Intermediate compounds, novel compounds, and methods and uses thereof

Abstract

The application discloses an intermediate compound, a new compound and a synthesis method and application thereof, and belongs to the technical field of drug synthesis. The application is based on the characteristics of normal substrate nucleotides of RNA polymerase simulated by simple small molecules (non-nucleoside inhibitors) and competitive occupation of nucleotide entry channels of viral RNA polymerase, so as to competitively block the entry of raw materials of viral nucleic acid replication, so that the RNA polymerase cannot exert normal biological functions, the viral life cycle is blocked, and the viral life elements are degraded by enzyme systems in host cells, thereby effectively exerting an antiviral effect. The compound developed by the application is a non-nucleoside inhibitor, and existing nucleoside inhibitors (such as ribavirin, remdesivir, favilavir and the like) have obvious adverse reactions such as mutagenicity, carcinogenicity and impairment of fertility.

Description

Technical Field

[0001] This invention relates to intermediate compounds, novel compounds, their synthetic methods, and applications. It belongs to the field of pharmaceutical synthesis technology. Background Technology

[0002] Both the novel coronavirus and influenza virus are viruses that have coexisted with humans for a long time. With the continuous application of corresponding antiviral drugs in clinical practice, drug-resistant mutant strains of both have emerged continuously. These include drug-resistant strains of the novel coronavirus that are resistant to remdesivir (RNA polymerase inhibitor), nematvir (main protease inhibitor), and leretvir (main protease inhibitor), as well as drug-resistant strains of influenza that are resistant to oseltamivir (influenza neuraminidase inhibitor) and baloxavir (influenza endonuclease inhibitor). Developing new antiviral drugs has positive practical significance.

[0003] Influenza viruses (especially type A) have a high mutation rate, and long-term or inappropriate use of drugs such as oseltamivir and mabaloxavir can easily induce drug-resistant strains, leading to decreased efficacy and drug resistance issues. Furthermore, during peak influenza seasons, drugs such as oseltamivir are often in short supply, affecting timely treatment. Therefore, there is an urgent need to develop new antiviral drugs for influenza and new drugs that inhibit the RNA polymerase of the novel coronavirus. Summary of the Invention

[0004] In view of the above-mentioned shortcomings of the prior art, the purpose of the present invention is to provide a new intermediate compound.

[0005] Meanwhile, this invention provides a novel method for synthesizing intermediate compounds.

[0006] Meanwhile, this invention provides applications for novel intermediate compounds.

[0007] Meanwhile, this invention provides new compounds C and E.

[0008] Meanwhile, this invention provides new methods for synthesizing compounds C and E.

[0009] Meanwhile, this invention provides new applications for compounds C and E.

[0010] To solve the above-mentioned technical problems, the technical solution adopted by the present invention is as follows: The structural formula of the intermediate compound is as follows:

[0011] The method for synthesizing compound 8 includes the following steps: Step 1: Synthesis of Compound 6 from Compound 4: Compound 5 (52.7 g, 436 mmol, 1.10 eq) and sodium hydroxide (15.8 g, 396 mmol, 1.00 eq) were added to an aqueous solution (500 mL) of Compound 4 (50.0 g, 396 mmol, 1.00 eq), and the mixture was stirred at 25 °C for 12 hours. LC-MS (EC11562-64-P1A1) showed that Compound 4 was completely consumed, and the content of the target product was 57.8%. The reaction solution was extracted with MTBE (200 mL × 3). The organic phase was dried over Na2SO4, filtered, and concentrated under reduced pressure to obtain the crude product. Compound 6 (yellow solid, 6.00 g, 36.11 mmol, 9.11% yield) was purified by column chromatography (eluent ratio: petroleum ether / ethyl acetate = 1 / 0 - 1 / 1; TLC plate: petroleum ether: ethyl acetate = 1:1, Rf = 0.30).

[0012] Step 2: Synthesis of Compound 7 from Compound 6: POCl3 (1.85 g, 12.0 mmol, 1.12 mL, 2.00 eq) was added to a DMF (20.0 mL) solution of Compound 6 (1.00 g, 6.02 mmol, 1.00 eq), and the mixture was stirred at 25 °C for 1 hour. Then, H2O (20.0 mL) was added. The mixture was then stirred at 80 °C for 1 hour. LC-MS (EC14440-1-P1A1) showed that Compound 6 was completely consumed and the main product peak was formed. The reaction mixture was diluted with water (50 mL), then extracted with EtOAc (50 mL × 3). The combined organic phases were washed with water (50 mL), saturated brine (50 mL), dried over anhydrous Na2SO4, filtered, and concentrated under reduced pressure to obtain Compound 7 (yellow oil, 1.10 g, 5.66 mmol, yield 94.13%).

[0013] Step 3: Synthesis of Compound 8 from Compound 7: Compound 7 (500 mg, 2.57 mmol, 1.00 eq), Compound 2 (430 mg, 2.83 mmol, 1.10 eq), and TEA (1.43 g, 14.1 mmol, 1.97 mL, 5.50 eq) were added to a three-necked round-bottom flask, followed by Ac₂O (1.58 g, 15.4 mmol, 1.45 mL, 6.00 eq). The mixture was stirred at 115 °C for 8 hours. The mixture was then quenched with water (30.0 mL), precipitating a large amount of solid. The solid was then filtered, and the filter cake was washed three times with distilled water (20.0 mL × 3). The solid was then dissolved in ethanol (5.00 mL), followed by the addition of 20% HCl aqueous solution (5.00 mL), and stirred at 80 °C for 3 hours. LC-MS (EC11586-104-P1C) showed that compound 7 was completely consumed, and the target product was a single main peak. Ice water (20 mL) was added to the reaction solution, resulting in the precipitation of a large amount of solid. The mixture was filtered, and the filter cake was washed with water until the pH of the filtrate was near neutral. The filter cake was then concentrated under reduced pressure to obtain compound 8 (yellow solid, 500 mg, 1.61 mmol, yield 62.5%), whose structure was confirmed by ¹H NMR (EC11586-104-P1C).

[0014] The method for synthesizing compound 10 includes the following steps: Step 1: Synthesis of Compound 6 from Compound 4: Compound 5 (52.7 g, 436 mmol, 1.10 eq) and sodium hydroxide (15.8 g, 396 mmol, 1.00 eq) were added to an aqueous solution (500 mL) of Compound 4 (50.0 g, 396 mmol, 1.00 eq), and the mixture was stirred at 25 °C for 12 hours. LC-MS (EC11562-64-P1A1) showed that Compound 4 was completely consumed, and the content of the target product was 57.8%. The reaction solution was extracted with MTBE (200 mL × 3). The organic phase was dried over Na2SO4, filtered, and concentrated under reduced pressure to obtain the crude product. Compound 6 (yellow solid, 6.00 g, 36.11 mmol, 9.11% yield) was purified by column chromatography (eluent ratio: petroleum ether / ethyl acetate = 1 / 0 - 1 / 1; TLC plate: petroleum ether: ethyl acetate = 1:1, Rf = 0.30).

[0015] Step 2: Synthesis of Compound 7 from Compound 6: POCl3 (1.85 g, 12.0 mmol, 1.12 mL, 2.00 eq) was added to a DMF (20.0 mL) solution of Compound 6 (1.00 g, 6.02 mmol, 1.00 eq), and the mixture was stirred at 25 °C for 1 hour. Then, H2O (20.0 mL) was added. The mixture was then stirred at 80 °C for 1 hour. LC-MS (EC14440-1-P1A1) showed that Compound 6 was completely consumed and the main product peak was formed. The reaction mixture was diluted with water (50 mL), then extracted with EtOAc (50 mL × 3). The combined organic phases were washed with water (50 mL), saturated brine (50 mL), dried over anhydrous Na2SO4, filtered, and concentrated under reduced pressure to obtain Compound 7 (yellow oil, 1.10 g, 5.66 mmol, yield 94.13%).

[0016] Step 3: Synthesis of Compound 10 from Compound 7: Compound 7 (500 mg, 2.57 mmol, 1.00 eq), Compound 9 (515 mg, 2.83 mmol, 1.10 eq), TEA (1.43 g, 14.1 mmol, 1.97 mL, 5.50 eq), and Ac₂O (1.58 g, 15.4 mmol, 1.45 mL, 6.00 eq) were added to a three-necked round-bottom flask. The mixture was stirred at 115 °C for 8 hours. The mixture was then quenched with water (30.0 mL), precipitating a large amount of solid. The solid was then filtered, and the filter cake was washed three times with distilled water (20.0 mL × 3). The solid was then dissolved in ethanol (5.00 mL), and 20% HCl aqueous solution (5.00 mL) was added. The mixture was then stirred at 80 °C for 3 hours. LC-MS (EC14091-7-P1A2) showed that compound 7 was completely consumed, with a main peak detected. After the reaction was complete, the reaction mixture was quenched with ice water (20 mL), precipitating a large amount of solid. The solid was filtered, and the filter cake was washed with water until the pH of the filtrate was close to neutral. The filter cake was concentrated under reduced pressure to give compound 10 (yellow solid, 850 mg, 1.87 mmol, 75.9% purity, 72.8% yield).

[0017] Application of intermediate compounds (compound 8 and compound 10) in the preparation of drugs that inhibit the RNA polymerase of the novel coronavirus.

[0018] Application of intermediate compounds (compound 8 and compound 10) in the preparation of anti-influenza virus drugs.

[0019] The structural formula of the new compound is as follows:

[0020] The synthesis of compound C includes the following steps: Compound C is prepared from compound 8 as described above. Specifically, under a nitrogen atmosphere, compound 8 (500 mg, 1.61 mmol, 1.00 eq) is added to an 8.00 mL MeOH solution of Pd / C (342 mg, 10.0% purity, commercial name Adamas-beta 80517D, CAS: 7440-05-3). The suspension is purged three times. The reaction mixture is stirred at 50 °C for 12 hours under hydrogen (50 Psi), and then at 55 °C for 24 hours. LC-MS (EC14509-1-P1A) shows that compound 8 is completely consumed and product formation is detected. The reaction mixture is filtered, and the filtrate is concentrated under reduced pressure to obtain the crude product. The compound was purified by Prep-HPLC (column: Phenomenex luna C18 150×25mm×10µm, mobile phase: [water (0.1% hydrochloric acid)-acetonitrile], gradient: 22%-52% acetonitrile, 10 min). Compound C (yellow solid, 40.0 mg, 127 µmol, yield 7.9%) was characterized by LC-MS (EC14509-1-P1B3), OR (EC14059-1-P1A), SFC (EC14509-1-P1B), and ¹H NMR (EC14509-1-P1A).

[0021] The synthesis of compound E includes the following steps: Compound E is prepared from compound 10 as described above. Specifically, Pd / C (200 mg, 10% purity) is added to a MeOH solution (4.00 mL) of compound 10 (400 mg, 1.18 mmol, 1.00 eq). The reaction solution is stirred at 50 °C for 6 hours under a hydrogen atmosphere (H2, 50 psi). LC-MS (EC14091-9-P1A) shows that compound 10 is completely consumed, and the product content is 57.4%. The reaction solution is filtered, and the filtrate is concentrated under reduced pressure to obtain the crude product. Compound E (40 mg, 116.16 μmol, 9.88% yield) was purified by Prep-HPLC (column: Phenomenex luna C18 150×25 mm ×10 µm, mobile phase: [water (0.1% hydrochloric acid)-acetonitrile], gradient: 20%-50% acetonitrile, 10 min). The compound was confirmed by LC-MS (EC11586-105-P1B1), OR (EC11586-105-P1B), SFC (EC11586-105-P1A), and ¹H NMR (EC1586-105-P1A).

[0022] Application of new compounds (compound C and compound E) in the preparation of drugs that inhibit the RNA polymerase of the novel coronavirus.

[0023] Application of new compounds (compound C and compound E) in the preparation of anti-influenza virus drugs.

[0024] Compounds 8, C, 10, and E of this invention can be used alone or in combination with drugs with different mechanisms of action (such as nematvir / cenostatin / leretevir, oseltamivir, etc.) in the treatment of novel coronavirus and influenza virus (experimentally tested as type A).

[0025] Compounds 8, C, 10, and E of this invention are used as follows: For mild / moderate cases of COVID-19 infection (diagnosed as mild / moderate according to the 10th edition of the COVID-19 treatment guidelines, characterized by a respiratory rate <30 breaths / min, oxygen saturation >93% at rest, and characteristic COVID-19 pneumonia manifestations on imaging), they can be used alone. For moderate / severe / critical cases (diagnosed as moderate / severe / critical according to the 10th edition of the COVID-19 treatment guidelines, characterized by a respiratory rate ≥30 breaths / min, oxygen saturation ≤93% at rest, characteristic COVID-19 pneumonia manifestations with significant lesion progression >50% within 24-48 hours on imaging, respiratory failure requiring mechanical ventilation, or other organ failure), they can be used in combination with main protease (Mpro) inhibitors such as nematvir / cenostatin / lereitvir.

[0026] For mild / moderate influenza virus (H1N1) infection (based on the 2025 edition of the Emergency Expert Consensus on the Diagnosis and Treatment of Adult Influenza, patients are classified as mild / moderate if they only present with upper respiratory tract infection, have cough, shortness of breath but respiratory rate <30 breaths / min, oxygen saturation >93% when breathing air at rest, and imaging shows characteristic influenza virus pneumonia), it can be used alone. For moderate / severe / critical cases (based on the 2025 edition of the Emergency Expert Consensus on the Diagnosis and Treatment of Adult Influenza, patients are classified as moderate / severe / critical if they have rapid breathing and respiratory rate ≥30 breaths / min, oxygen saturation ≤93% when breathing air at rest, imaging shows characteristic influenza virus pneumonia with significant lesion progression >50% within 24-48 hours, progressive respiratory failure, multiple organ dysfunction, etc.), it can be used in combination with oseltamivir neuraminidase inhibitors, mabaloxavir endonuclease inhibitors, etc.

[0027] Compared with the prior art, the present invention has the following beneficial effects: This invention synthesizes novel drugs for combating influenza viruses and inhibiting the RNA polymerase of the novel coronavirus.

[0028] This invention is based on a simple small molecule (non-nucleoside inhibitor) that mimics the characteristics of normal substrate nucleotides of RNA polymerase and competitively occupies the nucleotide entry channel of viral RNA polymerase. This competitively blocks the entry of raw materials for viral nucleic acid replication, preventing RNA polymerase from performing its normal biological function. The viral life cycle is blocked, and viral life elements are degraded by the host cell's enzyme system, thereby effectively exerting an antiviral effect. Attached Figure Description

[0029] Figure 1 The HCl spectrum of compound 8; Figure 2 The HCl spectrum of compound C; Figure 3 The mass spectrum of compound 10 is shown below. Figure 4 The 1H spectrum of compound E; Figure 5 This is a diagram showing the binding of compound C-(S) at the active site of RdRp; Figure 6 This is a diagram showing the binding of compound C-(R) at the active site of RdRp; Figure 7 This is a binding diagram of compound E-(S) at the active site of RdRp; Figure 8 This is a diagram showing the binding of compound E-(R) at the active site of RdRp. Detailed Implementation

[0030] The present invention will now be described in further detail with reference to the accompanying drawings and specific embodiments. The following embodiments are for illustrative purposes only and are not intended to limit the scope of the invention.

[0031] I. Pharmacological activity of inhibiting novel coronavirus RNA polymerase 1. Establishment of RdRp catalytic reaction system for novel coronavirus and detection of cell activity HEK293T cells were transfected with plasmids pCoV-Gluc, pCOVID19-nsp12, pCOVID19-nsp7, and pCOVID 19-nsp8 at a ratio of 1:20:60:60. Twelve hours after transfection, the cells were reseeded in 96-well plates (10T). 4Cells per well were added sequentially to each well using culture medium containing remdesivir, compound 8, compound C, compound 10, and compound E, respectively. Fluorescence values ​​were measured 24 hours later using the Gasussia luciferase activity assay. Inhibition rate (%) = [1 - (experimental group average / control group average)] × 100%. The results shown are the average of three independent experiments. Cell viability was also assessed according to the instructions of the CCK-8 Cell Counting Kit (Vazyme, cat# A311).

[0032] 2. Experimental Results Table 1 shows the results of the RdRp inhibition rate and CCK-8 cell activity assay obtained in the experiment.

[0033] In HEK293T cells, plasmids pCoV-Gluc, pCOVID19-nsp12, pCOVID19-nsp7, and pCOVID19-nsp8 expressed their respective target proteins and combined to form the novel coronavirus RNA polymerase replication system. This system recognizes and transcribes the substrate Gluc RNA, ultimately increasing Gluc protein expression. Targeting and inhibiting the core functional element nsp12 of RNA polymerase reduces Gluc protein expression. Remdesivir, compound 8, compound C, compound 10, and compound E are all targeted inhibitors of nsp12, mainly occupying the nucleotide substrate entry channel in nsp12 to inhibit its biological function. Remdesivir can be incorporated into the nascent RNA chain and further terminate the function of nsp12 in elongating the nascent RNA chain. This experiment found that remdesivir, compound 8, compound C, compound 10, and compound E all significantly and in a dose-dependent manner inhibited the novel coronavirus RdRp. Specific experimental data are shown in Table 1. Existing technology suggests that the novel coronavirus exonuclease NSP14 / NSP10 complex performs nucleic acid proofreading and effectively removes "mismatched" nucleotide analog inhibitors in the extended chain, significantly reducing the inhibitory activity of nucleotide analogs such as remdesivir and favipiravir. However, the novel coronavirus exonuclease NSP14 / NSP10 system was not introduced into the pharmacological model of this experiment; therefore, the measured inhibitory activity of the positive control remdesivir was higher than the real-world values. Furthermore, compounds 8, C, 10, and E showed lower cytotoxicity than remdesivir. In particular, compound E, with lower toxicity, exhibited stronger antiviral effects at a concentration of 50 μmol / L than remdesivir at a concentration of 25 μmol / L.

[0034] II. Pharmacological activity in inhibiting influenza virus Preliminary screening of anti-IAV compounds was conducted using 293T-GLUC cells as the viral host. The activity of the samples in inhibiting the luciferase activity carrying the viral reporter gene was measured. The specific experimental procedure is as follows: 293T-GLUC cells were seeded in 96-well plates, with 2.5 × 10⁶ cells per well. 4 Cells were cultured in 100 μL of DMEM medium containing 10% FBS and incubated at 37°C for 24 hours with 5% CO2. Drug was added before incubation for 2 hours, and then the viral protovirus was diluted and inoculated with the virus at an MOI of 0.3. After 24 hours of culture, 10 μL of supernatant was taken from each cell to measure the luciferase activity in the infected cells and the inhibition rate of each sample was calculated. The results shown are the average of three independent experiments.

[0035] Table 2 shows the results of the influenza virus inhibition rate and CCK-8 cell activity assay in the experiment.

[0036] III. Chemical Synthesis of the Target Compound The synthetic route for compound C is as follows:

[0037] Step 1: Synthesize compound 6 from compound 4:

[0038] Compound 5 (52.7 g, 436 mmol, 1.10 eq) and sodium hydroxide (15.8 g, 396 mmol, 1.00 eq) were added to an aqueous solution (500 mL) of compound 4 (50.0 g, 396 mmol, 1.00 eq), and the mixture was stirred at 25 °C for 12 hours. LC-MS (EC11562-64-P1A1) showed that compound 4 was completely consumed, and the content of the target product was 57.8%. The reaction mixture was extracted with MTBE (200 mL × 3). The organic phase was dried over Na2SO4, filtered, and concentrated under reduced pressure to obtain the crude product. Compound 6 (yellow solid, 6.00 g, 36.11 mmol, 9.11% yield) was purified by column chromatography (eluent ratio: petroleum ether / ethyl acetate = 1 / 0 - 1 / 1; TLC plate: petroleum ether: ethyl acetate = 1:1, Rf = 0.30) to obtain the crude product.

[0039] LC-MS of compound 6: EC11562-64-P1A1, [M+H] + : 167.2.

[0040] Step 2: Synthesize compound 7 from compound 6:

[0041] POCl3 (1.85 g, 12.0 mmol, 1.12 mL, 2.00 eq) was added to a DMF (20.0 mL) solution of compound 6 (1.00 g, 6.02 mmol, 1.00 eq), and the mixture was stirred at 25 °C for 1 hour. Then, H2O (20.0 mL) was added. The mixture was then stirred at 80 °C for 1 hour. LC-MS (EC14440-1-P1A1) showed that compound 6 was completely consumed and the main product peak was formed. The reaction mixture was diluted with water (50 mL), extracted with EtOAc (50 mL × 3), and the combined organic phases were washed with water (50 mL), saturated brine (50 mL), dried over anhydrous Na2SO4, filtered, and concentrated under reduced pressure to give compound 7 (yellow oil, 1.10 g, 5.66 mmol, yield 94.13%).

[0042] LC-MS of compound 7: EC14440-1-P1A1, [M+H] + : 195.2.

[0043] Step 3: Synthesize compound 8 from compound 7:

[0044] Compound 7 (500 mg, 2.57 mmol, 1.00 eq), compound 2 (430 mg, 2.83 mmol, 1.10 eq), and TEA (1.43 g, 14.1 mmol, 1.97 mL, 5.50 eq) were added to a three-necked round-bottom flask, followed by Ac₂O (1.58 g, 15.4 mmol, 1.45 mL, 6.00 eq). The mixture was stirred at 115 °C for 8 hours. The mixture was then quenched with water (30.0 mL), resulting in the precipitation of a large amount of solid. The solid was then filtered, and the filter cake was washed three times with distilled water (20.0 mL × 3). The solid was then dissolved in ethanol (5.00 mL), followed by the addition of 20% HCl aqueous solution (5.00 mL), and stirred at 80 °C for 3 hours. LC-MS (EC11586-104-P1C) showed that compound 7 was completely consumed, and the target product was a single main peak. Ice water (20 mL) was added to the reaction solution, and a large amount of solid precipitated. The mixture was filtered, and the filter cake was washed with water until the pH of the filtrate was close to neutral. The filter cake was concentrated under reduced pressure to obtain compound 8 (yellow solid, 500 mg, 1.61 mmol, yield 62.5%), and its structure was confirmed by 1H NMR (EC11586-104-P1C).

[0045] LC-MS of compound 8: EC11586-104-P1C, [M+H] + : 310.9.

[0046] ¹H NMR of compound 8: EC11586-104-P1C, (400 MHz, DMSO-d6). δ 10.47 (s, 1H), 10.29 (s, 1H), 9.58 (s, 1H), 7.96 (s, 1H), 7.50 (d, J =8.4 Hz, 2H), 6.80 (d, J = 8.4 Hz, 2H), 6.41 (s, 1H), 5.94 - 5.87 (m, 1H), 4.96 -4.92 (m, 2H), 3.35 (s, 2H).

[0047] The H NMR spectrum of compound 8 is shown below. Figure 1 .

[0048] Step four: Synthesize compound C from compound 8:

[0049] Compound 8 (500 mg, 1.61 mmol, 1.00 eq) was added to 8.00 mL of Pd / C (342 mg, 10.0% purity) MeOH solution under a nitrogen atmosphere. The suspension was purged three times. The reaction mixture was stirred at 50 °C for 12 h under hydrogen (50 Psi) atmosphere, followed by stirring at 55 °C for 24 h. LC-MS (EC14509-1-P1A) showed that compound 8 was completely consumed and product formation was detected. The reaction mixture was filtered, and the filtrate was concentrated under reduced pressure to obtain the crude product. Purification was performed using Prep-HPLC (column: Phenomenex luna C18 150×25 mm×10 µm, mobile phase: [water (0.1% hydrochloric acid)-acetonitrile], gradient: 22%-52% acetonitrile, 10 min). Compound C (yellow solid, 40.0 mg, 127 µmol, yield 7.9%) was characterized by LC-MS (EC14509-1-P1B3), OR (EC14059-1-P1A), SFC (EC14509-1-P1B), and ¹H NMR (EC14509-1-P1A).

[0050] LC-MS of compound C: EC14509-1-P1A, [M+H] + : 314.9.

[0051] ¹H NMR of compound C: EC14509-1-P1A, (400 MHz, DMSO-d6) δ 9.46 - 9.27 (m, 2H), 9.23 (s, 1H), 7.05 (d, J = 8.0 Hz, 2H), 6.69 (d,J = 8.0 Hz, 2H), 6.21 (s, 1H), 3.98 - 3.86 (m, 1H), 3.03 - 2.87 (m, 2H), 2.44 (t,J = 8.0 Hz, 2H), 1.54 - 1.24 (m, 2H), 0.85 (t,J = 8.0 Hz, 3H).

[0052] The H NMR of compound C is shown in [reference needed]. Figure 2 .

[0053] The synthetic route for compound E is as follows:

[0054] Step 1: Synthesize compound 10 from compound 7:

[0055] Compound 7 (500 mg, 2.57 mmol, 1.00 eq), compound 9 (515 mg, 2.83 mmol, 1.10 eq), TEA (1.43 g, 14.1 mmol, 1.97 mL, 5.50 eq), and Ac₂O (1.58 g, 15.4 mmol, 1.45 mL, 6.00 eq) were added to a three-necked round-bottom flask. The mixture was stirred at 115 °C for 8 hours. The mixture was then quenched with water (30.0 mL), resulting in the precipitation of a large amount of solid. The solid was then filtered, and the filter cake was washed three times with distilled water (20.0 mL × 3). The solid was then dissolved in ethanol (5.00 mL), followed by the addition of 20% HCl aqueous solution (5.00 mL), and stirred at 80 °C for 3 hours. LC-MS (EC14091-7-P1A2) showed that compound 7 was completely consumed, with a single main peak detected. After the reaction was complete, the reaction mixture was quenched with ice water (20 mL), and a large amount of solid precipitated. The mixture was filtered, and the filter cake was washed with water until the pH of the filtrate was close to neutral. The filter cake was concentrated under reduced pressure to obtain compound 10 (yellow solid, 850 mg, 1.87 mmol, 75.9% purity, yield 72.8%).

[0056] LC-MS of compound 10: EC14091-7-P1A2, [M+H] + 340.9, for details as follows Figure 3 As shown.

[0057] Step 2: Synthesize compound E from compound 10:

[0058] Pd / C (200 mg, 10% purity) was added to a 4.00 mL MeOH solution of compound 10 (400 mg, 1.18 mmol, 1.00 eq). The reaction mixture was stirred at 50 °C for 6 hours under a hydrogen atmosphere (H2, 50 psi). LC-MS (EC14091-9-P1A) showed that compound 10 was completely consumed, with a product purity of 57.4%. The reaction mixture was filtered, and the filtrate was concentrated under reduced pressure to obtain the crude product. Compound E (40 mg, 116.16 μmol, 9.88% yield) was purified by Prep-HPLC (column: Phenomenex luna C18 150×25 mm ×10 µm, mobile phase: [water (0.1% hydrochloric acid)-acetonitrile], gradient: 20%-50% acetonitrile, 10 min). The compound was confirmed by LC-MS (EC11586-105-P1B1), OR (EC11586-105-P1B), SFC (EC11586-105-P1A), and ¹H NMR (EC1586-105-P1A).

[0059] like Figure 4 As shown, the 1H NMR data for compound E are as follows: 1H NMR: EC11586-105-P1A, (400 MHz, DMSO- d6 ) δ 9.36 (s, 1H), 9.22 (s, 1H), 8.89 (s, 1H), 6.83 (d, J = 8.0 Hz, 1H), 6.72 - 6.59 (m, 2H), 6.20 (s, 1H), 3.93 - 3.88 (m, 1H), 3.73 (s, 3H), 3.01 - 2.87 (m, 2H), 2.44 (t, J = 8.0 Hz, 2H), 1.48 - 1.40 (m, 2H), 0.86 (t, J = 7.6 Hz, 3H).

[0060] LC-MS of compound E: EC14091-9-P1A, [MH] + : 342.8.

[0061]

[0062] Molecular docking steps: (1) Download the three-dimensional structure of the novel coronavirus RNA polymerase (ID: 7VB2) from the PDB database, process the protein structure with Mgtools 1.5.6, and save it as a pdbqt file after adding hydrogen, calculating charge, and combining nonpolar hydrogen. (2) Process the structure of the small molecule with Chemidraw 14.0, perform energy minimization optimization using MM2 force field and save it as a mo12 format, then process it with Mgtools 1.5.6, and save it as a pdbqt file after adding hydrogen, calculating charge, and combining nonpolar hydrogen. (3) Autodock Vina docking processing: Construct a 60×60×60 box centered on the active site of the protein (center x=92.517, center y=92.906, center z=101.245), and generate a pdbqt file through Vina operation. The generated pdbqt file was opened using Pymol 1.7.2.1 software, and the 7BV2 protein structure was opened, saved as a complex pdb file, and relevant plotting was performed.

[0063] Results of molecular docking experiments: Compound 8 enters the active site of the NTPs channel of RdRp and interacts with the amino acids (LYS545, ILE548, ARG555, VAL557, THR687, ASN691, etc.) at the active site of RdRp in a non-covalent manner. The carbonyl oxygen atom of the ILE548 skeleton forms a hydrogen bond with the hydrogen atom of the 4′ hydroxyl group, with a bond length of 2.431 Å and a free energy of 6.8 kcal / mol released by molecular docking.

[0064] Compound 10 enters the active site of the NTPs channel in RdRp and interacts with the amino acids (LYS545, ALA547, ILE548, ARG555, ARG836, etc.) at the active site of RdRp in a non-covalent manner. No hydrogen bonds were formed between the ligand and the acceptor. The free energy released by molecular docking is 7.1 kcal / mol.

[0065] like Figure 5As shown, compound C-(S) enters the active site of the NTPs channel of RdRp and interacts with amino acids (TRP617, ASP618, TYR619, PRO620, LYS621, CYS622, ASP623, ASP760, ASP761, etc.) at the active site of RdRp in a non-covalent manner. In CYS622, the hydrogen atom in the skeleton amide forms a hydrogen bond with the oxygen atom in the 5-position hydroxyl group (bond length 1.842 Å), and in ASP760, the oxygen atom in the carbonyl group forms a hydrogen bond with the hydrogen atom in the 4′ position hydroxyl group (bond length 2.630 Å). The free energy released by molecular docking is 6.6 kcal / mol.

[0066] like Figure 6 As shown, compound C-(R) enters the active site of the NTP channel of RdRp and interacts with the amino acids (ARG555, TRP617, ASP618, TYR619, LYS621, ASP623, ASP760, ASP761, etc.) at the active site of RdRp in a non-covalent manner. The hydrogen atom in the skeletal amide of TYR619 forms a hydrogen bond with the oxygen atom in the 5-position hydroxyl group (bond length 2.192 Å), and the oxygen atom in the carbonyl group of ASP760 forms a hydrogen bond with the hydrogen atom in the 5-position hydroxyl group (bond length 2.781 Å). The free energy released by molecular docking is 7.0 kcal / mol.

[0067] like Figure 7 As shown, compound E-(S) enters the active site of the NTPs channel of RdRp and interacts with amino acids (LYS551, ASP618, TYR619, CYS622, ASP760, ASP761, GLU811, etc.) at the active site of RdRp in a non-covalent manner. The oxygen atom in the carbonyl group of the TYR619 skeleton forms a hydrogen bond with the hydrogen atom in the 5-position hydroxyl group (bond length 2.436 Å), and the hydrogen atom in the amide skeleton of TYR619 forms a hydrogen bond with the oxygen atom in the 5-position hydroxyl group (bond length 2.191 Å). The free energy released by molecular docking is 6.7 kcal / mol.

[0068] like Figure 8As shown, compound E-(R) enters the active site of the NTPs channel of RdRp and interacts with amino acids (ARG553, TRP617, ASP618, TYR619, LYS621, CYS622, ASP623, ASP760, ASP761, GLU811, etc.) at the active site of RdRp in a non-covalent manner. The hydrogen atom in the skeletal amide of TYR619 forms a hydrogen bond with the oxygen atom in the 5-position hydroxyl group (bond length 2.128 Å), the oxygen atom in the carbonyl group at the end of ASP760 forms a hydrogen bond with the hydrogen atom in the 5-position hydroxyl group (bond length 2.096 Å), and the oxygen atom in the carbonyl group at the end of ASP623 forms a hydrogen bond with the hydrogen atom in the 3′ position hydroxyl group (bond length 1.973 Å). The free energy released by molecular docking is 7.3 kcal / mol.

[0069] It should be understood that, in order to simplify this disclosure and aid in understanding one or more of the various aspects of the invention, features of the invention are sometimes grouped together in a single embodiment, figure, or description thereof in the above description of exemplary embodiments of the invention. However, this method of disclosure should not be interpreted as reflecting an intention that the claimed invention requires more features than expressly recited in each claim. Rather, as reflected in the claims, inventive aspects lie in fewer than all the features of the foregoingly disclosed embodiments. Therefore, the claims, following the detailed description, are hereby expressly incorporated into that detailed description, wherein each claim itself is a separate embodiment of the invention.

[0070] Although the invention has been described with reference to a limited number of embodiments, those skilled in the art will understand from the foregoing description that other embodiments are conceivable within the scope of the invention described herein. Furthermore, it should be noted that the language used in this specification has been chosen primarily for readability and edibility purposes, and not for the purpose of interpreting or limiting the subject matter of the invention. Therefore, many modifications and variations will be apparent to those skilled in the art without departing from the scope and spirit of the appended claims. The disclosure of the invention is illustrative and not restrictive, and the scope of the invention is defined by the appended claims.

[0071] The above description is only a preferred embodiment of the present invention. It should be noted that for those skilled in the art, several improvements and modifications can be made without departing from the principle of the present invention, and these improvements and modifications should also be considered within the scope of protection of the present invention.

Claims

1. A novel compound, characterized in that, Its structural formula is as follows: 。 2. The method of synthesis of novel compounds as claimed in claim 1, wherein, The method for synthesizing compound C includes the following steps: ； Under a nitrogen atmosphere, 500 mg, 1.61 mmol, and 1.00 eq of compound 8 were added to 8.00 mL of 342 mg of 10.0% pure Pd / C MeOH solution. The suspension was purged with gas three times. The reaction solution was stirred at 50 °C for 12 hours under 50 Psi hydrogen atmosphere, and then stirred at 55 °C for 24 hours. After all compound 8 was consumed, the reaction solution was filtered, and the filtrate was concentrated under reduced pressure to obtain the crude product. The crude product was purified by Prep-HPLC to obtain compound C. The Prep-HPLC purification process was as follows: column: Phenomenex luna C18 150mm×25mm×10µm, mobile phase: 0.1% hydrochloric acid aqueous solution-acetonitrile, gradient: 22%-52% acetonitrile, elution for 10 min.

3. The method for synthesizing the new compound according to claim 1, characterized in that, The method for synthesizing compound E includes the following steps: ； 200 mg of 10% pure Pd / C was added to a 4.00 mL MeOH solution containing 400 mg of compound 10 at 1.18 mmol and 1.00 eq. The reaction solution was stirred at 50 °C for 6 hours under a 50 psi H2 hydrogen atmosphere. After all compound 10 was consumed, the reaction solution was filtered, and the filtrate was concentrated under reduced pressure to obtain the crude product. The crude product was purified by Prep-HPLC to obtain compound E. The Prep-HPLC purification process was as follows: column: Phenomenex luna C18 150 mm × 25 mm × 10 µm, mobile phase: 0.1% hydrochloric acid aqueous solution-acetonitrile, gradient: 20%-50% acetonitrile, elution for 10 min.

4. The use of the novel compound according to claim 1 in the preparation of a drug that inhibits RNA-dependent RNA polymerase of the novel coronavirus.

5. The use of the novel compound according to claim 1 in the preparation of drugs against influenza A virus.

Images