A co-crystal drug ZM849 and its application in resisting lung cancer

Abstract

The application relates to the technical field of biological medicine, and discloses a co-crystal drug ZM849 and application thereof in resisting lung cancer, and a preparation method of the co-crystal drug ZM849, which comprises the following steps: mixing osimertinib, L-proline and a mixed solvent, heating, stirring and dissolving, and then adding n-hexane; and yellow solid separated out is the co-crystal drug ZM849. The anti-tumor activity of the co-crystal molecule ZM849 is more than ten times that of a non-co-crystal mixture. In addition, in an in-vitro experiment, the ZM849 shows a significant inhibiting effect on an osimertinib-resistant cell strain; the co-crystal drug ZM849 and the preparation method thereof bring a new strategy to the field of lung cancer treatment, and are expected to play a key role in future clinical practice.

Description

Technical Field

[0001] This invention relates to the field of biomedical technology, and more specifically, to a eutectic drug ZM849 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 ZM849 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 ZM849, the chemical structural formula of which is as follows:

[0006] .

[0007] The present invention also provides a method for preparing the above-mentioned compound ZM849, comprising:

[0008] Osimertinib, L-proline, and a mixed solvent were mixed, heated, and stirred to dissolve. Then, n-hexane was added, and the precipitated yellow solid was the compound ZM849.

[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, L-proline, and the mixed solvent are heated and stirred to dissolve is at least 60°C.

[0012] The present invention also provides an application of the above-mentioned compound ZM849, the application comprising 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 ZM849.

[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 ZM849 exhibited more than 10 times the antitumor activity of its non-co-crystal mixture. Furthermore, in in vitro experiments, ZM849 showed a significant inhibitory effect on osimertinib-resistant cell lines.

[0023] The significant inhibitory effect of the cocrystallized drug ZM849 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 ZM849 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 ZM849 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 ZM8491 HNMR spectrum;

[0027] Figure 2 The XRD pattern of compound ZM849 is shown below.

[0028] Figure 3 The Fourier transform infrared spectrum of compound ZM849 is shown below.

[0029] Figure 4 The Raman spectrum of compound ZM849;

[0030] Figure 5 The TG and DSC spectra of compound ZM849 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 ZM849

[0044] Weigh osimertinib (200 mg, 0.4 mmol) and L-proline (92 mg, 0.8 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 268 mg of yellow powder, namely compound ZM849, 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 ZM849, 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 ZM849

[0061] like Figure 1 As shown, the 1H NMR spectrum of ZM849 reveals the 1H NMR data of osimertinib, specifically: 1H NMR (500MHz, DMSO-d6) δ 10.22 (s, 1H), 9.15 (s, 1H), 8.68 (s, 1H), 8.33 (d, J = 5.3Hz, 1H), 8.25 (d, J = 8.0Hz, 1H), 7.91 (s, 1H), 7.53 (d, J = 8.2Hz, 1H), 7.25 (t, J = 7.2Hz). ,2H),7.15(t,J=7.5Hz,1H),7.05(s,1H),6.49-6.39(m,1H),6.28(d,J=16.9Hz,1H),5.78(d,J=10.0Hz ,1H),3.92(s,3H),3.86(s,3H),2.90(t,J=5.8Hz,2H),2.72(s,3H),2.30(t,J=5.8Hz,2H),2.22(s,6H).

[0062] In addition, the 1H NMR data of L-proline were 1H NMR (500MHz, DMSO-d6) δ 3.66-3.61 (m, 1H), 3.26-3.15 (m, 1H), 3.06-2.97 (m, 2H), 2.09-1.89 (m, 3H), 1.83-1.63 (m, 3H). The molar ratio of API:CCF in the eutectic molecule was calculated to be 1:1.

[0063] The X-ray diffraction pattern (XRD pattern) of ZM849 is as follows: Figure 2As shown, osimertinib exhibits 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°. L-proline shows characteristic peaks at 2θ angles of 15.22°, 18.12°, and 24.8°. ZM849 shows new characteristic peaks at 2θ angles of 5.84°, 7.3°, 11.06°, 12.8°, 14.22°, 17.94°, 18.92°, and 19.4°, which preliminarily indicates the formation of a new crystal phase.

[0064] Fourier transform infrared (FT-IR) data of ZM849 are as follows: Figure 3 As shown in Table 1, the infrared characteristic peaks of the osimertinib / L-proline cocrystal molecule showed significant shifts, weakening, and changes compared to the active pharmaceutical ingredient (API). The amide characteristic peak of the osimertinib API includes 3428.64 cm⁻¹. -1 (ν N-H ), 1576.49cm -1 (ν C＝O ), 1533.96cm -1 (δ N-H ), 745.78cm -1 (γ N-H In eutectic molecules, these characteristic absorption peaks shift to lower frequencies, reaching 3410.86 cm⁻¹. -1 (ν N-H ), 1567.85cm -1 (ν C＝O ), 1513.11cm -1 (δ N-H ), 582.47cm -1 (γ N-H The characteristic peak of L-proline is 1622.59 cm⁻¹. -1 (ν C＝O ), 1560.03cm-1 (δ N-H ), 642.43cm-1(γ N-H In eutectic molecules, these characteristic absorption peaks also shift to lower frequencies, reaching 1567.85 cm⁻¹. -1 (ν C＝O ), 1513.11cm -1 (δ N-H ), 582.47cm -1 (γ N-HThis phenomenon occurs because, after the eutectic is formed, the -NH in osimertinib forms a strong NH…O bond with one of the -C=O ligands, which leads to a decrease in the vibrational frequencies of -C=O and -NH, resulting in a red shift of the characteristic peaks of the active pharmaceutical ingredient to varying degrees.

[0065] Table 1

[0066] Belonging <![CDATA[Osimertinib (cm -1 )]]> <![CDATA[L-proline (cm -1 )]]> <![CDATA[ZM849(cm -1 )]]> <![CDATA[ν N-H ]]> 3428.64 - 3410.86 <![CDATA[ν O-H ]]> - - - <![CDATA[ν C＝O ]]> 1576.49 1622.59 1567.85 <![CDATA[δ N-H ]]> 1533.96 1560.03 1513.11 <![CDATA[δ O-H ]]> - - - <![CDATA[γ N-H ]]> 745.78 642.43 582.47

[0067] like Figure 4 As shown in Table 2, the ZM849 Raman data revealed that the Raman characteristic peaks of the osimertinib / L-proline cocrystal molecule underwent significant shifts, intensity reductions, and even the disappearance of some peaks 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 ), 989.06cm -1 (γ N-H In eutectic molecules, these characteristic absorption peaks shift to lower frequencies, reaching 1468.6 cm⁻¹. -1 (δ C-N ), 1509.99cm -1 (δ N-H ), 1611.56cm -1 (ν C＝O The reason for this change may be that after the formation of the eutectic, the -NH group in osimertinib forms a strong NH…O hydrogen bond with the -C=O group in the ligand, which leads to a decrease in the vibrational frequencies of -C=O and -NH, and thus causes the characteristic peaks of the active pharmaceutical ingredient to redshift to varying degrees.

[0068] Table 2

[0069]

[0070]

[0071] like Figure 5As shown in the TG-DSC data, ZM849 exhibits five endothermic peaks. Two of these peaks, located between 88.8℃ and 106℃, represent the water loss process, while the other two peaks, between 209℃ and 224℃, correspond to the eutectic transformation process. Furthermore, the endothermic peak at 366℃ marks the melting peak, consistent with the three weight loss stages in the TG curve. In summary, ZM849 is a eutectic material with a eutectic molecular ratio of API:CCF = 1:1 and a melting point range of 209℃ to 224℃.

[0072] 2. Structural Characterization and Analysis of ZM852

[0073] like Figure 6 As 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.

[0074] The X-ray diffraction (XRD) pattern of ZM852 is as follows: Figure 7As 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.

[0075] like Figure 8 As 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.

[0076] Table 3

[0077] Belonging <![CDATA[Osimertinib (cm -1 )]]> <![CDATA[Gefitinib (cm -1 )]]> <![CDATA[ZM852(cm -1 )]]> <![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 ]]> - - -

[0078] like Figure 9As 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.

[0079] Table 4

[0080]

[0081]

[0082] 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.

[0083] 3. Structural Characterization and Analysis of ZM853

[0084] like Figure 11 As shown, the 1H NMR spectrum of the ZM853 eutectic presents the 1H NMR data of 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.

[0085] 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.

[0086] 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.

[0087] Table 5

[0088] 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

[0089] 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.

[0090] Table 6

[0091]

[0092] 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.

[0093] Based on the above analysis, it can be concluded that ZM853 did not form a eutectic.

[0094] 5. Summary

[0095] 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, ZM849, that forms a cocrystallization with osimertinib. Specifically, the molar ratio of the active pharmaceutical ingredient (API) to the cocrystallizing agent (CCF) in the ZM849 cocrystallization molecule is 1:1.

[0096] Example 3: In vitro antitumor activity study

[0097] 1. CCK-8 Experimental Principle

[0098] 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.

[0099] 2. Experimental methods and procedures

[0100] 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.

[0101] 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.

[0102] 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.

[0103] 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).

[0104] Table 7 Table 8

[0105]

[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 ZM849 exhibited significantly higher antitumor activity than osimertinib, with an IC50 value of 0.1252±0.0175 μM. In the H1975 cell line, osimertinib had an IC50 of 3.4837±1.4693 μM, while the cocrystal molecule ZM849 showed superior antitumor activity. For the inherently resistant H1650 cell line, osimertinib's IC50 was 4.8187 ± 0.1301 μM, and the cocrystal molecule ZM849 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 ZM849 was enhanced. In the HCT116 cell line, the antitumor activity of ZM849 was 1.1 times that of osimertinib. Compared with ZM849, the mixture showed approximately 25.5 times 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 co-crystal molecule and two non-co-crystal compounds. The results showed that the co-crystal molecule ZM849 exhibited approximately 5.78-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 co-crystal molecule ZM849 was more than 10 times higher than that of its non-co-crystal mixture. This preliminarily demonstrates that novel antitumor drugs can be developed to address osimertinib resistance through drug co-crystallization 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 ZM849 characterized by, The chemical structural formula of the compound ZM849 is: ； The compound ZM849 is a co-crystal compound, and its X-ray diffraction pattern has characteristic peaks at 2θ angles of 5.84°, 7.3°, 11.06°, 12.8°, 14.22°, 17.94°, 18.92°, and 19.4°.

2. A process for the preparation of the compound ZM849 according to claim 1, characterized in that, Comprise: After mixing osimertinib, L-proline and mixed solvent, heating and stirring to dissolve, then adding n-hexane, the yellow solid precipitated is the compound ZM849.

3. The production method according to claim 2, characterized by, The mixed solvent is acetone and water.

4. The production method according to claim 3, characterized by, The volume ratio of the acetone and water is 15:

1.

5. The preparation method according to claim 2, characterized in that, The temperature when mixing osimertinib, L-proline and mixed solvent, heating and stirring to dissolve is at least 60°C.

6. Use of a compound of claim 1, ZM849, wherein, The application at least includes any one of the following: A1) the application in preparing an anti-lung cancer drug; A2) the application in preparing a drug composition for preventing or treating drug-resistant lung cancer; A3) the application in preparing a drug resistance sensitizing agent for an anti-lung cancer drug.

7. A co-crystal pharmaceutical, characterized in that, The active ingredient of the co-crystal drug is the compound ZM849 of claim 1.

8. Use of the co-crystal drug of claim 7, characterized in that, The application at least includes any one of the following: A1) the application in preparing an anti-lung cancer drug; A2) the application in preparing a drug composition for preventing or treating drug-resistant lung cancer; A3) the application in preparing a drug resistance sensitizing agent for an anti-lung cancer drug.

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