HK40013281B - Novel pyrrolopyridine derivative, method for producing same, and use thereof - Google Patents

Description

Novel pyrrolopyridine derivative, preparation method and application thereof

Technical Field

The present invention relates to an antiviral compound, particularly to a compound having high selectivity and physiological activity against Human Immunodeficiency Virus (HIV), a preparation method and a use thereof.

Background

AIDS (AIDS) is caused by infection with Human Immunodeficiency Virus (HIV). HIV has two types, HIV-1 and HIV-2, and the type spreading worldwide is HIV-1. For the treatment of AIDS, enzyme inhibitors have been developed according to the mechanism of action of HIV, and these enzyme inhibitors are classified into Nucleoside Reverse Transcriptase Inhibitors (NRTI), Protease Inhibitors (PI), Fusion inhibitors (Fusion Inhibitor), Integrase inhibitors (Integrase Inhibitor) according to their site of action.

Integrase inhibitors are classified into catalytic site (catalytic site) inhibitors and non-catalytic site (non-catalytic site) inhibitors according to their mechanism, and studies related to catalytic site integrase inhibitors have been actively conducted, and 3 kinds of drugs have been developed and marketed. Letirasvir (Raltegravir) developed in 2008 is a typical drug. On the other hand, the mechanism of action of non-catalytic site integrase inhibitors is described by Ziger Debyser et al (Frauke Christ, Zeger Debyser et al, Nature Chemical Biology,2010, Vol.6,442), and development of inhibitors against this mechanism of action has been conducted actively.

In addition, in order to develop a drug for effectively treating a virus showing drug resistance, various studies have been conducted, and such a chemotherapeutic drug shows a significant effect of prolonging life by being administered in combination with 2 to 4 drugs, which inhibit mechanisms different from each other, called High Active Anti Retroviral Therapeutics (HAART). However, despite these efforts, aids cannot be completely cured, and even due to toxicity problems of drugs and the manifestation of resistance to existing therapeutic agents, development of new therapeutic agents is continuously required.

As a result of repeated studies for developing a novel therapeutic agent for aids as a means of solving such problems, the present inventors have found that a pyrrolopyridine derivative compound having a novel skeleton has an effect of inhibiting HIV proliferation, and have completed the present invention.

Disclosure of Invention

Technical subject

An object of the present invention is to provide a novel pyrrolopyridine derivative and a pharmaceutically acceptable salt thereof, which has an effect of inhibiting the proliferation of HIV-1 by inhibiting the activity of integrase of HIV-1 and shows excellent results in drug properties and basic toxicity tests.

It is another object of the present invention to provide a process for preparing the novel pyrrolopyridine derivatives and pharmaceutically acceptable salts thereof.

Another object of the present invention is to provide a pharmaceutical composition comprising the above compound as an active ingredient.

Means for solving the problems

A first aspect of the present invention provides a compound of formula I, a racemate, a stereoisomer, or a pharmaceutically acceptable salt thereof: [ chemical formula I ]

In the above formula, R₁Selected from C substituted or unsubstituted by halogen atoms_1-6Alkyl radical, C_1-3Alkyl or halogen substituted or unsubstituted benzyl, C₁-C₃Alkoxymethyl group, C₁-C₃Alkyl carbamates and C₁-C₃Alkyl-substituted or unsubstituted sulfonyl groups,

R₂and R₃Each independently is hydrogen, C_1-6An alkyl group or a halogen atom.

In one embodiment, the invention provides R₁Is C_1-6Alkyl compounds, racemates, stereoisomers thereof or pharmaceutically acceptable salts thereof.

In another embodiment, the invention provides R₁Is methyl, R₂And R₃A compound, racemate, stereoisomer or pharmaceutically acceptable salt thereof, each independently hydrogen, methyl or chlorine.

In another embodiment, the invention provides R₁Is methyl, R₂And R₃A compound which is hydrogen, a racemate, a stereoisomer thereof or a pharmaceutically acceptable salt thereof.

Specifically, the halogen atom means a chlorine, bromine or fluorine atom.

A second aspect of the present invention provides a method for preparing a compound represented by formula I based on the following reaction formula 1.

[ reaction formula 1]

Specifically, the method for preparing the compound represented by the chemical formula I comprises the following steps:

1) a first step of reacting a compound represented by the following chemical formula II with a compound represented by the following chemical formula III to prepare a compound represented by the following chemical formula IV; and

2) a second step of hydrolyzing the compound represented by chemical formula IV.

[ chemical formula I ]

[ chemical formula II ]

[ chemical formula III ]

[ chemical formula IV ]

In the above-mentioned formula, the compound of formula,

R₁selected from C substituted or unsubstituted by halogen atoms_1-6Alkyl radical, C_1-3Alkyl or halogen substituted or unsubstituted benzyl, C₁-C₃Alkoxymethyl group, C₁-C₃Alkyl carbamates and C₁-C₃Alkyl-substituted or unsubstituted sulfonyl groups,

R₂and R₃Each independently is hydrogen, C_1-6An alkyl group or a halogen atom,

R₄is C_1-6An alkyl group, a carboxyl group,

x is halogen, methylsulfonyl, tosyl or trifluoromethanesulfonyl.

Specifically, R is as defined above₄Can be methyl or ethyl, and X can be chlorine or p-toluenesulfonyl.

In the first step of the above method for preparing formula I, the molar ratio of the compound represented by formula II to the compound represented by formula III is preferably 1:2 to 1:5, but is not limited thereto.

In the first step, dichloromethane, dimethylformamide, tetrahydrofuran, or any combination thereof is used as a reaction solvent, but the reaction is not limited thereto.

The first step may be performed for 2 to 18 hours, but is not limited thereto.

The first step may be carried out in the presence of cesium carbonate, and dimethylformamide is preferably used as a solvent.

The molar ratio of cesium carbonate used in the above first step is preferably 2 to 5 equivalents with respect to chemical formula II.

At this time, the reaction temperature is preferably carried out at 40 ℃ to 100 ℃ and the reaction time is preferably carried out for 4 hours to 18 hours, but not limited thereto.

For example, the compound represented by formula II used as a starting material in the preparation of the compound represented by formula I of the present invention can be prepared according to the method disclosed in the preparation example of international patent (WO 2013/073875 a 1).

In the second step, i.e., the hydrolysis step, lithium hydroxide, calcium hydroxide, barium hydroxide, and potassium hydroxide may be used, but the method is not limited thereto, and potassium hydroxide or lithium hydroxide may be preferably used.

The molar ratio of potassium hydroxide or lithium hydroxide used in the hydrolysis step may be 3 to 8 equivalents with respect to formula IV, but is not limited thereto.

The above hydrolysis step is specifically carried out at normal temperature, more specifically at 35 to 50 ℃.

As the solvent in the above hydrolysis step, water, methanol, tetrahydrofuran, or any combination thereof may be used, but is not limited thereto.

In one embodiment, the hydrolysis is carried out using lithium hydroxide in a mixed solvent of, for example, 4N-sodium hydroxide/methanol or tetrahydrofuran/methanol/water.

The hydrolysis step is specifically performed for 6 hours to 18 hours, but is not limited thereto.

A third aspect of the present invention provides an antiviral composition comprising a compound represented by formula I, a racemate, a stereoisomer, or a pharmaceutically acceptable salt thereof.

Specifically, the composition is a composition for resisting Human Immunodeficiency Virus (HIV).

Specific examples of the compound represented by formula I of the present invention may be (S) -2- (tert-butoxy) -2- (4- (4-chlorophenyl) -2,3, 6-trimethyl-1- ((1-methyl-1H-pyrazol-4-yl) methyl) -1H-pyrrolo [2,3-b ] pyridin-5-yl) acetic acid, or

(S) -2- (tert-butoxy) -2- (1- ((5-chloro-1, 3-dimethyl-1H-pyrazol-4-yl) methyl) -4- (4-chlorophenyl) -2,3, 6-trimethyl-1H-pyrrolo [2,3-b ] pyridin-5-yl) acetic acid.

The compounds of formula I of the present invention thus prepared may form salts, especially pharmaceutically acceptable salts. Pharmaceutically acceptable salts are not particularly limited as long as they are salts generally used in the art, such as acid addition salts (see document [ J.pharm.Sci.,1977,66,1])

Preferred examples of acids for pharmaceutically acceptable acid addition salts include, for example, inorganic acids such as hydrochloric acid, hydrobromic acid, phosphoric acid, orthophosphoric acid or sulfuric acid; or an organic acid such as methanesulfonic acid, benzenesulfonic acid, toluenesulfonic acid, acetic acid, propionic acid, lactic acid, citric acid, fumaric acid, malic acid, succinic acid, salicylic acid, maleic acid, glycerophosphoric acid, or acetylsalicylic acid.

Further, a pharmaceutically acceptable metal salt can be obtained by a conventional method using a base. For example, the compound of formula I above may be dissolved in an excess of alkali metal hydroxide or alkaline earth metal hydroxide solution, the insoluble compound salt filtered, and the filtrate evaporated and dried to obtain a pharmaceutically acceptable metal salt of the compound.

Pharmaceutically unacceptable salts or solvates of the compounds of formula I may be used as intermediates in the preparation of the compounds of formula I, pharmaceutically acceptable salts or solvates thereof.

The compounds of the above formula I of the present invention include not only pharmaceutically acceptable salts thereof but also solvates and hydrates that can be prepared therefrom. Stereoisomers of the compounds of formula I and intermediates described above can be prepared using conventional methods.

In addition, the compound of formula I according to the present invention may be prepared in a crystalline form or an amorphous form, and in the case where the compound of formula I is prepared in a crystalline form, it may be optionally hydrated or solvated.

In addition, the present invention provides an antiviral composition comprising the compound of formula I, or a pharmaceutically acceptable salt, hydrate, or solvate thereof as an active ingredient. In this case, the above antiviral composition is particularly a composition for resisting Human Immunodeficiency Virus (HIV).

In the experimental examples of the present invention, it was confirmed that the compound of the formula I has low cytotoxicity, excellent HIV inhibitory effect, high physiological activity, safety in basic toxicity test results, and excellent solubility suitable for pharmaceutical properties.

The pharmaceutical composition of the invention can be prepared into oral administration or injection administration forms. Examples of oral administration forms include tablets, capsules and the like, and these forms contain, in addition to the active ingredient, a diluent (e.g., lactose, glucose, sucrose, mannitol, sorbitol, cellulose and/or glycine), a lubricant (e.g., silicon dioxide, talc, stearic acid and its magnesium or calcium salt or polyethylene glycol). Tablets may also contain binding agents such as magnesium aluminium silicate, starch paste, gelatin, tragacanth, methylcellulose, sodium carboxymethylcellulose or polyvinylpyrrolidone and, where appropriate, disintegrating agents or boiling mixtures and/or absorbents such as starch, agar, alginic acid or its sodium salt, colouring, perfuming and sweetening agents. As the formulation for injection, an isotonic aqueous solution or suspension is preferable.

The above compositions may be sterilized and/or contain adjuvants such as preserving, stabilizing, wetting or emulsifying agents, salts and/or buffers for regulating the osmotic pressure and other therapeutically useful substances.

The above dosage forms can be prepared by conventional mixing, granulating or coating methods, and may contain the active ingredient in an amount of about 0.1 to 75% by weight, and preferably may contain 1 to 50% by weight. A unit dosage form for about 50 to 70kg of a mammal contains about 10-200 mg of the active ingredient.

The preferred dose of the compound of the present invention varies depending on the state and body weight of the patient, the severity of the disease, the form of the drug, the route and time of administration, and can be appropriately selected by those skilled in the art. For administration, administration may be by oral or non-oral route 1 time or divided over 1 day.

The pharmaceutical composition of the present invention can be administered to mammals typified by rats, mice, livestock, humans, and the like by various routes. All modes of administration are envisioned, for example, administration may be by oral, rectal, intravenous, intramuscular, subcutaneous, epidural or intraventricular (intracerebroovirologic) injection.

Effects of the invention

The compound of formula I, its racemate, stereoisomer, pharmaceutically acceptable salt, hydrate, solvate thereof according to the present invention shows high selectivity and physiological activity against viruses, especially Human Immunodeficiency Virus (HIV) with low toxicity, and thus can be effectively used for the treatment of infection against viruses, especially Human Immunodeficiency Virus (HIV).

Detailed Description

The present invention will be described in more detail below with reference to the following preparation examples and examples. However, the following preparation examples and examples are only for illustrating the present invention and it is not intended that the scope of the present invention be limited only by these preparation examples and examples.

Preparation example 1: preparation of 4- (chloromethyl) -1-methyl-1H-pyrazole hydrochloride

To (1-methyl-1H-pyrazol-4-yl) methanol (380mg, 3.39mmol) prepared according to a known method (Frey, R.R; at al, J.Med.chem.,2008,51, 3777-one 3787) were added dichloromethane (1.8mL) and triethylamine (2 drops) and cooled to 0 ℃. A solution of thionyl chloride (0.62mL) dissolved in toluene (1.8mL) was added slowly and stirred at 30 ℃ for 2 hours. As for the reaction liquid, the solvent and excess thionyl chloride were removed under reduced pressure, thereby obtaining the objective compound. This material was used for the next reaction without purification.

Preparation example 2: preparation of 4- (bromomethyl) -5-chloro-1, 3-dimethyl-1H-pyrazole

(5-chloro-1, 3-dimethyl) -1H-pyrazol-4-yl) methanol (937mg, 5.8mmol) prepared according to the known method (Attadro, G.; Tripthy, S., PCT int. appl.2010, WO 2010-132999A 1) was dissolved in dichloromethane (40mL) and cooled to 0 ℃. A solution of phosphorus tribromide (0.54mL, 5.8mmol) diluted in dichloromethane (5mL) was slowly added thereto, followed by stirring at room temperature for 1.5 hours. The solvent was removed under reduced pressure from the reaction solution, thereby obtaining the objective compound. This material was used for the next reaction without purification.

Example 1: (S) -2- (tert-butoxy) -2- (4- (4-chlorophenyl) -2,3, 6-trimethyl-1- ((1-methyl-1H-pyrazol-4-yl) methyl) -1H-pyrrolo [2,3-b ] pyridin-5-yl) acetic acid

Step 1: methyl (S) -2- (tert-butoxy) -2- (4- (4-chlorophenyl) -2,3, 6-trimethyl-1H-pyrrolo [2,3-b ] pyridin-5-yl) acetate (700mg, 1.69mmol) was dissolved in dimethylformamide (14mL), cesium carbonate (2.75g, 8.45mmol) and 10 drops of triethylamine were added, the temperature was adjusted to 40 ℃, and then the compound obtained in preparation example 1 (560mg, 3.39mmol) was added separately over 1 hour. The reaction was terminated by stirring at the same temperature for 18 hours. After the reaction solution was cooled with ice water, water (50mL) was added and the mixture was stirred for 10 minutes. The resulting solid was filtered and washed with water. The obtained solid was purified by silica gel column chromatography without drying (eluent: ethyl acetate/n-hexane 1/2 and 1/1), so as to obtain the objective compound (430mg, 50%).

¹H-NMR(CDCl₃,500MHz)δ1.01(s,9H),1.49(s,3H),2.30(s,3H),2.75(s,3H),3.69(s,3H),3.83(s,3H),5.11(s,1H),5.32(s,2H),7.30(m,2H),7.44-7.47(m,4H):MS(EI,m/e)＝509(M⁺)。

Step 2: the compound (369mg, 0.724mmol) obtained in step 1 was dissolved in tetrahydrofuran (5.5mL), and a 4N-sodium hydroxide/methanol solution (0.98mL) was added, followed by stirring at 35 ℃ for 18 hours. After the reaction solution was cooled to 10 ℃, 4N-hydrochloric acid was added for neutralization. As for the reaction solution, the solvent was removed under reduced pressure, and the residue was purified by silica gel column chromatography (eluent: dichloromethane/methanol 95/5 and 90/10) to obtain the title compound (260mg, 73%) as a white solid.

¹H-NMR(CD₃OD,500MHz)δ1.00(s,9H),1.52(s,3H),2.31(s,3H),2.72(s,3H),3.80(s,3H),5.14(s,1H),5.37(bs,2H),7.34-7.53(m,6H):

MS(EI,m/e)＝495(M⁺)。

Example 2: (S) -2- (tert-butoxy) -2- (1- ((5-chloro-1, 3-dimethyl-1H-pyrazol-4-yl) methyl) -4- (4-chlorophenyl) -2,3, 6-trimethyl-1H-pyrrolo [2,3-b ] pyridin-5-yl) acetic acid

Methyl (S) -2- (tert-butoxy) -2- (4- (4-chlorophenyl) -2,3, 6-trimethyl-1H-pyrrolo [2,3-b ] pyridin-5-yl) acetate (200mg, 0.48mmol) and the compound obtained in preparation example 2 (432mg, 1.44mmol) were reacted in the same manner as in example 1 to obtain the title compound (30mg, 44%).

¹H-NMR(CD₃OD,500MHz)δ1.00(s,9H),1.49(s,3H),1.90(s,3H),2.19(s,3H),2.68(s,3H),3.75(s,3H),5.20(s,1H),2.28(dd,J＝40.7,15.8Hz,2H),7.24(d,J＝7.4Hz,1H),7.47-7.39(m,2H),7.63(d,J＝7.4Hz,1H):

MS(EI,m/e)＝544(M⁺)。

Experimental example 1: investigation of HIV-1 (Wild type/Mutant type) inhibitory Effect of the Compound of the present invention and cytotoxicity test

In order to know the HIV-1 (wild type/mutant type) inhibitory effect of the compound of the present invention, an in vitro HIV-1 (wild type/mutant type) inhibitory effect test was carried out as follows according to a known method (H.tanaka et al, J.Med.chem.,1991,34, 349). MT-4 cells were used as host cells to examine the degree of cytotoxicity of the compounds of the present invention against virus-infected MT-4 cells.

First, MT-4 cells were cultured at 1X 10 in a medium⁴The cells/wells were dispersed at a concentration of up to 500TCI₅₀(50% of the cells infected concentration)/well of HIV-1. Immediately after inoculation, the cell dispersion was transferred every 100. mu.L to a flat-bottom microtiter plate containing a sample of the compound of the present invention, and cultured at 17 ℃ for 4 to 5 days, and then the virus inhibitory effect was judged by the MTT method. In addition, cytotoxicity was judged by measuring the survival rate of cells infected with viruses through experiments using the MTT method. As comparative compounds, zidovudine (AZT), Raltegravir (Raltegravir), Dolutegravir (Dolutegravir) and Elvitegravir (Elvitegravir) were used. The results are shown in tables 1 and 2 below.

[ Table 1]

*EC₅₀: inhibiting HIV infection at a concentration of 50%

[ Table 2]

HIV-1 clones: letirasvir resistance mutants

(4736_2/4736_4/8070_1/8070_2/1556_1)

**IC₅₀: half inhibitory concentration

Experimental example 2: pharmacokinetic testing (Pharmacokinetics test) of the Compound of the invention

Experiments were conducted to confirm dynamic changes in vivo (in vivo) absorption, distribution, metabolism, excretion, and the like, for the compound of example 1 of the present invention. The jugular vein and the femoral vein of the rat were cannulated, and in the case of intravenous administration, administration was via the femoral vein, and in the case of oral administration, administration was via the oral cavity, and then blood was periodically collected from the jugular vein.

The administration concentration was 1mg/kg for intravenous administration and 2mg/kg for oral administration. For blood, plasma was separated by centrifugation, and plasma and urine samples were pretreated using a titration organic solvent, and then analyzed for concentration using LC-MS/MS. Non-compartmental pharmacokinetic parameters (noncompartmental pharmaceutical parameters) were calculated from the blood concentration-time data of the drug analyzed after oral and intravenous administration using WinNonlin (Pharsight, USA).

[ Table 3]

Experimental example 3: in vitro metabolic stability test of Compounds of the invention (in vitro metabolic stability test)

An in vitro metabolic stability assay for the compound of example 1 of the present invention was performed. In vitro metabolic stability, the stability of a drug was tested by measuring the half-life in liver microsomes (liver microsomes), reacting drug compounds with NADPH using species-specific (rat, dog, monkey, human) liver microsomes containing various metabolic enzymes, and then quantifying the drug in minutes by LC-MS/MS. It is clear that the compound of example 1 is a stable compound having a half-life of 2 or 3 hours or more.

[ Table 4]

Experimental example 4: CYP450 inhibition assay for Compounds of the invention

CYP450 inhibition assays were performed on example compounds of the present invention. To human liver microsomes (0.25mg/ml), 0.1M phosphoric acid buffer (pH 7.4) and 5 drug metabolizing enzyme substrates (CYP1A2, CYP2C9, CYP2D6, CYP3A4, CYPC19) drug cocktails (drug cocktails) (cocktail A: phenacetin 50. mu.M, S-mefenton 100. mu.M, dextromethorphan 5nM, imidazopyr 2.5. mu.M, cocktail B: tolbutamide 100. mu.M), the compound of example 1 was added at a concentration of 0, 10. mu.M each, and incubated at 37 ℃ for 15 minutes. Thereafter, in order to terminate the reaction, an acetonitrile solution containing an internal standard substance (chlorpropamide) was added, centrifugation was performed for 5 minutes (14,000rpm, 4 ℃), and then the supernatant was injected into an LC/MS/MS system while analyzing metabolites of the substrate drug, thereby evaluating the drug-metabolizing enzyme inhibitory activity based on the test substance. It was evaluated that the compound of example 1 showed no inhibitory activity against these 5 CYP enzymes.

[ Table 5]

Experimental example 5: hERG K of the Compounds of the invention⁺Channel activity assay

Implementation of hERG K capable of predicting cardiotoxicity of the compounds of the invention⁺Channel activity assay. Using a fully Automated planar patch clamp (Automated planar patch clamp) [ PatchXpress 7000A]HERG-HEK293 was used to determine the hERG activity of the compounds. This method is the most representative method for studying ion channels, and uses a voltage clamp (voltage clamp) to directly measure the flow of ions through a channel. hERG K for the Compound of example 1⁺IC of channel₅₀The value showed 66.7. mu.M, IC₅₀The value of 10 μ M or less is a criterion for judging that cardiotoxicity is likely to be exhibited, and the compound of example 1 is a safe compound because it is more than that.

[ Table 6]

Claims (3)

1. The following compounds or pharmaceutically acceptable salts thereof:

2. an antiviral composition comprising the compound of claim 1 or a pharmaceutically acceptable salt thereof.

3. Use of a compound according to claim 1 or a pharmaceutically acceptable salt thereof, or an antiviral composition according to claim 2, in the manufacture of a medicament for the treatment or inhibition of a viral infection, wherein the virus is a human immunodeficiency virus-1 wild type or mutant, wherein the human immunodeficiency virus-1 mutant is selected from the group consisting of the latifoliate virus resistant mutants 4736_2, 4736_4, 8070_1, 8070_2 and 1556_ 1.