty acids such as oleic acid find use in the preparation of injectables.

The compounds of the present invention may be administered in the form of suppositories for rectal administration. These pharmaceutical compositions can be prepared by mixing the drug with a suitable non-irritating excipient which is solid at ordinary temperatures but liquid in the rectum and therefore will melt in the rectum to release the drug. Such materials include cocoa butter, glycerogelatin, hydrogenated vegetable oils, polyethylene glycols of various molecular weights and mixtures of fatty acid esters of polyethylene glycols.

As is well known to those skilled in the art, the dosage of a drug to be administered depends on a variety of factors, including but not limited to: the activity of the particular compound employed, the age of the patient, the weight of the patient, the health of the patient, the diet of the patient, the time of administration, the mode of administration, the rate of excretion, the combination of drugs, etc.; in addition, the optimal treatment regimen, such as the mode of treatment, the daily amount of compound I of the formula or the type of pharmaceutically acceptable salt, can be verified according to conventional treatment protocols.

Unless otherwise indicated, the following terms used in the claims and specification have the following meanings:

"alkyl" means a saturated aliphatic radical of 1 to 20 carbon atoms, including straight and branched chain radicals (a numerical range referred to herein, e.g., "1 to 20", means that the radical, in this case alkyl, may contain 1 carbon atom, 2 carbon atoms, 3 carbon atoms, etc., up to and including 20 carbon atoms). The alkyl group in the present invention includes an "alkylene group". Alkyl groups having 1 to 6 carbon atoms are referred to as lower alkyl groups. When a lower alkyl group has no substituent, it is referred to as unsubstituted lower alkyl. More preferably, the alkyl group is a medium size alkyl group having 1 to 10 carbon atoms, such as methyl, ethyl, ethylene, propyl, propylene, 2-propyl, n-butyl, isobutyl, butylene, t-butyl, pentyl, and the like. Preferably, the alkyl group is a lower alkyl group having 1 to 4 carbon atoms, such as methyl, ethyl, propyl, 2-propyl, n-butyl, butylene, isobutyl, tert-butyl, etc. Alkyl groups may be substituted or unsubstituted.

"cycloalkyl" means a saturated or partially unsaturated monocyclic or polycyclic cyclic hydrocarbon substituent comprising 3 to 20 carbon atoms, preferably 3 to 12 carbon atoms, more preferably a cycloalkyl ring comprising 3 to 10 carbon atoms, most preferably a cycloalkyl ring comprising 3 to 6 carbon atoms, most preferably cyclopropyl. Non-limiting examples of monocyclic cycloalkyl groups include cyclopropyl, cyclobutyl, cyclopentyl, cyclopentenyl, cyclohexyl, cyclohexenyl, cyclohexadienyl, cycloheptyl, cycloheptatrienyl, cyclooctyl and the like, with cyclopropyl, cyclohexenyl being preferred. Polycyclic cycloalkyl groups include spiro, fused and bridged cycloalkyl groups. Cycloalkyl groups may be optionally substituted OR unsubstituted, and when substituted, the substituents are preferably one OR more groups independently selected from alkyl, alkenyl, alkynyl, alkoxy, alkylthio, alkylamino, halogen, mercapto, hydroxy, nitro, cyano, cycloalkyl, heterocyclyl, aryl, heteroaryl, cycloalkoxy, heterocycloalkoxy, cycloalkylthio, heterocycloalkylthio, oxo, amino, haloalkyl, hydroxyalkyl, carboxylic acid group, carboxylic acid ester group, -OR₁₁、-C(O)OR₁₁、-OC(O)R₁₁、-NHS(O)_mR₁₁、-C(O)R₁₁、-NHC(O)R₁₁、-NHC(O)OR₁₁、-NR₉R₁₀、-OC(O)NR₉R₁₀or-C (O) NR₉R₁₀。

"halogen" means fluorine, chlorine, bromine or iodine, preferably fluorine, chlorine or bromine.

"nitro" means-NO₂A group.

"alkoxy" means-O- (unsubstituted alkyl) and-O- (unsubstituted cycloalkyl). Representative examples include, but are not limited to, methoxy, ethoxy, propoxy, butoxy, cyclopropoxy, cyclobutoxy, cyclopentyloxy, cyclohexyloxy, and the like.

"hydroxyalkyl" refers to an alkyl group substituted with a hydroxy group, wherein alkyl is as defined above.

"aryl" means an all-carbon monocyclic or fused polycyclic group of 6 to 12 carbon atoms having a completely conjugated pi-electron system. Non-limiting examples of aryl groups are phenyl, naphthyl and anthracenyl. The aryl ring may be fused to a heteroaryl, heterocyclyl or cycloalkyl ring, wherein the ring attached to the parent structure is an aryl ring. The aryl group may be substituted or unsubstituted. When substituted, the substituents are preferably one or more, more preferably one, two or three, even more preferably one or two, independently selected from the group consisting of lower alkyl, trihaloalkyl, halogen, hydroxy, lower alkoxy, mercapto, (lower alkyl) thio, cyano, acyl, thioacyl, O-carbamoyl, N-carbamoyl, O-thiocarbamoyl, N-thiocarbamoyl, C-acylamino, N-acylamino, nitro, N-sulphonylamino, S-sulphonylamino. Preferably, aryl is 5-membered monocyclic aryl, 6-membered monocyclic aryl.

"heteroaryl" denotes a monocyclic or fused ring group of 5 to 12 ring atoms, containing one, two, three or four ring heteroatoms selected from N, O or S, the remaining ring atoms being C, and additionally having a completely conjugated pi-electron system. The heteroaryl ring may be fused to an aryl, heterocyclyl, or cycloalkyl ring, wherein the ring joined to the parent structure is a heteroaryl ring. Heteroaryl groups may be substituted or unsubstituted. When substituted, the substituents are preferably one or more, more preferably one, two or three, and still more preferably one or two. Non-limiting examples of unsubstituted heteroaryl groups are pyrrole, furan, thiophene, imidazole, oxazole, thiazole, pyrazole, pyrimidine, quinoline, isoquinoline, purine, tetrazole, triazine, and carbazole; preferably, the heteroaryl is a sulfur-containing 5-membered monocyclic heteroaryl, a nitrogen-containing 6-membered monocyclic heteroaryl.

The amino acid residue is a substitution residue formed after one hydrogen is absent from an amino group in the amino acid, the amino acid can be one of alanine (Ala), valine (Val), leucine (Leu), isoleucine (Ile), proline (Pro), phenylalanine (Phe), tryptophan (Trp), methionine (Met), glycine (Gly), tyrosine (Tyr), serine (Ser), threonine (Thr), cysteine (Cys), asparagine (Asn), glutamine (Gln), lysine (Lys), arginine (Arg), histidine (His), aspartic acid (Asp) and glutamic acid (Glu), and the hydrogen atom on the carboxyl group can be replaced by C_1～6The amino group of the amino acid substituted by the alkyl group lacks a hydrogen to form a substituted residue.

"pharmaceutically acceptable salts" refers to those salts that retain the biological effectiveness and properties of the parent compound. Such salts include:

(1) salts with acids are formed by reaction of the free base of the parent compound with inorganic acids such as, but not limited to, hydrochloric, hydrobromic, nitric, phosphoric, metaphosphoric, sulfuric, sulfurous, and perchloric acids or organic acids such as, but not limited to, acetic, propionic, acrylic, oxalic, (D) or (L) malic, fumaric, maleic, hydroxybenzoic, γ -hydroxybutyric, methoxybenzoic, phthalic, methanesulfonic, ethanesulfonic, naphthalene-1-sulfonic, naphthalene-2-sulfonic, p-toluenesulfonic, salicylic, tartaric, citric, lactic, mandelic, succinic, or malonic acids, and the like.

"pharmaceutical composition" refers to a mixture of one or more compounds described herein or their pharmaceutically acceptable salts and prodrugs with other chemical ingredients, such as pharmaceutically acceptable carriers and excipients. The purpose of the pharmaceutical composition is to facilitate the administration of the compound to an organism.

By "pharmaceutically acceptable carrier" is meant a carrier or diluent that does not cause significant irritation to the organism and does not interfere with the biological activity and properties of the compound being administered.

"excipient" refers to an inert substance added to a pharmaceutical composition to further facilitate administration of the compound. Examples of excipients include, without limitation, calcium carbonate, calcium phosphate, various sugars and types of starch, cellulose derivatives, gelatin, vegetable oils, and polyethylene glycols.

The invention also provides application of the compound or medicinal salt, hydrate and isomer thereof in the aspect of tumor resistance, and research shows that the compound has obvious anti-tumor activity. The results of the preliminary drug activity study show that: the compound has a good inhibition effect on lung cancer, colon cancer and ovarian cancer tumor cells, and simultaneously, SD rats are used as tested animals, the LC/MS/MS method is applied to determine the drug concentration in plasma of the rats at different times after the rats are gavaged with the compound, the pharmacokinetic behavior of the compound in the rats is researched, and the result shows that the compound has a remarkable drug absorption effect, and has a good application prospect in the field of antitumor drugs, especially the fields of lung cancer, colon cancer and ovarian cancer antitumor drugs.

Detailed Description

The following examples further illustrate the invention, but are intended to be illustrative only and not limiting as to the scope of the invention.

EXAMPLE 1 preparation of Compound I-1

Adding T-160g, adding 250ml of methanol for dissolving, and slowly adding NaBH₄TLC was used to monitor the completion of the reaction of the starting material while maintaining the temperature below 10 ℃ to give about 60.5g with a 99.8% yield.

T-260g is added, dichloromethane is added for dissolving, triethylamine is added, the temperature is kept below 0 ℃, MeSO2Cl is added dropwise, and TLC monitors that the raw materials are completely reacted. Work-up gave about 80g with a yield of 96%.

13.6g of T-3 was added, dissolved in 100ml of DMF, and 1.0eq of Compound T-4 and 1.5eq of Cesium carbonate were added, the mixture was heated to 100 ℃ for reaction, and HPLC was used to monitor the completion of the reaction of the starting materials. Column chromatography separation gave about 8g of T-5, 42.37% yield.

Adding 4g of T-5, dissolving with 40ml of ethanol, adding 40ml of water and 2eq of sodium hydroxide, stirring at room temperature for 12 hours, detecting complete reaction by TLC, adding 400ml of water, adjusting the pH value to 5-6, precipitating a large amount of white solid, filtering, drying to obtain 3.2g of white solid with the yield of 84 percent

0.31g of the compound T-6 is added, 20ml of DMF is added for dissolution, 2.0eq of ammonium bicarbonate is added for reaction at 100 ℃ for 4 hours, TLC monitors the complete reaction of the raw materials, the reaction is poured into 200ml of water to precipitate white solid, and the white solid is filtered and dried to obtain about 0.25g with the yield of 80 percent.

Adding 0.25g of compound T-7, dissolving the compound T-7 in 5ml of methanol, adding 2ml of concentrated hydrochloric acid, stirring the mixture at room temperature for 3 hours, detecting that the raw materials completely react by TLC (thin layer chromatography), carrying out spin-drying, dissolving the raw materials in water, adjusting the pH value to be 8-9, extracting the raw materials by ethyl acetate, drying the raw materials by anhydrous sodium sulfate, filtering the mixture, and carrying out spin-drying to obtain 0.17g of white solid with the yield of 94%.

¹HNMR(DMSO-d6):9.03(d,J＝2Hz,1H),8.78(d,J＝2Hz,1H),5.73～5.65(br,2H),5.19～5.11(m,1H),4.87～4.82(m,2H),4.41～4.35(m,2H),3.22～3.06(m,2H),2.55(s,3H)1.95～1.90(m,2H)。

m/z＝259(M+H⁺).

EXAMPLE 2 preparation of Compound I-2

Adding 0.31g of T-6, adding 20ml of DMF to dissolve, adding 1.5eq of methylamine hydrochloride, 3eq of triethylamine and 1.5eq of HBTU, stirring at room temperature for 24 hours to react, detecting by TLC that the reaction is complete, pouring the reaction into 200ml of water, extracting by ethyl acetate, drying by anhydrous sodium sulfate, filtering, spin-drying, passing through a column to obtain 0.21g of white solid, wherein the yield is 71 percent

Adding 0.21g of T-7, dissolving with 5ml of methanol, adding 2ml of concentrated hydrochloric acid, stirring at room temperature for 3h, detecting that the raw materials completely react by TLC, carrying out spin-drying, dissolving with water, adjusting the pH value to 8-9, extracting with ethyl acetate, drying with anhydrous sodium sulfate, filtering, and spin-drying to obtain 0.12g of white solid with the yield of 80%.

¹HNMR(DMSO-d6):9.04(d,J＝2Hz,1H),8.75(d,J＝2Hz,1H),5.80～5.63(br,2H),5.16～5.07(m,1H),4.33～4.24(m,2H),3.48(s,3H),3.16～3.10(m,2H),2.69(s,3H),2.47～2.32(m,2H),2.13～2.05(m,2H)。

m/z＝274(M+H⁺).

EXAMPLE 3 preparation of Compound I-3

Adding compound T-58g, adding 40ml of absolute ethyl alcohol, adding 16ml of concentrated hydrochloric acid, stirring at room temperature for 6h, detecting that raw materials are completely reacted by TLC, carrying out spin-drying, adding water for dissolving, adjusting the pH value to 8-9, extracting by ethyl acetate, drying by anhydrous sodium sulfate, filtering, and spin-drying to obtain 5.4g of white solid with the yield of 85%.

C-12.9g was added, dissolved in 30ml of DMSO, and then 1.5eq of potassium carbonate, 0.1eq of potassium iodide, 1.2eq of ethyl bromide and 0.1eq of tetrabutylammonium bromide were added, and after reaction with stirring at 60 ℃, about 1.9g was obtained by post-treatment with a yield of 60.1%.

Adding 1.5g of C-2, dissolving with 20ml of ethanol, adding 20ml of water and 2eq of sodium hydroxide, stirring at room temperature for 12 hours, detecting by TLC to complete the reaction, adding 200ml of water, adjusting the pH value to 5-6, precipitating a large amount of white solid, filtering, and drying to obtain 0.9g of white solid. 0.4g of C-3 is added, 10ml of DMF is added for dissolution, 2.0eq of ammonium bicarbonate is added for reaction at 100 ℃ for 4 hours, the reaction is poured into 200ml of water, white solid is separated out, and the white solid is filtered and dried to obtain about 0.25g, and the yield is 62%.

¹HNMR(DMSO-d6):9.04(d,J＝2Hz,1H),8.75(d,J＝2Hz,1H),5.55～5.39(br,2H),5.07(m,1H),3.18～3.11(m,2H),2.62～2.55(m,4H),2.35～2.19(m,5H),1.97～1.92(m,2H),1.14～1.06(m,3H).

m/z＝288(M+H⁺).

EXAMPLE 4 preparation of Compound I-4

0.4g of C-3 was added, 10ml of DCM was added and dissolved, 0.3ml of oxalyl chloride was added at room temperature, 1 drop of DMF was added, reflux reaction was carried out for 1 hour, concentration was carried out, the residue was dissolved in 10ml of THF, 1.0eq of hydroxylamine solution was added and reaction was carried out at room temperature for 2 hours, the reaction was poured into 200ml of water, extraction was carried out with ethyl acetate, drying was carried out with anhydrous sodium sulfate, filtration, spin-drying and column-chromatography gave about 0.21g, yield 50%.

¹HNMR(DMSO-d6):9.03(d,J＝2Hz,1H),8.74(d,J＝2Hz,1H),5.55(br,1H),5.39(br,1H),5.01(m,1H),3.18～3.11(m,2H),2.62～2.56(m,4H),2.35～2.19(m,5H),1.97～1.92(m,2H),1.14～1.06(m,3H)。

m/z＝304(M+H⁺).

EXAMPLE 5 preparation of Compound I-5

I-50.29g is put into the reactor, dissolved in 10ml of DMF, added with 1.5eq of triethylamine, 1.0eq of benzoic acid and 1.5eq of HBTU, stirred at room temperature for 12h, TLC detects the completion of the reaction of the raw materials, the reaction solution is poured into water, extracted with ethyl acetate, dried over anhydrous sodium sulfate, stirred and passed through the column to obtain 0.21g of off-white solid with a yield of 52%.

¹HNMR(DMSO-d6):9.02(d,J＝2Hz,1H),8.75(d,J＝2Hz,1H),7.68～7.66(m,2H),7.40～7.36(m,2H),5.83(br,1H),5.61(br,1H),5.16～5.07(m,1H),4.33～4.24(m,2H),3.40～3.28(s,3H),3.16～3.10(m,2H),2.79(s,3H),2.77～2.32(m,2H),2.13～2.05(m,2H)。

m/z＝378(M+H⁺).

Examples 6-50 were prepared according to the above scheme and the process for preparation of the compounds was similar to that described in the examples above. The following table gives the test data for the experimental target products of each example.

TABLE 1 EXAMPLES 6 to 51 List

EXAMPLE 51 preparation of Compound I-51

Adding raw materials I-60.29g, adding 10ml DCM, stirring at 0 ℃ for 10min, then dropping 1.5eq of triphosgene dissolved in DCM, stirring at low temperature for 3h, detecting by TLC to complete the reaction, washing the organic phase for 2 times, drying the organic phase with anhydrous sodium sulfate, filtering, and spin-drying to obtain 0.25g of off-white solid.

0.25ga is added and dissolved by 10ml dioxane, 1.0eq aniline and 1.5eq triethylamine are added, the mixture is placed at room temperature and stirred for 36h, the TLC detection reaction is complete, and the mixture is directly stirred and passed through a column to obtain 0.15g pale yellow solid.

¹HNMR(DMSO-d6):9.57(br,1H),9.46(br,1H),9.04(d,J＝2Hz,1H),8.69(d,J＝2Hz,1H),8.21～7.85(m,4H),7.55～7.08(m,5H),5.06(m,1H),3.37～3.26(m,2H),2.82～2.73(m,2H),2.59(s,3H),2.41～2.27(m,2H),2.12～1.99(m,2H).

m/z＝500(M+H⁺).

And (3) testing the pharmacological activity:

pharmacological tests prove that the compounds of the invention all have excellent antitumor activity. Therefore, the compound can be used for preparing the medicine for treating tumor diseases. The following are the pharmacodynamic tests and results of some of the compounds of the invention. The structural formula of the compound is shown in an example.

Growth inhibition of compounds on multiple human tumor cells

The compound in the specification of the invention is used for detecting the cytotoxicity test of 6 tumor cell strains (A549, NCI-H1299, NCI-H460, HCT-116, HT-29& SK-OV-3) by using a CCK-8(CellCountingkit) detection kit.

1. Cell lines:

a549 human non-small cell lung cancer cell line (ordered in Shanghai cell resource center of Chinese academy of sciences, Cat # TCTU 150)

NCI-H1299 human non-small cell lung cancer cell line (ordered in Shanghai cell resource center of Chinese academy of sciences, Cat # TCTU 160)

NCI-H460 human Large cell Lung cancer cell line (ordered in Shanghai cell resource center of Chinese academy of sciences, Cat # TCHU205)

HCT-116 human colon cancer cell line (ordered in Shanghai cell resource center of Chinese academy of sciences, Cat # TCTU 99)

HT-29 human colon cancer cell line (ordered in Shanghai cell resource center of Chinese academy of sciences, Cat # TCTU 103)

SK-OV-3 human ovarian cancer cell line (order from Shanghai cell resource center of Chinese academy of sciences, Cat # TCTU 185)

2. Reagents and consumables:

CellCountingKit-8(Cat#CK04-13，Dojindo)

96-well culture plate (Cat #3599, Corning Costar)

Culture medium and fetal bovine serum (GIBCO)

Desk type microplate reader SpectraMaxM5MicroplateReader (molecular devices)

Test Compounds the compounds of examples 1-32 were each tested.

3. Preparation of culture Medium

TABLE 2

Cell lines	Culture medium
		A549	RPMI 1640+10％FBS
NCI-H1299	RPMI 1640+10％FBS
		NCI-H460	RPMI1640+10％FBS
HCT-116	DMEM+10％FBS
		HT-29	Mccoy’s 5A+10％FBS
SK-OV-3	Mccoy’s 5A+10％FBS

IC50 experiment (CCK-8 test)

a) Cells in the logarithmic growth phase were collected, counted, resuspended in complete medium, adjusted to the appropriate concentration (as determined by the cell density optimization assay) and seeded into 96-well plates with 100. mu.l of cell suspension per well. Cells were incubated at 37 ℃ and 100% relative humidity, 5% CO₂Incubate in incubator for 24 hours.

b) The test compound was diluted with the medium to the set corresponding effect concentration and the cells were added at 25. mu.l/well. Dissolving in dimethyl sulfoxide according to the concentration required by the experiment to prepare a compound DMSO solution, wherein the final action concentration of the compound is 100 μ M, 25 μ M, 6.25 μ M, 1.5625 μ M, 0.390625 μ M, 0.097656 μ M, 0.024414 μ M, 0.006104 μ M and 0.001526 μ M;

c) cells were incubated at 37 ℃ and 100% relative humidity, 5% CO₂Incubate in incubator for 72 hours.

d) The medium was aspirated off, complete medium containing 10% CCK-8 was added and incubated in an incubator at 37 ℃ for 1-4 hours.

e) After gentle shaking, the absorbance at 450nm was measured on a SpectraMaxM5MicroplateReader, and the inhibition was calculated using the absorbance at 650nm as a reference.

5. Data processing

The inhibition rate of the drug on the growth of tumor cells was calculated according to the following formula: the inhibition rate of tumor cell growth [ (% A)_c-A_s)/(A_c-A_b)]×100％

A_sOA of the sample (cell + CCK-8+ test Compound)

A_cOA of negative control (cell + CCK-8+ DMSO)

A_bOA of positive control (Medium + CCK-8+ DMSO)

IC50 curve fits and IC50 values were calculated using software GraphpadPrism5 and using the calculation formula log (inhibitor) vs. normalizedrpossponse, as shown in the following table:

TABLE 3 IC of Compounds for various human tumor cell lines₅₀Value of

As a result: the compound of the invention has obvious inhibition effect on the growth of various human tumor cell strains.

Pharmacokinetic evaluation

SD rats are used as test animals, the drug concentrations in blood plasma of the rats at different times after the rats are administered with the compound I-2, the compound I-22, the compound I-36 and the compound I-50 by gavage are measured by an LC/MS/MS method, the pharmacokinetic behavior of the compound in the rats is researched, and the pharmacokinetic characteristics of the compound are evaluated.

The experimental scheme is as follows:

1. dose selection

The dose administered was 5 mg/kg.

2. Test animal

The test rats were given an environmental acclimation period of 2 days prior to the experiment.

24 SD rats were obtained, half male and half female, and the body weight was about 200 g. The groups of male/female rats were randomly divided into 4 test groups (6 per group, half male/female) based on average body weight.

3. Plasma sample processing

Collecting rat plasma 50 μ l, adding 200 μ l protein precipitant, shaking for 3min to precipitate protein, centrifuging at 20000rcf for 10min, transferring 80 μ l supernatant to sample injection bottle, and analyzing 2 μ l sample injection.

4. Procedure of experiment

Rats were fasted for 12h and 5mg/kg of Compound I-1, Compound I-11, Compound I-12, Compound I-20 and Olaparib were administered sequentially gavage (i.g.). After administration to rats, about 200. mu.l of venous blood from rats before (0) and 15min, 30min, 45min, 1h, 2h, 4h, 6h, 8h, 12h and 24h after administration were collected from orbital venous plexus into blood collection tubes (blood collection tubes were anticoagulated with 0.5% heparin sodium in advance), supernatant plasma was transferred after centrifugation at 4000rpm for 5min, plasma samples were pretreated with precipitated protein and analyzed to determine the chronological concentration of the compound in the plasma.

5. Pharmacokinetic parameter results

After 5mg/kg of the compound is administrated to SD rats by single intragastric administration, the pharmacokinetics parameters of the compound in SD rats are calculated by actually measuring the blood concentration of the drug in vivo by using DAS software, and the results are respectively listed in tables 3-7.

TABLE 4 pharmacokinetic parameters of Compound I-2 in rats (fasting)

TABLE 5 pharmacokinetic parameters of Compounds I-22 in rats (fasted)

TABLE 6 pharmacokinetic parameters of Compounds I-36 in rats (fasted)

TABLE 7 pharmacokinetic parameters of Compounds I-50 in rats (fasted)

And (4) conclusion: the compound of the invention has good drug absorption and obvious drug absorption effect.

Claims (10)

1. A compound having a structure represented by formula I and pharmaceutically acceptable salts, hydrates, isomers thereof:

wherein,

x represents-NH₂、-NHOH、-NHX¹Amino acid residues, a,

Y represents a hydrogen atom, an alkyl group, a cycloalkyl group, a benzyl group, an aryl group, a heteroaryl group, an alkylamino group or-COX²or-CONHX³Wherein aryl, heteroaryl, alkyl are optionally independently substituted with one or more substituents selected from the group consisting of halogen, trifluoromethyl, amino, alkylamino, hydroxy, hydroxyalkyl, alkoxy, cyano, nitro, aryl, and heteroaryl;

X¹the aryl-amino-substituted aryl-substituted heteroaryl group is selected from substituted or unsubstituted aryl, heteroaryl, alkyl, cycloalkyl, amino and alkylamino, and the substituent is independently selected from one or more of halogen, trifluoromethyl, amino, alkylamino, hydroxyl, hydroxyalkyl, alkoxy, cyano, nitro, aryl, ester group and heteroaryl;

the amino acid residue is a substitution residue formed after one hydrogen is absent in the amino group of the amino acid;

q is a substituted or unsubstituted 4-to 8-membered heterocycloalkyl group containing at least one N atom, wherein the 5-to 8-membered heterocycle contains one or more N, O, S atoms and the 4-to 8-membered heterocycle is further substituted with one or more groups selected from alkyl, aryl, heteroaryl, or cyano;

R¹、R²each independently selected from alkyl;

X²selected from the group consisting of cycloalkyl, aryl, heteroaryl, alkylamino, wherein aryl, heteroaryl, alkyl are optionally independently substituted with one or more groups selected from the group consisting of halogen, trifluoromethyl, amino, alkylamino, hydroxy, hydroxyalkyl, alkoxy, cyano, nitro, aryl, and heteroaryl;

X³the aryl is selected from substituted or unsubstituted alkyl, cycloalkyl, aryl or heteroaryl, and the substituent is one of halogen, nitro or alkyl.

2. The compound of claim 1, and pharmaceutically acceptable salts, hydrates, isomers thereof, wherein Y and X are¹In the group, the aryl is independently selected from 5-6 membered monocyclic aryl, and the heteroaryl is selected from 5-6 membered monocyclic heteroaryl containing nitrogen or S.

3. The compound of claim 1, wherein said alkyl is independently selected from the group consisting of C, and pharmaceutically acceptable salts, hydrates, and isomers thereof_1-6An alkyl group; cycloalkyl is independently selected from C_3-8A cycloalkyl group.

4. The compound of claim 1, wherein said aryl is independently selected from the group consisting of substituted and unsubstituted phenyl, and pharmaceutically acceptable salts, hydrates, and isomers thereof.

5. The compound of claim 1, wherein the heteroaryl group is independently selected from the group consisting of substituted or unsubstituted pyridyl or thienyl, and pharmaceutically acceptable salts, hydrates, and isomers thereof.

6. The compound of claim 1, wherein Q is substituted or unsubstituted piperazinyl and said substituent is C, and pharmaceutically acceptable salts, hydrates, and isomers thereof_1-6Alkyl, phenyl or pyridyl.

7. The compound of claim 1, and pharmaceutically acceptable salts, hydrates, isomers thereof, wherein the amino acid residues are: an amino acid residue formed by the amino group of the amino acid group of the carboxy-substituted or non-substituted alanine, phenylalanine, tyrosine, isoleucine, aspartic acid or valine which lacks one hydrogen, wherein the substituent is C_1-4An alkyl group.

8. The compound according to any one of claims 1 to 7, wherein the compound is selected from the group consisting of:

9. a pharmaceutical composition comprising as active ingredient a therapeutically effective amount of a compound as defined in any one of claims 1 to 8 in free form or in pharmaceutically acceptable salt form; one or more pharmaceutically acceptable carrier substances and/or diluents.

10. A pharmaceutical composition comprising a compound according to claim 1 for use as an antitumor agent in lung, colon and ovarian cancer.