YBM compound as well as preparation method and application thereof

Abstract

The invention relates to the field of medicines, in particular to a YBM compound as well as a preparation method and application thereof. The invention provides a YBM compound or a pharmaceutically acceptable salt and a metabolite thereof. The YBM compound has a structure as shown in a formula 1, formula 1. According to the invention, SAR405 is used as a core skeleton, in order to enhance in-vivo absorption or reduce the removal rate of SAR405, a ring structure and substituent groups on the ring are modified and replaced, and finally a brand new compound with different skeleton structures and modification groups is provided. The SAR405 can be used as a host-targeted antiviral drug for blocking virus replication by inhibiting virus-host interaction, and exerts the in-vivo antiviral ability that SAR405 cannot have. Therefore, the VPS34 autophagy inhibitor can be applied to preparation of a medicine for preparing a VPS34 autophagy inhibitor, preparation of a medicine for resisting the SARS-CoV-2 virus and preparation of a medicine for treating pneumonia caused by the SARS-CoV-2 virus.

Description

Technical Field

[0001] The present invention relates to the field of medicine, in particular to a YBM compound and a preparation method and application thereof. Background Art

[0002] Currently, treatments for COVID-19 primarily include antiviral drugs, immunomodulators, and glucocorticoids, but these remain limited in efficacy, exhibit significant side effects, and are susceptible to drug resistance. Current treatment options generally utilize anti-SARS-CoV-2 drugs, including namatevir / ritonavir combination tablets, azithromycin tablets, and monoclavir capsules. All three drugs target the virus's own proteins to block viral replication. Currently, no host-targeted antiviral drugs are used clinically for the treatment of COVID-19.

[0003] Specifically, the main mechanisms of action of anti-SARS-CoV-2 drugs can be divided into two aspects: targeting the virus by directly acting on the virus's own proteins or enzymes, interfering with the virus's life cycle, thereby inhibiting viral replication and spread; targeting the host by regulating key factors or signaling pathways in host cells to create an intracellular environment that is not conducive to viral replication, thereby indirectly inhibiting viral replication and spread. The former usually has high specificity and can accurately target the key functional proteins of SARS-CoV-2. However, as SARS-CoV-2 continues to evolve, the structure of its own replication enzymes may change accordingly. Traditional antiviral drugs targeting viral proteins will have problems such as reduced efficacy. The latter directly targets the key proteins required for viral replication and can exhibit a broader spectrum of anti-SARS-CoV-2 capabilities.

[0004] VPS34 (Vacuolar Protein Sorting 34), a key protein involved in SARS-CoV-2 replication, holds promise as a novel anti-SARS-CoV-2 drug target. VPS34 is a class III phosphatidylinositol 3-kinase (PI3K-III) that plays a central role in cellular processes such as autophagy, endocytosis, and vesicle trafficking. Studies have shown that after SARS-CoV-2 infects host cells, it relies on the autophagy-related functions of VPS34 to form double-membrane vesicles (DMVs) that serve as sites for its replication. Therefore, inhibiting VPS34 may be a potential anti-SARS-CoV-2 strategy. Regarding the use of autophagy inhibitors for the treatment of coronaviruses, autophagy inhibitors such as chloroquine and hydroxychloroquine have been tried for the treatment of COVID-19. These inhibitors indirectly inhibit autophagy by increasing lysosomal pH, but clinical studies have shown limited efficacy and significant side effects.

[0005] Currently, only a few studies have reported that the VPS34 inhibitors VPS34 IN-1, VPS34 IN-2, SAR405, and Autophinib can inhibit SARS-CoV-2 replication in vitro models. No studies have demonstrated that these inhibitors have significant inhibitory activity against SARS-CoV-2 in vivo. In particular, studies have found that SAR405, the best existing VPS34 inhibitor, has poor anti-SARS-CoV-2 activity in mice. Summary of the Invention

[0006] In view of this, the technical problem to be solved by the present invention lies in a YBM compound, a preparation method and application thereof. The YBM compound provided by the present invention has excellent ability to inhibit VPS34 autophagy and resist SARS-CoV-2 in vivo, and can be used as a host-targeted antiviral drug for the clinical treatment of COVID-19.

[0007] A YBM compound or a pharmaceutically acceptable salt or metabolite thereof, wherein the YBM compound has a structure of Formula 1;

[0008] Formula 1.

[0009] The YBM compound provided by the present invention is a brown powdery solid, light-sensitive, with a solubility of about 0.08 mg / mL in water and about 30 mg / mL in DMSO, and low hygroscopicity. The inventors of the present application used SAR405 as the core skeleton, modified and replaced its ring structure and the substituent groups on the ring, and ultimately provided a compound with a completely new structure, which was named YBM compound. It can be used as a host-targeted antiviral drug to block viral replication by inhibiting virus-host interaction. It has strong absorption and low clearance rate in the body, and can exert excellent in vivo antiviral ability, overcoming the disadvantage of SAR405 drug that it does not have in vivo antiviral ability.

[0010] The present invention also provides a method for preparing a YBM compound, comprising the following steps:

[0011] Reacting a compound having a structure of formula A, 3-methylmorpholine, and 2-isopropoxyethylamine to obtain a YBM compound having a structure of formula 1;

[0012] Formula A; Formula 1.

[0013] The present invention uses a compound having a structure of formula A as a main skeleton, and reacts it with 3-methylmorpholine and 2-isopropoxyethylamine, wherein the structure of the 3-methylmorpholine is , the structure of the 2-isopropoxyethylamine is By replacing the Cl group on the skeleton of the compound having the structure of Formula A with 3-methylmorpholine, a morpholine skeleton was introduced to construct a skeleton structure similar to but different from SAR405. 2-isopropoxyethylamine was used to introduce a new modifying group at the amino site originally belonging to SAR405 to adjust the chemical structure of the site. Combined with the optimization of the skeleton structure and the modifying group, the YBM compound of the present invention was obtained.

[0014] The present invention can first synthesize the compound having the structure of Formula A, and then use it to further prepare the YBM compound having the structure of Formula 1, which specifically includes the following steps:

[0015] S1) reacting a compound having the structure of formula A-0 with 3-oxopropionic acid to obtain a compound having the structure of formula A-1;

[0016] Formula A-0; Formula A-1;

[0017] S2) reacting the compound having the structure of formula A-1 obtained in step S1) with phosphorus oxychloride to obtain a compound having the structure of formula A;

[0018] S3) reacting the compound having the structure of formula A obtained in step S2), 3-methylmorpholine and 2-isopropoxyethylamine to obtain a YBM compound having the structure of formula 1.

[0019] In certain embodiments of the present invention, the preparation method of the YBM compound specifically includes the following steps: S1) reacting a compound having a structure of formula A-0 with 3-oxopropionic acid in an organic solvent under the action of DCC and HoBt to obtain a compound having a structure of formula A-1; S2) reacting the compound having a structure of formula A-1 obtained in step S1) with phosphorus oxychloride under the action of triethylamine to obtain a compound having a structure of formula A; S3) reacting the compound having a structure of formula A obtained in step S2) with 3-methylmorpholine and 2-isopropoxyethylamine in ethanol to obtain a YBM compound having a structure of formula 1.

[0020] In the method for preparing the YBM compound of the present invention, the reaction temperature in step S1) is room temperature, specifically 15°C to 30°C, and the reaction time in step S1) is 10 to 15 hours, preferably 12 hours. The reaction temperature in step S2) is 80°C to 100°C, preferably 90°C; the reaction time in step S2) is 10 to 15 hours, preferably 12 hours. The reaction temperature in step S3) is room temperature, specifically 15°C to 30°C, and the reaction time in step S3) is 4 to 8 hours, preferably 6 hours.

[0021] The present invention provides the use of any of the above-described YBM compounds, or pharmaceutically acceptable salts, metabolites thereof, or YBM compounds obtained by any of the above-described preparation methods, in the preparation of medicaments for VPS34 autophagy inhibitors. The YBM compounds provided by the present invention are compounds optimized based on SAR405. Like SAR405, they have the effect of inhibiting VPS34 autophagy, but differ in that the YBM compounds provided by the present invention also have the effect of inhibiting VPS34 autophagy in vivo. The YBM compounds of the present invention are prepared into VPS34 autophagy inhibitors, which can resist viruses that rely on the VPS34 autophagy function for replication, and are very suitable for use in the preparation of medicaments for VPS34 autophagy inhibitors.

[0022] The present invention also provides the use of any of the above-described YBM compounds, or pharmaceutically acceptable salts, metabolites thereof, or YBM compounds obtained by any of the above-described preparation methods, in the preparation of drugs against the SARS-CoV-2 virus. Specifically, after SARS-CoV-2 infects host cells, it relies on the autophagy-related function of VPS34 to form double-membrane vesicle structures to create a replication site for itself. The YBM compounds of the present invention can inhibit VPS34 autophagy. Therefore, the YBM compounds of the present invention also have excellent anti-SARS-CoV-2 virus ability and are very suitable for use in the preparation of drugs against the SARS-CoV-2 virus.

[0023] The present invention also provides the use of any of the above-mentioned YBM compounds, or pharmaceutically acceptable salts, metabolites thereof, or YBM compounds obtained by any of the above-mentioned preparation methods, in the preparation of drugs for treating pneumonia caused by the SARS-CoV-2 virus. The present invention further explored the pharmacokinetics of SAR405 and found that the absorption of SAR405 in the body and the distribution of SARS-CoV-2 in the lungs, the target organ of SARS-CoV-2, were poor, which may be the main reason for the lack of antiviral effect of SAR405 in the body; the YBM compound obtained by optimization based on the structure of SAR405 has enhanced in vivo absorption performance and reduced in vivo clearance rate, and has a more excellent therapeutic effect on pneumonia caused by the SARS-CoV-2 virus, and is very suitable for use in the preparation of drugs for treating pneumonia caused by the SARS-CoV-2 virus.

[0024] The present invention also provides a pharmaceutical preparation comprising an active compound and an excipient; the active compound is selected from any of the aforementioned YBM compounds, or a pharmaceutically acceptable salt, metabolite thereof, or a YBM compound obtained by any of the aforementioned preparation methods. The present invention does not particularly limit the excipient; any excipient acceptable in pharmaceuticals can be used. The pharmaceutical preparation of the present invention is in the form of an oral preparation or an injectable preparation.

[0025] The present invention discloses a YBM compound, a preparation method, and an application thereof. Specifically, the present invention provides a YBM compound or a pharmaceutically acceptable salt or metabolite thereof, wherein the YBM compound has a structure of Formula 1; Formula 1. The present invention uses SAR405 as the core skeleton, and modifies and replaces its ring structure and the substituent groups on the ring for the purpose of enhancing its absorption in the body or reducing its clearance rate, ultimately providing a new compound with a different skeleton structure and modified groups, which is named YBM compound. It can be used as a host-targeted antiviral drug that blocks viral replication by inhibiting virus-host interaction, and exerts an in vivo antiviral ability that SAR405 does not have, thereby having applications in the preparation of drugs for VPS34 autophagy inhibitors and in the preparation of drugs against SARS-CoV-2 viruses, and can treat pneumonia caused by SARS-CoV-2 virus, and thus has applications in the preparation of drugs for treating pneumonia caused by SARS-CoV-2 virus. BRIEF DESCRIPTION OF THE DRAWINGS

[0026] Figure 1 This is the test graph of the cell half-maximal toxicity concentration of SAR405;

[0027] Figure 2 This is the test graph of the half-maximal cytotoxic concentration of YBM;

[0028] Figure 3 This is a comparison of the antiviral ability and survival rates of SAR405 and YBM in vivo;

[0029] Figure 4 This is a comparison of the viral load of SAR405 and YBM in vivo antiviral ability;

[0030] Figure 5 Comparison of N gene copy numbers of SAR405 and YBM in vivo antiviral ability. DETAILED DESCRIPTION

[0031] The present invention discloses a YBM compound, its preparation method, and application. Those skilled in the art may refer to the contents herein and appropriately modify the process parameters to achieve the desired results. It should be noted that all similar substitutions and modifications obvious to those skilled in the art are considered to be included in the present invention. The methods and applications of the present invention have been described through preferred embodiments, and relevant persons will clearly be able to modify or appropriately alter and combine the methods and applications herein to implement and apply the technology of the present invention without departing from the content, spirit, and scope of the present invention.

[0032] Compounds used in the present invention , 3-oxopropionic acid, DMF (N,N-dimethylformamide), DCC (dicyclohexylcarbodiimide), HoBt (1-hydroxybenzotriazole), phosphorus oxychloride, triethylamine, ethanol, and 2-isopropoxyethylamine are all commercially available reagents.

[0033] The present invention will be further described below with reference to the embodiments:

[0034] Example 1

[0035] The YBM compound was synthesized according to the following reaction formula:

[0036]

[0037] The specific process is as follows:

[0038] Step 1: (100 mM, 18.56 g) and 3-oxopropionic acid (120 mM, 10.56 g) were dissolved in 800 mL of DMF, and DCC (120 mM, 24.72 g) and HoBt (120 mM, 16.2 g) were added. The mixture was stirred at room temperature of 20-25°C for 12 hours to obtain 20.48 g of light yellow solid intermediate B with a yield of 80.95% and a purity of 94%.

[0039] Step 2: Dissolve intermediate B (20 mmol, 5.08 g) in phosphorus oxychloride (200 mmol, 30.37 g). Add triethylamine (24 mmol, 2.43 g) dropwise to the mixture. Stir at 90°C for 12 h. Evaporate the reaction mixture to dryness and recrystallize from dichloromethane to obtain 3.78 g of yellow intermediate C. Yield: 51.20%, purity: 95%.

[0040] Step 3: Intermediate C (5 mmol, 1.36 g) and (5 mmol, 0.51 g) was dissolved in 100 mL of anhydrous ethanol and stirred at room temperature of 20-25 °C for 6 hours. 2-isopropoxyethylamine (6 mmol, 0.77 g) was added and stirred for 6 hours to obtain 0.7 g of a white product. , that is, the YBM compound of the present invention, with a yield of 35.61% and a purity of 98%.

[0041] The cytotoxicity and in vivo antiviral activity of the YBM compound prepared above were compared with those of SAR405, as follows:

[0042] 1. Comparison of cytotoxicity between YBM compounds and SAR405 cells.

[0043] Vero E6 cells were plated into 96-well plates with 1×10 cells per well. 4After the cells were cultured overnight, the original culture medium in the 96-well plate was discarded. YBM and SAR405 were diluted into different concentrations using DMDM ​​without serum and double antibodies, and transferred to the well plate, 100 μL per well. After that, 100 μL of DMEM containing 10% FBS was added to each well and cultured in a 37°C, 5% CO2 incubator for 48 hours. After that, the well plate was removed and the culture medium was discarded. The diluted CCK8 detection reagent was added to each well. After incubation at 37°C for 2 hours, the cell viability was calculated using a microplate reader. The data were processed using Graphpad Prism software and the corresponding cell half-toxic concentration (CC50) was fitted. 50 ), the results are as follows Figure 1 and Figure 2 As shown, Figure 1 This is the cell half toxicity concentration test chart of SAR405. Figure 2 The figure shows the median cytotoxic concentration of YBM. It can be seen that in Vero E6 cells, the drug toxicity of YBM is about 4.35 times lower than that of SAR405.

[0044] 2. Comparison of the antiviral ability of compound YBM and SAR405 in mice.

[0045] (1) Comparison of survival rates

[0046] Eight-month-old Balb / c mice were randomly divided into four groups (n=6 / group): (1) Mock group: PBS was dripped into the nasal cavity; (2) SARS-CoV-2+Vechicle group (solvent control group): SARS-CoV-2 was dripped into the nasal cavity and blank solvent was injected intraperitoneally every day; (3) SARS-CoV-2+SAR405 group: SARS-CoV-2 was dripped into the nasal cavity and SAR405 was injected intraperitoneally at 100 mg / kg every day; (4) SARS-CoV-2+YBM group: SARS-CoV-2 was dripped into the nasal cavity and YBM was injected intraperitoneally at 100 mg / kg every day. All groups were observed continuously for seven days after virus infection, and the survival curve was calculated. The results are shown as follows: Figure 3 As shown, Figure 3 The figure shows a comparison of the antiviral activity and survival rates of SAR405 and YBM in vivo. It shows that on day 5 after viral infection, all mice in the solvent control and SAR405 groups died, while the survival rate of mice in the YBM group was 50%.

[0047] (2) Comparison of viral load and N gene copy number

[0048] The model was treated in the same way as in the survival curve (n=3 / group). Mice were killed on the third day of virus infection and samples were collected for testing to detect the replication of the virus in the lung tissue. The results are shown in Figure 2. Figure 4 and Figure 5 As shown, Figure 4 This is a comparison chart of the antiviral capacity and viral load of SAR405 and YBM in vivo. Figure 5 The figure shows a comparison of the antiviral activity of SAR405 and YBM in vivo, and the N gene copy number. It can be seen that the viral load in the YBM-treated group was significantly lower than that in the control group, and the corresponding N gene copy number was also reduced.

[0049] (3) Comparison of pharmacokinetic parameters of compound YBM and SAR405 in mice

[0050] Pharmacokinetic parameters and target organ concentration distributions of SAR405 and TBM were measured at different time points after intraperitoneal injection. The results are shown in Tables 1 and 2. SAR405 reached a maximum plasma concentration of 5499.07±891.54 ng / mL 8 minutes after intraperitoneal injection, with a clearance rate of 3250±1081.29 mL / h / kg. YBM reached a maximum plasma concentration of 14327.9±1322.77 ng / mL 8 minutes after intraperitoneal injection, approximately 2.5-fold higher than that of SAR405. Its clearance was only 615.74±99.58 mL / h / kg, approximately 5-fold lower than that of SAR405.

[0051] Table 1 Drug concentrations in whole blood (Mean ± SD, n = 3)

[0052]

[0053] Table 2 Main pharmacokinetic parameters in whole blood (Mean ± SD, n = 3)

[0054]

[0055] The above description is only a preferred specific embodiment of the present invention, but the scope of protection of the present invention is not limited thereto. Any technician familiar with the technical field, within the technical scope disclosed by the present invention, who makes equivalent replacements or changes based on the technical solution and inventive concept of the present invention, should be covered by the scope of protection of the present invention.

Claims

1. A YBM compound or a pharmaceutically acceptable salt or metabolite thereof, characterized in that: The YBM compound has a structure of Formula 1; Formula 1.

2. A method for preparing a YBM compound, characterized in that: The following steps are involved: Reacting a compound having a structure of formula A, 3-methylmorpholine, and 2-isopropoxyethylamine to obtain a YBM compound having a structure of formula 1; Formula A; Formula 1.

3. The preparation method according to claim 2, characterized in that The specific steps include: S1) reacting a compound having the structure of formula A-0 with 3-oxopropionic acid to obtain a compound having the structure of formula A-1; Formula A-0; Formula A-1; S2) reacting the compound having the structure of formula A-1 obtained in step S1) with phosphorus oxychloride to obtain a compound having the structure of formula A; S3) reacting the compound having the structure of formula A obtained in step S2), 3-methylmorpholine and 2-isopropoxyethylamine to obtain a YBM compound having the structure of formula 1.

4. The preparation method according to claim 3, characterized in that The specific steps include: S1) reacting a compound having the structure of formula A-0 with 3-oxopropionic acid in an organic solvent under the action of DCC and HoBt to obtain a compound having the structure of formula A-1; S2) reacting the compound having the structure of formula A-1 obtained in step S1) with phosphorus oxychloride in the presence of triethylamine to obtain a compound having the structure of formula A; S3) reacting the compound having the structure of formula A obtained in step S2), 3-methylmorpholine and 2-isopropoxyethylamine in ethanol to obtain a YBM compound having the structure of formula 1.

5. The preparation method according to claim 3 or 4, characterized in that The reaction temperature in step S1) is 15°C to 30°C, and the reaction time in step S1) is 10 h to 15 h; The reaction temperature in step S2) is 80° C. to 100° C., and the reaction time in step S2) is 10 h to 15 h; The reaction temperature in step S3) is 15°C to 30°C, and the reaction time in step S3) is 4 h to 8 h.

6. Use of the YBM compound according to claim 1 or a pharmaceutically acceptable salt or metabolite thereof, or the YBM compound obtained by the preparation method according to any one of claims 2 to 5 in the preparation of a drug for a VPS34 autophagy inhibitor.

7. Use of the YBM compound according to claim 1 or a pharmaceutically acceptable salt, metabolite thereof, or the YBM compound obtained by the preparation method of any one of claims 2 to 5 in the preparation of a drug against SARS-CoV-2 virus.

8. Use of the YBM compound according to claim 1 or a pharmaceutically acceptable salt, metabolite thereof, or the YBM compound obtained by the preparation method of any one of claims 2 to 5 in the preparation of a medicament for treating pneumonia caused by SARS-CoV-2 virus.

9. A pharmaceutical preparation, characterized in that It includes active compounds and excipients; The active compound is selected from the YBM compound according to claim 1 or a pharmaceutically acceptable salt or metabolite thereof, or the YBM compound obtained by the preparation method according to any one of claims 2 to 5.

10. The pharmaceutical preparation according to claim 9, characterized in that Its dosage form is oral preparation or injection preparation.

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