CN116462719A - Process for preparing deoxynucleosides of alpha configuration - Google Patents

Abstract

The application provides a preparation method of alpha-configuration deoxynucleoside, belonging to the technical field of nucleoside synthesis. The preparation method of the deoxynucleoside with the alpha configuration comprises the following steps: reacting a mixed solution containing a deoxyribose raw material, a base raw material and a polar aprotic solvent. The preparation method of the alpha-configuration deoxynucleoside provided by the application takes the deoxyribose raw material with a specific structure as an initiator and is matched with the polar aprotic solvent, so that the purine base raw material and the deoxyribose raw material can react, the reaction is promoted to convert the target product alpha-configuration deoxynucleoside, the reaction is inhibited to convert the byproduct beta-configuration deoxynucleoside, the conversion rate of the target product alpha-configuration deoxynucleoside is improved, and the yield of the alpha-configuration deoxynucleoside is further improved. In addition, the preparation method of the alpha-configuration deoxynucleoside provided by the application is simple and feasible in synthesis process and is beneficial to industrial production.

Description

Preparation method of deoxynucleoside in alpha configuration

technical field

This application relates to the technical field of nucleoside synthesis, in particular, to a method for preparing deoxynucleosides in α configuration.

Background technique

Deoxynucleosides can be divided into α-configuration and β-configuration according to the type of glycosidic bond; among them, α-configuration purine deoxynucleosides are the most basic raw materials for synthesizing α-configuration antisense oligonucleotides. Antisense oligonucleotides in α-configuration are molecular drugs for gene level regulation, which can inhibit the expression of target genes through sequence-specific binding to target gene DNA or mRNA.

However, in the reaction process of the existing method for synthesizing the purine deoxynucleosides of the α configuration, the purine deoxynucleosides of the α configuration and the purine deoxynucleosides of the β configuration will be generated simultaneously, and the conversion rate of the purine deoxynucleosides of the target product α configuration is low (that is, the selectivity of the purine deoxynucleosides of the α configuration is low), resulting in a low yield of the purine deoxynucleosides of the α configuration; in addition, the existing method for synthesizing the purine deoxynucleosides of the α configuration Comparingly complicated, be unfavorable for industrialized production.

Contents of the invention

The purpose of this application is to provide a preparation method of α-configuration deoxynucleosides, which aims to improve the technical problem of low conversion rate of α-configuration purine deoxynucleosides in the existing preparation method of α-configuration purine deoxynucleosides.

The present application provides a method for preparing deoxynucleosides in α configuration, comprising: reacting a mixed solution containing deoxyribose raw materials, base raw materials and polar aprotic solvents.

Wherein, the structural formula of deoxyribose raw material is as follows:

R ₁ and R ₂ are each independently selected from protecting groups.

X is selected from leaving groups.

The base material is selected from any one of the compounds shown in the following structural formula:

R ₃ is selected from amino group, amino group substituted by protecting group, hydroxyl group, hydroxyl group substituted by protecting group, halogen atom, hydrogen atom, alkyl group or aryl group.

R ₄ and R ₆ are each independently selected from a hydrogen atom, an amino group substituted by a protecting group, a hydroxyl group substituted by a protecting group, a halogen atom, an alkyl group or an aryl group.

R ₅ and R ₇ are each independently selected from amino group, amino group substituted by protecting group, hydroxyl group, hydroxyl group substituted by protecting group, halogen atom, hydrogen atom, alkyl group or aryl group.

The preparation method of α-configuration deoxynucleosides provided by this application uses deoxyribose raw materials with specific structures as starting materials, and cooperates with polar aprotic solvents to make purine base raw materials react with deoxyribose raw materials, and promote the conversion of the reaction to the target product α-configuration deoxynucleosides, inhibit the conversion of the reaction to the by-product β-configuration deoxynucleosides (that is, increase the selectivity of the formation of α-configuration deoxynucleosides), realize the improvement of the conversion rate of the target product α-configuration deoxynucleosides, and then improve the α configuration. The yield of deoxynucleosides. In addition, the preparation method of the α-configuration deoxynucleosides provided by the present application has a simple synthesis process and is beneficial to industrial production.

In an optional embodiment of the present application, _R is selected from amino substituted by protecting group, hydroxyl substituted by protecting group, halogen atom, hydrogen atom, alkyl or aryl; and/or, R _{and R} are each independently selected from acetyl, benzoyl, benzyl or p-toluene; and/or, X is selected from halogen atom, trifluoromethanesulfonyloxy, methylsulfonyloxy, p-toluenesulfonyloxy or acetoxy.

In the above technical solution, R ₁ , R ₂ , R ₃ and X are selected from the above groups, which can increase the conversion rate of deoxynucleosides in the alpha configuration of the target product.

In an optional embodiment of the present application, _{both R1} and _R2 are p-toluyl; and/or, X is selected from a halogen atom; and/or, the base material is selected from any one of the compounds shown in the following structural formula:

And _R3 is selected from a hydroxyl group substituted by a protecting group, a halogen atom or a C1-C6 alkyl group, _R4 and _R6 are each independently selected from a hydrogen atom or a C1-C6 alkyl group, _R5 and _R7 are each independently selected from an amino group or an amino group substituted by a protecting group.

In an optional embodiment of the present application, X is selected from a fluorine atom, a chlorine atom, a bromine atom or an iodine atom; and/or, the base material is selected from any one of the compounds shown in the following structural formula:

And R ₃ is selected from a halogen atom, R ₄ and R ₆ are each independently selected from a hydrogen atom, R ₅ and R ₇ are each independently selected from an amino group or an amino group substituted by an isobutyryl group.

In an optional embodiment of the present application, the polar aprotic solvent includes at least one of N,N-dimethylformamide, dimethylacetamide, N-methylpyrrolidone and dimethylsulfoxide.

In the above technical scheme, the polar aprotic solvent is selected from the above-mentioned solvent, which is conducive to improving the conversion rate of the deoxynucleosides in the alpha configuration of the target product, inhibiting the conversion of the reaction to the deoxynucleosides in the beta configuration of the by-product, increasing the conversion rate of the deoxynucleosides in the alpha configuration of the target product, and then increasing the yield of the deoxynucleosides in the alpha configuration.

In an optional embodiment of the present application, the mixed solution also contains an accelerator, and the accelerator is a substance that can ionize ^I- .

In the above technical scheme, the mixed solution contains an accelerator, which can accelerate the reaction rate of the reaction between the base raw material and the deoxyribose raw material, and can also increase the conversion rate of the target product α-configuration deoxynucleosides, thereby increasing the yield of the α-configuration deoxynucleosides.

Optionally, the promoter includes at least one of sodium iodide, potassium iodide, ammonium iodide and tetra-n-butylammonium iodide, which is beneficial to accelerate the reaction rate of the reaction between the base material and the deoxyribose material and increase the conversion rate of the deoxynucleoside in the alpha configuration.

Optionally, the molar ratio of the deoxyribose raw material to the accelerator is 1:(1-3), which can further increase the reaction rate of the reaction between the base raw material and the deoxyribose raw material and increase the conversion rate of the α-configuration deoxynucleoside.

In an optional embodiment of the present application, the molar ratio of the deoxyribose raw material to the base raw material is 1:(1.5-4).

In the above technical scheme, the deoxyribose raw material and the base raw material are under the above ratio conditions, which can fully convert the deoxyribose raw material to the deoxynucleoside of the target product α configuration, thereby increasing the conversion rate of the deoxynucleoside of the target product α configuration.

In an optional embodiment of the present application, the base material is a dried base material; and/or, the polar aprotic solvent is a dried polar aprotic solvent; and/or, the reaction temperature is 10-40°C.

In the above technical scheme, the substances in the reacting mixed liquid are dried, which is beneficial to avoid the situation that the base raw materials participating in the reaction are less and the remaining deoxyribose raw materials not participating in the reaction are relatively large due to the presence of moisture in the reaction system, which in turn helps to improve the conversion rate of the deoxynucleosides of the alpha configuration of the target product. The temperature of the reaction is 10-40°C, which is beneficial to ensure the reaction rate and the conversion rate of the target product α-configuration deoxynucleosides, and improve the production efficiency and the yield of the target product α-configuration deoxynucleosides.

In an optional embodiment of the present application, the structural formula of the base raw material is as follows:

The preparation method of the α-configuration deoxynucleoside also includes: collecting the reacted organic phase, and drying the organic phase; mixing the dried material with acetonitrile, and then collecting the solid phase.

In the above technical solution, when the basic raw material has the above structure, high-purity target product deoxynucleosides in α-configuration can be separated by acetonitrile crystallization, and the separation and purification operation is simple and easy, which is beneficial to industrial production.

Optionally, the mixing temperature is 10-50° C., and the mixing time is 2-5 hours.

In an optional embodiment of the present application, R ₁ and R ₂ are both p-toluylbenzoyl. The preparation method of α-configuration deoxynucleosides also includes: after collecting the solid phase, removing R ₁ and R ₂ . The steps of removing _R1 and _R2 include: adding sodium methoxide to the mixed solution containing solid phase and methanol at -15 to -5°C to obtain a mixed system; deprotecting the mixed system at -10 to 0°C for 1 to 2 hours; cooling the system after the deprotection reaction to ≤ -15°C, and then adjusting the pH of the system after the deprotection reaction to 6-7.

In the above technical scheme, when _{both R1} and _R2 are p-toluyl, the p-toluyl protecting group can be effectively removed by the above method.

Description of drawings

In order to more clearly illustrate the technical solutions of the embodiments of the present application, the accompanying drawings used in the embodiments will be briefly introduced below. It should be understood that the following drawings only show some embodiments of the present application, and therefore should not be regarded as limiting the scope. For those of ordinary skill in the art, other related drawings can also be obtained according to these drawings without creative work.

Fig. 1 is the UPLC spectrogram of the intermediate prepared in Example 1 of the present application.

Fig. 2 is the ¹ H NMR spectrum of the intermediate prepared in Example 1 of the present application.

Fig. 3 is the ¹ H- ¹³ C HMBC spectrum of the intermediate prepared in Example 1 of the present application.

Figure 4 is the NOESY spectrum of the deoxynucleosides prepared in Example 1 of the present application.

Detailed ways

In the reaction process of the existing method for synthesizing purine deoxynucleosides in α configuration, purine deoxynucleosides in α configuration and purine deoxynucleosides in β configuration will be generated simultaneously during the reaction process, and the conversion rate of the target product α-configuration purine deoxynucleosides is low (generally less than 20%), resulting in a low yield of α-configuration purine deoxynucleosides; in addition, the existing method for synthesizing α-configuration purine deoxynucleosides is relatively complicated, which is not conducive to industrial production.

In order to solve the above problems, the present application provides a method for preparing deoxynucleosides in α-configuration, comprising: reacting a mixture containing deoxyribose raw materials, base raw materials and polar aprotic solvents.

Wherein, the structural formula of deoxyribose raw material is as follows:

R ₁ and R ₂ are each independently selected from protecting groups. X is selected from leaving groups.

The base material is selected from any one of the compounds shown in the following structural formula:

R ₃ is selected from amino group, amino group substituted by protecting group, hydroxyl group, hydroxyl group substituted by protecting group, halogen atom, hydrogen atom, alkyl group or aryl group. R ₄ and R ₆ are each independently selected from a hydrogen atom, an amino group substituted by a protecting group, a hydroxyl group substituted by a protecting group, a halogen atom, an alkyl group or an aryl group. R ₅ and R ₇ are each independently selected from amino group, amino group substituted by protecting group, hydroxyl group, hydroxyl group substituted by protecting group, halogen atom, hydrogen atom, alkyl group or aryl group.

For the situation that the structural formula of basic raw material is as follows:

The reaction path of base raw material and deoxyribose raw material reaction is as follows:

Among them, the substance A is a deoxynucleoside product of α configuration; in the above reaction path, the active hydrogen connected to the nitrogen atom on the base raw material reacts with the X atom on the deoxyribose raw material, so that the base raw material is connected with the deoxyribose raw material to form a deoxynucleoside of α configuration.

For the situation that the structural formula of basic raw material is as follows:

The reaction path of base raw material and deoxyribose raw material reaction is as follows:

Wherein, substance B is a deoxynucleoside product of α configuration; in the above reaction path, the active hydrogen connected to the nitrogen atom on the base raw material reacts with the X atom on the deoxyribose raw material, so that the base raw material is connected with the deoxyribose raw material to form a deoxynucleoside of α configuration.

The preparation method of α-configuration deoxynucleosides provided by this application uses deoxyribose raw materials with specific structures as starting materials, and cooperates with polar aprotic solvents to make purine base raw materials react with deoxyribose raw materials, and promote the conversion of the reaction to the target product α-configuration deoxynucleosides, inhibit the conversion of the reaction to the by-product β-configuration deoxynucleosides (that is, increase the selectivity of the formation of α-configuration deoxynucleosides), realize the improvement of the conversion rate of the target product α-configuration deoxynucleosides, and then improve the α configuration. The yield of deoxynucleosides.

In addition, the preparation method of the α-configuration deoxynucleosides provided by the present application has a simple synthesis process and is beneficial to industrial production.

In some optional embodiments of the present application, R _{and R} are each independently selected from acetyl, benzoyl, benzyl or p-toluyl; _R and _R are selected from the above groups, which can improve the conversion rate of deoxynucleosides in the alpha configuration of the target product.

In some optional embodiments of the present application, X is selected from a halogen atom, trifluoromethanesulfonyloxy (-OTf), methylsulfonyloxy (-OMs), p-toluenesulfonyloxy (-OTs) or acetyloxy (-OAc). X is selected from the above groups, which can increase the conversion rate of deoxynucleosides in the alpha configuration of the target product.

Further, in the embodiments of the present application, R ₁ and R ₂ are p-toluyl. Wherein, p-methylbenzoyl corresponds to the following structural formula:

Further, in the embodiments of the present application, X is selected from halogen atoms. Exemplarily, X is selected from fluorine atom, chlorine atom, bromine atom or iodine atom.

It should be noted that, in other feasible embodiments, R ₁ and R ₂ can also be independently selected from other protecting groups; X can also be selected from other leaving groups.

As an example, in the embodiment of the present application, the raw material of deoxyribose is 1-chloro-3,5-di-O-p-toluoyl-2-deoxy-D-ribofuranose, whose structural formula is as follows:

The above formula Tol is p-methylbenzoyl.

In some optional embodiments of the present application, R ₃ is selected from an amino group substituted by a protecting group, a hydroxyl group substituted by a protecting group, a halogen atom, a hydrogen atom, an alkyl group or an aryl group. R ₃ is selected from the above groups, which can improve the conversion rate of deoxynucleosides in the alpha configuration of the target product.

Further, when R ₄ , R ₅ , R ₆ and R ₇ are each independently selected from an alkyl group, the alkyl group is a C1-C6 alkyl group.

In some optional embodiments of the present application, the base material is selected from any one of the compounds shown in the following structural formula:

And _R3 is selected from a hydroxyl group substituted by a protecting group, a halogen atom or a C1-C6 alkyl group, _R4 and _R6 are each independently selected from a hydrogen atom or a C1-C6 alkyl group, _R5 and _R7 are each independently selected from an amino group or an amino group substituted by a protecting group.

Further, the base material is selected from any one of the compounds shown in the following structural formula:

And R ₃ is selected from a halogen atom, R ₄ and R ₆ are each independently selected from a hydrogen atom, R ₅ and R ₇ are each independently selected from an amino group or an amino group substituted by an isobutyryl group.

As an example, in the embodiments of the present application, the base material is selected from any one of the compounds shown in the following structural formula:

In some optional embodiments of the present application, the molar ratio of the deoxyribose raw material to the basic raw material is 1: (1.5-4); the deoxyribose raw material and the basic raw material are under the above ratio conditions, which can fully convert the deoxyribose raw material to the deoxynucleoside of the target product α configuration, thereby increasing the conversion rate of the deoxynucleoside of the target product α configuration.

As an example, the molar ratio of the deoxyribose raw material to the base raw material can be any one point value among 1:1.5, 1:1.7, 1:2, 1:2.2 and 1:2.5 or a range value between any two.

It should be noted that, in other feasible embodiments, the molar ratio of the deoxyribose raw material to the basic raw material is not limited to 1:(1.5-4), for example, the molar ratio of the deoxyribose raw material to the basic raw material can be 1:1 or 1:5, etc.

In some optional embodiments of the present application, the polar aprotic solvent includes at least one of N,N-dimethylformamide (DMF), dimethylacetamide (DMAc), N-methylpyrrolidone (NMP) and dimethylsulfoxide (DMSO). When the polar aprotic solvent is selected from the above-mentioned solvents, it is beneficial to improve the conversion rate of the deoxynucleosides in the alpha configuration of the target product, inhibit the conversion of the reaction to the deoxynucleosides in the beta configuration of the by-product, improve the conversion rate of the deoxynucleosides in the alpha configuration of the target product, and then increase the yield of the deoxynucleosides in the alpha configuration.

As an example, in the embodiments of the present application, the polar aprotic solvent is selected from DMF, which is beneficial to further increase the conversion rate of the target product deoxynucleosides in α configuration.

In some optional embodiments of the present application, in addition to the deoxyribose raw material, the base material and the polar aprotic solvent in the mixed solution for the reaction, the mixed solution for the reaction also contains a promoter, and the promoter is a substance that can ionize I ⁻ (i.e. iodide anion).

The mixed solution contains an accelerator, which can accelerate the reaction rate of the base raw material and the deoxyribose raw material, and can also increase the conversion rate of the target product α-configuration deoxynucleoside, thereby increasing the yield of the α-configuration deoxynucleoside.

Further, the accelerator includes at least one of sodium iodide, potassium iodide, ammonium iodide, and tetra-n-butylammonium iodide; the accelerator is selected from the above-mentioned substances, which is conducive to the realization of accelerating the reaction rate of the reaction between the base material and the deoxyribose material and improving the conversion rate of the deoxynucleoside in the α configuration.

As an example, in the embodiments of the present application, sodium iodide is selected as the accelerator, which is beneficial to further increase the conversion rate of deoxynucleosides in the α configuration.

In some optional embodiments of the present application, the molar ratio of the deoxyribose raw material to the accelerator is 1: (1-3); the deoxyribose raw material and the accelerator can further increase the reaction rate of the reaction between the base raw material and the deoxyribose raw material and increase the conversion rate of deoxynucleosides in the α configuration under the above-mentioned ratio conditions.

As an example, the molar ratio of the deoxyribose raw material to the accelerator can be any one point value in 1:0.1, 1:0.5, 1:1.0, 1:1.2, 1:1.5, 1:1.7, 1:2 and 1:2.2 or a range value between any two.

It should be noted that, in other feasible embodiments, the molar ratio of the deoxyribose raw material to the accelerator is not limited to 1:(1-3). For example, the molar ratio of the deoxyribose raw material to the accelerator can be 1:0.5 or 1:4, etc. In other possible embodiments, accelerators may also be omitted.

In order to further increase the conversion rate of the target product of deoxynucleosides in α configuration, in some optional embodiments of the present application, the substances in the mixed solution for the reaction are dried. For example, the base material in the mixed solution is a dried base material; and/or, the polar aprotic solvent in the mixed solution is a dried polar aprotic solvent.

The substances in the mixed solution for the reaction are dried, which is beneficial to avoid the situation that due to the presence of moisture in the reaction system, the base raw materials participating in the reaction are less and the deoxyribose raw materials that do not participate in the reaction remain more, which in turn helps to improve the conversion rate of the deoxynucleosides in the alpha configuration of the target product.

Further, in the case that there is an accelerator in the mixed solution for the reaction, the accelerator in the mixed solution is a dried accelerator.

As an example, the K.F.<150ppm of the mixed solution for the reaction is beneficial to avoid the presence of moisture in the reaction system, resulting in less base raw materials participating in the reaction and more deoxyribose raw materials not participating in the reaction remaining, which in turn is conducive to improving the conversion rate of the deoxynucleosides in the alpha configuration of the target product.

In some optional embodiments of the present application, the reaction temperature of the mixed liquid containing deoxyribose raw materials, basic raw materials and polar aprotic solvent is 10-40°C; the reaction temperature is within the above temperature range, which is beneficial to ensure the reaction rate and the conversion rate of the target product α-configuration deoxynucleosides, and improve the production efficiency and the yield of the target product α-configuration deoxynucleosides.

As an example, the reaction temperature can be any one of 10°C, 15°C, 18°C, 20°C, 22°C, 25°C, 27°C, 30°C, 35°C and 40°C or any range between them.

In some optional embodiments of the present application, the reaction time of the mixture containing deoxyribose raw materials, base raw materials and polar aprotic solvent is 5-24 hours; the reaction time is within the above time range, which is beneficial to ensure the progress of the reaction.

As an example, the reaction time may be any one of 5h, 7h, 10h, 12h, 15h, 17h, 20h, 22h and 24h or a range value between any two.

In some optional embodiments of the present application, the preparation method of α-configuration deoxynucleosides also includes: after reacting the mixture containing deoxyribose raw materials, base materials and polar aprotic solvents, isolating and obtaining α-configuration deoxynucleoside intermediates (that is, α-configuration deoxynucleosides protected by protective groups).

As an example, in the embodiment of the present application, the base material is N2-iBu guanine, and its structural formula is as follows:

The step of separating and obtaining the deoxynucleoside intermediate of α configuration includes: collecting the organic phase after the reaction (that is, after the reaction of the mixed solution), and drying the organic phase; mixing the dried material with acetonitrile, and then collecting the solid phase.

When the structural formula of the base raw material is selected from the corresponding material of the above structural formula (i.e. N2-iBu guanine), in the presence of a polar aprotic solvent, the reaction can form a deoxynucleoside intermediate with N9-α configuration (i.e. substance A), a deoxynucleoside intermediate of N7-α configuration, a deoxynucleoside intermediate of N9-β configuration and a system of deoxynucleoside intermediates of N7-β configuration, and the conversion rate of the deoxynucleoside intermediate of α configuration is much higher than The conversion rate of the deoxynucleoside intermediate of the β configuration, and the conversion rate of the deoxynucleoside intermediate of the N9-α configuration is also much higher than the conversion rate of the deoxynucleoside intermediate of the N7-α configuration; the deoxynucleoside intermediate of the N9-α configuration with high purity (98.5% or more) is separated and purified by acetonitrile crystallization, and the separation and purification operation is simple and easy, which is conducive to industrial production. Wherein, the structural formula of the deoxynucleoside intermediate of N7-α configuration is as follows:

The structural formula of the deoxynucleoside intermediate in the N9-β configuration is as follows:

The structural formula of the deoxynucleoside intermediate in the N7-β configuration is as follows:

Further, the above-mentioned mixing temperature of the dried substance and acetonitrile is 10-50° C., and the mixing time is 2-5 hours. The above mixing temperature and mixing time are beneficial to increase the amount of collected N9-α-configuration deoxynucleoside intermediates, and further help to increase the yield of N9-α-configuration deoxynucleosides.

Furthermore, after mixing the dried substance with acetonitrile, the step of "collecting the solid phase" includes: filtering the mixed system of the dried substance and acetonitrile, using 5 to 10 times the volume of acetonitrile at 10 to 50°C to stir and crystallize for 2 to 5 hours, and then filtering, and using 2 to 3 times the volume of acetonitrile at 10 to 50°C to suspend and beat the obtained filter cake and continue to purify for 2 to 3 times.

It should be noted that, in the case of using different base materials for the reaction, the method of separating the α-configuration deoxynucleoside intermediates from the post-reaction system can be purified by silica gel chromatography or reversed-phase chromatographic column (such as C18 reverse-phase chromatographic column).

As an example, when the base raw material is N2-iBuguanine, before mixing the dried substance with acetonitrile, the step of "collecting the reacted organic phase and drying the organic phase" includes: pouring the reacted system into a mixture of 35 to 40 times the volume of ethyl acetate and 120 to 170 times the volume of 0.3 to 0.7 wt% sodium bicarbonate solution, stirring thoroughly for 10 to 15 minutes, and filtering the filter cake with 10 to 30 times the volume of acetic acid Rinse once with ethyl ester, separate the organic phase in the filtrate, back-extract the aqueous phase twice with 20 to 40 times the volume of ethyl acetate, combine all the organic phases, wash with 70 to 90 times the volume of 1 to 3 wt% sodium chloride solution for 2 to 4 times, dry over anhydrous sodium sulfate, and concentrate under reduced pressure to obtain a solid crude product.

Further, in some optional embodiments of the present application, the preparation method of the α-configuration deoxynucleoside further includes: after separating the α-configuration deoxynucleoside intermediate, removing the protecting group on the α-configuration deoxynucleoside intermediate.

As an example, in the embodiments of the present application, the structural formula of the base raw material and the structural formula of the deoxyribose raw material are as follows:

Wherein Tol is p-methylbenzoyl. The step of breaking away from the protecting group (i.e., p-toluyl group) on the deoxynucleoside intermediate in the α configuration includes: after collecting the solid phase, adding sodium methoxide to the mixture containing the solid phase and methanol at -15 to -5°C to obtain a mixed system; deprotecting the mixed system at -10 to 0°C for 1 to 2 hours; cooling the deprotected system to ≤ -15°C, and then adjusting the pH of the deprotected system to 6 to 7.

Further, after adjusting the pH of the system after the deprotection reaction to 6-7, suspension and purification with acetonitrile and water can obtain the high-purity target product deoxynucleoside in α-configuration. As an example, after the pH of the system after the deprotection reaction is adjusted to 6-7, the pH-adjusted system is concentrated and dried under reduced pressure, and then concentrated once with a methanol belt, and 8-10 times the volume of acetonitrile is added, suspended and stirred at 10-25°C for 2-3 hours, filtered, the filter cake is rinsed once with a small amount of acetonitrile, and dried in vacuo to obtain a crude product. Suspend and stir the crude product with 3 to 5 times of deionized water at 10 to 45°C for 2 to 3 hours, then filter, and continue to suspend and stir the obtained filter cake with 1 to 2 times of deionized water at 10 to 45°C for 2 to 3 hours, then filter, rinse the filter cake once with a small amount of deionized water, and vacuum dry to obtain the high-purity target product α-configuration deoxynucleoside.

It should be noted that other conventional methods for removing the p-toluoyl protecting group on the deoxynucleoside intermediate of the α-configuration can also be selected; if the protecting group on the deoxynucleoside intermediate of the α-configuration is other groups, the corresponding protecting group can also be removed using the conventional method in the prior art, which is not limited in this application.

In order to make the purpose, technical solutions and advantages of the embodiments of the present application clearer, the technical solutions in the embodiments of the present application will be clearly and completely described below. Those who do not indicate the specific conditions in the examples are carried out according to the conventional conditions or the conditions suggested by the manufacturer. The reagents or instruments used were not indicated by the manufacturer, and they were all conventional products that could be purchased from the market.

Example 1

This embodiment provides a method for preparing deoxynucleosides in α configuration, comprising the following steps:

(1) Add 62.6g of N2-iBuguanine into a 5.0L round bottom flask, dehydrate with 1.0L of dry acetonitrile three times, add 2.5L of dry DMF, mix well, and let stand for 7min to obtain a mixed system, K.F.<150ppm.

(2) Add 28.9g of NaI solid and 50g of 1-chloro-3,5-di-O-p-toluoyl-2-deoxy-D-ribofuranose solid to the mixed system obtained in step (1) at 20°C under stirring. After continuing to stir for 10 hours, take the reaction mixture and send it to UPLC for monitoring. At a wavelength of 260nm, integrate all peaks except N2-iBuguanine, and calculate the main peak according to the peak area normalization method And impurity content, if the content of N9-α configuration deoxynucleoside product > 38%, and the total amount of impurities in MS 925 < 2%, the reaction is considered qualified, otherwise continue the reaction until the reaction is qualified; the initial product system is obtained.

(3) Stop stirring the reaction, pour the initial product system obtained in step (2) into the mixture formed by 2.0L of ethyl acetate and 7.5L of 0.5wt% NaHCO _aqueous solution, add 50.0g of diatomaceous earth, stir for 10min, and filter. The filter cake was rinsed with 1.0L of ethyl acetate, the organic phase in the filtrate was separated, the aqueous phase was back-extracted twice with 1.5L of ethyl acetate, all the organic phases were combined, washed 3 times with 4.0L of 2wt% NaCl aqueous solution, then dried with anhydrous sodium sulfate, concentrated and dried under reduced pressure to obtain a brown solid crude product.

(4) Concentrate the brown solid crude product obtained in step (3) once with 400 mL of acetonitrile, add 350 mL of acetonitrile, stir and crystallize at 45 ° C for 2 h, then cool naturally to 15 ° C, continue stirring for 5 h, filter, and obtain an off-white solid after vacuum drying the filter cake. The off-white solid was suspended and stirred with 150 mL of acetonitrile at 45°C for 4 hours, cooled to 15°C, filtered, the filter cake was rinsed with a small amount of acetonitrile, and suspended again to obtain the intermediate.

(5) Mix the intermediate obtained in step (4) with 250 mL of methanol to obtain a mixed solution; add 7.0 g of sodium methoxide to the mixed solution at -10°C; then react at -5°C for 1.5 h, cool to -15°C, and then adjust the pH of the system to 7. Concentrate and dry the pH-adjusted system under reduced pressure, use a methanol belt to dry concentrate once, add 250 mL of acetonitrile, suspend and stir at 20°C for 2.5 h, filter, rinse the filter cake once with a small amount of acetonitrile, and dry in vacuo to obtain a crude product. Suspend and stir the crude product with 108 mL of deionized water at 20°C for 2.5 h, then filter, and continue to suspend and stir the obtained filter cake with 27.5 mL of deionized water at 20°C for 2.5 h, then filter, rinse the filter cake once with a small amount of deionized water, and dry in vacuum to obtain deoxynucleosides.

Example 2

This embodiment provides a method for preparing deoxynucleosides in α configuration, comprising the following steps:

(1) Add 2.73g of 2-amino-6-chloropurine into a 100mL round-bottomed flask, remove water once with 50mL of dry acetonitrile, add 50mL of dry DMF, mix well, and let stand for 7min to obtain a mixed system.

(2) At 15°C, add 1.45 g of NaI solid and 2.5 g of 1-chloro-3,5-di-O-toluoyl-2-deoxy-D-ribofuranose solid to the mixed system obtained in step (1) under stirring, continue to stir and react for 12 hours, then take the reaction mixture and send it to UPLC for monitoring. At a wavelength of 260 nm, integrate all peaks and calculate the main peak and impurity content according to the peak area normalization method. If the content of the deoxynucleoside product is >30%, and the total amount of MS 873 impurities is <12%, the reaction is considered qualified, otherwise the reaction is continued until the reaction is qualified; the initial product system is obtained.

(3) stop stirring reaction, the initial product system that step (2) obtains is poured into the ethyl acetate of 120mL and 120mL 0.5wt% _NaHCO In the mixture that aqueous solution forms, stir 4min, filter, separate the organic phase in the filtrate, wash 5 times with the deionized water of 120mL, after drying over anhydrous sodium sulfate, concentrate under reduced pressure and dry to obtain yellow solid crude product.

Example 3

This example provides a method for preparing deoxynucleosides in α configuration. The difference between this example and Example 1 is that the mass of N2-iBu guanine is 71.1 g.

Example 4

This example provides a method for preparing deoxynucleosides in the α configuration. The difference between this example and Example 1 is that the mass of N2-iBu guanine is 42.7 g.

Example 5

This example provides a method for preparing deoxynucleosides in the α configuration. The difference between this example and Example 1 is that the mass of N2-iBu guanine is 28.5 g.

Example 6

This example provides a method for preparing deoxynucleosides in α configuration. The difference between this example and Example 1 is that the mass of N2-iBu guanine is 113.8 g.

Example 7

This example provides a method for preparing deoxynucleosides in α configuration. The difference between this example and Example 1 is that the mass of NaI is 42.4 g.

Example 8

This example provides a method for preparing deoxynucleosides in α-configuration. The difference between this example and Example 1 is that the mass of NaI is 19.3 g.

Example 9

This example provides a method for preparing deoxynucleosides in α-configuration. The difference between this example and Example 1 is that the mass of NaI is 3.9 g.

Example 10

This example provides a method for preparing deoxynucleosides in α-configuration. The difference between this example and Example 1 is that the mass of NaI is 57.8 g.

Example 11

This example provides a method for preparing deoxynucleosides in the α configuration. The difference between this example and Example 1 is that 28.9 g of NaI in Example 1 is replaced by 27.9 g of ammonium iodide.

Example 12

This example provides a method for preparing deoxynucleosides in α-configuration. The difference between this example and Example 1 is that NaI is not used in this example.

Example 13

This example provides a method for preparing deoxynucleosides in α-configuration. The difference between this example and Example 1 is that DMF in Example 1 is replaced with DMSO.

Example 14

This embodiment provides a method for preparing deoxynucleosides in α-configuration. The difference between this embodiment and Example 1 is that the step (1) is different. The step (1) of this embodiment is as follows:

Mix 62.6g of N2-iBuguanine and 2.5L of DMF thoroughly, and then let it stand for 7 minutes to obtain a mixed system with a K.F. of 2000ppm.

Example 15

This example provides a method for preparing deoxynucleosides in α-configuration. The difference between this example and Example 1 is that the reaction temperature in step (2) is 10°C.

Example 16

This example provides a method for preparing deoxynucleosides in α-configuration. The difference between this example and Example 1 is that the reaction temperature in step (2) is 40°C.

Example 17

This example provides a method for preparing deoxynucleosides in α-configuration. The difference between this example and Example 1 is that the reaction temperature in step (2) is 0°C.

Example 18

This example provides a method for preparing deoxynucleosides in α-configuration. The difference between this example and Example 1 is that the reaction temperature in step (2) is 60°C.

Experimental example 1

The intermediate prepared in Example 1 was characterized by ultra-high performance liquid chromatography (UPLC), and the characterization results are shown in FIG. 1 .

It can be seen from Figure 1 that in the intermediates prepared in step (4) of Example 1, the purity of the deoxynucleoside product in the N9-α configuration is 98.52%, and the ratios of the deoxynucleoside product in the N7-α configuration and the deoxynucleoside product in the N9-β configuration are 0.58% and 0.70% respectively; it shows that the preparation method of Example 1 of the present application can obtain a product with a higher purity in the N9-α configuration.

The intermediate prepared in Example 1 was characterized by hydrogen nuclear magnetic resonance spectrum ( ¹ H NMR) and ¹ H- ¹³ C heteronuclear multi-bond correlation spectrum ( ¹ H- ¹³ C HMBC), and the characterization results are shown in Fig. 2 and Fig. 3 respectively.

In Figure 2,¹H NMR (500MHz, CDCl₃): 12.20 (s, 1H, 1-NH), 10.32 (s, 1H, IBU-NH), 8.04 (s, 1H, 8-H), 7.85 (D, J = 8.0Hz, 2H, TOL-CH), 7.62 (D, J = 8.0Hz, 2H, TOL-CH), 7.17 (D, J = 8.0Hz, 2H, 2H, 2H, 2H, 2H, 2H. , TOL-CH), 7.13 (D, J = 8.0Hz, 2H, TOL-CH), 6.20 (DD, J = 5.0, 3.5Hz, 1h, 1 ’-H), 5.62-5.56 (m, 1H, 3’-H), 4.82-4.73 (m, 1H, 4’-H), 4.50 (d, j = 4.5Hz 4.5Hz 4.5Hz , 2H, 5’-H), 2.90 (SEP,

J＝7.0Hz, 1H, iBu-CH), 2.84-2.80(m, 2H, 2'-H), 2.351(s, 3H, Tol-CH ₃ ), 2.345(s, 3H, T

ol-CH ₃ ), 1.22 (d, J=7.0Hz, 3H, iBu-CH ₃ ), 1.21 (d, J=7.0Hz, 3H, iBu-CH ₃ ), from Figure 2

It can be seen that the intermediate prepared in Example 1 (without removing the toluyl protecting group) has the same structure as the expected intermediate.

In Figure 3, ^{the 1} H- ¹³ C HMBC spectrum shows that the 1'-H on the sugar ring in the obtained compound has no carbon-hydrogen long-distance correlation with the base 5-C, but there is a long-distance carbon-hydrogen correlation with the base 4-C, which shows that the N9 position of the base is connected to the sugar ring.

The deoxynucleosides prepared in Example 1 were characterized by NOESY spectra, and the characterization results are shown in FIG. 4 .

In Figure 4, there are NOE-related signals between the base 8-H and the 4'-H and 3'-OH on the sugar ring, but there is no NOE-related signal with the 3'-H on the sugar ring, so it can be known that the obtained deoxynucleoside (product, that is, the p-toluyl protecting group has been removed) is in the α-configuration.

Experimental example 2

In the initial product system obtained in Examples 1-18 (i.e., the system obtained in step (2)), the conversion rate of deoxynucleoside products of N9-α configuration (referred to as N9-α), deoxynucleoside products of N7-α configuration (referred to as N7-α), deoxynucleoside products of N9-β configuration (referred to as N9-β) and deoxynucleoside products of N7-β configuration (referred to as N7-β) was determined, and the measurement results are shown in Table 1.

Table 1

It can be seen from Table 1 that the conversion rate of α-configuration deoxynucleosides can be 29.5% or above by adopting the preparation method of α-configuration deoxynucleosides provided by the present application.

From the comparison of Example 1 and Example 3-6, it can be seen that compared to the molar ratio of deoxyribose raw material (i.e. 1-chloro-3,5-di-O-toluoyl-2-deoxy-D-ribofuranose) and base raw material (i.e. N2-iBuguanine) in Example 5 is 1:1, when the molar ratio of deoxyribose raw material and base raw material is 1: (1.5-4), the conversion rate of the deoxynucleoside of α configuration can be further improved to 36.3% and above.

From the comparison of Example 1 and Examples 7-10, it can be seen that compared to the molar ratio of the deoxyribose raw material (i.e. 1-chloro-3,5-di-O-toluoyl-2-deoxy-D-ribofuranose) to the accelerator (i.e. NaI) in Example 9 is 1:0.2, when the molar ratio of the deoxyribose raw material to the accelerator is 1:(1-3), the conversion rate of the deoxynucleoside in the α configuration can be further increased to 35.7% and above .

From the comparison of Example 1 and Example 11, it can be seen that, compared with the use of ammonium iodide in Example 11 as the accelerator, the NaI in Example 1 is used as the accelerator, which is conducive to further improving the conversion rate of deoxynucleosides in the α configuration.

From the comparison of Example 1 and Example 12, it can be seen that compared with the non-use of the accelerator NaI in Example 12, the use of the accelerator NaI in Example 1 is beneficial to further increase the conversion rate of deoxynucleosides in the α configuration.

From the comparison of Example 1 and Example 13, it can be seen that compared with the solvent in Example 13, DMSO is used, and the solvent in Example 1 is DMF, which is beneficial to further improve the conversion rate of deoxynucleosides in the α configuration.

From the comparison between Example 1 and Example 14, it can be seen that compared with Example 14, the base material (i.e. N2-iBuguanine) and the solvent (i.e. DMF) were not dehydrated or dried, and the base material was dehydrated in Example 1 and the use of a dry solvent could improve the conversion rate of deoxynucleosides in the α configuration.

From the comparison of embodiments 15-18, it can be seen that compared with deoxyuraogenic sugar raw materials (that is, 1-chloro-3,5-2 -o-Ol-2-deoxyrum-fuu nucleus) and alkaline-based raw materials (that is, N2-IBU birds), the temperature reacts is 0 ° C or 60 ° C. The degree of 10-40 ° C is conducive to further increasing the conversion rate of deoxyucleoside of α configuration.

In summary, the preparation method of α-configuration deoxynucleosides provided by this application uses deoxyribose raw materials with specific structures as starting materials, and cooperates with the selection of polar aprotic solvents, so that base raw materials can react with deoxyribose raw materials, and promote the conversion of the reaction to the target product α-configuration deoxynucleosides, inhibit the conversion of the reaction to the by-product β-configuration deoxynucleosides, increase the conversion rate of the target product α-configuration deoxynucleosides, and then increase the yield of α-configuration deoxynucleosides. In addition, the preparation method of the α-configuration deoxynucleoside provided by the present application is simple and easy to synthesize, which is beneficial to industrial production.

The embodiments described above are some of the embodiments of the present application, but not all of them. The detailed description of the embodiments of the application is not intended to limit the scope of the claimed application, but merely represents selected embodiments of the application. Based on the embodiments in this application, all other embodiments obtained by persons of ordinary skill in the art without making creative efforts belong to the scope of protection of this application.

Claims (10)

1. A method for preparing deoxynucleosides in the alpha configuration, comprising:

reacting a mixed solution containing a deoxyribose raw material, a base raw material and a polar aprotic solvent;

wherein the structural formula of the deoxyribose raw material is as follows:

R ₁ and R is ₂ Each independently selected from protecting groups;

x is selected from leaving groups;

the base raw material is selected from any one of compounds shown in the following structural formulas:

R ₃ selected from amino, amino substituted with a protecting group, hydroxyl substituted with a protecting group, halogen atom, hydrogen atom, alkyl or aryl;

R ₄ and R is ₆ Each independently selected from a hydrogen atom, a protecting group-substituted amino group, a protecting group-substituted hydroxyl group, a halogen atom, an alkyl group, or an aryl group;

R ₅ and R is ₇ Each independently selected from amino, protecting group substituted amino, hydroxyl, protecting group substituted hydroxyl, halogen atom, hydrogen atom, alkyl or aryl.

2. The method of claim 1, wherein R is ₃ Selected from the group consisting of a protecting group-substituted amino group, a protecting group-substituted hydroxyl group, a halogen atom, a hydrogen atom, an alkyl group, and an aryl group;

and/or, the R ₁ And said R ₂ Each independently selected from acetyl, benzoyl, benzyl or p-methylbenzoyl;

and/or, the X is selected from halogen atom, trifluoromethanesulfonyl group, methylsulfonyl group, p-toluenesulfonyloxy group or acetoxy group.

3. The preparation method according to claim 1, characterized in thatIn that the R ₁ And said R ₂ Are p-methylbenzoyl groups;

and/or, the X is selected from halogen atoms;

and/or the base raw material is selected from any one of compounds shown in the following structural formulas:

and said R is ₃ Selected from the group consisting of protecting group-substituted hydroxy groups, halogen atoms, and C1-C6 alkyl groups, said R ₄ And said R ₆ Each independently selected from a hydrogen atom or a C1-C6 alkyl group, R ₅ And said R ₇ Each independently selected from amino or amino substituted with a protecting group.

4. The method of claim 1, wherein X is selected from the group consisting of fluorine, chlorine, bromine, and iodine;

and/or the base raw material is selected from any one of compounds shown in the following structural formulas:

and said R is ₃ Selected from halogen atoms, said R ₄ And said R ₆ Each independently selected from hydrogen atoms, R ₅ And said R ₇ Each independently selected from amino or isobutyryl substituted amino.

5. The method according to claim 1, wherein the polar aprotic solvent comprises at least one of N, N-dimethylformamide, dimethylacetamide, N-methylpyrrolidone, and dimethylsulfoxide.

6. The method according to claim 1, wherein the mixed solution further comprises an accelerator, wherein the accelerator is ionizable I ^- Is a substance of (a);

optionally, the promoter comprises at least one of sodium iodide, potassium iodide, ammonium iodide, and tetra-n-butyl ammonium iodide;

optionally, the molar ratio of the deoxyribose raw material to the accelerator is 1 (1-3).

7. The method according to any one of claims 1 to 6, wherein the molar ratio of the deoxyribose raw material to the base raw material is 1 (1.5 to 4).

8. The method according to any one of claims 1 to 6, wherein the base raw material is a dried base raw material;

and/or the polar aprotic solvent is a polar aprotic solvent subjected to a drying treatment;

and/or the temperature of the reaction is 10-40 ℃.

9. The method according to claim 1, wherein the base material has the following structural formula:

the preparation method of the alpha-configuration deoxynucleoside further comprises the following steps: collecting the organic phase after the reaction, and drying the organic phase; mixing the dried material with acetonitrile, and then collecting a solid phase;

optionally, the temperature of the mixing is 10-50 ℃, and the time of the mixing is 2-5 h.

10. The method of claim 9, wherein R is ₁ And said R ₂ Are p-methylbenzoyl groups;

the preparation method of the alpha-configuration deoxynucleoside further comprises the following steps: after collecting the solid phase, the R is removed ₁ And said R ₂ ；

Removing the R ₁ And said R ₂ The method comprises the following steps: adding sodium methoxide into the mixed solution containing the solid phase and methanol at the temperature of minus 15 ℃ to minus 5 ℃ to obtain a mixed system; deprotection reaction is carried out on the mixed system for 1 to 2 hours at the temperature of minus 10 to 0 ℃; and cooling the system after the deprotection reaction to less than or equal to-15 ℃, and then regulating the pH value of the system after the deprotection reaction to 6-7.