CN112552361A

CN112552361A - Cytarabine crystal, cytarabine freeze-dried powder and freeze-drying method

Info

Publication number: CN112552361A
Application number: CN202011629349.3A
Authority: CN
Inventors: 杜加秋; 李湘萍; 赵技宇; 蔡邱华; 董福霞; 易芬芬; 夏莉
Original assignee: Hanhui Pharmaceutical Co ltd
Current assignee: Hanhui Pharmaceutical Co ltd
Priority date: 2020-12-30
Filing date: 2020-12-30
Publication date: 2021-03-26

Abstract

The invention provides cytarabine crystal, cytarabine freeze-dried powder and a freeze-drying method. The XRD diffraction pattern of the cytarabine crystal has characteristic peaks expressed in 2 theta at the following positions: 9.962 degrees +/-0.2 degrees, 12.680 degrees +/-0.2 degrees, 14.165 degrees +/-0.2 degrees, 15.993 degrees +/-0.2 degrees, 18.670 degrees +/-0.2 degrees, 19.491 degrees +/-0.2 degrees and 25.881 degrees +/-0.2 degrees. Experiments prove that compared with the amorphous structure of the existing cytarabine for injection, the cytarabine crystal form with the characteristic peak has the advantages that the stability is obviously improved, and the hydrolyzability is obviously reduced, so that the stability of cytarabine powder can be improved on the basis of not changing the formula of the cytarabine powder based on the cytarabine crystal form.

Description

Cytarabine crystal, cytarabine freeze-dried powder and freeze-drying method

Technical Field

The invention relates to the technical field of preparation of cytarabine, and particularly relates to a cytarabine crystal, cytarabine freeze-dried powder and a freeze-drying method.

Background

Cytarabine is an ancient anticancer drug, the earliest application of which began in 1969. It is also a versatile cancer therapeutic drug, such as cytarabine in combination with anthracycline, which can treat acute myeloid leukemia and lymphoma, among others, and is also common in alternatives for the treatment of other cancers. In particular, for certain chemotherapy regimens, the combination of chemotherapeutic agents with cytarabine may result in a cure rate of 40% to 50% for leukemia, but 0% if no cytarabine is involved in the treatment. The difference between the availability of cytarabine in these patients is life and death.

Chinese patent application No. 201911322514.8 discloses an cytarabine injection, which comprises 10-30 parts of cytarabine, 5-10 parts of benzyl alcohol, 1-5 parts of sodium chloride, 1-5 parts of sodium bicarbonate and water for injection with the constant volume of 1000 parts by mass. In the technology, the degradation product Ara-U of cytarabine is obviously reduced under the neutral alkalescent condition, the sodium bicarbonate and the sodium chloride are added, the condition that the raw material medicine is always kept at the neutral or slightly alkaline condition in the liquid preparation process can be controlled, the risk of acid damage of the raw material medicine is reduced, the stability of the product is increased, the permeation regulator is added, the preparation and development process of the preparation is optimized, the safety risk in the injection process can be effectively prevented, and the product with the stability superior to that of the original product is produced.

However, the above-mentioned arabinoside injection has complex components and is limited in the production and approval of medicines. Cytarabine, however, is susceptible to hydrolysis to form uridine arabinoside, and thus, there is a need to provide a means for increasing the stability of cytarabine.

Disclosure of Invention

The invention mainly aims to provide cytarabine crystals, cytarabine freeze-dried powder and a freeze-drying method, and aims to solve the problem of poor stability of cytarabine for injection in the prior art.

In order to achieve the above object, according to one aspect of the present invention, there is provided a cytarabine crystal having an XRD diffraction pattern with characteristic peaks expressed in 2 θ at the following positions: 9.962 degrees +/-0.2 degrees, 12.680 degrees +/-0.2 degrees, 14.165 degrees +/-0.2 degrees, 15.993 degrees +/-0.2 degrees, 18.670 degrees +/-0.2 degrees, 19.491 degrees +/-0.2 degrees and 25.881 degrees +/-0.2 degrees.

Further, the XRD diffraction pattern of the cytarabine crystalline substance described above also has characteristic peaks expressed in 2 θ at the following positions: 7.660 degrees +/-0.2 degree, 13.233 degrees +/-0.2 degree, 14.944 degrees +/-0.2 degree, 17.213 degrees +/-0.2 degree, 20.611 degrees +/-0.2 degree, 22.054 degrees +/-0.2 degree, 22.921 degrees +/-0.2 degree, 23.732 degrees +/-0.2 degree, 22.921 degrees +/-0.2 degree, 23.723 degrees +/-0.2 degree, 24.577 degrees +/-0.2 degree and 28.164 degrees +/-0.2 degree.

According to another aspect of the present invention, there is provided a cytarabine lyophilized powder comprising cytarabine crystals of any one of the above.

According to another aspect of the invention, a method for lyophilizing cytarabine is provided, the method comprising sequentially prefreezing, sublimation drying and analytical drying a cytarabine solution to obtain a lyophilized cytarabine powder, the prefreezing comprising sequentially performing the following steps: cooling to-40 to-30 ℃ in 30-50 min and preserving heat for 70-105 min; heating to-25 to-15 ℃ for 10-30 min and preserving heat for 70-105 min; reducing the temperature to-40 to-30 ℃ in 10-25 min and preserving the temperature for 100-150 min.

Further, the pre-freezing comprises the following steps which are carried out in sequence: cooling to-40 to-30 ℃ after 40 to 50min and preserving heat for 80 to 90 min; heating to-20-15 ℃ for 10-20 min and keeping the temperature for 80-100 min; reducing the temperature to-40 to-30 ℃ in 10-25 min and preserving the temperature for 120-130 min.

Further, the temperature of sublimation drying is-35-10 ℃, a gradient heating mode is preferably adopted for sublimation drying, and the preferred sublimation drying process comprises the following steps: under the first vacuum condition, heating the pre-frozen product obtained by pre-freezing to a first sublimation drying temperature for carrying out first-stage sublimation drying to obtain a first sublimation dried product; under a second vacuum condition, heating the first sublimation-dried product to a second sublimation-drying temperature, and carrying out second-stage sublimation drying to obtain a second sublimation-dried product; wherein the first sublimation drying temperature is-25 to-10 ℃, the second sublimation drying temperature is 0 to 10 ℃, and the gauge pressures of the first vacuum condition and the second vacuum condition are respectively and independently 0.1 to 0.5 mbar.

Further, the pre-frozen product is heated to the first sublimation drying temperature within 70-100 min and is kept warm for 270-330 min for carrying out the first stage sublimation drying.

Further, the first sublimation-dried product is heated to a second sublimation-drying temperature within 70-100 min and is kept warm for 100-150 min for carrying out second-stage sublimation-drying.

Further, the temperature for the desorption drying is 10 to 50 ℃.

Further, the analysis drying is performed by a gradient temperature rise method, and preferably the analysis drying includes: under the third vacuum condition, heating a sublimation-dried product obtained by sublimation drying to a first analysis drying temperature to carry out first-stage analysis drying to obtain a first analysis product; heating the first analytic product to a second analytic drying temperature under a fourth vacuum condition, and carrying out second-stage analytic drying to obtain cytarabine freeze-dried powder, wherein the first analytic drying temperature is 10-30 ℃, the second analytic drying temperature is 25-45 ℃, gauge pressures of the third vacuum condition and the fourth vacuum condition are respectively and independently 0.1-0.5 mbar, preferably, heating the sublimation-dried product to the first analytic drying temperature within 10-30 min, and carrying out heat preservation for 100-150 min to carry out first-stage analytic drying; preferably, the first analysis product is heated to the second analysis drying temperature within 10-30 min, and the temperature is kept for 200-250 min to perform the second stage analysis drying.

By applying the technical scheme of the invention, experiments prove that compared with the amorphous structure of the existing cytarabine for injection, the cytarabine crystal form with the characteristic peak has obviously improved stability and obviously reduced hydrolyzability, so that the stability of cytarabine powder can be improved on the basis of not changing the formula of the cytarabine powder.

Drawings

The accompanying drawings, which are incorporated in and constitute a part of this application, illustrate embodiments of the invention and, together with the description, serve to explain the invention and not to limit the invention. In the drawings:

FIG. 1 shows XRD diffraction patterns of cytarabine as a raw material used in examples according to the present invention and comparative examples;

figure 2 shows the XRD diffraction pattern of the lyophilisate obtained in example 1;

fig. 3 shows an XRD diffraction pattern of the freeze-dried product obtained in comparative example 1; and

figure 4 shows an XRD diffraction pattern corresponding to the reference formulation of table 15.

Detailed Description

It should be noted that the embodiments and features of the embodiments in the present application may be combined with each other without conflict. The present invention will be described in detail below with reference to the embodiments with reference to the attached drawings.

As analyzed in the background of the application, the aim of improving the stability of cytarabine is fulfilled by improving the formula of cytarabine for injection in the prior art, but the formula of the injection in the current drug approval is strictly approved, and the formula of the original medicine is generally not allowed to be changed. Based on the foregoing background, there is a need for improving the stability of cytarabine without changing its formulation.

In order to solve the above problems, the present application attempts to form cytarabine crystals having a specific crystal structure to improve the stability thereof. Based on the research on the crystal structure of cytarabine, the application provides a cytarabine crystal, cytarabine freeze-dried powder and a freeze-drying method.

In an exemplary embodiment of the present application, provided is a cytarabine crystalline substance having an XRD diffraction pattern with characteristic peaks expressed in 2 θ at the following positions:

9.962°±0.2°、12.680°±0.2°、14.165°±0.2°、15.993°±0.2°、18.670°±0.2°、19.491°±0.2°、25.881°±0.2°。

experiments prove that compared with the amorphous structure of the existing cytarabine for injection, the cytarabine crystal form with the characteristic peak has the advantages that the stability is obviously improved, and the hydrolyzability is obviously reduced, so that the stability of cytarabine powder can be improved on the basis of not changing the formula of the cytarabine powder based on the cytarabine crystal form.

In addition, the XRD diffraction pattern of the cytarabine crystalline substance preferably further has characteristic peaks expressed in 2 θ at the following positions:

7.660°±0.2°、13.233°±0.2°、14.944°±0.2°、17.213°±0.2°、20.611°±0.2°、22.054°±0.2°、22.921°±0.2°、23.732°±0.2°、22.921°±0.2°、23.723°±0.2°、24.577°±0.2°、28.164°±0.2°。

in another exemplary embodiment of the present application, there is provided a cytarabine lyophilized powder comprising cytarabine crystals of any one of the above. Because the cytarabine freeze-dried powder has the cytarabine crystal, the hydrolysis resistance and the stability of the cytarabine freeze-dried powder are improved, and further, when the cytarabine freeze-dried preparation is prepared, the water content of a rubber plug or other packaging materials does not need to be controlled in a customized mode, and the cytarabine can be prevented from being hydrolyzed.

In another exemplary embodiment of the present application, a method for lyophilizing cytarabine is provided, the method for lyophilizing cytarabine includes subjecting a cytarabine solution to prefreezing, sublimation drying, and analytical drying in sequence to obtain a lyophilized cytarabine powder, the prefreezing includes the following steps in sequence: cooling to-40 to-30 ℃ in 30-50 min and preserving heat for 70-105 min; heating to-25 to-15 ℃ for 10-30 min and preserving heat for 70-105 min; reducing the temperature to-40 to-30 ℃ in 10-25 min and preserving the temperature for 100-150 min.

Processes for preparing lyophilized formulations that are conventional in the art typically include prefreezing, sublimation drying, and/or analytical drying, but the effect on the formulation of the procedures for the various stages of the drug will vary. According to the method, an annealing process, namely a process of firstly cooling, then heating and then cooling, is adopted in the pre-freezing stage, in the process, part of cytarabine is transformed into a crystal structure with the characteristic peak, and then the cytarabine freeze-dried powder with hydrolytic stability is obtained by using the freeze-drying method.

In order to increase the crystallinity of cytarabine and control the grain size, it is preferred that the prefreezing comprises the following steps performed in sequence: cooling to-40 to-30 ℃ after 40 to 50min and preserving heat for 80 to 90 min; heating to-20-15 ℃ for 10-20 min and keeping the temperature for 80-100 min; reducing the temperature to-40 to-30 ℃ in 10-25 min and preserving the temperature for 120-130 min. Under the control of the pre-freezing parameters, not only can full crystallization be realized, but also the influence of undersize crystal grains caused by rapid cooling on subsequent drying can be avoided, and the grain size easy to redissolve can be obtained.

The freeze-drying process of this application adopts the freeze-drying equipment commonly used at present to realize, sets for the case temperature of advancing, the parameter of prefreezing, sublimation drying parameter and the analysis drying parameter of freeze-drying equipment before prefreezing usually. The box inlet temperature of the device can be the conventional box inlet temperature, such as between 5 and 20 ℃. The crystal structure of the cytarabine is formed after prefreezing, and the subsequent sublimation drying and the analytic drying mainly remove the solvent in the cytarabine solution, wherein the solvent in the cytarabine solution can be water for injection commonly used in the field, and the concentration of the cytarabine in the cytarabine solution is 25 mg/ml-85 mg/ml.

The sublimation in-process is mainly got rid of free solvent, and for control sublimation drying speed, preferred above-mentioned sublimation drying's temperature is-35 ~ 10 ℃, preferably adopts gradient intensification mode to carry out sublimation drying, and preferred sublimation drying's process includes: under the first vacuum condition, heating the pre-frozen product obtained by pre-freezing to a first sublimation drying temperature for carrying out first-stage sublimation drying to obtain a first sublimation dried product; under a second vacuum condition, heating the first sublimation-dried product to a second sublimation-drying temperature, and carrying out second-stage sublimation drying to obtain a second sublimation-dried product; wherein the first sublimation drying temperature is-25 to-10 ℃, the second sublimation drying temperature is 0 to 10 ℃, and the gauge pressures of the first vacuum condition and the second vacuum condition are respectively and independently 0.1 to 0.5 mbar. By the staged gradient temperature rise manner, the drying speed and the redissolution property can be adjusted as required.

In one embodiment, the pre-frozen product is preferably heated to the first sublimation drying temperature within 70-100 min and then kept at the first sublimation drying temperature for 270-330 min for the first sublimation drying. Preferably, the first sublimation-dried product is heated to the second sublimation-drying temperature within 70-100 min and is kept at the temperature for 100-150 min for the second stage sublimation-drying. The control of the sublimation process in the two stages realizes the gradual sublimation separation of the free solvent at a controllable speed, and avoids the problem that the agglomeration hardness caused by the excessive sublimation greatly affects the re-dissolution.

The desorption drying temperature is preferably 10 to 50 ℃ because the adsorption solvent on the surface of cytarabine crystals is mainly removed, and thus the temperature is higher than the sublimation drying temperature.

In some embodiments, the desorption drying is performed by using a gradient temperature rise method. For example, the desorption drying includes: under the third vacuum condition, heating a sublimation-dried product obtained by sublimation drying to a first analysis drying temperature to carry out first-stage analysis drying to obtain a first analysis product; and under the fourth vacuum condition, heating the first analysis product to a second analysis drying temperature for second-stage analysis drying to obtain cytarabine freeze-dried powder, wherein the first analysis drying temperature is 10-30 ℃, the second analysis drying temperature is 25-45 ℃, gauge pressures of the third vacuum condition and the fourth vacuum condition are respectively 0.1-0.5 mbar, and the adsorption solvent is gradually removed by the aid of the staged analysis drying. Preferably, the sublimation dried product is heated to the first analysis drying temperature within 10-30 min, and is kept warm for 100-150 min to carry out the first stage analysis drying; preferably, the first analysis product is heated to the second analysis drying temperature within 10-30 min, and the temperature is kept for 200-250 min to perform the second stage analysis drying. By utilizing the control of the conditions, the damage of the crystal structure in the resolving process is avoided, and the aim of fully removing the solvent is fulfilled.

The advantageous effects of the present application will be further described below with reference to examples and comparative examples.

In the following examples, the powder diffraction results of cytarabine are shown in FIG. 1. The cytarabine is mixed with water for injection, sterilized and filtered by a microporous filter membrane to obtain a cytarabine solution with the concentration of 50mg/ml in a sterile room, subpackaged in a penicillin bottle and half stoppered.

Example 1

The lyophilization process is shown in table 1.

TABLE 1

Example 2

The lyophilization process is shown in table 2.

TABLE 2

Example 3

The lyophilization process is shown in table 3.

TABLE 3

Example 4

The lyophilization process is shown in table 4.

TABLE 4

Example 5

The lyophilization process is shown in table 5.

TABLE 5

Example 6

The lyophilization process is shown in table 6.

TABLE 6

Example 7

The lyophilization process is shown in table 7.

TABLE 7

Example 8

The lyophilization process is shown in table 8.

TABLE 8

Example 9

The lyophilization process is shown in table 9.

TABLE 9

Example 10

The lyophilization process is shown in table 10.

Watch 10

Example 11

The lyophilization process is shown in table 11.

TABLE 11

Comparative example 1

The lyophilization process is shown in table 12.

TABLE 12

The freeze-dried products obtained by freeze-drying in each example are subjected to crystal form detection, and the XRD diffraction pattern of each freeze-dried product in each example is found to have characteristic peaks at 9.962 degrees +/-0.2 degrees, 12.680 degrees +/-0.2 degrees, 14.165 degrees +/-0.2 degrees, 15.993 degrees +/-0.2 degrees, 18.670 degrees +/-0.2 degrees, 19.491 degrees +/-0.2 degrees and 25.881 degrees +/-0.2 degrees, but the crystallinity is different. Wherein fig. 2 shows an XRD diffraction pattern of the freeze-dried product obtained in example 1, and fig. 3 shows an XRD diffraction pattern of the freeze-dried product obtained in comparative example 1, it can be seen that it is amorphous.

The stability of the freeze-dried powder obtained in each example and comparative example is tested by using a CP2020 cytarabine hydrochloride quality standard method for injection, the stability test result of example 1 is shown in Table 13, and the stability test result of comparative example 1 is shown in Table 14.

Watch 13

TABLE 14

In addition, the stability of the reference formulation (cytarabine for injection, size 0.1g, Pfizer Italia s.r.l., manufactured lot 7ND5111) was also determined by the above method and the results are shown in table 15, and the XRD diffractogram is shown in fig. 4, showing that it is an amorphous structure.

Watch 15

According to the comparison of the content of cytarabine in tables 13 and 15, the freeze-dried powder of the present application can hydrolyze the impurity uridine more slowly than the original developer under high temperature condition, so the stability of the prepared sample is improved.

From the diffraction patterns, the crystallinity of the crystals in the freeze-dried products obtained in each example was calculated and recorded in table 13, and the resolubility of each freeze-dried product was examined by the quality standard method using cytarabine hydrochloride for injection of CP2020 and the results are recorded in table 13. The uridine contents of the lyophilized powders of the respective examples and comparative examples measured at 0 days and the uridine contents (%) after treatment at 60 ℃ for 30 days are shown in Table 16.

TABLE 16

From the above description, it can be seen that the above-described embodiments of the present invention achieve the following technical effects:

experiments prove that compared with the amorphous structure of the existing cytarabine for injection, the cytarabine crystal form with the characteristic peak has the advantages that the stability is obviously improved, the hydrolyzability is obviously reduced, and other key quality control items are not influenced, so that the stability of powder can be improved on the basis of not changing the formula of cytarabine powder based on the cytarabine crystal form.

The above description is only a preferred embodiment of the present invention and is not intended to limit the present invention, and various modifications and changes may be made by those skilled in the art. Any modification, equivalent replacement, or improvement made within the spirit and principle of the present invention should be included in the protection scope of the present invention.

Claims

1. A cytarabine crystalline substance characterized by an XRD diffraction pattern having characteristic peaks expressed in 2 θ at the following positions:

2. the cytarabine crystalline substance of claim 1, wherein the XRD diffraction pattern of the cytarabine crystalline substance further comprises characteristic peaks expressed in 2 Θ at the following positions:

3. cytarabine lyophilized powder comprising the cytarabine crystals of claim 1 or 2.

4. The cytarabine freeze-drying method is characterized by comprising the steps of sequentially pre-freezing, sublimation drying and analytical drying a cytarabine solution to obtain cytarabine freeze-dried powder, wherein the pre-freezing comprises the following steps of sequentially:

cooling to-40 to-30 ℃ in 30-50 min and preserving heat for 70-105 min; heating to-25 to-15 ℃ for 10-30 min and preserving heat for 70-105 min; reducing the temperature to-40 to-30 ℃ in 10-25 min and preserving the temperature for 100-150 min.

5. Lyophilization process according to claim 4, characterized in that said prefreezing comprises the following steps carried out in sequence:

cooling to-40 to-30 ℃ after 40 to 50min and preserving heat for 80 to 90 min; heating to-20-15 ℃ for 10-20 min and keeping the temperature for 80-100 min; reducing the temperature to-40 to-30 ℃ in 10-25 min and preserving the temperature for 120-130 min.

6. The freeze-drying method according to claim 4, wherein the temperature of the sublimation drying is-35 to 10 ℃, the sublimation drying is preferably carried out by adopting a gradient temperature rise mode, and the process of the sublimation drying preferably comprises the following steps:

under a first vacuum condition, heating the pre-frozen product obtained by pre-freezing to a first sublimation drying temperature for first-stage sublimation drying to obtain a first sublimation dried product;

under a second vacuum condition, heating the first sublimation-dried product to a second sublimation-drying temperature, and carrying out second-stage sublimation drying to obtain a second sublimation-dried product;

wherein the first sublimation drying temperature is-25 to-10 ℃, the second sublimation drying temperature is 0 to 10 ℃, and gauge pressures of the first vacuum condition and the second vacuum condition are respectively and independently 0.1 to 0.5 mbar.

7. The freeze-drying method according to claim 6, wherein the pre-frozen product is heated to the first sublimation drying temperature within 70-100 min and is kept at the first sublimation drying temperature for 270-330 min to perform the first-stage sublimation drying.

8. The lyophilization method according to claim 6, wherein the first sublimation-dried product is heated to the second sublimation-drying temperature within 70 to 100min and is kept at the temperature for 100 to 150min to perform the second sublimation-drying.

9. The lyophilization method according to claim 4, wherein the temperature of the desorption drying is 10 to 50 ℃.

10. The lyophilization process according to claim 9, wherein the desorption drying is performed by a gradient temperature rising method, preferably the desorption drying comprises:

under a third vacuum condition, heating a sublimation dried product obtained by sublimation drying to a first analysis drying temperature to carry out first-stage analysis drying to obtain a first analysis product;

heating the first analytic product to a second analytic drying temperature under a fourth vacuum condition, carrying out second stage analytic drying to obtain cytarabine freeze-dried powder,

wherein the first analytical drying temperature is 10 to 30 ℃, the second analytical drying temperature is 25 to 45 ℃, gauge pressures of the third vacuum condition and the fourth vacuum condition are respectively and independently 0.1 to 0.5mbar,

preferably, the sublimation-dried product is heated to the first analysis drying temperature within 10-30 min, and is kept warm for 100-150 min to perform the first-stage analysis drying;

preferably, the temperature of the first analysis product is raised to the second analysis drying temperature within 10-30 min, and the temperature is maintained for 200-250 min to perform the second stage analysis drying.