TW201332946A - Benzamide derivatives with anti-cancer activity and preparation method and use thereof - Google Patents

Abstract

This invention relates to benzamide derivatives with anti-cancer activity. This invention also relates to the preparation of these compounds and their use in preparing anti-cancer drugs. In certain aspects, this invention is embodied by the poly(ADP-ribose) polymerase (PARP) inhibitors or their metabolites represented by compounds of formula (I), or pharmaceutically acceptable salts or prodrugs thereof, wherein R1, R2, R3 and R4 are defined in the specification.

Description

Benzylamine derivative having anticancer activity, preparation method and use thereof

The present invention relates to a benzamide derivative having an anticancer activity and a metabolite thereof. The invention also relates to the synthesis of such anticancer drugs and methods of using the compounds to treat cancer.

Cancer is undoubtedly a difficult and life-threatening disease, a disease that is common worldwide, and is therefore a serious global health problem. In the United States, among Americans under the age of 85, cancer is the leading cause of death; among older Americans, cancer is the second leading cause of death. According to statistics, about 1,500 people die of cancer every day in the United States, and there are about 3,400 new cancer cases every day. In China, about 1.8 million people die of cancer every year, and there are about 2.6 million new cancer cases every year. Cancer has become the number one cause of death for Chinese people.

Cancer is a disease that begins in the body's cells and is a malignant tumor. This tumor cell is extremely abnormal and undergoes random and disordered division, so its growth and proliferation are completely out of control. Cancer cells are highly aggressive, attacking and damaging surrounding tissues. They can also leave the original tumor and enter the blood or lymphatic system, creating new tumors in other parts of the body.

Although there are already many anticancer drugs on the market, the task is still very difficult to effectively treat cancer. Therefore, it is still a top priority to continue to develop anticancer drugs with higher activity and less side effects. Interestingly, halogenated nitro and nitrosoestrogamine compounds can be used to treat cancer, especially breast cancer; more interestingly, the structurally simple aromatic halogenated nitro and nitroso compounds have also been found to be very high. Antitumor activity, especially for breast cancer (Kun et al. US 5464871). This class of compounds is a poly(ADP-ribose) polymerase (PARP) inhibitor, which is involved in the repair of DNA damage. Therefore, inhibition of poly(ADP-ribose) polymerase can inhibit DNA repair, thereby enhancing the therapeutic effect of radiation and chemotherapy on cancer (Ossovskaya et al. US 20090149397; Sherman et al. US 20090131529 and 20090123419). In fact, aromatic nitro compounds are first converted to aromatic nitroso compounds in the body, while the latter are active compounds that inhibit tumor growth. Since aromatic nitroso compounds have relatively poor water solubility at physiological pH and limited stability, it is difficult to predict whether these compounds can reach cancer cells, and aromatic nitro compounds do not have these problems, so they are aromatic nitroso compounds. The ideal prodrug. Among these aromatic nitro compounds, 4-iodo-3-nitrobenzamide (INBA, code BSI-201) is a very promising anticancer drug. Currently, the compound has entered Phase III clinical trial and is being further evaluated for its anticancer effect in combination with other anti-tumor drugs. The cancer treated is a triple-negative breast cancer (mTNBC) that has spread.

Since organic iodides are sensitive to light and air and poor in water solubility, it has been expected to develop anticancer drugs with higher activity, better water solubility and no iodine. The present invention provides a novel benzamide derivative having anticancer activity, which is a class of iodine-free, more effective anticancer drug than INBA (BSI-201).

One of the objects of the present invention is to disclose a new class of anticancer drug-benzamide derivatives or pharmaceutically acceptable salts thereof.

The compounds of the present invention can be represented by formula ( I ):

Wherein: R ¹ and R ^{2 are} independently selected from the group consisting of hydrogen, substituted or unsubstituted (C ₁ -C ₈ )alkyl, substituted or unsubstituted (C ₃ -C ₈ )alkenyl, substituted or unsubstituted Substituted (C ₃ -C ₈ )alkynyl, substituted or unsubstituted (C ₃ -C ₇ )cycloalkyl, -COR ⁵ and -CO ₂ R ⁶ ; R ¹ and R ² may also be cyclized to form a substitution Or an unsubstituted four, five or six membered ring; R ³ is a nitro or nitroso group; R ^{4 is} selected from the group consisting of ethynyl, propynyl or cyano; and R ^{5 is} selected from the group consisting of: Unsubstituted (C ₁ -C ₈ )alkyl, substituted or unsubstituted (C ₃ -C ₈ )alkenyl, substituted or unsubstituted (C ₃ -C ₈ )alkynyl, substituted or unsubstituted (C _3- C ₇ ) cycloalkyl, substituted or unsubstituted aryl; R ^{6 is} selected from the group consisting of substituted or unsubstituted (C ₁ -C ₈ )alkyl, substituted or unsubstituted (C ₃ -C ₈ ) an alkenyl group, a substituted or unsubstituted (C ₃ -C ₈ ) alkynyl group, a substituted or unsubstituted (C ₃ -C ₇ )cycloalkyl group; wherein the above substituted or unsubstituted (C ₃ -C ₈ ) alkenyl group, a substituted or unsubstituted (C ₃ -C ₈₎ alkynyl group is preferably a double bond and a triple bond alkynyl group, an alkenyl group does not Amides of N, carbonyl or carbonyloxy directly Even.

Or a pharmaceutically acceptable salt thereof, or a prodrug thereof.

As used herein, "substituted", unless otherwise specified, may be substituted with a group selected from the group consisting of hydroxy, (C ₁ -C ₈ )alkyl, (C ₂ -C ₈ )alkenyl, ( C ₂ -C ₈ )alkynyl, (C ₃ -C ₇ )cycloalkyl, (C ₁ -C ₈ )alkoxy, amino, (C ₁ -C ₈ )alkylamino, di(C ₁ - C ₈ ) alkylamino group, (C ₁ -C ₈ )alkylthio group, halogen, preferably hydroxy group, methoxy group, amino group, methylamino group, dimethylamino group, methylthio group and halogen.

Wherein R ¹ and R ² above are preferably hydrogen, substituted or unsubstituted (C ₁ -C ₈ )alkyl, -COR ⁵ or -CO ₂ R ⁶ , and R ¹ and R ² may also be cyclized to form a substitution or not. a substituted four, five or six membered ring wherein R ⁵ is substituted or unsubstituted (C ₁ -C ₈ )alkyl, substituted or unsubstituted (C ₃ -C ₈ )alkenyl, substituted or unsubstituted ( C ₃ -C ₈ ) alkynyl, substituted or unsubstituted (C ₃ -C ₇ )cycloalkyl, substituted or unsubstituted aryl; R ⁶ is substituted or unsubstituted (C ₁ -C ₈ )alkyl , substituted or unsubstituted (C ₃ -C ₈ )alkenyl, substituted or unsubstituted (C ₃ -C ₈ )alkynyl, substituted or unsubstituted (C ₃ -C ₇ )cycloalkyl; The substituted or unsubstituted (C ₃ -C ₈ )alkenyl group, substituted or unsubstituted (C ₃ -C ₈ ) alkynyl group is preferably a double bond of an alkenyl group and a triple bond of an alkynyl group which is not bonded to a guanamine N or a carbonyl group. Or the carbonyloxy group is directly linked.

More preferably, R ¹ and R ^{2 are} independently selected from hydrogen, substituted or unsubstituted (C ₁ -C ₈ )alkyl, -COR ⁵ or -CO ₂ R ⁶ , and R ¹ and R ² may also be cyclized. Forming a substituted or unsubstituted tetra, five or six membered ring wherein R ⁵ is a substituted or unsubstituted (C ₁ -C ₈ )alkyl group, a substituted or unsubstituted (C ₃ -C ₇ )cycloalkyl group, a substituted or unsubstituted aryl group; R ⁶ is a substituted or unsubstituted (C ₁ -C ₈ )alkyl group, a substituted or unsubstituted (C ₃ -C ₇ )cycloalkyl group.

Most preferably, R ¹ and R ^{2 are} independently selected from hydrogen, methyl, ethyl, propyl, -COR ⁵ or -CO ₂ R ⁶ , and R ¹ and R ² may also be cyclized to form a substituted or unsubstituted a five- or six-membered ring, and the atom forming the ring may contain one or more heteroatoms, the heteroatom is a nitrogen, oxygen or sulfur atom, preferably a nitrogen atom; R ¹ and R ² are taken together to form a substituted or unsubstituted A five or six membered ring containing one or more heteroatoms, preferably a pyrrolidine ring, a piperidine ring, a morpholine ring, or a piperazine ring.

Wherein the above R ⁵ is preferably a substituted or unsubstituted (C ₁ -C ₈ )alkyl group, a substituted or unsubstituted aryl group, and most preferably a methyl group or a phenyl group.

Wherein R ⁶ is preferably substituted or unsubstituted (C ₁ -C ₈ )alkyl, more preferably methyl, ethyl, propyl, isopropyl, n-butyl or isobutyl, most preferably methyl, ethyl base.

Wherein R ³ is preferably a nitro group or a nitroso group, more preferably a nitro group.

Wherein R ⁴ is preferably an ethynyl group, a propynyl group or a cyano group, and more preferably an ethynyl group or a cyano group.

More specifically, the compound ( I ) wherein the compound is selected from the group consisting of:

4-ethynyl-3-nitrobenzamide ( I-1 )

N-methoxycarbonyl-4-ethynyl-3-nitrobenzamide ( I-2 )

N-ethoxycarbonyl-4-ethynyl-3-nitrobenzamide ( I-3 )

N-propoxycarbonyl-4-ethynyl-3-nitrobenzamide ( I-4 )

N-butoxycarbonyl-4-ethynyl-3-nitrobenzamide ( I-5 )

N-isopropyloxycarbonyl-4-ethynyl-3-nitrobenzamide ( I-6 )

N-isobutoxycarbonyl-4-ethynyl-3-nitrobenzamide ( I-7 )

4-cyano-3-nitrobenzamide ( I-8 )

N-methyl-4-ethynyl-3-nitrobenzamide ( I-9 )

N,N-Dimethyl-4-ethynyl-3-nitrobenzamide ( I-10 )

N-Ethyl-4-ethynyl-3-nitrobenzamide ( I-11 )

4-(1-propynyl)-3-nitrobenzamide ( I-12 )

N-benzylidene-4-ethynyl-3-nitrobenzamide ( I-13 )

N,N-Diethyl-4-ethynyl-3-nitrobenzamide ( I-14 )

N,N-dipropyl-4-ethynyl-3-nitrobenzamide ( I-15 )

N,N-dibutyl-4-ethynyl-3-nitrobenzamide ( I-16 )

N-ethyl-4-ethynyl-3-nitrobenzamide ( I-17 )

N-propyl-4-ethynyl-3-nitrobenzamide ( I-18 )

N-butyl-4-ethynyl-3-nitrobenzamide ( I-19 )

(4-ethynyl-3-nitrophenyl)(pyrrolidin-1-yl)methanone ( I-20 )

(4-ethynyl-3-nitrophenyl)(piperidin-1-yl)methanone ( I-21 )

(4-ethynyl-3-nitrophenyl)(morpholin-4-yl)methanone ( I-22 )

(4-ethynyl-3-nitrophenyl)(piperazin-1-yl)methanone ( I-23 )

(4-ethynyl-3-nitrophenyl)(4-methylpiperazin-1-yl)methanone ( I-24 )

(4-ethynyl-3-nitrophenyl)(azetidin-1-yl)methanone ( I-25 )

N-methyl-4-(1-propynyl)-3-nitrobenzamide ( I-26 )

N-methyl-4-cyano-3-nitrobenzamide ( I-27 )

4-ethynyl-3-nitrosobenzamide ( I-28 )

(4-ethynyl-3-nitrophenyl)(4-methylpiperazin-1-yl)methanone hydrochloride ( I-29 )

(4-ethynyl-3-nitrophenyl)(piperazin-1-yl)methanone hydrochloride ( I-30 ).

A second aspect of the invention also discloses the use of a compound of formula ( I ) and a pharmaceutical composition thereof for the manufacture of a medicament for the treatment of cancer. The cancer includes colon cancer, breast cancer, lung cancer, ovarian cancer, gastric cancer, acute leukemia, chronic leukemia, prostate cancer, uterine cancer, pancreatic cancer, liver cancer, brain cancer, CNS tumor, bladder cancer, kidney cancer, skin cancer, neck Cancer, myoma, lymphoma, bone cancer, and other types of cancer. Also disclosed is the use of a compound represented by the general formula ( I ) and a pharmaceutical composition for the preparation of a medicament for treating a condition caused, associated or concomitant with the destruction of cell proliferation and/or angiogenesis.

A further aspect of the invention resides in the use of a compound of formula ( I ) for the manufacture of a medicament for the treatment of a disease associated with a poly(ADP-ribose) polymerase (PARP) inhibitor. Diseases associated with poly(ADP-ribose) polymerase (PARP) inhibitors include cancer, Stroke, Myocardial infarction, Neurodegenerative Disease, and the like. Another aspect of the present invention provides a compound represented by the general formula (I) or the general formula (I) an effective amount of the compound represented by the pharmaceutical compositions alone or in combination for the treatment of cell proliferation and / or with other agents A method of causing, correlating, or accompanying a disorder of angiogenesis. Pharmaceutically acceptable carriers compatible with the compounds of formula ( I ) may also be included in the pharmaceutical compositions of the invention. The compound of the formula ( I ) can be administered in a usual dosage form, such as an injection form and an oral form, including a capsule, a tablet, a powder, a cachet, a suspension or a solution, preferably by injection. The dosage form and pharmaceutical compositions can be prepared using conventional pharmaceutically acceptable excipients and additives, as well as conventional techniques. The pharmaceutically acceptable excipients and additives include non-toxic compatible fillers, binders, disintegrants, buffers, preservatives, antioxidants, lubricants, flavoring agents, thickeners, colorants , emulsifiers, etc.

Another aspect of the present invention is to disclose a process for producing the benzamide derivative.

It is still another object of the present invention to provide a process for preparing a compound of the formula ( I ) (shown as the following Reaction Scheme 1). This method is applicable to the case where R ¹ , R ² are hydrogen, R ³ is a nitro group, and R ⁴ is an ethynyl group, a propynyl group or a cyano group in the compound of the formula ( I ), as used in the preparation of the compound I-1 , a compound. I-8 and compound I-12 .

Still another object of the present invention is to provide a process for preparing a compound of the formula ( I ) using Compound IX as a starting material (as shown in the following Reaction Scheme 2). This method is applicable to the compound of formula ( I ) wherein R ¹ is hydrogen, R ² is -COR ⁵ or -CO ₂ R ⁶ ; R ³ is nitro, R ⁴ is ethynyl, propynyl or cyano, R ⁵ and R ⁶ as defined above, as used in the preparation of compound I-2 , compound I-3 , compound I-4 , compound I- 5 , compound I-6 , compound I-7 , compound I-11 and compound I -13 .

A further object of the present invention is to provide a process for preparing a compound of the formula ( I ) (shown in the following Reaction Scheme 3). This method is applicable to the compound of formula ( I ) wherein R ¹ or R ² is hydrogen, substituted or unsubstituted (C ₁ -C ₈ )alkyl, substituted or unsubstituted (C ₃ -C ₈ )alkenyl, substituted or Unsubstituted (C ₃ -C ₈ )alkynyl, substituted or unsubstituted (C ₃ -C ₇ )cycloalkyl, R ¹ and R ² may also be cyclized to form a substituted or unsubstituted four, five or six a ring, but R ¹ , R ² cannot be hydrogen at the same time; R ³ is a nitro group, and when R ⁴ is an ethynyl group, a propynyl group or a cyano group, as used in the preparation of the compound I-9 , the compound I-10 , Compound I-14 , Compound I-15 , Compound I-16 , Compound I-17 , Compound I-18 , Compound I-19 , Compound I-20 , Compound I-21 , Compound I-22 , Compound I-23 , Compound I-24 , Compound I-25 , Compound I-26 and Compound I-27 .

A further object of the present invention is to provide a process for preparing a compound of the formula ( I ) (shown as the following Reaction Scheme 4). This method is applicable to the compounds of formula ( I ) wherein R ¹ and R ² are hydrogen, substituted or unsubstituted (C ₁ -C ₈ )alkyl, substituted or unsubstituted (C ₃ -C ₈ )alkenyl, substituted or Unsubstituted (C ₃ -C ₈ )alkynyl, substituted or unsubstituted (C ₃ -C ₇ )cycloalkyl, -COR ⁵ and -CO ₂ R ⁶ ; R ¹ and R ² may also be cyclized to form a substituted or unsubstituted tetra, five or six membered ring; R ³ is a nitroso group; R ⁴ is ethynyl, propynyl or cyano; and R ⁵ is substituted or unsubstituted (C ₁ -C ₈ )alkyl , substituted or unsubstituted (C ₃ -C ₈ )alkenyl, substituted or unsubstituted (C ₃ -C ₈ )alkynyl, substituted or unsubstituted (C ₃ -C ₇ )cycloalkyl, substituted or not Substituted aryl; R ⁶ is substituted or unsubstituted (C ₁ -C ₈ )alkyl, substituted or unsubstituted (C ₃ -C ₈ )alkenyl, substituted or unsubstituted (C ₃ -C ₈ ) In the case of alkynyl, substituted or unsubstituted (C ₃ -C ₇ )cycloalkyl, as used in the preparation of compound I-28 .

The present invention also provides a method for forming a salt of the compound of the formula ( I ) with an acid: the compound of the invention (I) is thoroughly mixed with a corresponding acid (for example, hydrochloric acid, sulfuric acid, etc.), and post-treated to obtain a corresponding salt. For example, compounds I-29 and I-30 were prepared .

Some of the terms used in the present invention are defined as follows:

"Halogen" means fluoro, chloro, bromo and iodo, preferably chloro, bromo and iodo.

"Alkyl" as a group or part of a group refers to a straight or branched aliphatic hydrocarbon group. Preference is given to C ₁ -C ₈ alkyl groups. Examples of alkyl groups include, but are not limited to, methyl, ethyl, n-propyl, 2-propyl, n-butyl, isobutyl, t-butyl, hexyl, and the like.

"Alkenyl" as a group or part of a group refers to an aliphatic hydrocarbon group containing a carbon-carbon double bond, either straight or branched. Preference is given to C ₃ -C ₈ alkenyl groups. Examples of alkenyl groups include, but are not limited to, allyl, 2-butenyl, and the like.

"Alkynyl" as a group or part of a group refers to an aliphatic hydrocarbon group containing a carbon-carbon triple bond, which may be straight or branched. Preference is given to C ₃ -C ₈ alkynyl groups. Examples of alkynyl groups include, but are not limited to, propargyl, 2-butynyl, and the like.

"Heterocyclyl" means an aromatic or non-aromatic heterocyclic group wherein one or more of the ring-forming atoms are heteroatoms such as oxygen, nitrogen, sulfur atoms and the like. An aromatic heterocyclic group, commonly referred to as "heteroaryl", preferably refers to an aromatic 5 to 6 membered monocyclic or 9 to 10 membered bicyclic ring which may contain 1, 2 or 3 selected from nitrogen, oxygen and/or An atom in sulfur, such as furyl, pyridyl, 2-oxo-1,2-dihydropyridyl, pyridazinyl, pyrimidinyl, pyrazinyl, thienyl, isoxazolyl, oxazolyl, cacao Diazolyl, imidazolyl, pyrrolyl, pyrazolyl, triazolyl, tetrazolyl, thiazolyl, isothiazolyl, 1,2,3-thiadiazolyl, benzodioxolyl Benzimidazolyl, fluorenyl, isodecyl, 1,3-dioxo-isoindenyl, quinolyl, oxazolyl, benzisothiazolyl, benzoxazolyl and benzo Isoxazolyl. A preferred heteroaryl group is pyridyl. The heteroaryl group may have the substituents described above with respect to the term "aryl". The non-aromatic heterocyclic group means a non-aromatic heterocyclic group, preferably having a 5- to 6-membered monocyclic ring or an 8- to 10-membered double- or tricyclic ring, which may contain 1, 2 or 3 selected from nitrogen and oxygen. And/or an atom in sulfur such as morpholinyl, thiomorpholinyl, tetrahydropyranyl, 1,1-dioxo-thiomorpholinyl, piperidinyl, 2-oxo-piperidine Base, pyrrolidinyl, 2-oxo-pyrrolidinyl, piperazin-2-one, 8-oxa-3-aza-bicyclo[3.2.1]octyl and piperazinyl. The heterocyclic group may optionally have a substituent as described above with respect to the term "aryl".

Furthermore, the term "pharmaceutically acceptable salt" refers to certain salts of the above compounds which retain their original biological activity and which are suitable for pharmaceutical use. The pharmaceutically acceptable salt of the compound represented by the formula ( I ) may be a metal salt, an amine salt formed with a suitable acid, a metal salt preferably an alkali metal or an alkaline earth metal salt, and suitable acids include inorganic acids and organic acids, for example, Acetic acid, benzenesulfonic acid, benzoic acid, camphorsulfonic acid, citric acid, ethanesulfonic acid, fumaric acid, gluconic acid, glutamic acid, hydrobromic acid, hydrochloric acid, isethionic acid, lactic acid, malic acid, Malay Acid, mandelic acid, methanesulfonic acid, nitric acid, phosphoric acid, succinic acid, sulfuric acid, tartaric acid, p-toluenesulfonic acid, and the like. Particularly preferred are hydrochloric acid, hydrobromic acid, phosphoric acid and sulfuric acid, and most preferred is the hydrochloride salt.

"Cycloalkyl" means a saturated or partially saturated monocyclic, fused or spiro carbon ring. A ring composed of 3 to 7 carbon atoms is preferred. Examples include, but are not limited to, cyclopropyl, cyclobutyl, cyclopentyl, cyclohexyl, and the like.

"Alkoxy" means a group of (alkyl-O-). Among them, the alkyl group is defined in the relevant definition herein. Alkoxy groups of C ₁ -C ₆ are preferred. Examples thereof include, but are not limited to, methoxy, ethoxy, n-propoxy, isopropoxy, n-butoxy, isobutoxy, tert-butoxy and the like.

The term "aryl", when used alone or in combination, refers to a carbocyclic aromatic system containing one or two rings, wherein the rings may be joined together in a fused manner. The term "aryl" includes aryl groups such as phenyl, naphthyl, tetrahydronaphthyl. Preferably, the aryl group is a C ₆ -C ₁₀ aryl group, more preferably the aryl group is a phenyl group. The "aryl" may carry one or more substituents such as C ₁ -C ₆ alkyl, hydroxy, halogen, haloalkyl, nitro, cyano, C ₁ -C ₆ alkoxy, C ₁ - C ₆ alkylamino group and the like.

We have found that the compounds of the invention are highly active tumor growth inhibitors.

Detailed description of the preferred embodiment

The invention is further illustrated by the following examples. The examples show the preparation of representative compounds represented by formula ( I ) and related structure identification data. It is to be understood that the following examples are intended to illustrate the invention and not to limit the invention.

In the following examples, all temperatures are in degrees Celsius unless otherwise indicated, and unless otherwise indicated, various starting materials and reagents are commercially available. Commercially available starting materials and reagents were used without further purification unless otherwise indicated.

The glassware is dried in an oven and/or dried by heating. The reaction was followed up for analysis on a glass silica gel-60 F254 plate (0.25 mm) (TLC). Analytical thin layer chromatography was carried out and developed in a suitable solvent ratio (v/v). The end point of the reaction was taken when the starting material was depleted on TLC.

^{The 1} H NMR spectrum was measured using a Bruker instrument (400 MHz) and the chemical shift was expressed in ppm. The internal standard (0.00 ppm) of tetramethyl decane was used. ¹ H NMR representation: s = singlet, d = doublet, t = triplet, m = multiplet, br = broadened, dd = doublet of doublet, dt = doublet of triplet. If a coupling constant is provided, its unit is Hz.

Mass spectrometry was measured by LC/MS, and the ionization method was ESI or APCI.

All melting points have not been corrected.

The following examples are merely illustrative of the synthetic methods of the particular compounds invented. However, there are no restrictions on the synthesis method. The compounds which are not listed below can also be prepared by the same synthetic route and synthesis method as below, by selecting an appropriate starting material, and adjusting the reaction conditions with appropriate common-sense conditions where necessary.

Example Example 1: Preparation of 4-iodo-3-nitrobenzoic acid (Compound V):

45 g (0.25 mol) of 4-amino-3-nitrobenzoic acid, 400 ml of water and 100 ml of concentrated hydrochloric acid were added to the reaction flask. The mixture was turned on, the mixture was cooled to 0 to 5 ° C, and 50 mL of an aqueous solution of 25.9 g (0.38 mol) of sodium nitrite was added dropwise, and the solid was gradually dissolved. After the dropwise addition, the reaction was carried out at 0 to 5 ° C for 1 hour, and at this temperature. 200 ml of an aqueous solution containing 88 g (0.5 mol) of potassium iodide was added dropwise, and the mixture was stirred at room temperature for 2 h to precipitate a solid. Filtration, washing with water and drying gave a solid 4-iodo-3-nitrobenzoic acid (Compound V ) 65 g (0.22 mol), yield 89.7%.

Example 2: Preparation of methyl 4-iodo-3-nitrobenzoate (Compound IV):

Add 55 g (0.19 mol) of 4-iodo-3-nitrobenzoic acid (Compound V ), 16.5 g (0.12 mol) of potassium carbonate and 550 ml of acetonitrile to the reaction flask, stir to cool, and cool to 0~5 °C, add 52.9 ml (0.38 mol) of triethylamine, the control temperature was below 10 ° C, 71 ml (1.12 mol) of methyl iodide was added, and the reaction was heated to about 40 ° C for 8 hours. After the reaction was completed, most of the acetonitrile was evaporated under reduced pressure. Then, it was extracted with 500 ml of ethyl acetate, washed with water three times, the organic layer was dried over anhydrous sodium sulfate, filtered, and the filtrate was concentrated at least, then 300 ml of petroleum ether was added, stirred for 30 min, filtered, and the filter cake was washed with petroleum ether and dried. Methyl 4-iodo-3-nitrobenzoate (Compound IV ) was about 41.2 g (0.13 mol), and the yield was 71.5%. ¹ H NMR (400 MHz, DMSO- d ₆ ): δ 3.91 (s, 3H), 7.87 (dd, 1H, J ₁ = 8.08 Hz, J ₂ = 1.88 Hz), 8.26 (d, 1H, J = 8.12 Hz) , 8.34 (d, 1H, J = 9.12 Hz); MS (m/z): 308 [M+H].

Example 3: Preparation of methyl 4-(2-trimethyldecyl)ethynyl-3-nitrobenzoate (Compound III):

Add 25 g (0.08 mol) of methyl 4-iodo-3-nitrobenzoate (Compound IV ), 20 ml (0.14 mol) of triethylamine and 200 ml of tetrahydrofuran, protect with nitrogen, stir and dissolve, then add 2.4 g (0.0034 mol) of bis(triphenylphosphine)palladium dichloride, 0.65 g (0.0034 mol) of cuprous iodide and 13.7 ml (0.096 mol) of trimethylsulfonium acetylene, reacted at room temperature for 1 hour, concentrated under reduced pressure The mixture was evaporated to dryness. EtOAc (EtOAc m. Ethyl ethynyl-3-nitrobenzoate (Compound III ) 15.1 g (0.055 mol), yield 66.9%. ¹ H NMR (400 MHz, DMSO- d ₆ ): δ 0.09 (s, 9 H), 3.74 (s, 3H), 7.69 (d, 1H, J = 8.08 Hz), 8.00 (dd, 1H, J ₁ = 8.08) Hz, J ₂ = 1.52 Hz), 8.30 (d, 1H, J = 1.36 Hz); MS (m/z): 278 [M+H].

Example 4: Preparation of methyl 4-ethynyl-3-nitrobenzoate (Compound II-1) Ready:

Add 15 g (0.054 mol) of methyl 4-(2-trimethyldecyl)ethynyl-3-nitrobenzoate (Compound III ) and 225 ml of tetrahydrofuran to the reaction flask, and stir to cool to -20~ At -25 ° C, 35 ml of a solution of 7.1 g (0.027 mol) of tetrabutylammonium fluoride in tetrahydrofuran was added dropwise, and the reaction was continued for 20 min after the completion of the dropwise addition. After the TLC detection reaction was completed, 0.5 M hydrochloric acid was added to adjust the pH to 4-5. The organic layer was extracted with EtOAc (3 mL). Methyl 4-ethynyl-3-nitrobenzoate (Compound II-1 ) 7.3 g (0.036 mol) was obtained in a yield of 65.8%. ¹ H NMR (400 MHz, DMSO- d ₆ ): δ 3.93 (s, 3H), 5.02 (s, 1H), 7.93 (d, 1H, J = 8.08 HZ), 8.21. (dd, 1H, J ₁ = 8.08 Hz , J ₂ =1.64 Hz), 8.50 (d, 1H, J = 1.48 Hz); MS (m/z): 206 [M+H].

Example 5: Preparation of 4-ethynyl-3-nitrobenzamide (Compound I-1):

To the reaction flask was added 7.3 g (0.036 mol) of methyl 4-ethynyl-3-nitrobenzoate (Compound II-1 ), 300 ml of methanol and 200 ml of tetrahydrofuran, and the mixture was stirred for 1 hour. After the reaction was carried out for 24 hours at room temperature, the reaction was completed by EtOAc. EtOAc (EtOAc m. To the dryness, column chromatography yielded 3.6 g (0.0019 mol) of 4-ethynyl-3-nitrobenzamide (Compound I-1 ) in a yield of 53.2%. ¹ H NMR (400 MHz, DMSO- d ₆ ): δ 4.93 (s, 1H), 7.82 (s, 1H), 7.92 (d, 1H, J = 8.08 Hz), 8.21. (dd, 1H, J ₁ = 8.04 Hz , J ₂ =1.68 Hz), 8.37 (s, 1H), 8.57 (d, 1H, J = 1.56 Hz); MS (m/z): 191 [M+H].

Example 6 Preparation of N-methoxycarbonyl-4-ethynyl-3-nitrobenzamide (Compound I-2)

In a 100 ml four-neck reaction flask, 3 g (0.016 mol) of 4-ethynyl-3-nitrobenzamide (Compound I-1 ), 45 ml of tetrahydrofuran and 6 ml of methyl chloroformate were added and stirred to dissolve. The temperature was lowered to -10 ° C, 7.5 g of sodium hydride was added, and the reaction was kept at about 0 ° C for 30 minutes. After the reaction was completed, the reaction solution was poured into crushed ice, and the pH was made acidic with hydrochloric acid, and then extracted with 200 ml of ethyl acetate. The organic layer was washed three times with water, dried over anhydrous sodium sulfate and filtered, and then filtered and evaporated to dryness to give the crude product about 3.5 g, which was obtained by column chromatography to give N-methoxycarbonyl-4-ethynyl-3-nitrobenzene Indoleamine (Compound I-2 ) 1.6 g (0.0065 mol), yield 40.8%. ¹ H NMR (400 MHz, DMSO- d ₆ ): δ 3.76 (s, 3H), 4.98 (s, 1H), 7.91 (d, 1H, J = 8.08 Hz), 8.18 (dd, 1H, J ₁ = 8.08 Hz , J ₂ =1.80 Hz), 8.58 (d, 1H, J = 1.72 Hz), 11.36 (s, 1H); MS (m/z): 249 [M+H].

The following compounds were prepared according to the method of Example 6 using Compound I-1 as a starting material and selecting a suitable reagent:

Example 12: Preparation of methyl 4-cyano-3-nitrobenzoate (Compound II-8):

5 g (0.016 mol) of methyl 4-iodo-3-nitrobenzoate (Compound IV-1 ), 1.77 g (0.02 mol) of cuprous cyanide and 15 ml of hexamethylphosphonium triamine were added to the reaction flask. After heating to 100 ° C for 1 hour, the reaction was completed, cooled to room temperature, then extracted with 100 ml of ethyl acetate. The organic layer was washed three times with water, dried over anhydrous sodium sulfate, filtered, and the filtrate was concentrated to dryness. Methyl 4-cyano-3-nitrobenzoate (Compound II-8 ) 2.7 g (0.013 mol), yield 80.4%. ¹ H NMR (400 MHz, DMSO- d ₆ ): δ 3.96 (s, 3H), 8.34 (d, 1H, J = 7.9 Hz), 8.42 (d, 1H, J = 7.7 Hz), 8.68 (s, 1H) MS (m/z): 229 [M+Na].

Example 13: Preparation of 4-cyano-3-nitrobenzamide (Compound I-8 ):

2 g (9.71 mmol) of methyl 4-cyano-3-nitrobenzoate (Compound II-8 ) and 60 ml of methanol were added to the reaction flask, stirred, and ammonia gas was passed for 1 hour, and then reacted at room temperature for 8 hours. After the reaction was completed, most of the methanol was evaporated under reduced pressure, and then extracted with ethyl acetate (100 ml). Recrystallization from ethyl acetate n-hexane gave 4-cyano-3-nitrobenzamide (Compound I-8 ) 0.6 g, yield 32.6%. ¹ H NMR (400 MHz, DMSO- d ₆ ): δ 7.97 (s, 1H), 8.32 (d, 1H, J = 7.96 Hz), 8.39 (dd, 1H, J ₁ = 7.96 Hz, J ₂ = 1.56 Hz) , 8.53 (s, 1H), 8.78 (d, 1H, J = 1.52 Hz); MS (m/z): 190 [MH].

Example 14: Preparation of N-methyl-4-iodo-3-nitrobenzamide (Compound VII-1):

Add 10 g (0.034 mol) of 4-iodo-3-nitrobenzoic acid (Compound VI-1 ) and 50 ml of N,N-dimethylformamide to the reaction flask, stir to dissolve, and cool to below 10 °C. 7.5 ml (0.10 mol) of hydrazine chloride was added, and the mixture was allowed to react to room temperature for one hour. The reaction solution was poured into a low temperature 200 ml of 30% aqueous methylamine solution, stirred for 5 minutes to precipitate a solid, and then 500 ml of ice was added. Water, stirred for 10 minutes, filtered, washed with water and dried to give N-methyl-4-iodo-3-nitrobenzamide (Compound VII-1 ) 6.5 g (0.021 mol), yield 62.3%. ¹ H NMR (400 MHz, DMSO- d ₆ ): δ 2.81 (d, 3H, J = 4.1 Hz), 7.84 (d, 1H, J = 7.8 Hz), 8.23 (d, 1H, J = 8.1 Hz), 8.33 (s, 1H), 8.75 (s, 1H); MS (m/z): 307 [M+H].

Example 15: Preparation of N-methyl-4-(2-trimethyldecyl)ethynyl-3-nitrobenzamide (Compound VIII-1):

6 g (0.02 mol) of N-methyl-4-iodo-3-nitrobenzamide (Compound VII-1 ), 60 ml of tetrahydrofuran and 4.2 ml of triethylamine were added to a 100 ml four-necked flask, and dissolved by stirring. Add 0.44 g (0.63 mmol) of bis(triphenylphosphine)palladium dichloride, 0.24 g (1.26 mmol) of cuprous iodide and 6 ml (0.042 mol) of trimethylsulfonium acetylene, and react at room temperature for 1 h. The concentrated tetrahydrofuran was evaporated under reduced pressure. EtOAc (EtOAc)EtOAc. (2-Trimethyldecyl)ethynyl-3-nitrobenzamide (Compound VIII-1 ) 3.8 g (0.014 mol), yield 70.2%. ¹ H NMR (400 MHz, DMSO- d ₆ ): δ 0.26 (s, 9 H), 2.81 (d, 3H, J = 4.5 Hz), 7.87 (d, 1H, J = 8.1 Hz), 8.14 (dd, 1H) , J ₁ =8.1 Hz, J ₂ =1.6 Hz), 8.52 (d,1H, J =1.5 Hz), 8.83 (d,1H, J =4.4 Hz); MS(m/z):277[M+H ].

Example 16: Preparation of N-methyl-4-ethynyl-3-nitrobenzamide (Compound I-9)

In a 100 ml reaction flask, 3.3 g (0.012 mol) of N-methyl-4-(2-trimethyldecyl)ethynyl-3-nitrobenzamide (Compound VIII-1 ) and 20 ml of methanol were added. The mixture was stirred and dissolved, and 0.53 g (0.0056 mol) of potassium fluoride dihydrate was added thereto, and the reaction was completed for 30 minutes. After completion of the reaction, 60 ml of water was added dropwise, and the mixture was filtered and dried to give a solid (2.2 g). N-methyl-4-ethynyl-3-nitrobenzamide (Compound I-9 ) 1.5 g (0.0074 mol), yield 61.7%. ¹ H NMR (400 MHz, DMSO- d ₆ ): δ 2.82 (d, 3H, J = 4.56 Hz), 4.92 (s, 1H), 7.92 (d, 1H, J = 8.08 Hz), 8.17 (dd, 1H, J ₁ =8.08 Hz, J ₂ =1.76 Hz), 8.54 (d, 1H, J = 1.64 Hz), 8.86 (d, 1H, J = 4.32 Hz); MS (m/z): 203 [MH].

Example 17: Preparation of N,N-dimethyl-4-ethynyl-3-nitrobenzamide (Compound I-10)

The compound I-10 was prepared according to the method of Example 14, Example 15 and Example 16 using the compound 4-iodo-3-nitrobenzoic acid (Compound VI-1 ) and dimethylamine as the starting material: ¹ HNMR (400 MHz, CDCl) ₃ ): δ 3.02 (s, 3H), 3.14 (s, 3H), 3.62 (s, 1H), 7.67 (dd, 1H, J ₁ = 7.96 Hz, J ₂ = 1.64 Hz), 7.74 (d, 1H, J = 7.92 Hz), 8.12 (d, 1H, J = 1.56 Hz); MS (m/z): 219 [M+H].

Example 18: Preparation of N-Ethyl-4-ethynyl-3-nitrobenzamide (Compound I-11)

4 g (0.021 mol) of 4-ethynyl-3-nitrobenzamide (Compound I-1 ), 30 ml (0.320 mol) of acetic anhydride and 20 ml of acetic acid were added to the reaction flask, stirred and heated to 120 ° C. After the reaction was carried out for 15 hours, the reaction was completed with EtOAc (EtOAc) eluted eluted eluted eluted eluted Ethyl-3-nitrobenzamide (Compound I-11 ) 1.1 g (0.0047 mol), yield 22.6%. ¹ H NMR (400 MHz, DMSO- d ₆ ): δ 2.36 (s, 3H), 4.99 (s, 1H), 7.94 (d, 1H, J = 8.08 Hz), 8.21. (dd, 1H, J ₁ = 8.12 Hz , J ₂ =1.80 Hz), 8.60 (d, 1H, J = 1.72 Hz), 11.32 (s, 1H); MS (m/z): 231 [MH].

Example 19: Preparation of 4-(1-propynyl)-3-nitrobenzamide (Compound I-12)

The compound I-12 was prepared by the method of Example 3, Example 4 and Example 5 using the compound 4-iodo-3-nitrobenzoic acid methyl ester (Compound IV ) and trimethylpropargyl decane as starting materials: ¹ HNMR (400 MHz, DMSO- d ₆ ): δ 2.16 (s, 3H), 7.74 (s, 1H), 7.79 (d, 1H, J = 8.16 Hz), 8.14 (dd, 1H, J ₁ = 8.13 Hz, J ₂ =1.77 Hz), 8.30 (s, 1H), 8.50 (d, 1H, J = 1.71 Hz); MS (m/z): 203 [MH].

Example 20: Preparation of N-benzimidyl-4-ethynyl-3-nitrobenzamide (Compound I-13)

Compound I-13 was prepared according to the method of Example 6 using compound I-1 and benzamidine chloride as the starting material: ¹ HNMR (400 MHz, DMSO- d ₆ ): δ 4.89 (s, 1H), 7.54 (t, 2H, J =7.5 Hz), 7.66 (t, 1H, J = 7.4 Hz), 7.91 - 7.96 (m, 3H), 8.20 (dd, 1H, J ₁ = 8.1 Hz, J ₂ = 1.6 Hz), 8.58 (d, 1H) , J = 1.5 Hz), 11.52 (s, 1H); MS (m/z): 293 [MH].

Starting from the compound 4-iodo-3-nitrobenzoic acid (Compound VI-1 ), the appropriate amine and trimethylsulfonium acetylene were selected, and the compound I was prepared according to the method of Example 14, Example 15 and Example 16 in turn. 14. Compound I-15 , compound I-16 , compound I-17 , compound I-18 , compound I-19 , compound I-20 , compound I-21 , compound I-22 , compound I-23 , compound I- 24 and compound I-25 ; using the compound N-methyl-4-iodo-3-nitrobenzamide (compound VII-1 ) and trimethylpropargyl decane as starting materials, followed by Example 15 and Example 16 Preparation of Compound I-26:

Example 34: Preparation of N-methyl-4-cyano-3-nitrobenzamide (Compound I-27)

2 g (6.53 mmol) of N-methyl-4-iodo-3-nitrobenzamide (Compound VII-1 ), 0.09 g (10.0 mmol) of cuprous cyanide and 10 ml were added to the reaction flask. Hexamethylphosphonium triamine, heated to 100 ° C for 40 minutes, the reaction is completed, cooled to room temperature, then extracted with ethyl acetate, the organic layer was washed three times with water, dried over anhydrous sodium sulfate, decolorized with activated carbon, filtered, concentrated To dryness, ethyl acetate was recrystallized to give N-methyl-4-cyano-3-nitrobenzamide (Comp. I-27 ) (yield: ¹ H NMR (400 MHz, DMSO- d ₆ ): δ 2.84 (d, 3H, J = 4.6 Hz), 8.27 (d, 1H, J = 8.0 Hz), 8.32 (dd, 1H, J ₁ = 8.0 Hz, J ₂ = 1.5 Hz), 8.72 (d, 1H, J = 1.4 Hz), 8.90 (m, 1H); MS (m/z): 204 [MH].

Example 35: Preparation of 4-ethynyl-3-nitrosobenzamide (Compound I-28)

Step 1: Add 5 g (26.3 mmol) of 4-ethynyl-3-nitrobenzamide (Compound XII-1 ) and 75 ml of tetrahydrofuran to a 150 ml four-necked flask, dissolve by mechanical stirring, and dissolve. 10 g (186.9 mmol) of aqueous ammonium chloride solution 10 ml, then add 10 g (178.5 mmol) of iron powder in portions, react at room temperature for 4 hours, complete the reaction, filter, wash the filter cake with tetrahydrofuran, the filtrate with acetic acid The organic phase is washed with water, dried over anhydrous sodium sulfate, decolorized with activated carbon, filtered, concentrated to precipitated solid, and dried by filtration to give 4-ethynyl-3-aminobenzamide (Compound XI-1 , gray solid) 2.4 g The yield was 59.4%. ¹ H NMR (400 MHz, DMSO- d ₆ ): δ 4.55 (s, 1H), 7.25 (dd, 1H, J ₁ = 8.0 Hz, J ₂ = 1.6 Hz), 7.31 (d, 1H, J = 7.9 Hz) , 7.35 (s, 1H), 7.61 (d, 1H, J = 1.4 Hz), 7.97 (s, 1H), 8.21 (s, 1H), 8.65 (s, 1H); MS (m/z): 161 [ M+H].

Step 2: Add 0.46 g (2.87 mmol) of 4-ethynyl-3-aminobenzamide (Compound XI-1 ) to a 50 ml single-mouth bottle, 10 ml of water, add 1.2 g of 58% sulfuric acid, and add 0.92. The solution of 18.5% potassium dichromate was stirred for 3 minutes, the reaction was completed, ethyl acetate was extracted, dried over anhydrous sodium sulfate and separated by column chromatography to give 4-ethynyl-3-nitrosobenzamide (Compound I-28) , yellow green solid) 0.06 g, yield 12%, stored at -20 ° C. ¹ H NMR (400 MHz, DMSO- d ₆ ): δ 4.97 (s, 1H), 6.99 (d, 1H, J = 1.3 Hz), 7.70 (s, 1H), 8.09 (d, 1H, J = 8.0 Hz) , 8.27 (s, 1H), 8.34 (dd, 1H, J ₁ = 8.0 Hz, J ₂ = 1.6 Hz); MS (m/z): 219 [M+HCOO ^- ].

Example 36: Preparation of (4-ethynyl-3-nitrophenyl)(4-methylpiperazin-1-yl)methanone hydrochloride (Compound I-29)

0.75 g (2.74 mmol) of (4-ethynyl-3-nitrophenyl)(4-methylpiperazin-1-yl)methanone (Compound I-24 ) and 7.5 ml were added to a 50 ml vial Anhydrous methanol was dissolved by magnetic stirring. The pH was adjusted to 2 with concentrated hydrochloric acid. The white solid was separated by cooling with ice water, filtered, washed with anhydrous ethanol and dried to give (4-ethynyl-3-nitrophenyl) (4) -Methylpiperazin-1-yl)methanone hydrochloride (Compound I-29 ) 0.46 g, Yield: 54.1%. ¹ H NMR (400 MHz, DMSO- d ₆ ): δ 2.76 (s, 3H), 3.16 (br s, 4H), 3.66 (br s, 4H), 4.91 (s, 1H), 7.85 (dd, 1H, J ₁ = 8.0 Hz, J ₂ = 1.4 Hz), 7.88 (d, 1H, J = 8.0 Hz), 8.21 (d, 1H, J = 1.3 Hz); MS (m/z): 274 [M+H-HCl ].

Example 37: Preparation of (4-ethynyl-3-nitrophenyl)(piperazin-1-yl)methanone hydrochloride (Compound I-30)

Prepared by the method of Example 36 using (4-ethynyl-3-nitrophenyl)(piperazin-1-yl)methanone (Compound I-23 ) as the starting material (4-ethynyl-3-nitrophenyl) (Piperazin-1-yl)methanone hydrochloride (Compound I-30 ). ¹ H NMR (400 MHz, D ₂ O): δ 3.32 (br s, 2H), 3.45 (br s, 2H), 3.78 (br s, 2H), 4.03 (br s, 2H), 4.11 (s, 1H) , 7.80 (dd, 1H, J ₁ = 8.0 Hz, J ₂ = 1.6 Hz), 7.91 (d, 1H, J = 8.0 Hz), 8.24 (d, 1H, J = 1.4 Hz); MS (m/z) : 260 [M+H-HCl].

The pharmacodynamic screening of the compounds of the invention is carried out as follows: One. In vitro pharmacodynamic screening Cell culture

Human colon cancer cell lines (Colo205 and HCT-116), human breast cancer cell lines (MCF-7 and MDA-MB435), human lung cancer cell lines (A549, 95D and NCI460), human ovarian cancer cell line (OVCAR-5, OVCAR-8, HO8910 and SKOV-3), human leukemia (HL-60 and K562), human prostate cancer (DU145 and BXPC-3), human uterine cancer (Hela), human pancreatic cancer (PANC-1), human liver cancer (HepG 2), both obtained from ATCC. Colo205, HL-60, K562 cells were cultured in RPMI 1640 containing 2 mM/L-glutamine, 10% FBS, 1.0 mM sodium pyruvate. HCT-116, MCF-7, A549, 95D, HO8910, NCI460, OVCAR-5, OVCAR-8, DU145, BXPC-3, Hela, PANC-1, HepG 2 cells were cultured in 2 mM/L-glutamine Indoleamine, 10% FBS in DMEM. MDA-MB435 cells were cultured in L-15 containing 2 mM/L-glutamine, 10% FBS. SKOV-3 cells were cultured in McCoy's containing 2 mM/L monoglutamine, 10% FBS. Colo205, HL-60 and K562 cells were seeded in 96-well plates, 150 μL/well, 8000 cells per well, and 96-well plates were pre-incubated for 24 hours at 37 ° C, 5% CO ₂ , 100% relative humidity incubator. HCT-116, MCF-7, NCI460, OVCAR-5, OVCAR-8, Hela, HepG 2 and MDA-MB435 cells were seeded in 96-well plates at 5000 cells per well; A549, 95D, HO8910, DU145, BXPC-3, PANC-1 and SKOV-3 cells were seeded in 96-well plates at 10,000 cells per well, and 96-well plates were pre-incubated for 24 hours at 37 ° C, 5% CO ₂ , 100% relative humidity incubator. The cells are attached.

2. Screening compounds

50 μL of pre-cooled 50% (mass/volume) TCA fixed cells were added to the time zero control wells of each cell line. Add 50 μL of different concentrations of compound to other wells for 48 h, set 3 replicate wells for each drug concentration, and set blank control (cell culture medium, no cells), no drug control well (no drug, plus equivalent) Complete medium), positive drug control BSI-201, was placed in a 5% CO ₂ incubator at 37 ° C for 48 h under total humidity (100% relative humidity).

3. Cell detection

50 μL of pre-cooled 50% (mass/volume) TCA fixed cells were added to the culture solution. Then, it was allowed to stand at 4 ° C for 1 h, the supernatant was discarded, and each well was washed 5 times with deionized water to remove TCA and serum protein. After drying in air, add a sufficient amount of 0.4% SRB (prepared with 1% acetic acid) to each well for about 100 μL and leave it at room temperature for 20 to 30 minutes. The liquid in each well was discarded and quickly washed 5 times with 1% acetic acid to remove unbound dye until the unbound dye was rinsed clean. Dry in air until no moisture is visible, dissolve in 200 μL Tris base, shake on a plate shaker for 5 min or beat up and down with a Tip head and mix on the multifunction meter (M5 detection system, MD Group Ltd. The 690 nm blank was zeroed and the detection wavelength was 515 nm.

Using XL-fit Dose response curves plotted to determine its value ₅₀ GI.

4. Screening results

BSI-201 ( Iniparib ) was used as a positive drug by in vitro pharmacodynamic screening experiments, and the results showed that compounds I-1 , I-2 , I-3 , I-4 , I-5 , I-6 , I-7 , I -8 , I-9 , I-10 , I-11 , I-12 , I-13 , I-14 , I-15 , I-16 , I-17 , I-18 , I-19 , I-20 , I-21 , I-22 , I-23 , I-24 , I-25 , I-26 , I-27 , I-28 , I-29 , I-30 inhibit tumor cell proliferation, among which, compounds I-1 was more than 10 times more potent than the positive drug BSI-201 on the 18 tumor cell lines tested; I-2 , I-3 , I-14 , I-15 , I-17 , I-20 , I-21 , I-23 , I-24 , I-29 , I-30 are slightly less effective than I-1 ; I-4 , I-5 , I-6 , I-7 , I-8 , I- 9 , I-10 , I-11 , I-12 , I-13 , I-16 , I-18 , I-19 , I-25 , I-27 , I-28 also have a good inhibitory effect on tumor cell proliferation . The screening results are shown in Table 1.

Table 1 Partial data of compound (I) on tumor cell inhibitory activity

Table 1 (continued) Partial data of compound (I) on tumor cell inhibitory activity

two. In vivo pharmacodynamic screening 1. Establishment of animal models

5-6 week old female or male BALB/C nude mice weighing approximately 18-20 grams were housed. Construction of human cancer nude mouse xenograft tumor models: human colon cancer cell lines HCT-116 and Colo205, human breast cancer cell line MDA-MB435, human prostate cancer cell line BXPC-3, human ovarian cancer cell line SKOV-3, human Lung cancer cell line A549, human pancreatic cancer cell line PANC-1, and human uterine cancer cell line Hela were all derived from ATCC. Culture, the monolayer cultured tumor cells are digested and dissected, collected and resuspended in serum-free culture medium, adjusted to a concentration of 2×10 ⁶ /0.2 mL, placed in an ice box and carried to the animal room, directly with a belt The syringe of the 6th needle was transplanted into the subcutaneous scapula of the left axilla of the nude mouse with a 0.2 mL cell suspension, 2×10 ⁶ /0.2 mL/only, and the tumor-forming volume was measured every 2-3 days. After two weeks, the tumor growth was selected and In nude mice without ulceration, the tumor was removed under aseptic conditions, and the tumor tissue was cut into a diameter of about 2-3 mm and inoculated into the scapula of the left axilla of the nude mouse. After three passages, the tumor volume grew to 100 mm. ^{At 3} o'clock, the nude mice with too large or too small tumors were randomly divided into groups.

2. Screening compounds

They were randomly divided into 5 groups, including the negative control group (media), the positive control group (BSI-201, 80 mg/kg), and the high, medium and low doses of the three treatment groups (40 mg/kg, 30 mg/kg, 20, respectively). Mg/kg, of which high dose was lower than MTD), 8 nude mice in each group, of which 16 were negative control group, administered intraperitoneally once a day for 3 weeks. Animal body weight, tumor volume, and number of animal deaths were recorded every 2 days. Animals were sacrificed 24 hours after the last administration, tumor volume, tumor weight, nude mouse body weight were measured, tumor volume growth curve, nude mouse weight growth curve and tumor inhibition rate, animal mortality, and relative tumor growth rate T/C were calculated. %), according to the formula T/C (%) = TRTV / CRTV * 100%. (TRTV: treatment group RTV; CRTV: negative control group RTV, relative tumor volume RTV = V _t / V _0, wherein V ₀ is a packet when administered tumor volume, V _t is the tumor volume after administration).

3. Screening results

BSI-201 ( Iniparib ) was used as a positive drug in the in vivo drug screening test. The results showed that the dose of Compound I-1 30 mg/kg was equivalent to that of BSI-201 80 mg/kg, which had significant pharmacodynamic effects. See Table 2.

Table 2 Compound (I) inhibition rate in nude mice tumor model

All documents mentioned in the present application are hereby incorporated by reference in their entirety in their entireties in the the the the the the the the In addition, it should be understood that various modifications and changes may be made by those skilled in the art without departing from the scope of the invention.

Claims (24)

A compound represented by the formula (I), or a pharmaceutically acceptable salt thereof, or a prodrug thereof: Wherein: R ¹ and R ^{2 are} independently selected from the group consisting of hydrogen, substituted or unsubstituted (C ₁ -C ₈ )alkyl, substituted or unsubstituted (C ₃ -C ₈ )alkenyl, substituted or unsubstituted Substituted (C ₃ -C ₈ )alkynyl, substituted or unsubstituted (C ₃ -C ₇ )cycloalkyl, -COR ⁵ and -CO ₂ R ⁶ ; R ¹ and R ² may also be cyclized to form a substitution Or an unsubstituted four, five or six membered ring; R ³ is a nitro or nitroso group; R ^{4 is} selected from the group consisting of ethynyl, propynyl or cyano; and R ^{5 is} selected from the group consisting of: Unsubstituted (C ₁ -C ₈ )alkyl, substituted or unsubstituted (C ₃ -C ₈ )alkenyl, substituted or unsubstituted (C ₃ -C ₈ )alkynyl, substituted or unsubstituted (C _3- C ₇ ) cycloalkyl, substituted or unsubstituted aryl; and R ^{6 is} selected from the group consisting of substituted or unsubstituted (C ₁ -C ₈ )alkyl, substituted or unsubstituted (C ₃ - C ₈ ) alkenyl, substituted or unsubstituted (C ₃ -C ₈ ) alkynyl, substituted or unsubstituted (C ₃ -C ₇ )cycloalkyl; wherein the above substituted or unsubstituted (C ₃ -C ₈₎ alkenyl group, a substituted or unsubstituted (C ₃ -C ₈₎ alkynyl group is preferably a double bond and a triple bond alkynyl group, an alkenyl group does not Amides of N, carbonyl or carbonyloxy Then connected. The compound of claim 1, wherein R ¹ and R ^{2 are} independently selected from hydrogen, substituted or unsubstituted (C ₁ -C ₈ )alkyl, -COR ⁵ or -CO ₂ R ⁶ , R ¹ and R ² may also be cyclized to form a substituted or unsubstituted tetra, five or six membered ring wherein R ⁵ is substituted or unsubstituted (C ₁ -C ₈ )alkyl, substituted or unsubstituted (C _3- C ₈ )alkenyl, substituted or unsubstituted (C ₃ -C ₈ )alkynyl, substituted or unsubstituted (C ₃ -C ₇ )cycloalkyl, substituted or unsubstituted aryl; R ⁶ is Substituted or unsubstituted (C ₁ -C ₈ )alkyl, substituted or unsubstituted (C ₃ -C ₈ )alkenyl, substituted or unsubstituted (C ₃ -C ₈ )alkynyl, substituted or unsubstituted (C ₃ -C ₇ )cycloalkyl. The compound of claim 2, wherein R ¹ and R ^{2 are} independently selected from hydrogen, substituted or unsubstituted (C ₁ -C ₈ )alkyl, -COR ⁵ or -CO ₂ R ⁶ , R ¹ and R ² may also be cyclized to form a substituted or unsubstituted tetra, five or six membered ring wherein R ⁵ is substituted or unsubstituted (C ₁ -C ₈ )alkyl, substituted or unsubstituted (C _3- C ₇ ) cycloalkyl, substituted or unsubstituted aryl; R ⁶ is substituted or unsubstituted (C ₁ -C ₈ )alkyl, substituted or unsubstituted (C ₃ -C ₇ )cycloalkyl . The compound of claim 3, wherein R ¹ and R ^{2 are} independently selected from the group consisting of hydrogen, methyl, ethyl, propyl. The compound of claim 3, wherein R ¹ and R ² may also be cyclized to form a substituted or unsubstituted five- or six-membered ring, and the atom forming the ring may contain one or more heteroatoms. Wherein the hetero atom is a nitrogen, oxygen or sulfur atom, preferably a nitrogen atom. The compound of claim 5, wherein R ¹ and R ^{2 are} cyclized to form a substituted or unsubstituted five or six membered ring containing one or more heteroatoms, preferably a pyrrolidine ring, a piperidine ring, Morpholine ring, piperazine ring. A compound according to any one of the preceding claims, wherein R ^{5 is} selected from substituted or unsubstituted (C ₁ -C ₈ )alkyl, preferably methyl. The compound of any one of claims 1 to 6, wherein R ^{5 is} selected from a substituted or unsubstituted aryl group, preferably a phenyl group. A compound according to any one of the preceding claims, wherein R ^{6 is} selected from substituted or unsubstituted (C ₁ -C ₈ )alkyl. The compound of claim 9, wherein R ^{6 is} selected from the group consisting of methyl, ethyl, propyl, isopropyl, n-butyl or isobutyl, preferably methyl or ethyl. A compound according to any one of the preceding claims, wherein R ³ is a nitro group or a nitroso group, preferably a nitro group. A compound according to any one of the preceding claims, wherein R ^{4 is} selected from the group consisting of ethynyl, propynyl, preferably ethynyl. The compound of any one of claims 1 to 11, wherein R ⁴ is a cyano group. A compound according to claim 1, which is selected from the group consisting of: 4-ethynyl-3-nitrobenzamide; N-methoxycarbonyl-4-ethynyl-3-nitrobenzamide Amine; N-ethoxycarbonyl-4-ethynyl-3-nitrobenzamide; N-propoxycarbonyl-4-ethynyl-3-nitrobenzamide; N-butoxycarbonyl-4- Ethynyl-3-nitrobenzamide; N-isopropoxycarbonyl-4-ethynyl-3-nitrobenzamide; N-isobutoxycarbonyl-4-ethynyl-3-nitrobenzene Methionine; 4-cyano-3-nitrobenzamide; N-methyl-4-ethynyl-3-nitrobenzamide; N,N-dimethyl-4-ethynyl- 3-nitrobenzamide; N-acetamido-4-ethynyl-3-nitrobenzamide; 4-(1-propynyl)-3-nitrobenzamide; N- Benzylmercapto-4-ethynyl-3-nitrobenzamide; N,N-diethyl-4-ethynyl-3-nitrobenzamide; N,N-dipropyl-4 -ethynyl-3-nitrobenzamide; N,N-dibutyl-4-ethynyl-3-nitrobenzamide; N-ethyl-4-ethynyl-3-nitrobenzene Formamide; N-propyl-4-ethynyl-3-nitrobenzamide; N-butyl-4-ethynyl-3-nitrobenzamide; (4-ethynyl-3- Nitrophenyl)(pyrrolidin-1-yl)methanone (4-ethynyl-3-nitrophenyl)(piperidin-1-yl)methanone; (4-ethynyl-3-nitrophenyl)(morpholin-4-yl)methanone; (4 -ethynyl-3-nitrophenyl)(piperazin-1-yl)methanone; (4-ethynyl-3-nitrophenyl)(4-methylpiperazin-1-yl)methanone; (4-ethynyl-3-nitrophenyl)(azetidin-1-yl)methanone; N-methyl-4-(1-propynyl)-3-nitrobenzamide N-methyl-4-cyano-3-nitrobenzamide; 4-ethynyl-3-nitrosobenzamide; (4-ethynyl-3-nitrophenyl) (4 -methylpiperazin-1-yl)methanone hydrochloride; and (4-ethynyl-3-nitrophenyl)(piperazin-1-yl)methanone hydrochloride. A process for the preparation of a compound of formula ( I ) as described in claim 1, wherein the process comprises: reacting a compound of formula ( II ) with a compound of formula ( X ) to give: Wherein R ¹ and R ² are hydrogen, R ³ is a nitro group, and R ⁴ is an ethynyl group, a propynyl group or a cyano group. A process for the preparation of a compound of formula ( I ) as described in claim 1, wherein the process comprises: formulating a compound of formula ( IX ) with (R ⁵ CO) ₂ O, R ⁵ COX or XCO ₂ R ⁶ reaction to get: Wherein R ¹ is hydrogen, R ² is -COR ⁵ or -CO ₂ R ⁶ , R ² is a nitro group, R ⁴ is an ethynyl group, a propynyl group or a cyano group, and R ⁵ and R ^{6 are} as in the scope of claim 1 As defined by the item, X is a halogen. A process for the preparation of a compound of formula ( I ) as described in claim 1 wherein the process comprises: deamination of the compound of formula VIII : Wherein R ¹ and R ^{2 are} independently selected from hydrogen, substituted or unsubstituted (C ₁ -C ₈ )alkyl, substituted or unsubstituted (C ₃ -C ₈ )alkenyl, substituted or unsubstituted (C ₃ -C ₈ ) alkynyl, substituted or unsubstituted (C ₃ -C ₇ )cycloalkyl, R ¹ and R ² may also be cyclized to form a substituted or unsubstituted tetra, five or six-membered ring, but R ¹ , R ^{2 may} not be hydrogen at the same time; R ³ is a nitro group, and R ⁴ is an ethynyl group or a propynyl group. A process for the preparation of a compound of formula ( I ) as described in claim 1 of the patent application, which is as follows: Wherein R ¹ and R ^{2 are} independently selected from hydrogen, substituted or unsubstituted (C ₁ -C ₈ )alkyl, substituted or unsubstituted (C ₃ -C ₈ )alkenyl, substituted or unsubstituted (C ₃ -C ₈ ) alkynyl, substituted or unsubstituted (C ₃ -C ₇ )cycloalkyl, R ¹ and R ² may also be cyclized to form a substituted or unsubstituted tetra, five or six-membered ring, but R ¹ , R ² cannot be hydrogen at the same time; R ³ is a nitro group, and R ⁴ is a cyano group. A process for the preparation of a compound of formula ( I ) as described in claim 1 of the patent application, which is as follows: Wherein R ¹ and R ^{2 are} independently selected from hydrogen, substituted or unsubstituted (C ₁ -C ₈ )alkyl, substituted or unsubstituted (C ₃ -C ₈ )alkenyl, substituted or unsubstituted (C ₃ -C ₈ ) alkynyl, substituted or unsubstituted (C ₃ -C ₇ )cycloalkyl, -COR ⁵ and -CO ₂ R ⁶ ; R ¹ and R ² may also be cyclized to form a substituted or unsubstituted a four, five or six membered ring; R ³ is a nitroso group; R ⁴ is ethynyl, propynyl or cyano; R ⁵ is substituted or unsubstituted (C ₁ -C ₈ )alkyl, substituted or unsubstituted (C ₃ -C ₈ )alkenyl, substituted or unsubstituted (C ₃ -C ₈ )alkynyl, substituted or unsubstituted (C ₃ -C ₇ )cycloalkyl, substituted or unsubstituted aryl; R ⁶ is a substituted or unsubstituted (C ₁ -C ₈ )alkyl group, a substituted or unsubstituted (C ₃ -C ₈ )alkenyl group, a substituted or unsubstituted (C ₃ -C ₈ )alkynyl group, a substituted or Unsubstituted (C ₃ -C ₇ )cycloalkyl. A pharmaceutical composition comprising an effective amount of a compound of formula ( I ), or a pharmaceutically acceptable salt thereof, or a prodrug thereof, according to any one of claims 1 to 14. Use of a compound according to any one of claims 1 to 14 for the manufacture of a medicament for the treatment of a disease associated with a poly(ADP-ribose) polymerase (PARP) inhibitor. The use according to claim 21, wherein the disease associated with a poly(ADP-ribose) polymerase (PARP) inhibitor is selected from the group consisting of cancer, stroke, myocardial infarction, and neurodegenerative disease. Use of a compound according to any one of claims 1 to 14 for the preparation of a medicament for the treatment of cancer. The application according to claim 23, wherein the cancer comprises colon cancer, breast cancer, lung cancer, ovarian cancer, gastric cancer, acute leukemia, chronic leukemia, prostate cancer, uterine cancer, pancreatic cancer, liver cancer, brain cancer, CNS tumor, bladder cancer, kidney cancer, skin cancer, neck cancer, muscle tumor, lymphoma and bone cancer.