KR100467312B1 - Antiviral isoxazoline and isoxazole nucleosides and method of their manufacture - Google Patents

Abstract

The present invention provides an isooxazolin (Formula 1) or isoxazole (Formula 2) having excellent antiviral efficacy having the formula:

Wherein R is hydrogen or an alkyl group of C1-C5, R 'is a hydrogen, alkyl or aryl group of C1-C10, X and Y are hydrogen or a protecting group;

Wherein R is hydrogen, a halogen element, or an alkyl group of C1-C5, R 'is hydrogen, an alkyl or aryl group of C1-C10, and X and Y are hydrogen or a protecting group.

Description

Antiviral isoxazoline and isoxazole nucleosides and method of their manufacture}

FIELD OF THE INVENTION The present invention relates to antiviral isoxazolines and isoxoxol nucleosides and methods for their preparation, and more particularly, to antiviral sugar-modified nucleosides containing isoxazoline and isoxazole part and It relates to a manufacturing method.

Viral infections are an important factor causing mortality and economic loss in humans. The number of antiviral therapies available today for viral infections is limited in number and not sufficient to cure all viruses. Currently less than 20 antiviral therapeutic agents (egidoxuridine, trifluridine, acyclovir, valacyclovir, famcyclovir, bidaribine, gangcyclovir, poscarnet, sipofovir, grapefilox, etc.) Has been approved by the US FDA to treat a small number of non-HIV viral diseases. Most of these drugs are modified nucleosides.

Since most of these drugs have the same mechanism of action, even simple variations make them resistant to the entire family of drugs. For example, the appearance of thymidine kinase deficiency variants of herpes simplex virus (HSV) in immunocompromised patients renders three acyclic nucleosides for HSV infection ineffective in treating such patients. Amantadine and rimatadine share the same mechanism of action, and influenza viruses quickly become resistant to the two drugs even in immunocompromised patients. In addition to being resistant to the virus, in AIDS patients, three drugs approved for cutomegaly virus (CMV) retinitis, i.e. sipofovir, gangcyclovir, and poscarnet, have significant toxicity. Therefore, the development of clinically effective and safe antiviral therapies is an inherent problem in suppressing viral infections.

Viruses are intracellular pathogens that share most of the nutrient requirements and synthetic pathways of the host cells they infect. Therefore, an ideal antiviral therapeutic agent effectively inhibits viral infection as a result of interaction with the virus's only target.

In principle, such selective antiviral activity can be achieved by developing a drug that is specific for the gene, enzyme, or cellular receptor specific to the virus. Selectivity can also be achieved by disrupting the replication of the virus through host mechanisms. Most of the currently approved antiviral agents have been developed by random screening, but as molecular-level details on the structure and replication of viruses have evolved in recent years, it is possible to logically design and develop antiviral agents using such information. Is making it possible.

Recent developments in chemistry and biology have led to a variety of new species that can be easily used in drug development, with molecules generated from combinatorial libraries, isolated from natural products, and collected in chemical databases. Searching those sources on the basis of knowledge can lead to the discovery of active compounds with unexpected structures.

It is known that many potentially effective antiviral agents are inherently active but appear to have unacceptable toxicity. By using new controlled dosage systems, optimal therapeutic responses, extended efficacy, and reduced toxicity can be achieved.

There are many viral diseases in the world and the number of infected patients is on the rise. Every year, 2.8 million people die of AIDS, and the number of people living with HIV was 34.3 million by the end of 2000, and an alarming 16,000 daily increase. Thus, HIV is one of the most dangerous viral diseases. There are a variety of drugs, but none can kill HIV, but only inhibit the growth of the virus. In addition, the virus has a high variability and is quickly immune to the same type of drug. HIV is one of the most cursed viruses now known. The second thing to mention is the herpes virus. This virus is also highly contagious. 80% of humans are positive for HSV-1 and 20 to 40% positive for HSV-2. Therefore, standard biopsy is done for HIV and herpes virus.

HIV has three different enzymes: protease, integrase and reverse transcriptase. Reverse transcriptase produces double stranded DNA molecules from one strand of RNA. All nucleoside drugs are inhibitors of reverse transcriptase. The virus does not have a repair system like eukaryotic cells. Therefore, the virus cannot replace misassembled nucleosides. Most commercially available antiviral nucleosides are free of hydroxyl groups at the 3 'position of the sugar moiety. This shows that RNA replication of viruses can occur from 5 'to 3'. Thus, once the viral reverse transcriptase is assembled to a nucleoside that does not have a hydroxyl group at the 3 'position, the enzyme can no longer grow viral DNA. The function of nucleoside drugs is to eliminate the chain reaction.

Since 1987, the year of the approval of the antiviral drug (3'-Azido-3'-deoxythymidine (AZT)), many laboratories have synthesized new modified nucleosides to find new antiviral drugs. Some of these have good antiviral effects, but only a few have received drug approval from the FDA. Most of these drugs are sugar-modified nucleosides. McGeegan's lab reports a variety of nucleoside drugs. For example, the lab synthesized two rings of pyrimidine, which has potent potency against the varella-zoster virus (VSV). Optimum alkyl chain lengths were C ₄ -C ₇ , C ₈ -C ₁₁ with p -phenyl rings (McGuigan et al., J. Med . Chem . 2000 , 43, 4993). The most active compound was 10,000 times more potent than the acylovir drug, and the selectivity index value was over 1,000,000. They also synthesized derivatives without p -phenyl rings (McGuigan et al., J. Med . Chem . 1999 , 42, 4479). The optimal chain length was C ₈ -C ₁₀ . The mechanism of action of this compound and the function of the p -phenyl ring are not clear, but the presence of the p -phenyl ring increased the activity 100-fold.

The first paper published on Ixoxazole nucleoside was in 1995. Up to this point, it had gone through the synthesis of widely distributed compounds in nature that simply replaced one carbon with oxygen to produce a furanose system. Many good anti-HIV drugs such as AZT, d4T (2 ', 3'-Didehydro-3'-deoxythymidine) and ddI (2', 3'-Dideoxyinosine) are drugs in the furanose form. By introducing a second heteroatom into the furanose ring, several new classes of nucleoside analogues can be synthesized, one of which is the isoxazole class.

Professor Zhao's lab published a paper on 1996 isoxoxazole nucleosides (Zhao and coworkers, Tetrahedron Lett. 1996 , 37, 4877). The efficacy of isoxazole nucleosides was not high. However, it was relatively easy to synthesize 6-chloropurine with oxime using a 1,3-2 polar cycloaddition reaction.

The technical problem to be achieved by the present invention is to design and synthesize an antiviral nucleoside variant that can be a novel leading material showing excellent efficacy by using a ringer reaction.

In order to achieve the above object, the present invention provides isoxazoline represented by Formula 1:

<Formula 1>

Wherein R is hydrogen or an alkyl group of C1-C5, R 'is hydrogen, an alkyl or aryl group of C1-C10, and X and Y are hydrogen or a protecting group.

Preferably, R is selected from the group consisting of hydrogen or methyl group, R 'is selected from the group consisting of methyl, benzyl, isobutyl, 2-methylpropyl, X and Y are independently of each other hydrogen, benzoyl (Bz) , t-butoxycarbonyl (BOC), and acetyl (Ac).

The present invention also provides isoxazole represented by the formula 2:

<Formula 2>

Wherein R is hydrogen, a halogen element, or an alkyl group of C1-C5, R 'is hydrogen, an alkyl or aryl group of C1-C10, and X and Y are hydrogen or a protecting group.

Preferably, R 'is selected from the group consisting of hydrogen, methyl, and fluorine, R' is selected from the group consisting of methyl, benzyl, isobutyl, 2-methylpropyl, X and Y are independently of each other hydrogen, Bz, BOC, and Ac.

For another object of the present invention,

(a) synthesizing an N-protected amino ester from an amino acid, reducing the ester to synthesize an N-protected amino aldehyde, from which synthesize an N-protected amino aldoxime;

(b) protecting both amino groups N ¹ and N ³ in the pyrimidine base with a protecting group followed by reaction with a weak acid to remove one protecting group to synthesize an N ³ -protected pyrimidine base;

(c) from pyrimidine base with allyl alcohol under Mitsunobu reaction conditions and with N ¹ -allyl N ³ -benzoyl pyrimidine (uracil / thymine) or with propargyl alcohol under similar reaction conditions N ¹ -propar Synthesizing the length N ³ -benzoyl pyrimidine (uracil / thymine / 5-fluorouracil); And

(d) the N ¹ -allyl N ³ -benzoyl pyrimidine (uracil / thymine) or N ¹ -propargyl N ³ -benzoyl pyrimidine (uracil / thymine / 5-fluorouracil) with the amino aldooxime [ 3 + 2] there is provided a method for synthesizing isooxazolin of formula (1) or isoxazole of formula (2) comprising the step of reacting with a ringer:

<Formula 1>

Wherein R is hydrogen or an alkyl group of C1-C5, R 'is hydrogen, an alkyl or aryl group of C1-C10, and X and Y are hydrogen or a protecting group.

<Formula 2>

Wherein R is hydrogen, a halogen element, or an alkyl group of C1-C5, R 'is hydrogen, an alkyl or aryl group of C1-C10, and X and Y are hydrogen or a protecting group.

In step (a), the N-protected amino ester is reduced with DIBAL in toluene at -78 ° C to -20 ° C, and the resulting N-protected amino aldehyde is dissolved in the presence of Na ₂ CO ₃ in aqueous methanol solution. Preference is given to synthesizing the aldooxime by treatment with hydroxyl amine hydrochloride.

In the step (b), the protecting group is a benzoyl group, and it is preferable to use 0.5 MK ₂ CO ₃ in water at the time of deprotection of the N ¹ -protecting group.

In the step (d), the ring polymerization reaction is preferably carried out in the presence of NaOCl.

Hereinafter, the present invention will be described in more detail step by step.

1.N-protected amino Aldooxime ( aldoxime ) of Synthesizing

This step is shown in Scheme 1 below. N-protected amino aldooxime (5a-5e) was synthesized in four steps from five amino acids (1a-1e). N-protected amino aldehydes are important in organic synthesis and are used in a variety of ways. In particular they have been used primarily to synthesize various peptide analogs. N-protected amino aldehydes are relatively unstable and readily turn into racemic mixtures in solution and during chromatography on silica gel. It is therefore recommended that these compounds be used soon after synthesis. Generally, amino aldehydes are synthesized from the corresponding amino acid derivatives by reduction methods. In this example, amino aldehydes (4a-4e) were synthesized from N-protected amino esters with diisobutylaluminum hydride (DIBAL, 2.3 equivalents) in toluene at -78 ° C for 5 minutes. In DIBAL reduction, small amounts of amino alcohol caused by overreduction were also isolated. Because amino aldehydes are unstable, they were converted directly to the corresponding amino aldooximes (5a-5e) by treatment with hydroxylamine hydrochloride in the presence of Na ₂ CO ₃ in aqueous methanol. Aldooxime was isolated as a white solid in high yield. They turned out to be mixtures of E and Z isomers.

2. N ³ -Protected Pyrimidine Synthesis of base

This step is illustrated by Scheme 2 below. The pyrimidine base has two amino groups N ¹ and N ³ and therefore one must be protected. In most cases the amine functionality of the pyrimidine base is protected with benzoyl (Bz) or trimethyl silyl (TMS) groups. TMS groups are used to increase the nucleophilicity of pyridine bases. In this example, a Bz group was used, and an excess of benzoyl chloride was used to protect two amine groups. The N ¹ benzoyl group of the resulting compound was stirred with a weak acid (0.5 MK ₂ CO ₃ in water) to selectively remove the protecting group. Pyrimidine with one N-protecting group removed also yielded a high yield, but there were always small amounts of two N-benzoyl derivatives. However, for the 5-fluoro uracil 6c, the reaction was complete.

3. Above Pyrimidine N from base ^One Allyl N ³ - Benzoyl Pyrimidine ( Uracil Of Thymine Step)

This step is illustrated by Scheme 3. The pyrimidine bases (6a-6c) were treated with allyl alcohol in the presence of diethylazodicarbonylate (DEAD) and triphenyl phosphine (Mitsunobu reaction conditions) to N ¹ -allyl N ³ -benzoyl pyrimidine Switched. N ¹ -allyl N ³ -benzoyl pyrimidine was isolated from diethyl hydrazinedicarbonylate (DEHD) and by-products, and triphenyl phosphine oxide was also formed in this reaction. The mixture was clarified twice by flash column chromatography with a different solvent system.

4. N from the pyrimidine base ^One Propargyl N ³ -Benzoyl pyrimidine (uracil / thymine) / 5-fluorouracil)

This step is illustrated by Scheme 4. N ¹ -propargyl N ³ -benzoyl pyrimidine (8a-8c) was synthesized from N ³ -benzoyl pyrimidine (6a-6c) under similar reaction conditions used for allyl derivatives. However, unlike allyl derivatives, because of the high solubility of these products, they could be well purified in a relatively simple manner. 5-fluoro uracil is a known inhibitor of enzymes that methylate uridine bases, namely thymidylate synthetase.

5. N above ^One Allyl N ³ -Benzoyl pyrimidine (uracil / thymine) or N ^One Propargyl N ³ 3- [1-N- () from benzoyl pyrimidine (uracil / thymine / 5-fluorouracil) tert -Butyloxycarbonyl ) Amino-3-R ']-5-[(R / S)-(N ³ -Benzoyl) -methylene-base] -isoxazoline (9) or 3- [1- N- ( tert -Butyloxycarbonyl) amino-3-R ']-5-[(N ³ -Benzoyl) -methylene-base] -isoxazole Synthesizing 10, respectively

The yield of the cycloaddition reaction depends on many factors such as solubility of the reactants, protecting groups present on the pyrimidine. The yield of the cycloaddition reaction is increased by changing the solvent from methylene chloride to THF. Another important observation is that the yield of the cycloaddition reaction from the double bond dipolaropropyl (N ¹ -allyl N ³ -benzoyl pyrimidine) is its triple bond analog (N ¹ -propargyl N ³ -benzoyl pyrimidine). It is always higher than the yield of ring reaction. In this example, a library of cyclization reaction products was synthesized in combination with five different oximes and four dipolar profiles (7a, 7b, 8a, 8b).

6. 3- [1-N-amino-2- Phenylethyl ] -5- (N3- Benzoyl )- Methylene - Thymine - Ixoxazole Synthesis of 11

All of the aforementioned cycloaddition products have two protecting groups, Boc and Bz, which can protect the functional groups. Thus, it is necessary to deprotect one or two protecting groups, which may increase their antiviral activity. Unfortunately, the antiviral activity of the compound from which Boc was removed completely lost that activity. These observations indicate that protecting groups are of great antiviral drug candidates. Other observations indicate that compounds with large R groups (phenylalanine, leucine) have better activity than compounds with small R groups (alanine, valine). This relationship between structure and activity can be explained by the different mechanisms of action of these compounds from the general mechanism of virus growth inhibition.

7. 3- [1-N-amino-2-phenylethyl] -5- [methylene-5-fluoro-uracil] -isoxazole (12a) and 3- [1-N-amino-3-phenylethyl ] -5- [methylene-5-fluoro-uracil] -isoxazole Synthesis of 12b

In accordance with the above observations, it was investigated whether deprotecting the protecting groups of all of these functional groups (N ³ of the pyrimidine base and amino groups of the amino acid portion) had a positive or negative effect on antiviral activity. Completely deprotected compound (11) was synthesized as follows.

8. 3- [1-N- (benzoyl) amino-2-phenylethyl] -5-[(N ³ -Benzoyl) -methylene-thymine] -isoxazole Synthesis of 13

After recognizing that compound (11) from which the Boc protecting group was taken off showed surprising activity, the Boc group was cleaved and protected the free amine with the benzoyl group. Compound (13) showed no activity against polio virus. This shows that different protecting groups with completely different 3D structures can have an antiviral effect. Observing the antiviral data of the various compounds obtained so far, it can be concluded that the properties of the amine protecting group (Boc or benzoyl) play an important role.

The following table shows the bioassay data of the compounds produced in the above synthesis steps. Antiviral activity (anti-HIV, anti-HSV, anti-Polio activity) is expressed as EC ₅₀ value. Antiviral activity and cytotoxicity values were measured by the Korea Research Insitute of Chemical Technology in Daejeon. EC ₅₀ is defined as the effective concentration of the drug that inhibits 50% of the growth rate of the virus in μg / ml. SI values are the ratio of cytotoxic CC ₅₀ and EC ₅₀ . ND = Not detected.

In the table, ROD and IIIB are a type of HIV virus, Cox.B3 is Coxsakie B3, and PV-1 represents Poliovirus-1.

In the table it can be seen that the excellent antiviral efficacy of compound 10b.

In the table it can be seen that the excellent antiviral efficacy of compound 10d.

In the table it can be seen that the excellent antiviral efficacy of compound 9c.

(Example)

N- BOC Amino acids (2)

Amino acid (10 mmol) was dissolved in a solution containing 1 N NaOH (20 ml) and dioxane (20 ml). To this solution, (BOC) ₂ O (11 mmol) was added and stirred at room temperature for 4 hours. After completion of the reaction, the solution was cooled in ice water, acidified to pH 3 with saturated potassium bisulfite solution and extracted between EA (Ethyl acetate) and water. The organic phase was dried over magnesium sulfite and concentrated under reduced pressure to give a colorless oil. The resulting protected amino acid was used in the next step without further purification.

N- BOC -L-amino acid methyl Ester (3)

Method I: phenylalanine, alanine

To the N-BOC-L-amino acid solution was added dry methanol (3.5 ml per mmol of N-BOC-L-amino acid) and sulfuric acid (0.025 equiv). The reaction mixture was stirred at room temperature for 24 hours. After addition of EA the solution was cooled in ice water. After neutralization with 5% sodium bicarbonate solution, the solution was extracted with EA and the organic phase was dried over sodium sulfate. Concentrated under reduced pressure, and the residue was purified by flash column chromatography (eluent: hexane-EA 3: 1).

Method II: Leucine, Isoleucine, Valine

To the N-BOC-L-amino acid solution in DMF was added iodomethane (3 equiv) and potashite carbonate (2.5 equiv) and the mixture was stirred at room temperature for 5 hours. The solution was extracted with ether. The organic phase was washed five times with water, dried over magnesium sulphate and concentrated under reduced pressure. The residue was purified by flash column chromatography (eluent: hexane-EA 3: 1).

N- BOC -L-amino aldehyde (4)

Diisobutylaluminum hydride (1.5 M solution in toluene) in pre-cooled (-78 ° C.) N-BOC-L-amino acid methyl ester in dry toluene (2 ml per mmol of N-BOC-L-amino acid methyl ester) 2 equivalents) was added dropwise over 30 minutes. Methanol was carefully added after 5 minutes until no reaction of diisobutylaluminum hydride was observed. The mixture was poured into a pre-cooled (0 ° C.) aqueous citric acid solution. After stirring for 1 hour, the mixture was extracted with EA. The organic phase was collected, dried over magnesium sulphate, filtered and concentrated under reduced pressure. The resulting amino aldehyde was used for the next step without further purification.

N- BOC -L-amino Aldooxime (5)

To a pre-cooled solution of aldehyde (4 ° C.) in methanol / water (1: 1 volume / volume) was added sodium carbonate (1 equiv) and hydroxylamine hydrochloride (2 equiv). After stirring for 8 hours, the mixture was concentrated under reduced pressure to reduce the volume in half. The mixture was then extracted with EA, dried over magnesium sulphate and filtered. The oxime was recrystallized and purified.

N- ( tert - Butyloxycarbonyl ) -L- Alanial Oxime (5a)

(Yield 79%), ¹ H NMR (CDCl ₃ ) δ 7.03 (d, J = 4.4 Hz, 1H), 7.3 (br, 1H), 4.80 (br, 1H), 4.3 (br, 1H), 1.39 ( s, 9H), 1.25 (d, J = 6.8 Hz, 3H); ¹³ C NMR (CD ₃ OD) δ 157.9, 154.6, 80.5, 43.9, 29.0, 17.8; MS (m / e) 188 (M ⁺ ); mp 144.5-145.5 ° C.,

N- ( tert - Butyloxycarbonyl ) -L- Phenylalanial Oxime (5b)

(Yield 77%), IR (CHCl ₃ , cm ⁻¹ ) 3338, 2977, 1690, 1506, 1367, 1253, 1168, 775; ¹ H NMR (CDCl ₃ ) δ [7.45 (d, J = 4.1 Hz), 5.92 (br), (1H)], 7.33-7.12 (m, 5H), 5.08 (br, 1H), 4.55 (br, 1H ), 3.04 (br, 2H), 1.42 (s, 9H); MS (m / e) 264 (M ⁺ ); mp 164.5-165.5 ° C.

N- ( tert - Butyloxycarbonyl ) -L- Ballinal Oxime (5c)

(Yield 69%), ¹ H NMR (CD ₃ OD) δ6.52 (d, J = 6.8 Hz, 1H), 4.66 (m, 1H), 1.91 (m, 1H), 1.43 (s, 9H), 0.93 (d, J = 6.8 Hz, 3H), 0.91 (d, J = 6.8 Hz, 3H); ¹³ C NMR (CD ₃ OD) δ 158.3, 152.0, 80.4, 52.4, 32.4, 29.0, 19.5, 18.9; mp 154-155 ° C.

N- ( tert - Butyloxycarbonyl ) -L- Isoryu Signal Oxime (5d)

(Yield 70%), ¹ H NMR (CDCl ₃ ) δ 7.44 (d, J = 7.8 Hz, 1H), 7.3 (br, 1H), 6.2 and 5.2 (2d, J ₁ = 7.5 Hz, J ₂ = 6.9 Hz, 1H), 4.3 (m, 1H), 1.44 (s, 9H), 1.13 (m, 1H), 0.92 (m, 8H).

N- ( tert - Butyloxycarbonyl ) -L- Ryusinnal Oxime (5e)

(Yield 65%), ¹ H NMR (CDCl ₃ ) δ7.78 (br, 1H), 7.39 (d, J = 4.8 Hz, 1H), 4.76 (br, 1H), 4.31 (br, 1H), 1.73- 1.64 (m, 2H), 1.42 (s, 10H), 0.92 (d, J = 6.3 Hz, 6H); MS (m / e) 230 (M ⁺ ); mp 150-151 ° C.

N ³ - Benzoyl - Thymine /- Uracil / -5 FU

A solution of pyrimidine base (20 mmol), benzoyl chloride (55 mmol), dry toluene (20 ml) and dry pyridine (20 ml) was stirred at room temperature for 24 hours and then concentrated under reduced pressure at 80 ° C. The resulting residue was separated between MC (Methylene Chloride) and water. The organic phase was collected and concentrated. The resulting residue was dissolved in a mixture of aqueous potassium carbonate solution (0.5 M, 40 ml) and dioxane (80 ml) and stirred at room temperature for 1 hour. Then, the pH was lowered to about 5 by adding acetic acid on ice. The mixture was concentrated under reduced pressure and the residue was stirred with saturated aqueous sodium carbonate solution (100 ml). After 1 hour, the white solid product was filtered off and washed with cold water. The residue was purified by flash column chromatography (eluent: MC: methanol 50: 1) to give a white solid (yield 75-85%) compound (6a-6c).

N ³ - Bz - Thymine (6a)

(Yield 85%), 1 H NMR [(CD ₃ ) ₂ SO] δ 11.40 (br, 1 H), 7.94 (m, 2 H), 7.77 (t, J = 7.3, 1 H), 7.58 (m, 3 H), 1.82 (s, 3 H); ¹³ C NMR [(CD ₃ ) ₂ SO] δ 170.2, 163.6, 150.0, 138.8, 135.4, 131.4, 130.2, 129.5, 107.9, 11.7; mp 178-180 ° C.

N ³ - Bz - Uracil (6b)

(Yield 81%), ¹ H NMR [(CD ₃ ) ₂ SO] δ 11.62 (br, 1H), 7.96 (m, 2H), 7.77 (t, J = 7.4 Hz, 1H), 7.67 (d, J = 7.7 Hz, 1H), 7.6 (t, J = 7.8 Hz, 2H), 5.75 (d, J = 7.7 Hz, 1H); ¹³ C NMR [(CD ₃ ) ₂ SO] δ 170.4, 163.3, 150.4, 143.7, 135.8, 131.7, 130.6, 129.8, 100.4; mp 173.5-175 ° C.

N ^One Allyl N ³ - Benzoyl - Thymine /- Uracil

To a pre-cooled (0 ° C.) triphenylphosphine (18 mmol) solution in dry THF (150 ml) was added diethyl azodicarbonylate (DEAD) (18 mmol) and stirred for 30 minutes. Thereafter, a solution of N ³ -Bz protected pyrimidine base (10 mmol) and allyl alcohol (30 mmol) in dry THF was added to the solution by a syringe and stirred at room temperature for 8 hours. The reaction mixture was separated between MC and water. The organic phase was concentrated in MC and redissolved. After 24 hours, most of the byproduct of the reaction, diethyl hydrazinecarbonylate (DEHD), was formed. After filtration, the filtrate was concentrated and purified twice by flash column chromatography (first eluent: hexane: EA: MC 5: 1: 1) (second eluent: MC: methanol 100: 1) to give a white solid (80). -87% yield) Compounds (7a, 7b) were obtained.

N ³ - Bz -N ^One Allyl Thymine (7a)

(Yield 87%), ¹ H NMR (CDCl ₃ ) δ7.90 (br, 2H), 7.63 (br, 1H), 7.47 (br, 2H), 7.11 (s, 1H), 5.92 (m, 1H), 5.38 (br, 2H), 4.37 (d, J = 7.17 Hz, 2H), 2.0 (s, 2H); ¹³ C NMR (CDCl ₃ ) δ 169.4, 163.5, 150.2, 139.7, 135.3, 132.1, 131.9, 130.8, 129.5, 120.3, 111.4, 50.5, 12.8; mp 127-128.5 ° C.

N ³ - Bz -N ^One Allyl Uracil (7b)

(Yield 80%), ¹ H NMR (CDCl ₃ ) δ 7.81 (br, 2H), 7.62 (br, 1H), 7.47 (br, 2H), 5.80 (m, 1H), 5.32 (br, 2H), 4.35 (d, J = 5.88 Hz, 2H).

N ³ - Bz -N ^One - Propargyl - Thymine /- Uracil / 5- Fluoro Uracil

DEAD (18 mmol) was added to a pre-cooled (0 ° C.) triphenylphosphine (18 mmol) solution in dry THF (150 ml) and stirred for 30 minutes. Thereafter, a solution of N ³ -Bz protected pyrimidine base (10 mmol) and propargyl alcohol (30 mmol) in dry THF was added to the solution by syringe and stirred at room temperature for 8 hours. The reaction mixture was separated between MC and water. The filtrate was concentrated and purified twice by flash column chromatography (first eluent: hexane: EA: MC 5: 1: 1) (second eluent: MC: methanol 100: 1) to give a white solid (76-82%). Yield) Compound (8a-8c) was obtained.

N ³ - Bz -N ^One - Propargyl - Thymine (8a)

(Yield 82%), ¹ H NMR (CDCl ₃ ) δ7.91 (br, 2H), 7.62 (br, 1H), 7.48 (br, 2H), 7.35 (s, 1H), 4.55 (d, J = 2.61 Hz, 2H), 2.49 (t, J = 2.57 Hz, 1H), 1.98 (d, J = 1.09 Hz, 3H); ¹³ C NMR (CDCl ₃ ) δ 169.1, 163.3, 149.8, 138.5, 135.4, 132.0, 130.9, 129.6, 126.3, 111.9, 76.5, 76.0, 37.3, 12.8; mp 185.5-188 ° C.

N ³ - Bz -N ^One - Propargyl - Uracil (8b)

(Yield 76%), ¹ H NMR (CDCl ₃ ) δ 7.89 (br, 2H), 7.63 (br, 1H), 7.49 (br, 2H), 7.44-7.54 (d, 1H), 5.81 (d, J = 8.09 Hz, 1H), 4.52 (d, J = 2.58, 2H), 2.50 (t, J = 2.55 Hz, 1H); ¹³ C NMR (CDCl ₃ ) δ 169.0, 162.6, 149.7, 143.0, 135.7, 131.7, 130.9, 129.6, 103.1, 76.5, 76.1, 37.7.

3-[(1S) -N- ( tert -Butyloxycarbonyl) amino-3-R ']-5-[(R / S)-(N ³ -Benzoyl) -methylene -Pyrimidine] -isoxazoline (9) and 3-[(1S) -N ( tert - Butyloxycarbonyl ) Amino-3-R ']-5-[(N ³ - Benzoyl )- Methylene - Pyrimidine ]- Ixoxazole 10

To an aqueous solution of allyl- (7) or propargyl-base (8) (1 mmol) in THF (20 ml) was added dropwise an aqueous amino aldooxime (5) and 4% NaOCl solution over very slowly. The reaction mixture was extracted with MC and water, and the organic phase was dried over magnesium sulphate, filtered and concentrated under reduced pressure. The residue was purified by flash column chromatography (eluent: hexane: EA: MC 3: 1: 1) to give a white solid. Recrystallized from MC / hexane. (Yield 60-92%).

3- [1-N- ( tert - Butyloxycarbonyl ) Amino-ethyl] -5-[(R / S)-(N ³ - Benzoyl )- Methylene - Thymine ]- Isoxazoline (9a)

(Yield 45%), ¹ H NMR (CDCl ₃ ) δ 7.89 (br, 2H), 7.62 (br, 1H), 7.48 (br, 2H), 7.24-7.21 (br, 1H), 4.90-4.75 (m) , 2H), 4.50 (m, 1H), 4.08 (m, 1H), 3.59 (m, 1H), 3.14 (dd, J ₁ = 10.53 Hz, J ₂ = 17.49 Hz, 1H), 2.77 (m, 1H) , 1.94 (d, J = 1.12 Hz, 3H), 1.43-1.38 (br, 12H); ¹³ C NMR (CDCl ₃ ) δ 165.7, 164.1, 160.0, 150.5, 141.6, 135.4, 130.9, 129.6, 127.1, 121.2, 51.5, 38.6, 28.7, 19.2, 15.9, 12.8; Calcd for MS (m / e) [M ⁺ + Na] 479.19, found 479.34.

3- [1-N- ( tert - Butyloxycarbonyl Amino-2- Methylbutyl ] -5-[(R / S)-(N ³ - Benzoyl )- Methylene - Thymine ]- Isoxazoline (9c)

(Yield 91%), ¹ H NMR (CDCl ₃ ) δ7.93 (br, 2H), 7.63 (br, 1H), 7.53 (br, 2H), 7.26 (br, 1H), 4.88 (m, 2H), 4.39 (m, 1H), 4.1 (m, 1H), 3.64 (m, 1H), 3.12 (m, 1H), 2.74 (m, 1H), 1.96 (d, J = 1.12 Hz, 3H), 1.75 (m , 1H), 1.59 (m, 2H), 1.45 (d, J = 1.32 Hz, 9H), 1.26 (m, 1H), 0.94 (m, 6H); Calcd for MS (m / e) [M ⁺ + Na] 521.238, experimental 521.36; mp 158-160 ° C.

3- [1-N- ( tert - Butyloxycarbonyl Amino-2- Methylpropyl ] -5-[(R / S)-(N ³ - Benzoyl )- Methylene - Thymine ]- Isoxazoline (9e)

(Yield 82%), 1 H NMR (CDCl 3) δ7.90 (br, 2H), 7.62 (br, 1H), 7.47 (br, 2H), 7.22 (s, 1H), 4.96 (br, 2H), 4.31 ( br, 1H), 4.05 (m, 1H), 3.63 (m, 1H), 3.11 (m, 1H), 2.74 (m, 1H), 2.05 (m, 1H), 1.95 (d, J = 6.81, 3H) , 1.43 (s, 9 H), 0.95 (m, 6 H); Calcd for MS (m / e) [M ⁺ + Na] 507.223, found 507.37; mp 167-170 ° C.

3- [1-N- ( tert - Butyloxycarbonyl ) Amino-ethyl] -5-[(R / S)-(N ³ - Benzoyl )- Methylene - Uracil ]- Isoxazoline (9f)

(Yield 34%), ¹ H NMR (CDCl ₃ ) δ7.92 (br, 2H), 7.64 (br, 1H), 7.49 (br, 2H), 7.41 (m, 1H), 5.78 (m, 1H), 5.11-4.96 (m, 1H), 4.49 (m, 1H), 4.08 (m, 1H), 3.65 (m, 1H), 3.13 (m, 1H), 2.76 (m, 1H), 1.42 (s, 9H) , 1.37 (d, J = 6.87, 3H); Calcd for MS (m / e) [M ⁺ + Na] 465.175, found 465.37; mp 77-78 ° C.

3- [1-N- ( tert - Butyloxycarbonyl Amino-2- Methylbutyl ] -5-[(R / S)-(N ³ - Benzoyl )- Methylene - Thymine ]- Isoxazoline (9h)

(Yield 88%), ¹ H NMR (CDCl ₃ ) δ7.94 (br, 2H), 7.63 (br, 2H), 7.52 (br, 2H), 7.39 (d, J = 7.98, 1H), 5.79 (d , J = 8.01, 1H), 4.89 (br, 2H), 4.37 (br, 1H), 4.04 (m, 1H), 3.69 (dd, J ₁ = 7.34, J ₂ = 14.42, 1H), 3.11 (dd, J ₁ = 10.62, J ₂ = 17.63, 1H), 2.75 (m, 1H), 1.55 (m, 1H), 1.42 (s, 9H), 1.24 (m, 1H), 1.12 (m, 1H), 0.94 ( m, 6H); calcd for MS (m / e) [M ⁺ + Na] 541.207, found 541.36; mp 107-108 ° C.

3- [1-N- ( tert - Butyloxycarbonyl Amino-2- Methylpropyl ] -5-[(R / S)-(N ³ - Benzoyl )- Methylene - Uracil ]- Isoxazoline (9j)

(Yield 70%), ¹ H NMR (CDCl ₃ ) δ 7.89 (br, 2H), 7.64 (br, 1H), 7.49 (br, 2H), 7.39 (m, 1H), 5.77 (m, 1H), 4.90 (br, 2H), 4.30 (br, 1H), 4.03 (m, 1H), 3.66 (m, 1H), 3.10 (m, 1H), 2.73 (m, 1H), 2.20 (br, 1H), 1.42 (s, 9H), 0.94 (m, 6H); Calcd for MS (m / e) [M ⁺ + Na] 493.207, found 493.38.

3- [1-N- ( tert - Butyloxycarbonyl ) Amino-ethyl] -5-[(R / S)-(N ³ - Benzoyl )- Methylene - Thymine ]- Ixoxazole (10a)

(Yield 63%), ¹ H NMR (CDCl ₃ ) δ7.91 (br, 2H), 7.67 (br, 1H), 7.51 (br, 2H), 6.30 (s, 1H), 4.99 (d, J = 5.16 Hz, 2H) 5.01-4.93 (br, 1H), 1.98 (s, 3H), 1.51 (d, J = 6.69, 3H), 1.51 (s, 9H); Calcd for MS (m / e) [M ⁺ + Na] 479.199, found 470.34.

3- [1-N- ( tert - Butyloxycarbonyl Amino-2- Phenylethyl ] -5-[(R / S)-(N ³ - Benzoyl )- Methylene - Thymine ]- Ixoxazole (10b)

(Yield 73%), ¹ H NMR (CDCl ₃ ) δ7.91 (br, 2H), 7.66 (br, 1H), 7.51 (br, 2H), 7.24 (br, 3H), 7.13 (br, 2H), 7.27-7.12 (s, 1H), 6.07 (s, 1H), 4.96 (d, J = 3.63,2H), 5.10-4.90 (br, 2H), 3.15 (d, J = 6.33, 2H), 1.98 (d , J = 0.99, 3H), 1.58 (s, 3H), 1.39 (s, 9H); mp 167-169 ℃

3- [1-N- ( tert - Butyloxycarbonyl Amino-2- Methylbutyl ] -5-[(R / S)-(N ³ - Benzoyl )- Methylene - Thymine ]- Ixoxazole (10c)

(Yield 41%), ¹ H NMR (CDCl ₃ ) δ 7.89 (br, 2H), 7.66 (br, 1H), 7.50 (br, 2H), 7.25 (d, J = 1.11, 1H), 6.23 (s , 1H), 5.08 (br, 1H), 4.99 (d, J = 4.89, 2H), 4.79 (br, 1H), 1.97 (d, J = 0.87, 3H), 1.85 (br, 1H), 1.3-1.15 (m, 2H), 1.43 (s, 9H), 0.92 (m, 6H); Calcd for MS (m / e) [M ⁺ + Na] 519.236, found 519.34; mp 184.5-185.8 ° C.

3- [1-N- ( tert - Butyloxycarbonyl Amino-3- Methylbutyl ] -5-[(R / S)-(N ³ - Benzoyl )- Methylene - Thymine ]- Ixoxazole (10d)

(Yield 62%), ¹ H NMR (CDCl ₃ ) δ7.91 (br, 2H), 7.66 (br, 1H), 7.50 (br, 2H), 7.25 (d, J = 1.02, 1H), 6.27 (s , 1H), 4.99 (d, J = 3.57, 2H), 4.90-4.80 (br, 2H), 1.98 (s, 3H), 1.67 (m, 2H), 1.44 (s, 9H), 0.96 (m, 6H ); Calcd for MS (m / e) [M ⁺ + Na] 519.236, found 519.33; mp 168-170 ° C.

3- [1-N- ( tert - Butyloxycarbonyl Amino-2- Methylpropyl ] -5-[(R / S)-(N ³ - Benzoyl )- Methylene - Thymine ]- Ixoxazole (10e)

(Yield 71%), ¹ H NMR (CDCl ₃ ) δ 7.89 (br, 2H), 7.66 (br, 1H), 7.50 (br, 2H), 7.24 (d, J = 0.99, 1H), 6.24 (s , 1H), 4.99 (d, J = 3.57, 2H), 5.10-4.90 (br, 1H), 4.75 (br, 1H), 1.99 (br, 1H), 1.44 (s, 9H), 0.94 (m, 6H ); Calcd for MS (m / e) [M ⁺ + Na] 505.207, found 505.26; mp 181.5-183.1 ° C.

3- [1-N- ( tert - Butyloxycarbonyl ) Amino-ethyl] -5-[(R / S)-(N ³ - Benzoyl )- Methylene - Uracil ]- Ixoxazole (10f)

(Yield 66%), ¹ H NMR (CDCl ₃ ) δ7.91 (br, 2H), 7.64 (br, 1H), 7.51 (br, 2H), 7.42 (d, J = 8.01, 1H), 6.29 (s , 1H), 5.86 (d, J = 8.01, 1H), 5.04 (m, 1H), 5.00 (d, J = 4.68, 2H), 4.93 (m, 1H), 1.49 (d, J = 6.9, 3H) , 1.43 (s, 9 H); Calcd for MS (m / e) [M ⁺ + Na] 463.173. Found 468.28.

3- [1-N- ( tert - Butyloxycarbonyl Amino-2- Methylbutyl ] -5-[(R / S)-(N ³ - Benzoyl )- Methylene - Thymine ]- Ixoxazole (10h)

(Yield 69%), ¹ H NMR (CDCl ₃ ) δ7.91 (br, 2H), 7.67 (br, 2H), 7.51 (br, 2H), 7.42 (d, J = 8.04, 1H), 6.24 (s , 1H), 5.89 (d, J = 8.04, 1H), 5.02 (d, J = 4.86, 2H), 5.10-4.90 (br, 1H), 4.82 (br, 1H), 1.89 (m, 1H), 1.44 (s, 9H), 1.5-1.1 (m, 2H), 0.92 (m, 6H); Calcd for MS (m / e) [M ⁺ + Na] 505.207, found 505.36; mp 145-146.6 ° C.

3- [1-N- ( tert - Butyloxycarbonyl Amino-2- Methylpropyl ] -5-[(R / S)-(N ³ - Benzoyl )- Methylene - Uracil ]- Ixoxazole (10j)

(Yield 74%), ¹ H NMR (CDCl ₃ ) δ 7.89 (br, 2H), 7.64 (br, 1H), 7.48 (br, 2H), 7.39 (d, J = 8.03, 1H), 6.22 (s , 1H), 5.87 (d, J = 8.03, 1H), 5.00 (d, J = 3.14, 2H), 5.06-4.94 (br, 1H), 4.71 (br, 1H), 2.09 (m, 1H), 1.41 (s, 9H), 0.90 (m, 6H); Calcd for MS (m / e) [M ⁺ + Na] 491.205, found 491.47; mp 79-80.5 ° C.

3- [1-N- ( tert - Butyloxycarbonyl Amino-3- Methylbutyl ] -5-[(R / S)-(N ³ - Benzoyl )- Methylene -5- Fluoro - Uracil ]- Ixoxazole (10n)

(Yield 58%), ¹ H NMR (CDCl ₃ ) δ 7.88 (br, 2H), 7.65 (br, 1H), 7.50 (br, 2H), 7.51-4.45 (s, 1H), 7.23 (br, 3H ), 7.12 (m, 2H), 6.11 (s, 1H), 5.26 (br, 1H), 5.08 (br, 1H), 4.91 (s, 2H), 3.11 (m, 2H), 1.40 (s, 9H) .

3- [1-amino-2- Phenylethyl ] -5-[(N ³ - Benzoyl )- Methylene - Thymine ]- Ixoxazole (11)

Trifluoroacetic acid (TFA) (1.5 ml) was added to a 3- [1-amino-2-phenylethyl] -5-[(N ³ -benzoyl) -methylene-thymine] -isoxazole solution in MC (10 ml) It was. The solution was stirred at room temperature for 8 hours. The reaction mixture was then neutralized with saturated sodium hydrogen carbonate solution, extracted with MC, the organic phase was dried over magnesium sulfate, filtered and concentrated under reduced pressure. The residue was purified by flash column chromatography (eluent: MC: methanol 20: 1) to give a white solid. (Yield 55%). ¹ H NMR (MeOH) δ 7.69 (br, 2H), 7.52 (br, 1H), 7.52-7.51 (s, 1H), 7.42 (br, 2H), 7.26 (br, 3H), 7.18 (br, 2H ), 6.34 (s, 1H), 5.55 (m, 1H), 5.04 (s, 2H), 3.15-3.35 (m, 2H), 1.87 (s, 3H); ¹³ C NMR [(CH ₃ ) ₂ O] δ 167.5, 167.4, 164.8, 164.6, 150.7, 140.5, 137.3, 133.8, 131.2, 128.9, 128.0, 127.9, 126.3, 126.2, 110.1, 101.5, 42.3, 10.9.

3- [1-N-amino-2 Phenylethyl ] -5- [ Methylene -5- Fluoro - Uracil ]- Ixoxazole And N-amino-3- Methylbutyl ] -5- [ Methylene -5- Fluoro - Uracil ]- Ixoxazole (12)

3 [(1S) -N- ( tert -butyloxycarbonyl) amino-benzyl ']-5-[(R / S)-(N3-benzoyl) -methylene-5-fluoro- in MC (30 ml) Hydrogen fluoride pyridine (6 ml) was added to a solution of uracil] -isoxazole (1.68 mmol). The reaction mixture was stirred at room temperature for 24 hours. The reaction mixture was then neutralized with sodium carbonate and concentrated under reduced pressure. The residue was purified by flash column chromatography on reversed phase silica (eluent: water: acetonitrile 20: 1) to give a white solid.

3- [1-N-amino-2 Phenylethyl ] -5- [ Methylene -5- Fluoro - Uracil ]- Ixoxazole (12a)

(Yield 85%), ¹ H NMR (D ₂ O) δ 7.39 (d, J = 5.58 Hz, 1H), 7.08 (br, 3H), 6.63 (br, 2H), 6.07 (s, 1H), 4.78 (s, 2H), 4.13 (t, 1H), 2.84 (m, 2H); ¹³ C NMR (D ₂ O) δ 169.2, 168.3, 167.7, 158.2, 144.0, 140.8, 137.5, 129.8, 128.9, 127.1, 101.9, 49.6, 44.6, 42.7.

3- [1-N-amino-3- Methylbutyl ] -5- [ Methylene -5- Fluoro - Uracil ]- Ixoxazole (12b)

(Yield 68%), ¹ H NMR (D ₂ O) δ 7.57 (d, J = 5.55 Hz, 1H), 6.27 (s, 1H), 4.91 (s, 2H), 3.97 (t, 1H), 1.46 (m, 2H), 1.36 (m, 1H), 0.75 (m, 6H)

3 [(1S) -N- (benzoyl) amino-3-R ']-5-[(R / S)-(N ³ -Benzoyl) -methylene-thymine] -isoxazole (13)

To the solution of compound (10) in pyridine was added benzoyl chloride (2 equiv). The reaction mixture was stirred for 24 hours and concentrated under reduced pressure. The residue was separated between MC and water. The organic phase was washed with water, dried over magnesium sulfate and filtered. The residue was purified by flash column chromatography (eluent: hexane: MC: EA 3: 1: 1) to give a white solid. (Yield 60%). (Yield 84%), ¹ H NMR (CDCl ₃ ) δ 7.87 (br, 2H), 7.68 (br, 2H), 7.62 (br, 1H), 7.45 (br, 5H), 7.26-7.22 (s, 1H ), 7.26-7.22 (br, 2H), 7.15 (br, 3H), 6.59 (d, 1H), 6.08 (s, 1H), 5.60 (m, 1H), 4.94 (d, J = 3.82, 1H), 3.29 (dd, J ₁ = 6.68 Hz, J ₂ = 6.7 Hz, 2H), 1.96 (d, J = 1.11, 3H); ¹³ CNMR (CDCl ₃ ) δ 187, 164, 135.6, 130.9, 129.9, 129.1, 127.4, 104.1, 48, 43, 13.

Although the present invention has been described through its preferred embodiments, it is merely exemplary, and those skilled in the art will appreciate that various modifications and equivalent other embodiments are possible. Therefore, the true technical protection scope of the present invention should be defined by the technical spirit of the appended claims.

The anti-isoxazolin and isoxazole nucleosides synthesized as described above show excellent antiviral activity. In particular, some compounds are excellent in their antiviral activity and are expected to derive antiviral agents.

Claims (9)

delete delete delete delete (a) synthesizing an amino ester from which N is protected by a tert-butoxycarbonyl group, and reducing the ester to synthesize an N-protected amino aldehyde, from which an N-protected amino aldoxime is synthesized Doing; (b) protecting both amino groups N ¹ and N ³ in the pyrimidine base with benzyl or trimethylsilyl and then reacting with a weak acid to remove one protecting group to synthesize an N ³ -protected pyrimidine base; (c) from pyrimidine base with allyl alcohol under Mitsunobu reaction conditions and with N ¹ -allyl N ³ -benzoyl pyrimidine (uracil / thymine) or with propargyl alcohol under similar reaction conditions N ¹ -propar Synthesizing the length N ³ -benzoyl pyrimidine (uracil / thymine / 5-fluorouracil); And (d) the N ¹ -allyl N ³ -benzoyl pyrimidine (uracil / thymine) or N ¹ -propargyl N ³ -benzoyl pyrimidine (uracil / thymine / 5-fluorouracil) with the amino aldooxime [ 3 + 2] a method for synthesizing isoxazoline of formula (1) or isoxazole of formula (2) comprising the step of reacting with a ringer: <Formula 1> Wherein R is hydrogen or an alkyl group of C1-C5, R 'is hydrogen, an alkyl or aryl group of C1-C10, and X and Y are hydrogen or a protecting group. <Formula 2> Wherein R is hydrogen, a halogen element, or an alkyl group of C1-C5, R 'is hydrogen, an alkyl or aryl group of C1-C10, and X and Y are hydrogen or a protecting group. The method of claim 5, wherein the N-protected amino ester in step (a) is reduced with DIBAL in toluene at -78 ° C to -20 ° C, and the resulting N-protected amino aldehyde in Na _{2 in} aqueous solution Treating said aldooxime by treatment with hydroxyl amine hydrochloride in the presence of CO ₃ . The method according to claim 5, wherein the protecting group in step (b) is a benzoyl group, and 0.5 MK ₂ CO ₃ in water is used for deprotection of the N ¹ -protecting group. 6. The method according to claim 5, wherein in the step (d), the cycloaddition reaction is carried out in the presence of NaOCl. delete