WO2018028541A1 - Substituted quinoline compound and pharmaceutical composition and use thereof - Google Patents

Abstract

Provided is a substituted quinoline compound and a pharmaceutical composition and a use thereof.The quinoline compound is a compound as shown in formula (I), or a crystal form, a pharmaceutically acceptable salt, a hydrate or a solvate thereof. The compound can be used as a hepatitis C virus inhibitor and has better hepatitis C virus protein NS3/4A inhibitory activity, has better pharmacodynamic/pharmacokinetic properties, has good applicability and a high level of safety, and can be used in the preparation of drugs for treating hepatitis C virus infection, and has good market development prospects.

Description

Substituted quinoline compound and pharmaceutical composition thereof and application thereof Technical field

The invention belongs to the technical field of medicine, and in particular relates to a hepatitis C virus inhibitor, a pharmaceutical composition and application thereof.

Background technique

Hepatitis C is a blood-borne infectious liver disease that, if left untreated, can cause significant damage to the liver. About 185 million people worldwide are infected with hepatitis C virus. The rate of hepatitis C infection in China is about 3.2%, and the number of patients is about 40 million. In the United States, there are about 3.2 million hepatitis C patients, and about 15,000 people die each year. Mostly die from hepatitis C-related diseases such as cirrhosis and liver cancer.

HCV (Hepatitis C Virus) is an RNA virus belonging to the genus Hepacivirus genus in the Flaviviridae family. The encapsulated HCV virion contains a positive-stranded RNA genome that encodes all known virus-specific proteins in a single uninterrupted open reading frame. The open reading frame comprises approximately 9500 nucleotides and encodes a single large polyprotein of approximately 3000 amino acids. Polyproteins include core proteins, envelope proteins E1 and E2, membrane-bound protein P7, and non-structural proteins NS2, NS3/4A, NS4A, NS4B, NS5A, and NS5B.

Sofabru is the first medicine in the world that can completely cure hepatitis C in the short term. It takes the oral route directly to the lesion, and the method has simple side effects and is highly sought after by patients. Sofibuvir is produced by Gilead, USA, and is marketed in the United States in 2013. It has been clinically proven to be effective in treating hepatitis C, type 1, 2, 3, and 4, including liver transplantation, liver cancer, and HCV/HIV-1 co-infection. Clinical Trials. This breakthrough has brought the gospel to hepatitis C patients around the world.

HCV infection is associated with progressive liver disease, including cirrhosis and hepatocellular carcinoma. Simeprevir is a new generation of NS3/4A protease inhibitors, a daily oral medication developed by Medivir and Janssen for the treatment of compensatory liver disease in adult patients with chronic hepatitis C. Including liver fibrosis at various stages, its working principle is to inhibit the replication of HCV in liver cells by blocking proteases. On January 22, 2013, hepatitis C, a new drug, dexamethavir was approved by the FDA, combined with peginterferon and ribavirin for genotype 1 chronic hepatitis C adult patients with compensatory liver disease ( Treatment including cirrhosis.

Therefore, there is still a need in the art to develop compounds having inhibitory activity or better pharmacodynamic properties against the hepatitis C virus protein NS3/4A.

Summary of the invention

In view of the above technical problems, the present invention discloses a hepatitis C virus inhibitor, a pharmaceutical composition and use thereof, which have better hepatitis C virus protein NS3/4A inhibitory activity and/or have better pharmacodynamics/pharmacokinetics. Kinetic performance.

In this regard, the technical solution adopted by the present invention is:

A hepatitis C virus inhibitor, such as a quinoline compound of the formula (I), or a crystalline form thereof, a pharmaceutically acceptable salt, a hydrate or a solvent compound,

Wherein R ¹ , R ² , R ³ , R ⁴ , R ⁵ , R ⁶ , R ⁷ , R ⁸ , R ⁹ , R ¹⁰ , R ¹¹ , R ¹² , R ¹³ , R ¹⁴ , R ¹⁵ , R ¹⁶ , R ¹⁷ , R ¹⁸ , R ¹⁹ , R ²⁰ , R ²¹ , R ²² , R ²³ , R ²⁴ , R ²⁵ , R ²⁶ , R ²⁷ , R ²⁸ , R ²⁹ , R ³⁰ , R ³¹ , R ³² , R ³³ And R ³³ , R ³⁴ , R ³⁵ , R ³⁶ , R ³⁷ , R ³⁸ , R ³⁹ , R ⁴⁰ , R ⁴¹ , R ⁴² , R ⁴³ , R ⁴⁴ , R ⁴⁵ are each independently hydrogen, deuterium, halogen or trisole Fluoromethyl;

Additional conditions are R ¹ , R ² , R ³ , R ⁴ , R ⁵ , R ⁶ , R ⁷ , R ⁸ , R ⁹ , R ¹⁰ , R ¹¹ , R ¹² , R ¹³ , R ¹⁴ , R ¹⁵ , R ¹⁶ And R ¹⁷ , R ¹⁸ , R ¹⁹ , R ²⁰ , R ²¹ , R ²² , R ²³ , R ²⁴ , R ²⁵ , R ²⁶ , R ²⁷ , R ²⁸ , R ²⁹ , R ³⁰ , R ³¹ , R ³² , R ^{33. At least one} of R ³³ , R ³⁴ , R ³⁵ , R ³⁶ , R ³⁷ , R ³⁸ , R ³⁹ , R ⁴⁰ , R ⁴¹ , R ⁴² , R ⁴³ , R ⁴⁴ and R ⁴⁵ is deuterated.

With this technical solution, the shape and volume of the ruthenium in the drug molecule are substantially the same as those of the hydrogen. If the hydrogen in the drug molecule is selectively replaced with hydrazine, the deuterated drug generally retains the original biological activity and selectivity. At the same time, the inventors have confirmed through experiments that the binding of carbon-germanium bonds is more stable than the combination of carbon-hydrogen bonds, which can directly affect the absorption, distribution, metabolism and excretion of some drugs, thereby improving the efficacy, safety and tolerability of the drugs.

Preferably, the strontium isotope content of the cerium in the deuterated position is at least greater than the natural strontium isotope content (0.015%), preferably greater than 30%, more preferably greater than 50%, more preferably greater than 75%, and even more preferably greater than 95. %, more preferably greater than 99%.

Specifically, in the present invention, R ¹ , R ² , R ³ , R ⁴ , R ⁵ , R ⁶ , R ⁷ , R ⁸ , R ⁹ , R ¹⁰ , R ¹¹ , R ¹² , R ¹³ and R ¹⁴ , R ¹⁵ , R ¹⁶ , R ¹⁷ , R ¹⁸ , R ¹⁹ , R ²⁰ , R ²¹ , R ²² , R ²³ , R ²⁴ , R ²⁵ , R ²⁶ , R ²⁷ , R ²⁸ , R ²⁹ , R ³⁰ , R ³¹ And R ³² , R ³³ , R ³³ , R ³⁴ , R ³⁵ , R ³⁶ , R ³⁷ , R ³⁸ , R ³⁹ , R ⁴⁰ , R ⁴¹ , R ⁴² , R ⁴³ , R ⁴⁴ and R ^{45 are} each in the metamorphic position The strontium isotope content is at least 5%, preferably more than 10%, more preferably more than 15%, more preferably more than 20%, more preferably more than 25%, more preferably more than 30%, more preferably more than 35%, More preferably more than 40%, more preferably more than 45%, more preferably more than 50%, more preferably more than 55%, more preferably more than 60%, more preferably more than 65%, more preferably more than 70%, more Preferably, it is greater than 75%, more preferably greater than 80%, more preferably greater than 85%, more preferably greater than 90%, more preferably greater than 95%, and even more preferably greater than 99%.

Preferably, R ¹ , R ² , R ³ , R ⁴ , R ⁵ , R ⁶ , R ⁷ , R ⁸ , R ⁹ , R ¹⁰ , R ¹¹ , R ¹² , R ¹³ , R of the compound of formula (I) ¹⁴ , R ¹⁵ , R ¹⁶ , R ¹⁷ , R ¹⁸ , R ¹⁹ , R ²⁰ , R ²¹ , R ²² , R ²³ , R ²⁴ , R ²⁵ , R ²⁶ , R ²⁷ , R ²⁸ , R ²⁹ , R ³⁰ , R ³¹ , R ³² , R ³³ , R ³³ , R ³⁴ , R ³⁵ , R ³⁶ , R ³⁷ , R ³⁸ , R ³⁹ , R ⁴⁰ , R ⁴¹ , R ⁴² , R ⁴³ , R ⁴⁴ and R ⁴⁵ , at least One R contains ruthenium, more preferably two R contains ruthenium, more preferably three R contains ruthenium, more preferably four R contains ruthenium, more preferably five R contains ruthenium, more preferably six R contains ruthenium, more preferably six R-containing ruthenium, More preferably, seven R contains ruthenium, more preferably eight R contains ruthenium, more preferably nine R contains ruthenium, more preferably ten R contains ruthenium, more preferably eleven R contains ruthenium, more preferably ten The two Rs contain ruthenium, more preferably thirteen R 氘, more preferably fourteen R 氘, more preferably fifteen R 氘, more preferably sixteen R 氘, more preferably ten The seven Rs contain ruthenium, more preferably eighteen R 氘, more preferably nineteen R 氘, more preferably twenty R 氘, more preferably twenty-one R 氘, more preferably Twenty-two R contains 氘, better two The three Rs contain 氘, preferably twenty-four R 氘, more preferably twenty-five R 氘, more preferably twenty-six R 氘, more preferably twenty-seven R 氘, More preferably twenty-eight R 氘, more preferably twenty-nine R 氘, more preferably thirty R 氘, more preferably thirty-one R 氘, more preferably thirty-two R contains 氘, more preferably thirty-three R 氘, more preferably thirty-four R 氘, more preferably thirty-five R 氘, more preferably thirty-six R 氘, better Thirty-seven R contains 氘, more preferably thirty-eight R contains 氘, more preferably thirty-nine R contains 氘, more preferably forty R contains 氘, more preferably forty-one R Preferably, forty-two R 氘, more preferably forty-three R 氘, more preferably forty four R 氘, more preferably forty-five R 氘.

As a further improvement of the present invention, R ¹⁹ , R ²⁰ and R ²¹ are each independently hydrazine or hydrogen.

In another preferred embodiment, R ¹⁹ , R ²⁰ and R ²¹ are 氘.

As a further improvement of the present invention, R ³² , R ³³ and R ³⁴ are each independently hydrazine or hydrogen.

In another preferred embodiment, R ³² , R ³³ , and R ³⁴ are deuterium.

As a further improvement of the present invention, R ³⁵ , R ³⁶ and R ³⁷ are each independently hydrazine or hydrogen.

In another preferred embodiment, R ³⁵ , R ³⁶ , and R ³⁷ are 氘.

As a further improvement of the present invention, the compound is selected from the group consisting of the following compounds or a pharmaceutically acceptable salt thereof:

In another preferred embodiment, the compound does not include a non-deuterated compound.

The present invention also discloses a pharmaceutical composition comprising a pharmaceutically acceptable carrier and the hepatitis C virus inhibitor as described above, or a crystalline form, a pharmaceutically acceptable salt, a hydrate or a solvent thereof A pharmaceutical composition of a compound, stereoisomer, prodrug or isotopic variation.

As a further improvement of the present invention, it further comprises other active compounds including, but not limited to, other HCV antiviral agents, anti-infective agents, immunomodulators, antibiotics or vaccine combinations.

As a further improvement of the present invention, the immunomodulator is an interferon drug compound.

As a further improvement of the invention, the quinoxaline macrocycles of the invention may be used in combination therapies involving one or more additional therapeutic agents. Additional therapeutic agents include those that also target HCV, target different pathogenic agents, or enhance the immune system. Agents that enhance the immune system include those that normally enhance the function of the immune system and those that produce a specific immune response against HCV. Additional therapeutic agents that target HCV include agents that target NS3/4A and agents that target other HCV activities, such as NS5A and NS5B, and agents that target host cell activity involved in HCV replication.

Other examples of therapeutic agents that may be present in the combination include ribavirin, levovirin, viramidine, thymosin alpha-1, interferon-beta, interferon-alpha, PEGylated interferon-alpha (peg interferon-alpha) a combination of interferon-[alpha] and ribavirin, a combination of peg interferon-[alpha] and ribavirin, a combination of interferon-[alpha] and levovirin, and a combination of peg interferon-[alpha] and levovirin. Interferon-α includes recombinant interferon-α2a (such as Roferon interferon available from Hoffmann-LaRoche, Nutley, NJ), PEGylated interferon-α2a (Pegasys), interferon-α2b (eg available from Schering Corp) .Kenilworth, NJ's Intron-A interferon), PEGylated interferon-α2b (PegIntron), recombinant complex interferon (such as interferon alphacon-1), and purified interferon-α product. The individual components of the combination may be administered separately at different times during the course of the treatment or simultaneously in separate or individual combinations.

For the treatment of HCV infection, the compounds of the invention may also be administered in combination with the antiviral agent amantadine (1-aminoadamantane). For a full description of the agent, see J. Kirschbaum, 12 Anal. Profiles Drug Subs. 1-36 (1983).

For the treatment of HCV infection, the compounds of the invention may also be administered in combination with the antiviral polymerase inhibitor R7128 (Roche).

For the treatment of HCV infection, the compounds of the invention may also be administered in combination with an HCV NS5B polymerase inhibitor. Such HCV NS5B polymerase inhibitors that can be used as a combination therapy include, but are not limited to, International Patent Application Publication No. WO 02/057287, WO 02/057425, WO 03/068244, WO 2004/000858, WO 04/003138, and WO Those disclosed in U.S. Patent No. 6,777,392 and U.S. Patent Application Publication No. 2004/0067901, the entire contents of each of each of Other such HCV polymerase inhibitors include, but are not limited to, valopicitabine (NM-283; Idenix) and 2'-F-2'-beta-methylcytidine (see also WO 2005/003147).

As a further improvement of the present invention, the pharmaceutically acceptable carrier includes a glidant, a sweetener, a diluent, a preservative, a dye/colorant, a flavor enhancer, a surfactant, a wetting agent, a dispersant At least one of a disintegrant, a suspending agent, a stabilizer, an isotonic agent, a solvent or an emulsifier.

As a further improvement of the present invention, the pharmaceutical composition is a tablet, a pill, a capsule, a powder, a granule, an ointment, an emulsion, a suspension, a solution, a suppository, an injection, an inhalant, a gel, a microsphere or Aerosol.

Typical routes of administration of the pharmaceutical compositions of the invention include, but are not limited to, oral, rectal, transmucosal, enteral, or topical, transdermal, inhalation, parenteral, sublingual, intravaginal, intranasal, intraocular, intraperitoneal , intramuscular, subcutaneous, intravenous administration. Oral administration or injection administration is preferred.

The pharmaceutical composition of the present invention can be produced by a method known in the art, such as a conventional mixing method, a dissolution method, a granulation method, a sugar-coating method, a pulverization method, an emulsification method, a freeze-drying method, and the like.

The present invention also provides a method of preparing a pharmaceutical composition comprising the steps of: administering a pharmaceutically acceptable carrier to a hepatitis C virus inhibitor as described above, or a crystalline form thereof, a pharmaceutically acceptable salt, or a hydrate thereof Or the solvate is mixed to form a pharmaceutical composition.

The invention also discloses the use of a hepatitis C virus inhibitor as described above, characterized in that it is used for the preparation of a medicament for the treatment of hepatitis C virus infection.

NS3/4A inhibitors are also useful in the preparation and implementation of screening assays for antiviral compounds. For example, such compounds can be used to isolate enzyme mutants, which are excellent screening tools for more potent antiviral compounds. In addition, these compounds can be used to establish or measure binding sites for other antiviral agents to HCV protease, for example by competitive inhibition.

In order to inhibit the HCV NS3/4A protease and to treat HCV infection and/or reduce the likelihood or severity of HCV infection symptoms, the compounds of the invention, optionally in salt form, can be administered by contacting the active agent with the site of action of the drug. medicine. They can be administered as a separate therapeutic or combination of therapeutic agents by conventional means which can be used in combination with the drug. They can be administered alone, but they are It is often administered with a pharmaceutical carrier selected according to the chosen route of administration and standard pharmaceutical practice.

The HCV includes a plurality of genotypes thereof and a plurality of gene subtypes, such as 1a, 1b, 2a, 2b, 3a, 3b, 4a, 5a, 6a.

It is to be understood that within the scope of the present invention, the various technical features of the present invention and the various technical features specifically described hereinafter (as in the embodiments) may be combined with each other to constitute a new or preferred technical solution. Due to space limitations, we will not repeat them here.

Herein, "halogen" means F, Cl, Br, and I unless otherwise specified. More preferably, the halogen atom is selected from the group consisting of F, Cl and Br.

As used herein, unless otherwise specified, "deuterated" means that one or more hydrogens in the compound or group are replaced by deuterium; deuteration may be monosubstituted, disubstituted, polysubstituted or fully substituted. The terms "one or more deuterated" are used interchangeably with "one or more deuterated".

As used herein, unless otherwise specified, "non-deuterated compound" means a compound containing a proportion of germanium atoms not higher than the natural helium isotope content (0.015%).

Pharmaceutically acceptable salts include inorganic and organic salts. A preferred class of salts are the salts of the compounds of the invention with acids. Suitable acids for forming salts include, but are not limited to, mineral acids such as hydrochloric acid, hydrobromic acid, hydrofluoric acid, sulfuric acid, nitric acid, phosphoric acid; formic acid, acetic acid, trifluoroacetic acid, propionic acid, oxalic acid, malonic acid, succinic acid, Organic acids such as fumaric acid, maleic acid, lactic acid, malic acid, tartaric acid, citric acid, picric acid, benzoic acid, methanesulfonic acid, ethanesulfonic acid, p-toluenesulfonic acid, benzenesulfonic acid, naphthalenesulfonic acid; Amino acids such as amino acid, phenylalanine, aspartic acid, and glutamic acid. Another preferred class of salts are the salts of the compounds of the invention with bases, such as alkali metal salts (for example sodium or potassium salts), alkaline earth metal salts (for example magnesium or calcium salts), ammonium salts (for example lower alkanolammonium). Salts and other pharmaceutically acceptable amine salts), such as methylamine, ethylamine, propylamine, dimethylamine, trimethylamine, diethylamine, triethylamine, tert-butyl A base amine salt, an ethylenediamine salt, a hydroxyethylamine salt, a dihydroxyethylamine salt, a trihydroxyethylamine salt, and an amine salt formed of morpholine, piperazine, and lysine, respectively.

The term "solvate" refers to a complex of a compound of the invention that is coordinated to a solvent molecule to form a specific ratio. "Hydrate" means a complex formed by the coordination of a compound of the invention with water.

The beneficial effects of the present invention compared to the prior art are: First, the compound of the present invention has excellent inhibitory effect on the hepatitis C virus protein NS3/4A. Second, by deuteration this technique changes the metabolism of the compound in the organism, giving the compound better pharmacokinetic parameter characteristics. In this case, the dosage can be changed and a long-acting preparation can be formed to improve the applicability. Third, replacing the hydrogen atom in the compound with hydrazine increases the drug concentration of the compound in the animal due to its strontium isotope effect, thereby improving the drug efficacy. Fourth, replacing the hydrogen atom in the compound with hydrazine can inhibit certain metabolites and improve the safety of the compound.

detailed description

The preparation of the structural compound of the formula (I) of the present invention is more specifically described below, but these specific methods do not constitute any limitation to the present invention. The compounds of the invention may also optionally combine various synthetic methods described in the specification or known in the art. It is convenient to make such a combination that can be easily carried out by those skilled in the art to which the present invention pertains.

Usually, in the preparation scheme, each reaction is usually carried out in an inert solvent at room temperature to reflux temperature (e.g., 0 ° C to 100 ° C, preferably 0 ° C to 80 ° C). The reaction time is usually from 0.1 to 60 hours, preferably from 0.5 to 24 hours.

Example 1. Synthesis of Intermediate 5.

Step 1. Synthesis of Compound 2.

2.68 g of benzyl bromide was dissolved in 100 mL of acetonitrile, and 4.19 g of cinchonidine was added under nitrogen atmosphere, and the mixture was stirred overnight in the dark. Filtration, the residue was washed three times with EtOAc and EtOAc (EtOAc)EtOAc.

Step 2. Synthesis of Compound 5.

10 g of glycine methyl ester hydrochloride and anhydrous sodium sulfate were dispersed in 200 mL of methyl tert-butyl ether, and benzaldehyde and triethylamine were added successively, and the mixture was stirred at room temperature overnight. Filter and concentrate and the resulting oily liquid was used directly in the next step.

435 mg of the compound 2 was dispersed in 14 mL of toluene, and the above-mentioned 3.98 g of an oily liquid in toluene (16 mL) was added dropwise thereto in an ice water bath, and the mixture was allowed to react at room temperature for 15 minutes. A solution of 4.0 g of 1,4-dibromo-2-butene in toluene (16 mL) was slowly added dropwise and stirred for 45 min. A 20% NaOH solution was slowly added dropwise, and after the completion of the dropwise addition, the mixture was stirred at 10 ° C overnight. The mixture was separated and the organic layer was filtered, washed with brine, dried ¹ H NMR (400 MHz, CDCl ₃ ) δ 5.71 (ddd, J = 17.1, 10.3, 9.2 Hz, 1H), 5.22 (ddd, J = 17.2, 1.9, 0.7 Hz, 1H), 5.05 (ddd, J = 10.3, 1.8, 0.6 Hz, 1H), 3.73 (s, 3H), 2.08–1.99 (m, 1H), 1.56 (dd, J=7.5, 4.8 Hz, 1H), 1.35 (dd, J=9.3, 4.7 Hz) , 1H).

Example 2. Synthesis of Intermediate 11.

Step 1. Synthesis of Compound 8.

Take N-methyltrifluoroacetamide 5.0 g dissolved in 28 mL DMF, add 60% NaH 1.656 g in a cold water bath, and stir at room temperature for 1 hour. Take 6-bromo-1-hexene 6.42 g dissolved in 25 mL DMF. It was added dropwise to the above solution, and after the completion of the dropwise addition, it was stirred at room temperature at 70 ° C overnight. The reaction mixture was poured into water (200 mL), evaporated, evaporated, evaporated, evaporated

The above yellow oily liquid was dissolved in 50 mL of methanol, and 66 mL of a 25 M KOH solution was added dropwise thereto, and stirred at room temperature overnight. The reaction mixture was extracted with EtOAc (EtOAc)EtOAc. ¹ H NMR (400 MHz, CDCl ₃ ) δ 5.81 (dd, J = 16.9, 10.2, 6.6 Hz, 1H), 5.08 - 4.85 (m, 2H), 2.62 - 2.53 (m, 2H), 2.43 (s, 3H), 2.13–2.01 (m, 2H), 1.66–1.31 (m, 4H), 1.26 (br, 1H).

Step 2. Synthesis of Compound 9.

Take (1R,4R,5R)-3-oxo-2-oxabicyclo[2.2.1]hexane-5-carboxylic acid cinchonidine salt 2.27 g and 0.71 g of compound 8 dissolved in 25 mL DMF, ice bath 2-(7-Azobenzotriazole)-N,N,N',N'-tetramethyluronium hexafluorophosphate (HATU, 2.30 g), N,N-diisopropyl Ethylamine (DIPEA, 0.78 g) was stirred in an ice bath for 1 hour. The reaction mixture was poured into 100 mL of water, and ethyl acetate was evaporated, and ethyl acetate was evaporated. ¹ H NMR (400 MHz, CDCl ₃ ) δ 6.01 - 5.58 (m, 1H), 5.17 - 4.84 (m, 3H), 3.51 - 3.17 (m, 2H), 2.99 (t, J = 8.0 Hz, 2H) , 2.93 (s, 1H), 2.30 - 2.00 (m, 6H), 1.70 - 1.31 (m, 4H). LC-MS (APCI): m/z = 252.3 (M + 1) ⁺ .

Step 3. Synthesis of Compound 10.

1.04 g of the compound 9 was dissolved in 8 mL of tetrahydrofuran, and 4.5 mL of a 1 M LiOH solution was added dropwise thereto in an ice water bath, and the mixture was stirred for 1 hour under ice-cold bath. 4N HCl solution was adjusted to pH 1-2, ethyl acetate was evaporated, and ethyl acetate was evaporated. LC-MS (APCI): m / z = 268.2 (M + 1) +.

Step 4. Synthesis of Compound 11.

163 mg of compound 5 was dissolved in 3 mL of anhydrous DMF, and 253 mg of HATU, 90 mg of compound 10, 287 μL of DIPEA was added under ice bath, and the mixture was stirred for 2 hours under ice bath. The reaction mixture was poured into 20 mL of water and extracted with EtOAc. EtOAc (EtOAc) Yellow oily liquid. LC-MS (APCI): m / z = 393.3 (M + 1) +.

Example 3. Synthesis of Intermediate 17.

Step 1. Synthesis of Compound 13.

2.6 g of methyl isopropyl ketone was dissolved in absolute ethanol, and 4.87 g of liquid bromine was added dropwise to the ice salt bath, and the mixture was reacted at room temperature for 3 hours after the completion of the dropwise addition. Add 60 mL of petroleum ether to the reaction system, wash twice with water, strip the aqueous phase with petroleum ether, combine with petroleum ether, wash twice with ice-cold saturated sodium carbonate solution, wash once with saturated brine, dry over anhydrous sodium sulfate, filter Concentration gave a colorless oily liquid, 3.04 g. ¹ H NMR (300 MHz, CDCl ₃ ) δ 3.99 (s, 2H), 2.99 (hept, J = 6.9 Hz, 1H), 2.44 (s, 0H), 2.14 (s, 0H), 1.94 (s, 0H) ), 1.86 (s, 0H), 1.17 (d, J = 6.9 Hz, 6H).

Step 2. Synthesis of Compound 15.

0.75 g of Compound 13 and 0.49 g of thiooxalinate (Compound 14) were dissolved in absolute ethanol and heated to reflux for 3 hours. The ice water bath was cooled, and the reaction was quenched with ethyl acetate and water. The mixture was combined with ethyl acetate and water, and the mixture was evaporated. Column chromatography gave a pale yellow oily liquid 0.383 g. ^{1 H NMR (300MHz, DMSO-} d6) δ7.72 (s, 1H), 4.35 (q, J = 7.1 Hz, 2H), 3.17-3.05 (m, 1H), 1.31 (t, J = 7.1 Hz, 3H ), 1.25 (d, J = 6.9 Hz, 6H).

Step 3. Synthesis of Compound 16.

383 mg of Compound 15 was dissolved in 1.1 mL of MeOH and 3.6 mL of THF (tetrahydrofuran), and LiOH·H ₂ O 77 mg and 50 μL of water were added, and the mixture was reacted at 50 ° C for 2 hours. The reaction mixture was concentrated, and the mixture was diluted with EtOAc EtOAc EtOAc EtOAc EtOAc. LC-MS (APCI): m / z = 172.0 (m + 1) +.

Step 4. Synthesis of the compound without 17.

261 mg of Compound 16 was dissolved in 3.6 mL of dry dichloromethane, 7.2 μL of DMF (dimethylformamide) was added, and 280 μL of oxalyl chloride was added thereto under ice-cooling, and the mixture was stirred at room temperature for 1 hour. Concentration gave a brownish yellow oily liquid which was taken directly to next.

Example 4. Synthesis of intermediate 21.

Step 1. Synthesis of Compound 19.

Take 5.0 g of 2-methyl-3-methoxyaniline in 55 mL of xylene, add 38.5 mL of 1M BCl ₃ /DCM solution at 0 ° C, react for 5 minutes at 5 ° C, add 2.5 mL of acetonitrile, and react at 5 ° C. In a minute, the reaction solution was transferred to a dropping funnel, and added dropwise to a solution of 5.14 g of aluminum trichloride in 25 mL of xylene at 0 ° C, and reacted at 5 ° C for 45 minutes, then heated, and the separator was separated by dichloromethane at 75 ° C. The reaction was overnight. The mixture was cooled in an ice water bath, and the reaction solution was poured into 10 mL of ice water and refluxed for 2 hours. After cooling to room temperature, the mixture was separated with methylene chloride. EtOAc (EtOAc)EtOAc. The ice-cold isopropyl ether was added thereto, and the mixture was stirred at 0 ° C for 2 hours to precipitate a gray solid, which was filtered, and the residue was washed with ice-cold isopropyl ether. The residue was collected and dried under vacuum at 45 ° C overnight to obtain 1.86 g of a gray solid powder. . ¹ H NMR (500 MHz, CDCl ₃ ) δ 7.66 (d, J = 9.0 Hz, 1H), 6.31 (d, J = 9.0 Hz, 1H), 3.88 (s, 3H), 2.55 (s, 3H), 2.02 (s, 3H).

Step 2. Synthesis of Compound 20.

206 mg of the above-mentioned fresh compound 17 was dissolved in 3 mL of dioxane, and a solution of compound 19 (245 mg) in dioxane (3 mL) was added dropwise, and the mixture was stirred at room temperature for 1 hour. The reaction solution was concentrated, with 3 mL of saturated NaHCO ₃ solution and 3 mL of dichloromethane, and the aqueous phase extracted with dichloromethane. The combined dichloromethane was washed with brine, dried over anhydrous sodium sulfate and filtered and evaporated ¹ H NMR (300 MHz, CDCl ₃ ) δ 11.30 (s, 1H), 7.76 (d, J = 8.7 Hz, 1H), 7.17 (d, J = 0.9 Hz, 1H), 6.79 (d, J = 8.7 Hz, 1H), 3.93 (s, 3H), 3.32 - 3.13 (m, 1H), 2.59 (s, 3H), 2.17 (s, 3H), 1.41 (d, J = 6.9 Hz, 6H).

Step 3. Synthesis of Compound 21.

255 mg of Compound 20 was dissolved in 5 mL of tert-butanol, and 180 mg of potassium t-butoxide was added thereto, followed by heating under reflux for 6 hours. The reaction mixture was concentrated, dichloromethane (DCM) and EtOAc (EtOAc) was evaporated. . ¹ H NMR (300 MHz, CDCl ₃ ) δ 9.54 (s, 1H), 8.24 (d, J = 9.0 Hz, 1H), 7.09 (d, J = 0.9 Hz, 1H), 7.01 (d, J = 9.1 Hz, 1H), 6.72 (d, J = 1.8 Hz, 1H), 3.97 (s, 3H), 3.28 - 3.09 (m, 1H), 2.42 (s, 3H), 1.39 (d, J = 6.9 Hz, 6H) ).

Example 5. Preparation of Substituted Quinoline Compound A-1

The specific synthesis steps are as follows:

Step 1. Synthesis of Compound 23.

5.0 g of 2-methyl-3-hydroxynitrobenzene was dissolved in 40 mL of DMF, 6.30 g of potassium carbonate solid was added, and 2.24 mL of deuterated iodomethane (CD ₃ I) was added under ice water bath, and stirred at room temperature for 3 hours. The reaction mixture was poured into water (300 mL), EtOAc (EtOAc m. ¹ H NMR (300 MHz, CDCl ₃ ) δ 7.40 (dd, J = 8.2, 1.1 Hz, 1H), 7.32 - 7.22 (m, 2H), 7.04 (dd, J = 8.2, 1.1 Hz, 1H).

Step 2. Synthesis of Compound 24.

8.4 g of compound 23 was dissolved in 40 mL of ethanol (EtOH) and 10 mL of water (H ₂ O), and 8.9 g of reduced iron powder and 3.5 g of ammonium chloride solid were added, and the mixture was heated under reflux for 2 hours. Filtration and filtration of the residue with ethyl acetate until the filtrate was colourless. The filtrate was concentrated to give a colorless oil by column chromatography, liquid 4.7 g, LC-MS (APCI ): m / z = 170.2 (M + 1) +.

Step 3. Synthesis of Compound 25.

Take 4.7 g of compound 24 dissolved in 55 mL of xylene, add 38.85 mL of 1M BCl ₃ /DCM solution at 0 ° C, react at 5 ° C for 30 minutes, add 4.01 mL of acetonitrile, react at 5 ° C for 45 minutes, transfer the reaction solution to the dropping solution. In a funnel, the solution was added dropwise to a solution of 4.56 g of aluminum trichloride in 25 mL of xylene at 0 ° C, and the mixture was heated at 5 ° C for 45 minutes, and then heated. The dichloromethane was separated by a water separator and reacted at 75 ° C overnight. The mixture was cooled in an ice water bath, and the reaction solution was poured into 10 mL of ice water and refluxed for 2 hours. After cooling to room temperature, the mixture was separated with methylene chloride. EtOAc (EtOAc)EtOAc. The ice-cold isopropyl ether was added thereto, and the mixture was stirred at 0 ° C for 2 hours to precipitate a gray solid, which was filtered, and the residue was washed with ice-cold isopropyl ether. The residue was collected and dried under vacuum at 45 ° C overnight to obtain 2.992 g of a gray solid powder. . ¹ H NMR (500 MHz, CDCl ₃ ) δ 7.65 (d, J = 9.0 Hz, 1H), 6.30 (d, J = 9.0 Hz, 1H), 2.55 (s, 3H), 2.01 (s, 3H).

Step 4. Synthesis of Compound 26.

500 mg of the compound 17 was dissolved in 5 mL of dioxane, and a solution of the compound 25 (450 mg) in dioxane (5 mL) was added dropwise, and the mixture was stirred at room temperature for 1 hour. The reaction solution was concentrated, with 3 mL of saturated NaHCO ₃ solution and 3 mL of dichloromethane, and the aqueous phase extracted with dichloromethane. The combined dichloromethane was washed with brine, dried over anhydrous sodium sulfate

Step 5. Synthesis of Compound 27.

2.85 g of Compound 26 was dissolved in 30 mL of tert-butanol, 2.37 g of potassium t-butoxide was added, and the mixture was heated under reflux for 6 hours. The reaction mixture was concentrated, DCM and EtOAc was evaporated, evaporated, evaporated, evaporated, evaporated. ¹ H NMR (300 MHz, CDCl ₃ ) δ 9.55 (s, 1H), 8.25 (d, J = 9.0 Hz, 1H), 7.10 (d, J = 0.9 Hz, 1H), 7.01 (d, J = 9.1 Hz, 1H), 6.72 (d, J = 1.8 Hz, 1H), 3.28 - 3.12 (m, 1H), 2.43 (s, 3H), 1.39 (d, J = 6.9 Hz, 6H).

Step 6. Synthesis of Compound 28.

500 mg of compound 11, 486 mg of compound 27 and 669 mg of triphenylphosphine were dissolved in 10 mL of freshly distilled anhydrous THF, and 552 μL of diisopropyl azodicarboxylate (DIAD) was added dropwise at -15 ° C, and at -15 The mixture was stirred at ° C for 5 hours, and naturally stirred at room temperature overnight. The reaction mixture was quenched with EtOAc (EtOAc)EtOAc. LC-MS (APCI): m / z = 692.4 (M + 1) +.

Step 7. Synthesis of Compound 29.

485 mg of compound 28, 18 mg of first-generation gerab catalyst (Hoveyda-Grubbs catalyst) was dissolved in 420 mL of anhydrous dichloroethane, degassed under nitrogen for 30 minutes, and stirred at 75 ° C for 17 hours. The reaction mixture was concentrated to give 234 mg (yield of pale yellow solid). LC-MS (APCI): m / z = 665.4 (M + 1) +.

Step 8. Synthesis of Compound 30.

477 mg of compound 29 was dissolved in 15 mL of methanol and 26 mL of tetrahydrofuran, and 2.89 g of lithium hydroxide monohydrate and 7.5 mL of water were added thereto under ice water, and stirred at room temperature overnight. 1N Hydrochloric acid was adjusted to pH 2~3, EA was extracted, EA phase was combined and washed with saturated brine, dried over anhydrous sodium sulfate and filtered and evaporated to give 315 mg of pale yellow solid powder, LC-MS (APCI): m/z=650.4 (M +1) ⁺ .

Step 9. Synthesis of Compound A-1.

496 mg of compound 30 was dissolved in 15 mL of freshly distilled anhydrous tetrahydrofuran, and 308 mg of N,N'-carbonyldiimidazole (CDI) was added under a nitrogen atmosphere, and the mixture was heated under reflux for 2 hours. After cooling to 50 ° C, 400 mg of cyclopropylsulfonamide and 286 mg of 1,8-diazabicyclo [5.4.0]undec-7-ene (DBU) were added and stirred at 50 ° C overnight. The reaction mixture was concentrated, and 2 mL of 1N hydrochloric acid and 5 mL of dichloromethane was added. The organic phase was washed with saturated brine, dried over anhydrous sodium sulfate and filtered and evaporated to give 277 mg of pale yellow solid powder as compound A-1 , LC-MS (APCI): m/z = 753.4 (M + 1) ⁺ .

Example 6. Preparation of Substituted Quinoline Compound A-2

The specific synthesis steps are as follows:

Step 1. Synthesis of Compound 32.

4.0 g of 2-chloro-6-nitrotoluene was dissolved in 40 mL of MeOD, 1.0 g of MeONa was added, and the mixture was stirred at 100 ° C for 2 hours in a microwave. After cooling, the pH was adjusted to pH 3-4, and the reaction mixture was poured into 20 mL of brine. The yellow solid is compound 32. ¹ H NMR (500 MHz, CDCl ₃ ) δ 7.71 (dd, J = 8.2, 1.3 Hz, 1H), 7.61 (dd, J = 8.0, 1.3 Hz, 1H), 7.29 (d, J = 8.2, 1H) .

Step 2. Synthesis of Compound 33.

4.24 g of compound 32, 4.52 g of sodium tetramethoxyborate, 223 mg of tris(dibenzylideneacetone)dipalladium, and 205 mg of 2-di-tert-butylphosphino-2',4',6'-triisopropyl The phenylbenzene was mixed, and 22 mL of anhydrous DMF was added under nitrogen atmosphere, and the mixture was stirred at 100 ° C for 2 hours. After the reaction was cooled, the mixture was poured into EtOAc (3 mL).

Step 3. Synthesis of Compound 34.

3.25 g of compound 33, 6.35 g of reduced iron powder, and 2.43 g of ammonium chloride solid were mixed, and 20 mL of ethanol and 5 mL of water were added, and the mixture was heated under reflux for 2 hours. The reaction mixture was cooled and filtered, and the residue was washed with methanol, and the filtrate was evaporated.

Step 4. Synthesis of Compound 35.

4.7 g of compound 34 was dissolved in 55 mL of xylene, and 38.85 mL of 1 M BCl ₃ /DCM solution was added at 0 ° C, reacted at 5 ° C for 30 minutes, 4.01 mL of acetonitrile was added, and reacted at 5 ° C for 45 minutes to transfer the reaction solution to the dropping solution. In a funnel, the solution was added dropwise to a solution of 4.56 g of aluminum trichloride in 25 mL of xylene at 0 ° C, and the mixture was heated at 5 ° C for 45 minutes, and then heated. The dichloromethane was separated by a water separator and reacted at 75 ° C overnight. The mixture was cooled in an ice water bath, and the reaction solution was poured into 10 mL of ice water and refluxed for 2 hours. After cooling to room temperature, the mixture was separated with methylene chloride. EtOAc (EtOAc)EtOAc. The ice-cold isopropyl ether was added thereto, and the mixture was stirred at 0 ° C for 2 hours to precipitate a gray solid, which was filtered, and the residue was washed with ice-cold isopropyl ether. The residue was collected and dried under vacuum at 45 ° C overnight to obtain 2.992 g of a gray solid powder. . ¹ H NMR (500 MHz, CDCl ₃ ) δ 7.65 (d, J = 9.0 Hz, 1H), 6.31 (d, J = 9.0 Hz, 1H), 3.87 (s, 3H), 2.55 (s, 3H).

Step 5. Synthesis of Compound 36.

500 mg of the compound 17 was dissolved in 5 mL of dioxane, and a solution of the compound 35 (450 mg) in dioxane (5 mL) was added dropwise, and the mixture was stirred at room temperature for 1 hour. The reaction solution was concentrated, with 3 mL of saturated NaHCO ₃ solution and 3 mL of dichloromethane, and the aqueous phase extracted with dichloromethane. The combined dichloromethane was washed with brine, dried over anhydrous sodium sulfate

Step 6. Synthesis of Compound 37.

451 mg of compound 36 was dissolved in 10 mL of t-butanol, and 540 mg of potassium t-butoxide was added thereto, followed by heating under reflux for 6 hours. The reaction mixture was concentrated, DCM and EtOAc was evaporated, evaporated, evaporated, evaporated, evaporated. ¹ H NMR (400 MHz, CDCl ₃ ) δ 9.53 (s, 1H), 8.30 - 8.19 (m, 1H), 7.10 (d, J = 0.9 Hz, 1H), 7.02 (d, J = 9.1 Hz, 1H) ), 6.72 (d, J = 1.8 Hz, 1H), 3.98 (s, 3H), 3.24 - 3.16 (m, 1H), 1.40 (d, J = 6.9 Hz, 6H).

Step 7. Synthesis of Compound 38.

500 mg of compound 11, 486 mg of compound 37 and 669 mg of triphenylphosphine were dissolved in 10 mL of freshly distilled anhydrous THF, and 552 μL of DIAD was added dropwise at -15 ° C, and stirred at -15 ° C for 5 hours, and stirred at room temperature overnight. . The reaction was quenched by the addition of ice water, and the aqueous layer was evaporated. EtOAc was evaporated. LC-MS (APCI): m / z = 692.4 (M + 1) +.

Step 8. Synthesis of Compound 39.

481 mg of compound 38, 18 mg of Hoveyda-Grubbs first generation catalyst was dissolved in 420 mL of anhydrous dichloroethane, degassed for 30 minutes under nitrogen, and stirred at 75 ° C for 15 hours. The reaction mixture was concentrated and purified by column chromatography to yieldd to afforded Compound Compound Compound LC-MS (APCI): m / z = 664.4 (M + 1) +.

Step 9. Synthesis of Compound 40.

220 mg of Compound 39 was dissolved in 20 mL of methanol and 30 mL of tetrahydrofuran, and 2.89 g of lithium hydroxide monohydrate and 15 mL of water were added under ice water, and stirred at room temperature overnight. 1N Hydrochloric acid was adjusted to pH 2~3, EA was extracted, EA phase was combined and washed with saturated brine, dried over anhydrous sodium sulfate and evaporated to give 186 mg of pale yellow solid powder, LC-MS (APCI): m/z=650.4 (M +1) ⁺ .

Step 11. Synthesis of Compound A-2.

150 mg of compound 40 was dissolved in 15 mL of freshly distilled anhydrous tetrahydrofuran, and 120 mg of CDI was added under nitrogen atmosphere, and the mixture was heated under reflux for 2 hours. After cooling to 50 ° C, 145 mg of cyclopropylsulfonamide and 83 mg of DBU were added and stirred at 50 ° C overnight. The reaction mixture was concentrated, and 2 mL of 1N hydrochloric acid and 5 mL of dichloromethane was added, and the organic phase was washed with saturated brine, dried over anhydrous sodium sulfate, LC-MS (APCI): m / z = 753.4 (M + 1) +.

Example 7. Preparation of Substituted Quinoline Compound A-3

The specific synthesis steps are as follows:

Step 1. Synthesis of Compound 41.

0.749 g of 6-bromo-1-ethene was dissolved in 7.4 mL of DMSO, and 448 mg of sodium azide was slowly added in portions, and the mixture was stirred at room temperature for 2 hours. The reaction solution was poured into 25 mL of water and extracted with EtOAc (3 mL, 3). To the ether solution, 2.41 g of triphenylphosphine, 0.5 mL of water was added, and the mixture was stirred at room temperature overnight. The reaction solution was washed three times with 1N hydrochloric acid (25 mL×3), The mixture was adjusted to pH 8 to 9 with NaOH solids, and extracted with diethyl ether. The mixture was combined with diethyl ether, washed with saturated brine, dried over anhydrous sodium sulfate and filtered to give 450 mg of colorless oil.

Step 2. Synthesis of Compound 42.

450 mg of compound 41 was dissolved in 25 mL of diethyl ether, Boc ₂ O 1.0 g was added dropwise thereto in an ice water bath, and DIPEA 974 μL was added dropwise thereto, and stirred at room temperature overnight. The reaction mixture was concentrated and purified by column chromatography eluting 881

Step 3. Synthesis of Compound 43.

899 mg of compound 42 was dissolved in 3 mL of DMF, and 130 mg of 60% NaH was added in portions, stirred at room temperature for 30 minutes, and 655 mg of deuterated iodomethane was added dropwise, and the mixture was stirred at room temperature for 2 hours. The reaction mixture was poured into water (20 mL), EtOAc (EtOAc)EtOAc. _{^{1 H NMR (400MHz, CDCl 3}} ) δ5.79 (ddt, J = 16.9,10.2,6.7 Hz, 1H), 5.13-4.87 (m, 2H), 3.19 (s, 2H), 2.06 (dt, J = 8.6 , 4.3 Hz, 2H), 1.51 (dt, J = 14.8, 7.2 Hz, 2H), 1.44 (s, 9H), 1.36 (dt, J = 14.7, 7.2 Hz, 2H).

Step 4. Synthesis of Compound 44.

756 mg of compound 43 was dissolved in 10 mL of a DCM/TFA (V/V, 8/2) mixed solution and stirred at room temperature for 2 hours. The reaction mixture was concentrated, and the mixture was evaporated, evaporated, evaporated, evaporated. The oily liquid 336 g is the compound 44.

Step 5. Synthesis of Compound 45.

Take (1R,4R,5R)-3-oxo-2-oxabicyclo[2.2.1]hexane-5-carboxylic acid cinchonidine salt 3.51 g and 1.09 g of compound 44 dissolved in 10 mL DMF, ice HATU 3.57 g, DIPEA 10 g was added under cooling with a bath and stirred for 1 hour in an ice bath. The reaction mixture was poured into 100 mL of water, and ethyl acetate was evaporated, and ethyl acetate was evaporated. ¹ H NMR (500 MHz, CDCl ₃ ) δ 5.77 (ttd, J = 13.1, 6.7, 2.8 Hz, 1H), 5.13 - 4.86 (m, 3H), 3.43 - 3.23 (m, 2H), 3.08 - 2.94 ( m, 2H), 2.29–2.02 (m, 6H), 1.69–1.46 (m, 2H), 1.46–1.31 (m, 2H). LC-MS (APCI): m/z=255.2 (M+1) ⁺ .

Step 6. Synthesis of Compound 46.

1.86 g of the compound 45 was dissolved in 13 mL of tetrahydrofuran, and 5.4 mL of a 1 M LiOH solution was added dropwise thereto in an ice water bath, and the mixture was stirred for 1 hour under ice-cold bath. 4N HCl solution was adjusted to pH 1-2, ethyl acetate (10×3), and ethyl acetate was evaporated. APCI): m/z = 252.3 (M + 1) ⁺ .

Step 7. Synthesis of Compound 47.

1.26 g of compound 46 was dissolved in 10 mL of anhydrous DMF, and 1.94 g of HATU and 789 mg of compound 5, 920 μL of DIPEA were added under ice bath, and the mixture was stirred for 1 hour under ice bath. The reaction solution was poured into 100 mL of water, extracted with EA, and the EA phase was combined, washed with 1N HCl solution, water, saturated aqueous sodium chloride solution, dried over anhydrous sodium sulfate, and concentrated by filtration. . LC-MS (APCI): m / z = 396.3 (M + 1) +.

Step 8. Synthesis of Compound 48.

500 mg of compound 47, 477 mg of compound 21 and 664 mg of triphenylphosphine were dissolved in 10 mL of freshly distilled anhydrous THF, and 552 μL of DIAD was added dropwise at -15 ° C, and stirred at -15 ° C for 5 hours, and stirred at room temperature overnight. . The reaction was quenched by the addition of ice water, and the mixture was evaporated. LC-MS (APCI): m / z = 692.4 (M + 1) +.

Step 9. Synthesis of Compound 49.

529 mg of compound 48, 24 mg of Hoveyda-Grubbs first generation catalyst was dissolved in 480 mL of anhydrous dichloroethane, degassed under nitrogen for 30 minutes, and stirred at 75 ° C for 15 hours. The reaction mixture was concentrated and purified by column chromatography to afford 327 g of pale yellow solid. LC-MS (APCI): m / z = 664.4 (M + 1) +.

Step 10. Synthesis of Compound 50.

689 mg of Compound 49 was dissolved in 20 mL of methanol and 30 mL of tetrahydrofuran, and 2.89 g of lithium hydroxide monohydrate and 15 mL of water were added thereto under ice water, and stirred at room temperature overnight. 1N Hydrochloric acid was adjusted to pH 2~3, EA was extracted, EA phase was combined and washed with saturated brine, dried over anhydrous sodium sulfate and filtered to give 686mg of pale yellow solid powder, LC-MS (APCI): m/z=650.4 (M+ 1) ⁺ .

Step 11. Synthesis of Compound A-3.

686 mg of compound 50 was dissolved in 17 mL of freshly distilled anhydrous tetrahydrofuran, and 308 mg of CDI was added under nitrogen atmosphere, and the mixture was heated under reflux for 2 hours. After cooling to 50 ° C, 400 mg of cyclopropylsulfonamide and 286 mg of DBU were added and stirred at 50 ° C overnight. The reaction mixture was concentrated, and 2 mL of 1N hydrochloric acid and 5 mL of dichloromethane was added, and the organic phase was washed with saturated brine, dried over anhydrous sodium sulfate, filtered, and concentrated to give 335 mg of pale yellow solid powder as compound A- 3, LC-MS (APCI): m / z = 753.4 (M + 1) ⁺ .

Example 8. Preparation of Substituted Quinoline Compound A-4

The specific synthesis steps are as follows:

Step 1. Synthesis of Compound 51.

4.41 g of compound 33 was dissolved in 30 mL of dichloromethane, and 4.69 mL of boron trichloride was slowly added dropwise in an ice salt bath, and the reaction was naturally carried out for 30 minutes. 10 mL of ice water was added to the ice water bath. The organic layer was washed with brine, dried over anhydrous sodium sulfate

Step 2. Synthesis of Compound 52.

3.56 Compound 51 was dissolved in 20 mL of DMF, and 6.30 g of potassium carbonate solid was added, stirred at room temperature for 30 minutes, and 3.97 g of deuterated iodomethane was added dropwise, and stirred at room temperature for 2 hours. The reaction mixture was poured into 150 mL of water and EtOAc (3 mL, EtOAc)

Step 3. Synthesis of Compound 53.

3.63 g of compound 52, 6.40 g of reduced iron powder, and 2.41 g of ammonium chloride solid were mixed, and 20 mL of ethanol and 5 mL of water were added, and the mixture was heated under reflux for 2 hours. The reaction mixture was cooled and filtered, and the residue was washed with methanol, and the filtrate was evaporated. The filtrate was concentrated and concentrated to give a colorless liquid 3.36 g.

Step 4. Synthesis of Compound 54.

Take 4.7 g of compound 53 dissolved in 55 mL of xylene, add 38.85 mL of 1M BCl ₃ /DCM solution at 0 ° C, react at 5 ° C for 30 minutes, add 4.01 mL of acetonitrile, react at 5 ° C for 45 minutes, transfer the reaction solution to the dropping solution. In a funnel, the solution was added dropwise to a solution of 4.56 g of aluminum trichloride in 25 mL of xylene at 0 ° C, and the mixture was heated at 5 ° C for 45 minutes, and then heated. The dichloromethane was separated by a water separator and reacted at 75 ° C overnight. The mixture was cooled in an ice water bath, and the reaction solution was poured into 10 mL of ice water and refluxed for 2 hours. After cooling to room temperature, the mixture was separated with methylene chloride. EtOAc (EtOAc)EtOAc. The ice-cold isopropyl ether was added thereto, and the mixture was stirred at 0 ° C for 2 hours to precipitate a gray solid, which was filtered, and the residue was washed with ice-cold isopropyl ether. The residue was collected and dried in vacuo at 45 ° C overnight to give the compound . ¹ H NMR (400 MHz, CDCl ₃ ) δ 7.66 (d, J = 9.0 Hz, 1H), 6.31 (d, J = 9.1 Hz, 1H), 2.56 (s, 3H).

Step 5. Synthesis of Compound 55.

500 mg of the compound 54 was dissolved in 5 mL of dioxane, and a solution of the compound 17 (450 mg) in dioxane (5 mL) was added dropwise, and the mixture was stirred at room temperature for 1 hour. The reaction solution was concentrated, with 3 mL of saturated NaHCO ₃ solution and 3 mL of dichloromethane, and the aqueous phase extracted with dichloromethane. The combined dichloromethane was washed with brine, dried over anhydrous sodium sulfate

Step 6. Synthesis of Compound 56.

445 mg of Compound 55 was dissolved in 10 mL of t-butanol, and 540 mg of potassium t-butoxide was added thereto, followed by heating under reflux for 6 hours. The reaction mixture was concentrated, DCM and EtOAc was evaporated. ¹ H NMR (400 MHz, CDCl ₃ ) δ 9.53 (s, 1H), 8.30 - 8.19 (m, 1H), 7.10 (d, J = 0.9 Hz, 1H), 7.02 (d, J = 9.1 Hz, 1H) ), 6.72 (d, J = 1.8 Hz, 1H), 3.24 - 3.16 (m, 1H), 1.40 (d, J = 6.9 Hz, 6H).

Step 7. Synthesis of Compound 57.

500 mg of compound 11, 486 mg of compound 56 and 669 mg of triphenylphosphine were dissolved in 10 mL of freshly distilled anhydrous THF, and 552 μL of DIAD was added dropwise at -15 ° C, and stirred at -15 ° C for 5 hours, and stirred at room temperature overnight. . The reaction was quenched by the addition of ice water, and the mixture was evaporated. LC-MS (APCI): m/z = 695.3 (M+1 ⁾ .

Step 8. Synthesis of Compound 58.

473 mg of compound 57, 11 mg of Hoveyda-Grubbs first generation catalyst was dissolved in 400 mL of anhydrous dichloroethane, degassed under nitrogen for 30 minutes, and stirred at 75 ° C for 15 hours. The reaction mixture was concentrated and purified by column chromatography to yield 288 g of pale yellow solid. LC-MS (APCI): m / z = 666.4 (M + 1) +.

Step 9. Synthesis of Compound 59.

250 mg of Compound 58 was dissolved in 20 mL of methanol and 30 mL of tetrahydrofuran, and 2.89 g of lithium hydroxide monohydrate and 15 mL of water were added under ice water, and stirred at room temperature overnight. 1N Hydrochloric acid was adjusted to pH 2~3, EA was extracted, EA phase was combined and washed with saturated brine, dried over anhydrous sodium sulfate, and filtered and evaporated to give a pale yellow solid powder, LC-MS (APCI): m/z=652.4 (M +1) ⁺ .

Step 10. Synthesis of Compound A-4.

200 mg of compound 59 was dissolved in 15 mL of freshly distilled anhydrous tetrahydrofuran, 210 mg of CDI was added under nitrogen, and the mixture was heated under reflux for 2 hours. After cooling to 50 ° C, 235 mg of cyclopropylsulfonamide and 189 mg of DBU were added and stirred at 50 ° C overnight. The reaction mixture was concentrated, and 2 mL of 1N hydrochloric acid and 5 mL of dichloromethane was added, and the organic phase was washed with saturated brine, dried over anhydrous sodium sulfate, and filtered and concentrated to give 168 mg of pale yellow solid powder as compound A-4 . LC-MS (APCI): m / z = 755.3 (M + 1) +.

Example 9. Preparation of substituted quinoline compound A-5.

The specific synthesis steps are as follows:

Step 1. Synthesis of Compound 60.

500 mg of compound 26, 482 mg of compound 47 and 664 mg of triphenylphosphine were dissolved in 10 mL of freshly distilled anhydrous THF, and 552 μL of DIAD was added dropwise at -15 ° C, and stirred at -15 ° C for 5 hours, and stirred at room temperature overnight. . The reaction was quenched by the addition of ice water, and the aqueous layer was evaporated. EtOAc was evaporated. LC-MS (APCI): m / z = 695.4 (M + 1) +.

Step 2. Synthesis of Compound 61.

534 mg of compound 60, 11 mg of Hoveyda-Grubbs first generation catalyst was dissolved in 300 mL of anhydrous dichloroethane, degassed under nitrogen for 30 minutes, and stirred at 75 C for 15 hours. The reaction mixture was concentrated and purified by column chromatography to afford 365 g of pale yellow solid. LC-MS (APCI): m / z = 666.4 (M + 1) +.

Step 3. Synthesis of Compound 62.

689 mg of Compound 61 was dissolved in 20 mL of methanol and 30 mL of tetrahydrofuran, and 2.89 g of lithium hydroxide monohydrate and 15 mL of water were added under ice water, and stirred at room temperature overnight. 1N Hydrochloric acid was adjusted to pH 2~3, EA was extracted, EA phase was combined and washed with saturated brine, dried over anhydrous sodium sulfate, and filtered to give 686 mg of pale yellow solid powder, LC-MS (APCI): m/z=653.4 (M +1) ⁺ .

Step 4. Synthesis of Compound A-5.

500 mg of compound 62 was dissolved in 15 mL of freshly distilled anhydrous tetrahydrofuran, and 520 mg of CDI was added under a nitrogen atmosphere, and the mixture was heated under reflux for 2 hours. After cooling to 50 ° C, 515 mg of cyclopropylsulfonamide and 412 mg of DBU were added and stirred at 50 ° C overnight. The reaction mixture was concentrated, and 2 ml of 1N hydrochloric acid and 5 mL of dichloromethane was added. The organic phase was washed with saturated brine, dried over anhydrous sodium sulfate 5. LC-MS (APCI): m / z = 755.3 (M + 1) +.

Evaluation of biological activity.

To verify the effect of the compounds described herein on HCV, the inventors used the HCV Replicon System as an evaluation model. Since its first report in Science in 1999, the HCV replication system has become one of the most important tools for studying HCV RNA replication, pathogenicity and viral persistence, for example, the use of replicons has successfully demonstrated the 5' required for HCV RNA replication. - NCR minimum region, and the HCV replication subsystem has been successfully used as an evaluation model for antiviral drugs. The inventors of the present invention verified according to the methods described in Science, 1999, 285 (5424), 110-3, and J. Virol, 2003, 77(5), 3007-19.

(1) Detection of compound anti-HCV 1a and 1b genotype replicon activity

The inhibitory activities of the recombinant hepatitis C virus genotype 1a and 1b replicons were detected by stable transfection of replicon cells with HCV-1a and HCV-1b. This experiment will use the NS3/4A inhibitor Simeprevir as a positive control compound.

Step 1: The compound was diluted 1:3 in 8 series points, double-replicated, and added to a 96-well plate. The DMSO was set to no compound control. The final concentration of DMSO in the cell culture was 0.5%.

Step 2: HCV-1a and 1b cells were separately suspended in a culture medium containing 10% FBS, and seeded into a 96-well plate containing the compound at a density of 8,000 cells per well. The cells were cultured for 3 days at 5% CO ₂ at 37 °C.

Step 3: The cytotoxicity of the compound against GT1b replicon was determined using CellTiter-Fluor (Promega).

Step 4: Detection of luciferase assay by Bright-Glo (Promega) for anti-hepatitis C virus activity.

Step Five: using GraphPad Prism data analysis software, the curve fitting and EC ₅₀ values _were calculated and the ₅₀ value CC.

The compounds A-1 to A-5 of Examples 1 to 5 were analyzed in the above procedure to calculate EC ₅₀ values and CC ₅₀ values, and the results are shown in Table 1.

Table 1 Comparison of anti-HCV genotype replicon activities of the compounds of Examples A-1 to A-5 and the control Simeprevir

Numbering GT1a EC ₅₀ (nM) GT1b EC ₅₀ (nM) CC ₅₀ (nM) Simeprevir 22.59 11.41 >1000 Compound A-1 8.07 3.79 >1000 Compound A-2 12.97 6.53 >1000 Compound A-3 9.48 3.96 >1000 Compound A-4 11.84 6.28 >1000 Compound A-5 10.29 8.43 >1000

The experimental results show that the compounds of the present invention can inhibit the multiple genotypes of HCV and can be used for the inhibition of hepatitis C virus. The compound of the present invention exhibited more excellent inhibitory activity against the GT1a and GT1b replicons than the non-deuterated compound Simeprevir.

(2) Determination of compound NS 3/4A inhibitory activity

For the test method, the HCV NS3/4A protease time-resolved fluorescence (TRF) assay described in Anal. Biochem. 373: 1-8, 2008 and International Patent Application Publication No. WO 2006/102087.

Numbering EC ₅₀ (nM) replicon assay Ki(nM) Compound A-1 <10 <10 Compound A-2 <10 <10 Compound A-3 <10 <10 Compound A-4 <10 <10

Compound A-5 <10 <10

As shown in the above table, the experimental results indicate that the compound of the present invention can be used as an inhibitor of HCV NS3/4A protease, and can be used in a drug against hepatitis C virus.

(3) Metabolic stability evaluation

Microsomal experiments: human liver microsomes: 0.5 mg/mL, Xenotech; rat liver microsomes: 0.5 mg/mL, Xenotech; coenzyme (NADPH/NADH): 1 mM, Sigma Life Science; magnesium chloride: 5 mM, 100 mM phosphate Buffer (pH 7.4).

Preparation of Stock Solution: A certain amount of Compound Example Compounds A-1 to A-5 and the positive control Simeprevir powder were accurately weighed and dissolved to 5 mM with DMSO, respectively.

Preparation of phosphate buffer (100 mM, pH 7.4): Mix 150 mL of pre-formed 0.5 M potassium dihydrogen phosphate and 700 mL of 0.5 M potassium dihydrogen phosphate solution, and adjust the mixture with 0.5 M potassium dihydrogen phosphate solution. The pH of the solution was adjusted to 7.4, diluted 5 times with ultrapure water before use, and magnesium chloride was added to obtain a phosphate buffer (100 mM) containing 100 mM potassium phosphate, 3.3 mM magnesium chloride, and a pH of 7.4.

A solution of NADPH regeneration system (containing 6.5 mM NADP, 16.5 mM G-6-P, 3 U/mL G-6-P D, 3.3 mM magnesium chloride) was prepared and placed on wet ice before use.

Formulation stop solution: acetonitrile solution containing 50 ng/mL propranolol hydrochloride and 200 ng/mL tolbutamide (internal standard). Take 25057.5 μL of phosphate buffer (pH 7.4) into a 50 mL centrifuge tube, add 812.5 μL of human liver microsomes, and mix to obtain a liver microsome dilution with a protein concentration of 0.625 mg/mL. Take 25057.5 μL of phosphate buffer (pH 7.4) into a 50 mL centrifuge tube, add 812.5 μL SD rat liver microsomes, and mix to obtain a liver microsome dilution with a protein concentration of 0.625 mg/mL.

Incubation of the sample: The stock solution of the corresponding compound was diluted to 0.25 mM with an aqueous solution containing 70% acetonitrile, and used as a working solution. 398 μL of human liver microsomes or rat liver microsome dilutions were added to 96-well incubation plates (N=2), and 2 μL of 0.25 mM working solution was added and mixed.

Determination of metabolic stability: 300 μL of pre-cooled stop solution was added to each well of a 96-well deep well plate and placed on ice as a stop plate. The 96-well incubation plate and the NADPH regeneration system were placed in a 37 ° C water bath, shaken at 100 rpm, and pre-incubated for 5 min. 80 μL of the incubation solution was taken from each well of the incubation plate, added to the stopper plate, and mixed, and 20 μL of the NADPH regeneration system solution was added as a 0 min sample. Then, 80 μL of the NADPH regeneration system solution was added to each well of the incubation plate to start the reaction and start timing. The corresponding compound had a reaction concentration of 1 μM and a protein concentration of 0.5 mg/mL. 100 μL of the reaction solution was taken at 10, 30, and 90 min, respectively, and added to the stopper plate, and the reaction was terminated by vortexing for 3 min. The plate was centrifuged at 5000 x g for 10 min at 4 °C. 100 μL of the supernatant was taken into a 96-well plate to which 100 μL of distilled water was previously added, mixed, and sample analysis was performed by LC-MS/MS.

Data analysis: The peak area of the corresponding compound and the internal standard was detected by LC-MS/MS system, and the ratio of the peak area of the compound to the internal standard was calculated. The slope is measured by the natural logarithm of the percentage of the remaining amount of the compound versus time, and t _1/2 and CL _{int are} calculated according to the following formula, where V/M is equal to 1/protein concentration.

The metabolic stability of human and rat liver microsomes was evaluated by simultaneously testing the compounds of the present invention and their compounds without deuteration. The half-life and liver intrinsic clearance as indicators of metabolic stability are shown in Table 2. The undeuterated compound Simeprevir was used as a control sample in Table 2. As shown in Table 2, the compound of the present invention can significantly improve metabolic stability by comparison with the undeuterated compound Simeprevir, and is thus more suitable as a hepatitis C virus inhibitor.

Table 2 Comparison of Metabolic Stability of Example Compounds A-1 to A-5 with Simeprevir Control

The compounds of the examples were analyzed according to the above procedure, and the results of the experiments showed that the compounds of the present invention exhibited excellent metabolic stability in both human liver microsomes and rat liver microsomes.

(4) Rat pharmacokinetic experiments

EXPERIMENTAL OBJECTIVE: To investigate the pharmacokinetic behavior of the compounds of the present invention after administration of Simeprevir to the compounds of the examples.

Experimental animals:

Type and strain: SD rat grade: SPF grade

Gender and quantity: male, 6

Weight range: 180 ~ 220g (actual weight range is 187 ~ 197g)

Source: Shanghai Xipuer Bikai Experimental Animal Co., Ltd.

Laboratory and animal certificate number: SCXK (Shanghai) 2013-0016.

experiment procedure:

Before the blood sample was collected, 20 L of 2M sodium fluoride solution (esterase inhibitor) was added to the EDTA-K2 anticoagulant tube, dried in an 80 degree oven, and stored in a 4 degree refrigerator.

Rats, males, weighing 187-197 g, were randomly divided into 2 groups. They were fasted overnight in the afternoon before the experiment but were free to drink water and given food 4 h after administration. Group A was given 3 mg/kg of Simeprevir, and group B was given compound A-1 to A-53 mg/kg of the formula, respectively, from 15 min, 30 min, 1, 2, 3, 5, 8, 10 h after administration. The mouse eye vein was taken from a blood of about 100-200 L, placed in an EDTA-K2 anticoagulated 0.5 mL Eppendorf tube, and immediately mixed. After anticoagulation, the tube was gently inverted and mixed 5-6 times as soon as possible. After taking it, place it in an ice box, centrifuge the blood sample at 4000 rpm, 10 min, 4 °C for 30 min, collect all the plasma and store at -20 °C immediately. Plasma concentrations in plasma at each time point were determined after sample collection at all time points.

According to the average blood drug concentration-time data obtained after the above, male Sprague-Dawley rats were administered ig to Simeprevir (3 mg/kg) and Example Compounds A-1 to A according to the non-compartmental statistical moment theory using Winnonin software. Pharmacokinetic related parameters after -5 (3 mg/kg).

The experimental results show that compared with Simeprevir, the present inventors have found that the compound has superior activity than Simeprevir and has excellent pharmacokinetic properties, and thus is more suitable as a compound for inhibiting the hepatitis C virus protein NS3/4A, and is suitable for the compound. Preparation of a medicament for treating hepatitis C virus infection.

It is to be understood that the examples are merely illustrative of the invention and are not intended to limit the scope of the invention, and the experimental methods in which the specific conditions are not indicated in the examples, usually in accordance with conventional conditions or in accordance with the conditions suggested by the manufacturer. Parts and percentages are parts by weight and percentage by weight unless otherwise stated.

The above is a further detailed description of the present invention in connection with the specific preferred embodiments, and the specific embodiments of the present invention are not limited to the description. It will be apparent to those skilled in the art that the present invention may be made without departing from the spirit and scope of the invention.

Claims (9)

A substituted quinoline compound characterized by the compound of formula (I), or a crystalline form thereof, a pharmaceutically acceptable salt, a hydrate or a solvent compound,

Wherein R ¹ , R ² , R ³ , R ⁴ , R ⁵ , R ⁶ , R ⁷ , R ⁸ , R ⁹ , R ¹⁰ , R ¹¹ , R ¹² , R ¹³ , R ¹⁴ , R ¹⁵ , R ¹⁶ , R ¹⁷ , R ¹⁸ , R ¹⁹ , R ²⁰ , R ²¹ , R ²² , R ²³ , R ²⁴ , R ²⁵ , R ²⁶ , R ²⁷ , R ²⁸ , R ²⁹ , R ³⁰ , R ³¹ , R ³² , R ³³ And R ³³ , R ³⁴ , R ³⁵ , R ³⁶ , R ³⁷ , R ³⁸ , R ³⁹ , R ⁴⁰ , R ⁴¹ , R ⁴² , R ⁴³ , R ⁴⁴ , R ⁴⁵ are each independently hydrogen, deuterium, halogen or trisole Fluoromethyl; Additional conditions are R ¹ , R ² , R ³ , R ⁴ , R ⁵ , R ⁶ , R ⁷ , R ⁸ , R ⁹ , R ¹⁰ , R ¹¹ , R ¹² , R ¹³ , R ¹⁴ , R ¹⁵ , R ¹⁶ And R ¹⁷ , R ¹⁸ , R ¹⁹ , R ²⁰ , R ²¹ , R ²² , R ²³ , R ²⁴ , R ²⁵ , R ²⁶ , R ²⁷ , R ²⁸ , R ²⁹ , R ³⁰ , R ³¹ , R ³² , R ^{33. At least one} of R ³³ , R ³⁴ , R ³⁵ , R ³⁶ , R ³⁷ , R ³⁸ , R ³⁹ , R ⁴⁰ , R ⁴¹ , R ⁴² , R ⁴³ , R ⁴⁴ and R ⁴⁵ is deuterated. The hepatitis C virus inhibitor according to claim 1, wherein each of R ¹⁹ , R ²⁰ and R ^{21 is} independently hydrazine or hydrogen. The hepatitis C virus inhibitor according to claim 1, wherein each of R ³² , R ³³ and R ^{34 is} independently hydrazine or hydrogen. The hepatitis C virus inhibitor according to claim 1, wherein each of R ³⁵ , R ³⁶ and R ^{37 is} independently hydrazine or hydrogen. [Correct according to Rule 26 14.09.2017]
The hepatitis C virus inhibitor according to claim 1, wherein the compound is selected from the group consisting of the following compounds or a pharmaceutically acceptable salt thereof:

[Correct according to Rule 26 14.09.2017]
A pharmaceutical composition comprising a pharmaceutically acceptable carrier and a quinoline compound according to any one of claims 1 to 6, or a crystalline form thereof, a pharmaceutically acceptable salt, a hydrate or A pharmaceutical composition of a solvate, stereoisomer, prodrug or isotopic variation. [Correct according to Rule 26 14.09.2017]
The pharmaceutical composition according to claim 7, characterized in that it further comprises other active compounds which may be selected from the group consisting of HCV protease inhibitors, HCV NS5A inhibitors and HCV NS5B polymerase inhibitors. [Correct according to Rule 26 14.09.2017]
Use of a quinoline compound according to any one of claims 1 to 6 for use in the manufacture of a medicament for the treatment of hepatitis C virus infection. [Correct according to Rule 26 14.09.2017]
Use of a pharmaceutical composition according to any one of claims 7 and 8, characterized in that it is used for the preparation of a medicament for inhibiting the activity of HCV NS3/4A protease in a subject in need thereof.