CN111170911A

CN111170911A - Pleuromutilin compounds for treating bacterial infectious diseases

Info

Publication number: CN111170911A
Application number: CN201910934475.0A
Authority: CN
Inventors: 朱孝云; 蒋维平
Original assignee: Antikang Wuxi Biotechnology Co ltd
Current assignee: Antikang Wuxi Biotechnology Co ltd
Priority date: 2019-09-29
Filing date: 2019-09-29
Publication date: 2020-05-19

Abstract

The present invention relates to novel pleuromutilin compounds, their pharmaceutical compositions and methods of use. In addition, the invention relates to therapeutic methods for treating bacterial infections, including infections caused by drug-resistant microorganisms, including multidrug-resistant microorganisms.

Description

Pleuromutilin compounds for treating bacterial infectious diseases

Technical Field

Pleuromutilin (pleuromutilin), a compound of the formula,

antibiotics which are naturally occurring, are produced, for example, by The basidiomycetes pleuromutilis (Basidomycetes Pleurotusmutilus) and pleurotus palustris (p. passeckeriana), see The Merck lndex, 13 th edition, item 7617. Many other pleuromutilins have been developed as antimicrobial agents, having the main ring structure of pleuromutilin and being substituted on the side chain primary alcoholic hydroxyl group. Due to their pronounced antimicrobial activity, a series of pleuromutilin derivatives, such as amino-hydroxy-substituted cyclohexylsulfanylacetamidelins disclosed in WO2008/113089, have been found to have superior antimicrobial activity. The 14-O- { [ (4-amino-2-hydroxy-cyclohexyl) sulfanyl ] acetyl) -thimerosides disclosed in WO2008/11089 show good application prospects, they have good inhibitory activity against gram-positive and gram-negative pathogens, which are associated with pathogenic bacteria associated with infections of the respiratory tract and the skin and skin structures.

These disclosed pleuromutilin derivatives are simple substitutions on side chain hydroxyl groups and are not modified on the main structural mother ring. Therefore, the prior side chain modified pleuromutilin derivative has the disadvantages of insufficient activity against drug-resistant bacteria, poor in vivo stability, low oral absorption and utilization rate, diarrhea side effect and even cardiotoxicity.

The invention provides a novel pleuromutilin derivative by modifying a main structural mother ring. They have the advantages of strong activity of resisting drug-resistant bacteria, high in-vivo stability, good oral absorption utilization rate, good lung targeting property, low cardiac toxicity and no diarrhea side effect.

Detailed Description

I. Compound (I)

Briefly, the present invention relates to novel pleuromutilin derivatives. The compounds of the present invention have the following structural formula (I) or are stereoisomers, prodrugs, active metabolites or pharmaceutically acceptable salts, solvates or crystalline forms thereof:

wherein,

R¹is hydroxy, hydrogen or deuterium;

R²is hydroxy, hydrogen, deuterium or fluorine;

R^2‘is hydroxy, hydrogen, deuterium or fluorine;

R³is hydroxy, hydrogen, deuterium or fluorine;

R^3’is hydroxy, hydrogen, deuterium or fluorine;

R⁴is ethyl, pentadeuteroethyl, 1, 2-dideuteroethyl, vinyl, trideuterovinyl, 2, 2-dideuterovinyl;

m is hydrogen or deuterium;

a is methylene, oxygen, -NH-, sulfur, sulfoxide, or sulfone;

l is

When L is

In which

n is 0 to 4;

m is 0 or 1;

R⁵is hydrogen or (C)_1-6) An alkyl group;

R⁶is hydrogen, (C)_3-6) Cycloalkyl radicals, unsubstituted (C)_1-6) Alkyl, substituted by one or more of hydroxy, methoxy, halogen, (C)_3-6) Cycloalkyl-substituted (C)_1-6) Alkyl, or

R⁵And R⁶Hydrogen together with the nitrogen atom to which they are attached form a 5-7 membered heterocyclic ring comprising at least one nitrogen atom or one nitrogen atom and one further heteroatom, e.g. selected from N or O, or

R⁵Is hydroxy and R⁶Is a formyl group;

R⁷is OH, OR⁸A halogen atom, or

R⁸Is unsubstituted (C)_1-6) Alkyl or (C)_3-6) Cycloalkyl radical, C_2-18Fatty acyl, PEG (1-20).

In another aspect, the invention relates to a compound of formula (II),

wherein,

n is 0 to 4;

R¹is hydroxy, hydrogen or deuterium;

R²is hydroxy, hydrogen, deuterium or fluorine;

R^2‘is hydroxy, hydrogen, deuterium or fluorine;

R³is hydroxy, hydrogen, deuterium or fluorine;

R^3’is hydroxy, hydrogen, deuterium or fluorine;

m is hydrogen or deuterium;

R⁵is hydrogen or (C)_1-6) An alkyl group;

R⁵Is hydroxy and R⁶Is formyl.

In another aspect, the invention relates to a compound of formula (III),

wherein,

n is 0 to 4;

R¹is hydroxy, hydrogen or deuterium;

R²is hydroxy, hydrogen, deuterium or fluorine;

R^2‘is hydroxy, hydrogen, deuterium or fluorine;

R³is hydroxy, hydrogen, deuterium or fluorine;

R^3’is hydroxy, hydrogen, deuterium or fluorine;

m is hydrogen or deuterium.

In another aspect, the invention relates to compounds of formula (IV),

wherein,

n is 0 to 4;

R¹is hydroxy, hydrogen or deuterium;

R²is hydroxy, hydrogen, deuterium or fluorine;

R^2‘is hydroxy, hydrogen, deuterium or fluorine;

R³is hydroxy, hydrogen, deuterium or fluorine;

R^3’is hydroxy, hydrogen, deuterium or fluorine;

R⁴is ethyl, 1, 2-dideuteroethyl, vinyl, 2, 2-dideuterovinyl.

In yet another aspect, the pleuromutilin derivatives of formula (I), formula (II), formula (III) or formula (IV), stereoisomers, prodrugs, active metabolites or pharmaceutically acceptable salts, solvates or crystalline forms thereof, according to the present invention, are represented by any one of the following structural formulae:

the invention also relates to a pharmaceutical composition comprising a therapeutically effective amount of a compound as described above in unit dosage form and one or more pharmaceutically acceptable carriers.

The invention also relates to a pharmaceutical combination composition comprising a therapeutically effective amount of a compound according to the above claims and another agent having therapeutic activity against a microbial mediated disease.

The compounds of the present invention, their N-oxides or prodrugs thereof, may be used for the manufacture of a medicament for the treatment of a condition mediated by a microorganism.

The microorganism-mediated disease condition can be selected from systemic infectious diseases, ocular infectious diseases, upper respiratory infectious diseases, lower respiratory infectious diseases, skin and soft tissue infectious diseases, acute sinusitis, chronic bronchitis, community-acquired pneumonia and hospital-acquired pneumonia.

The compounds of the invention are generally used in the form of the free acid or free base. Alternatively, the compounds of the present invention may be used in the form of acid or base salts. Acid addition salts of the free amino compounds of the present invention may be prepared by methods well known in the art and may be prepared from organic and inorganic acids. Suitable acids include maleic acid, fumaric acid, benzoic acid, ascorbic acid, succinic acid, methanesulfonic acid, acetic acid, trifluoroacetic acid, oxalic acid, propionic acid, tartaric acid, salicylic acid, citric acid, gluconic acid, lactic acid, mandelic acid, phenylacetic acid, aspartic acid, stearic acid, palmitic acid, glycolic acid, glutamic acid, benzoic acid, adipic acid, benzoic acid, benzenesulfonic acid, butyric acid, camphoric acid, camphorsulfonic acid, cyclohexylsulfamic acid, ethanesulfonic acid, fumaric acid, glutamic acid, glycolate, hemisulfuric acid, 2-hydroxyethanesulfonic acid, heptanoic acid, hexanoic acid, hydroxymaleic acid, lactic acid, malic acid, maleic acid, methanesulfonic acid, 2-naphthalenesulfonic acid, oxalic acid, bishydroxynaphthoic acid, phenylacetic acid, diphosphonic acid, picric acid, trimethylacetic acid, propionic acid, quinic acid, salicylic acid, stearic acid, succinic acid, sulfamic acid, sulfanilic acid, tartaric acid, succinic acid, methanesulfonic acid, benzoic acid, benzenesulfonic acid, toluene sulfonic acid (p-toluene sulfonic acid), trifluoroacetic acid, and undecanoic acid. Suitable inorganic acids include hydrochloric, hydrobromic, sulfuric, phosphoric, nitric, hydrogen carbonate, hydroiodic, persulfuric, and hydrogen sulfate. Base salts include salts with carboxylate anions and include salts with anions such as those selected from alkali metal salts such as sodium, lithium and potassium; alkaline earth metal salts such as aluminum, calcium, and magnesium salts; salts with organic bases such as choline, meglumine, piperazine, dicyclohexylamine salts, and N-methyl-D-glucamine; and salts with amino acids such as arginine, lysine, ornithine, etc. Thus, the term "pharmaceutically acceptable salt" of formula (I), formula (II), formula (III) or formula (IV) shall include and all acceptable salt forms.

Basic amino-containing groups can be quaternized with such agents as: lower alkyl halides, such as methyl, ethyl, propyl and butyl halides: dialkyl sulfates such as dimethyl, diethyl, dibutyl, diamyl sulfate; long chain halides such as decyl, lauryl, myristyl and stearyl halides; arylalkyl halides such as benzyl bromide and others. Non-toxic physiologically acceptable salts are preferred, although other salts may be used, for example for isolating or purifying the product. The salt may be formed by conventional means, for example by reacting the product in free base form with one or more equivalents of a suitable acid in a solvent or medium in which the salt is insoluble, removing the solvent in vacuo or by freeze drying, or by exchanging an existing anion of the salt for another anion on a suitable ion exchange resin.

Representative examples of prodrugs include, but are not limited to, acetate, formate and benzoate derivatives of alcohol and amine functional groups of compounds of the general formula (I), formula (II), formula (III) or formula (IV). Further, in the case of carboxylic acid (-COOH), esters such as methyl ester, ethyl ester, propyl ester, and the like may be included. In the case of hydroxyl groups, mixed anhydrides such as methoxy, ethoxy, propoxy, tert-butoxy, and the like may be included. The effect of long-acting is achieved by the compounds of formula (I) and their metabolites forming chain fatty ethers, long-chain fatty esters or long-chain fatty amides via hydroxyl or amino groups with long-chain fatty alcohols or acids, such as pentanol or acid, hexanol or acid, heptanol or acid, octanol or acid, nonanol or acid, decanol or acid, lauryl alcohol or acid, stearyl alcohol or acid, and the like.

The compounds of formula (I), formula (II), formula (III) or formula (IV) have one or more chiral centers, it being understood that the invention includes all such stereoisomers, including enantiomers and diastereomers. It is therefore to be understood that the present invention includes within its definition any such optically active or racemic form having the above-mentioned activity, provided that certain compounds of formula (I), formula (II), formula (III) or formula (IV) may exist in optically active or racemic form due to one or more asymmetric carbon atoms. The present invention includes all such stereoisomers having activity as defined herein.

The synthesis of the optically active form can be carried out by standard techniques of organic chemistry well known in the art, for example by synthesis from optically active starting materials, or by resolution of the racemic form. The racemates can be separated into the individual enantiomers using known methods (see, for example, Advanced Organic Chemistry: third edition: author J March, pages 104 to 107). Suitable methods include the formation of diastereomeric derivatives by reaction of the racemic material with a chiral auxiliary, followed by separation of the enantiomers, for example by chromatography, followed by cleavage of the auxiliary species. Similarly, the above activities can be assessed using standard laboratory techniques as described below.

Thus, throughout the specification, when reference is made to a compound of formula (I), formula (II), formula (III) or formula (IV), it is to be understood that the term compound includes isomers, mixtures of isomers and stereoisomers which are derivatives of pleuromutilin.

The configuration of the substituents attached to the asymmetric carbon atom of the thimerosal-tricycles (tricyclus) is preferably the same as the configuration of the native pleuromutilins.

Stereoisomers may be separated using conventional techniques, for example chromatography or fractional crystallisation. The enantiomers can be separated, for example, by separation of the racemates by fractional crystallization, resolution or HPLC. Diastereomers may be separated by separation by different physical properties of the diastereomers, for example, by fractional crystallization, HPLC, or flash chromatography. Specific stereoisomers may alternatively be made by chiral synthesis from chiral starting materials or by derivatization using chiral reagents under conditions that do not cause racemisation or epimerisation.

When a particular stereoisomer is provided (whether by separation, by chiral synthesis, or by other means), it may be advantageous to provide the stereoisomer substantially separately from other stereoisomers of the same compound. In one aspect, a mixture containing a particular stereoisomer of a compound of formula (I), formula (II), formula (III) or formula (IV) may contain less than 30% by weight, particularly less than 20% by weight and more particularly less than 10% by weight of other stereoisomers of the same compound. In another aspect, a mixture containing a particular stereoisomer of a compound of formula (I), formula (II), formula (III) or formula (IV) may contain less than 6% by weight, particularly less than 3% by weight and more particularly less than 2% by weight of other stereoisomers of the compound. In another aspect, a mixture containing a particular stereoisomer of a compound of formula (I), formula (II), formula (III) or formula (IV) may contain less than 1% by weight, particularly less than 0.5% by weight and more particularly less than 0.1% by weight of other stereoisomers of the compound.

It is to be understood that the present invention includes within its definition any such tautomeric form having the aforementioned activity, provided that certain compounds of formula (I), formula (II), formula (III) or formula (IV) as hereinbefore defined may exist in tautomeric form. Thus, the present invention relates to all tautomeric forms of the compounds of formula (I), formula (II), formula (III) or formula (IV), whether or not explicitly detailed in the specification.

It is also understood that certain compounds of formula (I), formula (II), formula (III) or formula (IV) and pharmaceutically acceptable salts thereof may exist in solvated as well as unsolvated forms such as hydrated forms. It is to be understood that the invention encompasses all such solvated forms. For the sake of clarity, this includes solvated (e.g. hydrated) forms of the free form of the compound, as well as solvated (e.g. hydrated) forms of the salt of the compound.

For the sake of clarity, it is understood that formula (I), formula (II)The atom of the compound of formula (II), (III) or (IV) and any examples or embodiments disclosed herein is intended to encompass all isotopes of that atom. For example, H (or hydrogen) includes any isotopic form of hydrogen, including 1H,²H (D) and³h (T); c includes any isotopic form of carbon, including¹²C、¹³C and¹⁴c; o includes any isotopic form of oxygen, including¹⁶O、¹⁷O and¹⁸o; n includes any isotopic form of nitrogen, including¹³N、¹⁴N and¹⁵n; p includes any isotopic form of phosphorus, including³¹P and³²p; s includes any isotopic form of sulfur, including³²S and³⁵s; f includes any isotopic form of fluorine, including¹⁹F and¹⁸f; cl includes any isotopic form of chlorine, including³⁵Cl、³⁷Cl and³⁶cl, and the like. In one aspect, the compound of formula (I), formula (II), formula (III) or formula (IV) comprises an isotope of an atom encompassed therein in an amount corresponding to its naturally occurring abundance. However, in certain instances, it is desirable to have atoms enriched in one or more particular isotopes that are typically naturally present in lower abundance. For example,¹h is typically present in an abundance of greater than 99.98%, however, in one aspect, the compounds of the invention may be enriched at one or more positions where H is present¹H or³H. In another aspect, when the compounds of the invention are enriched in radioisotopes, for example³H and¹⁴c, the compounds are useful in drug and/or substrate tissue distribution assays. It is to be understood that the present invention encompasses all such isotopic forms which are useful in the treatment of bacterial infections.

II. Application method

The compounds of the present invention have the general formula (I), formula (II), formula (III) or formula (IV) or are stereoisomers, prodrugs, active metabolites or pharmaceutically acceptable salts, solvates or crystalline forms thereof, exhibit antimicrobial activity.

For example, the following antibacterial activities: against gram-positive bacteria, for example, coagulase-positive Staphylococci (Staphylococi) such as Staphylococcus aureus (Staphyloccocus aureus), coagulase-negative Staphylococci such as Staphylococcus epidermidis (Staphyloccocus epidermidis), Staphylococcus haemolyticus (Staphyloccocus haemolyticus), and Streptococcus (Streptococcus) such as Streptococcus pyogenes (Streptococcus pyogenes), Streptococcus pneumoniae (Streptococcus pneumae), Enterococci (Enterococci) such as enterococcus faecium (Enterococcus faecium), and Listeria monocytogenes (Listeria monocytogenes); and anti-gram-negative bacteria such as Moraxella (Moraxella) such as Moraxella catarrhalis (Moraxella catarrha1is), and Haemophilus such as Haemophilus influenzae (Haemophilus influenzae), and Legionella such as Legionella pneumophila (Legionella), Neisseria such as Neisseria gonorrhoeae (Neisseria gonorrhoeae); and anti-mycoplasmas (mycoplasmas), Chlamydia (Chlamydia), and obligate anaerobes such as bacteroides fragilis (bacteroides fragilis), Clostridium difficile (Clostridium difficile), Clostridium (Fusobacterium spp.), and Propionibacterium (Propionibacterium spp.).

In vitro Activity against aerobic bacteria by agar dilution test or microdilution test according to the Clinical Laboratory Standards Committee (CLSI, formerly NCCLS) document No. M7-A7Vo1.26, No.2: "Methods for dilution of Antimicrobial Succinity Tests for bacteria that Grow aerobic-applied Standard; measurement was performed in the seventh edition (2006) ": anti-anaerobe assays were performed according to the clinical laboratory standards Committee (CLSI, Pre-NCCLS) document No. Mll-A6, Vo1.24, No.2: "Methods for anti-microbial Suceptibility Testing of anti-aerobic Bacteria-applied Standard; sixth edition (2004) ": and in vivo activity was tested against staphylococcus aureus in a mouse model of septicemia.

The compounds of the invention are therefore suitable for the treatment and prophylaxis of diseases which are mediated by microorganisms, for example bacteria. Diseases that can be treated also include, for example, Helicobacter (Helicobacter pylori) mediated diseases and Mycobacterium tuberculosis (Mycobacterium tuberculosis) mediated diseases. Diseases that can be treated also include inflammatory diseases in general, where the inflammation, including acne for example, is mediated by microorganisms.

In another aspect, the present invention provides a compound of the invention for use as a medicament, preferably as an antimicrobial such as an antibiotic and e.g. an anti-anaerobic medicament.

In another aspect, the invention provides a compound of the invention for use in the treatment of acne.

In a further aspect, the present invention provides a compound of the invention for use in the manufacture of a medicament for the treatment of a disease mediated by a microorganism, such as a bacterium, for example a disease mediated by a bacterium, for example selected from staphylococci, streptococci, enterococci; diseases mediated by bacteria, for example selected from moraxella, haemophilus, legionella, neisseriaceae; diseases mediated by helicobacter; diseases mediated by mycobacterium tuberculosis; diseases mediated by mycoplasma, chlamydia and obligate anaerobes.

Use of a compound of the invention having the general formula (I), formula (II), formula (III) or formula (IV) or being a stereoisomer, prodrug, active metabolite or pharmaceutically acceptable salt, solvate or crystalline form thereof, in the manufacture of a medicament for the treatment of a human disease state mediated by a bacterial infection.

The compounds of the invention are also suitable for use as veterinary, e.g. veterinary active compounds, e.g. for the prevention and treatment of microbial, e.g. bacterial diseases, in animals such as poultry, pigs and cattle: and dilution liquids such as are used in artificial insemination and egg soaking techniques. In another aspect, the present invention provides a compound of the invention for use as a veterinary drug. In a further aspect, the present invention provides a compound of the invention for use in the preparation of a veterinary composition for use as a veterinary drug. In another aspect, the present invention provides a veterinary method of prevention and treatment of microbial, e.g. bacterial, diseases, which comprises administering to a subject in need of such treatment an effective amount of a compound of the present invention, e.g. in the form of a veterinary composition.

In one aspect, the terms "infection" and "bacterial infection" may refer to a gynecological infection. In another aspect, the terms "infection" and "bacterial infection" may refer to a Respiratory Tract Infection (RTI). In yet another aspect, the terms "infection" and "bacterial infection" may refer to a sexually transmitted disease. In yet another aspect, the terms "infection" and "bacterial infection" may refer to Urinary Tract Infection (UTI). In a further aspect, the terms "infection" and "bacterial infection" may refer to an acute episode of chronic bronchitis (ACEB). In a still further aspect, the terms "infection" and "bacterial infection" may refer to acute otitis media. In one aspect, the terms "infection" and "bacterial infection" may refer to acute sinusitis. In another aspect, the terms "infection" and "bacterial infection" may refer to an infection by a drug-resistant bacterium. In yet another aspect, the terms "infection" and "bacterial infection" may refer to catheter-related sepsis. In yet another aspect, the terms "infection" and "bacterial infection" may refer to chancroid. In a further aspect, the terms "infection" and "bacterial infection" may refer to chlamydia. In a still further aspect, the terms "infection" and "bacterial infection" may refer to community-acquired pneumonia (CAP). In still a further aspect, the terms "infection" and "bacterial infection" may refer to a complex skin and skin structure infection. In one aspect, the terms "infection" and "bacterial infection" may refer to uncomplicated skin and skin structure infections. In another aspect, the terms "infection" and "bacterial infection" may refer to endocarditis. In yet another aspect, the terms "infection" and "bacterial infection" may refer to febrile neutropenia. In yet another aspect, the terms "infection" and "bacterial infection" may refer to gonococcal meningitis. In a further aspect, the terms "infection" and "bacterial infection" may refer to gonococcal urethritis. In a still further aspect, the terms "infection" and "bacterial infection" may refer to nosocomial pneumonia (HAP). In yet another aspect, the terms "infection" and "bacterial infection" may refer to osteomyelitis. In a further aspect, the terms "infection" and "bacterial infection" may refer to sepsis. In a still further aspect, the terms "infection" and "bacterial infection" may refer to syphilis. In a further aspect, the terms "infection" and "bacterial infection" may refer to intra-abdominal infection (IAI).

In one embodiment of the present invention, the terms "infection" and "bacterial infection" refer to an infection by a gram-negative bacterium, also referred to as a "gram-negative infection". In one aspect of this embodiment, the gram-negative infection is an infection resistant to one or more antibiotics. In one aspect of this embodiment, the gram-negative infection is an infection that is resistant to multiple drugs.

In one embodiment of the invention, the terms "infection" and "bacterial infection" refer to an infection by a gram-positive bacterium, also referred to as a "gram-positive infection". In one aspect of this embodiment, the gram-positive infection is an infection resistant to one or more antibiotics. In one aspect of this embodiment, the gram-positive infection is an infection that is resistant to multiple drugs.

the compounds of the invention may be administered in combination with any antibiotic belonging to, but not limited to, the group of patulin (clavams), carbapenems, macrolides, penicillins and/or cephalosporins, or being a stereoisomer, prodrug, active metabolite or pharmaceutically acceptable salt, solvate or crystalline form thereof, in one aspect of the invention, in combination with one or more of penicillin, methicillin, oxacillin, nafcillin, cloxacillin, dicloxacillin, flucloxacillin, temocillin, amoxicillin, ampicillin, amoxicillin, aroline, carbenicillin, cocaine, meropenem, piperacillin, cephalexin, CXA-101, cefazolin, cefaclor, cefuroxime, cefamandole, cefotetan, cefoxitin, ceftriaxone, cefotaxime, cefpodoxime, phacidoxime, cefepime, or (paminolate, or cefepime, or p, cefepime, or p, or cefepime, or.

In one aspect of the invention, the compounds of the invention are combined with one or more of the following: aminoglycosides, spectinomycin, macrolides, ketolides, streptogramins, oxazolidinones, tetracyclines, fluoroquinolones, coumarin antibiotics, glycopeptides, lipoglycopeptides, nitroimidazoles, ansamycins, lichol, mupirocin (mupirocyn), fosfomycin, tobramycin, linezolid, daptomycin, vancomycin, tazobactam, avibactam, clavulanic acid, LK-157, LK-176, SA-1-204, SA-2-13, BLI-489(Pfizer/Wyeth), BAL0029880(Baselea), and/or relebatam (MK 7655).

III、Preparation

Pure forms or suitable pharmaceutical compositions of the compounds of the present invention or pharmaceutically acceptable salts thereof may be administered by any acceptable mode of administration of agents that serve similar utilities. The pharmaceutical compositions of the present invention may be prepared by combining a compound of the present invention with a suitable pharmaceutically acceptable carrier, diluent or excipient, and may be formulated into solid, semi-solid, liquid or gaseous form preparations such as tablets, capsules, powders, granules, ointments, solutions, suppositories, injections, inhalants, gels, microspheres and aerosols. Typical routes of administration for the pharmaceutical compositions include, but are not limited to, oral, topical, transdermal, inhalation, parenteral, sublingual, buccal, rectal, vaginal, and intranasal administration. As used herein, the term parenteral includes subcutaneous injections, intravenous, intramuscular, intrasternal injection or infusion techniques. The pharmaceutical compositions of the present invention are formulated to allow the active ingredients contained therein to be bioavailable upon administration of the composition to a patient. One or more dosage units in the composition to be administered to a subject or patient, wherein, for example, a capsule can be a single dosage unit and a container containing a compound of the invention in aerosol form can hold a plurality of dosage units. The actual methods of preparing the dosage forms are known, or will be known, to those skilled in the art. The compositions to be administered will contain, in any event, a therapeutically effective amount of a compound of the present invention, or a pharmaceutically acceptable salt thereof, in order to treat the disease or condition of interest in accordance with the teachings of the present invention.

Pharmaceutical compositions

The pharmaceutical compositions of the present invention may be in solid or liquid form. In one aspect, the carrier is a microparticle, such that the composition is in the form of, for example, a tablet or powder. The carrier can be a liquid and the composition can be, for example, an oral syrup, an injectable liquid, or an aerosol suitable for administration, for example, by inhalation. When intended for oral administration, the pharmaceutical composition is preferably selected from solid or liquid forms, wherein semi-solid, semi-liquid, suspension and gel forms are included in the forms considered herein to be solid or liquid. For oral solid compositions, the pharmaceutical compositions can be formulated in the form of powders, granules, compressed tablets, pills, capsules, chewable tablets, powder tablets, and the like. Such solid compositions typically contain one or more inert diluents or edible carriers. In addition, one or more binders such as carboxymethyl cellulose, ethyl cellulose, microcrystalline cellulose, xanthan gum or gelatin; excipients, such as starch, lactose or dextrin; disintegrating agents, such as alginic acid, sodium alginate, Primogel, corn starch, and the like; lubricants, such as magnesium stearate or hydrogenated vegetable oil (Sterotex); glidants, such as colloidal silicon dioxide; sweetening agents, such as sucrose or saccharin; a flavoring agent, such as peppermint, methyl salicylate, or orange flavoring; and a colorant.

When the pharmaceutical composition is in the form of a capsule, for example a gelatin capsule, it may contain, in addition to materials of the above type, a liquid carrier, for example polyethylene glycol or an oil. The pharmaceutical composition may be in liquid form, such as a tincture, syrup, solution, emulsion, or suspension. Such liquids may be administered orally, or delivered by injection, as two examples. When intended for oral administration, the compositions preferably contain one or more of sweeteners, preservatives, dyes/colorants and flavor enhancers in addition to the compounds of the present invention. In compositions intended for administration by injection, one or more of surfactants, preservatives, wetting agents, dispersing agents, suspending agents, buffering agents, stabilizing agents and isotonicity agents may be included.

The liquid pharmaceutical compositions of the present invention, whether in solution, suspension or other similar form, may include one or more adjuvants selected from sterile diluents such as water for injection, physiological saline solutions, preferably physiological saline, Ringer's solution, isotonic sodium chloride, fixed oils (e.g., synthetic mono-or diglycerides which may be used as a solvent or suspending medium), polyethylene glycols, glycerol, propylene glycol and the like; antibacterial agents such as benzyl alcohol or methyl paraben; antioxidants, such as ascorbic acid or sodium bisulfite; chelating agents, such as ethylenediaminetetraacetic acid; buffers such as acetates, citrates or phosphates, and agents for adjusting tonicity such as sodium oxide or dextrose. Parenteral formulations can be packaged in ampules, disposable syringes or multi-dose vials made of glass or plastic. Physiological saline is a preferred adjuvant. The injectable pharmaceutical composition is preferably sterile.

Liquid pharmaceutical compositions of the invention intended for parenteral or oral administration should contain an amount of a compound of the invention such that a suitable dosage is obtained. The pharmaceutical compositions of the invention may be intended for topical administration, in which case the carrier preferably comprises a solution, emulsion, ointment or gel base. For example, the base may comprise one or more of paraffin oil, lanolin, polyethylene glycol, beeswax, mineral oil, diluents (e.g., water and alcohol), and emulsifiers and stabilizers. Thickeners may be present in the pharmaceutical compositions for topical administration. If transdermal administration is intended, the composition may comprise a transdermal patch or an iontophoretic device.

The pharmaceutical compositions of the invention may be intended for rectal administration, in the form of suppositories for example, which will melt in the rectum and release the drug. Compositions for rectal administration may contain an oily base as a suitable non-irritating excipient. Such bases include, but are not limited to, lanolin, cocoa butter, and polyethylene glycols.

The pharmaceutical compositions of the present invention may include a variety of substances that alter the physical form of the solid or liquid dosage unit. For example, the composition may include a material that forms a coating shell around the active ingredient. The material forming the coating shell is generally inert and may be selected from, for example, sugars, shellac, and other enteric coating agents. Alternatively, the active ingredient may be encapsulated in a gelatin capsule.

The pharmaceutical compositions of the invention in solid or liquid form may include an agent that binds to the compound of the invention and thereby aids in the delivery of the compound. Suitable agents with this capability include monoclonal or polyclonal antibodies, proteins or liposomes.

The pharmaceutical compositions of the present invention may be comprised of dosage units that can be administered in the form of an aerosol. The term aerosol is used to denote a variety of systems ranging from the jelly-like variety to systems consisting of pressurized packs. Delivery may be by liquefied or compressed gas, or by a suitable pump system that dispenses the active ingredient. Aerosols of the compounds of the invention may be delivered as single, two, or three phase systems to deliver the active ingredient. The delivery side of the aerosol comprises the necessary containers, actuators, valves, sub-containers, etc., which together may form a kit. Preferred aerosols are determined by one of skill in the art without undue experimentation.

The pharmaceutical compositions of the present invention can be prepared by methods well known in the pharmaceutical arts. For example, pharmaceutical compositions to be administered by injection may be prepared by combining a compound of the invention with sterile distilled water to form a solution. Surfactants may be added to facilitate the formation of a homogeneous solution or suspension. Surfactants are compounds that non-covalently interact with the compounds of the present invention, thereby facilitating dissolution or uniform suspension of the compounds in an aqueous delivery system.

For the carrier of the injectable preparation, for example, distilled water, or a physiological salt solution, a glucose solution, other isotonic solution is used. The carrier for capsules, powders, granules or tablets is used in admixture with known pharmaceutical excipients (for example, starch, maltose, sucrose, calcium carbonate or calcium phosphate), binders (for example, starch, gum arabic, carboxymethylcellulose, hydroxypropylcellulose or crystalline cellulose), lubricants (for example, magnesium stearate, talc) and disintegrants (for example, carboxymethylcellulose and talc). Thus, the pharmaceutical and antibacterial compositions for use in this specification may contain formula (I), formula (II), formula (III) or formula (IV) alone or in stereoisomeric, prodrug, active metabolite or pharmaceutically acceptable salt, solvate or crystalline form thereof, or may contain a carrier as described above, or may contain an appropriate amount of any other suitable antibacterial compound.

The compounds of the present invention, or pharmaceutically acceptable salts thereof, are administered in a therapeutically effective amount, which will vary depending on a variety of factors, including the activity of the particular compound employed; metabolic stability and length of action of the compound; the age, weight, general health, sex, and diet of the patient; mode and time of administration; the rate of excretion; a pharmaceutical composition; the severity of the particular disorder or condition; and individuals undergoing therapy.

The compounds of the present invention or pharmaceutically acceptable derivatives thereof can also be administered simultaneously with, before, or after the administration of one or more other therapeutic agents. Such combination therapy includes the administration of a single pharmaceutical dosage formulation containing a compound of the present invention and one or more other active agents, as well as the administration of separate pharmaceutical dosage formulations of a compound of the present invention with each active agent itself. For example, a compound of the invention and another active agent can be administered to a patient in a single orally administered composition (e.g., a tablet or capsule), or each agent can be administered as a separate orally administered formulation. In the case of separate administration formulations, the compound of the invention and the one or more additional active agents may be administered substantially at the same time (i.e., simultaneously) or at separately staggered times (i.e., sequentially); combination therapy is understood to include all such regimens.

The pharmaceutical combination of the present invention can be administered in a dose adjusted depending on the disease state, the administration route, the age or body weight of the patient. For oral administration to adults, the dosage is usually 0.1-100 mg/kg/day, preferably 1-60 mg/kg/day. The appropriate dose of the present invention should be set in consideration of the age, body weight, condition, administration route, etc. of the patient, and the oral administration is usually in the range of 0.5 to 100 mg/kg/day, preferably 1 to 60 mg/kg/day. For parenteral administration, the dosage of the pharmaceutical composition of the present invention varies widely depending on the route of administration, but is usually in the range of 0.05 to 50 mg/kg/day, preferably 1 to 20 mg/kg/day.

The dose of the compound of the present invention varies depending on the administration method, age, body weight, condition of the patient and the kind of the disease, but in general, in the case of oral administration, the dose is about 0.5mg to 3000mg, preferably about 100mg to 2000mg per 1 day for an adult. Can be administered separately as required. In addition, in the case of parenteral administration, the dose is about 1mg to 10000mg, preferably about 10mg to 1000mg per 1 day for an adult. It is to be understood that in the present invention, combinations of substituents and/or variables of the formulas are permissible only if they result in stable compounds.

V. examples

The colloquial name refers to the IUPAC system name (1S, 2R, 3S, 4S, 6R, 7R, 8R, 14R) -3, 6-dihydroxy-2, 4, 7, 14-tetramethyl-4-vinyl-tricyclo [5.4.3.0 ]^1,8]Tetradecan-9-one. In these examples, pleuromutilin derivatives, such as BC-3781, are numbered similarly to the numbering system for the gimelin described in H.Berner (Berner, H.; Schulz, G.; Schneider H.tetrahedron 1980,36, 1807-1811):

14-O- { [ (1R,2R,4R) -4-amino-2-hydroxycyclohexyl-sulfanyl ] acetyl } -mordenin (BC-3781) was prepared according to Chinese patent CN201410406131. X.

Certain preferred embodiment aspects of the invention are shown by way of illustration in the following non-limiting examples. The reagents are commercially available or prepared according to literature procedures.

The novel pleuromutilin derivatives disclosed herein may be prepared by microbial transformation or by methods well known in the art of organic chemistry. If desired, the reaction product is optionally purified using conventional techniques including, but not limited to, filtration, distillation, crystallization, chromatographic separation and purification. Such materials are optionally characterized by conventional means including physical constants and spectral data.

Mass spectrum: electrospray mass spectra were recorded on a Waters single quadrupole mass spectrometer using flow injection in alternating positive and negative ion mode. The mass range was 120-2000Da, the capillary voltage was set at 4500V, and nitrogen was used for atomization.

LC-MS spectrometer detector, Waters, PDA (200-320nm), mass detector: ZQ

Eluent: a is acetonitrile containing 0.05 percent of trifluoroacetic acid; acetonitrile/water-1/9 (v/v) with 0.05% trifluoroacetic acid.

Example 1.14-O- { [ (1R,2R,4R) -4-amino-2-hydroxycyclohexyl-sulfanyl ] acetyl } -2, 2, 4-trideuterometrine the synthetic route is as follows:

example 114-O- { [ (1R,2R,4R) -4-amino-2-hydroxycyclohexyl-sulfanyl ] acetyl } -2, 2, 4-trideuterometrine

19.8g of intermediate 5 are dissolved in 40ml of isopropanol and stirred at room temperature. Adding 35g of 85% phosphoric acid, heating to 50 ℃, reacting for 18hr, and detecting complete reaction by TLC. Cooled to room temperature and 150ml dichloromethane were added. Cooling the reaction solution to 5-10 ℃ and controlling the temperature, dropwise adding a saturated sodium bicarbonate solution, adjusting the pH to 7-8, extracting the water layer once again by using a dichloromethane layer, combining the dichloromethane layers, washing for 1 time by using water, and washing for one time by using a saturated sodium chloride solution. Concentrating under reduced pressure to viscous state, adding 200ml MTBE, and pulping for 1hr to obtain the compound of example 1. LC-Ms ESI +: 511.3[ M + H]⁺,533.3[M+Na]⁺The deuteration rate is 98.5%.

¹H-NMR(CDCl₃400MHz, partial characteristic peaks 0.75(3H, d), 0.88 to 0.90(3H, d),1.23(3H, s),1.46(3H, s),1.90 to 1.94(1H, m),2.18 to 2.23(1H, m),2.93 to 2.81(1H, m),3.23 to 3.26(2H, AB),3.38(1H, m),3.42 to 3.48(1H, m),5.19 to 5.23(2H, m),5.82 to 5.84(1H, d),6.49 to 6.53(1H, m).

Intermediate 1: 2, 2, 4-Trideuterome forest

10g of pleuromutilin (Jiangsu Fengyuan bioengineering Co., Ltd.), 80mL of deuterated methanol was added and dissolved by stirring, 40g of deuterium water (purchased from Sahn chemical technology (Shanghai) Co., Ltd.) and 7.5g of 40% NaOD solution (purchased from Sahn chemical technology (Shanghai) Co., Ltd.) were added and stirred at 40 ℃ for 20hr, and the reaction was detected by TLC to be complete. The methanol was evaporated under reduced pressure, the pH was adjusted to 5-6 with dilute hydrochloric acid, and extraction was carried out with dichloromethane. The dichloromethane layer was spin dried to give 8.6g of a white solid. LC-Ms ESI +: 346.2[ M + Na ]]⁺And the deuteration rate is 98.6 percent.

¹H-NMR(CDCl₃300MHz, partial characteristic peaks 0.93-0.99(6H, m),1.12-1.18(4H, m),1.31-1.78(14H, m),1.89-1.93(1H, m),2.19-2.22(1H, m),3.42(1H, m),4.36(1H, m),5.29-5.40(2H, m),6.13-6.21(1H, m).

Intermediate 2: (R) - (3-Cyclohexenyl) -carbamic acid tert-butyl ester

61g of (R) -3-cyclohexenecarboxylic acid (available from Sahn's chemical technology, Shanghai, Ltd.) and 600mL of toluene were added to the reaction flask at room temperature and stirred. 53.8g of triethylamine is added dropwise and stirred for 30min after the dropwise addition. Then, 140g of DPPA (diphenylphosphoryl azide) was added dropwise, the temperature was slowly raised to 95 ℃ and the stirring was maintained for 15min, followed by heating and refluxing. TLC detection of the disappearance of the starting material, the reaction solution was cooled to 80 ℃ and 180g of tert-butanol was added dropwise, followed by 1.6g of CuCl. All mixtures were then warmed to 100 ℃ and stirred for an additional 50min, and the reaction was complete by TLC. The reaction solution was cooled, and 4L of a saturated aqueous solution of sodium hydrogencarbonate was added. The mixture was stirred for 2 hr. Filtering, extracting mother liquor, washing the organic layer with water for 2 times, and evaporating the mother liquor under reduced pressure to obtain a crude product. Purifying the crude product by flash silica gel column chromatography, and eluting the crude product with an eluent: ethyl acetate/petroleum ether to yield 81.1g of a solid.

Intermediate 3: (1R, 3R, 6S) - (7-oxa-bicyclo [4.1.0] hept-3-yl) -carbamic acid tert-butyl ester

135g m-CPBA (m-chloroperoxybenzoic acid) and 730mL of methylene chloride were added to the reactor and the mixture was cooled to 10-15 ℃. Slowly adding 90g (R) - (3-cyclohexenyl) -carbamic acid tert-butyl ester solution in dichloromethane (150ml) dropwise, maintaining the reaction system at below 25 ℃, adding 45ml dichloromethane, stirring for 50min after the addition is finished, then heating to reflux, carrying out reflux reaction for 2hr, and detecting the reaction completion by TLC. Cooling the reaction solution to-5-0 deg.C, stirring for 2hr, filtering, washing the filtrate with 10% sodium thiosulfate solution for 2 times, washing with 10% sodium bicarbonate solution for 2 times, washing with water for 2 times, concentrating under reduced pressure to minimum volume, adding toluene, distilling under reduced pressure again, repeating the operation for 1 time to obtain toluene solution (content: 75%) of intermediate 3.

Intermediate 4 { (1R,2R,4R) -4- [ (tert-Butoxycarbonyl) -amino ] -2-hydroxy-cyclohexyl-sulfanyl } -acetic acid

27.3g of a toluene solution (content: 75%) of the above intermediate 3 was added with 30ml of toluene, and stirred at room temperature. Below 30 ℃, 9g of thiothioglycolic acid/10 ml of toluene solution are added dropwise. Adding 0.85g tetrabutylammonium chloride monohydrate, heating to 40-45 deg.C, stirring for 4hr, and detecting by TLC to obtain the final product. Cooling the reaction solution to 15 ℃, dropwise adding a 5% sodium bicarbonate aqueous solution into the reaction solution, extracting under the condition that the pH value is 6-7, and washing with water for 2 times. The organic layer was concentrated to give an oil, which was recrystallized from toluene/heptane and dried in vacuo to give intermediate 4.

Intermediate 14-O- { [ (1R,2R,4R) -4-tert-Butoxycarbonylamino-2-hydroxycyclohexyl-sulfanyl ] acetyl } -2, 2, 4-trideuterourea

14g of the intermediate 4 are dissolved in 60ml of dichloromethane, the temperature is reduced to 0 to 5 ℃, 4.6g of oxalyl chloride is added dropwise, 0.7g of DMF is added dropwise, the temperature is raised to room temperature after the addition, and the mixture is stirred for 30 min. The solvent was evaporated under reduced pressure and 20ml of dichloromethane were taken up 3 times. The acid chloride intermediate was dissolved in 40ml of dichloromethane for further use.

14.8g 2, 2, 4-three deuterated precursor forest dissolved in 100ml dichloromethane, cooled to 5-10 ℃, slowly added with 5.6g triethylamine, slowly added with the acyl chloride intermediate solution (triethylamine and acyl chloride solution are simultaneously added). After the addition, the temperature is raised to 15-20 ℃. The reaction was carried out for 2hr, and the reaction was terminated by TLC. Adding water into the reaction solution, extracting to obtain a dichloromethane layer, washing with water for 2 times, distilling under reduced pressure, purifying by rapid silica gel column chromatography, and taking ethyl acetate/petroleum ether as an eluting solvent. 16.9g of a white solid, 14-O- { [ (1R,2R,4R) -4-tert-butoxycarbonylamino-2-hydroxycyclohexyl-sulfanyl, are obtained]Acetyl } -2, 2, 4-trideuterometrine. LC-Ms ESI +: 656.3[ M +2Na]⁺。

Example 2.14-O- { [ (1R,2R,4R) -4-amino-2-hydroxycyclohexyl-sulfanyl ] acetyl } -2, 2-dideuterourea-Line

The synthetic route is as follows:

example 214-O- { [ (1R,2R,4R) -4-amino-2-hydroxycyclohexyl-sulfanyl ] acetyl } -2, 2-dideuterohumol

19.8g of intermediate 7 are dissolved in 40ml of isopropanol and stirred at room temperature. Adding 35g of 85% phosphoric acid, heating to 50 ℃, reacting for 18hr, and detecting complete reaction by TLC. Cooled to room temperature and 150ml dichloromethane were added. Cooling the reaction solution to 5-10 deg.C, controlling temperature, adding saturated sodium bicarbonate solution dropwise, adjusting pH to 7-8, extracting, collecting dichloromethane layer againThe aqueous layer was extracted once, the dichloromethane layers were combined, washed 1 time with water and once with saturated sodium chloride solution. Concentrating under reduced pressure to viscous state, adding 200ml MTBE, and pulping for 1hr to obtain the compound of example 2. LC-Ms ESI +: 510.3[ M + H]⁺The deuteration rate is 95.1%.

¹H-NMR(CDCl₃400MHz, partial characteristic peaks 0.76(3H, d), 0.89-0.91(3H, d),1.22(3H, s),1.46(3H, s),1.92-1.95(1H, m),2.19-2.24(1H, m),2.66(1H, s),2.81-2.92(1H, m),3.25-3.27(2H, AB),3.39(1H, m),3.41-3.49(1H, m),5.20-5.23(2H, m),5.83-5.86(1H, d),6.50-6.55(1H, m).

Intermediate 6: 2, 2-dideuterothimers

10g of pleuromutilin is added with 80ml of methanol and stirred to be dissolved, 40g of water is added, 7.5g of 40 percent NaOH solution is added, the mixture is stirred for 8 hours at 40 ℃, and the TLC detection reaction is completed. Adjusting ph to 5-6 with 2N hydrochloric acid, extracting with dichloromethane, and washing with water once. The organic layer was evaporated under reduced pressure to remove the solvent, and 50ml of methanol was added thereto and evaporated again. Then adding 80ml of deuterated methanol, stirring and dissolving, adding 10g of deuterium oxide, adding 0.125g of 40% NaOD solution, stirring at 40 ℃ for 10hr, dilute hydrochloric acid acidifying, dichloromethane extracting, spin-drying the dichloromethane layer to obtain a white solid, dissolving the white solid in 60ml of deuterated methanol again, stirring and dissolving, adding 6g of deuterium oxide, adding 0.10g of 40% NaOD solution, stirring at 40 ℃ for 10hr, dilute hydrochloric acid acidifying, dichloromethane extracting, and spin-drying the dichloromethane layer to obtain 6.5g of white solid. LC-Ms ESI +: 345.2[ M + Na ]]⁺And the deuteration rate is 95.8 percent.

¹H-NMR(CDCl₃300MHz, partial characteristic peaks 0.90-0.98(6H, m),1.10-1.18(4H, m),1.29-1.76(14H, m),1.91-2.00(1H, m),2.18-2.21(1H, m),2.65(1H, s),3.44(1H, m),4.35(1H, m),5.28-5.38(2H, m),6.11-6.19(1H, m).

Intermediate 7-14-O- { [ (1R,2R,4R) -4-tert-Butoxycarbonylamino-2-hydroxycyclohexyl-sulfanyl ] acetyl } -2, 2-dideuterourea forest

14.8g of 2, 2-dideuterourea in 100ml of dichloromethane, cooling to 5-10 ℃, slowly adding 5.6g of triethylamine dropwise, and slowly adding the acyl chloride intermediate solution dropwise (adding triethylamine and the acyl chloride solution dropwise at the same time). After the addition, the temperature is raised to 15-20 ℃. The reaction was carried out for 2hr, and the reaction was terminated by TLC. Adding water into the reaction solution, extracting to obtain a dichloromethane layer, washing with water for 2 times, distilling under reduced pressure, purifying by rapid silica gel column chromatography, and taking ethyl acetate/petroleum ether as an eluting solvent. 15.3g of a white solid, 14-O- { [ (1R,2R,4R) -4-tert-butoxycarbonylamino-2-hydroxycyclohexyl-sulfanyl, are obtained]Acetyl } -2, 2-dideuterometrine. LC-Ms ESI +: 632.4[ M + Na ]]⁺。

Example 3.14-O- { [ (1R,2R,4R) -4-ethylamino-2-hydroxycyclohexyl-sulfanyl ] acetyl } -2, 2, 4-trideuterometrine

14-O- { [ (1R,2R,4R) -4-amino-2-hydroxycyclohexyl-sulfanyl]Acetyl } -2, 2, 4-trideuteromer 1.8g was dissolved in 20ml of dichloromethane, and 5.6ml of acetaldehyde and 0.14ml of concentrated sulfuric acid were added thereto, followed by stirring at room temperature for 3 hr. To the reaction mixture was added 1.5g of sodium triacetoxyborohydride, and stirred for 4 hr. Quenching the solution with water, extracting with dichloromethane, concentrating the organic layer under reduced pressure, purifying by flash silica gel column chromatography, and performing gradient elution with ethyl acetate/methanol/ammonia water. 0.36g of the expected product is obtained. LC-Ms ESI +: 561.3[ M + Na ]]⁺And the deuteration rate is 98.6 percent.

¹H-NMR(CDCl₃400MHz, partial characteristic peaks 0.71(3H, d), 0.88 to 0.90(3H, d),1.13(t,3H, NCH2CH3), 1.20(3H, s),1.41(3H, s),1.89 to 1.92(1H, m),2.15 to 2.21(1H, m),2.65(q,2H, NCH2), 2.90 to 2.79(1H, m),3.21 to 3.26(2H, AB),3.35(1H, m),3.40 to 3.45(1H, m),5.18 to 5.24(2H, m),5.81 to 5.83(1H, d),6.45 to 6.51(1H, m).

Example 4.14-O- { [ (1R,2R,4R) -4-diethylamino-2-hydroxycyclohexyl-sulfanyl ] acetyl } -2, 2, 4-trideuterourea forest

14-O- { [ (1R,2R,4R) -4-diethylamino-2-hydroxycyclohexyl-sulfanyl]Acetyl } -2, 2, 4-trideuterosomalene (3 g) was dissolved in 30ml of acetone/acetonitrile (1: 1), and 1.73g of sodium bicarbonate was added thereto, followed by stirring at room temperature for 30 min. 2.75g of iodoethane was added to the reaction mixture, and stirred for 18 hr. Quenching the solution with water, extracting with dichloromethane, concentrating the organic layer under reduced pressure, purifying by flash silica gel column chromatography, and performing gradient elution with ethyl acetate/methanol/ammonia water. 1.5g of the expected product are obtained. LC-Ms ESI +: 589.4[ M + Na ]]⁺And the deuteration rate is 98.4%.

¹H-NMR(CDCl₃400MHz, partial characteristic peaks 0.70(3H, d), 0.88 to 0.91(3H, d),0.98(t,6H, NCH2CH3), 1.15(3H, s),1.40(3H, s),1.89 to 1.92(1H, m),2.15 to 2.21(1H, m),2.55(m,4H, NCH2), 2.79 to 2.81(1H, m),3.22 to 3.25(2H, AB),3.36(1H, m),3.41 to 3.44(1H, m),5.19 to 5.26(2H, m),5.80 to 5.84(1H, d),6.44 to 6.52(1H, m).

Example 5.14-O- { [ (1R,2R,4R) -4-amino-2-hydroxycyclohexyl-sulfanyl ] acetyl } -19, 20-dihydro-2, 2, 4-trideuterometrine

Adding 14-O- { [ (1R,2R,4R) -4-amino-2-hydroxycyclohexyl-sulfanyl]Acetyl } -2, 2, 4-trideuteromer forest 1g, dissolved in 20ml of methanol. 0.6g of 10% palladium on charcoal was added and hydrogenation was continued at 25 ℃ for 8 hr. After the reaction, the mixture was filtered, and the filter cake was washed with methanol. Spin-drying the filtrate, purifying by flash silica gel column chromatography, and performing gradient elution with ethyl acetate/methanol/ammonia water. This gives 0.8g of 14-O- { [ (1R,2R,4R) -4-amino-2-hydroxycyclohexyl-sulfanyl]Acetyl } -19, 20-dihydro-2, 2, 4-trideuterometrine. LC-Ms ESI +: 535.3[ M + Na ]]⁺And the deuteration rate is 98.4%.

Example 6.14-O- { [ (1S,2S,4S) -4-amino-2-hydroxycyclohexyl-sulfanyl ] acetyl } -2, 2, 4-trideuterometrine

The synthetic route is as follows:

example 614-O- { [ (1S,2S,4S) -4-amino-2-hydroxycyclohexyl-sulfanyl ] acetyl } -2, 2, 4-trideuterometrine

18.8g of intermediate 11 are dissolved in 40ml of isopropanol and stirred at room temperature. Adding 35g of 85% phosphoric acid, heating to 50 ℃, reacting for 18hr, and detecting complete reaction by TLC. Cooled to room temperature and 150ml dichloromethane were added. Cooling the reaction solution to 5-10 ℃ and controlling the temperature, dropwise adding a saturated sodium bicarbonate solution, adjusting the pH to 7-8, extracting the water layer once again by using a dichloromethane layer, combining the dichloromethane layers, washing for 1 time by using water, and washing for one time by using a saturated sodium chloride solution. Concentrate to a viscous state under reduced pressure, add 260ml MTBE, and slurry for 2hr to give the compound of example 16. LC-Ms ESI +: 511.3[ M + H]⁺,533.3[M+Na]⁺The deuteration rate is 98.5%.

Intermediate 8: (R) - (3-Cyclohexenyl) -carbamic acid tert-butyl ester

The procedure for the preparation of intermediate 8 was as described for the preparation of intermediate 2, except that (R) -3-cyclohexenecarboxylic acid was replaced by (S) -3-cyclohexenecarboxylic acid (available from Sahn' S chemical technology, Shanghai, Ltd.).

Intermediate 9: (1S, 3S, 6R) - (7-oxa-bicyclo [4.1.0] hept-3-yl) -carbamic acid tert-butyl ester

The procedure for the preparation of intermediate 9 was as for the preparation of intermediate 3, except that (S) - (3-cyclohexenyl) -carbamic acid tert-butyl ester was replaced with (R) - (3-cyclohexenyl) -carbamic acid tert-butyl ester.

Intermediate 10 { (1S,2S,4S) -4- [ (tert-butoxycarbonyl) -amino ] -2-hydroxy-cyclohexyl-sulfanyl } -acetic acid

The procedure for the preparation of intermediate 10 is as for the preparation of intermediate 4, except that intermediate 3 is replaced by (1S, 3S, 6R) - (7-oxa-bicyclo [4.1.0] hept-3-yl) -carbamic acid tert-butyl ester.

Intermediate 11-14-O- { [ (1S,2S,4S) -4-tert-Butoxycarbonylamino-2-hydroxycyclohexyl-sulfanyl ] acetyl } -2, 2, 4-trideuterome-rine

The procedure for the preparation of intermediate 11 was as for the preparation of intermediate 5 except that intermediate 4 was replaced with { (1S,2S,4S) -4- [ (tert-butoxycarbonyl) -amino ] -2-hydroxy-cyclohexyl-sulfanyl } -acetic acid.

LC-Ms ESI+：656.3[M+2Na]⁺。

Table compounds:

using a similar protocol to that above, the following table compounds and their single enantiomers were prepared.

Table 1: structure of table compound

Evaluation of biological Activity:

1. culture medium and reagent material

MH broth medium, MH agar medium, nutrient broth medium, nutrient agar medium, blood plates, cyclophosphamide, sterile plastic plates, sterile 96-well plates, azithromycin (purchased from shanghai chemical technology, ltd.), linezolid (purchased from shanghai chemical technology, ltd.), levofloxacin (purchased from shanghai chemical technology, ltd.), moxifloxacin (purchased from shanghai chemical technology, ltd.), BC-3781 (homemade).

2. Main instrument

The system comprises a constant-temperature incubator, a biological safety cabinet, a vertical pressure steam sterilizer, a high performance liquid chromatography-quadrupole rod tandem mass spectrometer (including a Japanese Shimadzu high performance liquid chromatography system (LC-20A), an American AB mass spectrometry system (API4000), an electrospray ion source and an Analyst 1.5.1 workstation).

3. Test strains and animals

All strains were purchased from ATCC and CVCC: aureus (staphylococcus aureus); MRSA (methicillin-resistant staphylococcus aureus); pyogens, group a streptococcus; agalactiae, group Bstreptococci; e.faecalium; s. pneumoniae; influenzae; catarralis; pneumoniae.

The test animals are SPF-grade Balb \ c mice purchased from the center of laboratory animals of Yangzhou university.

MIC (minimum inhibitory concentration) determination

Minimum Inhibitory Concentration (MIC) was according to M7-A7[2006 ]]Determined by the gravy dilution method referred to the Clinical and Laboratory Standards Institute (CLSI). The quality control range of microorganisms using different bacteria, and the standard of interpretation of the comparison substances, are shown in CLSI M100-S1[2007 ]]As disclosed in (1). Briefly, serial 2-fold dilutions of the test compound were prepared at 2X concentration in mullerxinton broth (Mueller Hinton Borth). Compound dilutions were mixed at 1:1 ratio ofMixed with bacterial inoculum in 96-well assay plates. The inoculum was prepared from a suspension of colonies in agar plates prepared the previous day. Bacteria were suspended in sterile saline and added to each assay plate to obtain a final concentration of 5X 10⁵CFU/mL. The plates were incubated at 35 ℃ in ambient air for 20 hours. The MIC determined was the lowest concentration of test compound that resulted in no observable bacterial growth when compared to the untreated control group. Each group was repeated 3 times.

5. Animal drug efficacy testing

In vivo infection mouse model

Selecting SPF-grade Balb \ c mice with the weight of 20-25g into groups, feeding 6 female mice in each group for 7 days, and then carrying out the test. 150mg/kg is injected 4 days before the experiment, and 100mg/kg of cyclophosphamide injection is injected 1 day before the experiment. Two bacteria, S.pneumoniae and S.aureus, were treated in 50. mu.L 10⁶The CFU dose was inoculated into the abdominal cavity of mice (TatedaKet. al; AAC1996), 5 mice were inoculated per strain per group, and drug solutions of the corresponding concentration (1.25-320 mg/kg/day in vivo) were injected subcutaneously 120min later and administered twice a day. After 24 hours of treatment, mice were scored for diarrhea (no diarrhea, mild diarrhea, moderate diarrhea + +, dehydration with severe diarrhea + +), then the lung tissue of the mice was aseptically removed, and the number of bacteria at the infected site was determined by agar plate. PRSP (penillilin-resistant s. pneumoniae); MRSA (methicillin-resistant s.

Pharmacokinetic parameter testing

Mice given a single dose of 35-70mg/kg, subcutaneous injection, non-neutropenic mice (no cyclophosphamide injection given), mice were tested for pharmacokinetic parameters 0hr dosing to 5.5hr post dose, data were recorded for the middle 7 points of plasma and alveolar lavage, PK samples were tested using LC-MS/MS. The alveolar lavage epithelial mucus layer drug concentration (ELF) was obtained by calculating the ratio of the urine drug concentration to the immediate plasma concentration. The method of in vitro equilibrium dialysis measures approximately 20% free drug in plasma.

Data analysis

Exposure levels (plasma and baseline) to each isolate using Hill-type model (Hill-type)ELF) and 24h post-treatment response (Δ CFU/lung), compared to the relationship between free drug plasma and ELF exposure. And this relationship was further evaluated by data collected based on s.pneumoniae and s.aureu. By using the above relationship, 1-and 2-log with each pathogen was determined₁₀CFU reduction decreased the associated AUC from baseline: MIC ratio. 1-log after first administration₁₀CFU reduction; 2-log after second administration₁₀Drug lung targeting index ═ AUC (ELF): AUC (plasma free drug).

hERG assay

To assess the risk of prolongation of the QT interval of the electrocardiogram, HEK293 cells expressing the human ether-a-go-go related gene (hERG) channel were used to study the delayed rectifier K which plays an important role in the ventricular repolarization process⁺Electric current (I)_Kr) The function of (1).

Using a full-automatic patch-clamp system, I induced upon administration of depolarizing stimulus of +50mV for 2 seconds, further depolarizing stimulus of-50 mV for 2 seconds after cell maintenance at-80 mV membrane potential was recorded by the full-cell patch-clamp method_Kr. After the generated current was stabilized, an extracellular fluid (NaCl:137mmol/L, KCl:4mmol/L, CaCl) in which the test substance was dissolved at a desired concentration was allowed to flow₂:1.8mmol/L,MgCl₂-6H₂1mmol/L O, 10mmol/L, HEPES:10mmol/L glucose, pH7.4) was applied to the cells at room temperature for 10 minutes. From the obtained I_KrThe absolute value of the maximum tail current was measured using analysis software with reference to the current value of the resting membrane potential. Further, the inhibition ratio with respect to the maximum tail current before application of the test substance was calculated, and the test substance was evaluated for I in comparison with the medium application group (0.1% DMSO solution)_KrThe influence of (c).

7. Results of bioactivity test

The compounds disclosed herein, including the example compounds and the table compounds, all exhibited MIC for their in vitro antibacterial activity₉₀MIC of less than 100. mu.g/ml, preferably of the compound₉₀MIC of less than 50. mu.g/ml, further preferred compounds₉₀Less than 10. mu.g/ml, wherein the biological activities of representative examples are as follows.

Examples representative compounds in vitro activity test results are shown in table 2, and all of the compounds of examples have significant inhibitory effects on gram-positive bacteria (e.g., s.aureus, s.pneumoniae, MRSA, s.pyogenes, group a streptococcus, s.agalactiae, group B streptococcus), gram-negative bacteria (e.g., h.influenzae, m.catarrhalis), atypical pathogenic microorganisms (m.pneumoniae), and the like. Among them, the example compounds 1 and 2MIC in all strains tested₉₀Is more than BC-37811 times. Especially against s.agalactiae, group B streptococcus, example compound 1MIC₉₀0.02. mu.g/ml, Compound 2MIC₉₀Is 0.01 mu g/ml, and the in vitro activity is BC-3781 (MIC)₉₀0.05. mu.g/ml) 2.5 times and 5 times. The test results indicate that the compound of the embodiment has better antibacterial curative effect in clinic than the existing clinical mainstream antibiotics such as azithromycin, linezolid, moxifloxacin and BC-3781. In vitro tests show that 9 strains can cause systemic infectious diseases or infectious diseases of partial organs and are common strains of respiratory tract infection and lung infection, and the compound of the embodiment is expected to be clinically used for treating the systemic infectious diseases and the infectious diseases of the partial organs, and is particularly used for treating upper respiratory tract infection, lower respiratory tract infection and lung infectious diseases.

TABLE 2 in vitro antibacterial Activity test results for the example Compounds

Ratio (plasma free drug AUC: MIC)₉₀) By drug concentration in plasma and MIC₉₀The ratio indicates the ability of inhibiting bacteria in the blood vessel of the drug, and the larger the value, the stronger the in vivo antibacterial ability. Ratio (ELF AUC: MIC) through bloodDrug concentration in plasma and MIC₉₀The ratio indicates the ability of the drug to inhibit bacteria in the blood vessel, and the larger the value, the stronger the in vivo antibacterial ability. Pulmonary targeting index ═ AUC (ELF): AUC (plasma free drug) describes to some extent the drug distribution/blood distribution in the lung, the larger the value, the higher the drug lung targeting.

EXAMPLES in vivo antibacterial Activity test results of representative Compounds in PRSP (penillin-resistant S. pneumoconiae) Lung infection mouse model in vivo antibacterial Activity are shown in Table 3.BC-3781 ratio (plasma free drug AUC: MIC) after first and second dosing₉₀) Less than 1 indicates weak antibacterial ability in blood. After the compound 1 and the compound 2 of the embodiment are administrated for the first time, the ratio is more than 1, and the obvious effect of inhibiting bacteria is achieved. After the second administration, the ratio of example compounds 1 and 2 was 6-fold and 5-fold higher than that of BC-3781, respectively, suggesting that the example compounds had better therapeutic effects and lower doses in clinical applications.

After the second administration, the lung targeting indexes of the example compounds 1 and 2 are 16 and 13.6 respectively, and the reference substance BC-3781 is 9.7, which shows that the respiratory tract and lung targeting of the example compounds 1 and 2 are better, and the clinical treatment effect is better, the dosage is lower and the side effect is lower in the aspect of treating upper respiratory tract infection, lower respiratory tract infection and lung infectious diseases under the same dosage.

TABLE 3 in vivo antibacterial Activity test results of the example Compounds in PRSP Lung infected mouse model

Examples representative compounds the results of in vivo antimicrobial activity testing in a mouse model of MRSA (methicillin-resistant s. aureus) pulmonary infection are shown in table 4. MRSA-infected respiratory and pulmonary diseases are currently clinically very problematic and control approaches are limited. The compound has very obvious effect of resisting MRSA in vivo. As shown in table 4, the example compounds 1 and 2 had 3.6-fold and 4-fold higher ratios, respectively, than BC-3781 after the second administration, suggesting that the example compounds had better therapeutic effects and lower dosages in clinical applications.

After the second administration, the lung targeting indexes of the example compounds 1 and 2 are 15.4 and 14.2 respectively, and the reference substance BC-3781 is 10.7, which shows that the respiratory tract and lung targeting of the example compounds 1 and 2 are better, and the clinical treatment effect is better, the dosage is lower and the side effect is lower in the aspect of treating upper respiratory tract infection, lower respiratory tract infection and lung infectious diseases under the same dosage.

TABLE 4 results of in vivo antibacterial Activity test in MRSA Lung infection mouse model

The diarrhea of the mice with 2 doses of the representative compound in the examples is shown in Table 5, and the diarrhea of the example compound occurs at a low rate, even if mild diarrhea occurs, and severe diarrhea with dehydration is not observed. The control group is administered with BC-3781, mice are found to have diarrhea to different degrees, and the MRSA lung infection mice treatment group even has severe diarrhea with dehydration phenomenon. The phenomenon shows that the compound of the embodiment improves the clinical curative effect of the pleuromutilin antibiotics, overcomes the side effect of diarrhea of the pleuromutilin antibiotics and has bright clinical application prospect.

TABLE 5 diarrhea in mice with pulmonary infection after 2 doses

In order to evaluate the risk of QT interval prolongation of the drug electrocardiogram, the compounds of the examples were subjected to the hERG test, the test results are shown in Table 6. from the results of the inhibition rates in Table 6, the inhibition rates of the compounds 1 and 2 of the examples are significantly less than that of BC-3781, especially the inhibition rates of 1 and 3 are less than one fifth of BC-3781. The test results show that 1 and 2 are less cardiotoxic than the compound BC-3781. In clinical application, the tolerance is higher, the patient has better compliance, and the application range of the patient population is also expanded.

TABLE 6 inhibition of the example compounds at 0.3-10. mu. mol/L

It will be appreciated by those skilled in the art that the present disclosure is not limited to the foregoing illustrative embodiments, but may be embodied in other specific forms without departing from the essential attributes thereof. It is therefore intended that the embodiments described herein are to be considered in all respects as illustrative and not restrictive, the embodiments referenced by the appended claims being other than the foregoing embodiments, the references being made to the appended claims rather than to the foregoing examples, and all changes which come within the meaning and range of equivalency of the claims are therefore intended to be embraced therein.

All patents, patent applications, and literature references cited in this specification are hereby incorporated by reference in their entirety. In the event of inconsistencies, the present disclosure, including definitions, will be convincing.

Claims

1. The present invention provides a compound of formula I, or a stereoisomer, prodrug, active metabolite, or pharmaceutically acceptable salt, solvate, or crystalline form thereof:

wherein,

R¹is hydroxy, hydrogen or deuterium;

R²is hydroxy, hydrogen, deuterium or fluorine;

R^2‘is hydroxy, hydrogen, deuterium or fluorine;

R³is hydroxy, hydrogen, deuterium or fluorine;

R^3’is hydroxy, hydrogen, deuterium or fluorine;

m is hydrogen or deuterium;

a is methylene, oxygen, -NH-, sulfur, sulfoxide, or sulfone;

l is

When L is

In which

n is 0 to 4;

m is 0 or 1;

R⁵is hydrogen or (C)_1-6) An alkyl group;

R⁵Is hydroxy and R⁶Is a formyl group;

R⁷is OH, OR⁸A halogen atom, or

2. The present invention provides a compound of formula II, or a stereoisomer, prodrug, active metabolite, or pharmaceutically acceptable salt, solvate, or crystalline form thereof:

wherein,

n is 0 to 4;

R¹is hydroxy, hydrogen or deuterium;

R²is hydroxy, hydrogen, deuterium or fluorine;

R^2‘is hydroxy, hydrogen, deuterium or fluorine;

R³is hydroxy, hydrogen, deuterium or fluorine;

R^3’is hydroxy, hydrogen, deuterium or fluorine;

m is hydrogen or deuterium;

R⁵is hydrogen or (C)_1-6) An alkyl group;

R⁵Is hydroxy and R⁶Is formyl.

3. The present invention provides a compound of formula III, or a stereoisomer, prodrug, active metabolite, or pharmaceutically acceptable salt, solvate, or crystalline form thereof:

wherein,

n is 0 to 4;

R¹is hydroxy, hydrogen or deuterium;

R²is hydroxy, hydrogen, deuterium or fluorine;

R^2‘is hydroxy, hydrogen, deuterium or fluorine;

R³is hydroxy, hydrogen, deuterium or fluorine;

R^3’is hydroxy, hydrogen, deuterium or fluorine;

m is hydrogen or deuterium.

4. The present invention provides a compound of formula IV, or a stereoisomer, prodrug, active metabolite, or pharmaceutically acceptable salt, solvate, or crystalline form thereof:

wherein,

n is 0 to 4;

R¹is hydroxy, hydrogen or deuterium;

R²is hydroxy, hydrogen, deuterium or fluorine;

R^2‘is hydroxy, hydrogen, deuterium or fluorine;

R³is hydroxy, hydrogen, deuterium or fluorine;

R^3’is hydroxy, hydrogen, deuterium or fluorine;

R⁴is ethyl, 1, 2-dideuteroethyl, vinyl, 2, 2-dideuterovinyl.

5. The compound as provided in claims 1 to 4, or a stereoisomer, prodrug, active metabolite, or pharmaceutically acceptable salt, solvate, or crystalline form thereof, wherein the compound is represented by any one of the following structural formulae:

6. a pharmaceutical composition comprising a therapeutically effective amount of a compound of any one of claims 1 to 5 in unit dosage form and one or more pharmaceutically acceptable carriers.

7. A pharmaceutical combination composition comprising a therapeutically effective amount of a compound according to any one of claims 1 to 5 and another agent having therapeutic activity against a microbial-mediated disease.

8. Use of a compound of any one of claims 1 to 5, an N-oxide thereof or a prodrug thereof, for the manufacture of a medicament for the treatment of a condition mediated by a microorganism.

9. The use of claim 8, wherein the microorganism-mediated disorder is selected from the group consisting of systemic infectious diseases, ocular infectious diseases, upper respiratory infectious diseases, lower respiratory infectious diseases, skin and soft tissue infectious diseases, acute sinusitis, chronic bronchitis, community-acquired pneumonia, and hospital-acquired pneumonia.