WO2023140691A1 - Oral formulation containing 5-aza-4'-thio-2'-deoxycytidine and preparation method therefor - Google Patents

Abstract

The present invention relates to an oral formulation containing 5-aza-4'-thio-2'-deoxycytidine and a preparation method therefor. The present invention is for solving the problem of very great variations among patients in terms of drug efficacy or drug exposure with respect to a cytidine-based anticancer drug such as decitabine.

Description

Oral dosage form containing 5-aza-4'-thio-2'-deoxycytidine and preparation method thereof

The present invention relates to an oral dosage form containing 5-aza-4'-thio-2'-deoxycytidine and a preparation method thereof.

Decitabine (also called Dacogen® or 5-aza-2'-deoxycytidine) is a pyrimidine nucleoside analog of cytidine that inhibits DNA methyltransferase, leading to DNA hypomethylation. Specifically, decitabine functions by incorporating into DNA strands during replication, and when DNA methyltransferases (DNMTs), such as DNMT1, bind DNA and replicate methylation to daughter strands, DNMTs bind irreversibly to decitabine and cannot be separated. Thus, decitabine action is dependent on cell division. Cells must divide for the drug to work. Therefore, cells that divide much faster than most other cells in the body (such as cancer cells) are more affected by decitabine. That is, decitabine is used in the treatment of cancers such as leukemias, including myelodysplastic syndrome (MDS) and acute myelogenous leukemia (AML), in which DNA hypermethylation is important for development.

5-aza-4'-thio-2'-deoxycytidine ("aza-T-dCyd") is a thio-substituted derivative of decitabine that has received early clinical evaluation by the National Cancer Institute (NCI). This DNMT1 inhibitor has recently attracted attention due to its high DNMT elimination and cell inhibitory activity, reduced rate of cytidine deaminase degradation, and relatively low production of toxic by-products compared to existing compounds with a 5-azacytidine backbone. Like decitabine, aza-T-dCyd can be manufactured in a variety of forms and crystal structures.

U.S. Patent No. 5,591,722 relates to 2'-deoxy-4'-thioribonucleosides and intermediates useful for treating viral diseases and describes a general formula comprising 5-azacytidine compounds. US Patent Publication No. 2006/0014949 reports polymorphs of decitabine. Thottassery, et al. (Cancer Chemother Pharmacol, 2014) reports aza-T-dCyd. Clinical trial NCT04167917 reports a phase I trial of Aza-T-dCyd in MDS and AML expected to be completed in 2025. However, the polymorph of aza-T-dCyd has so far remained elusive.

On the other hand, DNA methyltransferase (DNMT) inhibitors based on cytidine analogs such as decitabine and azacitidine have excellent efficacy in the treatment of elderly patients with myelodysplastic syndrome (MDS) and acute myeloid leukemia (AML). However, there are many limitations. For example, many patients do not respond to decitabine or azacytidine. In addition, even in patients who respond to this, the treatment effect is limited due to side effects such as neutropenia and cytopenia. Finally, patient compliance is low because the treatment is commonly used as an injectable drug due to a poor pharmacokinetic (PK) profile.

In addition, anticancer drugs of the cytidine analogue family, such as decitabine or azacytidine, commonly have difficulties in being developed as oral anticancer drugs due to individual differences in absorption and metabolism rates in the human body.

That is, even if the same amount of anticancer drugs of the cytidine analog series is administered, the variation in exposure among individuals based on the average drug exposure is very high, so that the optimal dose for one individual is a dose that cannot show a therapeutic effect for another individual.

Therefore, there is still a need for additional research on formulations capable of enhancing the anticancer effect of Aza-T-dCyd.

Gastrointestinal Tract refers to all organs of the digestive system from the mouth to the anus.

In the small intestine, most nutrients are absorbed into the blood, and fat-soluble nutrients are absorbed into the lymphatic system.

Absorption of a drug means the movement of substances into or across tissues, in particular, the movement of a drug to the wall of the gastrointestinal tract and then to the bloodstream. Absolute bioavailability is the rate at which a drug is systemically utilized after oral, rectal, subcutaneous, transdermal, intranasal, or extravascular administration of the drug.

It is important to maintain an appropriate blood level of the drug in order to exert a therapeutic effect. A dosage form is a means of controlling the action of a drug by adjusting variable factors affecting dissolution and absorption characteristics of a drug, which are drug-derived factors.

When designing an appropriate dosage form in the pharmaceutical manufacturing process, the physical state of the pharmaceutical raw material, that is, whether it is crystalline or amorphous, is very important. In general, the amorphous form has high solubility and is helpful in increasing the efficacy and showing the fast effect, but it is unstable and the shelf life is shortened, and it is also difficult to release the drug and control the blood concentration. However, since the crystalline form has low solubility and low bioavailability per unit weight, stability is secured and there is an advantage in making a continuous controlled release formulation, crystalline raw materials are used in most pharmaceutical formulations except for special cases.

The crystalline form of a drug affects the physical and chemical stability, hygroscopicity, and dissolution rate of a compound in water. Chemical instability causes restrictions on the manufacturing, management, transportation, and shelf life of pharmaceutical raw materials. It also affects the stability after formulation as an oral preparation. If the crystal form is different, the crystal shape, purity, yield, etc. are different, and it greatly affects the formulation process, manufacturing environment, and manufacturing cost.

When the present invention is developed as an oral anticancer agent, the individual drug exposure of the aza-T-dCyd drug varies greatly due to the difference in the degree of absorption for each individual, so that the optimally designed dose for anticancer efficacy may be a dose that cannot show a therapeutic effect for each individual or a dose that causes serious toxicity. do

To this end, an object of the present invention is to provide an oral dosage form in which the ratio (wt%) of crystalline Form A in aza-T-dCyd is adjusted to a known value within an acceptable error range, so as to calculate a single dose that exhibits a desired therapeutic effect from the highest blood concentration (Cmax) of the aza-T-dCyd drug, and to minimize individual differences in absorption during oral administration to stably achieve the highest blood concentration (Cmax) of the aza-T-dCyd drug.

A first aspect of the present invention provides a method for preparing an oral dosage form containing aza-T-dCyd as an active ingredient, the efficacy of which is dependent on the highest blood concentration (Cmax), characterized by precisely designing a single dose that exhibits a desired therapeutic effect from the highest blood concentration (Cmax) of the aza-T-dCyd drug within an acceptable error range.

A second aspect of the present invention provides an oral dosage form in which the ratio (wt%) of crystalline Form A in the aza-T-dCyd drug is controlled, wherein the crystalline raw material containing crystalline Form A in a desired ratio (wt%) is prepared from crude materials of aza-T-dCyd, which is a synthesized product of the aza-T-dCyd compound, and then formulated into an oral dosage form.

At this time, by controlling the dosage of crystalline form A, the exposure amount of the aza-T-dCyd drug in the blood can be stably implemented within an acceptable error range.

A third aspect of the present invention provides a method for preparing an oral dosage form containing aza-T-dCyd as an active ingredient, wherein the ratio (wt%) of crystalline Form A and/or crystalline Form F in the crystalline raw material of aza-T-dCyd is confirmed and then formulated into an oral dosage form.

A fourth aspect of the present invention provides an oral dosage form wherein a single dose of the aza-T-dCyd drug is designed such that the ratio of Form A is greater than or equal to the ratio of Form A corresponding to the inflection point of the Cmax phase at which the maximum blood concentration (Cmax) change value according to the change in the ratio of Form A in the same dose of the aza-T-dCyd drug is increased.

A fifth aspect of the present invention provides an oral dosage form containing at least 70% of Form A in the aza-T-dCyd drug.

A sixth aspect of the present invention provides an oral dosage form characterized by containing 30 mpk to 70 mpk of crystalline Form A when designing a single dose of the aza-T-dCyd drug.

A seventh aspect of the present invention is a method for preparing an oral dosage form containing an aza-T-dCyd drug whose efficacy is dependent on the maximum blood concentration (Cmax), comprising the steps of crystallizing the aza-T-dCyd compound in the presence of a solvent and then removing the solvent to convert it into a non-solvate crystalline form; and a second step of preparing an oral dosage form designed so that the non-solvate crystalline form prepared in the first step can be dissolved in the stomach.

Hereinafter, the present invention will be described.

As used herein, a drug is any substance used to diagnose, cure, alleviate, treat, or prevent disease (excluding food or devices) or to affect the structure or function of the body. For example, any chemical or biological substance that affects the body and its metabolism. The chemical name of a drug indicates the drug's atomic or molecular structure.

In the present specification, the aza-T-dCyd drug includes a compound represented by Formula 1 below as well as pharmaceutically acceptable salts thereof. "Pharmaceutically acceptable salts" include non-toxic acid and base addition salts of the compounds to which the term refers.

As used herein, "solvate" refers to a compound provided herein or a salt thereof that further comprises a stoichiometric or non-stoichiometric amount of a solvent bound by non-covalent intermolecular forces. When the solvent is water, the solvate is a hydrate.

aza-T-dCyd of Chemical Formula 1 is a DNMT1 inhibitor based on a 4-Thio-2-deoxyribose backbone, and has both a sugar structure change (4'-thiodeoxyribose structure) and an aza-cytosine group.

[Formula 1]

aza-T-dCyd is activated by triphosphate in cells and used instead of some dC (deoxycytidine) during DNA synthesis. It is a nucleoside anticancer drug that induces cancer cell death by trapping DNMT1 after DNA synthesis and activating various epigenetic action mechanisms.

Unlike Decitabine/Azacytidine, aza-T-dCyd, due to its Thio-nucleoside structure, can exhibit strong anticancer efficacy by simultaneously blocking endonuclease that participates in DNA damage repair, an existing mechanism of resistance development, along with strong inhibition of DNMT1, the main drug target.

Aza-T-dCyd compounds can be rapidly activated by triphosphate in cancer cells compared to normal cells. The Aza-T-dCyd compound delays DNA replication by causing base excision repair and/or mismatch repair through DNA insertion of aza-T-dCTP, a triphosphate of the aza-T-dCyd compound, thereby inducing replication stress and increasing DNA damage response.

The Aza-T-dCyd compound suppresses the expression of RRM1 protein, a ribonucleotide reductase that is important for dNTP de novo synthesis, induces DNA replication stress by reducing the amount of dCTP and dTTP in cells, and can generate a strong DNA damage response.

In the case of Decitabine/Azacytidine, it is vulnerable to metabolism by Cytidine Deaminase present in our body, and its PK profile is unstable, which limits its effectiveness. Surprisingly, compared to Decitabine/Azacytidine, it was found that aza-T-dCTP was degraded by cytidine deaminase at a slower rate when administered orally, thus overcoming metabolic resistance by cytidine deaminase in cancer cells as well as having an excellent PK profile even when administered orally.

In addition, aza-T-dCyd significantly lowers the activation rate by dCK (deoxycytidine kinase) in normal cells due to its Thio-nucleoside structure, thereby selectively delivering the active ingredient of the drug to cancer cells. As a result, it is possible to secure an excellent safety profile and secure a wide therapeutic window, so it can be administered at a high dose that prevents the mechanism of resistance development from operating.

Furthermore, through the pharmacokinetic analysis of Example 6, (1) the anti-cancer treatment effect of aza-T-dCyd is Cmax dependent rather than AUC dependent; and (2) exposure to higher doses of the aza-T-dCyd drug for a short period of time is an effective anti-cancer therapy.

In addition, (3) through the dissolution rate profile analysis at various pH points in Example 7, Form A and Form F show similar dissolution rates at pH 1.2 (pH conditions of the stomach and large intestine), but Form A shows a consistent dissolution rate profile compared to Form F, and Form F has a higher dissolution rate than Form A at pH 6.5 and pH 5 (pH conditions of the appendix and small intestine) (Table 6 and FIGS. 20 to 22); (4) In Example 12, it was found that the aza-T-dCyd drug was not absorbed in the caecum through the analysis of the absorption profile in the gastrointestinal tract.

The present invention was completed based on these findings.

In order to solve the problem that the individual drug exposure variation of the aza-T-dCyd drug is very high due to the difference in the degree of absorption between individuals, the optimally designed dose for anticancer efficacy may be a dose that cannot show a therapeutic effect or a dose that causes serious toxicity depending on the individual. To provide a design technology for an oral dosage form containing aza-T-dCyd drug that is precisely controlled in

For example, in an oral dosage form targeting release of the active ingredient of the drug at about pH 1.2 (above), the proportion (wt%) of Form A in the drug aza-T-dCyd can be precisely controlled.

Therefore, according to one embodiment of the present invention, a single dose that exhibits a desired therapeutic effect is calculated from the highest blood concentration (Cmax) of the aza-T-dCyd drug, and the ratio (wt%) of Form A in the aza-T-dCyd drug is adjusted to a known value within an acceptable error range to stably exert the highest blood concentration (Cmax) of the aza-T-dCyd drug by minimizing individual differences in absorption during oral administration. can design

The method for preparing an oral dosage form containing aza-T-dCyd as an active ingredient according to the present invention is characterized by precisely designing a single dose that exhibits a desired therapeutic effect from the highest blood concentration (Cmax) of the aza-T-dCyd drug within an acceptable error range.

In the present specification, "precisely designing a single dose that exhibits a desired therapeutic effect from the highest blood concentration (Cmax) of the aza-T-dCyd drug within an acceptable error range" means, for example, designing a dosage form capable of controlling a constant dissolution rate profile and absorption of the drug within a predictable error range to minimize the variation in drug exposure, and based on this, a single administration of the aza-T-dCyd drug that achieves the highest blood concentration (Cmax) that exhibits the desired therapeutic effect It could be capacity design.

The pharmacokinetic parameters for evaluation of bioavailability are the amount of drug that the active ingredient or its active metabolites reach the systemic circulation from the preparation and the time it takes for them to reach the systemic circulation.

The area under the plasma level-time curve (AUC) refers to the degree of drug exposure, that is, the degree of bioabsorption of the drug, and reflects the total amount of active drug that reaches the systemic circulation. The unit of AUC is expressed as concentration·time (eg, μg hr/mL). AUC is a measure of the degree of drug bioavailability. It represents the extent of total systemic exposure.

Cmax is the highest blood concentration after drug administration and is an indicator indicating whether the drug is sufficiently absorbed into the systemic circulation to show a therapeutic response, as well as providing information on whether or not it can cause toxic effects.

t _max is the time at which the blood concentration reaches the highest value after drug administration, and refers to the moment at which the absorption rate of the drug and the excretion rate become the same as the time point at which the drug absorption reaches the highest value. After t _max , drug absorption continues but slows down. Therefore, when comparing the absorption of drugs, it is an index for the rate of absorption.

IC ₅₀ values are related to AUC. Usually, the dose can be calculated from AUC and bioavailability.

However, with respect to the blood drug concentration, surprisingly, even if the aza-T-dCyd drug had the same AUC, the efficacy of aza-T-dCyd decreased significantly as the exposure time to the cells increased, and rather, the size of the tumor increased rapidly (Example 6). This suggests that exposure to a higher dose of aza-T-dCyd drug for a short period of time can provide efficient anticancer treatment.

Therefore, in order to maximize the efficacy of the aza-T-dCyd drug that is Cmax dependent rather than AUC, the present invention is characterized by designing a single dose of the aza-T-dCyd drug so as to exert a desired therapeutic effect from the highest blood concentration (Cmax) of the aza-T-dCyd drug.

For example, in the present invention, a single dose of the aza-T-dCyd drug may be 5-70 mg/m ² . For non-cancer use, it may be 5-10 mg/m ² . For anticancer use, it may be preferably 5 to 55 mg/m ² , more preferably about 5 to 30 mg/m ² , and still more preferably 5 to 20 mg/m ² . For example, in combination therapy, it may be 5 to 10 mg/m ² .

From the toxicity profile of the non-clinical animal model, the non-toxic amount (No-observed-adverse-effect level, NOAEL) or the highest non-severe toxic dose (HNSTD) that does not cause serious toxicity can be confirmed. NOAEL refers to the maximum amount at which no effect appears on animals even if the test drug is continuously supplied during the toxicity test period (usually 1-2 years).

Therefore, when calculating the appropriate dosage for humans, it can be adjusted to be less than the no-observed-adverse-effect level (NOAEL) or the highest non-severe toxic dose (HNSTD) that does not cause serious toxicity, and the dose of the anticancer agent can be compared with the non-toxic amount (NOAEL) or the highest dose that does not cause serious toxicity (HNSTD), and the risk factor (risk) during administration can be predicted.

The peak blood concentration (Cmax) after administration of a drug is not only an indicator of whether a drug has been sufficiently absorbed into the systemic circulation to produce a therapeutic response, but also provides information on the toxicity profile.

Characterization of the pharmacokinetics (PK) of aza-T-dC compound in mice after intravenous (IV) and oral (PO) dosing revealed that the optimally efficacious dose of aza-T-dC compound when administered orally was 2.0 mpk (mpk = milligram per kilogram). In addition, the GLP Tox experiment was performed on dogs, and the non-toxic amount (NOAEL) was 0.15mpk (< 3mg/m ² /day), and the highest dose (HNSTD) that did not cause serious toxicity was 0.5mpk (10mg/m ² /day).

In Example 11, when comparing the results of 2mpk treatment and 1mpk treatment of mice twice, in consideration of the fact that the weight loss of mice was severe in the latter, the present invention provides that the highest blood concentration (Cmax) of the aza-T-dCyd drug calculated from a single dose of the aza-T-dCyd drug is a non-toxic level (No-observed-adverse-effect level, NOAEL) or the highest that does not cause serious toxicity. A single dose of the aza-T-dCyd drug can be designed to be lower than the value corresponding to the highest non-severe toxic dose (HNSTD). In addition, since the present invention can design a single dose of the aza-T-dCyd drug to be lower than the value corresponding to the non-toxic level (NOAEL) or the highest dose that does not cause serious toxicity (HNSTD) based on Cmax rather than AUC, it is possible to expand the range of a very narrow therapeutic window of the aza-T-dCyd drug.

For example, a database comparing in vitro data IC ₅₀ , IC ₆₀ , IC ₇₀ , IC ₈₀ and IC ₉₀ with side effect values (IC ₅₀ based on NOAEL and/or HNSTD) in a non-clinical large animal model can be constructed. In addition, a single dose of the aza-T-dCyd drug is calculated from this, and then the maximum blood concentration (Cmax) of the aza-T-dCyd drug calculated therefrom is compared with the Cmax value corresponding to the non-toxic amount (NOAEL) or the highest dose that does not cause serious toxicity (HNSTD), and various therapeutic efficacy and/or side effect prediction information derived from a single dose of the aza-T-dCyd drug in a specific oral dosage form can be provided. .

On the other hand, due to the presence of cytidine deaminase (CDA) in the blood and the difference in concentration from person to person, the problem due to the individual metabolic rate difference in cytidine analog-based anticancer drugs is that aza-T-dCyd has a low rate of degradation of cytidine deaminase compared to other cytidine analog-based anticancer drugs. Not only is the effect not significant, but the maximum blood concentration (Cmax) of the desired aza-T-dCyd drug can be maintained for a longer time there is

In addition, the present invention is characterized in that the ratio (wt%) of crystalline form A in the aza-T-dCyd drug is closely controlled to a known value within an error range, thereby solving problems caused by individual differences in absorption.

This is based on the discovery that a desired Cmax value of an aza-T-dCyd drug can be achieved in a state that is less dependent on various factors by controlling the dosage (wt%) of the highly bioavailable crystalline Form A in the aza-T-dCyd polymorph or the ratio (wt%) of the crystalline Form A in the aza-T-dCyd before formulation into an oral dosage form. Accordingly, the present invention can provide an oral dosage form containing an aza-T-dCyd drug in which absorption of the drug and control of the maximum blood concentration (Cmax) are precisely controlled within an acceptable error range.

aza-T-dCyd is an analog of decitabine represented by Formula 1 and, like decitabine, can be prepared in various forms and crystal structures.

[Formula 1]

In the present specification, polymorphs of aza-T-dCyd include (pseudo-)polymorphs of aza-T-dCyd.

Polymorphism is a phenomenon in which one compound has more than one molecular arrangement structure, that is, a crystal structure, in a solid state, and has chemically identical but physically different characteristics. Here, the crystal structure means the internal structure of the crystal. The most important characteristics of a solid are a specific distance between molecules and a specific bonding force, and polymorphs have different melting points because of the different distance and bonding strength between molecules, and therefore have different solubility.

Since polymorphs have different physical properties, solubility and dissolution rate are different, and as a result, there is a difference in bioavailability, resulting in a problem of bioequivalence.

When a sample is measured with an X-ray powder diffractometer, X-rays are diffracted by the crystal planes, and the distance of the peaks from the 2θ angle of the crystal planes can be known. A direct proportional relationship is established between the intensity of the diffraction peaks and the concentration of the sample.

Pseudo-polymorphism is chemically different and physically different. This is mainly in the case of solvates, in which solvent molecules enter the crystal lattice to form crystals. When the solvent is water, it is called a hydrate.

For example, the synthesized product of the aza-T-dCyd compound mainly contains crystalline forms A and F, and may include various other crystalline forms (Example 1).

In one embodiment of the present invention, crystalline form A, crystalline form F, or a combination (mixed) form of crystalline form A and crystalline form F of aza-T-dCyd is disclosed.

Crystalline Form A of aza-T-dCyd can be defined as having peaks at 2θ diffraction angles of about 8º, about 13º, about 15º, about 17º, about 19º, about 22º, about 23º, about 26º, about 28º, about 29º, about 31º, about 33º, and about 37º in a powder X-ray diffraction spectrum.

For example, crystalline form A of aza-T-dCyd has 2θ diffraction angles of 7.7°±0.3°, 13.02°±0.3°, 15.34°±0.3°, 16.78°±0.3°, 18.62°±0.3°, 19.42°±0.3°, 21.94°±0.3°, and ±22.90°. .3°, 25.70°±0.3°, 26.64°±0.3°, 27.86°±0.3°, 28.63°±0.3°, 29.45°±0.3°, 31.42°±0.3°, 32.70°±0.3°, 34.72 ±0.3, 35.97°±0.3° and 37. It may have a peak of 46 ° ± 0.3 °. For example, it may have peaks at 7.7º, 13.02º, 15.34º, 16.78º, 18.62º, 19.42º, 21.94º, 22.90º, 25.70º, 27.86º, 28.70º, 31.42º, 32.70º, and 37.46º. there is

Crystalline form F of aza-T-dCyd has diffraction angles in 2θ of about 6º, about 12º, about 13º, about 14º, about 16º, about 18º, about 20º, about 21º, about 22º, about 26º, about 27º, about 29º, about 30º, about 33º, about 35º, about 36º in the powder X-ray diffraction spectrum. º, about 39º, and about 41º.

For example, crystalline form F of aza-T-dCyd has 2θ diffraction angles of 6.06º ± 0.3°, 12.10º ± 0.3°, 13.02º ± 0.3°, 14.38º ± 0.3°, 15.94º ± 0.3°, 17.50º ± 0.3°, 19.62º ± 0.3°, 21.18º ± 0.3°, 22.34º ± 0.3°, 26.18º ± 0.3°, 27.42º ± 0.3°, 28.50º ± 0.3°, 29.90º ± 0.3°, 32.66º ± 0.3°, 35.02º ± 0.3°, 36.30º ± 0.3 °, 38.94º ± 0.3 °, and 41.06º ± 0.3 °. For example, 6.06º, 12.10º, 13.02º, 14.38º, 15.94º, 17.50º, 19.62º, 21.18º, 22.34º, 26.18º, 27.42º, 28.50º, 29.90º, 32.66º, 35.02º, It may have peaks at 36.30º, 38.94º, and 41.06º.

A method for preparing an oral dosage form containing an aza-T-dCyd drug according to the present invention,

A first step of crystallizing the aza-T-dCyd compound in the presence of a solvent and then converting the aza-T-dCyd compound into a non-solvate crystalline form by removing the solvent; and

The second step of preparing an oral dosage form designed so that the non-solvate crystalline form prepared in the first step can be dissolved in the stomach.

includes

The first step may be to convert the crystalline polymorph containing the solvate to a single crystalline Form A by removing the solvent. Thus, an aza-T-dCyd drug consisting only of crystalline Form A can be provided.

The second step is to provide an oral dosage form containing an aza-T-dCyd drug that is precisely designed within an acceptable error range for a single dose that exhibits a desired therapeutic effect from the highest blood concentration (Cmax) of the aza-T-dCyd drug.

In the case of the amorphous form, it dissolves quickly and shows a quick effect, and the duration is short. In the case of the crystalline form, it dissolves slowly, so the effect appears slowly but the duration is long.

Therefore, the physicochemical properties (physical properties) of the aza-T-dCyd drug directly affect formulation development. Since polymorphs differ in solubility and stability, polymorphs are very important pharmaceutically, and in particular, crystalline forms that are well soluble are preferred in terms of bioavailability. However, simply selecting a form that dissolves well does not end all problems, and it is necessary to study whether or not the form is converted. Therefore, the present invention was completed through polymorph screen and characterization of aza-T-dCyd in Examples 4 and 5.

Through experiments, the present inventors found that crystalline form A and crystalline form F are stable anhydride forms in physical and chemical terms. Therefore, constructing the aza-T-dCyd drug raw material in crystalline form A and/or crystalline form F with high physicochemical stability not only has advantages such as storage stability and purity control during formulation development, but more importantly, the crystalline form can affect the biological activity of the drug.

In addition, the dosage of the aza-T-dCyd drug may vary within a range depending on the dosage form (crystal form) and route of administration.

For example, an oral dosage form containing aza-T-dCyd may be designed to dissolve 90% or more of Form A and/or Form F in the stomach. In addition, the maximum blood concentration (Cmax) and/or the area under the blood drug concentration-time curve (AUC) of the aza-T-dCyd drug may be adjusted by adjusting the composition ratio of Form A and/or Form F of the aza-T-dCyd drug.

Through experiments, it was found that Form A exhibits a high dissolution rate/solubility in strong acidic conditions and exhibits a consistent dissolution rate profile compared to Form F. The present inventors prepared various crystalline forms of aza-T-dCyd and confirmed that the anticancer effect in crystalline form A was the most excellent. Therefore, it is a feature of the present invention to apply this to the design of an oral dosage form containing an Aza-T-dCyd drug whose efficacy is dependent on Cmax.

Surprisingly, it has been found that the desired Cmax value of the aza-T-dCyd drug can be stably achieved within an acceptable error range by adjusting the dose of Form A and/or the ratio of the aza-T-dCyd drug, and by applying this, the aza-T-dCyd drug can be formulated to exert a desired therapeutic effect by minimizing individual variation.

That is, in view of the fact that the conventional aza-T-dCyd compound exhibits drug efficacy more stably by having a specific crystalline form than when it is in a mixed form of several crystalline forms, by designing an oral dosage form with a certain ratio or more of crystalline form A, for example, 70% or more of the Aza-T-dCyd drug, high Cmax efficacy could be realized at the same single dose of the aza-T-dCyd drug (Example 10).

For example, when a crystalline drug having low solubility compared to an amorphous drug is formulated into an oral dosage form, (partially) amorphous and highly water-soluble additives may be added and mixed thereto to improve solubility and increase bioavailability. Non-limiting examples of the additive include starch, lactose, PVP, and microcrystalline cellulose.

The oral dosage form of the present invention may contain a disintegrating carrier so that at least 80% of the aza-T-dCyd drug is dissolved in an acidic stomach.

The amorphous form has high solubility, which helps to increase the efficacy of the drug and show rapid action, but it is difficult to release the drug and control the blood concentration; no absorption of the aza-T-dCyd drug from the cecum (Example 12); Form A and Form F have high dissolution rate/solubility and similar dissolution rates at pH 5 (pH condition of the small intestine) and pH 1.2 (pH condition of the stomach). However, taking advantage of the fact that Form A shows a consistent dissolution rate profile compared to Form F, the present invention provides (1) a single dose of aza-T-dCyd drug and (2) aza-T-d by controlling the ratio of Form A to aza-T-dCyd drug showing a consistent dissolution profile in the stomach, Despite the very narrow therapeutic window of Cyd drugs, high Cmax can be easily achieved within an acceptable margin of error.

In addition, taking advantage of the fact that dissolution is a rate-limiting step in absorption of solid dosage forms, absorption of most drugs occurs in the small intestine, and crystalline form A has relatively high solubility and dissolution rate at the pH of the stomach, the present invention designs an oral dosage form that is decomposed by gastric acid so that, for example, 90% or more of crystalline form A dissolves in the stomach, thereby absorbing most of the aza-T-dCyd drug at the beginning of the small intestine, thereby exhibiting the desired therapeutic effect with a lower single dose than other crystalline forms. A high Cmax can be precisely controlled with significantly reduced potential for toxic side effects.

Furthermore, the present invention provides a single dose of the aza-T-dCyd drug and a crystalline form of the aza-T-dCyd drug to precisely control (i) the highest blood concentration (Cmax) of the drug Aza-T-dCyd that exerts the desired therapeutic effect, optionally (ii) the amount of the drug aza-T-dCyd that reaches the systemic circulation from the oral dosage form, and optionally (iii) the time it takes for the drug drug aza-T-dCyd to reach the systemic circulation. It is characterized by designing the ratio of A.

The ratio of crystalline form A in the aza-T-dCyd drug may be wt% or mole%, but is not limited thereto.

The present invention can design a single dose of aza-T-dCyd drug and the ratio of crystalline form A in aza-T-dCyd drug in order to easily control the maximum blood concentration (Cmax) after drug administration, which indicates whether the drug is sufficiently absorbed into the systemic circulation to produce a therapeutic response.

In addition, by adjusting the dosage of the aza-T-dCyd drug and the ratio (wt%) of the crystalline form A in the aza-T-dCyd drug, that is, the amount of the drug reaching the systemic circulation from the oral dosage form and the time taken to reach the systemic circulation can be easily controlled by adjusting the dosage of the aza-T-dCyd drug and the dosage of the crystalline form A.

In the case of crystalline Form A, by designing an oral dosage form that is degraded by gastric acid so that, for example, 90% or more of Form A is dissolved in the stomach, focusing on the fact that the dissolution rate is the highest in the stomach compared to the small intestine and is not absorbed in the caecum, the present invention can easily control the amount of the drug reaching the systemic circulation from the oral formulation and the time taken to reach the systemic circulation by adjusting the dose of Form A or the ratio (wt%) of the aza-T-dCyd drug.

For example, in order to control the ratio (wt%) of crystalline form A in the aza-T-dCyd drug in the oral dosage form of the present invention, a crystalline raw material containing crystalline form A in a desired ratio (wt%) can be prepared from aza-T-dCyd crude materials, which are synthesized products of the aza-T-dCyd drug, and then formulated into an oral dosage form.

Accordingly, the present invention provides an oral dosage form in which the ratio (wt%) of crystalline Form A in the aza-T-dCyd drug is controlled, wherein the crystalline raw material containing Form A in a desired ratio (wt%) is prepared from crude materials of aza-T-dCyd, which is a synthetic product of the aza-T-dCyd drug, and then formulated into an oral dosage form.

In the present specification, preparing a crystalline raw material containing crystalline form A in a desired ratio (wt%) from aza-T-dCyd crude materials, which are the synthetic product of the aza-T-dCyd drug, can be a process of controlling the desired ratio (wt%) of crystalline form A within a ±10% error range, preferably within a ±5% error range, and the desired ratio (wt%) of crystalline form A is within a ±10% error range, preferably ±5% error range. It may be a verification process.

In addition, the oral dosage form of the present invention is characterized in that a single dosage of the aza-T-dCyd drug is designed so that the dosage of the aza-T-dCyd drug contains more than the ratio of Form A corresponding to the inflection point of the Cmax phase at which the maximum blood concentration (Cmax) change value increases according to the change in the ratio of Form A in the same single dose of the aza-T-dCyd drug.

The Cmax inflection point considered in designing the ratio of Form A in the drug Aza-T-dCyd may appear when the ratio of Form A in the drug Aza-T-dCyd is 50% to 80%.

As illustrated in FIG. 25 , among the drugs of aza-T-dCyd, crystalline form A exhibited a high Cmax beyond prediction at 70% or more. Therefore, based on the PK data illustrated in FIG. 25 , the ratio of crystalline Form A in the aza-T-dCyd drug may be 70% or more.

For example, as illustrated in FIG. 25 , if the inflection point is 70% in terms of the ratio of Form A in the aza-T-dCyd drug, the oral dosage form can be designed such that the ratio of Form A in the aza-T-dCyd drug is 70% or more. Preferably, the ratio of crystalline Form A in the aza-T-dCyd drug in the oral dosage form may be 100%. In this case, as suggested in Examples 6 and 11, the bioavailability of the aza-T-dCyd drug whose therapeutic effect/toxic side effect depends on Cmax can be precisely controlled during oral administration. At this time, in order to set the ratio of crystalline form A in the aza-T-dCyd drug in the oral dosage form to 100%, after synthesizing the aza-T-dCyd compound and converting all of the aza-T-dCyd to crystalline form A, the oral dosage form may be formulated.

As used herein, “cancer” refers to or describes a physiological condition in mammals that is typically characterized by unregulated cell growth. Examples of cancers include, but are not limited to, blood-borne tumors (eg, multiple myeloma, lymphoma, and leukemia) and solid tumors. Non-limiting examples of hematological cancers include non-Hodgkin's lymphoma, Hodgkin's lymphoma, multiple myeloma, leukemia, lymphoma, myelodysplastic syndrome, acute granulocytic leukemia, acute myelogenous leukemia, chronic myeloid leukemia, etc. Non-limiting examples of solid cancers include gastric cancer, kidney cancer, ovarian cancer, cervical cancer, uterine cancer, prostate cancer, lung cancer, colon cancer, breast cancer, melanoma, and pancreatic cancer.

As used herein, “patient” and “subject” refer to animals such as mammals. In certain embodiments, the patient is a human. In other embodiments, the patient is a non-human animal, such as a dog, cat, livestock (eg, horse, pig or donkey), chimpanzee or monkey.

In the present specification, the anticancer effect or therapeutic effect of an anticancer agent may refer to an action that reduces the severity of cancer, reduces the size of a tumor, or delays or slows down the progression of cancer, which occurs while a patient is suffering from a specific cancer.

For example, the anticancer effect of an anticancer agent may be Cell Viability (a change in the degree of cytotoxicity or the number of cells) of cancer cells after treatment with the anticancer agent in vitro and/or in vivo. For example, it can be confirmed indirectly through a drug response test through a cell line or a non-clinical animal model (xenograft). In addition, even in cancer patients, the anticancer effect of the anticancer agent can be directly confirmed, and related data can be derived and used as a database. In addition, when designing an anticancer drug dosage guideline, animal model PK parameters and/or toxicity profile may be considered in parallel.

The anticancer effect of an anticancer agent may be inferred from in-vitro data, such as the % maximum effect of the anticancer agent, such as IC ₅₀ , IC ₆₀ , IC ₇₀ , IC ₈₀ and IC ₉₀ , and the highest blood concentration of the drug (Cmax) and / or blood drug concentration - It can also be confirmed in non-clinical animal models and clinical cancer patients through in-vivo data such as area under the time curve (AUC). .

Reactivity of an anticancer agent means clinical sensitivity in terms of anticancer effect.

"Sensitivity" and "susceptibility" when referring to treatment with an anti-cancer agent are relative terms that refer to the degree of effectiveness of a compound in alleviating or reducing the progression of the tumor or disease being treated.

An "effective patient's anticancer effect/response" can be, for example, a 5%, 10%, 15%, 20%, 25%, 30%, 40%, 50%, 60%, 70%, 80%, 90%, 100%, 200%, or more inhibition of a patient's response, as measured by any suitable means, such as gene expression, cell counts, assays, and the like.

In the present specification, the dose is the dose at which drug efficacy is expected. In the present invention, the medicinal effect may be an anticancer effect. The reactivity (anti-cancer effect) of an anti-cancer agent is the degree of response, and may be the % maximum effect of the anti-cancer agent, such as IC ₅₀ , IC ₆₀ , IC ₇₀ , IC ₈₀ and IC ₉₀ , and a value that exhibits toxicity to normal cells (LC ₅₀ ).

For example, dosage forms for oral use can be formulated using a variety of formulation techniques known in the art. For example, it may include a biodegradable (hydrolyzable) polymeric carrier used to adhere to the oral mucosa. It is designed to slowly erode over a predetermined period of time, wherein drug delivery is provided essentially entirely.

Drug delivery in an oral dosage form avoids the weaknesses encountered with oral drug administration, such as slow absorption, degradation of the active agent by fluid present in the gastrointestinal tract and/or first pass and inactivation in the liver, as recognized by those skilled in the art. For biodegradable (hydrolysable) polymeric carriers, it will be appreciated that virtually any such carrier may be used provided that the desired drug release profile is not compromised, and that the carrier is compatible with aza-T-dCyd and any other ingredient present in an oral dosage unit. Generally, polymeric carriers include hydrophilic (water-soluble and water-swellable) polymers that adhere to the wet surface of the oral mucosa. Examples of polymeric carriers useful herein include acrylic acid polymers and co, eg, those known as “carbomers” (Carbopol ® (available from B.F. Goodrich) is one such polymer). In some embodiments, non-limiting examples of other ingredients that can be incorporated into an oral dosage form include disintegrants, diluents, binders, lubricants, flavoring agents, coloring agents, preservatives, and the like. In some embodiments, it may be in the form of a conventionally formulated tablet, lozenge, or gel for buccal or sublingual administration.

In some embodiments, administration of the compound is continued at the physician's discretion when the patient's condition improves; Alternatively, the dose of drug to be administered may be temporarily reduced or temporarily discontinued for some length of time (ie, a "holiday"). The length of the washout can vary between 2 days and 1 year, by way of example only, 2 days, 3 days, 4 days, 5 days, 6 days, 7 days, 10 days, 12 days, 15 days, 20 days, 28 days, 35 days, 50 days, 70 days, 100 days, 120 days, 150 days, 180 days, 200 days, 250 days. , 280 days, 300 days, 320 days, 350 days, or 365 days. In some embodiments, the dose reduction during the washout is 10%-100%, by way of example only, 10%, 15%, 20%, 25%, 30%, 35%, 40%, 45%, 50%, 55%, 60%, 65%, 70%, 75%, 80%, 85%, 90%, 95%, or 100 contains %.

When improvement of the patient's condition occurs, a maintenance dose is administered, if necessary. Subsequently, the dosage or frequency of administration, or both, can be reduced as a function of symptoms, to a level at which improved disease, disorder or condition is maintained. However, patients require intermittent treatment over a long period of time upon any recurrence of symptoms.

The amount of a given agent that will correspond to such amount will vary with factors of the subject or host in need of treatment such as the particular compound, severity of the disease, identity (e.g., body weight), but can nevertheless be routinely determined in a manner known in the art, depending on, for example, the particular agent to be administered, the route of administration, and the particulars surrounding the case involving the subject or host to be treated. In general, however, doses used for adult human treatment will typically range from 0.02-5000 mg/day, or about 1-1500 mg/day.

A single dose herein may be given as a single dose or in divided doses administered simultaneously, for example as 2, 3, 4 or more sub-doses.

In some embodiments, oral formulations are unit dosage forms suitable for single administration of precise dosages. In unit dosage form, the formulation is divided into unit doses containing appropriate amounts of one or more compounds. In some embodiments, the unit dose is in the form of patches containing discrete amounts of the formulation. Non-limiting examples are packaged tablets or capsules, and powders in vials or ampoules. Aqueous suspension compositions can be packaged in single-dose non-reclosable containers. Alternatively, multi-dose reclosable containers may be used, in which case it is typical to include a preservative in the composition.

By way of example only, in some embodiments, formulations for parenteral injection are presented in unit dosage form, including but not limited to ampoules, or in multi-dose containers, with an added preservative.

In order for a drug to act effectively in vivo, the concentration of the drug in the body must be maintained within a therapeutic range for a certain period of time or longer. When a drug is present in excess in the body, it exhibits toxicity, and when the amount is too small, the therapeutic effect does not appear. Therefore, the bioavailability of the aza-T-dCyd drug can be controlled by adjusting the polymorphism of the aza-T-dCyd drug at the crystalline level. In addition, the present invention uses the polymorphism of the aza-T-dCyd crystalline form to control the release and absorption of the aza-T-dCyd drug, thereby reducing the side effects of the aza-T-dCyd drug that is Cmax dependent rather than the AUC and maximizing the efficacy of the aza-T-dCyd drug.

Furthermore, the present invention can provide an oral dosage form in which the ratio (wt%) of crystalline Form A among polymorphs of aza-T-dCyd drug is adjusted in order to achieve the highest blood concentration (Cmax) within a single dose of the aza-T-dCyd drug, which is related to toxicity in the body, which is a side effect, based on a single dose. In addition, by adjusting the dosage of the aza-T-dCyd drug and the ratio (wt%) of crystalline Form A in the Aza-T-dCyd drug, the amount of the drug reaching the systemic circulation from the oral formulation and the time it takes to reach the systemic circulation can be precisely and easily controlled.

Figure 1 shows representative HT-XRPD and HR-XRPD patterns for aza-T-dCyd starting material (SM: aza-T-dCyd for which no specific crystallization conditions have yet been applied).

Figure 2 shows representative simulated XRPD and HR-XRPD of aza-T-dCyd crystalline Form A.

Figure 3 shows a representative TGMS analysis of aza-T-dCyd starting material (SM).

Figure 4 shows a representative DSC trace of aza-T-dCyd starting material (SM).

5 shows representative simulated XRPD and HT-XRPD of aza-T-dCyd Form A after second cycling DSC.

Figure 6 shows the cycle DSC of aza-T-dCyd starting material (SM).

7A and 7B show representative results of LCMS of aza-T-dCyd starting material (SM). Specifically, FIG. 7A shows a representative LC chromatogram of aza-T-dCyd starting material (SM). 7B shows a representative MS spectrum of aza-T-dCyd from liquid chromatography.

Figures 8A-C show representative results of LCMS of aza-T-dCyd starting material (SM) after forming a solution in water. Specifically, FIG. 8A shows an LC chromatogram of aza-T-dCyd formulated in water. Figure 8b shows the MS spectrum of the impurity eluted at 3.8 minutes. Figure 8c shows the MS spectrum of aza-T-dCyd eluted at 4.4 min.

Figure 9 presents representative data showing the chemical stability of aza-T-dCyd in various solutions.

10 shows representative data showing the chemical stability of aza-T-dCyd in various solutions over time.

11 shows a representative XRPD pattern of Form A of aza-T-dCyd.

12 shows representative XRPD patterns of Forms A, B, A+C1, A+C2, A+D1, and A+D2 of aza-T-dCyd and aza-T-dCyd starting material (SM).

13A-C show representative chemical analyzes of Form A. 13A shows the TGMS analysis of Form A. 13B shows the DSC analysis of Form A. 13C shows the LCMS analysis of Form A.

14 shows representative XRPD patterns of crystalline Forms E, F, G1, G2, H, I, J, F+K and L of aza-T-dCyd and aza-T-dCyd starting material (SM).

15A-C show representative chemical analyzes of Form F. 15A shows the TGMS analysis of Form F. 15B shows the DSC analysis of Form F. 15C shows the LCMS analysis of Form F.

16 shows a representative XRPD pattern of Form F of aza-T-dCyd.

17 and 18 present in vivo luciferase activity data showing tumor size when aza-T-dCyd starting material (SM) was administered to female NOD-SCID mice.

Fig. 19 shows the half-maximal inhibitory concentration (IC50) when hematological malignant cells (Mv4-11) were treated with aza-T-dCyd starting material (SM).

20 shows the dissolution rate profiles of Form A and Form F at pH 1.2.

21 shows the dissolution rate profiles of Form A and Form F at pH 5.0.

22 shows the dissolution rate profiles of Form A and Form F at pH 6.5.

Figure 23 shows IC ₅₀ values when the K562 cell line was treated with crystalline Form A or SM.

Figure 24 shows the IC ₅₀ values when the HL-60 cell line was treated with Form A or SM.

25 shows PK analysis results obtained by blood sampling at 0.25, 0.5, 1, 2, 4, 6, 8, and 24 hr after oral administration by adjusting the ratio (wt%) of crystalline form A in Aza-T-dCyd drug.

26A and B show dissolution rate profiles of PO and IC administration of 1mpk and 3mpk of aza-T-dCyd compound, respectively.

Hereinafter, the present invention will be described in more detail through examples. However, the following examples are only intended to clearly illustrate the technical features of the present invention, but do not limit the protection scope of the present invention.

In order to design the formulation of the drug Aza-T-dCyd and to evaluate the bioavailability of the polymorphism of the crystalline form of the drug Aza-T-dCyd, pharmacokinetic parameters were measured in in vitro tests and animal models through the following examples.

Example 1: Characterization of starting materials

Approximately 4.0 g of aza-T-dCyd was prepared and analyzed by X-ray powder diffraction (XRPD), differential scanning calorimetry (DSC), thermogravimetry-mass spectrometry (TGMS), and liquid chromatography/mass spectrometry (LCMS). The starting material (SM) is aza-T-dCyd that has not yet been subjected to specific crystallization conditions. Figure 1 shows high-throughput XRPD (HT-XRPD) and high-resolution XRPD (HR-XRPD) in the top and bottom patterns, respectively. The starting materials include crystals suitable for single crystal structure analysis. As a result of crystallization of the starting material, a crystalline form having an asymmetric monoclinic P21 space group was obtained, which is referred to as crystalline form A. Table 1 provides the relevant dimensions of Form A.

The HR-XRPD pattern of the starting material was compared with the HR-XRPD pattern, which is a simulated pattern of a single crystal of crystalline Form A, and is shown in FIG. 2 . The crystal A has peaks in 7.7 °, 13.02 °, 15.34 °, 16.78 °, 18.62 °, 19.42 °, 21.94 °, 22.90 °, 25.70 °, 27.86 °, 28.70 °, 31.42 °, 32.70 °, and 37.46 ° 2 θ. Based on this comparison, the starting material is calculated to contain about 70% of crystalline Form A and about 30% of the other crystalline forms of aza-T-dCyd.

TGMS analysis of the starting material between 25-300 °C (10 °C/min) showed a mass loss of 11.7% between 100-170 °C, most likely due to organic solvents (Fig. 3). Simultaneously with the mass loss, the heat flow signal showed two endotherms with an exotherm in between. A third endotherm was observed around 195 °C due to the onset of melting and decomposition.

DSC analysis of the starting material between 25-300 °C (10 °C/min) showed two endotherms at 131 °C and 162 °C with an exotherm at 141 °C, consistent with the heat flow signal observed during the TGMS analysis. A third endothermic event was observed at T _peak of 196.4 °C, which was related to the melting of a non-solvated anhydrous phase (Fig. 4).

XRPD and single crystal structure analysis revealed that the starting material consisted of a mixture of crystalline phases. To further investigate the nature of the thermal phenomena, two cyclic DSC experiments were performed on the starting materials. One sample was heated to 170°C and cooled back to room temperature. As a result of analyzing the obtained solid by XRPD, it was consistent with the simulated pattern of crystalline Form A (FIG. 5). In a second cycling DSC experiment, the starting material was heated to 170 °C, cooled to 25 °C and then heated to 300 °C (Fig. 6). No thermal phenomena were observed during cooling, and only endothermic melting was observed at 194 °C in the second heating cycle, confirming the melting temperature of Form A.

Example 2: Production of amorphous material

For the polymorph screen, an amorphous material was generated from the starting material via a lyophilized solution method of aza-T-dCyd. Water, water/1,4-dioxane (50/50), water/THF (50/50) and water/tert-butyl alcohol (50/50% (v/v)) were added to aza-T-dCyd to obtain a solution of aza-T-dCyd in organic solvent for freeze-drying experiments. Lyophilization of the aza-T-dCyd solution resulted in poor crystalline materials containing impurities.

Example 3: Solubility Study

The thermodynamic solubility of aza-T-dCyd was determined according to the shake flask method. A suspension of crystalline aza-T-dCyd was prepared in 25 pure solvents. A small amount of solvent was added to aza-T-dCyd until thin suspensions were obtained. The sample was then equilibrated at room temperature under continuous stirring for 24 hours. After equilibration, a small amount of mother liquor was filtered and analyzed by HPLC. Concentrations of solutes were determined against a calibration curve of aza-T-dCyd. The solubility values of aza-T-dCyd at room temperature are listed in Table 3 according to the United States Pharmacopoeia classification (USP29). aza-T-dCyd was dissolved in high-boiling solvents such as DMF and DMA. In general, aza-T-dCyd is slightly or very slightly soluble in polar solvents and sparingly soluble in non-polar solvents.

Example 4: Polymorph Screen

A polymorph screen was performed with various pure organic solvents and solvent mixtures of various compositions, combining six different crystallization methods. Considering the poor thermal stability of aza-T-dCyd in solution and the limited stability of aza-T-dCyd in water and ketones, the screening experimental conditions were chosen as follows: (1) the experiment was started with a crystalline starting material; (2) the compound stayed in solution for a limited time (<5 days); (3) avoided high temperatures (<50 °C); (4) solid aza-T-dCyd is handled in a glovebox under as dry conditions as possible (relative humidity about 20%) to avoid moisture absorption; (5) avoided water and limited ketone use; and (6) moderate stress conditions to evaluate the physical stability of the resulting solid.

The following crystallization method was applied.

Solvent equilibration experiments .

Solvent equilibration experiments were performed at two temperatures: RT for 1 day and 5 °C for 5 days. Suspensions of aza-T-dCyd were prepared in different solvents along with the crystalline starting material and the solid was separated from the mother liquor upon completion of the equilibration time.

Evaporative crystallization experiments .

Evaporative crystallization experiments were set up using solvent equilibration experiments performed at room temperature and filtered mother liquors recovered from solvent mixtures. The solvent was slowly evaporated at ambient conditions and then further dried under vacuum (10 mbar) at 50 °C.

Anti-solvent experiments .

Anti-solvent experiments were performed using 10 solvent and anti-solvent combinations by reverse addition. A small amount of a highly concentrated solution of aza-T-dCyd was added to 20 mL of anti-solvent (Step 1).

Thermocycling experiments .

Thermocycling experiments were performed by preparing aza-T-dCyd suspensions in various solvents and solvent mixtures at room temperature. The resulting suspension was subjected to a temperature profile of 5 to 50 °C.

Sonication experiments .

Sonication experiments were performed by sonicating crystalline starting materials in the presence of small amounts of solvent.

Vapor diffusion into solution experiments .

Vapor diffusion experiments into solution were performed with the slow method of anti-solvent crystallization. A saturated aza-T-dCyd solution was exposed to anti-solvent vapor for one week at room temperature.

All solids obtained were analyzed by HT-XRPD after drying overnight in a glovebox at RT and 20% relative humidity and after drying overnight under vacuum (10 mbar) at RT. When the mother liquor was recovered, the mother liquor (ML) was evaporated and the recovered solid was analyzed by HT-XRPD. All solids were then exposed to accelerated aging conditions (25°C/60% RH) for 2 days and then reanalyzed by HT-XRPD.

Form A was the most abundant crystalline phase recovered from the screening experiment. Form A was found in all crystallization methods and in various solvents and solvent mixtures. From solvent equilibrium experiments, it was observed that Form A was obtained as a pure phase from solvents in which aza-T-dCyd was slightly or very slightly soluble.

In some solids, in addition to the XRPD pattern of Form A, the presence of peaks previously observed in the starting material was detected and described above. The aza-T-dCyd batch contained 70% Form A and 30% other crystalline phases. The presence of 30% of the other phases was highlighted most clearly at 16.0°, 17.6°, 24.8°, 26.3° and 34.1° 2θ, peaks appearing in the XRPD pattern. The solids recovered in the polymorph screen experiments were evaluated to attempt assignment and classification of these impurity peaks. An overview of the XRPD patterns of the starting materials, Form A, Form B, Form A+C1, Form A+C2, Form A+D1, and Form A+D2, is shown in FIG. 11 .

The peak at 26.3° 2θ belonged to Form B. The observed peak at 16.0° 2θ represents Form C1 and the peaks at 16.0 and 17.6° 2θ were attributed to Form C2. The observed peak at 24.8° 2θ is attributed to Form D1 and the peaks at 24.8 and 34.1° 2θ are attributed to Form D2. According to this assignment, some solids were classified as crystalline forms A+D1/D2, A+C1/C2 or A+B+D2.

Form B was obtained as a pure phase by solvent equilibration in DMA and DMF both at room temperature and 5° C. and also from thermocycling experiments in DMSO/2-ethyl-1-hexanol (50/50). Form B was physically unstable and converted to Form A after storage at 25ºC and 60% relative humidity.

Classes C and D were not observed as pure crystalline phases but were always admixed with Form A. In most cases, these mixtures converted to Form A after storage at 25° C. and 60% relative humidity.

A new crystalline form was discovered in a solution-based crystallization method in which no seeds of crystalline form A were present. These new forms have been classified as crystalline forms E, F, G1, G2, H, I, J, and K. Form E was obtained by antisolvent addition of DMA/chloroform or evaporative crystallization of DMA/TBME (80/20). Form E was converted to Form A after storage at 25ºC and 60% relative humidity.

Form F was obtained from vapor diffusion or evaporative crystallization in various solvents. Form F was physically stable. The peaks of Form F are 6.06°, 12.10°, 13.02°, 14.38°, 15.94°, 17.50°, 19.62°, 21.18°, 22.34°, 26.18°, 27.42°, 28.50°, 29.90°, 32.66°, 35.0 2°, 36.30°, 38.94°, and 41.06° 2θ.

Crystalline forms G1 and G2 have similar XRPD patterns, where some peaks shift between the two forms. Form G1 was obtained from avoiding solvent addition or from sonication. Form G2 was obtained from evaporative crystallization using DMA/EtOH. Both Form G1 and Form G2 were converted to Form A after storage at 25°C and 60% relative humidity.

Form H was obtained from evaporative crystallization in several solvent mixtures. This form is unstable. When obtained from NMP, Form H was converted to Form F. Form H was converted to Form A when obtained from other solvents.

Form I was obtained from evaporative crystallization from DMSO/IPA. Form I was converted to Form A after storage at 25°C and 60% relative humidity.

Form J was obtained from vapor diffusion into a solution with DMF as solvent and THF as anti-solvent. Form J was converted to Form A after storage at 25°C and 60% relative humidity.

Form K was observed in mixtures with Form F after evaporative crystallization from DMF. Form K was converted to form F after storage at 25°C and 60% relative humidity.

Form L was observed in the solid after storage at 25° C. and 65% relative humidity.

The XRPD patterns for each of these new forms are shown in FIG. 12 .

Example 5: Characterization of a novel form of AZA-T-DCYD

Each unique form identified in the screen was further characterized by TGMS and LCMS. Forms A and F appear to be anhydrous, while the other forms are solvated. Table 4 summarizes the crystallization conditions for the aforementioned crystalline forms of aza-T-dCyd, and Table 5 summarizes the properties of the various aza-T-dCyd crystalline forms (AAC indicates storage at 25 °C, 60% relative humidity).

Form A obtained from solvent equilibration experiments of RT in TFE was used for analytical characterization. TGMS results showed residual solvent release of about 0.7% in the temperature range of 30 - 190 °C (FIG. 12A). An endotherm was observed in the DSC trace at 205 °C due to melting and decomposition (Fig. 12B). LCMS analysis confirmed the integrity of 100% (area %) purity of Form A (FIG. 12C).

Form F obtained from evaporative crystallization experiments using DMF/acetonitrile (80/20, v/v) was used for characterization. The TGMS results showed a small loss of 1.1% between 30 and 140 °C, which may be mostly due to residual solvent (FIG. 15A). The DSC trace showed one endothermic event at about 170 °C due to melting and decomposition (Fig. 15B). LCMS analysis confirmed the integrity of the API with 100% purity (area %) (FIG. 15C).

Form A has a higher melting temperature than Form F and can be considered a more thermodynamically stable form. Both crystalline forms A and F are anhydrous.

Forms B, C2, D2, E, G1, G2, H, I, J and K, respectively, convert to Form A when solvated and stored at 25° C., 60% relative humidity for 2 days.

Form B obtained from solvent equilibration experiments in DMA at room temperature was further characterized. The TGMS results showed a gradual mass loss upon heating between 30 and 170 °C with a mass loss of 25.0%. The temperature at which decomposition begins is not clear due to the gradual loss of mass on heating. Form B can be a non-stoichiometric solvate that can be formed with different solvents. LCMS analysis indicated a solid purity of 97.3% aza-T-dCyd and the presence of impurities of 2.7% (area %).

Form C2 showed two additional peaks observed in the XRPD pattern in mixtures with other forms. TGMS analysis showed a mass loss of 0.7% over the temperature range of 30 - 160 °C. The heat flow signal showed only one endothermic event around 190 °C, which could be related to the melting and decomposition of Form A. The investigation of form C2 is inconclusive because form C2 is only present in trace amounts in mixtures with crystalline form A. Therefore, the nature of this form is still unclear. However, it appears to be a true (pseudo-)polymorph of aza-T-dCyd, as the chemical purity of the total solid sample was 100% (area %).

Form D2 exhibited two additional peaks observed in the XRPD pattern in mixtures with Form A. TGMS analysis of Form A+D2 showed that Form D2 was most likely the solvated form. A mass loss of 5.1% was observed between 90 and 170 °C. The results were inconclusive with respect to the released solvent. LCMS analysis of the crystalline mixture confirmed the integrity of aza-T-dCyd with a chemical purity of 100% (area %).

Form E from evaporative crystallization experiments using DMA was further analyzed by TGMS and LCMS. The TGMS results showed a 25.8% mass loss of DMA, which corresponds to 1 molar equivalent of solvent. The solvent was released in a stepwise manner between 90 and 160 °C, suggesting that Form E is a mono-DMA solvate. After desolvation, an endothermic event was recorded at 200 °C, probably corresponding to melting of Form A. Compound integrity was confirmed by LCMS analysis.

Class G is an isostructural class of solvates. Crystalline forms G1 and G2 were further characterized by TGMS and LCMS. LCMS analysis confirmed compound integrity (area % of 100%). Form G1 obtained from antisolvent addition experiments using NMP and cyclohexane was used for characterization. TGMS results showed a stepwise mass loss of 27.5% between 90 and 160 °C. The 27.5% mass loss corresponds to approximately one molecule of NMP per molecule of aza-T-dCyd, so Form G1 may be a mono-NMP solvate. The DSC signal recorded two endotherms around 110 and 150 °C due to solvent loss and a third endotherm at 200 °C, which may correspond to the melting of Form A. Form G2 was obtained by evaporative crystallization in DMA/ethanol (80/20, v/v). The 14.6% mass loss observed by TGMS between 70 and 120 °C corresponds to 0.5 molar equivalents of DMA. This suggested that Form G2 may be a hemi-DMA solvate. In the DSC signal, two endotherms were observed around 80 and 90 °C due to solvent loss, and a third endotherm around 195 °C due to melting and decomposition.

Form H obtained from evaporative crystallization from NMP/THF (80/20, v/v) was used for characterization of Form H. The gradual mass loss observed by TGMS analysis was 15.3% from 30 to 180°. C corresponds to about 0.5 molar equivalent of NMP. At the same time, an extensive endotherm was observed around 130 °C. Form H was observed in experiments with other solvents and is therefore most likely a non-stoichiometric solvate capable of incorporating other solvent molecules into the crystal structure. A second extensive endotherm was observed in the DSC trace due to decomposition at around 220 °C. From the TGMS data, it is not clear where solvent loss ends and where thermal decomposition begins. Events may partially overlap. To obtain dry samples, the solids had to be vacuum dried at 50 °C for 24 hours. This may have affected the purity as the LCMS data indicated that the solid was 82% (area %) pure.

Form I was obtained by evaporative crystallization from DMSO/IPA (80/20, v/v). TGMS data showed a gradual mass loss of 14.7% between 30 and 170 °C. A mass loss of 14.7% corresponds to about 0.5 molar equivalent of DMSO. Form I may be a hemi-DMSO solvate. The DSC trace showed two broad endotherms at 70ºC and 110 °C due to mass loss, and a third endotherm around 190 °C due to melting and decomposition processes.

Form J, precipitated by vapor diffusion from solution using DMF and THF, was further characterized. TGMS data showed a 7.6% mass loss of THF stepwise between 120 and 170 °C. The mass loss corresponds to about 0.3 molar equivalents of THF and Form J is therefore most likely a non-stoichiometric solvate. The DSC trace recorded two endotherms at 120 and 150 °C due to solvent loss, and the third endotherm recorded at 200 °C, consistent with the melting/decomposition event of Form A.

Form K was observed once in a mixture with Form F and was obtained by evaporation from a DMF solution. The mixture was further characterized. TGMS analysis showed a mass loss of 6.3% between 30 and 160 °C, probably due to the loss of DMF. The mass loss was accompanied by a small endotherm around 110 °C. Two large endotherms were observed at 180 and 195 °C. The endotherm at 195 °C may be due to melting and decomposition of Form A. Since Form K is a mixture with Form F (the unsolvated form), Form K is most likely the solvated form.

Form L was a poor crystalline solid observed only after storage at 25° C., 60% relative humidity and very low yield. TGMS analysis observed a mass loss of 2.8% between 30 and 170 °C followed by decomposition. The absence of thermal events in the DSC traces may be due to the small amount of sample used for analysis. It is not clear whether the mass loss is due to solvent trapped in the crystal structure or residual solvent. The nature of Form L is unclear as no further characterization could be performed.

Attempts to grow single crystals from aza-T-dCyd solutions in acetonitrile have shown that the resulting crystals are acetonitrile solvates. This step was not observed in any of the screening experiments. The solvate crystallized into a monoclinic P21 space group with cell dimension of a = 9.2948(15), b = 7.3509(9), c = 10.2312(15) Å, β = 107.661(2)°, V = 666.10(17) Å ³ , Z = 2 and density of 1.423 g/cm ³ . Since only single crystals were formed (very low yield), no further characterization was performed on this form and its physical stability should also be investigated.

Example 6: Pharmacokinetics of aza-T-dCyd

The pharmacokinetic properties of aza-T-dCyd (starting material; SM; aza-T-dCyd not yet subjected to specific crystallization conditions) were studied as follows.

Aza-T-dCyd starting material (SM) was administered to 6 female NOD-SCID mice divided into 4 groups. Group 1 was the vehicle control group. Group 2 was administered with 2.0 mg/kg of aza-T-dCyd starting material (SM) once a day, and group 3 was administered with 1.0 mg/kg of aza-T-dCyd starting material (SM) twice a day. In Groups 2 and 3, the aza-T-dCyd starting material (SM) was administered in the above amount for 5 days, followed by a 2-day break, another 5 days of administration, and then a 9-day break. This cycle repeated itself. Group 4 administered aza-T-dCyd starting material (SM) 1.0 mg/kg once a day for 5 days and then repeated this cycle with a 2-day break. The tumor size of the mouse was measured using a fluorescent agent, and the results are shown in FIG. 17 .

As shown in FIG. 17 , tumor size increased in group 1 (vehicle control group). In addition, it was confirmed that the increase in tumor size was most significantly suppressed in group 2. In contrast, despite the expectation that the AUC of SM in group 3 would be the same as in group 2, it was observed that the size of the tumor rapidly increased after 40 days of administration. From this, it was found that aza-T-dCyd was Cmax dependent rather than AUC dependent.

In addition, as shown in FIG. 18, the tumor size of group 2 (2.0 mg/kg, once a day) was significantly smaller than that of group 1 (1.0 mg/kg, twice a day), demonstrating the results on day 43.

In addition, hematological malignant cells (Mv4-11) were treated with aza-T-dCyd starting material (SM), and the half-maximal inhibitory concentration (IC ₅₀ ) was measured at 1 hour, 2 hours, and 4 hours. The results are shown in FIG. 19 . The measured IC ₅₀ at 1 hour was about 160 nM, so the IC ₅₀ at 2 hours was expected to be 80 nM and the IC ₅₀ at 4 hours to be 20 nM. However, the IC ₅₀ measured at 2 h was about 120 nM, much higher than the expected value of 80 nM. In addition, the IC ₅₀ measured at 4 hours was about 80 nM, much higher than the expected value of 20 nM. Accordingly, it was confirmed that the efficiency of the aza-T-dCyd starting material (SM) decreased significantly as the exposure time of the compound to cells increased. This suggests that exposure to a higher amount of aza-T-dCyd starting material (SM) for a short period of time can provide an efficient treatment.

Thus, the data suggest that crystalline polymorphs with large dissolution profiles, such as Form A or Form F in Example 7 described below, are superior to aza-T-dCyd starting material (SM) and other crystalline polymorphs with inferior dissolution profiles. Also for the same reason, the data suggest that crystalline polymorphs such as Form A or Form F exhibit improved PK profiles over the aza-T-dCyd starting material (SM) and other crystalline polymorphs.

That is, although the crystalline form usually has disadvantages in solubility compared to the amorphous form, in the case of crystalline forms A and F, although they are physicochemically stable forms, the dissolution characteristics according to the pH condition are very uniform and rapidly. Since they dissolve stably, the crystalline form can be controlled and used according to the need for formulation development.

Example 7: Dissolution Profiles of Form A and Form F at Various pH Points

The dissolution rates of Form A and Form F were measured at pH 1.2, pH 6.5 and pH 5.0 and are shown in Table 6 and FIGS. 20-22.

As shown in Table 6 and FIGS. 20 to 22, at pH 1.2 (gastric and colonic pH conditions), Form A and Form F exhibited similar dissolution rates, whereas Form A exhibited a more consistent dissolution profile compared to Form F. At pH 6.5 and pH 5 (pH conditions of the appendix and small intestine), Form F showed a higher dissolution rate than Form A.

This suggests that Form A can be prepared into a variety of drug forms that target release of the drug's active ingredient at about pH 1.2 (e.g., stomach or large intestine). Further, this suggests that Form F can be made into various drug forms that target release of the drug's active ingredient at about pH 5.0-6.5 (eg, small intestine).

Example 8: Pharmacokinetic Comparison of AZA-T-DCYD Starting Material, Form A and Form F

The pharmacokinetic properties of aza-T-dCyd (starting material, SM, aza-T-dCyd not yet subjected to specific crystallization conditions), Form A and Form F were studied as follows.

The aza-T-dCyd starting material (SM), crystalline form A, and crystalline form F were prepared in the form of capsules mixed with microcrystalline cellulose in an 8:92 (w/w) ratio, respectively, at 2 mg/kg of SM, crystalline form A or crystalline form F. Each of the SM capsules, crystalline A capsule, and crystalline Form F capsule was administered at a dose of 2 mg/kg to 2 male SD rats (i.e., a total of 6 male SD rats). As shown in Tables 7-9, plasma concentrations of each of SM, Form A and Form F in tested SD rats were measured at 0.25, 0.5, 1, 2, 4, 6, 8 and 24 hours after capsule administration.

coefficient of variation ( CV )

standard deviation

In addition, pharmacokinetic parameters were obtained as shown in Tables 10-12 below.

As shown above, Form A and Form F showed higher Cmax values than SM. In particular, Form A showed a Cmax value about 1.3 times higher than SM, and Form F showed a Cmax value about 1.4 times higher than SM. In addition, both crystalline form A and crystalline form B showed about 30% higher AUC values than SM.

Example 9: AZA-T-DCYD Comparison of half maximal inhibitory concentration (IC ₅₀ ) of starting material and Form A

K562 and HL-60 cell lines were cultured and maintained in RPMI (10% FBS, 1% penicillin-streptomycin) medium at 37°C, 95% air and 5% CO ₂ . K562 and HL-60 cell lines were each seeded in 96-well plates at a density of 3000 cells/well (90 μl). Crystalline A and SM were treated in 10 μl using a 3-fold dilution, and each well was treated at a final concentration of 10 μM. Cells were incubated for 3 days at 37°C, 95% air and 5% CO ₂ . The 96-well plate was left at room temperature for 30 minutes to equilibrate. Then, 100 μl of CellTiter-Glo® Luminescent Cell Viability Assay Reagent was added to the 96-well and incubated for 10 minutes at room temperature. Luminescence was measured using a Luminometer, and IC ₅₀ values were analyzed using GraphPrism.

As shown in FIGS. 23 and 24 , Form A exhibits an IC ₅₀ value about 5% lower than SM, providing a greater effect.

Example 10: PK experiment in rats according to the ratio (wt%) of crystalline form A in drug Aza-T-dCyd

When 1 capsule/rat of the capsule formulation prepared by varying the content (wt%) of Aza-T-dCyd drug was orally administered to male SD rats, the result of Cmax was confirmed through a PK experiment. As shown in Table 13, in the Aza-T-dCyd drug whose therapeutic effect is dependent on Cmax, the optimal therapeutic effect when administered orally is an important factor in the ratio (wt%) of crystalline form A in the Aza-T-dCyd drug.

Specifically, samples for PK experiments upon oral administration were prepared as follows. A drug raw material of Aza-T-dCyd with 100% pure crystalline form A was prepared. After synthesizing the Aza-T-dCyd compound, pure crystalline Form F could not be obtained, and the obtained Aza-T-dCyd drug substance was a polycrystalline form composed of 52% of crystalline form F, 15% of crystalline form A, and 33% of other undefined forms. This polycrystalline Aza-T-dCyd drug substance was named polycrystalline form F'. 100% pure crystalline Aza-T-dCyd drug substance and polycrystalline form F' named Aza-T-dCyd drug substance were mixed in various composition ratios, and microcrystalline cellulose was added as an excipient to prepare oral dosage form capsule samples for PK experiments. At this time. The ratio of Aza-T-dCyd drug raw material and excipient is 8:92 (w/w).

The experimental group for the PK experiment consisted of a total of 11 groups, and the ratio (mole %) of Aza-T-dCyd in each experimental group (crystal form A: other forms) of Aza-T-dCyd is shown in Table 13 below.

PK analysis was performed by blood sampling at 0.25, 0.5, 1, 2, 4, 6, 8, and 24 hr after oral administration, and the results are shown in Table 14 and FIG. 25.

Aza-T-dCyd significantly increased Cmax to 852-897 ng/mL when the ratio of crystalline form A was 70% or more. Specifically, when the ratio of crystalline form A in the Aza-T-dCyd drug is 70% or more in rats, the Cmax stably shows 850 ng/mL, but when the ratio is less than 70%, the Cmax has 442-703 ng/mL.

This suggests that it is very important to design and quality control the ratio of crystalline form A in the Aza-T-dCyd drug to 70 wt% or more due to the characteristics of the Cmax-dependent Aza-T-dCyd drug in terms of therapeutic effect.

Example 11: Side Effect Control of Drug Aza-T-dCyd

As described above, when comparing the results of treating mice with 2mpk and 1mpk twice, the weight loss of mice was severe in the latter. In addition, when the in vivo luciferase activity was measured, when the treatment was divided into two times, the rate of increase was steeper, so it can be predicted that the tumor inhibitory effect is more excellent when 2mpk is treated at one time.

That is, rather than exposure to the Aza-T-dCyd drug several times, a desired therapeutic effect can be exerted by sufficiently exhibiting the drug effect through rapid treatment in a short period of time.

Therefore, the present invention can solve the problem that the variance in drug exposure among individual patients is very high, so that the optimal administration dose for one individual becomes a dose that cannot show a therapeutic effect for another individual, and for another individual, a dose that causes severe toxicity, resulting in a very narrow therapeutic window.

Example 12: Absorption profile in the gastrointestinal tract when formulated for oral dosage form

To confirm the absorption profile in the gastrointestinal tract, the Aza-T-dCyd drug was orally administered (PO) to the mice, and after inserting the Aza-T-dCyd drug into the caecum with a cannula, plasma concentration graphs as shown in FIG. 26 were obtained.

26A and 26B show plasma concentration distribution graphs showing drug absorption through the gastrointestinal tract in the case of PO as a result of PO and IC administration of 1mpk and 3mpk, respectively. However, in the case of IC, a very small amount of drug appears to be present in the plasma, that is, it shows a very low exposure to plasma compared to PO. That is, when administered orally, Aza-T-dCyd is not absorbed from the caecum (the digestive organ that swells like a pouch at the beginning of the large intestine).

That is, since it can be inferred that the aza-T-dCyd drug is not absorbed in the postcecal region, which is the starting point of the large intestine, toxic side effects can be controlled in a narrow treatment window of the aza-T-dCyd drug.

In the case of the Aza-T-dCyd drug, it can be inferred that formulation so that the drug is dissolved in the stomach and absorbed at the beginning of the small intestine is highly stable and is closely related to improving the efficacy of the drug.

In addition, since there is no additional dissolution in the small intestine, Cmax can be implemented within the therapeutic window range, and toxic side effects due to the narrow therapeutic window can also be solved.

The foregoing description of embodiments has been presented for purposes of illustration and description. It is not intended to be exhaustive or limiting the disclosure. Individual elements or features of a particular embodiment are generally not limited to that particular embodiment, but, where applicable, are interchangeable and can be used in a selected embodiment, even if not specifically shown or described. The same thing can be different in many ways. Such variations are not to be regarded as a departure from this disclosure, and all such modifications are intended to be included within the scope of this disclosure.

Claims (26)

1. In the method for preparing an oral dosage form containing 5-aza-4'-thio-2'-deoxycytidine (aza-T-dCyd), the efficacy of which is dependent on the highest blood concentration (Cmax), as an active ingredient,

   A method for preparing an oral dosage form characterized by precisely designing a single dose that exhibits a desired therapeutic effect from the highest blood concentration (Cmax) of an aza-T-dCyd drug within an acceptable error range.
2. The method of claim 1, wherein the therapeutic effect is an anti-cancer effect.
3. The method of claim 1, wherein the single dose of aza-T-dCyd is designed so that the highest blood concentration (Cmax) of the aza-T-dCyd drug calculated from the single dose of the aza-T-dCyd drug is lower than the value corresponding to the no-observed-adverse-effect level (NOAEL) or the highest non-severe toxic dose (HNSTD) that does not cause serious toxicity. Preparation of oral dosage forms.
4. The method of claim 1 , wherein: (i) the highest blood concentration (Cmax) of the aza-T-dCyd drug that exerts the desired therapeutic effect, optionally (ii) the amount of the aza-T-dCyd drug that reaches the systemic circulation from the oral dosage form, and optionally (iii) the time taken for the aza-T-dCyd drug to reach the systemic circulation; A method for preparing an oral dosage form characterized by designing the ratio of Form A.
5. The method of claim 1, wherein in order to adjust the ratio (wt%) of crystalline form A in the aza-T-dCyd drug in the oral dosage form, a crystalline raw material containing crystalline form A in a desired ratio is prepared from aza-T-dCyd crude materials, which are synthesized products of the aza-T-dCyd compound, and then formulated into an oral dosage form.
6. The preparation method of an oral dosage form according to claim 5, wherein preparing the crystalline raw material containing crystalline Form A in a desired ratio from aza-T-dCyd crude materials, which are synthesized products of the aza-T-dCyd compound, is (i) a process of controlling the desired ratio (wt%) of Form A within ±10% error range or (ii) a process of confirming that the desired ratio (wt%) of Form A is within ±10% error range.
7. An oral dosage form in which the ratio (wt%) of crystalline form A in 5-aza-4'-thio-2'-deoxycytidine (aza-T-dCyd) drug is controlled, wherein the oral dosage form is formulated into an oral dosage form after preparing a crystalline raw material containing crystalline form A in a desired ratio from aza-T-dCyd crude materials, which is a synthetic product of an aza-T-dCyd compound.
8. [Claim 8] The oral dosage form according to claim 7, wherein the dose of crystalline Form A stably realizes the exposure amount of aza-T-dCyd in the blood within an acceptable error range.
9. 8. The oral dosage form according to claim 7, which is designed to dissolve 90% or more of Form A in the stomach.
10. [Claim 8] The oral dosage form according to claim 7, which contains a carrier disintegrating so that at least 80% of the aza-T-dCyd drug is dissolved in an acidic stomach.
11. [Claim 8] The oral dosage form according to claim 7, wherein the amount of crystalline form A in the aza-T-dCyd drug is 70wt% or more.
12. 8. The oral dosage form according to claim 7, which contains 5-70 mg/m ² , preferably 5-55 mg/m ² , more preferably about 5-30 mg/m ² , even more preferably 5-20 mg/m ² of Form A in a single dose of the drug aza-T-dCyd.
13. The method of claim 7, wherein the preparation of the crystalline raw material containing the crystalline form A in a desired ratio (wt%) from the aza-T-dCyd crude materials, which are synthesized products of the aza-T-dCyd compound, means that (i) the desired ratio (wt%) of the crystalline form A is within ±10% error range or (ii) the desired percentage (wt%) of Form A within a ±10% margin of error. An oral dosage form characterized in that it is a process for confirming that it is within.
14. In the method for preparing an oral dosage form containing 5-aza-4'-thio-2'-deoxycytidine (aza-T-dCyd) as an active ingredient,

    A method for preparing an oral dosage form characterized in that the ratio of crystalline form A and/or crystalline form F in the crystalline raw material of aza-T-dCyd is confirmed and then formulated into an oral dosage form.
15. 15. The method of claim 14, wherein the aza-T-dCyd-containing oral dosage form is designed to dissolve 90% or more of Form A and/or Form F in the stomach.
16. The method according to claim 14, wherein the maximum blood concentration (Cmax) and/or the area under the drug concentration-time curve (AUC) of the aza-T-dCyd drug is adjusted by adjusting the composition ratio of Form A and/or Form F of the aza-T-dCyd drug.
17. An oral dosage form characterized in that a single dose of the aza-T-dCyd drug is designed to contain more than the ratio of Form A corresponding to the Cmax phase inflection point at which the maximum blood concentration (Cmax) change value according to the change in the ratio of Form A in the same dose of aza-T-dCyd drug is increased.
18. 18. The oral dosage form according to claim 17, which contains 5-70 mg/m ² , preferably 5-55 mg/m ² , more preferably about 5-30 mg/m ² , even more preferably 5-20 mg/m ² of Form A in a single dose of the drug aza-T-dCyd.
19. [Claim 18] The oral dosage form according to claim 17, wherein the ratio (wt%) of crystalline Form A in the drug of aza-T-dCyd is 70% or more.
20. 5-aza-4'-thio-2'-deoxycytidine (aza-T-dCyd) An oral dosage form containing at least 70 wt % of Form A as defined by a powder X-ray diffraction spectrum in the drug.
21. 5-70 mg/m ² , preferably 5 to 55 mg/m ² , more preferably about 5 to 30 mg/m ² , still more preferably 5 to 20 mg/m ² of crystalline Form A defined by powder X-ray diffraction spectrum when designing a single dose of 5-aza-4'-thio-2'-deoxycytidine (aza-T-dCyd). .
22. In the method for preparing an oral dosage form containing 5-aza-4'-thio-2'-deoxycytidine (aza-T-dCyd) drug whose efficacy is dependent on the highest blood concentration (Cmax),

    A first step of crystallizing the aza-T-dCyd compound in the presence of a solvent and then converting the aza-T-dCyd compound into a non-solvate crystalline form by removing the solvent; and

    The second step of preparing an oral dosage form designed so that the non-solvate crystalline form prepared in the first step can be dissolved in the stomach.

    A method for preparing an oral dosage form containing an aza-T-dCyd drug comprising:
23. 23. The preparation method of an aza-T-dCyd drug-containing oral dosage form according to claim 22, wherein the non-solvate crystalline form prepared in the first step is crystalline form A defined by a powder X-ray diffraction spectrum.
24. 24. The preparation method of the aza-T-dCyd drug-containing oral dosage form according to claim 23, wherein the first step is to convert the crystalline polymorph containing the solvate into a single crystalline form A by removing the solvent.
25. 24. The method of claim 23, wherein the aza-T-dCyd drug is composed only of crystalline Form A.
26. 23. The preparation method of an oral dosage form containing aza-T-dCyd according to claim 22, wherein the second step is to provide an oral dosage form containing an aza-T-dCyd drug that is precisely designed within an acceptable error range for a single dose that exhibits a desired therapeutic effect from the highest blood concentration (Cmax) of the aza-T-dCyd drug.

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