KR20180083094A - Carbocyclic nucleosides derivatives and the antivirus compositions containing the same - Google Patents

Abstract

Provided is a carbocyclic nucleoside derivative or a pharmaceutically acceptable salt thereof. The carbocyclic nucleoside derivative or the pharmaceutically acceptable salt thereof is represented by the following chemical formula A. In the chemical formula A, R_1and R_2 are independently hydrogen or fluorine, and B is the following chemical formula B-1 or B-2. In the chemical formula B-1, X_1 is chlorine, a hydroxyl group, an amino group or an alkylamino group, and Y_1 is hydrogen or an amino group. In the chemical formula B-2, X_2 is a hydroxyl group or an amino group, and Y_2 is hydrogen, a methyl group, or halogen.

Description

[0001] CARBOCYCLIC NUCLEOSIDES DERIVATIVES AND THE ANTIVIRUS COMPOSITIONS CONTAINING THE SAME [0002] The present invention relates to carbocyclic nucleoside derivatives,

The present invention relates to a carbocyclic nucleoside derivative and an antiviral agent comprising the same.

Viruses are one of the leading causes of many intractable diseases that pose a threat to the health of human beings, and they are investing heavily in capital to prevent or treat them worldwide. In recent years, intractable RNA viruses such as Chickungunya virus, SARS virus, MERS virus and Zika virus have posed a serious risk to mankind and inhibited their proliferation. Although there is an urgent need for the development of antiviral drugs, currently there are no antiviral drugs or prophylactics that can effectively inhibit these viruses.

Antiviral agents, on the other hand, can inhibit the virus through a variety of mechanisms, including the following two mechanisms that can be used to effectively suppress the virus.

The first is to inhibit S-adenosylhomocysteine (hereinafter, SAH) hydrolase.

SAH hydrolytic enzyme is a tetramer type enzyme that uses NAD ⁺ as a coenzyme. It plays a role of reversible hydrolysis of SAH to adenosine (Ado) and homocysteine (Hcy) Lipid and nucleic acids as well as methylation of substances in the body such as histamine and norepinephrine.

Inhibition of SAH hydrolytic enzymes induces accumulation of SAH. Excess SAH sequentially inhibits the inhibition of S-adenosylmethionine (AdoMet) -dependent transmethylase and capping of virus mRNA, It prevents the virus from being properly produced, resulting in an antiviral effect.

Since viral mRNA guanosine N7-methytransferases (O-2'-methytransferases) are essential for mRNA encapsulation as well as most animal DNA viruses, SAH hydrolase is considered to be an essential element in the development of broad- spectrum antiviral agents do. That is, the development of a therapeutic agent for RNA viruses and the development of a SAH hydrolase inhibitor are considered to have a high correlation.

The second is to inhibit the viral RNA polymerase.

RNA viruses are replicated by inserting the substrate Nucleoside-5'-triphosphate (NTP) into the RNA chain by RNA polymerase. Therefore, a substance that inhibits RNA polymerase can also act as an antiviral agent and can be converted into triphosphate in the body to selectively inhibit viral RNA polymerase, or directly inserted into a viral RNA chain to perform chain termination ) Can be used to develop an effective antiviral agent.

However, since it is difficult to increase selectivity when an antiviral agent also has an inhibitory function of an RNA polymerase, it may exhibit toxicity to normal cells as well as viruses.

Accordingly, a problem to be solved by the present invention is to provide a pharmaceutical composition which is capable of effectively inhibiting a virus because it has a function of inhibiting SAH hydrolase, has little toxicity in the body, is well suited for the inhibition of viruses and for the prevention and treatment of viral diseases And to provide novel carbocyclic nucleoside derivatives that can be utilized.

The present invention has been made in view of the above problems, and it is an object of the present invention to provide a method of manufacturing the same.

In order to solve the above-mentioned problems, a carbocyclic nucleoside derivative or a pharmaceutically acceptable salt thereof according to an embodiment of the present invention is represented by the following formula (A).

≪ Formula (A)

In the formula (A), R ₁ and R ₂ are each independently hydrogen or fluorine, and B is B-1 or B-2.

<Formula B-1>

In the above formula (B-1), X ₁ is chlorine, a hydroxyl group, an amino group or an alkylamino group, and Y ₁ is a hydrogen atom or an amino group.

<Formula B-2>

In the above formula (B-2), X ₂ is a hydroxyl group or an amino group, and Y ₂ is hydrogen, a methyl group or a halogen.

The R &lt; ₁ &gt; may be fluorine.

The formula (A) may be represented by the following formula (5a) or (5b).

&Lt; Formula 5a &

&Lt; Formula 5b >

An antiviral agent comprising a carbocyclic nucleoside derivative or a pharmaceutically acceptable salt thereof according to another embodiment of the present invention for solving the above-mentioned problems is represented by the following formula (A).

&Lt; Formula (A)

In the formula (A), R ₁ and R ₂ are each independently hydrogen or fluorine, and B is B-1 or B-2.

<Formula B-1>

In the above formula (B-1), X ₁ is chlorine, a hydroxyl group, an amino group or an alkylamino group, and Y ₁ is a hydrogen atom or an amino group.

<Formula B-2>

In the above formula (B-2), X ₂ is a hydroxyl group or an amino group, and Y ₂ is hydrogen, a methyl group or a halogen.

The R &lt; ₁ &gt; may be fluorine.

The formula (A) may be represented by the following formula (5a) or (5b).

&Lt; Formula 5a &

&Lt; Formula 5b >

Advantages and features of the present invention and methods of achieving them will become apparent with reference to the embodiments described in detail below. The present invention may, however, be embodied in many different forms and should not be construed as being limited to the embodiments set forth herein. Rather, these embodiments are provided so that this disclosure will be thorough and complete, and will fully convey the scope of the invention to those skilled in the art. Is provided to fully convey the scope of the invention to those skilled in the art, and the invention is only defined by the scope of the claims.

Hereinafter, the present invention will be described in detail.

A carbocyclic nucleoside derivative according to an embodiment of the present invention may be represented by the following formula (A).

&Lt; Formula (A)

In the formula (A), R ₁ and R ₂ are each independently hydrogen or fluorine, and B may be represented by the following formula (B-1) or (B-2).

<Formula B-1>

In the above formula (B-1), X ₁ is chlorine, a hydroxy group, an amino group or an alkylamino group, and Y ₁ may be hydrogen or an amino group.

The alkylamino group may be a monoalkyl amino group (-NHR), or -NMe _2, -NMeEt, -NEt _2, such as a dialkyl amino group (-NR _2), such as -NHMe, -NHEt.

<Formula B-2>

In the above formula (B-2), X ₂ is a hydroxyl group or an amino group, and Y ₂ may be hydrogen, a methyl group or a halogen.

Specifically, in the above formula (A), R ₁ may be fluorine, and B may be the above formula (B-1).

More specifically, the carbocyclic nucleoside derivative represented by the above formula (A) may be a compound represented by the following formula (5a) or a compound represented by the following formula (5b).

&Lt; Formula 5a &

&Lt; Formula 5b >

The carbocyclic nucleoside derivative represented by the above formula (A) may be provided in the form of a pharmaceutically acceptable salt. Salts are useful as acid addition salts formed by various pharmaceutically acceptable organic or inorganic acids. Suitable organic acids include, for example, organic acids such as saturated carboxylic acids, phosphonic acids, sulfonic acids, acetic acid, propionic acid, octanoic acid, decanoic acid, glycolic acid, lactic acid, fumaric acid, succinic acid, adipic acid, malic acid, tartaric acid, There may be used maleic acid, benzoic acid, salicylic acid, phthalic acid, phenylacetic acid, benzenesulfonic acid, 2-naphthalenesulfonic acid, methylsulfuric acid, ethylsulfuric acid and dodecylsulfuric acid. Suitable inorganic acids include, for example, Or phosphoric acid can be used.

However, the present invention is not limited thereto. The carbocyclic nucleoside derivative of the present invention may be provided in the form of all salts, hydrates and solvates which can be prepared by conventional methods.

The antiviral agent according to one embodiment of the present invention may include the above-mentioned carbocyclic nucleoside derivative as an active ingredient.

An antiviral agent means a pharmaceutical composition which can be used not only for inhibiting the activity and replication of a virus but also for preventing and / or treating all diseases caused by viruses.

The virus may be, for example, Chickungunya Virus (CHIKV), Semliki Forrest Virus (SFV), Middle East Respiratory Syndrome Coronavirus (MERS-CoV), Severe Acute Viruses such as Respiratory Syndrome Coronavirus (SARS-CoV), Equine Arteritis Virus (EAV), Mouse Hepatitis Virus (MHV), and Sindbis Virus (SINV).

The antiviral agent comprising the above-described carbocyclic nucleoside derivative may be a pharmaceutical composition that effectively inhibits the virus by inhibiting the SAH hydrolase.

The antiviral agent may be administered systemically or locally and may be formulated using excipients (or diluents) such as fillers, extenders, binders, wetting agents, disintegrants, and surfactants that can be generally used for oral or parenteral administration . Hereinafter, the excipient and the formulation method will be specifically exemplified, but are not limited thereto.

Solid formulations for oral administration may include tablets, pills, powders, granules, capsules and the like, which may contain one or more excipients such as starch, calcium carbonate, sucrose sucrose), lactose, gelatin, and the like. In addition to simple excipients, lubricants such as magnesium stearate, talc, and the like may also be used. Examples of liquid formulations for oral administration include suspensions, solutions, emulsions, syrups, and the like. The liquid formulations may contain various excipients such as water, liquid paraffin, which are simple diluents commonly used, Flavoring agents, fragrances, preservatives, and the like.

Formulations for parenteral administration may include injections, emulsions, inhalants, suppositories, and the like. The injectable preparation may contain sterilized aqueous solvents such as propylene glycol, polyethylene glycol, vegetable oils such as olive oil, esters such as ethyl oleate, non-aqueous solvents and suspensions, and the suppositories may contain witepsol, Macrogol, tween 61, cacao paper, laurin, glycerogelatin, and the like. In addition, the antiviral agent of the present invention may be formulated into ointments or creams for topical application.

The preferred dosage of the antiviral agent varies depending on a number of factors such as the condition and weight of the patient, the severity of the disease, the type of the drug, the route of administration and the period of time, but may be suitably selected by those skilled in the art. The route of administration may also vary depending on the condition of the patient and its severity.

Since the carbocyclic nucleoside derivative of the present invention has antiviral efficacy against RNA viruses and has biocompatible properties with little toxicity in the body (see Experimental Example), it is possible to inhibit viruses and prevent and / Or as a pharmaceutical composition that is well suited for treatment.

Manufacturing example  One

(CuBr / Me ₂ S complex) (933 mg, 4.54 mmol) was dissolved in anhydrous THF (300 ml) at -78 ° C. and then a 1.0 M solution of vinylmagnesium bromide in THF 69.3 ml, 69.3 mmol) and hexamethylphosphoramide (HMPA) (33.4 ml, 192 mmol). D-cyclopentenone (5 g, 32.4 mmol) and chlorotrimethylsilane (TMSCl) (20.6 ml, 162 mmol) were dissolved together in anhydrous THF (180 ml), and the mixture was slowly added dropwise to the reaction flask. Lt; / RTI &gt; for 2 h. After completion of the reaction was confirmed by TLC, an aqueous solution of saturated ammonium chloride (NH ₄ Cl) was added thereto. The aqueous layer was extracted with ethyl acetate, and the organic layer was separated. The separated organic layer was washed thoroughly with water and brine, dried over magnesium sulfate (MgSO ₄ ), and filtered under reduced pressure to remove the remaining solid and then concentrated under reduced pressure. The concentrated residue was purified by silica gel chromatography to obtain a first intermediate (4.48 g, 76%).

The first intermediate (4.7 g, 26.0 mmol) was dissolved in methanol (100 mL) and sodium borohydride (NaBH ₄ ) (1.1 g, 28.6 mmol) was added at 0 ° C. The reaction mixture was stirred at the same temperature for 30 minutes, and neutralized with water and a small amount of acetic acid. The aqueous layer was extracted with ethyl acetate, and the organic layer was separated. The separated organic layer was washed thoroughly with water and brine, dried over magnesium sulfate (MgSO ₄ ), and filtered under reduced pressure to remove the remaining solid and then concentrated under reduced pressure. The concentrated residue was purified by silica gel chromatography to obtain a second intermediate (4.6 g, 97%).

The imidazole (8.6 g, 126.2 mmol) and chlorotetrabutyldiphenylsilane (TBDPSCl) (10.4 g, 37.8 mmol) were dissolved in DMF (30 mL), and the second intermediate (4.6 g, 25.2 mmol) At 0 &lt; 0 &gt; C. The reaction mixture was stirred at room temperature for 1 hour, water was added, and the mixture was extracted with diethyl ether and the organic layer was separated. The separated organic layer was washed thoroughly with water and brine, dried over magnesium sulfate (MgSO ₄ ), and filtered under reduced pressure to remove the remaining solid and then concentrated under reduced pressure. The concentrated residue was purified by silica gel chromatography to obtain a third intermediate (10.2 g, 92%).

The tertiary intermediate (10.0 g, 23.6 mmol) was dissolved in THF (125 mL), and a borane-dimethyl sulfide complex 2.0 M solution in THF (35.5 mL, 71.0 mmol) Lt; 0 &gt; C. The reaction mixture was stirred at room temperature for 4 hours, then sodium perborate (7.2 g, 71.0 mmol) was added at 0 ° C and water (30 mL) was slowly added thereto. After the reaction mixture was stirred at room temperature for 12 hours, the aqueous layer was extracted with ethyl acetate, and the organic layer was separated. The separated organic layer was washed thoroughly with water and brine, dried over magnesium sulfate (MgSO ₄ ), and filtered under reduced pressure to remove the remaining solid and then concentrated under reduced pressure. The concentrated residue was purified by silica gel chromatography to obtain a fourth intermediate (7.5 g, 72%).

The fourth intermediate (7.5g, 17.0mmol) of was dissolved in THF (200mL) at 0 ℃ 1.0M tetra - n - butylammonium fluoride / THF solution (tetra- n -butylammonium fluoride 1.0M solution in THF) (51.0 mL, 51.1 mmol). The reaction mixture was stirred at room temperature for 12 hours, and the solvent was removed under reduced pressure. The concentrated residue was separated and purified by silica gel chromatography to obtain a fifth intermediate (3.3 g, 93%).

4-Dimethylaminopyridine (DMAP) (0.2 g, 1.6 mmol) and triethylamine (6.8 mL, 48.9 mmol) were dissolved in 40 mL of methylene chloride. mmol) and chlorotetrabutyldiphenylsilane (TBDPSCl) (4.9 g, 17.9 mmol). The reaction mixture was stirred at room temperature for 12 hours. After water was added to terminate the reaction, the aqueous layer was extracted with methylene chloride, and the organic layer was separated. The organic layer was washed thoroughly with water and brine, dried over magnesium sulfate (MgSO ₄ ), and filtered under reduced pressure to remove the remaining solid, which was then concentrated under reduced pressure. The concentrated residue was purified by silica gel chromatography to obtain a sixth intermediate (5.7 g, 80%).

After dissolving the sixth intermediate (5.0 g, 11.3 mmol) in methylene chloride (25 mL), N-methylmorpholine N-oxide (2.0 g, 17.0 mmol) and 4 molecular sieves (1.0 g) and tetrapropylammonium perruthenate (0.2 g, 0.5 mmol) were added. The reaction mixture was stirred at room temperature for 30 minutes, then filtered through Celite and silica gel pad under reduced pressure to remove the remaining solid and then concentrated under reduced pressure. The concentrated residue was separated and purified by silica gel chromatography to obtain 4.8 g (98%) of a compound represented by the following formula (1) having a ¹ H-NMR spectrum as described below.

¹ H NMR (400 MHz, CDCl ₃ )? 7.64 (m, 4 H), 7.41 (m, 6 H), 4.57 (d, J = 5.6 Hz, m, 3 H), 2.73 ( dd, J = 8.8, 18.4 Hz, 1 H), 2.61 (m, 1 H), 2.03 (dt, J = 1.6, 17.6 Hz, 1 H), 1.68 (m, 1 H ), 1.43 (s, 3 H), 1.33 (s, 3 H), 1.05 (s, 9 H)

&Lt; Formula 1 >

Manufacturing example  2-1

Compound (1) (4.2 g, 9.5 mmol) synthesized in Preparation Example 1 was dissolved in anhydrous THF, and chlorotriethylsilane (TESCl) (16.7 mL, 99.2 mmol) was added dropwise and cooled to -78 ° C. To the cooled mixture, a 1.0 M lithium bis (trimethylsilyl) amide / THF solution (LiHMDS 1.0M solution in THF) (50 mL, 50 mmol) was slowly added dropwise and the reaction was stirred for 30 minutes under the same conditions for silylenol etherification. After confirming that the reaction was completed by TLC, the reaction mixture was added with an aqueous solution of saturated ammonium chloride (NH ₄ Cl), the aqueous layer was extracted with ethyl acetate, and the organic layer was separated. The separated organic layer was washed thoroughly with water and brine, dried over magnesium sulfate (MgSO ₄ ), and filtered under reduced pressure to remove the remaining solid and then concentrated under reduced pressure. The concentrated residue was dissolved in anhydrous acetonitrile and then cooled to 0 &lt; 0 &gt; C. To the cooled mixture was added a solution of 1-chloromethyl-4-fluoro-1,4-diazoniabicyclo [2.2.2] octane bis (tetrafluoroborate) (Selectfluor ^® ) (10.15 g, 28.65 mmol) The mixture was stirred for 15 hours under the same conditions to obtain a mixture of fluorinated compounds.

After confirming that the reaction for producing the mixture was completed by TLC, an aqueous solution of saturated ammonium chloride (NH ₄ Cl) was added to the reaction mixture, the aqueous layer was extracted with ethyl acetate, and the organic layer was separated. The separated organic layer was washed thoroughly with water and brine, dried over magnesium sulfate (MgSO ₄ ), and filtered under reduced pressure to remove the remaining solid, which was then concentrated under reduced pressure. The concentrated residue was separated and purified by silica gel chromatography to obtain the compound of the following formula (2a) (3.7 g, 84%) having the ¹ H NMR spectrum as described below.

¹ H NMR (300 MHz, CDCl ₃ )? 7.65 (m, 4 H), 7.39 (m, 6 H), 5.43 (dd, J = 50.6, 7.14 Hz, 3H), 1.29 (s, 3H), 1.03 (m, 2H), 1.46 s, 9 H) (mixed with a peak estimated to be equilibrated in diol form).

&Lt; EMI ID =

Manufacturing example  2-2

The same procedure was followed as in Production Example 2-1, except that the compound of Formula 2a (2.5 g, 5.4 mmol) synthesized in Production Example 2-1, which was not the compound of Formula 1, was used as the starting material, To obtain the compound of the following formula (2b) (1.6 g, 61%) having ¹ H NMR spectrum.

¹ H NMR (400 MHz, CDCl ₃ )? 7.69 (m, 4 H), 7.40 (m, 6 H), 4.35 (dd, J = 3.6, 7.6 Hz, (Br s, 1H), 2.65 (m, 1H), 1.96 (m, 1 H), 3.78 (ddd, J = 6.4, 10.4, 3.2 Hz, 2H) H), 1.71 (m, 1H), 1.54 (s, 3H)

(2b)

Manufacturing example  3-1

Compound (2a) (2.5 g, 5.4 mmol) synthesized in Preparation Example 2-1 was dissolved in methanol and cooled to -40 ° C. Sodium borohydride (NaBH ₄ ) (1.45 g, 38.4 mmol) was slowly added to the cooled solution. Thereafter, the reaction mixture was stirred under the same conditions, and after completion of the reaction was confirmed by TLC, an aqueous solution of saturated ammonium chloride (NH ₄ Cl) was added to the reaction mixture, the aqueous layer was extracted with ethyl acetate, and the organic layer was separated. The separated organic layer was washed thoroughly with water and brine, dried over magnesium sulfate (MgSO ₄ ), and filtered under reduced pressure to remove the remaining solid, which was then concentrated under reduced pressure. The resulting alcohol compound was separated and purified by silica gel chromatography to obtain an intermediate (2.54 g, 76%) which was dissolved in pyridine anhydride and cooled to 0 ° C. Trifluoromethanesulfonic anhydride (3.26 ml, 19.36 mmol) was added dropwise to the cooled mixture, and the mixture was stirred for 30 minutes under the same conditions to perform trifluoromethylsulfone oxidation. After completion of the reaction was confirmed by TLC, an aqueous solution of saturated ammonium chloride (NH ₄ Cl) was added to the reaction mixture, the aqueous layer was extracted with ethyl acetate, and the organic layer was separated. The separated organic layer was washed thoroughly with water and brine, dried over magnesium sulfate (MgSO ₄ ), and filtered under reduced pressure to remove the remaining solid and then concentrated under reduced pressure. The resulting trifluoromethylsulfone compound and 10 equivalents of sodium azide (NaN ₃ ) were mixed in anhydrous DMF, and the mixture was stirred under heating at 100 ° C. After stirring for about 4 hours, it was confirmed that the reaction was terminated by TLC. After adding water, the aqueous layer was extracted with ethyl acetate, and the organic layer was separated. The separated organic layer was washed thoroughly with water and brine, dried over magnesium sulfate (MgSO ₄ ), and filtered under reduced pressure to remove the remaining solid, which was then concentrated under reduced pressure. The azide compound thus obtained was isolated and purified by silica gel chromatography, and the resulting compound was dissolved in methanol. An appropriate amount of palladium / carbon was added to the solution, and the hydrogenation reaction was carried out by hydrogen substitution in the reaction vessel to reduce the azide group to an amine group. After the completion of the reaction was confirmed by TLC, the remaining solid was removed by filtration under reduced pressure, and the filtrate was concentrated under reduced pressure to obtain 1.6 g (61%) of a compound of the following formula (3a) having the following ¹ H NMR spectrum.

^{1 H NMR (400 MHz, CDCl} 3) δ 7.69 (m, 4 H), 7.40 (m, 6 H) 4.75 (dt, J = 2.8, 53.2 Hz, 1 H), 4.36 (m, 2 H), 3.78 (s, 3 H), 1.30 (s, 3 H), 1.06 (s, 3 H) , 9 H)

&Lt; EMI ID =

Manufacturing example  3-2

(2.117 g, 4.636 mmol) synthesized in Preparation Example 2-2, which was not the compound of Formula 2a, was used as the starting material, the temperature condition of the sodium borohydride reduction was adjusted to 0 ° C or lower, The procedure of Preparation Example 3-1 was repeated except that 1.89 g of sodium azide was used as the azide and 29.04 mmol of the azide was used and the temperature was heated to 60 ° C to obtain the following ¹ H NMR spectrum To obtain the compound of Formula 6b (1.28 g, 60%).

¹ H NMR (400 MHz, CDCl ₃ )? 7.69 (m, 4 H), 7.40 (m, 6 H), 4.35 (dd, J = 3.6, 7.6 Hz, (Br s, 1H), 2.65 (m, 1H), 1.96 (m, 1 H), 3.78 (ddd, J = 6.4, 10.4, 3.2 Hz, 2H) H), 1.71 (m, 1H), 1.54 (s, 3H)

&Lt; EMI ID =

Manufacturing example  4-1

(300 mg, 0.65 mmol) synthesized in Production Example 3-1 was reacted with 5-amino-4,6-dichloropyrimidine (1.85 g, 11.27 mmol) and di diisopropylethylamine (DIPEA) (6.54mL, 37.57mmol) and n - butanol was heated to 170 ℃ using that later, microwave mixing the (n -butanol) was stirred for 12 hours. After confirming that the reaction was completed by TLC, the reaction mixture was mixed with methanol and concentrated under reduced pressure. The residue was separated and purified by silica gel chromatography to obtain an intermediate (964 mg, 66%). The intermediate was dissolved in diethoxymethyl acetate and stirred at 140 ° C for 3 hours using a microwave. After that the checking by TLC the reaction was completed, the reaction mixture was mixed with methanol to having a ¹ H NMR spectrum as described below, to obtain separation in the back was concentrated under reduced pressure, silica gel chromatography, compounds of formula 8a (240mg, 61.6%) &Lt; / RTI &gt;

¹ H NMR (400 MHz, CDCl ₃ )? 8.78 (s, 1H), 8.32 (d, J = 2.4 Hz, 1H), 7.68 1 H), 1.95 (m, 2H), 1.59 (s, 1H), 5.08 (m, 2H), 4.62 3 H), 1.34 (s, 3 H), 1.06 (s, 9 H)

&Lt; Formula 4a &

Manufacturing example  4-2

The compound of Formula 3b (250 mg, 0.52 mmol) synthesized in Preparation Example 3-2 was dissolved in 1,4-dioxane (50 mL), and 4,6-dichloro-5- (120 mg, 0.65 mmol) and trimethylamine (0.55 ml, 3.86 mmol) were added. After the reaction mixture was refluxed for 2 days, the reaction was terminated by TLC, and the reaction mixture was concentrated under reduced pressure. The residue was separated and purified by silica gel chromatography to obtain an intermediate. Diethoxymethyl acetate (5 mL) was added to the intermediate, followed by stirring at 140 ° C for about 12 hours. After that the checking by TLC the reaction was completed, a mixture as the reaction mixture with methanol to having a ¹ H NMR spectrum as described below, to obtain separated then concentrated under reduced pressure, silica gel chromatography, compounds of formula 4b (100mg, 31%) &Lt; / RTI &gt;

¹ H NMR (400 MHz, CDCl ₃ )? 8.80 (s, 1H), 8.25 (d, J = 2.4 Hz, 1H), 7.70 m, 1 H), 5.16 (superimposed dd, J = 6.8 Hz, 1H), 4.54 (superimposed dd, J = 6.4 Hz, 1H), 3.80 (S, 3 H), 1.33 (s, 3 H), 1.07 (s, 9 H)

<Formula 4b>

Manufacturing example  5-1

The compound of Formula 4a (200 mg, 0.336 mmol) synthesized in Production Example 4-1 was dissolved in methanol, and a solution of trifluoroacetic acid (TFA) (5 mL) and water at 1: 1 was added dropwise at 0 ° C . After the reaction mixture was stirred at room temperature for 2 hours, it was confirmed by TLC that the reaction was terminated and concentrated under reduced pressure. The concentrated residue was separated and purified by silica gel chromatography to obtain an intermediate (80 mg, 75.4%).

Tert after the intermediate (70mg, 0.22mmol) was dissolved in butanol (tert -butanol), was transferred to a high-temperature closed container (stainless steel bomb reactor). Saturated ammonia / butanol was added dropwise to the mixed solution, and the reaction vessel was sealed and stirred at 120 ° C for 15 hours. After confirming that the reaction was completed by TLC, the reaction mixture was diluted with methanol and concentrated under reduced pressure. The concentrated residue was separated and purified by silica gel chromatography to obtain the following compound of formula (5a) (55 mg, 84.6%) having ¹ H NMR spectrum as described below.

^{1 H NMR (400 MHz, CD} 3 OD) δ 8.26 (d, J = 2.0 Hz, 1 H), 8.20 (s, 1 H), 5.11 (dt, J = 3.6, 55.2 Hz, 1 H), 4.99 ( ddd, J = 3.2, 9.6, 26.4 Hz, 1 H), 4.75 (m, 1 H), 4.04 (m, 1 H), 3.71 (m, 2 H), 2.38 (m, 1 H), 1.89 (m , 2 H)

&Lt; Formula 5a &

Manufacturing example  5-2

Except that the compound of the formula 4b (100 mg, 0.3 mmol) synthesized in Production Example 4-2, which was not the compound of the formula 4a, was used as a starting material and the mixture was stirred at 90 ° C for about 3 hours during the amination reaction The procedure of Preparation Example 5-1 was repeated to give the compound of Formula 5b (30 mg, 32%) having the ¹ H NMR spectrum as described below.

^{1 H NMR (300 MHz, CD} 3 OD) δ 8.27 (d, J = 2.1 Hz, 1H), 8.20 (s, 1H), 5.28 (m, 1H), 4.76 (m, 1H), 4.06 , 1H), 1.98 (m, 1H), 1.79 (m, 1H)

&Lt; Formula 5b >

Experimental Example  1 - Antiviral activity and cytotoxicity experiments

In order to examine the antiviral activity and cytotoxicity of the carbocyclic nucleoside derivative of the present invention, the following experiment was conducted.

Vero E6 cells were cultured and infected with CHIKV (Chikungunya Virus) LS3 and SFV (Semliki Forrest Virus) and titrated to obtain CHIKV inoculum and SFV inoculum, respectively.

The compound of Chemical Formula 5a synthesized in Preparation Example 5-1 was diluted with 200 μM in IMD (Iscove's modified Dulbecco's medium) containing 1% fetal calf serum (FCS) and an antibiotic. Vero E6 cells were cultured in 96-well plates containing 2 x 10 ⁴ cells per well. The cells were incubated overnight at 37 ° C, then 50 μl of the compound dilution was injected into each well, mixed with 100 μl of EMEM (Eagle's minimal essential medium) containing 2% FCS and 50 μl of the CHIKV inoculum. Subsequently, the cell-to-cell survival difference caused by virus-induced CPP (cytopathic effect) and compound side effects at 3 dpi (day postinfection) was analyzed using the CellTiter 96 Aqueous nonradioactive cell proliferation assay (Promega). The cytotoxicity by the compound treatment was confirmed by culturing the cells in the same manner as above except that a general medium was injected instead of the inoculated virus.

Using Graphpad Prism 5 software, cell viability for CPE and cell viability for compound adverse events were calculated for 50% effective concentration (EC ₅₀ ) and 50% cytotoxic concentration (CC ₅₀ ), respectively, based on the respective results . EC ₅₀ represents the minimum concentration of the compound for which 50% of the pathogen (virus) is inhibited, and CC ₅₀ represents the concentration of the compound when 50 pro of the host cell is killed. The degree of relative efficacy of the compounds against a particular virus was defined as selective (SI; CC ₅₀ / EC ₅₀ ).

Further, experiments were carried out in the same manner as above except that the compounds were mixed with the inoculum of SFV to calculate the EC ₅₀ , CC ₅₀ and SI values for the SFV of the compound of the formula 5a. The experiment for CHIKV above proceeded in the same way except that the compound of 5b was used to calculate the EC ₅₀ , CC ₅₀ and SI values of the compound of formula 5b.

Table 1 shows the EC ₅₀ , CC ₅₀ and SI values calculated by the above method.

Virus Type compound EC ₅₀ ( μM ) CC ₅₀ ( μM ) SI (CC ₅₀ / EC ₅₀ ) CHIKV  LS3 5a 0.2 > 250 > 1,000 5b 4.2 > 250 60 SFV 5a 6.4 > 250 > 30

As shown in Table 1, since the carbocyclic nucleoside derivative of the present invention has a low EC ₅₀ for the RNA viruses CHIKV and SFV and a high CC ₅₀ for the host cells, the inhibition of the virus and the prevention of viral diseases And as a pharmaceutical composition that is well suited for treatment.

Experimental Example  2 - SAH  Hydrolytic enzyme inhibitory effect experiment

In order to examine the SAH hydrolase inhibitory effect of the carbocyclic nucleoside derivative of the present invention, the following experiment was conducted.

A reaction mixture (250 μl) of 50 mM sodium phosphate (pH 8.0) and an AdoHcy hydrolase (2 μM as a monomer; 5 μM as a tetramer) was mixed with the compound of Formula 5a synthesized in Preparation Example 5-1 and the compound Preincubated with a compound of formula 5b for 10 minutes at 37 &lt; 0 &gt; C. After the pre-incubation, 100 μM AdoHcy was added to initiate the reaction and the reaction was allowed to proceed for 20 minutes. Thereafter, the reaction mixture was further cultured in 200 [mu] M DTNB and the maximum absorbance of the resulting product, TNB (5-thio-2-nitrobenzoic acid) at 412 nm, was measured with a spectrophotometer. The IC ₅₀ values of the compounds of formulas (5a) and (5b) were calculated using the maximum absorbance measured and the molar extinction coefficient of TNB (? 412 = 13700 M ^{- 1} cm ^{- 1} ).

compound IC ₅₀ ( μM ) 5a 0.1 5b 0.09

As shown in Table 2, since the carbocyclic nucleoside derivative of the present invention has a low IC ₅₀ value against the SAH hydrolase, it is suitable as a pharmaceutical composition having an antiviral effect through the SAH hydrolase inhibiting mechanism It can be seen that it can be used as a material.

While the present invention has been described in connection with certain exemplary embodiments, it is to be understood that the invention is not limited to the disclosed exemplary embodiments. It is therefore to be understood that the above-described embodiments are illustrative in all aspects and not restrictive.

Claims (6)

A carbocyclic nucleoside derivative represented by the following formula (A) or a pharmaceutically acceptable salt thereof.
&Lt; Formula (A)

(In the above formula (A)
R ₁ and R ₂ are each independently hydrogen or fluorine,
B is the following formula (B-1 or B-2)
<Formula B-1>

(In the above formula (B-1), X ₁ is a chlorine, a hydroxyl group, an amino group or an alkylamino group, and Y ₁ is a hydrogen atom or an amino group)
<Formula B-2>

(In the above formula (B-2), X ₂ is a hydroxyl group or an amino group, and Y ₂ is hydrogen, a methyl group or a halogen) The method according to claim 1,
Wherein R &lt; _{1 &gt;} is fluorine, or a pharmaceutically acceptable salt thereof. The method according to claim 1,
Wherein the formula (A) is a carbocyclic nucleoside derivative of the following formula (5a) or (5b): or a pharmaceutically acceptable salt thereof.
&Lt; Formula 5a &

&Lt; Formula 5b >

An antiviral agent comprising a carbocyclic nucleoside derivative represented by the following formula (A) or a pharmaceutically acceptable salt thereof.
&Lt; Formula (A)

(In the above formula (A)
R ₁ and R ₂ are each independently hydrogen or fluorine,
B is the following formula (B-1 or B-2)
<Formula B-1>

(In the above formula (B-1), X ₁ is a chlorine, a hydroxyl group, an amino group or an alkylamino group, and Y ₁ is a hydrogen atom or an amino group)
<Formula B-2>

(In the above formula (B-2), X ₂ is a hydroxyl group or an amino group, and Y ₂ is hydrogen, a methyl group or a halogen) 5. The method of claim 4,
Wherein R &lt; _{1 &gt;} is fluorine. 5. The method of claim 4,
Wherein the formula (A) is represented by the following formula (5a) or (5b).
&Lt; Formula 5a &

&Lt; Formula 5b >