Pharmaceutically acceptable salts of mor receptor agonists, polymorphs thereof and uses thereof

Abstract

Disclosed are pharmaceutically acceptable salts of MOR receptor agonists, polymorphs thereof and uses thereof. Specifically, disclosed are pharmaceutically acceptable salts of (4S,6S)-6-isopropyl-N-(2-((R)-9-(pyridin-2-yl)-6-oxaspiro[4.5]dec-9-yl)ethyl)-5,6-dihydro-4H-pyrrolo[1,2-b]pyrazol-4-amine, polymorphs thereof and uses thereof. In addition, disclosed are pharmaceutical compositions comprising the pharmaceutically acceptable salts of the compound or polymorphs thereof and uses thereof.

Description

[0001] Cross-references to related applications

[0002] This application claims priority to Chinese Patent Application No. 2021107887124, filed on July 13, 2021, entitled “Pharmaceutically Acceptable Salts of MOR Receptor Agonists, Polymorphs thereof and Uses thereof”, the entire contents of which are incorporated herein by reference. Technical Field

[0003] This application belongs to the field of pharmaceutical technology, and more specifically, this application relates to a pharmaceutically acceptable salt of a MOR receptor agonist, its polymorphs, and its uses. Background Technology

[0004] Opioid receptors are an important class of G protein-coupled receptors (GPCRs) and are targets for the binding of endogenous opioid peptides and opioid drugs. Endogenous opioid peptides are naturally occurring opioid-like active substances in mammals. Currently known endogenous opioid peptides are broadly classified into enkephalins, endorphins, dynorphins, and neophorphins. Corresponding opioid receptors exist in the central nervous system, including μ(MOR), δ(DOR), and κ(KOR) receptors. Studies have found that the strength of the analgesic effect of endogenous opioid peptides mainly depends on the expression level of opioid receptors, which are the targets of opioid drugs and the analgesic effects of endogenous opioid peptides.

[0005] Current research suggests that GPCRs mediate and regulate physiological functions primarily through two pathways: the G protein pathway and the β-arrestin pathway. Traditional GPCR agonists, upon binding to receptors, activate G protein signaling pathways, including second messenger systems such as calcium ions, adenylate cyclase (AC), and mitogen-activated protein kinases (MAPKs). β-arrestin-preferred ligands, however, primarily activate the β-arrestin pathway. β-arrestin-mediated GPCR responses involve three main aspects: 1) acting as a negative regulator, interacting with G protein-coupled receptor kinases (GRKs) to induce receptor desensitization in GPCRs, thus halting G protein signal transduction; 2) acting as a scaffold protein, recruiting endocytic proteins and inducing GPCR endocytosis; and 3) acting as an adaptor protein, forming complexes with downstream GPCR signaling molecules to activate signal transduction molecules such as MAPK, Src protein tyrosine kinase, and Akt in a G protein-independent manner. The differences in ligand-stimulated G protein signaling and / or β-arrestin signaling ultimately determine the ligand-specific cellular biological effects of GPCRs.

[0006] MOR is the target of endogenous enkephalins and opioid analgesics such as morphine. Early studies showed that endogenous enkephalins and the opioid etorphine could stimulate G proteins and induce receptor endocytosis, but morphine did not induce receptor endocytosis at all. This is because morphine's ability to stimulate MOR phosphorylation is too weak, and it can only recruit trace amounts of β-arrestin to the membrane (Zhang et al., Proc Natl Acad Sci USA, 1998, 95(12):7157-7162). Such ligands exert their physiological functions entirely through the G protein signaling pathway rather than the β-arrestin pathway. Studies have found that after injecting morphine into β-arrestin2 gene knockout mice, the analgesic effect mediated by G protein signaling was stronger and lasted longer (Bohn et al., Science, 1999). It is evident that if such ligands exhibit a stronger negative β-arrestin preference, they may even escape β-arrestin-mediated receptor desensitization, leading to prolonged G protein signaling time and a more potent analgesic effect.

[0007] Currently published patent applications for MOR agonists include WO2017106547, WO2017063509, WO2012129495, and WO2017106306.

[0008] Long-term use of opioids can lead to tolerance and side effects such as respiratory depression and constipation, which have been shown to be closely related to the function of β-arrestin. To reduce the side effects of opioids, drugs can be designed based on the negative β-arrestin-preferred ligands of MOR, thereby reducing β-arrestin-mediated side effects and enhancing therapeutic efficacy.

[0009] Given the aforementioned background, and considering the unsatisfactory properties of the free base form of the already developed MOR receptor agonists, it is necessary to further develop pharmaceutically acceptable salts and polymorphs of MOR receptor agonists to facilitate further drug development. Summary of the Invention

[0010] In a first aspect of this application, a pharmaceutically acceptable salt of formula X and a polymorph of the pharmaceutically acceptable salt of formula X are provided:

[0011]

[0012] The pharmaceutically acceptable salt is selected from maleate, L-tartrate and succinate.

[0013] In some embodiments, the pharmaceutically acceptable salt of the compound of formula X and the polymorph of the pharmaceutically acceptable salt of the compound of formula X are in anhydrous form, hydrate form or solvate form.

[0014] In some embodiments, the polymorph is a type A crystal of maleate of formula X, i.e., crystal form A, whose X-ray powder diffraction pattern has peaks at at least the following diffraction angles 2θ (°): 10.06±0.2, 10.45±0.2, 11.13±0.2, 13.03±0.2, 16.81±0.2, 17.29±0.2, 19.45±0.2, 20.05±0.2, 24.04±0.2.

[0015] In some embodiments, the polymorph is a type A crystal of maleate of formula X, i.e., crystal form A, whose X-ray powder diffraction pattern has peaks at at least the following diffraction angles 2θ (°): 10.06±0.2, 10.45±0.2, 11.13±0.2, 13.03±0.2, 16.81±0.2, 17.29±0.2, 19.45±0.2, 20.05±0.2, 24.04±0.2, 37.51±0.2.

[0016] In some embodiments, the X-ray powder diffraction pattern of crystal form A further includes two or more peaks selected from the group consisting of the following: 11.98±0.2, 14.19±0.2, 15.79±0.2, 18.52±0.2, 20.95±0.2, 21.97±0.2, and 25.57±0.2.

[0017] In some embodiments, the X-ray powder diffraction pattern of crystal form A further includes two or more peaks selected from the group consisting of the following: 11.98±0.2, 14.19±0.2, 15.79±0.2, 18.52±0.2, 20.95±0.2, 21.97±0.2, 25.57±0.2, and 43.66±0.2.

[0018] In some embodiments, the X-ray powder diffraction pattern of crystal form A has peaks at the following diffraction angles 2θ (°): 10.06±0.2, 10.45±0.2, 11.13±0.2, 11.98±0.2, 13.03±0.2, 14.19±0.2, 15.79±0.2, 16.81±0.2, 17.29±0.2, 18.52±0.2, 19.45±0.2, 20.05±0.2, 20.95±0.2, 21.97±0.2, 24.04±0.2, 25.57±0.2.

[0019] In some embodiments, the X-ray powder diffraction pattern of crystal form A has peaks at the following diffraction angles 2θ (°): 10.06±0.2, 10.45±0.2, 11.13±0.2, 11.98±0.2, 13.03±0.2, 14.19±0.2, 15.79±0.2, 16.81±0.2, 17.29±0.2, 18.52±0.2, 19.45±0.2, 20.05±0.2, 20.95±0.2, 21.97±0.2, 24.04±0.2, 25.57±0.2, 37.51±0.2, 43.66±0.2.

[0020] In some embodiments, the X-ray powder diffraction pattern of crystal form A has a peak at the 2θ (°) values ​​shown in Table A1, and the relative intensities of each peak are shown in Table A1:

[0021] Table A1

[0022] 2θ(°) <![CDATA[I / I0]]> 2θ(°) <![CDATA[I / I0]]> 2θ(°) <![CDATA[I / I0]]> 1.99 W 2.23 W 7.14 W 10.06 VS 10.45 VS 11.13 S 11.98 M 13.03 S 14.19 M 14.64 W 15.13 W 15.79 M 16.81 VS 17.29 VS 18.52 M 19.45 VS 20.05 S 20.95 M 21.97 M 24.04 VS 25.57 M 26.08 W 26.91 W 27.58 W 29.32 W 29.62 W 30.25 W 30.58 W 31.09 W 32.65 W 33.73 W 35.58 W 37.51 S 38.97 W 40.91 W 41.03 W 42.63 W 43.66 M 43.96 W

[0023] In some embodiments, the X-ray powder diffraction pattern of crystal form A is substantially as follows: Figure 1 What it represents.

[0024] In some implementations, crystal form A is in anhydrous form.

[0025] In some embodiments, the crystal form A further has one or more features selected from the group consisting of:

[0026] (i) In the differential scanning calorimetry (DSC) spectrum, the peak temperature is 177.62 ± 2 °C; in some embodiments, the DSC spectrum is basically as follows: Figure 2 What it represents;

[0027] (ii) The basic thermogravimetric analysis spectrum is as follows Figure 3 What it represents.

[0028] In some embodiments, the polymorph is a B-type crystal of the compound L-tartrate of formula X, i.e., crystal form B, whose X-ray powder diffraction pattern has peaks at at least the following diffraction angles 2θ (°): 13.82±0.2, 14.87±0.2, 20.55±0.2, and 23.15±0.2.

[0029] In some embodiments, the polymorph is a B-type crystal of the compound L-tartrate of formula X, i.e., crystal form B, whose X-ray powder diffraction pattern has peaks at at least the following diffraction angles 2θ (°): 13.82±0.2, 14.87±0.2, 20.55±0.2, 23.15±0.2, and 38.00±0.2.

[0030] In some embodiments, the X-ray powder diffraction pattern of crystal form B further includes two or more peaks with diffraction angle 2θ (°) values ​​selected from the group consisting of: 10.41±0.2, 15.75±0.2, 16.28±0.2, 17.21±0.2, 17.63±0.2, 19.98±0.2, 20.22±0.2, 21.08±0.2, 22.22±0.2, 24.56±0.2, and 28.91±0.2.

[0031] In some embodiments, the X-ray powder diffraction pattern of crystal form B further includes two or more peaks with diffraction angle 2θ (°) values ​​selected from the group consisting of: 10.41±0.2, 15.75±0.2, 16.28±0.2, 17.21±0.2, 17.63±0.2, 19.98±0.2, 20.22±0.2, 21.08±0.2, 22.22±0.2, 24.56±0.2, 28.91±0.2, and 44.21±0.2.

[0032] In some embodiments, the X-ray powder diffraction pattern of crystal form B has peaks at the following diffraction angles 2θ (°): 10.41±0.2, 13.82±0.2, 14.87±0.2, 15.75±0.2, 16.28±0.2, 17.21±0.2, 17.63±0.2, 19.98±0.2, 20.22±0.2, 20.55±0.2, 21.08±0.2, 22.22±0.2, 23.15±0.2, 24.56±0.2, 28.91±0.2.

[0033] In some embodiments, the X-ray powder diffraction pattern of crystal form B has peaks at the following diffraction angles 2θ (°): 10.41±0.2, 13.82±0.2, 14.87±0.2, 15.75±0.2, 16.28±0.2, 17.21±0.2, 17.63±0.2, 19.98±0.2, 20.22±0.2, 20.55±0.2, 21.08±0.2, 22.22±0.2, 23.15±0.2, 24.56±0.2, 28.91±0.2, 38.00±0.2, 44.21±0.2.

[0034] In some embodiments, the X-ray powder diffraction pattern of crystal form B has a peak at the 2θ (°) values ​​shown in Table B1, and the relative intensities of each peak are shown in Table B1:

[0035] Table B1

[0036] 2θ(°) <![CDATA[I / I0]]> 2θ(°) <![CDATA[I / I0]]> 2θ(°) <![CDATA[I / I0]]> 2.12 W 6.50 W 9.56 W 9.98 W 10.41 M 11.37 W 11.82 W 13.82 VS 14.87 VS 15.75 M 16.28 M 16.82 W 17.21 M 17.63 M 18.86 W 19.31 W 19.98 M 20.22 M 20.55 S 21.08 M 21.77 W 22.22 M 23.15 S 23.71 W 24.56 M 27.05 W 28.91 M 30.11 W 32.45 W 33.26 W 35.13 W 38.00 VS 41.90 W 44.21 M 48.32 W

[0037] In some embodiments, the X-ray powder diffraction pattern of crystal form B is substantially as follows: Figure 4 What it represents.

[0038] In some implementations, the crystal form B is in anhydrous form.

[0039] In some embodiments, the crystal form B further has one or more features selected from the group consisting of:

[0040] (i) In the differential scanning calorimetry (DSC) spectrum, the peak temperature is 143.31 ± 2 °C; in some embodiments, the DSC spectrum is basically as follows: Figure 5 What it represents;

[0041] (ii) The basic thermogravimetric analysis spectrum is as follows Figure 6 What it represents.

[0042] In some embodiments, the infrared spectrum of crystal form B is at approximately 3457 cm⁻¹. -1 3073cm -1 2959cm -1 2873cm -1 1716cm -1 1589cm -1 1571cm -1 1468cm -1 1440cm -1 1377cm -1 1287cm -1 1131cm -1 1105cm -1 1062cm -1 799cm -1 There is an absorption peak at that point.

[0043] In some embodiments, the infrared spectrum of crystal form B is substantially as follows: Figure 15 What it represents.

[0044] In some embodiments, the polymorph is a C-1 type crystal of the succinate of formula X, i.e., crystal form C-1, whose X-ray powder diffraction pattern has peaks at at least the following diffraction angles 2θ (°): 6.52±0.2, 10.37±0.2, 10.84±0.2, 13.66±0.2, 14.11±0.2, 15.61±0.2, 17.35±0.2, 20.92±0.2, 23.20±0.2.

[0045] In some embodiments, the polymorph is a C-1 type crystal of the succinate of formula X, i.e., crystal form C-1, whose X-ray powder diffraction pattern has peaks at at least the following diffraction angles 2θ (°): 6.52±0.2, 10.37±0.2, 10.84±0.2, 13.66±0.2, 14.11±0.2, 15.61±0.2, 17.35±0.2, 20.92±0.2, 23.20±0.2, 37.51±0.2, 43.61±0.2.

[0046] In some embodiments, the X-ray powder diffraction pattern of the crystal form C-1 also includes peaks with diffraction angle 2θ (°) values ​​selected from the group consisting of: 19.33±0.2, 25.09±0.2.

[0047] In some embodiments, the X-ray powder diffraction pattern of the crystal form C-1 has peaks at the following diffraction angles 2θ (°): 6.52±0.2, 10.37±0.2, 10.84±0.2, 13.66±0.2, 14.11±0.2, 15.61±0.2, 17.35±0.2, 19.33±0.2, 20.92±0.2, 23.20±0.2, 25.09±0.2.

[0048] In some embodiments, the X-ray powder diffraction pattern of the crystal form C-1 has peaks at the following diffraction angles 2θ (°): 6.52±0.2, 10.37±0.2, 10.84±0.2, 13.66±0.2, 14.11±0.2, 15.61±0.2, 17.35±0.2, 19.33±0.2, 20.92±0.2, 23.20±0.2, 25.09±0.2, 37.51±0.2, 43.61±0.2.

[0049] In some embodiments, the X-ray powder diffraction pattern of crystal form C-1 has a peak at the 2θ (°) values ​​shown in Table (C-1), and the relative intensities of each peak are shown in Table (C-1):

[0050] Table (C-1)

[0051] 2θ(°) <![CDATA[I / I0]]> 2θ(°) <![CDATA[I / I0]]> 2θ(°) <![CDATA[I / I0]]> 1.96 W 5.98 W 6.52 S 10.37 S 10.84 S 12.34 W 12.80 W 13.66 S 14.11 VS 14.94 W 15.61 S 17.35 S 18.37 W 19.33 M 20.03 W 20.92 S 23.20 VS 24.83 W 25.09 M 25.88 W 26.77 W 28.51 W 29.59 W 31.72 W 32.80 W 34.58 W 36.20 W 37.51 VS 38.41 W 43.61 S

[0052] In some embodiments, the X-ray powder diffraction pattern of the crystal form C-1 is substantially as follows: Figure 7 What it represents.

[0053] In some embodiments, the polymorph is a C-2 type crystal of succinate of formula X, i.e., crystal form C-2, whose X-ray powder diffraction pattern has peaks at at least the following diffraction angles 2θ (°): 37.51±0.2, 43.64±0.2.

[0054] In some embodiments, the X-ray powder diffraction pattern of the crystal form C-2 further includes two or more peaks with diffraction angle 2θ (°) values ​​selected from the group consisting of: 9.67±0.2, 12.55±0.2, 13.66±0.2, 14.26±0.2, 14.92±0.2, 15.70±0.2, 17.05±0.2, 17.56±0.2, 21.46±0.2, and 23.12±0.2.

[0055] In some embodiments, the X-ray powder diffraction pattern of the crystal form C-2 has peaks at the following diffraction angles 2θ (°): 9.67±0.2, 12.55±0.2, 13.66±0.2, 14.26±0.2, 14.92±0.2, 15.70±0.2, 17.05±0.2, 17.56±0.2, 21.46±0.2, 23.12±0.2, 37.51±0.2, 43.64±0.2.

[0056] In some embodiments, the X-ray powder diffraction pattern of crystal form C-2 has peaks at the 2θ (°) values ​​shown in Table (C-2), and the relative intensities of each peak are shown in Table (C-2):

[0057] Table (C-2)

[0058] 2θ(°) <![CDATA[I / I0]]> 2θ(°) <![CDATA[I / I0]]> 2θ(°) <![CDATA[I / I0]]> 1.96 W 3.71 W 9.67 M 10.13 W 11.59 W 12.55 M 13.66 M 14.26 M 14.92 M 15.70 M 17.05 M 17.56 M 17.98 W 18.26 W 19.31 W 19.78 W 21.46 M 21.88 W 23.12 M 23.80 W 24.82 W 27.02 W 28.75 W 33.96 W 37.51 VS 43.64 VS

[0059] In some embodiments, the X-ray powder diffraction pattern of the crystal form C-2 is substantially as follows: Figure 8 What it represents.

[0060] In some implementations, the crystal form C-2 is in anhydrous form.

[0061] In some embodiments, the crystal form C-2 further has one or more features selected from the group consisting of:

[0062] (i) In the differential scanning calorimetry (DSC) spectrum, the peak temperature is 117.14 ± 2 °C; in some embodiments, the DSC spectrum is basically as follows: Figure 9 What it represents;

[0063] (ii) The basic thermogravimetric analysis spectrum is as follows Figure 10 What it represents.

[0064] A second aspect of this application provides a method for preparing a pharmaceutically acceptable salt of a compound of formula X, comprising the following steps:

[0065] S100: A pharmaceutically acceptable salt of compound X is prepared by mixing a compound of formula X, an acid, and a solvent and carrying out a salt-forming reaction.

[0066] Compound X has the following structure:

[0067]

[0068] The acid is selected from maleic acid, L-tartaric acid and succinic acid.

[0069] A third aspect of this application provides a method for preparing a polymorph of a pharmaceutically acceptable salt of compound X, comprising the following steps:

[0070] S210: Mix compound X, acid and solvent to carry out salt formation reaction to obtain reaction solution;

[0071] S220: Crystallize the reaction solution to obtain a polymorph;

[0072] Compound X has the following structure:

[0073]

[0074] The acid is selected from maleic acid, L-tartaric acid and succinic acid.

[0075] In some embodiments, in step S210, the molar ratio of acid to compound X is about (1.1-1.3):1, and in other embodiments it is 1.2:1.

[0076] In some embodiments, in step S210, the compound of formula X, the acid and the solvent are mixed and reacted at a temperature of about 25°C to about 55°C for about 5 to 120 min, then reacted at a temperature of about 25°C to about 45°C for about 5 to 120 min, and then reacted at a temperature of about 25°C to about 35°C for about 5 to 120 min.

[0077] In some embodiments, in step S210, the compound of formula X, the acid and the solvent are mixed and reacted at a temperature of about 50°C for about 60 min, then reacted at a temperature of about 40°C for about 60 min, and then reacted at a temperature of about 30°C for about 60 min.

[0078] In some implementations, in step S220, crystallization is performed by cooling, suspension and shaking, or adding an antisolvent.

[0079] In some implementations, in step S220, crystallization is performed by natural cooling.

[0080] In some embodiments, the solvent in step S210 is selected from one or more of acetonitrile, water, methanol, ethanol, isopropanol, acetone, ethyl acetate, methyl tert-butyl ether, tetrahydrofuran, and n-heptane.

[0081] In some embodiments, the polymorph is polymorph A, the acid is maleic acid, and the solvent in step S210 is one or more of acetonitrile, water, methanol, ethanol, isopropanol, acetone, ethyl acetate, methyl tert-butyl ether, tetrahydrofuran, and n-heptane; in other embodiments, it is ethyl acetate.

[0082] In some embodiments, the polymorph is polymorph B, the acid is L-tartaric acid, and the solvent in step S210 is one or more of acetonitrile, water, methanol, ethanol, isopropanol, acetone, ethyl acetate, methyl tert-butyl ether, tetrahydrofuran, and n-heptane; in other embodiments, it is acetone.

[0083] In some embodiments, the polymorph is crystal form C-1, the acid is succinic acid, and the solvent in step S210 is acetone.

[0084] In some embodiments, the polymorph is C-2, the acid is succinic acid, and the solvent in step S210 is ethyl acetate.

[0085] In some implementations, the polymorph is the polymorph described in the first aspect of this application.

[0086] The fourth aspect of this application provides a method for preparing crystal form A, comprising the steps of:

[0087] S310: Mix compound X, maleic acid and solvent to carry out a salt formation reaction to obtain a reaction solution;

[0088] S320: Crystallization treatment of the reaction solution by cooling, evaporation, suspension and shaking, or addition of antisolvent.

[0089] In some embodiments, in step S310, the solvent is selected from one or more of acetonitrile, water, methanol, ethanol, isopropanol, acetone, ethyl acetate, methyl tert-butyl ether, tetrahydrofuran, and n-heptane.

[0090] In some embodiments, in step S310, the solvent is ethyl acetate.

[0091] In some implementations, in step S320, crystal form A is obtained by natural cooling.

[0092] A fifth aspect of this application provides a method for preparing crystal form B, comprising the steps of:

[0093] S410: Mix compound X, L-tartaric acid and solvent to carry out a salt formation reaction to obtain a reaction solution;

[0094] S420: Crystallization treatment of the reaction solution by cooling, suspension and shaking, or addition of antisolvent.

[0095] In some implementations, in step S420, crystallization is performed by cooling or suspension shaking.

[0096] In some embodiments, in step S410, the solvent is selected from one or more of acetonitrile, water, methanol, ethanol, isopropanol, acetone, ethyl acetate, methyl tert-butyl ether, tetrahydrofuran, and n-heptane.

[0097] In some embodiments, in step S420, the product obtained after crystallizing the reaction solution is dissolved again in a second solvent, and crystallization is performed by cooling to obtain crystal form B. The second solvent is selected from acetonitrile, acetone, ethyl acetate, methyl tert-butyl ether, tetrahydrofuran, n-heptane, and combinations thereof.

[0098] In some implementations, step S420 includes the following steps:

[0099] S421: Cool the reaction solution to approximately -10°C to approximately 10°C, collect the solid after it precipitates;

[0100] S422: Dissolve the solid in a second solvent and crystallize it by cooling to obtain the crystal form B.

[0101] In some embodiments, the second solvent is acetonitrile, and step S422 includes the following steps:

[0102] S4221: Dissolve the solid in acetonitrile at approximately 30-70°C (60°C in some embodiments) to prepare a near-saturated solution, filter, and collect the filtrate;

[0103] S4222: Add acetonitrile to the filtrate and cool it to about -10°C to about 20°C. Crystals will precipitate. Collect the precipitated crystals to obtain crystal form B.

[0104] The sixth aspect of this application provides a method for preparing crystal form C-1, comprising the steps of:

[0105] S510: Mix compound X, succinic acid and solvent to carry out a salt formation reaction to obtain a reaction solution;

[0106] S520: Crystallization treatment of the reaction solution by cooling, evaporation, suspension and shaking, or addition of antisolvent.

[0107] In some implementations, in step S510, the solvent is selected from acetone.

[0108] In some implementations, step S520 involves a cooling process for crystallization.

[0109] The seventh aspect of this application provides a method for preparing crystal form C-2, comprising the steps of:

[0110] S610: Mix compound X, succinic acid and solvent to carry out a salt formation reaction to obtain a reaction solution;

[0111] S620: Crystallization treatment of the reaction solution by cooling, evaporation, suspension and shaking, or addition of antisolvent.

[0112] In some embodiments, in step S610, the solvent is selected from ethyl acetate.

[0113] In some implementations, step S620 involves a cooling process for crystallization.

[0114] An eighth aspect of this application provides a pharmaceutical composition comprising:

[0115] (a) The pharmaceutically acceptable salt of formula X or the polymorph of the pharmaceutically acceptable salt of formula X as described in the first aspect of this application, the pharmaceutically acceptable salt of formula X prepared by the preparation method described in the second aspect, the polymorph of the pharmaceutically acceptable salt of formula X prepared by the preparation method described in the third aspect, crystal form A prepared by the preparation method described in the fourth aspect, crystal form B prepared by the preparation method described in the fifth aspect, crystal form C-1 prepared by the preparation method described in the sixth aspect, or crystal form C-2 prepared by the preparation method described in the seventh aspect; and

[0116] (b) Pharmaceutically acceptable carrier.

[0117] The ninth aspect of this application provides the use of a pharmaceutically acceptable salt or polymorph of a pharmaceutically acceptable salt of a compound of formula X as described in the first aspect of this application, a pharmaceutically acceptable salt of a compound of formula X prepared by the preparation method of the second aspect, a polymorph of a pharmaceutically acceptable salt of a compound of formula X prepared by the preparation method of the third aspect, crystal form A prepared by the preparation method of the fourth aspect, crystal form B prepared by the preparation method of the fifth aspect, crystal form C-1 prepared by the preparation method of the sixth aspect, crystal form C-2 prepared by the preparation method of the seventh aspect, or a pharmaceutical composition as described in the eighth aspect of this application in the preparation of a medicament for the prevention and / or treatment of MOR receptor agonist-mediated diseases.

[0118] In some implementations, the MOR receptor agonist-mediated diseases are selected from pain, immune dysfunction, inflammation, esophageal reflux, neurological and psychiatric diseases, urinary and reproductive diseases, cardiovascular diseases, and respiratory diseases.

[0119] In some implementations, the MOR receptor agonist-mediated disease is pain.

[0120] In some implementations, the pain is selected from postoperative pain, cancer-related pain, neuropathic pain, traumatic pain, and inflammatory pain.

[0121] In some implementations, the cancer in the pain caused by the cancer is selected from breast cancer, endometrial cancer, cervical cancer, skin cancer, prostate cancer, ovarian cancer, fallopian tube tumors, ovarian tumors, hemophilia, and leukemia.

[0122] The tenth aspect of this application provides a method for preventing and / or treating MOR receptor agonist-mediated diseases, comprising administering to a desired patient a therapeutically effective amount of a pharmaceutically acceptable salt of the first aspect or a polymorph of a pharmaceutically acceptable salt of the first aspect, a pharmaceutically acceptable salt of the second aspect prepared by the preparation method, a polymorph of a pharmaceutically acceptable salt of the third aspect prepared by the preparation method, crystal form A prepared by the preparation method of the fourth aspect, crystal form B prepared by the preparation method of the fifth aspect, crystal form C-1 prepared by the preparation method of the sixth aspect, crystal form C-2 prepared by the preparation method of the seventh aspect, or a pharmaceutical composition as described in the eighth aspect of this application.

[0123] It should be understood that, within the scope of this application, the above-described technical features of this application and the technical features specifically described below (such as in the embodiments) can be combined with each other to form new or preferred technical solutions. Due to space limitations, they will not be described in detail here.

[0124] The pharmaceutically acceptable salts of the compound of formula X and their polymorphs not only have good physicochemical stability, but also good in vivo and in vitro related pharmacological activities, thus having the potential to be further developed into drugs. Attached Figure Description

[0125] Figure 1 The X-ray powder diffraction pattern is for crystal form A in Example 2 of this application.

[0126] Figure 2 This is the differential scanning calorimetry (DSC) spectrum of crystal form A in Example 2 of this application.

[0127] Figure 3 This is the thermogravimetric analysis spectrum of crystal form A in Example 2 of this application.

[0128] Figure 4 The X-ray powder diffraction pattern of crystal B in Example 3a of this application is shown.

[0129] Figure 5 This is the differential scanning calorimetry (DSC) spectrum of crystal B in Example 3a of this application.

[0130] Figure 6The thermogravimetric analysis spectrum of crystal B in Example 3a of this application is shown.

[0131] Figure 7 The X-ray powder diffraction pattern of crystal form C-1 in Example 4 of this application is shown.

[0132] Figure 8 The X-ray powder diffraction pattern of crystal form C-2 in Example 5 of this application is shown.

[0133] Figure 9 This is the differential scanning calorimetry (DSC) spectrum of crystal form C-2 in Example 5 of this application.

[0134] Figure 10 This is the thermogravimetric analysis spectrum of crystal form C-2 in Example 5 of this application.

[0135] Figure 11 This is a single-crystal structure diagram of compound 1e.

[0136] Figure 12 The X-ray powder diffraction patterns of crystal form A at 7 days and 30 days under conditions of 60℃ and 40℃ and 75%RH are shown.

[0137] Figure 13 The X-ray powder diffraction patterns of crystal form B at 7 days and 30 days under conditions of 60℃ and 40℃ and 75%RH are shown.

[0138] Figure 14 X-ray powder diffraction patterns of crystal form C-1 at 7 days and 30 days under conditions of 60℃ and 40℃ at 75%RH.

[0139] Figure 15 This is the infrared spectrum of crystal form B. Detailed Implementation

[0140] Through extensive and in-depth research, the inventors discovered compound X, which exhibits high inhibitory activity against cAMP and a high Emax value, while showing a low Emax value against β-arrestin, indicating good bias. Further research unexpectedly revealed a pharmaceutically acceptable salt of compound X and a series of its polymorphs. These salts and polymorphs not only possess good physicochemical stability but also exhibit good in vivo and in vitro pharmacological activities, thus demonstrating the potential for further development into drugs.

[0141] Compound X

[0142] In this application, compound X is (4S,6S)-6-isopropyl-N-(2-((R)-9-(pyridin-2-yl)-6-oxaspiro[4.5]decane-9-yl)ethyl)-5,6-dihydro-4H-pyrrolo[1,2-b]pyrazole-4-amine, which has high inhibitory activity against cAMP and a low Emax value against β-arrestin, exhibiting excellent bias.

[0143] This application also includes pharmaceutically acceptable salts of compound X and polymorphs of pharmaceutically acceptable salts of compound X, wherein the pharmaceutically acceptable salts are selected from maleates, L-tartrates, and succinates. In this application, polymorphs include polymorphs of pharmaceutically acceptable salts of compound X (such as maleates, L-tartrates, or succinates) and their various solvates, as well as different polymorphs of the same salt or solvate. The polymorphs of this application include (but are not limited to) crystal form A, crystal form B, crystal form C-1, and crystal form C-2.

[0144] Polymorphs

[0145] Solids include both amorphous and crystalline forms. In the crystalline form, molecules are located within three-dimensional lattice sites. When a compound crystallizes from a solution or slurry, it can crystallize using different spatial lattice arrangements (a property known as "polymorphism"), forming crystals with different crystalline forms, which are called "polymorphs." Different polymorphs of a given substance can differ from each other in one or more physical properties (such as solubility and dissolution rate, true specific gravity, crystal form, packing order, fluidity, and / or solid-state stability).

[0146] crystallization

[0147] Production-scale crystallization can be achieved by manipulating the solution to exceed the solubility limit of the target compound. This can be done in various ways, such as dissolving the compound at a relatively high temperature and then cooling the solution below its saturation limit. Alternatively, the liquid volume can be reduced by boiling, evaporation at atmospheric pressure, vacuum drying, or other methods. The solubility of the target compound can be reduced by adding an antisolvent or a solvent in which the compound has low solubility, or a mixture of such solvents. Another alternative method is to adjust the pH value to reduce solubility. For a detailed description of crystallization, see Crystallization, 3rd Edition, JW Mullens, Butterworth-Heineman Ltd., 1993, ISBN 0750611294.

[0148] The "suspension shaking" described in this application refers to a method of obtaining crystals by mixing a compound of formula X with a corresponding acid or a solution of the corresponding acid in a suitable solvent to form a turbid liquid and then shaking it. A suitable solvent may be water or an organic solvent.

[0149] The “slow evaporation” described in this application refers to a method of slowly evaporating the solvent from a solution containing a pharmaceutically acceptable salt of compound X or a solution containing compound X and the corresponding acid at a certain temperature to obtain crystals.

[0150] The "antisolvent addition" or "addition of antisolvent" described in this application refers to a method of precipitating crystals by adding another suitable solvent to a solution of a directional compound X.

[0151] This application relates to a technique for the simultaneous formation and crystallization of salts. Specifically, if the salt has a lower solubility in the reaction medium than the reactants, the addition of an appropriate acid or base can lead to the direct crystallization of the desired salt. Similarly, in a medium where the solubility of the final product is lower than that of the reactants, the completion of the synthesis reaction can allow the final product to crystallize directly.

[0152] Optimization of crystallization may include seeding the crystallizer with the desired crystal form as a seed crystal in the crystallization medium. Additionally, many crystallization methods utilize combinations of the strategies described above. One embodiment involves dissolving the compound of interest in a solvent at a high temperature, followed by the controlled addition of an appropriate volume of antisolvent to bring the system just below saturation. At this point, the desired crystal form can be added (while maintaining the integrity of the seed crystal), and the system is cooled to complete crystallization.

[0153] As used herein, the term “room temperature” (RT) generally refers to about 4–30°C, and in some embodiments to about 20 ± 5°C.

[0154] Identification and properties of polymorphs

[0155] The polymorphs of compound X and their preparation process can be tested and studied using a variety of methods and instruments.

[0156] X-ray powder diffraction

[0157] The polymorph of the pharmaceutically acceptable salt of compound X of this application has a specific crystal morphology and exhibits specific characteristic peaks in its X-ray powder diffraction (XRPD) pattern. In some embodiments, the XRPD patterns were acquired using an ARL Equinox 3000 X-ray powder diffractometer, and the XRPD parameters are shown in the table below:

[0158] parameter XRPD X-ray source Cu K (λ = 1.54056 Angstrom) Light tube settings 40 kV, 30 mA detector PSD Scan range, 2θ (°) 0°～108° Scan step size, 2θ (°) 0.05 Scan rate 1 second / step

[0159] In X-ray powder diffraction patterns, the positions of each peak are determined by 2θ (°). It is understood that different instruments and / or conditions can lead to slightly different data, and variations in the positions and relative intensities of each peak. Those skilled in the art should understand that when determining crystal form based on XRPD patterns, the peaks at low diffraction angles, their intensity, and peak shape integrity are relatively more meaningful. When the crystal form of this application was tested using the aforementioned instrument, strong background peaks of the blank metal disk appeared in two high diffraction angle ranges: 37.5-38 and 43.5-44. Figure 1 , 4 The same positions in 7 and 8 correspond to peaks. In this application, each crystal form uses its highest diffraction peak as the base peak, and its relative intensity is defined as 100%, as I0. The intensity division of the peaks only reflects the approximate size of the peaks at each position. (The peak with a 2θ (°) value of 17.29 for crystal form A is the base peak, the peak with a 2θ (°) value of 13.82 for crystal form B is the base peak, the peak with a 2θ (°) value of 14.11 for crystal form C-1 is the base peak, and the peak with a 2θ (°) value of 37.51 for crystal form C-2 is the base peak). The ratio of the peak height of each other peak to the peak height of the base peak is used as its relative intensity I / I0. The division of the relative intensity of each peak is defined as shown in the table below:

[0160] <![CDATA[Relative intensity I / I0 (%)]]> definition 50～100 VS (Very strong) 25～50 S (Strong) 10～25 M (Medium) 1～10 W(weak)

[0161] Differential scanning calorimetry (DSC) and thermogravimetric analysis (TGA)

[0162] DSC and TGA spectra were acquired using a DSC25A differential scanning calorimeter and a TGA550 thermogravimetric analyzer, respectively. The test parameters are shown in the table below:

[0163] parameter DSC TGA method linear heating linear heating Sample tray Aluminum tray, open Platinum plate, cap Temperature range 25℃ – Set temperature 25℃ – Set temperature Scan rate (°C / min) 10 10 Protective gas Nitrogen Nitrogen

[0164] It is understood that other values ​​may be obtained when using other types of instruments that have the same function as the instruments described above or when using test conditions different from those used in this application. Therefore, the values ​​cited should not be regarded as absolute values.

[0165] Due to instrument errors or differences in operators, those skilled in the art will understand that the parameters used to characterize the physical properties of crystals may have slight differences. Therefore, the above parameters are only used to assist in characterizing the polymorphs provided in some embodiments of this application, and should not be regarded as a limitation on the polymorphs of this application.

[0166] Infrared spectroscopy (IR)

[0167] The tests were performed using an Agilent Cary 630 FTIR spectrometer and the solid KBr pellet method.

[0168] Pharmaceutical compositions of compound X and their applications

[0169] Typically, the compound of formula X or a pharmaceutically acceptable salt thereof can be formulated with one or more pharmaceutical carriers to form suitable dosage forms for administration. These dosage forms are suitable for oral, rectal, topical, intraoral, and other non-gastrointestinal administration (e.g., subcutaneous, intramuscular, intravenous, etc.). For example, suitable dosage forms for oral administration include capsules, tablets, granules, and syrups. The compound of this application contained in these formulations can be a solid powder or granules; a solution or suspension in an aqueous or non-aqueous liquid; an oil-in-water or water-in-oil emulsion, etc. The above dosage forms can be prepared from the active compound and one or more carriers or excipients using common pharmaceutical methods. The carriers mentioned above need to be compatible with the active compound or other excipients. For solid dosage forms, commonly used non-toxic carriers include, but are not limited to, mannitol, lactose, starch, magnesium stearate, cellulose, glucose, sucrose, etc. Carriers used for liquid dosage forms include water, physiological saline, glucose aqueous solution, ethylene glycol, and polyethylene glycol, etc. The active compound can form solutions or suspensions with the above carriers.

[0170] The compositions of this application are formulated, quantified, and administered in accordance with medical practice guidelines. The “effective amount” of the compound administered is determined by factors such as the specific condition to be treated, the individual being treated, the cause of the condition, the target of the drug, and the route of administration.

[0171] This application provides that a pharmaceutically acceptable salt of compound X, or a polymorph of a pharmaceutically acceptable salt of compound X, as described in the first aspect of this application, can be used to prepare medicaments for the prevention and / or treatment of MOR receptor agonist-mediated diseases. The MOR receptor agonist-mediated diseases are selected from pain, immune dysfunction, inflammation, esophageal reflux, neurological and psychiatric diseases, urinary and reproductive diseases, cardiovascular diseases, and respiratory diseases. The pain is selected from postoperative pain, cancer-related pain, neuropathic pain, traumatic pain, and inflammatory pain. The cancer in the cancer-related pain is selected from breast cancer, endometrial cancer, cervical cancer, skin cancer, prostate cancer, ovarian cancer, fallopian tube tumors, ovarian tumors, hemophilia, and leukemia.

[0172] As used in this article, "therapeutic effective amount" means an amount that is functional or active in humans and / or animals and is acceptable to humans and / or animals.

[0173] As used herein, “pharmaceuticalally acceptable carrier” means a non-toxic, inert, solid, or semi-solid substance or liquid filling machine, diluent, encapsulation material, or excipient or any type of excipient that is compatible with patients, preferably mammalian, more preferably human, and suitable for delivering an active agent to a target site without terminating the agent’s activity.

[0174] As used in this article, "patient" refers to an animal, preferably a mammal, and more preferably a human. The term "mammal" refers to warm-blooded vertebrate mammals, including animals such as cats, dogs, rabbits, bears, foxes, wolves, monkeys, deer, rats, pigs, and humans.

[0175] As used in this article, “treatment” means to reduce, slow the progression of, diminish, prevent, or maintain an existing disease or condition (such as cancer). Treatment also includes curing, preventing the development of, or reducing to some extent one or more symptoms of a disease or condition.

[0176] The therapeutically effective amount of the pharmaceutically acceptable salt of formula X or the polymorph of the pharmaceutically acceptable salt of formula X contained in the pharmaceutical composition of this application is about 0.1 mg / kg to about 5 g / kg (body weight). Specific Implementation

[0178] The present application is further illustrated below with reference to specific embodiments. It should be understood that these embodiments are for illustrative purposes only and are not intended to limit the scope of the application. Experimental methods in the following embodiments that do not specify specific conditions are generally performed under conventional conditions or as recommended by the manufacturer. Unless otherwise stated, percentages and parts are by weight.

[0179] Reagents and Instruments

[0180] In the following embodiments of this application, the structure and purity of the compound were determined by nuclear magnetic resonance (NMR). 1 It can be determined by ¹H NMR and / or liquid chromatography-mass spectrometry (LC-MS). 1 HNMR: Bruker AVANCE-400 NMR spectrometer, internal standard: tetramethylsilane (TMS). LC-MS: Agilent 1290HPLC System / 6130 / 6150MS LC-MS / MS spectrometer (manufacturer: Agilent), Waters BEH / CHS column, 50×2.1mm, 1.7μm.

[0181] Single-crystal detection was performed using a Bruker D8 Venture X-ray single-crystal diffractometer. Preparative high-performance liquid chromatography (pre-HPLC): GX-281 (manufacturer: Gilson).

[0182] An ISCO Combiflash-Rf75 or Rf200 automatic column chromatography system was used, along with Agela disposable silicone columns in 4g, 12g, 20g, 40g, 80g, and 120g sizes.

[0183] The starting materials known in the following examples can be synthesized using or according to methods known in the art, or can be purchased from companies such as ABCR GmbH & Co. KG, Acros Organics, Aldrich Chemical Company, AccelaChemBio Inc, and Darui Chemicals.

[0184] Unless otherwise specified in the examples, all reactions were carried out under a nitrogen or argon atmosphere. As used herein, DCE is 1,2-dichloroethane, THF is tetrahydrofuran, EA is ethyl acetate, PE is petroleum ether, DCM is dichloromethane, n-BuLi is n-butyllithium, HATU is 2-(7-azobenzotriazole)-N,N,N',N'-tetramethylurea hexafluorophosphate, DMF is dimethylformamide, DMSO is dimethyl sulfoxide, DIEA or DIPEA is N,N-diisopropylethylamine, DBU is 1,8-diazabicyclo[5.4.0]undec-7-ene, and DIBAL-H is diisobutylaluminum hydride. Acetonitrile is ACN, methanol is MeOH, ethanol is EtOH, isopropanol is IPA, acetone is ACE, ethyl acetate is EA, methyl tert-butyl ether is MTBE, tetrahydrofuran is THF, and water is H2O.

[0185] Intermediate 1a

[0186]

[0187] (R)-2-(9-(pyridin-2-yl)-6-oxaspiro[4.5]decyl-9-yl)acetonitrile (11 g, 43 mmol, purchased from Shanghai Yucheng Pharmaceutical Technology Co., Ltd., CAS No. 1401031-38-6) was dissolved in DCM (100 ml) solution. Diisobutylaluminum hydride solution (1.0 M, 86 ml) was added dropwise at -78 °C, and the mixture was stirred at -78 °C for 1 hour. Sodium sulfate decahydrate (30 g) was added to the reaction solution, and the mixture was stirred at room temperature for half an hour. The mixture was filtered, and the filtrate was concentrated under reduced pressure to obtain a yellow oily intermediate. The intermediate was dissolved in ethanol (100 ml) solution, and hydrochloric acid (6 N, 100 ml) solution was added. The mixture was stirred at 100 °C for 36 hours. The residue was concentrated under reduced pressure and purified by preparative liquid chromatography to give (R)-2-(9-(pyridin-2-yl)-6-oxaspiro[4.5]decyl-9-yl)acetaldehyde 1a (6.5 g), yield 59%. MS m / z (ESI): 260.2 [M+1].

[0188] Intermediate 1b

[0189]

[0190] Step 1: Add triethylphosphonoacetate (476 mL, 2.4 mol), DBU (365 g, 2.4 mol), lithium chloride (127 g, 3 mol), and acetonitrile (1.2 L) to a 3 L single-necked flask. Stir at room temperature for 20 minutes under argon protection. Cool to 0 °C (internal temperature) and slowly add isobutyraldehyde (144 g, 2 mol). Stir at room temperature for 12 hours. LCMS showed complete reaction. Filter, wash the filter cake twice with EA (100 mL). Add water (1 L), extract twice with EA (1.5 L), wash with saturated sodium chloride, dry to anhydrous sodium sulfate, and evaporate to dryness to obtain compound 1b-1 (175 g, colorless liquid).

[0191] Step 2: DMF (1L) was added to a 3L single-necked flask containing compound 1b-1 (300g, 2.1mol). While stirring, potassium carbonate (579g, 4.2mol) and pyrazole (287g, 4.2mol) were added. The mixture was stirred at 65℃ for 18 hours. LC-MS showed the reaction was complete. The mixture was directly evaporated to dryness. The remaining solid was slurried with acetonitrile (150ml), and the filtered white solid was filtered. The filter cake was washed with EA (500mL x 2). It was then washed with saturated brine (300mL x 3), dried over anhydrous sodium sulfate, evaporated to dryness, and purified by column chromatography (PE containing 5% EA as the mobile phase) to obtain compound 1b-2 (258g, Y: 58%, colorless liquid). MS m / z (ESI): 211.1 [M+1].

[0192] Step 3: Dissolve 200 mL of water in potassium hydroxide (133 g, 3.32 mol) and pre-cool to 5 °C. Add compound 1b-2 (465 g, 2.21 mol), methanol (0.5 L), and THF (0.5 L) to a 3 L flask, then add the pre-cooled potassium hydroxide solution. Stir for 2 hours, adjust the pH to approximately 3 with concentrated hydrochloric acid, and extract with DCM (800 mL x 2). Combine all organic phases, wash with saturated brine, dry, and concentrate to obtain compound 1b-3 (410.5 g, Y: 100%, white solid). MS m / z (ESI): 183.1 [M+1].

[0193] Step 4: Under nitrogen protection, add compound 1b-3 (10.2 g, 0.055 mol) and THF (200 mL) to a three-necked flask (500 mL), and cool to -75 °C. Slowly add a 2.5 M solution of n-butyllithium (55 mL, 0.137 mol) in THF. After the addition is complete, slowly raise the temperature to -10 °C and continue stirring for 3 hours. Quench with saturated ammonium chloride, add water (100 mL), extract with EA (400 mL x 2), combine the organic phases, wash with saturated brine, dry, and concentrate to obtain a brown liquid. Purify by column chromatography (PE containing 30% EA as the mobile phase) to obtain compound 1b-4 (4 g, Y: 22.2%, white solid), MS m / z (ESI): 165.1 [M+1].

[0194] Step 5: Compound 1b-4 (17 g, 103.5 mmol) was dissolved in 250 mL of toluene, and (R)-2-methylpropane-2-sulfinamide (18.8 g, 155 mmol) and tetraisopropyl titanate (117 g, 414 mmol) were added. The mixture was stirred at 110 °C for 16 hours. After cooling to room temperature, 80 mL of water and 600 mL of DCM were added to the reaction solution. The mixture was filtered, and the filter cake was washed with 100 mL of DCM. The layers were separated, and the organic phase was washed with 100 mL of saturated brine. The mixture was dried over anhydrous sodium sulfate, concentrated under reduced pressure, and purified by column chromatography (PE containing 30% EA as the mobile phase) to obtain compound 1b (20.3 g, Y: 73.5%, brown liquid). MS m / z (ESI): 268.1 [M+1].

[0195] Compound 1e

[0196]

[0197] Compound 1d (0.11 g, 0.67 mmol) was dissolved in dichloromethane (8 mL), and triethylamine (0.2 g, 2 mmol) and 4-bromobenzenesulfonyl chloride (0.204 g, 0.804 mmol) were added at 0 °C. The mixture was stirred at 0 °C for 1 hour, and then dichloromethane (10 mL) was added. The mixture was washed with saturated sodium chloride solution, dried over anhydrous sodium sulfate, and the organic phase was concentrated under reduced pressure. The residue was purified by preparative liquid chromatography to give a white solid 4-bromo-N-((4S,6S)-6-isopropyl-5,6-dihydro-4H-pyrrolo[1,2-b]pyrazol-4-yl)benzenesulfonamide 1e (0.06 g) in 23.3% yield. MS m / z (ESI): 384.1 [M+1].

[0198] Preparation of single crystals of compound 1e: A small amount of compound 1e was placed in a small beaker and dissolved in ethyl acetate. The small beaker was sealed with a thin film, and small holes were punched in the film. Petroleum ether was placed in a large beaker, and then the small beaker was placed inside the large beaker. The large beaker was sealed and allowed to stand at room temperature for crystallization. Crystals precipitated from the solution. Single-crystal X-ray analysis of the crystal using a Bruker D8venture X-ray single-crystal diffractometer confirmed that its absolute configuration was (4S, 6S), and its structural diagram is shown below. Figure 11 As shown. Therefore, the absolute configuration of compound 1d can be deduced from the single-crystal structure of compound 1e as (4S, 6S) configuration.

[0199] Instrument parameters:

[0200]

[0201] Example 1: Preparation of Compound X

[0202]

[0203] Step 1: Compound 1b (0.39 g, 1.45 mmol) and methanol (5 mL) were added to a single-necked flask. The mixture was cooled to 0°C, and sodium borohydride (0.11 g, 2.9 mmol) was slowly added. The mixture was stirred at room temperature for 1.5 hours. After cooling to 0°C, 20 mL of ice water was added, and the mixture was extracted with DCM (50 mL x 2). The extract was washed with saturated brine (20 mL), dried over anhydrous sodium sulfate, and evaporated to dryness. The extract was prepared by preparative chromatography (preparative conditions: column: 21.2 x 250 mm C18 column, system: 10 mM NH4HCO3H2O, wavelength: 254 / 214 nm gradient: 28%-30% acetonitrile) to obtain a single diastereomer compound 1c (150 mg, yield: 38.4%, white solid, retention time: 9.5 min), MS m / z (ESI): 270.0 [M+1).

[0204] Step 2: Add compound 1c (120 mg, 0.45 mmol) and methanol (3 mL) to a single-necked flask, cool to 0°C, and slowly add methanol hydrochloric acid gas (1.2 mL, 4.5 mmol, 4 mol / L). Stir at room temperature for 1.5 hours. Adjust the pH to approximately 8 with saturated sodium bicarbonate, extract with DCM (50 mL x 2), combine all organic phases, wash with saturated brine, dry, and concentrate to obtain compound 1d (70 mg, yield: 95.12%, yellow liquid). MS m / z (ESI): 166.1 [M+1].

[0205] Step 3: Compound 1d (50 mg, 0.3 mmol) was dissolved in 5 mL of methanol. Compound 1a (78.3 mg, 0.3 mmol) and sodium cyanoborohydride (94 mg, 1.5 mmol) were added, and the mixture was stirred at 20 °C for 3 hours. 20 mL of water was added to the reaction mixture, and the mixture was extracted with DCM (30 mL × 2). The extract was dried over anhydrous sodium sulfate, concentrated under reduced pressure, and purified by preparative chromatography (preparative column: 21.2 × 250 mm C18 column; system: 10 mM NH4HCO3H2O; wavelength: 254 / 214 nm; gradient: 30%–60% acetonitrile change) to obtain compound X (10.12 mg, yield: 8.26%), a white solid. The absolute configuration of compound X was determined by inference from the absolute configuration of compound 1e.

[0206] MS m / z(ESI): 409.2 [M+1]; 1 H NMR (400MHz, CD3OD) δ8.51(ddd,J=4.9,1.8,0.8Hz,1H),7.79–7.71(m,1H),7.50(d,J=8.1Hz,1H),7.41(d,J=1. 9Hz,1H),7.22(ddd,J=7.5,4.9,1.0Hz,1H),5.87(dd,J=1.9,0.6Hz,1H),4.07(ddd,J=21.0,11.7,6.2Hz,2H),3 .81–3.69(m,2H),2.73–2.58(m,2H),2.52(dd,J=14.0,2.3Hz,1H),2.48–2.35(m,2H),2.12–1.97(m,2H),1.95– 1.82(m,2H),1.78–1.65(m,3H),1.64–1.36(m,5H),1.14–1.04(m,1H),0.99(d,J=7.0Hz,3H),0.79–0.65(m,4H).

[0207] Example 2 Preparation of maleate

[0208] 500 mg of compound X was weighed using the weight reduction method and added to a 20 mL glass sample vial. 10 mL of ethyl acetate was added, and the mixture was sonicated to dissolve. Then, 1 mol / L maleic acid solution was added at an acid-base molar ratio of 1.2:1. The reaction was carried out at 50 °C for 1 h. The temperature was then adjusted to 40 °C and stirring continued for 1 h. The temperature was then adjusted again to 30 °C and stirring continued for 1 h. Finally, the heating was turned off, and the mixture was stirred overnight. After the reaction was complete, the temperature was slowly lowered to 0 °C. The amount of solid precipitate increased. The solid was separated by centrifugation, and the solvent was evaporated to obtain the maleate salt solid product, which was a crystalline powder, defined as crystal form A. The X-ray powder diffraction pattern of crystal form A is shown below. Figure 1As shown (the 2θ angle is marked in the figure), the differential scanning calorimetry (DSC) spectrum and thermogravimetric analysis (TGA) spectrum are respectively as follows: Figure 2 and 3 As shown, the endothermic peak is 177.62℃, and the weight loss is 3.282% when heated to 150℃.

[0209] Example 3a Preparation of L-Tartrate

[0210] 300 mg of compound X was weighed using the weight reduction method and added to a 20 mL glass sample vial. 1.5 mL of acetone was added, and the mixture was sonicated to dissolve. Then, 1 mol / L L-tartaric acid solution was added at an acid-base molar ratio of 1.2:1. The reaction was carried out at 50 °C for 1 h. The temperature was then adjusted to 40 °C and stirring was continued for 1 h. The temperature was then adjusted again to 30 °C and stirring was continued for 1 h. Finally, the heating was turned off, and the mixture was stirred overnight. After the reaction was complete, the temperature was slowly lowered to 0 °C, resulting in an increase in the amount of solid precipitate. The solid was separated by centrifugation, and the solvent was evaporated to obtain the L-tartaric acid salt solid product a of compound X. Weigh approximately 40-50 mg of solid product a into a glass vial. Add an appropriate amount of acetonitrile in a 60°C water bath and stir until dissolved to obtain a nearly saturated solution. Filter the solution and add 20-200 μL of the corresponding solvent, acetonitrile, to the clear solution. Turn off the heating button and allow it to cool slowly. After reaching room temperature, place it in an ice bath to further cool to approximately 4°C. Collect the suspension and centrifuge at 12000 r / min for 15 min. Decant the supernatant and allow the solid to slowly evaporate overnight at room temperature. Collect the L-tartrate solid product b, which is a crystalline powder, defined as crystal form B. The X-ray powder diffraction pattern of crystal form B is shown below. Figure 4 As shown (the 2θ angle is marked in the figure), the differential scanning calorimetry (DSC) spectrum and thermogravimetric analysis (TGA) spectrum are respectively as follows: Figure 5 and 6 As shown, the endothermic peak is at 143.31℃, with a weight loss of 0.700%. The infrared spectrum is shown below. Figure 15 As shown.

[0211] The L-tartaric acid content of crystal form B was determined by HPLC. The measured tartaric acid content was 26.0%, and the theoretical tartaric acid content was 26.9%. The tartaric acid content was consistent with the theoretical value, indicating that the salt ratio of compound X to L-tartaric acid was 1:1.

[0212] Example 3b: Preparation of L-Tartrate

[0213] Six suspension-shaking experiments were conducted at 25°C using different solvent systems. Approximately 20 mg of the solid product a obtained in Example 3a was weighed into a glass vial, 1 mL of the organic reagent listed in Table 1 was added, the vial was tightly capped, sealed with sealing film to prevent evaporation, and then shaken at 25°C and 25 rpm. The vial was then centrifuged at 4°C and 14000 rpm for 15 min, the supernatant was discarded, and the solid was allowed to slowly evaporate overnight at room temperature. The solid product was collected and subjected to XRPD testing. The test results are shown in Table 1.

[0214] Table 1

[0215] solvent Solid crystal form after shaking for 1 day Solid crystal form after 7 days of shaking Tetrahydrofuran Crystal form B Crystal form B Acetonitrile Crystal form B Crystal form B Ethyl acetate Crystal form B Crystal form B acetone Crystal form B Crystal form B n-Heptane Crystal form B Crystal form B MTBE Crystal form B Crystal form B

[0216] Example 3c Preparation of L-Tartrate

[0217] Six suspension-shaking experiments were conducted at 50°C using different solvent systems. Approximately 20 mg of the solid product a obtained in Example 3a was weighed into a glass vial, 1 mL of the organic reagent listed in Table 2 was added, the vial was tightly capped, sealed with sealing film to prevent evaporation, and then shaken at 50°C and 225 rpm for one day. Afterward, the vial was removed, centrifuged at 4°C and 14000 rpm for 15 min, the supernatant was discarded, and the solid was allowed to slowly evaporate overnight at room temperature. The resulting solid was collected and subjected to XRPD testing. The test results are shown in Table 2.

[0218] Table 2

[0219] solvent Solid crystal form after shaking for 1 day Acetonitrile Crystal form B Ethyl acetate Crystal form B acetone Crystal form B n-Heptane Crystal form B MTBE Crystal form B

[0220] Example 4 Preparation of Succinate

[0221] 200 mg of compound X was weighed using the weight reduction method and added to a 20 mL glass sample vial. 10 mL of acetone was added, and the mixture was sonicated to dissolve. Then, 1 mol / L succinic acid solution was added at an acid-base molar ratio of 1.2:1. The reaction was carried out at 50 °C for 1 h, then the temperature was adjusted to 40 °C and stirring was continued for 1 h. The temperature was then adjusted again to 30 °C and stirring was continued for 1 h. Finally, the heating was turned off, and the mixture was stirred overnight. After the reaction was complete, the temperature was slowly lowered to 0 °C, resulting in an increase in the amount of solid precipitate. The solid was separated by centrifugation, and the solvent was evaporated to obtain the succinate solid product, which was a crystalline powder, defined as crystal form C-1. The X-ray powder diffraction pattern of the obtained crystal form C-1 is shown below. Figure 7 As shown (angle 2θ is marked in the figure).

[0222] Example 5 Preparation of Succinate

[0223] 300 mg of compound X was weighed using the weight reduction method and added to a 20 mL glass sample vial. 1.5 mL of ethyl acetate was added, and the mixture was sonicated to dissolve. Then, 1 mol / L succinic acid solution was added at an acid-base molar ratio of 1.2:1. The reaction was carried out at 50 °C for 1 h, then the temperature was adjusted to 40 °C and stirring was continued for 1 h. The temperature was then adjusted again to 30 °C and stirring was continued for 1 h. Finally, the heating was turned off, and the mixture was stirred overnight. After the reaction was complete, the temperature was slowly lowered to 0 °C, resulting in an increase in the amount of solid precipitate. The solid was separated by centrifugation, and the solvent was evaporated to obtain the succinate solid product, which was a crystalline powder, defined as crystal form C-2. The X-ray powder diffraction pattern of the obtained crystal form C-2 is shown below. Figure 8 As shown (the 2θ angle is marked in the figure), the differential scanning calorimetry (DSC) spectrum and thermogravimetric analysis (TGA) spectrum are respectively as follows: Figure 9 and 10 As shown, the endothermic peak is at 117.14℃, and the weight loss is 2.717% when heated to 120℃.

[0224] Example 6: Determination of Solubility

[0225] 10 mg of maleate crystal form A, L-tartrate crystal form B, succinate crystal form C-1, and crystal form C-2 were weighed separately, and 1 ml of solvent was added to each. The solubility of the samples in buffer solutions at pH 4.5 (acetic acid-sodium acetate system), pH 6.8 (sodium dihydrogen phosphate-sodium hydroxide system), and water was tested at room temperature. The phenomena were observed, and the test results are shown in Table 3 (the unit of concentration is mg / ml).

[0226] Table 3

[0227]

[0228] Example 7 Stability Experiment

[0229] Appropriate amounts of maleate crystal form A, L-tartrate crystal form B, and succinate crystal form C-1 samples were weighed and placed at 60℃ and 40℃ 75% RH, respectively. Meanwhile, another group of samples was sealed and stored at 5℃ as a control. Changes in crystal form and purity were detected at 7 days and 30 days, respectively. Figures 12 to 14 It can be seen that crystal forms A, B, and C-1 exhibit good physical stability. Table 4 shows that crystal forms A, B, and C-1 also exhibit good chemical stability.

[0230] Table 4. Chemical stability data for the three crystal forms

[0231]

[0232] - Indicates that the level is below the detection limit.

[0233] Example 8 Hygroscopicity Test

[0234] According to the "Guidelines for Hygroscopicity Testing of Drugs in Part IV, 9103 of the 2020 Edition of the Chinese Pharmacopoeia", the hygroscopicity of maleate crystal form A, L-tartrate crystal form B, succinate crystal form C-1 and crystal form C-2 samples were tested. The results showed that under the condition of 95% RH, crystal form C-1 and crystal form C-2 were slightly hygroscopic, crystal form A was slightly hygroscopic, and crystal form B was slightly hygroscopic.

[0235] Example 9 Biological Testing

[0236] The cell lines used in the following test cases are CHO-K1 OPRM1 β-Arrestin CellLine, Source: DiscoverX, Catalog Number: 93-0213C2, Lot Number: 13K0402.

[0237] The reagents used, their suppliers, product numbers, and storage temperatures are as follows:

[0238] Assay Complete TM Cell Culture Kit 107, DiscoverX, 92-3107G, -20°C;

[0239] AssayComplete TM Thawing Reagent, DiscoverX, 92-4002TR, -20℃;

[0240] AssayComplete TM Cell Detachment Reagent, DiscoverX, 92-0009, -20℃;

[0241] Assay Complete TM Cell Plating Reagent, DiscoverX, 93-0563R2, -20℃;

[0242] PathhunterDetection Kit, DiscoverX, 93-0001, -20℃;

[0243] PBS(1×)0.0067M(PO4), Hyclone, SH30256.01, 4℃;

[0244] DMSO, Sigma, D5879-100ML, room temperature;

[0245] NKH477, Sigma, 1603, -20℃;

[0246] IBMX, Tocris, I5879, -20℃.

[0247] The instruments used, their models, and suppliers are as follows:

[0248] Countsatr BioMed, IM1200, ALIT;

[0249] Microscope, IX51, OLYMPUS;

[0250] Centrifuge, 5804, Eppendorf;

[0251] Thermostatic Water Bath, DK-S420, ShanghaiShenxian thermostatic equipment factory;

[0252] Cell Incubator, 3111, Thermo;

[0253] Biological Safety Cabinet, BSC-1300IIA2, AIRTECH;

[0254] OptiPlate-384White Opaque, 6007290, Perkin Elmer;

[0255] Multimode plate Reader, Victor X5, PerkinElmer;

[0256] Culture Plate-384White Opaque,TC-treated, 6007680, PerkinElmer.

[0257] (1) HTRF-cAMP cell experiment

[0258] Experimental methods and procedures

[0259] I. Cell resuscitation

[0260] 1. Remove the resuscitation fluid from the 4°C refrigerator and preheat it in a 37°C water bath for 15 minutes.

[0261] 2. Remove the P6 generation cells from the liquid nitrogen tank, and quickly place the frozen cell cryovials into a 37°C water bath and gently shake for 30 seconds to 1 minute, until small ice crystals are visible or the cells are about to completely thaw.

[0262] 3. Thoroughly disinfect and dry with 70% alcohol.

[0263] 4. Centrifuge to remove the cryopreservation solution, and resuspend the cells in preheated fresh resuscitation solution.

[0264] a. Transfer 3 ml of preheated cell resuscitation solution into a 15 ml centrifuge tube.

[0265] b. Centrifuge at 1300 rpm for 3 minutes.

[0266] c. Remove the supernatant cryopreservation solution and resuspend the cells in 4 ml of preheated resuscitation solution.

[0267] 5. Transfer the cell suspension to a T25 cell culture flask and incubate for 24 hours at 37°C and 5% CO2.

[0268] 6. After culturing for 24 hours, replace the resuscitation solution in the cell culture flask with preheated cell culture medium.

[0269] II. Cell Passaging

[0270] 1. When the cell growth density in the T25 culture flask is >70%, digest and passage the cells using cell digestion solution.

[0271] a. Remove the culture medium from the culture flask, add 4 ml of preheated PBS, gently shake to rinse the cells, and then discard the PBS.

[0272] b. Add 1 ml of cell digestion solution to a T25 culture flask.

[0273] c. Shake the culture flask repeatedly to ensure the digestion solution completely covers the flask, and place it in a 37°C, 5% CO2 incubator for 5 minutes.

[0274] d. Remove the cell culture flask and observe the cells under a microscope to see if the cells have been separated.

[0275] e. Add 3ml of preheated cell culture medium to stop digestion.

[0276] f. Rinse the culture flask gently and repeatedly with cell culture medium, and collect the cell suspension into a 15ml centrifuge tube.

[0277] g. Centrifuge at 1300 rpm for 3 minutes and remove the supernatant.

[0278] h. Resuspend in 3 ml of cell culture medium.

[0279] 2. Passage cells at a ratio of 1:3 (add 1 ml of cell resuspension solution + 3 ml of cell culture medium to each flask and pass them to flask T25).

[0280] III. Cell Seeding Plate

[0281] 1. Repeat the cell passage steps ah until the cells reach passage P8. Count the cells and then resuspend them in 2× / 1mM IBMX stimulation buffer to achieve a cell density of 1.2*10^6 / ml.

[0282] 2. Using a multichannel pipette, seed a cell solution of 1.2*10^6 / ml into a 384-well plate at a volume of 10μl per well (i.e., 12,000 cells per well).

[0283] IV. c-AMP Experiment

[0284] 1. Prepare the relevant reagents and prepare the compounds according to the drug dilution preparation table.

[0285] a. 1×Stimulation buffer solution: Add 1 ml of 5×Stimulation buffer solution to 4 ml of distilled water and mix well.

[0286] b. 5 ml of 2×1 mM IBMX stimulation buffer: Add 10 μl of 500 mM IBMX storage solution to 4990 μl of cell culture medium and gently pipette to mix.

[0287] c. Gradient dilution preparation table for morphine, a positive control drug:

[0288]

[0289]

[0290] d. Before diluting the compound, dissolve it in DMSO to achieve a storage concentration of 10 mM.

[0291] Dilution preparation table for the positive control drug TRV130 and its compounds:

[0292]

[0293] e. 50 μM NK477 1 ml: Add 1 μl of 50 mM NKH477 storage solution to 999 μl of 1× Stimulation buffer solution and vortex to mix.

[0294] f. Test reagents

[0295] A. cAMP-Cryptate (donor, lyophilized) reaction solution: Add 1 ml of 5×cAMP-Cryptate storage solution to 4 ml of 1×Lysis & Detection Buffer solution and mix gently.

[0296] B. Anti-cAMP-d2 (acceptor, lyophilized) reaction solution: Add 1 ml of 5× Anti-cAMP-d2 storage solution to 4 ml of 1× Lysis & Detection Buffer solution and mix gently.

[0297] 2. cAMP experimental procedure

[0298] a. Seed 12,000 cells in 10 μl of 2x IBMX stimulation buffer per well.

[0299] b. Add 8 μl of compound sample diluent to each well of cells.

[0300] c. Add 2 μl of the prepared 10xNKH477 solution to each well.

[0301] d. Incubate at 37℃ for 45 mins.

[0302] e. Add 10 μl of cAMP-d2 and 10 μl of anti-cAMP Cryptate reaction solution.

[0303] f. Incubate at room temperature in the dark for 60 minutes.

[0304] g. HTRF reading board.

[0305] 3. RFU detection reader

[0306] After 60 minutes of incubation, all samples will be read using homogeneous time-resolved fluorescence.

[0307] Data Analysis

[0308] Data, including signal values ​​at 665nm and 620nm, was exported from the computer connected to the multi-functional plate reader. The ratio was calculated using the formula: Ratio = 665nm signal value / 620nm signal value × 10000. The data was analyzed using GraphPad Prism software. The best-fit curve was determined using log(agonist) vs. response. The EC50 value of the compound was determined using computer-aided dose-response curve nonlinear regression analysis; pEC50 = -logEC50 (EC50 value is in moles); the maximum effect value of % morphine = (compound sample ratio - blank well ratio) / TOP × 100 (Note: the TOP value is the top value of the fitted curve obtained by analyzing the morphine sample ratio - blank well ratio using GraphPad Prism software). The results are shown in Table 5.

[0309] Table 5. Activities of compounds against cAMP

[0310]

[0311] The structure of the comparative compound D1 is shown below, and it can be prepared according to the method described in CN111662284A.

[0312]

[0313] (2) β-Arrestin cell experiment

[0314] Experimental methods and procedures

[0315] I. Cell resuscitation

[0316] 1. Remove the resuscitation fluid from the 4°C refrigerator and preheat it in a 37°C water bath for 15 minutes.

[0317] 2. Remove the P6 generation cells from the liquid nitrogen tank, and quickly place the frozen cell culture tubes in a 37°C water bath and gently shake for 30 seconds to 1 minute, until you see small ice crystals or the cells are about to completely melt.

[0318] 3. Thoroughly disinfect and dry with 70% alcohol.

[0319] 4. Centrifuge to remove the cryopreservation solution, and resuspend the cells in preheated fresh resuscitation solution.

[0320] a. Transfer 3 ml of preheated cell resuscitation solution into a 15 ml centrifuge tube.

[0321] b. Centrifuge at 1300 rpm for 3 minutes.

[0322] c. Remove the supernatant and resuspend the cells in 4 ml of preheated resuscitation solution.

[0323] 5. Transfer the cell suspension to a T25 cell culture flask and incubate for 24 hours at 37°C and 5% CO2.

[0324] 6. After culturing for 24 hours, replace the resuscitation solution in the cell culture flask with preheated cell culture medium.

[0325] II. Cell Passaging

[0326] 1. When the cell growth density in the T25 culture flask is >70%, digest and passage the cells using cell digestion solution.

[0327] a. Aspirate the culture medium from the culture flask, add 4 ml of preheated PBS, gently shake to rinse the cells, and then discard the PBS.

[0328] b. Add 1 ml of cell digestion solution to a T25 culture flask.

[0329] c. Shake the culture flask repeatedly to ensure the digestion solution completely covers the flask, and place it in a 37°C, 5% CO2 incubator for 5 minutes.

[0330] d. Remove the cell culture flask and observe the cells under a microscope to see if the cells have been separated.

[0331] e. Add 3 ml of preheated cell culture medium to stop digestion.

[0332] f. Rinse the culture flask gently and repeatedly with cell culture medium, and finally transfer the cell suspension to a 15ml centrifuge tube.

[0333] Centrifuge at 1300 rpm for 3 minutes and remove the supernatant.

[0334] h. Resuspend in 3 ml of cell culture medium.

[0335] 2. Passage cells at a ratio of 1:3 (add 1 ml of cell resuspension solution + 3 ml of cell culture medium to each flask and pass them to flask T25).

[0336] 3. Repeat the cell passage steps ah until the cells are passaged to P8 generation.

[0337] III. Cell Seeding Plate

[0338] 1. Use a pipette to take 20 μl of cell suspension and measure the cell count using a cell counter.

[0339] 2. Centrifuge at 1300 rpm for 3 minutes to precipitate the cells.

[0340] 3. Remove the supernatant and add the appropriate cell plating medium to achieve a cell concentration of 2 × 10⁻⁶ cells / mL. 5 / ml.

[0341] 4. Using a multichannel pipette, according to the experimental design, pipette 2×10 5 A 20 μL / ml cell solution was seeded in each well of a 384-well plate (i.e., 4000 cells per well).

[0342] 5. Place the seeded 384-well plate in a 37°C, 5% CO2 incubator and incubate for 24 hours.

[0343] IV. β-arrestin test

[0344] 1. Prepare the compounds according to the following dilution table.

[0345] a. Gradient dilution preparation table for morphine, a positive control drug:

[0346]

[0347] b. Before diluting the compound, dissolve it in DMSO to achieve a storage concentration of 10 mM.

[0348] Dilution preparation table for the positive control drug TRV130 and its compounds:

[0349]

[0350] 2. Take 5 μl of the prepared diluent for each compound sample and add it to a 384-well plate.

[0351] 3. After adding the samples, place the 384-well plate back into a 37°C, 5% CO2 incubator and incubate for 90 minutes.

[0352] V. Relative Light Unit (RLU) Detection

[0353] 1. Before the end of compound incubation, prepare the Working Detection solution according to the following ratio (avoid light). Then add 12.5 μl to each well and incubate for 1 hour at room temperature in the dark on a shaker.

[0354]

[0355] 2. After the compound incubation is complete, add 12.5 μl of the above working solution to each well and incubate for 1 hour at room temperature and 80 rpm in the dark.

[0356] 3. After incubation, use a multi-functional plate reader to read the plates.

[0357] Data Analysis

[0358] Data was exported from the computer connected to the multi-functional plate reader and analyzed using GraphPad Prism software. The best-fit curve was determined using log(agonist) vs. response. The EC50 value of the compound was determined using computer-aided dose-response nonlinear regression analysis; pEC50 = -logEC50 (EC50 value is in moles); the maximum effect value of % morphine = (RLU value of the compound sample - RLU value of the blank well) / TOP × 100 (Note: TOP value is the TOP value of the fitted curve obtained by analyzing the RLU value of the morphine sample - RLU value of the blank well using GraphPad Prism software). The results are shown in Table 6.

[0359] Table 6 shows the test results of the compounds on β-arrestin.

[0360]

[0361] As shown in Tables 5 and 6, compared with the control compound D1, compound X exhibits higher inhibitory activity against cAMP and a higher Emax value. Furthermore, compound X shows a lower Emax value for β-arrestin, indicating better bias.

[0362] All references to this application are incorporated herein by reference as if each reference were individually incorporated herein by reference. Furthermore, it should be understood that after reading the foregoing teachings of this application, those skilled in the art can make various alterations or modifications to this application, and these equivalent forms also fall within the scope defined by the appended claims.

Claims

1. A pharmaceutically acceptable salt of a compound of formula X, said compound X having the following structure: in, The pharmaceutically acceptable salts are maleate, L-tartrate, or succinate.

2. A pharmaceutically acceptable salt of compound X as claimed in claim 1, wherein, The pharmaceutically acceptable salt of compound X is a polymorph, which is selected from any of the following polymorphs: The A-type crystal of maleate of compound X, i.e., crystal form A, has an X-ray powder diffraction pattern with peaks at at least the following diffraction angles 2θ (°): 10.06±0.2, 10.45±0.2, 11.13±0.2, 13.03±0.2, 16.81±0.2, 17.29±0.2, 19.45±0.2, 20.05±0.2, 24.04±0.2; The B-type crystal of compound X, L-tartrate, i.e. crystal form B, has an X-ray powder diffraction pattern with peaks at at least the following diffraction angles 2θ (°): 13.82±0.2, 14.87±0.2, 20.55±0.2, and 23.15±0.

2. The C-1 type crystal of the succinate of formula X, i.e., crystal form C-1, has an X-ray powder diffraction pattern with peaks at at least the following diffraction angles 2θ (°): 6.52±0.2, 10.37±0.2, 10.84±0.2, 13.66±0.2, 14.11±0.2, 15.61±0.2, 17.35±0.2, 20.92±0.2, and 23.20±0.

2. The C-2 crystal of succinate of formula X, i.e., crystal form C-2, has an X-ray powder diffraction pattern with peaks at at least the following diffraction angles 2θ (°): 9.67±0.2, 12.55±0.2, 13.66±0.2, 14.26±0.2, 14.92±0.2, 15.70±0.2, 17.05±0.2, 17.56±0.2, 21.46±0.2, 23.12±0.2, 37.51±0.2, and 43.64±0.

2.

3. A pharmaceutically acceptable salt of compound X as described in claim 2, wherein, The X-ray powder diffraction pattern of crystal form A is basically characterized by Figure 1; The X-ray powder diffraction pattern of crystal form B is basically characterized as shown in Figure 4. The X-ray powder diffraction pattern of the crystal form C-1 is basically characterized in Figure 7; The X-ray powder diffraction pattern of the crystal form C-2 is basically characterized as shown in Figure 8.

4. A pharmaceutically acceptable salt of compound X as described in claim 2, wherein, The X-ray powder diffraction pattern of crystal form B also contains two or more peaks with diffraction angle 2θ (°) values ​​selected from the following group: 10.41±0.2, 15.75±0.2, 16.28±0.2, 17.21±0.2, 17.63±0.2, 19.98±0.2, 20.22±0.2, 21.08±0.2, 22.22±0.2, 24.56±0.2, 28.91±0.

2.

5. A pharmaceutically acceptable salt of compound X as described in claim 2, wherein, The crystal form B also has one or more features selected from the group consisting of: (i) In the differential scanning calorimetry (DSC) spectrum, the peak temperature is 143.31 ± 2 °C; the DSC spectrum is basically represented as shown in Figure 5. (ii) The basic thermogravimetric analysis spectrum is shown in Figure 6; (iii) The basic infrared spectrum is characterized as shown in Figure 15.

6. A method for preparing a pharmaceutically acceptable salt of a compound of formula X, comprising the following steps: A pharmaceutically acceptable salt of compound X is prepared by mixing a compound of formula X, an acid, and a solvent and carrying out a salt-forming reaction. The compound of formula X has the following structure: The acid is selected from maleic acid, L-tartaric acid and succinic acid.

7. A method for preparing a polymorph of a pharmaceutically acceptable salt of compound X according to any one of claims 2-5, comprising the following steps: (i) Mix the compound of formula X, the acid and the solvent to carry out a salt formation reaction to obtain a reaction solution; (ii) The reaction solution is subjected to crystallization treatment to obtain the polymorph; The compound of formula X has the following structure: The acid is selected from maleic acid, L-tartaric acid and succinic acid.

8. The preparation method according to claim 6 or 7, wherein, The molar ratio of the acid to the compound of formula X is (1.1-1.3):

1.

9. The preparation method according to claim 6 or 7, wherein, The molar ratio of the acid to the compound of formula X is 1.2:

1.

10. The preparation method according to claim 6 or 7, wherein, The salt-forming reaction of mixing compound X, acid, and solvent includes the following steps: The compound of formula X, acid and solvent are mixed and reacted at 25℃-55℃ for 5-120 min, then at 25℃-45℃ for 5-120 min, and then at 25℃-35℃ for 5-120 min.

11. The preparation method according to claim 7, wherein, The crystallization process can be carried out by means of cooling, suspension and shaking, or adding antisolvent; or by means of natural cooling.

12. The preparation method according to claim 7, wherein, The polymorph is crystal form A, the acid is maleic acid, and the solvent is acetonitrile, water, methanol, ethanol, isopropanol, acetone, ethyl acetate, methyl tert-butyl ether, tetrahydrofuran, n-heptane, or a combination thereof. or The polymorph is crystal form B, the acid is L-tartaric acid, and the solvent is acetonitrile, water, methanol, ethanol, isopropanol, acetone, ethyl acetate, methyl tert-butyl ether, tetrahydrofuran, n-heptane, or a combination thereof. or The polymorph is C-1, the acid is succinic acid, and the solvent is acetone; or The polymorph is C-2, the acid is succinic acid, and the solvent is ethyl acetate.

13. The preparation method according to claim 7, wherein, The polymorph is crystal form B, and the crystallization process includes the following steps: Cool the reaction solution to -10°C to 10°C and collect the solid. The solid is dissolved in a second solvent and crystallized by cooling to obtain crystal form B. The second solvent is selected from tetrahydrofuran, acetonitrile, ethyl acetate, acetone, n-heptane, and methyl tert-butyl ether.

14. The preparation method according to claim 13, wherein, The second solvent is acetonitrile. The step of dissolving the solid in the second solvent and crystallizing it by cooling to obtain crystal form B includes the following steps: The solid was dissolved in acetonitrile at 30℃-70℃ to prepare a near-saturated solution, which was then filtered and the filtrate was collected. Acetonitrile was added to the filtrate, and the temperature was lowered to -10°C to 20°C. The precipitated crystals were collected to obtain crystal form B.

15. A pharmaceutical composition comprising: (a) A pharmaceutically acceptable salt of the compound of formula X as claimed in claim 1, a polymorph of any one of claims 2 to 5, or a pharmaceutically acceptable salt of the compound of formula X prepared by the method of claim 6, or a polymorph of the compound of formula X prepared by the method of any one of claims 7 to 14; and (b) Pharmaceutically acceptable carrier.

16. Use of a pharmaceutically acceptable salt of the compound of formula X according to claim 1, a polymorph of any one of claims 2 to 5, or a pharmaceutically acceptable salt of the compound of formula X prepared by the method of claim 6, or a polymorph of the compound of formula X prepared by the method of any one of claims 7 to 14, or the pharmaceutical composition of claim 15 in the preparation of a medicament for the prevention and / or treatment of MOR receptor agonist-mediated diseases.

17. The use as described in claim 16, wherein, The diseases mediated by the MOR receptor agonists are selected from pain, immune dysfunction, inflammation, esophageal reflux, neurological and psychiatric diseases, urinary and reproductive diseases, cardiovascular diseases, and respiratory diseases.

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