Novel crystal form of key amino acid induced trelagliptin based on protein-drug interaction and preparation method thereof

Abstract

The invention discloses a key amino acid induced trelagliptin novel crystal form based on protein-drug interaction structure excavation and a preparation method thereof. According to the technical scheme, key amino acid is introduced into a solvent crystallization system of trelagliptin to induce trelagliptin to generate five new crystal forms; the preparation method does not need to use a large amount of organic solvent, has the characteristics of simplicity, accuracy, high efficiency and the like, has strong crystal form regulation and control capability, is energy-saving, environment-friendly and safe, and is easy to realize green industrial production.

Description

Technical Field

[0001] The present invention belongs to the technical field of drug preparation, and in particular relates to a method for regulating the crystal form of Trelagliptin drug by using key amino acids. Background Art

[0002] The discovery and regulation of new drug crystal forms is the core of drug crystallography research. It provides a basis for screening out superior crystal forms with excellent physical and chemical properties, significant efficacy and high safety, and has great value and practical significance for drug crystal form research. Differences in drug crystal forms will affect the efficacy, safety, stability and formulation process of drugs.

[0003] The essence of crystallization is the process of molecular assembly. The intermolecular interactions (such as hydrophobic interaction, hydrogen bonding, van der Waals force, and electrostatic force) will directly affect the assembly of molecules, and then affect the crystallization and crystal structure of the substance. Existing studies have reported that there is an interaction between trelagliptin and human serum albumin (HSA) and pepsin (PEP). This interaction is also the intermolecular interaction between drugs and proteins. Based on this interaction, key amino acids that have strong interactions with trelagliptin have been mined. The difference in the interaction between key amino acids and drug molecules will change the assembly of drug molecules, thereby affecting drug crystallization and the crystal structure of drugs. At present, studies on the induction of drug crystallization by key amino acids mined based on protein-drug interactions have not been reported. At the same time, proteins are essentially peptides formed by the dehydration condensation of amino acids, and the peptide chains are folded to form the three-dimensional structure of proteins. Therefore, it is necessary to go deep into the protein, dig out key amino acids, and use the strong interaction between key amino acids and drugs to study the effects of key amino acids on the crystallization and crystal form of trelagliptin drugs, and accurately induce new crystal forms of drugs.

[0004] In this study, the key amino acids that play a strong interaction in the interactions between trelagliptin and human serum albumin (HSA) and trelagliptin and pepsin (PEP) were selected as the research objects, and the key amino acids represented by glutamic acid (GLU), arginine (ARG), alanine (ALA), threonine (THR), tyrosine (TYR) were studied to explore the formation of a new crystal form of trelagliptin induced by the key amino acids in protein-drug interaction and its preparation method.

[0005] Trelagliptin (TRE), chemical name: 2-[[6-[(3R)-3-amino-1-piperidinyl]-3,4-dihydro-3-methyl-2,4-dioxo-1(2H)-pyrimidinyl]methyl]-4-fluoro-benzonitrile, chemical formula: C 18 H 20FN5O2. Trelagliptin was developed by Takeda Pharmaceuticals of Japan. It is a long-acting dipeptidyl peptidase-4 (DPP-4) inhibitor and also a very good oral hypoglycemic drug for the treatment of type 2 diabetes. The structural formula is shown below.

[0006]

[0007] According to known reports, TRE has multiple crystalline forms. For example, world patent WO2014127735-A1 (invention name: Crystal form A of trelagliptin useful for eg preparing medicine for treating disease mediated by dipeptidyl peptidase IV, exhibits powder X-ray diffraction pattern, and has diffraction peak at specified angle) discloses a trelagliptin crystal A. The 2θ values ​​of the X-ray powder diffraction pattern of this trelagliptin crystal A are 5.7±0.2°, 11.4±0.2°, 12.5±0.2°, 16.8±0.2°, 17.1±0.2°, 19.4±0.2°, 19.9±0.2°, 20.5±0.2°, 22.5±0.2°, 22.9±0.2°, and 29.1±0.2°. Secondly, Chinese patent CN105384724 (invention name: a fluorinated crystal form and its preparation method) discloses trelagliptin crystal form II and crystal form IV and their preparation methods; wherein, the X-ray powder diffraction spectrum 2θ values ​​of crystal form II are 20.04°, 20.85°, 21.86°, and the X-ray powder diffraction spectrum 2θ values ​​of crystal form IV are 16.28°, 20.03°, 22.30°, 27.55°. For example, Chinese patent CN105693691 (invention name: a new crystal form of high-purity trelagliptin and its preparation) discloses a method for preparing a new crystal form of high-purity trelagliptin, and the X-ray powder diffraction spectrum 2θ values ​​of this new crystal form of trelagliptin are 4.8±0.2°, 9.6±0.2°, 18.4±0.2°, 18.9±0.2°. For example, Chinese invention patent CN104003975 (invention name: new solid form of trelagliptin, preparation method and use thereof) discloses five crystal forms of trelagliptin and preparation methods thereof, including crystal form A, crystal form B, crystal form C, crystal form D, crystal form E and amorphous form. For example, Chinese invention patent (application publication number) CN115785066 (invention name: new crystal form F of trelagliptin and preparation method thereof) discloses a preparation method of new crystal form F of trelagliptin, which uses protein grinding induction to prepare a new crystal form of trelagliptin, and the preparation method is green and environmentally friendly.

[0008] However, most of the above methods have harsh conditions, and no safe and green amino acids are used as regulators for regulating the crystallization of trelagliptin. This patent is based on the three-dimensional structure of the interaction between trelagliptin-human serum albumin (HSA) and trelagliptin-pepsin (PEP), and has discovered key amino acids that have strong interactions with the drug. It has innovatively introduced key amino acids into the trelagliptin solvent system, induced the formation of multiple new crystal forms of trelagliptin, and designed its preparation method. The relevant experiments were conducted using amino acids represented by glutamic acid (GLU), phenylalanine (PHE), arginine (ARG), threonine (THR), and tyrosine (TYR). Summary of the invention

[0009] The purpose of the present invention is to provide a method for preparing a variety of new crystal forms of trelagliptin by introducing key amino acids into a trelagliptin solvent crystallization system and obtaining the key amino acids through the interaction between the key amino acids and trelagliptin.

[0010] The second object of the present invention is to provide five new crystal forms of trelagliptin.

[0011] To this end, the first technical solution provided by the present invention is as follows:

[0012] A method for preparing new crystal forms of trelagliptin induced by key amino acids in protein-drug interaction, wherein key amino acids are introduced into the solvent crystallization system of trelagliptin to induce trelagliptin to produce a variety of new crystal forms.

[0013] Furthermore, the above-mentioned method for preparing a new crystal form of Trelagliptin induced by key amino acids based on protein-drug interaction comprises the following steps in sequence:

[0014] 1) slowly adding the solvent to trelagliptin until trelagliptin is just dissolved to saturation, to obtain a trelagliptin solution for use;

[0015] 2) Add the amino acid solution into deionized water until the amino acid is dissolved to saturation to obtain an amino acid solution;

[0016] 3) Then, using a rubber-tipped dropper, the amino acid solution prepared in 2) is slowly dripped into the trelagliptin solution prepared in 1) in sequence, and the whole process is carried out in a constant temperature water bath at 25° C.-50° C.;

[0017] 4) After the addition is completed, seal it with plastic wrap, and poke 30-35 holes in the seal, place it in a fume hood and wait for crystals to precipitate, so as to induce trelagliptin to produce multiple new crystal forms;

[0018] The molar ratio of the amino acid to trelagliptin is 1:2 to 1:30.

[0019] Furthermore, in the above-mentioned method for preparing a new crystal form of trelagliptin induced by key amino acids based on protein-drug interaction, the amino acid is one of glutamic acid, arginine, threonine, and tyrosine or any combination thereof; and the amino acid is one of ethyl acetate, isopropanol, acetone, methanol, ethanol, and acetonitrile.

[0020] The second technical solution provided by the present invention is the above-mentioned key amino acid-induced new crystal form of trelagliptin based on protein-drug interaction, and the new crystal form of trelagliptin is TRE-Ⅰ, TRE-Ⅱ, TRE-Ⅲ, TRE-Ⅳ, TRE-Ⅴ, and TRE-Ⅵ.

[0021] Furthermore, the above-mentioned key amino acid-induced new crystal form of trelagliptin based on protein-drug interaction, the 2θ values ​​of the X-ray powder diffraction spectrum of the new crystal form TRE-Ⅰ are 5.66°±0.2°, 11.35°±0.2°, 12.47°±0.2°, 17.08°±0.2°, 19.88°±0.2°, 22.48°±0.2°, 22.87°±0.2°, and 27.54°±0.2°.

[0022] Furthermore, the above-mentioned key amino acid-induced new crystal form of trelagliptin based on protein-drug interaction, the 2θ value of the new crystal form TRE-Ⅱ is 8.07°±0.2°, 13.31°±0.2°, 13.64°±0.2°, 16.29°±0.2°, 16.79°±0.2°, 17.40°±0.2°, 19.40°±0.2°, 21.33°±0.2°, 22.83°±0.2°, 26.26°±0.2°, 26.57°±0.2°, 26.79°±0.2°, 27.49°±0.2°, 27.90°±0.2°, and 33.33°±0.2°.

[0023] Furthermore, the above-mentioned key amino acid-induced new crystal form of trelagliptin based on protein-drug interaction, the 2θ values ​​of the new crystal form TRE-Ⅲ are 9.99°±0.2°, 13.29°±0.2°, 15.27°±0.2°, 22.62°±0.2°, 26.79°±0.2°, 27.90°±0.2°, 30.36°±0.2°, and 40.73°±0.2°.

[0024] Furthermore, the above-mentioned key amino acid-induced new crystal form of trelagliptin based on protein-drug interaction, the 2θ values ​​of the new crystal form TRE-Ⅳ are 5.47°±0.2°, 11.02°±0.2°, 12.88°±0.2°, 16.59°±0.2°, 20.08°±0.2°, 20.91°±0.2°, 22.17°±0.2°, 23.21°±0.2°, 27.84°±0.2°, and 33.55°±0.2°.

[0025] Furthermore, the above-mentioned key amino acid-induced new crystal form of trelagliptin based on protein-drug interaction, the 2θ values ​​of the new crystal form TRE-V are 5.49°±0.2°, 11.04°±0.2°, 12.47°±0.2°, 12.86°±0.2°, 16.59°±0.2°, 20.06°±0.2°, 20.88°±0.2°, 22.19°±0.2°, 28.08°±0.2°, 28.98°±0.2°, 30.47°±0.2°, and 33.54°±0.2°.

[0026] Compared with the prior art, the technical solution provided by the present invention has the following technical advantages:

[0027] 1. The technical solution provided by the present invention is firstly to innovatively introduce key amino acids into the trelagliptin solvent crystallization system, and only combine with trace amounts of organic solvents to obtain a variety of new crystal forms of trelagliptin and preparation methods through the interaction of different solvent systems.

[0028] 2. The amino acids in the technical solution provided by the present invention are essential components of the human body and are essential components for synthesizing all proteins in the human body. When combined in drugs, they are not only non-toxic but also necessary for human growth. At the same time, the use of a large amount of toxic and harmful organic solvents is avoided, and not only a new crystal form of Trelagliptin is prepared, but the preparation process is also accurate, environmentally friendly and safe, which is in line with the national concept of green production.

[0029] 3. The technical solution provided by the present invention is based on the interaction between key amino acids and trelagliptin, and 5 new crystal forms are obtained, which broadens the research on drug crystal forms and provides certain guidance for the production of subsequent drug preparations. BRIEF DESCRIPTION OF THE DRAWINGS

[0030] Figure 1 It is an X-ray powder diffraction spectrum of five new crystalline forms of Trelagliptin induced by key amino acids based on protein-drug interaction structure mining and the original crystalline form of Trelagliptin;

[0031] Figure 2 is the X-ray powder diffraction pattern of the new crystal form I prepared in Example 1;

[0032] Figure 3 is the X-ray powder diffraction pattern of the new crystal form II prepared in Example 2;

[0033] Figure 4 is the X-ray powder diffraction pattern of the new crystal form III prepared in Example 3;

[0034] Figure 5 is the X-ray powder diffraction pattern of the new crystal form IV prepared in Example 4;

[0035] Figure 6 is the X-ray powder diffraction pattern of the new crystalline form V prepared in Example 5;

[0036] Figure 7 is a microscopic observation picture of the new crystal form I prepared in Example 1;

[0037] Figure 8 is a microscopic observation picture of the new crystal form II prepared in Example 2;

[0038] Fig. 9 is a microscopic observation picture of the new crystal form III prepared in Example 3;

[0039] Fig.10 is an observation diagram of the preparation of new crystal form IV in Example 4;

[0040] Fig.11 is an observation diagram of the new crystal form V prepared in Example 5; DETAILED DESCRIPTION

[0041] The following describes various new crystal forms of Trelagliptin induced by key amino acids in a solvent system and their preparation methods in combination with specific implementation methods and the content of the invention, but is not limited to these types, and all those related to this patent are protected.

[0042] Example 1

[0043] 1) Accurately weigh 100 mg of trelagliptin and add it to a 100 mL conical flask. Then add 4 mL of methanol solvent to the conical flask and slowly add dropwise until trelagliptin is just dissolved and saturated. Keep it for later use.

[0044] 2) Take another 6.6 mg of threonine (Thr), add it to the conical flask, and add 2 mL of deionized water to dissolve it until it is saturated.

[0045] 3) Using a rubber-tipped dropper, slowly drip the amino acid solution prepared in step 2) into the solution prepared in step 1) in sequence, and the whole process is carried out in a constant temperature water bath at 37°C.

[0046] 4) After adding, seal with plastic wrap and poke 30-35 holes in the seal, and place in a fume hood to wait for crystals to precipitate.

[0047] 5) After the crystals are precipitated, the growth state of the crystals is observed and recorded using an inverted microscope. The crystals are separated from the solution by filtration, and the collected crystals are placed in a dry environment at T = 30°C for drying, and a portion of the obtained crystals are sent for XPRD measurement.

[0048] The X-ray powder diffraction pattern of crystalline trelagliptin was recorded under the following conditions: room temperature 25°C, relative humidity <60%; XPert PRO polycrystalline X-ray diffractometer (PANalytical, the Netherlands), Cu Kα radiation The light tube voltage is 40 kV, the tube current is 40 mA, the 2θ scanning range is 4 to 50°, the step length is 0.01313°, and the counting time is 30 ms / step. The test results are as follows Figure 2 As shown, the 2θ values ​​of the new crystal form TRE-Ⅰ are 5.66°±0.2°, 11.35°±0.2°, 12.47°±0.2°, 17.08°±0.2°, 19.88°±0.2°, 22.48°±0.2°, 22.87°±0.2°, and 27.54°±0.2°. Compared with the existing crystal forms, there are differences and it is determined to be a new crystal form.

[0049] Example 2

[0050] 1) Accurately weigh 150 mg of trelagliptin and add it to a 100 mL conical flask. Then add 6 mL of methanol solvent to the conical flask and slowly add dropwise until trelagliptin is just dissolved and saturated. Keep it for later use.

[0051] 2) Take 10 mg of tyrosine (Tyr) in a conical flask and add 3 mL of 20% hydrochloric acid solution to dissolve it until it is just saturated.

[0052] 3) Using a rubber-tipped dropper, slowly drip the amino acid solution prepared in step 2) into the solution prepared in step 1) in sequence, and the whole process is carried out in a constant temperature water bath at 50°C.

[0053] 4) After adding, seal with plastic wrap and poke 30-35 holes in the seal, and place in a fume hood to wait for crystals to precipitate.

[0054] 5) After the crystals are precipitated, the growth state of the crystals is observed and recorded using an inverted microscope. The crystals are separated from the solution by filtration, and the collected crystals are placed in a dry environment at T = 30°C for drying, and a portion of the obtained crystals are sent for XPRD measurement.

[0055] The X-ray powder diffraction pattern of crystalline trelagliptin was recorded under the following conditions: room temperature 25°C, relative humidity <60%; XPert PRO polycrystalline X-ray diffractometer (PANalytical, the Netherlands), Cu Kα radiation The light tube voltage is 40 kV, the tube current is 40 mA, the 2θ scanning range is 4 to 50°, the step length is 0.01313°, and the counting time is 30 ms / step. The test results are as follows Figure 3 As shown, the 2θ values ​​of the new crystal form TRE-Ⅱ are 8.07°±0.2°, 13.31°±0.2°, 13.64°±0.2°, 16.29°±0.2°, 16.79°±0.2°, 17.40°±0.2°, 19.40°±0.2°, 21.33°±0.2°, 22.83°±0.2°, 26.26°±0.2°, 26.57°±0.2°, 26.79°±0.2°, 27.49°±0.2°, 27.90°±0.2°, and 33.33°±0.2°. Compared with the existing crystal forms, there are differences and it is determined to be a new crystal form.

[0056] Example 3

[0057] 1) Accurately weigh 250 mg of trelagliptin and add it to a 100 mL conical flask. Then add 8 mL of acetonitrile or 6 mL of acetone solvent to the conical flask and slowly add dropwise until trelagliptin is just dissolved and saturated. Keep it for later use.

[0058] 2) Take 10 mg of tyrosine (Tyr) in a conical flask and add 3 mL of 20% hydrochloric acid solution to dissolve it until it is just saturated.

[0059] 3) Using a rubber-tipped dropper, slowly drip the amino acid solution prepared in step 2) into the solution prepared in step 1) in sequence, and the whole process is carried out in a constant temperature water bath at 37°C.

[0060] 4) After adding, seal with plastic wrap and poke 30-35 holes in the seal, and place in a fume hood to wait for crystals to precipitate.

[0061] 5) After the crystals are precipitated, the growth state of the crystals is observed and recorded using an inverted microscope. The crystals are separated from the solution by filtration, and the collected crystals are placed in a dry environment at T = 30°C for drying, and a portion of the obtained crystals are sent for XPRD measurement.

[0062] The X-ray powder diffraction pattern of crystalline trelagliptin was recorded under the following conditions: room temperature 25°C, relative humidity <60%; XPert PRO polycrystalline X-ray diffractometer (PANalytical, the Netherlands), Cu Kα radiation The light tube voltage is 40 kV, the tube current is 40 mA, the 2θ scanning range is 4 to 50°, the step length is 0.01313°, and the counting time is 30 ms / step. The test results are as follows Figure 5As shown, the 2θ values ​​of the new crystal form TRE-Ⅲ are 9.99°±0.2°, 13.29°±0.2°, 15.27°±0.2°, 22.62°±0.2°, 26.79°±0.2°, 27.90°±0.2°, 30.36°±0.2°, and 40.73°±0.2°. Compared with the existing crystal forms, the differences are determined to be new crystal forms.

[0063] Example 4

[0064] 1) Accurately weigh 100 mg of trelagliptin and add it to a 100 mL conical flask. Then add 4 mL of ethyl acetate solvent to the conical flask and slowly add dropwise until trelagliptin is just dissolved and saturated. Keep it for later use.

[0065] 2) Take 3.3 mg of arginine or 2.7 mg of glutamic acid or a mixture of 3.3 mg of arginine and 2.7 mg of glutamic acid, add one of them to a conical flask, and add 2 mL of deionized water to dissolve until it is just saturated.

[0066] 3) Using a rubber-tipped dropper, slowly drip the amino acid solution prepared in step 2) into the solution prepared in step 1) in sequence, and the whole process is carried out in a constant temperature water bath at 25°C.

[0067] 4) After adding, seal with plastic wrap and poke 30-35 holes in the seal, and place in a fume hood to wait for crystals to precipitate.

[0068] 5) After the crystals are precipitated, the growth state of the crystals is observed and recorded using an inverted microscope. The crystals are separated from the solution by filtration, and the collected crystals are placed in a dry environment at T = 30°C for drying, and a portion of the obtained crystals are sent for XPRD measurement.

[0069] The X-ray powder diffraction pattern of crystalline trelagliptin was recorded under the following conditions: room temperature 25°C, relative humidity <60%; XPert PRO polycrystalline X-ray diffractometer (PANalytical, the Netherlands), Cu Kα radiation The light tube voltage is 40 kV, the tube current is 40 mA, the 2θ scanning range is 4 to 50°, the step length is 0.01313°, and the counting time is 30 ms / step. The test results are as follows Figure 6 As shown, the 2θ values ​​of the new crystal form TRE-Ⅳ are 5.47°±0.2°, 11.02°±0.2°, 12.88°±0.2°, 16.59°±0.2°, 20.08°±0.2°, 20.91°±0.2°, 22.17°±0.2°, 23.21°±0.2°, 27.84°±0.2°, and 33.55°±0.2°. Compared with the existing crystal forms, there are differences and it is determined to be a new crystal form.

[0070] Example 5

[0071] 1) Accurately weigh 100 mg of trelagliptin and add it to a 100 mL conical flask. Then add 4.5 mL of isopropanol solvent to the conical flask and slowly add dropwise until trelagliptin is just dissolved and saturated. Keep it for later use.

[0072] 2) Take 3.3 mg of arginine or 2.7 mg of glutamic acid or a mixture of 3.3 mg of arginine and 2.7 mg of glutamic acid, add one of them to a conical flask, and add 2 mL of deionized water to dissolve until it is just saturated.

[0073] 3) Using a rubber-tipped dropper, slowly drip the amino acid solution prepared in step 2) into the solution prepared in step 1) in sequence, and the whole process is carried out in a constant temperature water bath at 40°C.

[0074] 4) After adding, seal with plastic wrap and poke 30-35 holes in the seal, and place in a fume hood to wait for crystals to precipitate.

[0075] 5) After the crystals are precipitated, the growth state of the crystals is observed and recorded using an inverted microscope. The crystals are separated from the solution by filtration, and the collected crystals are placed in a dry environment at T = 30°C for drying, and a portion of the obtained crystals are sent for XPRD measurement.

[0076] The X-ray powder diffraction pattern of crystalline trelagliptin was recorded under the following conditions: room temperature 25°C, relative humidity <60%; XPert PRO polycrystalline X-ray diffractometer (PANalytical, the Netherlands), Cu Kα radiation The light tube voltage is 40 kV, the tube current is 40 mA, the 2θ scanning range is 4 to 50°, the step length is 0.01313°, and the counting time is 30 ms / step. The test results are as follows Figure 7 As shown, the 2θ values ​​of the new crystal form TRE-Ⅴ are 5.49°±0.2°, 11.04°±0.2°, 12.47°±0.2°, 12.86°±0.2°, 16.59°±0.2°, 20.06°±0.2°, 20.88°±0.2°, 22.19°±0.2°, 28.08°±0.2°, 28.98°±0.2°, 30.47°±0.2°, and 33.54°±0.2°. Compared with the existing crystal forms, there are differences and it is determined to be a new crystal form.

[0077] Example 6

[0078] 1) Accurately weigh 200 mg of trelagliptin and add it to a 100 mL conical flask. Then add 6 mL of ethanol solvent to the conical flask and slowly drop it until the trelagliptin is just dissolved and saturated. Keep it for later use.

[0079] 2) Take 10 mg of tyrosine (Tyr) in a conical flask and add 3 mL of 20% hydrochloric acid solution to dissolve it until it is just saturated.

[0080] 3) Using a rubber-tipped dropper, slowly drip the amino acid solution prepared in step 2) into the solution prepared in step 1) in sequence, and the whole process is carried out in a constant temperature water bath at 40°C.

[0081] 4) After adding, seal with plastic wrap and poke 30-35 holes in the seal, and place in a fume hood to wait for crystals to precipitate.

[0082] 5) After the crystals are precipitated, the growth state of the crystals is observed and recorded using an inverted microscope. The crystals are separated from the solution by filtration, and the collected crystals are placed in a dry environment at T = 30°C for drying, and a portion of the obtained crystals are sent for XPRD measurement.

[0083] The X-ray powder diffraction pattern of crystalline trelagliptin was recorded under the following conditions: room temperature 25°C, relative humidity <60%; XPert PRO polycrystalline X-ray diffractometer (PANalytical, the Netherlands), Cu Kα radiation The light tube voltage is 40 kV, the tube current is 40 mA, the 2θ scanning range is 4 to 50°, the step length is 0.01313°, and the counting time is 30 ms / step. The test results are as follows Figure 4 As shown, the 2θ values ​​of the new crystal form TRE-Ⅱ are 8.07°±0.2°, 13.31°±0.2°, 13.64°±0.2°, 16.29°±0.2°, 16.79°±0.2°, 17.40°±0.2°, 19.40°±0.2°, 21.33°±0.2°, 22.83°±0.2°, 26.26°±0.2°, 26.57°±0.2°, 26.79°±0.2°, 27.49°±0.2°, 27.90°±0.2°, and 33.33°±0.2°. Compared with the existing crystal forms, there are differences and it is determined to be a new crystal form.

[0084] Example 7

[0085] 1) Accurately weigh 100 mg of trelagliptin and add it to a 100 mL conical flask. Then add 4 mL of acetone to the conical flask at 25°C-50°C until the trelagliptin is just dissolved and saturated. Keep it for later use.

[0086] 2) Take another 10 mg of tyrosine (TYR) and put it into a conical flask, and add 3 mL of 20% hydrochloric acid to dissolve it until it is saturated.

[0087] 3) Using a rubber-tipped dropper, slowly drip the amino acid solution prepared in step 2) into the solution prepared in step 1) in sequence, and the whole process is carried out in a constant temperature water bath at 50° C.;

[0088] 4) After adding, seal with plastic wrap and poke 30-35 holes in the seal, and place in a fume hood to wait for crystals to precipitate.

[0089] 5) After the crystals are precipitated, the growth state of the crystals is observed and recorded using an inverted microscope. The crystals are separated from the solution by filtration, and the collected crystals are placed in a dry environment at T = 30°C for drying, and a portion of the obtained crystals are sent for XPRD measurement.

[0090] The X-ray powder diffraction pattern of crystalline trelagliptin was recorded under the following conditions: room temperature 25°C, relative humidity <60%; XPert PRO polycrystalline X-ray diffractometer (PANalytical, the Netherlands), Cu Kα radiation The light tube voltage is 40 kV, the tube current is 40 mA, the 2θ scanning range is 4 to 50°, the step length is 0.01313°, and the counting time is 30 ms / step. The test results are as follows Figure 6 As shown, the 2θ values ​​of the new crystal form TRE-Ⅲ are 9.99°±0.2°, 13.29°±0.2°, 15.27°±0.2°, 22.62°±0.2°, 26.79°±0.2°, 27.90°±0.2°, 30.36°±0.2°, and 40.73°±0.2°. Compared with the existing crystal forms, the differences are determined to be new crystal forms.

[0091] Example 8

[0092] 1) Accurately weigh 100 mg of trelagliptin and add it to a 100 mL conical flask. Then add 5.5 mL of isopropanol solvent to the conical flask and slowly add dropwise until trelagliptin is just dissolved and saturated. Keep it for later use.

[0093] 2) Take 4.9 mg of arginine or 4.1 mg of glutamic acid or a mixture of 4.9 mg of arginine and 4.1 mg of glutamic acid, add one of them to a conical flask, and add 3 mL of deionized water to dissolve until it is just saturated.

[0094] 3) Using a rubber-tipped dropper, slowly drip the amino acid solution prepared in step 2) into the solution prepared in step 1) in sequence, and the whole process is carried out in a constant temperature water bath at 40°C.

[0095] 4) After adding, seal with plastic wrap and poke 30-35 holes in the seal, and place in a fume hood to wait for crystals to precipitate.

[0096] 5) After the crystals are precipitated, the growth state of the crystals is observed and recorded using an inverted microscope. The crystals are separated from the solution by filtration, and the collected crystals are placed in a dry environment at T = 30°C for drying, and a portion of the obtained crystals are sent for XPRD measurement.

[0097] The X-ray powder diffraction pattern of crystalline trelagliptin was recorded under the following conditions: room temperature 25°C, relative humidity <60%; XPert PRO polycrystalline X-ray diffractometer (PANalytical, the Netherlands), Cu Kα radiation The light tube voltage is 40 kV, the tube current is 40 mA, the 2θ scanning range is 4 to 50°, the step length is 0.01313°, and the counting time is 30 ms / step. The test results are as follows Figure 7 As shown, the 2θ values ​​of the new crystal form TRE-Ⅴ are 5.49°±0.2°, 11.04°±0.2°, 12.47°±0.2°, 12.86°±0.2°, 16.59°±0.2°, 20.06°±0.2°, 20.88°±0.2°, 22.19°±0.2°, 28.08°±0.2°, 28.98°±0.2°, 30.47°±0.2°, and 33.54°±0.2°. Compared with the existing crystal forms, there are differences and it is determined to be a new crystal form.

[0098] Example 9

[0099] 1) Accurately weigh 200 mg of trelagliptin and add it to a 100 mL conical flask. Then add 5 mL of ethanol solvent to the conical flask and slowly drop it until the trelagliptin is just dissolved and saturated. Keep it for later use.

[0100] 2) Take another 20 mg of tyrosine (Tyr) in a conical flask, add 6 mL of 20% hydrochloric acid solution and dissolve until it is just saturated.

[0101] 3) Use a rubber-tipped dropper to slowly drip the amino acid solution prepared in 2) into the solution prepared in 1) in sequence, and the whole process is carried out in a constant temperature water bath at 37°C.

[0102] 4) After adding, seal with plastic wrap and poke 30-35 holes in the seal, and place in a fume hood to wait for crystals to precipitate.

[0103] 5) After the crystals are precipitated, the growth state of the crystals is observed and recorded using an inverted microscope. The crystals are separated from the solution by filtration, and the collected crystals are placed in a dry environment at T = 30°C for drying, and a portion of the obtained crystals are sent for XPRD measurement.

[0104] The X-ray powder diffraction pattern of crystalline trelagliptin was recorded under the following conditions: room temperature 25°C, relative humidity <60%; XPert PRO polycrystalline X-ray diffractometer (PANalytical, the Netherlands), Cu Kα radiation The light tube voltage is 40 kV, the tube current is 40 mA, the 2θ scanning range is 4 to 50°, the step length is 0.01313°, and the counting time is 30 ms / step. The test results are as follows Figure 4 As shown, the 2θ values ​​of the new crystal form TRE-Ⅱ are 8.07°±0.2°, 13.31°±0.2°, 13.64°±0.2°, 16.29°±0.2°, 16.79°±0.2°, 17.40°±0.2°, 19.40°±0.2°, 21.33°±0.2°, 22.83°±0.2°, 26.26°±0.2°, 26.57°±0.2°, 26.79°±0.2°, 27.49°±0.2°, 27.90°±0.2°, and 33.33°±0.2°. Compared with the existing crystal form, there are differences and it is determined to be a new crystal form.

[0105] Example 10

[0106] 1) Accurately weigh 100 mg of trelagliptin and add it to a 100 mL conical flask. Then add 4.5 mL of isopropanol solvent to the conical flask and slowly add dropwise until trelagliptin is just dissolved and saturated. Keep it for later use.

[0107] 2) Take 10 mg of arginine or 8 mg of glutamic acid or a mixture of 10 mg of arginine and 8 mg of glutamic acid, add one of them to a conical flask, and add 2 mL of deionized water to dissolve until it is just saturated.

[0108] 3) Using a rubber-tipped dropper, slowly drip the amino acid solution prepared in step 2) into the solution prepared in step 1) in sequence, and the whole process is carried out in a constant temperature water bath at 45°C.

[0109] 4) After adding, seal with plastic wrap and poke 30-35 holes in the seal, and place in a fume hood to wait for crystals to precipitate.

[0110] 5) After the crystals are precipitated, the growth state of the crystals is observed and recorded using an inverted microscope. The crystals are separated from the solution by filtration, and the collected crystals are placed in a dry environment at T = 30°C for drying, and a portion of the obtained crystals are sent for XPRD measurement.

[0111] The X-ray powder diffraction pattern of crystalline trelagliptin was recorded under the following conditions: room temperature 25°C, relative humidity <60%; XPert PRO polycrystalline X-ray diffractometer (PANalytical, the Netherlands), Cu Kα radiation (λ = 1.540

[0112] ), light tube voltage 40kV, tube current 40mA, 2θ scanning range 4~50°, step length 0.01313°, counting time 30ms / step, the detection results are as follows Figure 7As shown, the 2θ values ​​of the new crystal form TRE-Ⅴ are 5.49°±0.2°, 11.04°±0.2°, 12.47°±0.2°, 12.86°±0.2°, 16.59°±0.2°, 20.06°±0.2°, 20.88°±0.2°, 22.19°±0.2°, 28.08°±0.2°, 28.98°±0.2°, 30.47°±0.2°, and 33.54°±0.2°. Compared with the existing crystal forms, there are differences and it is determined to be a new crystal form.

[0113] Embodiment 11

[0114] 1) Accurately weigh 100 mg of trelagliptin and add it to a 100 mL conical flask. Then add 5.5 mL of ethyl acetate solvent to the conical flask and slowly add dropwise until trelagliptin is just dissolved and saturated. Keep it for later use.

[0115] 2) Take 10 mg of arginine or 8 mg of glutamic acid or a mixture of 10 mg of arginine and 8 mg of glutamic acid, add one of them to a conical flask, and add 2 mL of deionized water to dissolve until it is just saturated.

[0116] 3) Using a rubber-tipped dropper, slowly drip the amino acid solution prepared in step 2) into the solution prepared in step 1) in sequence, and the whole process is carried out in a constant temperature water bath at 50°C.

[0117] 4) After adding, seal with plastic wrap and poke 30-35 holes in the seal, and place in a fume hood to wait for crystals to precipitate.

[0118] 5) After the crystals are precipitated, the growth state of the crystals is observed and recorded using an inverted microscope. The crystals are separated from the solution by filtration, and the collected crystals are placed in a dry environment at T = 30°C for drying, and a portion of the obtained crystals are sent for XPRD measurement.

[0119] The X-ray powder diffraction pattern of crystalline trelagliptin was recorded under the following conditions: room temperature 25°C, relative humidity <60%; XPert PRO polycrystalline X-ray diffractometer (PANalytical, the Netherlands), Cu Kα radiation The light tube voltage is 40 kV, the tube current is 40 mA, the 2θ scanning range is 4 to 50°, the step length is 0.01313°, and the counting time is 30 ms / step. The test results are as follows Figure 6As shown, the 2θ values ​​of the new crystal form TRE-Ⅳ are 5.47°±0.2°, 11.02°±0.2°, 12.88°±0.2°, 16.59°±0.2°, 20.08°±0.2°, 20.91°±0.2°, 22.17°±0.2°, 23.21°±0.2°, 27.84°±0.2°, and 33.55°±0.2°. Compared with the existing crystal forms, there are differences and it is determined to be a new crystal form.

[0120] Example 12

[0121] 1) Accurately weigh 1 portion of 250 mg of trelagliptin and add it to a 100 mL conical flask. Then add 6 mL of acetonitrile to the conical flask at 25°C-50°C until the trelagliptin is just dissolved and saturated. Keep it for later use.

[0122] 2) Take another 12.8 mg of tyrosine (TYR) and add it into a conical flask, then add 6.5 mL of 20% hydrochloric acid to dissolve it until it is saturated.

[0123] 3) Using a rubber-tipped dropper, slowly drip the amino acid solution prepared in step 2) into the solution prepared in step 1) in sequence, and the whole process is carried out in a constant temperature water bath at 50°C.

[0124] 4) After adding, seal with plastic wrap and poke 30-35 holes in the seal, and place in a fume hood to wait for crystals to precipitate.

[0125] 5) After the crystals are precipitated, the growth state of the crystals is observed and recorded using an inverted microscope. The crystals are separated from the solution by filtration, and the collected crystals are placed in a dry environment at T = 30°C for drying, and a portion of the obtained crystals are sent for XPRD measurement.

[0126] The X-ray powder diffraction pattern of crystalline trelagliptin was recorded under the following conditions: room temperature 25°C, relative humidity <60%; XPert PRO polycrystalline X-ray diffractometer (PANalytical, the Netherlands), Cu Kα radiation The light tube voltage is 40 kV, the tube current is 40 mA, the 2θ scanning range is 4 to 50°, the step length is 0.01313°, and the counting time is 30 ms / step. The test results are as follows Figure 6 As shown, the 2θ values ​​of the new crystal form TRE-Ⅲ are 9.99°±0.2°, 13.29°±0.2°, 15.27°±0.2°, 22.62°±0.2°, 26.79°±0.2°, 27.90°±0.2°, 30.36°±0.2°, and 40.73°±0.2°. Compared with the existing crystal forms, the differences are determined to be new crystal forms.

[0127] In order to demonstrate the effects of the five new crystalline forms of Trelagliptin provided in the present application, the solubility of the five new crystalline forms and the original crystalline form in pH=1.2 hydrochloric acid solution, pH=7 aqueous solution, and pH=6.8 and pH=7.4 PBS solutions were determined below.

[0128] The specific operation is: take samples of 5 new crystal forms and the original crystal form after reaching dissolution equilibrium, filter with a 0.22um filter membrane, take an appropriate amount of filtrate, and measure the absorbance value at a wavelength of 279nm according to the UV-Vis spectrophotometry (Chinese Pharmacopoeia 2020 edition). Substitute it into the standard curve to obtain the solubility of the 5 new crystal forms and the original crystal form, as shown in Table 1.

[0129] Table 1 Solubility table

[0130]

[0131] The results in Table 1 show that the solubility of the five new crystalline forms of Trelagliptin found in pH=1.2 hydrochloric acid solution, deionized water (pH=7.0), pH=6.8 sodium phosphate buffer solution, and pH=7.4 sodium phosphate buffer solution is significantly higher than that of the original crystalline form. The high solubility of the new crystalline form can make up for the defect of poor solubility of the original crystalline form of Trelagliptin.

[0132] The above description is only a specific implementation mode of the present invention, not all implementation modes. Any equivalent transformations made by other technicians in the field based on the description of the present invention are all covered by the claims of the present invention and are also the contents protected by the present invention.

Claims

1. A method for preparing a new crystal form of Trelagliptin based on key amino acids inducing protein-drug interaction, characterized in that: Amino acids were introduced into the solvent crystallization system of trelagliptin to induce the production of five new crystal forms of trelagliptin.

2. The method for preparing a new crystal form of Trelagliptin induced by key amino acids based on protein-drug interaction according to claim 1, characterized in that: The steps are as follows: 1) slowly adding a solvent dropwise to trelagliptin until trelagliptin is just dissolved to saturation, thereby obtaining a trelagliptin solution for use; 2) Add deionized water or 20% hydrochloric acid solution to the amino acid until the amino acid is saturated to obtain an amino acid solution; 3) Then, using a rubber-tipped dropper, the amino acid solution prepared in 2) is slowly dripped into the trelagliptin solution prepared in 1) in sequence, and the whole process is carried out in a constant temperature water bath at 25° C.-50° C.; 4) After the addition is completed, seal it with plastic wrap, and poke 20-35 holes in the seal, place it in a fume hood and wait for crystals to precipitate, thereby inducing the production of multiple new crystal forms of trelagliptin; The molar ratio of the amino acid to trelagliptin is 1:2 to 1:

30.

3. The method for preparing a new crystal form of Trelagliptin induced by key amino acids based on protein-drug interaction according to claim 1 or 2, characterized in that: The amino acid is one of glutamic acid, arginine, threonine, and tyrosine, or any combination thereof; the solvent for dissolving trelagliptin is one of ethyl acetate, isopropanol, acetone, methanol, ethanol, and acetonitrile.

4. A novel crystal form of Trelagliptin induced by the key amino acid based on protein-drug interaction according to claim 1 or 2, characterized in that: The new crystal forms of Trelagliptin are TRE-Ⅰ, TRE-Ⅱ, TRE-Ⅲ, TRE-Ⅳ and TRE-Ⅴ.

5. The new crystal form of Trelagliptin induced by key amino acids based on protein-drug interaction according to claim 4, characterized in that: The 2θ values ​​of the X-ray powder diffraction pattern of the new crystal form TRE-Ⅰ are 5.66°±0.2°, 11.35°±0.2°, 12.47°±0.2°, 17.08°±0.2°, 19.88°±0.2°, 22.48°±0.2°, 22.87°±0.2°, and 27.54°±0.2°.

6. The new crystal form of Trelagliptin induced by key amino acids based on protein-drug interaction according to claim 4, characterized in that: The 2θ values ​​of the new crystal form TRE-Ⅱ are 8.07°±0.2°, 13.31°±0.2°, 13.64°±0.2°, 16.29°±0.2°, 16.79°±0.2°, 17.40°±0.2°, 19.40°±0.2°, 21.33°±0.2°, 22.83°±0.2°, 26.26°±0.2°, 26.57°±0.2°, 26.79°±0.2°, 27.49°±0.2°, 27.90°±0.2°, and 33.33°±0.2°.

7. The new crystal form of Trelagliptin induced by key amino acids based on protein-drug interaction according to claim 4, characterized in that: The 2θ values ​​of the new crystal form TRE-Ⅲ are 9.99°±0.2°, 13.29°±0.2°, 15.27°±0.2°, 22.62°±0.2°, 26.79°±0.2°, 27.90°±0.2°, 30.36°±0.2°, and 40.73°±0.2°.

8. The new crystal form of Trelagliptin induced by key amino acids based on protein-drug interaction according to claim 4, characterized in that: The 2θ values ​​of the new crystal form TRE-Ⅳ are 5.47°±0.2°, 11.02°±0.2°, 12.88°±0.2°, 16.59°±0.2°, 20.08°±0.2°, 20.91°±0.2°, 22.17°±0.2°, 23.21°±0.2°, 27.84°±0.2°, and 33.55°±0.2°.

9. The new crystal form of Trelagliptin induced by key amino acids based on protein-drug interaction according to claim 4, characterized in that: The 2θ values ​​of the new crystal form TRE-V are 5.49°±0.2°, 11.04°±0.2°, 12.47°±0.2°, 12.86°±0.2°, 16.59°±0.2°, 20.06°±0.2°, 20.88°±0.2°, 22.19°±0.2°, 28.08°±0.2°, 28.98°±0.2°, 30.47°±0.2°, and 33.54°±0.2°.

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