A co-crystal drug ZM851 and its application in anti-lung cancer

Abstract

The application relates to the field of biological medicine, and discloses a co-crystal drug ZM851 and application thereof in resisting lung cancer, and a preparation method of the co-crystal drug ZM851, which comprises the following steps: mixing osimertinib, dihydroartemisinin and a mixed solvent, heating, stirring and dissolving, and then adding n-hexane; and yellow solid separated out is the co-crystal drug ZM851. The anti-tumor activity of the co-crystal molecule ZM851 is more than ten times that of a non-co-crystal mixture. In addition, in an in-vitro experiment, the ZM851 shows a significant inhibitory effect on an osimertinib-resistant cell strain. The significant inhibitory effect of the co-crystal drug ZM851 on the osimertinib-resistant cell strain in the in-vitro experiment indicates that the co-crystal drug ZM851 can provide a new treatment scheme for lung cancer patients in clinical treatment.

Description

Technical Field

[0001] This invention relates to the field of biomedical technology, and more specifically, to a eutectic drug ZM851 and its application in the treatment of lung cancer. Background Technology

[0002] Lung cancer is the leading cause of cancer morbidity and mortality worldwide, with non-small cell lung cancer (NSCLC) accounting for 80% to 85% of cases, while small cell lung cancer (SCLC) accounts for 10% to 15%. Epidermal growth factor receptor (EGFR) is one of the key targets for treating NSCLC, and first- to third-generation NSCLC therapies targeting this target have been successfully launched. Osimertinib, as a third-generation epidermal growth factor receptor tyrosine kinase inhibitor (EGFR-TKI), not only overcomes the resistance problem caused by methionine (TM790) mutations in first-generation EGFR-TKIs but also addresses the insufficient selectivity of second-generation EGFR-TKIs for wild-type EGFR. However, patients may develop resistance after 9 to 14 months of osimertinib treatment.

[0003] Therefore, developing new drugs that can inhibit osimertinib-resistant cell lines is particularly crucial. Summary of the Invention

[0004] In view of this, the present invention proposes a eutectic drug ZM851 and its application in the treatment of lung cancer, aiming to solve the problem of drug resistance in existing treatment regimens.

[0005] This invention proposes a compound ZM851, the chemical structural formula of which is:

[0006]

[0007] The present invention also provides a method for preparing the above-mentioned compound ZM851, comprising:

[0008] Osimertinib, dihydroartemisinin, and a mixed solvent were mixed, heated, stirred, and dissolved. Then, n-hexane was added, and the precipitated yellow solid was the compound ZM851.

[0009] Preferably, the mixed solvent is acetone and water.

[0010] Preferably, the volume ratio of acetone to water is 15:1.

[0011] Preferably, the temperature at which osimertinib, dihydroartemisinin, and the mixed solvent are heated and stirred to dissolve them is at least 60°C.

[0012] The present invention also provides an application of the above-mentioned compound ZM851, the application of which includes at least one of the following:

[0013] A1) Applications in the preparation of antitumor drugs;

[0014] A2) Use in the preparation of pharmaceutical compositions for the prevention or treatment of drug-resistant tumors;

[0015] A3) Application in the preparation of drug resistance sensitizers for antitumor drugs.

[0016] The present invention also provides a cocrystal drug, wherein the active ingredient of the cocrystal drug is the above-mentioned compound ZM851.

[0017] The present invention also provides an application of the above-described cocrystallized drug, the application comprising at least one of the following:

[0018] A1) Applications in the preparation of antitumor drugs;

[0019] A2) Use in the preparation of pharmaceutical compositions for the prevention or treatment of drug-resistant tumors;

[0020] A3) Application in the preparation of drug resistance sensitizers for antitumor drugs.

[0021] Compared with the prior art, the beneficial effects of the present invention are as follows:

[0022] This invention successfully synthesized one co-crystal molecule and two non-co-crystal compounds. In particular, the co-crystal molecule ZM851 exhibited more than 10 times the antitumor activity of its non-co-crystal mixture. Furthermore, in in vitro experiments, ZM851 showed a significant inhibitory effect on osimertinib-resistant cell lines.

[0023] The significant inhibitory effect of the cocrystallized drug ZM851 on osimertinib-resistant cell lines in in vitro experiments suggests that it may provide a new treatment option for lung cancer patients in clinical practice. The application of the cocrystallized drug ZM851 is expected to improve treatment efficacy, prolong progression-free survival, and may have a positive impact on overall survival.

[0024] In summary, the eutectic drug ZM851 and its preparation method of the present invention bring new strategies to the field of lung cancer treatment and are expected to play a key role in future clinical practice. Attached Figure Description

[0025] Various other advantages and benefits will become apparent to those skilled in the art upon reading the following detailed description of preferred embodiments. The accompanying drawings are for illustrative purposes only and are not intended to limit the invention. Furthermore, the same reference numerals denote the same parts throughout the drawings. In the drawings:

[0026] Figure 1 For compound ZM8511 HNMR spectrum;

[0027] Figure 2 The XRD pattern of compound ZM851 is shown below.

[0028] Figure 3 The Fourier transform infrared spectrum of compound ZM851 is shown below.

[0029] Figure 4 The Raman spectrum of compound ZM851;

[0030] Figure 5 The TG and DSC spectra of compound ZM851 are shown.

[0031] Figure 6 The image shows the 1H NMR spectrum of compound ZM852.

[0032] Figure 7 The XRD pattern of compound ZM852 is shown below.

[0033] Figure 8 The Fourier transform infrared spectrum of compound ZM852 is shown below.

[0034] Figure 9 The Raman spectrum of compound ZM852;

[0035] Figure 10 The TG and DSC spectra of compound ZM852 are shown.

[0036] Figure 11 The image shows the 1H NMR spectrum of compound ZM853.

[0037] Figure 12 The XRD pattern of compound ZM853 is shown below.

[0038] Figure 13 The Fourier transform infrared spectrum of compound ZM853 is shown below.

[0039] Figure 14 The Raman spectrum of compound ZM853 is shown below.

[0040] Figure 15 The images show the TG and DSC spectra of compound ZM853. Detailed Implementation

[0041] Exemplary embodiments of the present disclosure will now be described in more detail with reference to the accompanying drawings. While exemplary embodiments of the present disclosure are shown in the drawings, it should be understood that the present disclosure may be implemented in various forms and should not be limited to the embodiments set forth herein. Rather, these embodiments are provided to enable a more thorough understanding of the present disclosure and to fully convey the scope of the disclosure to those skilled in the art. It should be noted that, unless otherwise specified, embodiments and features in the embodiments of the present invention can be combined with each other. The present invention will now be described in detail with reference to the accompanying drawings and embodiments.

[0042] Example 1

[0043] I. Preparation of compound ZM851

[0044] Weigh osimertinib (200 mg, 0.4 mmol) and dihydroartemisinin (114 mg, 0.4 mmol) into a reaction tube, add 1.5 mL of a mixed solvent (acetone:water = 15:1), heat to 60 °C and stir to dissolve. Then add 3 mL of n-hexane, precipitating a yellow solid. Filter and wash to obtain 251 mg of yellow powder, namely compound ZM851, with the following chemical structural formula:

[0045]

[0046] II. Preparation of compound ZM852

[0047] Weigh osimertinib (200 mg, 0.4 mmol) and gefitinib (179 mg, 0.4 mmol) into a reaction tube, add 1.5 mL of mixed solvent (ethanol:water = 15:1), heat to 80 °C and stir to dissolve. Then cool to 60 °C, add 3 mL of n-hexane, and a light brown solid precipitates. Filter and wash to obtain 361 mg of light brown powder, namely compound ZM852, with the following chemical structural formula:

[0048]

[0049] III. Preparation of Compound ZM853

[0050] Weigh osimertinib (200 mg, 0.4 mmol) and ibrutinib (176 mg, 0.4 mmol) into a reaction tube, add 1.5 mL of mixed solvent (ethanol:water = 15:1), heat to 80 °C and stir to dissolve. Then cool to 60 °C, add 3 mL of n-hexane, and slowly cool to room temperature, precipitating a light brown solid. Filter and wash to obtain 323 mg of light brown powder, namely compound ZM853, with the following chemical structural formula:

[0051]

[0052] Example 2: Structural characterization analysis of the compound prepared in Example 1.

[0053] The following is a characterization analysis of ZM851, ZM852 and ZM853.

[0054] I. Process

[0055] ¹H NMR: ¹H NMR data were acquired at room temperature using a DSX-500 NMR spectrometer from Bruker Bayespen GmbH, Switzerland, with DMSO-d6 as the solvent and TMS as the internal standard. Powder X-ray diffraction (PXRD): PXRD data were obtained using a Rigaku Ultrama IV diffractometer from Rigaku Ltd., Japan. The Rigaku Ultrama IV powder Xray Cu Ka diffractometer was set with the following parameters: wavelength 1.5418 Å, voltage 40 kV, current 40 mA, sample form: powder, target material: copper, scanning range: 5–90° (2θ), scanning speed: 10° / min. All sample data were obtained at room temperature.

[0056] Fourier transform infrared spectroscopy (FT-IR): FT-IR data were obtained using a Nicoleti N10 spectroscopy unit manufactured by Thermo Fisher Scientific, USA. The maximum number of axes for routine testing ranges from 400 to 4000 cm⁻¹, and is used for routine powder compression and liquid cell tests. The experimental data in this paper were obtained using the KBr pelleting method, where KBr was ground to 2 μm using an agate mortar. A certain amount of sample (2-5 mg) and KBr (100-120 mg) were added, ground and mixed until no obvious particles remained. The mixture was then placed in a mold and pressed into a transparent or translucent pellet.

[0057] Raman spectroscopy (RM): Raman data was obtained using a Thermo Fisher Scientific DXR detector. The sample should be placed as flat as possible on the sample holder to ensure the accuracy of the spectral data. Appropriate laser wavelength, power, and detector parameters should be selected according to the sample properties and experimental requirements to ensure the clarity and stability of the spectral signal. A background scan must be performed before sample testing to eliminate interference from the instrument's background signal.

[0058] Simultaneous thermal analysis (TG-DSC): The test data for simultaneous thermal analysis were obtained using a Hitachi 200 instrument from Japan. TG-DSC testing can simultaneously obtain TG, DSC, and DTG curves. The sample temperature range is from room temperature to below 800℃ (inclusive). The specific test temperature is 30-600℃, the test gas atmosphere is N2, and the heating rate is 10℃ / min.

[0059] II. Results

[0060] 1. Structural Characterization and Analysis of ZM851

[0061] like Figure 1 As shown, the 1H NMR spectrum of the ZM851 eutectic exhibits the following data: 1H NMR (500MHz, DMSO-d6) δ 10.20 (s, 1H), 9.15 (s, 1H), 8.68 (s, 1H), 8.33 (d, J = 5.3Hz, 1H), 8.24 (d, J = 8.0Hz, 1H), 7.89 (s, 1H), 7.52 (d, J = 8.2Hz, 1H), 7.26–7.22 (m, 2H), 7.15 (t, J = 5.3Hz, 1H). The values ​​are 7.5 Hz (1H), 7.04 (s, 1H), 6.46–6.40 (m, 1H), 6.29 (s, 1H), 5.77 (d, J = 10.1 Hz, 1H), 3.92 (s, 3H), 3.86 (s, 3H), 2.89 (t, J = 5.8 Hz, 2H), 2.72 (s, 3H), and 2.21 (s, 6H). These data correspond to the 1H NMR spectrum of osimertinib. On the other hand, the ¹H NMR (500MHz, DMSO-d⁶) values ​​were δ 6.22 (d, J = 4.3Hz, ¹H), 5.43 (s, ¹H), 4.97 (t, J = 3.9Hz, ¹H), 2.19–2.13 (m, ¹H), 2.01–1.96 (m, ¹H), 1.88–1.76 (m, 2H), 1.68–1.62 (m, ¹H), 1.59–1.53 (m, ¹H), and 1.42–1.3. The 1H NMR spectra of dihydroartemisinin are as follows: 1 (m, 3H), 1.27 (d, J = 6.8 Hz, 3H), 1.16–1.09 (m, 1H), 0.89 (t, J = 6.6 Hz, 3H), 0.83 (d, J = 7.3 Hz, 3H), and 0.78 (d, J = 7.2 Hz, 1H). Two of the three hydrogen atoms at 2.29 (t, J = 5.8 Hz, 3H) belong to osimertinib, and one belongs to dihydroartemisinin. Calculations show that the molar ratio of API (active pharmaceutical ingredient) to CCF (cocrystallizing agent) in the cocrystallization molecule is 1:1.

[0062] The X-ray diffraction (XRD) pattern of ZM851 is shown below. Figure 2 As shown, the X-ray diffraction pattern of osimertinib shows characteristic peaks at 2θ angles of 9.16°, 10.34°, 15.88°, 16.6°, 17.58°, 20.44°, 20.82°, 22.74°, 23.14°, 24.1°, 25.98°, 26.54°, and 27.3°. The X-ray diffraction pattern of dihydroartemisinin shows characteristic peaks at 2θ angles of 7.72°, 9.28°, and 11.16°. However, the X-ray diffraction pattern of ZM851 exhibits new characteristic peaks at 2θ angles of 5.88°, 15.06°, 18.36°, 24.62°, and 25.26°, which preliminarily indicates the formation of a new crystal phase.

[0063] like Figure 3 As shown in Table 1, ZM851 FT-IR analysis revealed significant changes in the infrared characteristic peaks of the osimertinib / dihydroartemisinin cocrystal compared to the active pharmaceutical ingredient (API). Specifically, some characteristic peaks of the API showed shifts, decreased intensity, and morphological changes. For example, the amide spectrum characteristic peak of the osimertinib API was located at 745.78 cm⁻¹. -1 (γ N-H ) and 1533.96cm -1 (δ N-H ), which shifted to 733.81 cm in the eutectic. -1 (γ N-H ) and 1571.71cm -1 (δ N-H Similarly, the characteristic peak of the -OH band in dihydroartemisinin is at 3380.76 cm⁻¹. -1( ν O-H ) and 661.99cm -1 (δ O-H At ), while in eutectic molecules, these characteristic peaks shift to lower frequencies to 3375.97 cm⁻¹. -1 (ν O-H ) and 594.96cm -1 (δ O-H The reason for this change is that after the eutectic is formed, the -NH in osimertinib forms a strong NH…O hydrogen bond with the -OH in the ligand, which leads to a decrease in the vibrational frequency of -OH and -NH, a broadening of the spectral band, and consequently a red shift of the characteristic peaks of the active pharmaceutical ingredient to varying degrees.

[0064] Table 1

[0065] Belonging <![CDATA[Osimertinib (cm -1 )]]> <![CDATA[Dihydroartemisinin (cm -1 )]]> <![CDATA[ZM851(cm -1 )]]> <![CDATA[v N-H ]]> 3428.64 - - <![CDATA[v O-H ]]> - 3380.76 3375.97 (Peak becomes blunt) vC=0 1576.49 - 1676.24 <![CDATA[δ N-H ]]> 1533.96 - 1571.71 <![CDATA[δ O-H ]]> - 661.99 594.96 <![CDATA[γ N-H ]]> 745.78 - 733.81

[0066] like Figure 4As shown in Table 2, the Raman spectra of ZM851 revealed that the Raman characteristic peaks of the osimertinib / dihydroartemisinin cocrystal molecule were significantly shifted, weakened, or even disappeared compared to the active pharmaceutical ingredient (API). Specifically, the amide characteristic peak of the osimertinib API was located at 3205.13 cm⁻¹. -1 (ν N-H ), 1621.07cm -1 (ν C＝O ), 989.06cm -1 (γ N-H ) and 1555.4cm -1 (δ N-H In eutectic molecules, these characteristic absorption peaks shift to lower frequencies, reaching 3102.13 cm⁻¹. -1 (ν N- H), 1576.94cm -1 (ν C＝O ), 976.54cm -1 (γ N-H ) and 1513.97cm -1 (δ N-H This phenomenon is caused by the formation of a eutectic, where the -NH in osimertinib forms a strong NH…O hydrogen bond with the -OH in the ligand, leading to a decrease in the vibrational frequency of -OH and -NH, which in turn causes the characteristic peaks of the active pharmaceutical ingredient to undergo varying degrees of red shift.

[0067] Table 2

[0068] Belonging <![CDATA[Osimertinib (cm -1 )]]> <![CDATA[Dihydroartemisinin (cm -1 )]]> <![CDATA[ZM851(cm -1 )]]> <![CDATA[ν N-H ]]> 3205.13 - 3102.13 <![CDATA[ν O-H ]]> - 3245.76 - <![CDATA[v C-N ]]> 1353.63 - 1400.89 <![CDATA[δ C-N ]]> 1538.3 - 1464.09 <![CDATA[δ N-H ]]> 1555.4 - 1513.97 <![CDATA[v C＝O ]]> 1621.07 - 1576.94 <![CDATA[γ O-H ]]> - 722.22 - <![CDATA[γ N-H ]]> 989.06 - 976.54

[0069] Furthermore, the ZM851's thermogravimetric-differential scanning calorimetry (TG-DSC) is as follows: Figure 5 The four-step weight loss behavior of TGA is revealed. The initial weight loss is likely due to the release of water molecules. The second weight loss is attributed to the decomposition of dihydroartemisinin. After losing the cocrystal formation, the osimertinib molecules in osimertinib-dihydroartemisinin begin to decompose upon further heating. The DSC curve shows four endothermic peaks in ZM851, located at 63℃, 83℃, and 142℃. The first two peaks correspond to the endothermic process of solvent and water removal from the cocrystal, while the peak at 142℃ corresponds to the melting peak of osimertinib-dihydroartemisinin. The four weight loss steps in thermogravimetric analysis (TG) indicate that the osimertinib-dihydroartemisinin cocrystal corresponds to the DSC curve. Based on the above analysis, ZM851 is a cocrystal with an API:CCF ratio of 1:1 and a melting point of 142℃.

[0070] 2. Structural Characterization and Analysis of ZM852

[0071] like Figure 6As shown, the 1H NMR spectrum of the ZM852 eutectic exhibits the following data: 1H NMR (500MHz, DMSO-d6) δ 10.20 (s, 1H), 9.15 (s, 1H), 8.6 (s, 1H), 8.33 (d, J = 5.3Hz, 1H), 8.24 (d, J = 8.0Hz, 1H), 7.89 (s, 1H), 7.52 (d, J = 8.2Hz, 1H), 7.26–7.21 (m, 2H), 7.18–7.12 (m, 1H) The values ​​are 7.04 (s, 1H), 6.47–6.40 (m, 1H), 6.30–6.24 (m, 1H), 5.79–5.75 (m, 1H), 3.91 (s, 3H), 3.86 (s, 3H), 2.89 (t, J = 5.9 Hz, 2H), 2.72 (s, 3H), 2.30 (s, 2H), and 2.21 (s, 6H). These data correspond to the proton NMR spectrum of osimertinib. The ¹H NMR (500MHz, DMSO-d6) δ9.55(s, ¹H), 8.50(s, ¹H), 8.13–8.10(m, ¹H), 7.82–7.77(m, 2H), 7.44(t, J = 9.1Hz, ¹H), 7.20(s, ¹H), 4.18(t, J = 6.3Hz, 2H), 3.94(s, 3H), 3.58(t, J = 4.6Hz, 4H), 2.47(d, J = 7.0Hz, 2H), 2.39(s, 4H), and 1.99(m, J = 6.6Hz, 2H) correspond to the ¹H NMR spectrum of gefitinib. Calculations revealed that the molar ratio of API (active pharmaceutical ingredient) to CCF (cocrystallizing agent) in the cocrystallized molecule is 1:1.

[0072] The X-ray diffraction (XRD) pattern of ZM852 is as follows: Figure 7 As shown, the X-ray diffraction patterns of osimertinib exhibit characteristic peaks at 2θ angles of 10.34°, 15.88°, 16.6°, 17.58°, 20.44°, 20.82°, 22.74°, 23.14°, 24.1°, 25.98°, 26.54°, and 27.3°. The X-ray diffraction patterns of gefitinib exhibit characteristic peaks at 2θ angles of 15.8°, 17.62°, 18.6°, 19.3°, 20.6°, 22.44°, 24°, and 26.34°. The X-ray diffraction patterns of ZM852 exhibit characteristic peaks at 2θ angles of 10.4°, 20.52°, 20.9°, 23.24°, 24.26°, and 26.36°. Since no obvious new characteristic peaks appeared in the product, it is impossible to determine whether a new crystal phase has been formed.

[0073] like Figure 8As shown in Table 3, ZM852 FT-IR analysis revealed that the characteristic peaks of the osimertinib / gefitinib cocrystal molecules underwent significant shifts, intensities, and changes compared to the active pharmaceutical ingredient (API). Specifically, the amide characteristic peak of the osimertinib API was located at 1533.96 cm⁻¹ (δ). N-H ) and 1576.49cm -1 (ν C＝O (Stretching vibrations), while in eutectic molecules, these characteristic absorption peaks shift to the high-frequency region to 1622.59 cm⁻¹. -1 (δ N-H ) and 1669.51cm -1 (ν C＝O Similarly, the secondary amine band of gefitinib API appeared at 1575.67 cm⁻¹. -1 (δ N-H ) and 1630.41cm -1 (ν C＝O In eutectic molecules, these characteristic absorption peaks also shift to higher frequencies, reaching 1622.59 cm⁻¹. -1 (δ N-H ) and 1669.51cm -1 (ν C＝O This change is attributed to the hydrogen bonding that occurs after the formation of the eutectic, which causes the characteristic peaks of the active pharmaceutical ingredient to shift to varying degrees.

[0074] Table 3

[0075] Belonging <![CDATA[Osimertinib (cm -1 )]]> <![CDATA[Gefitinib (cm -1 )]]> <![CDATA[ZM852(cm -1 ) <!-- 6 -->]]> <![CDATA[v N-H ]]> 3428.64 3397.83 3431.72 <![CDATA[v O-H ]]> - - - <![CDATA[ν C＝O ]]> 1576.49 1630.41 1669.51 <![CDATA[δ N-H ]]> 1533.96 1575.67 1622.59 <![CDATA[δ O-H ]]> - - - <![CDATA[γ N-H ]]> - - -

[0076] like Figure 9 As shown in Table 4, the ZM852 Raman spectroscopy revealed significant shifts, weakening, and even disappearances in the Raman characteristic peaks of the osimertinib / gefitinib cocrystal compared to the active pharmaceutical ingredient (API). Specifically, the amide characteristic peak of the osimertinib API is located at 1538.3 cm⁻¹. -1 (δ C-N ), 1555.4cm -1 (δ N-H ), 1621.07cm -1 (ν C＝O In eutectic molecules, these characteristic absorption peaks shift to lower frequencies, reaching 1535.56 cm⁻¹. -1 (δ C-N ), 1552.67cm -1 (δ N-H ), 1612.85cm -1 (ν C＝O For gefitinib raw material, its secondary amine spectrum characteristic peak is located at 1342 cm⁻¹. -1 (ν C-N), 1616.96cm -1 (δ N-H ), 784.54cm -1 (γ N-H In the eutectic, these characteristic absorption peaks red-shift to 1340.62 cm⁻¹. -1 (ν C-N ), 1552.67cm -1 (δ N-H ), 774.28cm -1 (γ N-H This phenomenon is caused by the fact that after the eutectic is formed, the -C=O in osimertinib forms a strong NH…O hydrogen bond with one of the -NH in the ligand, which leads to a decrease in the vibrational frequency of -C=O and -NH, and thus causes the characteristic peaks of the active pharmaceutical ingredient to redshift to varying degrees.

[0077] Table 4

[0078] Belonging <![CDATA[Osimertinib (cm -1 )]]> <![CDATA[Gefitinib (cm -1 )]]> <![CDATA[ZM852(cm -1 )]]> <![CDATA[v N-H ]]> - - - <![CDATA[v O-H ]]> - - - <![CDATA[v C-N ]]> 1353.63 1342 1340.62 <![CDATA[δX C-N ]]> 1538.3 - 1535.56 <![CDATA[δ N-H ]]> 1555.4 1616.96 1552.67 vc=o 1621.07 - 1612.85 <![CDATA[γ N-H ]]> 989.06 784.54 774.28

[0079] like Figure 10 As shown, TG-DSC analysis of ZM852 revealed a two-stage weight loss process. The first step corresponds to the desolventizing and dehydrating processes, represented by exothermic peaks at 86.3℃ and 106℃ in the DSC spectrum, respectively. The second step may be related to the decomposition of osimertinib in ZM852, with an exothermic peak appearing at 169℃. Furthermore, the endothermic peak at 351℃ indicates the melting process of ZM852. Comprehensive analysis suggests that ZM852 did not form a eutectic.

[0080] 3. Structural Characterization and Analysis of ZM853

[0081] like Figure 11As shown, the 1H NMR spectrum of the ZM853 eutectic presents the 1H NMR data for osimertinib and ibrutinib. The 1H NMR spectrum of osimertinib... NMR (500MHz, DMSO-d6) data are as follows: δ 10.21 (s, 1H), 9.16 (s, 1H), 8.68 (s, 1H), 8.33 (d, J = 5.3Hz, 1H), 7.90 (s, 1H), 7.52 (d, J = 8.2Hz, 1H), 7.26–7.22 (m, 2H), 7.04 (s, 1H), 6.46–6.40 (m, 1H), 6.28 (d, J = 15.8Hz, 1H), 5.79–5.75 (m, 1H), 3.92 (s, 3H), 3.86 (s, 3H), 2.89 (t, J = 5.8Hz, 2H), 2.72 (s, 3H), 2.29 (t, J = 5.9Hz, 3H), 2.21 (s, 6H). Ibrutinib 1H NMR (500MHz, DMSO-d6) data are as follows: δ 7.66 (t, J = 6.8Hz, 2H), 7.43 (t, J = 7.8Hz, 2H), 6.89–6.83 (m, 1H), 6.75–6.68 (m, 1H), 6.17–6.03 (m, 1H), 5.71 (d, J = 10.5Hz, 1H), 4.70 (d, J = 23.5Hz, 1H), 4.56 (d, J = 12.6Hz, 1H), 4.19 (d, J = 12.2Hz, 1H), 4.07 (d, J = 13.7Hz, 1H), 3.20 (q, J = 14.0, 13.2Hz, 1H), 2.13 (s, 1H), 1.92 (s, 1H), 1.59 (s, 1H). In the region 8.28–8.21 (m, 2H), the two hydrogen atoms belong to osimertinib and ibrutinib, respectively. In the region 7.21–7.10 (m, 7H), one of the seven hydrogen atoms comes from osimertinib, and the remaining six come from ibrutinib. Calculations show that the molar ratio of API to CCF in the eutectic molecule is 1:1.

[0082] like Figure 12As shown, in the X-ray diffraction pattern of ZM853, the characteristic peaks of osimertinib at the 2θ angle appear at 10.34°, 15.88°, 16.6°, 17.58°, 20.44°, 20.82°, 22.74°, 23.14°, 24.1°, 25.98°, 26.54°, and 27.3°. The characteristic peaks of irutinib at the 2θ angle appear at 10.58°, 16.02°, 18.22°, 18.9°, 19.72°, 20.46°, 21.22°, 21.64°, 22.32°, 23.44°, and 26.44°. The X-ray diffraction pattern of ZM853 shows characteristic peaks at the 2θ angle at 18.56°, 19.86°, 20.6°, 20.98°, 21.36°, 26.1°, and 27.48°. No obvious new characteristic peaks were observed in the product, therefore it is impossible to determine whether a new crystal phase has formed.

[0083] Fourier transform infrared (FT-IR) data of ZM853 are as follows: Figure 13 As shown in Table 5, the infrared characteristic peaks of the osimertinib / ibrutinib cocrystal molecules showed significant shifts, weakening, and changes compared to the active pharmaceutical ingredient (API). For example, the amide spectrum characteristic peak of the osimertinib API is located at 745.78 cm⁻¹. -1 (γ N-H ) and 1576.49cm -1 (ν C＝O In eutectic molecules, these characteristic absorption peaks shift to lower frequencies, reaching 611.72 cm⁻¹. -1 (γ N-H ) and 1636.34cm -1 (ν C＝O The characteristic peak of ilutinib is 618.10 cm⁻¹. -1 (γ N-H ) and 1620.38cm -1 (δ N-H In the eutectic, these characteristic peaks shift to lower frequencies, reaching 611.72 cm⁻¹. -1 (γ N-H ) and 1614.00cm -1 (δ N-H This phenomenon is caused by the formation of hydrogen bonds, which leads to a decrease in vibrational frequency, resulting in a redshift of the characteristic peaks of the active pharmaceutical ingredient to varying degrees.

[0084] Table 5

[0085] Belonging <![CDATA[Osimertinib (cm -1 )]]> <![CDATA[Ibrutinib (cm -1 )]]> <![CDATA[ZM853(cm -1 )]]> <![CDATA[v N-H ]]> 3428.64 - - <![CDATA[v O-H ]]> - - - <![CDATA[ν C＝O ]]> 1576.49 1639.53 1636.34 <![CDATA[δ N-H ]]> 1533.96 1620.38 1614.00 <![CDATA[δ O-H ]]> - - - <![CDATA[γ N-H ]]> 745.78 618.10 611.72

[0086] like Figure 14As shown in Table 6, ZM853 Raman data revealed significant shifts, weakening, and changes in the Raman characteristic peaks of the osimertinib / ibrutinib cocrystal compared to the active pharmaceutical ingredient (API). Specifically, the amide characteristic peak in the osimertinib API was located at 1538.3 cm⁻¹. -1 (δ C-N ) and 1555.4cm -1 (δ N-H In eutectic molecules, these characteristic absorption peaks shift to higher frequencies, reaching 1554.13 cm⁻¹. -1 (δ C-N ) and 1576.01cm -1 (δ N-H Furthermore, the C=O absorption peak in osimertinib is located at 1621.07 cm⁻¹. -1 (ν C＝O In the eutectic, the characteristic peak of C=O is at 1612.85 cm⁻¹. -1 (ν C＝O The primary amine characteristic peak of ibrutinib API is at 1608 cm⁻¹. -1 (δ N-H In the eutectic, this characteristic absorption peak red-shifts to 1576 cm⁻¹. -1 (δ N-H The reason for this change is that after the eutectic is formed, the -C=O in osimertinib forms a strong NH…O bond with one of the -NH in the ligand.

[0087] Table 6

[0088]

[0089]

[0090] Further analysis of the ZM853 TG-DSC data (see...) Figure 15 Three endothermic peaks can be observed in the DSC plot. The peak at 104℃ represents the dehydration process of the eutectic molecules, while the peak at 293℃ corresponds to the melting peak of the eutectic molecules. The TG plot shows that the weight loss is 36.56% between 293℃ and 483℃, corresponding to the melting process of ZM853 reaching its melting point. The weight loss before 104℃ in the TG plot is attributed to the removal of water molecules from the eutectic molecules.

[0091] Based on the above analysis, it can be concluded that ZM853 did not form a eutectic.

[0092] 5. Summary

[0093] This invention utilizes drug cocrystallization technology to successfully design and synthesize three compounds. Through a series of characterization techniques, including 1H NMR, XRD, FT-IR, Raman, and TG-DSC analysis, we identified one molecule, ZM851, that forms a cocrystallization with osimertinib. Specifically, the molar ratio of the active pharmaceutical ingredient (API) to the cocrystallizing agent (CCF) in the ZM851 cocrystallization molecule is 1:1.

[0094] Example 3: In vitro antitumor activity study

[0095] 1. CCK-8 Experimental Principle

[0096] Cell Counting Kit-8 (CCK-8) is a simple and accurate method for cell proliferation and toxicity analysis. Its basic principle is based on WST-8, chemically known as 2-(2-methoxy-4-nitrophenyl)-3-(4-nitrophenyl)-5-(2,4-disulfonic acid benzene)-2H-tetrazole monosodium salt, contained in the reagent. Under the action of the electron carrier 1-methoxy-5-methylphenazineonium sulfate dimethyl ester (1-Methoxy PMS), intracellular dehydrogenases reduce WST-8 to a highly water-soluble yellow formazan product. The amount of formazan produced is directly proportional to the number of viable cells, a property that allows CCK-8 to be directly applied to cell proliferation and toxicity analysis.

[0097] 2. Experimental methods and procedures

[0098] Cell source: All cell lines used in this experiment were purchased from the Chinese Academy of Sciences. PC-9, NCI-H1975, and NCI-H1650 cells were cultured in RPMI-1640 complete medium. The RPMI-1640 complete medium used for PC-9 cells consisted of basal medium, 10% fetal bovine serum (FBS), and 1% 100 U / mL penicillin and 100 μg / mL streptomycin (P / S). The medium formulations used for NCI-H1975 and NCI-H1650 cells were the same. The complete medium for NCI-HCT116 cells consisted of Hepes-free RPMI-1640 medium, 10% fetal bovine serum, and 1% penicillin-dipeptide antibiotics. The cell cryopreservation solution consisted of 55% basal medium, 40% FBS, and 5% DMSO, and was stored in liquid nitrogen. Cells were cultured in an environment of 95% air, 37°C, and 5% CO2, ensuring freedom from mycoplasma and chlamydia contamination.

[0099] In vitro cytotoxicity assay: Cells in the logarithmic growth phase were selected and cultured at a density of 7 × 10⁶ cells per well. 3Cells were seeded at a density of [insert density here] in 96-well plates and incubated at 37°C and 5% CO2 for 24 hours to maintain normal physiological pH. After cell adhesion, different concentrations of the test compound were added to three groups of wells, with 0.1% DMSO as a control. After 72 hours, 10 μL of CCK-8 reagent was added to each well, and the cells were incubated at 37°C for 0.5 hours. The absorbance was measured at 450 nm using a microplate reader, and the 50% inhibitory concentration (IC50) was calculated using GraphPad Presm software. All data were repeated three times.

[0100] Establishment of drug-resistant cell lines: Osimertinib-resistant cell lines were established by inducing drug resistance through a progressive drug concentration method. PC-9 cell lines were used as parental cells, and their IC50 values ​​were tested. Concentrations close to this value were selected for culture. After determining the optimal starting concentration, the drug concentration was gradually increased to induce resistance in the PC-9 cell lines. The culture medium was changed regularly, and cell growth was observed under a microscope. Once the cells exhibited stable resistance, the drug concentration was appropriately increased. Through repeated changes in culture medium, passages, and increases in drug concentration, the drug resistance index of the cell line to osimertinib exceeded 15, indicating successful construction of drug resistance. Successfully constructed drug-resistant cell lines are denoted as PC-9OR.

[0101] The IC50 test is used to assess the toxicity and tolerability of drugs to cells, with a lower IC50 value indicating a stronger inhibitory effect. In this paper, we assessed the sensitivity of cells to osimertinib by calculating the resistance index (RI). The resistance index (RI) is calculated by dividing the IC50 value of resistant cells by the IC50 value of parental cells. Based on the RI value, resistance is graded as follows: 1-5 for low resistance; 5-15 for moderate resistance; and above 15 for high resistance. We tested the activity of five cell lines, including PC-9, PC-9OR (osimertinib acquired resistance), H1975 (osimertinib highly sensitive cell line), H1650 (osimertinib inherently resistant cell line), and HCT116 (human colon cancer cell line).

[0102] Table 7

[0103]

[0104]

[0105] Table 8

[0106]

[0107] Table 9

[0108]

[0109] We investigated the in vitro antitumor activity of one newly synthesized cocrystal molecule and two non-cocrystal compounds. The results showed that osimertinib significantly reduced the sensitivity of the resistant PC-9OR cell line compared to PC-9, with IC50 values ​​of 0.0283±0.0075 μM and 0.7232±0.2166 μM, respectively, and a resistance index of 25.6, a statistically significant difference (P<0.01). Further comparison revealed that compound ZM851 exhibited significantly higher antitumor activity than osimertinib, with an IC50 value of 0.1237±0.0359 μM, also showing a statistically significant difference (P<0.01). In the H1975 cell line, the IC50 of osimertinib was 3.4837±1.4693 μM, while the cocrystal molecule ZM851 showed superior antitumor activity. For the inherently resistant H1650 cell line, osimertinib's IC50 was 4.8187 ± 0.1301 μM, and the cocrystal molecule ZM851 also exhibited superior antitumor activity. In the HCT116 human colon cancer cell line, osimertinib's IC50 was 0.4954 ± 0.1443 μM, while the antitumor activity of the cocrystal molecule ZM851 was enhanced. In the HCT116 cell line, the antitumor activity of ZM851 was 1.2 times that of osimertinib. Compared with ZM851, the mixture showed approximately 26.1-fold lower activity against the resistant PC-9OR cell line. These results indicate that the osimertinib cocrystal molecule exhibits superior activity against the resistant PC-9OR cell line, preliminarily revealing the potential advantages of cocrystal molecules in antitumor activity.

[0110] 3. Summary

[0111] This invention evaluated the in vitro anti-non-small cell lung cancer activity of one cocrystal molecule and two non-cocrystal compounds. The results showed that the cocrystal molecule ZM851 exhibited approximately 5.85-fold enhanced activity against the acquired resistance cell line PC-9OR compared to osimertinib. For the acquired resistance cell line PC-9OR, the antitumor activity of the cocrystal molecule ZM851 was more than 10 times higher than that of its non-cocrystal mixture. This preliminarily demonstrates that novel antitumor drugs can be developed to address osimertinib resistance through drug cocrystallization technology.

[0112] Finally, it should be noted that the above embodiments are only used to illustrate the technical solutions of the present invention and not to limit it. Although the present invention has been described in detail with reference to the above embodiments, those skilled in the art should understand that modifications or equivalent substitutions can still be made to the specific implementation of the present invention. Any modifications or equivalent substitutions that do not depart from the spirit and scope of the present invention should be covered within the scope of protection of the claims of the present invention.

Claims

1. A compound ZM851, characterized in that, The chemical structural formula of the compound ZM851 is as follows: ； The X-ray diffraction pattern of the compound ZM851 shows characteristic peaks at 2θ angles of 5.88°, 15.06°, 18.36°, 24.62°, and 25.26°. The preparation method of the compound ZM851 includes: mixing osimertinib, dihydroartemisinin and a mixed solvent, heating and stirring to dissolve, then adding n-hexane, and the precipitated yellow solid is the compound ZM851; The mixed solvent is acetone and water; The volume ratio of acetone to water is 15:1; The temperature at which osimertinib, dihydroartemisinin, and the mixed solvent are heated and stirred to dissolve them is at least 60°C.

2. A method for preparing compound ZM851 according to claim 1, characterized in that, include: Osimertinib, dihydroartemisinin and a mixed solvent were mixed, heated and stirred to dissolve, and then n-hexane was added. The precipitated yellow solid was the compound ZM851. The mixed solvent is acetone and water; The volume ratio of acetone to water is 15:1; The temperature at which osimertinib, dihydroartemisinin, and the mixed solvent are heated and stirred to dissolve them is at least 60°C.

3. An application of the compound ZM851 according to claim 1, characterized in that, The application includes at least one of the following: A1) Applications in the preparation of antitumor drugs; A2) Use in the preparation of pharmaceutical compositions for the prevention or treatment of drug-resistant tumors; A3) Application in the preparation of drug resistance sensitizers for antitumor drugs; The tumor is either adenocarcinoma of the lung or colon cancer cells.

4. A eutectic drug, characterized in that, The active ingredient of the eutectic drug is compound ZM851 as described in claim 1.

5. The application of the eutectic drug according to claim 4, characterized in that, The application includes at least one of the following: A1) Applications in the preparation of antitumor drugs; A2) Use in the preparation of pharmaceutical compositions for the prevention or treatment of drug-resistant tumors; A3) Application in the preparation of drug resistance sensitizers for antitumor drugs; The tumor is either adenocarcinoma of the lung or colon cancer cells.

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