KR101203118B1 - 18-membered macrocycles and analogs thereof - Google Patents

Abstract

The present invention relates to an 18-atomic macrocyclic antimicrobial agent called Tiacumicin, specifically OPT-80 (most of which consists of R-Tiacumicin B), pharmaceutical compositions comprising OPT-80, and OPT-80 It relates to a method using. In particular, the compound is a potential pharmaceutical to treat bacterial infections, especially C. difficile (difficile) infection.

Macrocycles, tiacumicins, optical isomers, bacteria.

Description

18-atomic macrocycles and their analogs {18-MEMBERED MACROCYCLES AND ANALOGS THEREOF}

The present invention relates to an 18-atom macrocyclic antimicrobial agent called Tiacumicin, in particular R-Tiacumicin B or Tiacumicin B and related compositions. More specifically, bacterial infections, specifically, Clostridium difficile ( C. difficile ), methicillin resistant Staphylococcus aureus (MRSA), including Staphylococcus aureus S. aureus and Clostridium perprisence perfringens : C. perfringens ) is substantially pure R-thiacumycin B as a potential antibiotic for the treatment of GI infections caused by toxin-induced varieties of C. perfringens ).

Giant rings are an important therapeutic class of antibiotics. Such compositions are often produced as a family of closely related biogenic analogs. Tiacumicins are a family of 18-atomic macrocyclic antibiotics in which the macrocycles are glycosidically linked to one or two sugars. 7-carbon sugars are esterified with small fatty acids at various positions. All other sugars, when present, are esterified with the isomers of substituted benzoic acid, everninic acid (Journal of Liquid Chromatography, 1988, 11: 191-201).

Tiacumicins are a family associated with compounds containing the 18-membered ring shown in the following formula (I).

Several unique tiacumicins have now been identified, of which six (tiacumicin AF) are characteristic patterns of their substituents R ¹ , R ² and R ³ as shown in Table 1 (US Pat. No. 4,918, ® J). Antibiotics, 1987, 40: 575-588).

[Table 1]

Tiacumicin AF has been characterized spectroscopically and by other physical methods. The chemical structure of tiacumicins is based on spectroscopy: UV-visible light, IR and NMR of ¹ H and ¹³ C, for example see J. Antibiotics, 1987, 40: 575-588. It can be seen that certain members within the observation series of Table 1 are structurally related isomers and / or distinguished by the presence or absence of certain moieties. Others differ in the properties of their ester groups.

Tiacumicins are known as Dactylosporangium ( Dactylosporangium). Produced by bacteria, including aurantiacum subspecies hamdenensis , which can be obtained from the ARS patent collection bureau of the US Department of Agriculture's Northern Research Center (1815 North University Street, Peoria, IL 61604) under NRRL 18085. The characteristics of the varieties AB 718C-41 are shown in J. Antibiotics, 1987, 40: 575-588 and US Pat. No. 4,918,174.

C. Clostridium difficile- associated diarrhea (CDAD) is a disease characterized by severe and painful diarrhea. C. difficile is involved in approximately 20% of antibiotic-associated diarrhea (AAD) and the majority of antibiotic-associated colitis (AAC). These diseases are Clostridium difficile: Ste Philo nose kusu aureus (Staphylococcus aureus), including the (Clostridium difficile C. difficile), methicillin-resistant pillow Ste nose kusu aureus (MRSA, Methicillin-resistant Staphylococcus aureus ) And toxin-inducing varieties of Clostridium perfringens ( C. perfringens ). The AAD estimates annual hospital costs of roughly $ 3–6 billion, in the United States alone, making it a major economic burden on the health care system.

Providing a continuous source of intestinal colonization is infection, vancomycin-resistant enterococci (VRE, Vacomycin-resistant Enterococci) was also a major pathogenic agents associated with increased healthcare costs and mortality. VRE can appear as a complication in patients infected with C. difficile, or more commonly in certain high-risk patients, such as hematologic and neoplastic patients, patients in intensive care units, and transplant patients. .

Methicillin-resistant Staphylococcus aureus (Staphylococci), such as MRSA are prevalence (prevalence) increases in both hospital and community settings. Staphylococcus is found in the skin and in the digestive and respiratory tracts but can infect open wounds and burns and develop into serious systemic infections. In particular, the occurrence of various drug-resistant staphylococci in hospitals where the use of antibiotics and high selective pressure on drug-resistant organisms has been found to be an obstacle in treating these patients. The presence of MRSA in the skin of patients and healthcare workers promotes the transmission of various drug resistant organisms.

Although not limited to these, similar diseases including Clostridium enterocolitis, neonatal diarrhea, antibiotic-associated enterocolitis, sporadic enterocolitis, and in-hospital enterocolitis have become important problems in some animal species.

AAD is a serious problem in community hospitals and long-term care homes. C. difficile is the leading cause of AAD in hospitals, approximately 20% of AAD and the major cause of antibiotic-associated colitis (AAC). Increasing incidence of Clostridium difficile- related diarrhea (CDAD) is seen as a result of frequent prescription of a wide range of antibiotics for inpatients.

The most severe form of the disease is pseudomembranous colitis (PMC), which is manifested histologically by colitis of the mucosa and clinically by severe diarrhea, abdominal cramps, and systemic toxicity. Although overall mortality from CDAD is low, it is very high in patients who develop severe colitis or systemic toxicity. Recent studies show that even when mortality is not directly attributable to C. difficile, mortality rates in CDAD patients are much higher than in comparable controls.

Diarrhea and colitis are caused by the assimilation of one or more C. difficile toxins. The organism proliferates in the colon of patients with extensive antibiotics or, less commonly, cancer chemotherapy. CDAD is diagnosed in approximately 20% of inpatients who develop diarrhea after being treated with this agent.

There are currently two major treatments for CDAD: vancomycin and metronidazole. Vancomycin is not recommended as a priority treatment for CDAD because it is primarily an antibiotic that acts on some seriously life-threatening, multi-drug resistant bacteria. Therefore, in an effort to minimize the appearance of vancomycin-resistant enterococci (VRE) or vancomycin-resistant staphylococcus (S. aureus) (VRSA), the medical community restrains the use of these medications except when desperately needed. .

Metronidazole is recommended as an initial treatment without worrying about the promotion and selection of vancomycin-resistant enteric fungi, especially enterococci. Despite reports that the frequency of C. difficile resistance may be> 6% in some countries, metronidazole, which is considered almost as effective as vancomycin, is much cheaper and can be used orally or intestinally. Metronidazole is associated with serious side effects including nausea, neuropathy, leukopenia, seizures, and toxic reactions to alcohol. Moreover, it is not safe to use for infants or pregnant women. Clinical relapse occurs in up to 20% of cases after treatment with vancomycin or metronidazole. Metronidazole therapy has been reported as an important risk factor for VRE colonization and infection. Recent treatment policies for gastrointestinal infections, such as Clostridium difficile related diarrhea (CDAD), are rather burdensome requiring up to 500 mg four times a day for 10 to 14 days. Therefore, better treatment is needed for CDAD cases as well as other antibiotic-associated diarrheal diseases (AAD) and antibiotic-related colitis (AAC).

Tiacumicins, especially Tiacumicin B, are active against a variety of bacterial pathogens, particularly C. difficile, Gram-positive bacterium (Antimicrob. Agents Chemother. 1991, 1108-1111). C. difficile is an anaerobic spore forming bacterium that causes intestinal infections. Diarrhea may be the most common symptom, but abdominal pain and fever may develop. C. difficile is a major cause of colitis (inflammation of the colon) and diarrhea that can occur following antibiotic intake. The fungus is mainly obtained in hospitals and long-term care facilities. Since tiacumicin B exhibits promising activity against C. difficile, it is expected to be effective in the treatment of bacterial infections in mammals, particularly in the case of the gastrointestinal tract. Examples of such treatments include, but are not limited to, treatment of colitis and irritable bowel syndrome. Tiacumicin can also be used for the treatment of gastrointestinal cancer.

Tiacumicin antibiotics are described in US Pat. No. 4,918,174 (published Apr. 17, 1990), J. Antibiotics 1987, 40: 575-588, J. Antibiotics 1987, 40: 567-574, J. Liquid Chromatography 1988, 11: 191. -201, antimicrobials and chemotherapy 1991, 35: 1108-1111, U.S. Patent No. 5,583,115 (published Dec. 10, 1996), and U.S. Patent No. 5,767,096 (published June 16, 1998). Incorporated herein by reference. Related compounds include Lipiamycin antibiotics (cf, Chem. Soc. Perkin Trans. I, 1987, 1353-1359 and J. Antibiotics 1988, 41: 308-315) and Closomicin antibiotics (J. Antibiotics). 1986, 39: 1407-1412, which are incorporated herein by reference.

The present invention relates to novel pharmaceutical compositions containing R-tiacumicins, in particular optically pure R-tiacumicin B, and to the novel compositions for treating infections caused by Gram-positive anaerobes. It is about using a combination of existing medicines.

One embodiment of the present invention is based on the discovery that the chiral center of C-19 of tiacumicin B has a great impact on biological activity. The preparation of substantially pure, highly active R-tiacumicin B having an R-hydroxy group in C-19 is significantly lower than the optically pure S-isomer of tiacumicin B and other thiacumicin B related compounds. It has a MIC value.

In another embodiment of the present invention, substantially pure R-tiacumicin B has exceptional long-term post antibiotic activity (PAE).

Dactylosporangium microorganisms Dactylosporangium aurantithiacum subspecies Hamdenensis aurantiacum subspecies hamdenensis ) and include novel antibiotics containing substantially pure R- tyacumicins by aerobic immersion flotation. The recipe is supported by WO 2004/014295 A2, incorporated herein by reference.

The term "antibiotic-related symptoms" occurs when antibiotic therapy interferes with the balance of microbial flora in the stomach, causing pathogens such as C. difficile, S. aureus and C. perprisence to thrive. Refers to the symptoms that become. These organisms can cause diarrhea, false membrane colitis, and colitis and are identified by abnormal conditions between diarrhea, urgency, abdominal cramps, tenesmus, and other symptoms. In severe cases, diarrhea causes dehydration and medical complications associated with dehydration.

The term "asymmetrically substituted" refers to a molecular structure with atoms having four tetrahedral balances in which four different atoms or groups are bonded. The most common case involves carbon atoms. In this case, two optical isomers (D- and L-enantiomers or R- and S-enantiomers) do not overlap each mirror image per carbon. Many compounds have one or more asymmetric carbons. This raises the possibility of many optical isomers whose number is determined by equation 2 ⁿ where n is the number of asymmetric carbons.

As used herein, the term "culture liquid" refers to a fluid culture medium obtained during or after fermentation. Cultures comprise a mixture of water, certain antibiotic (s), unused nutrients, live or dead organisms, metabolic products, and absorbents with or without absorbed output.

The term "C-19 ketone" refers to Tiacumacin B associated with the compound shown in Formula II:

The term "diastereomer" refers to the structural isomer of each non-enantiomer.

The term "enantiomer" refers to one's own non-overlapping mirror image. The enantiomer of the optically active isomer rotates the planar polarization by the same in the opposite direction of the original isomer. Solutions containing the same amount of optically active isomers and their enantiomers are known as racemic solutions and have a total rotation of polarized light on a plane of zero. Enantiomers each have the opposite prefix: D- becomes L- or R- becomes S-. Often in the biological world only one enantiomer is active because most biological reactions are enzymatic and the enzymes are bound to only one enantiomer.

The term "excipient" refers to an inert substance added to the pharmacological compound to make the compound easier to take. Examples of excipients include, but are not limited to, calcium carbonate, calcium phosphate, various sugars and starch forms, cellulose derivatives, gelatin, vegetable oils and polyethylene glycols.

The term "halogen" includes F, Cl, Br and I.

The term “isomer mixture” refers to a mixture of two or more species having the same chemical formula, with specific spatial arrangements. An isomeric mixture is a genus containing individual isomeric species. Examples of isomeric mixtures include, but are not limited to, stereoisomers (enantiomers and diastereomers), regioisomers, which can be produced, for example, from cyclic co-reactions. Compounds of the present invention include asymmetrically substituted carbon atoms. Such asymmetrically substituted carbon atoms can give rise to single stereoisomers, or mixtures of stereoisomers, especially at asymmetrically substituted carbon atoms. As a result, the racemic mixtures of the present invention, mixtures of diastereomers as well as single diastereomers are included in the present invention.

The term "Lipiarmycin A4" refers to Tiacumicin B associated with the following structural formula III.

The term "lower alkyl" may be used alone or in combination with one to eight carbons (eg, C ₁ , C ₂ , C ₃ , C ₄ , C ₅ , C ₆ , C ₇ and C ₈ ), preferably Refers to an optionally substituted straight chain or optionally substituted branched chain having 1 to 4 carbons (eg, C ₁ , C ₂ , C ₃ and C ₄ ). Examples of alkyl radicals include methyl, ethyl, n-propyl, isopropyl, n-butyl, isobutyl, sec-butyl, tert-butyl. "Lower alkyl" is generally one containing short alkyl, eg, 1 to 4 carbon atoms (eg, C ₁ , C ₂ , C ₃ and C ₄ ).

The term " macrocycle " refers to organic molecules having a macrocyclic structure usually containing at least 10 atoms.

The term "18-atom macrocyclic ring" refers to organic molecules having a ring structure containing 18 atoms.

The term "-atomic ring" may encompass any ring structure including cyclic carbons and heterocycles as described above. The term "-atom" is meant to symbolize the number of skeletal atoms that make up the ring. Thus, for example, pyridine, pyran and thiopyran are 6-membered rings and pyrrole, furan, and thiophene are 5-atomic rings.

The term "MIC" or "minimum inhibitory concentration" refers to the lowest concentration of antibiotic needed to inhibit the growth of bacteria in vitro. The most common method of measuring the MIC of an antibiotic is to prepare several tubes containing a series of dilutions of the antibiotic and inject it into the isolate of the bacteria of interest. The MIC of the antibiotic is determined from the lowest concentration tube showing no turbidity (no growth).

The term “MIC ₅₀ ” refers to the lowest concentration of antibiotic required to inhibit the growth of 50% of the bacterial strains tested in a given bacterial species.

The term "MIC ₉₀ " refers to the lowest concentration of antibiotic required to inhibit the growth of 90% of the bacterial strains tested in a given bacterial species.

The term “OPT-80” refers to approximately 70-100%, preferably 90% (optical) of optically pure R-tiacumicin B (having an R-hydroxy group at position C-19, see Formula IV). For antibiotic substances, by means of HPLC analysis). The remaining portion consists essentially of small amounts of thiacumacin B related compounds, including but not limited to ripamycin A4 and C-19 ketones. Formulations of this type are described in detail in PCT application PCT / US03 / 21977, International Publication No. WO 2004/014295 A2, which documents and formulations are incorporated herein by reference. However, as an exclusive use for non-humans, a natural “OPT-80” containing less than 70% (by HPLC analysis, for total antibiotic material) may be used which is optically pure.

The term "ORTEP" refers to Oak Ridge's Thermal Ellipsoid Plot computer program written in Fortran to create crystal structure figures. Ball-and-stick shaped figures suitable for printing are fabricated into spherical or thermodynamic probability ellipsoids and derived from anisotropic temperature factor parameters on the atomic site. The program also provides three-dimensional pairs of shapes to help visualize the complex arrangement of atoms and their associated thermal motion patterns.

The term "PAE" or "post-antibiotic effect" refers to well-established pharmacological parameters that reflect the continued suppression of bacterial growth following antibiotic exposure.

The term "patient" refers to a human or animal requiring medical practice. For the purposes of the present invention, human patients are those typically accommodated in primary care facilities such as hospitals or nursing homes. However, treatment of diseases associated with antibiotic use or cancer chemotherapy or antimicrobial treatment may not be associated with the primary clinic and may be done at an outpatient basis away from the primary clinic or may be prescribed by a physician for outpatient treatment. Animals requiring medical care are usually treated by a veterinarian.

The term "pharmaceutically acceptable carrier" refers to a pharmaceutically acceptable carrier or diluent.

The term "pharmaceutically acceptable salts" refers to those derived from pharmaceutically acceptable inorganic and organic bases. Salts derived from suitable bases include alkali metals (eg sodium or potassium), alkaline earth metals (eg magnesium), ammonium and N (C ₁ -C ₄ alkyl) ₄ ⁺ salts, and the like. Include. Some clear examples of these include sodium hydroxide, potassium hydroxide, choline hydroxide, sodium carbonate, and the like.

The term "pharmaceutical composition" refers to the mixing of one or more tiacumicins, or physically acceptable salts thereof, described herein with other chemical components such as physically acceptable carriers and / or excipients.

The term "physically acceptable carriers" refers to carriers or diluents that do not cause serious irritation to the organs and do not inhibit the biological activity and properties of the administered compound.

The term “false colitis” or “colitis” refers to the formation of false membrane material (ie, a substance consisting of fibrin, mucus, necrotic epithelial cells and leukocytes) due to inflammation of the mucosa of both the small and large intestine.

The terms "R" and "S" arrays as used herein refer to Section E, IUPAC's 1974 Recommendations for Basic Stereochemistry ( Pure). Appl . Chem . (1976) 45, 13-30 ). Chiral molecules may be named based on the atoms or groups of atoms attached to the chiral center, and the atomic number of the ligands. Ligands are given a rank (the higher the atomic number, the higher the rank) and is called R- if the rank increases clockwise. Conversely, if the rank increases counterclockwise, it is called S-.

The term "R-thiacumycin B" refers to an optically pure (R) -isomer having an (R) -hydroxy group at the C-19 position, as shown in the following formula IV:

The term " S-thiacumycin B " refers to an optically pure (S) -isomer having a (S) -hydroxy group at the C-19 position, as shown in Formula V below:

The term “stereoisomers” refers to compounds having the same number and type of atoms and the same atomic arrangement but with different spatial arrangements.

The term "sugar" generally refers to mono-, di- or oligosaccharides. Sugars include, for example, glucosamine, galactosamine, acetylglucose, acetylgalactose, N-acetylglucosamine, N-acetyl-galactosamine, galactosyl-N-acetylglucosamine, N-acetylneuraminic acid (salicylic acid), and the like. It can be substituted as well as sulfided and phosphorylated sugars. For the purposes of the present invention, the sugars are in the form of pyranose or furanose.

As used herein, the term "tiacumicin" refers to the collection of all compounds, including the 18-atom macrocycle shown in Formula I:

As used herein, the term “tiacumicin B” refers to the 18-atomic macrocycle shown in the following structure VI.

The term "yield" as used herein refers to the amount of raw tiacumicin reproduced in methanol for the same volume as the initial fermentation broth. Yields are determined using standard HPLC techniques. Yield is in mg / L.

Dactylosporangium microorganisms Dactylosporangium aurantithiacum subspecies Hamdenensis Tiacumicin, a composition of novel antibiotic agents due to aerobic immersion of aurantiacum subspecies hamdenensis ). This production method is supported by WO 2004/014295 A2.

The present invention provides a novel antimicrobial composition comprising R-thiacumycin, in particular R-thiacumycin B (having R-hydroxy at position C-19), and to treat infections caused by gram positive anaerobes. It relates to the use of the new compositions in combination with existing medicaments.

The present invention also relates to a novel OPT-80 formulation containing 70-100%, preferably 90% (by HPLC analysis, for antibiotic substance premise). The remaining portion consists essentially of small amounts of tiacumacin B related compounds, including but not limited to ripamycin A4 and C-19 ketones. Formulations of this type are described in detail in PCT application PCT / US03 / 21977, International Publication No. WO 2004/014295 A2. However, as an exclusive use for non-humans, natural OPT-80 may be used which contains less than 70% (by HPLC analysis) for optically pure R-tiacumicin B (for total antibiotic material).

According to the present invention there is provided a compound having the structure of formula VII:

here:

X is selected from lower alkyl and the term "lower alkyl" as used herein is a branched or straight chain comprising one or two carbon atoms, including methyl, ethyl, n-propyl, isopropyl, and the like. Refers to an alkyl group;

Y is selected from OH or ketone (= O);

Z is selected from H or lower alkyl, and the term "lower alkyl" as used herein refers to one to five carbon atoms including methyl, ethyl, propyl, isopropyl, n-butyl, t-butyl, and the like. Refers to a branched or straight chain alkyl group comprising.

Preferred compounds of the invention are compounds of formula VII, wherein X is methyl or ethyl, Y is ketone (= O) or OH and Z is isopropyl.

More preferred compounds of the present invention are compounds of formula VII, wherein X is ethyl, Y is ketone (= O) or OH and Z is isopropyl.

Most preferred compounds of the present invention are compounds of formula VII, wherein X is ethyl, Y is OHR and Z is isopropyl.

One embodiment of the present invention is due to the discovery that the chiral center of C-19 of Tiacumicin B has a great impact on biological activity. R-Tiacumicin B with an R-hydroxy group at the C-19 position has significantly higher activity than S-Tiacumicin B and other Tiacumicin B related compounds (ripiamycin A4 and C-19 ketones) Was found. High activity appears at by far lower MIC, which C. difficile, S. aureus, E. faecalis par (faecalis), and then carried out for several strains of E. par when Titanium (faecium) Example 3, Table It can be seen from 3 and 4. The effect of these C-19 chiral centers on biological activity is unexpected and new findings.

In another embodiment of the present invention, OPT-80 (mostly consisting of R-tiacumicin B) has exceptional long-term post antibiotic activity (PAE). This will be discussed in Example 4 below showing that the OPT-80 has a PAE of 24 hours or more. These PAEs are unexpectedly longer than 1-5 hours, a typical antibiotic PAE.

The invention also relates to the description of a pharmaceutical composition comprising a compound of the invention in combination with a pharmaceutically acceptable carrier.

Another aspect of the invention discloses a method of inhibiting or treating a bacterial infection in a human comprising administering to a patient a therapeutically effective amount of a compound of the invention alone or in combination with other antimicrobial or antifungal agents. have.

Produce

18-atom macrocycles and their analogues are prepared by fermentation. Dactylosporangium Dactylosporangium for the preparation of tiacumicins aurantiacum subspecies hamdenensis ) AB 718C-41 NRRL 18085 is incubated with appropriate aeration conditions and in a sterile environment with one or more absorbents in a medium containing a carbon source, inorganic salts and other organic additives.

Microorganisms for producing active antimicrobial agents have been identified as belonging to the Actinoplanaceae and Dactylosporangium species (Journal of Antibiotics, 1987, 40: 567-574 and US Patent 4,918,174). . Dactylosporanthium auranticumum subspecies Hamdenensis 718C-41. A subculture was obtained from the ARS Patent Collection Bureau of the US Department of Agriculture's Northern Research Center (1815 North University Street, Peoria, IL. 61604, USA), with the filing number NRRL 18085. The characteristics of the AB 718C-41 strain are given in Journal of Antibiotics, 1987, 40: 567-574 and US Pat. No. 4,918,174.

Dactylosporangium microorganisms Dactylosporangium aurantithiacum subspecies Hamdenensis Tiacumicin, a composition of novel antibiotic agents due to aerobic immersion of aurantiacum subspecies hamdenensis ). This production method is supported by WO 2004/014295 A2.

Pharmaceutical Formulations and Administration

The pharmaceutical compositions of the tiacumicin compounds of the invention, in particular OPT-80 (mostly consisting of R-tiacumicin B), can be administered either substantially immediately after or at any predetermined time or time after administration of the antibiotic according to the invention. Can be prepared for post-release.

The latter form of the composition is generally known as a controlled release formulation, which produces a substantially constant concentration of medicament in the gastrointestinal tract for an extended period of time, and M.J. Rathbone, J. Hodgraft and M.S. Agents that have controlled release characteristics based on time or environmental criteria, as described in Roberts' "Controlled Release Pharmaceutical Dosing Technique" (Marcel Dekker, Inc.).

Any biologically acceptable oral dosage form, or combination thereof, may be employed in the methods of the present invention. Examples of such dosage forms include chewable tablets, rapid eluting tablets, bubble tablets, processable powders, elixirs, liquids, suppositories, creams, solutions, suspensions, emulsions, tablets, multilayer tablets, multilayer tablets, capsules, Light gelatin capsules, hard gelatin capsules, osmotic tablets, osmotic capsules, caplets, lozenge, chewable rosin, beads, powders, granules, particles, microparticles, dispersible particles, ingestibles , Juices, healthy bars, candy, animal feed, cereals, cereal coatings, foods, nutritional foods, functional foods, and combinations thereof. The preparation of any of these dosage forms is well known to those of ordinary skill in the art. In addition, pharmaceutical formulations may be designed to provide either immediate or controlled release of antibiotics, depending on the desired site. The choice of immediate or controlled release composition depends on the species and antibiotic sensitivity of the Gram-positive bacteria to be treated and the bacteriostatic / sterilizing properties of the therapeutic agent. Well known methods for making formulations are described, for example, in Remington: The Science and Practice of Pharmacy (20th ed.), Ed. (A.R. Gennaro, 2000, Lippincott Williams & Wilkins, Philadelphia), or Encyclopedia of Pharmaceutical Technology, eds. (J. Swarbrick and J.C. Boylan, 1988-1999, Marcel Dekker, New York).

Immediate release formulations for oral use include tablets or capsules containing the active ingredients mixed with pharmaceutically acceptable nontoxic excipients. Such excipients are for example inert diluents or fillers (e.g., sucrose, sorbitol, sugars, mannitol, microcrystalline cellulose, starch containing potato starch, calcium carbonate, sodium chloride, lactose, calcium phosphate, calcium sulfate or phosphoric acid). salt); Granulating and disintegrating agents (for example cellulose derivatives comprising microcrystalline cellulose, starch comprising potato starch, croscarmellose sodium, alginate or alginic acid); Binders (e.g. sucrose, glucose, mannitol, sorbitol, acacia, alginic acid, sodium alginate, gelatin, starch, gelatinized starch, microcrystalline cellulose, magnesium aluminum silicate, carboxymethylcellulose sodium, methylcellulose, hydroxypropyl methyl Cellulose, ethylcellulose, polyvinylpyrrolidone, or polyethylene glycol); And lubricants, glidants, and antiadhesives (eg, magnesium stearate, zinc stearate, stearic acid, silica, hydrogenated vegetable oils, or talc). Other pharmaceutically acceptable excipients include colorants, sweeteners, plasticizers, wetting agents, buffers, and the like, for example, in The Handbook of Pharmaceutical Excipients, published by Arthur H. Kibbe of the American Pharmaceutical Society of Washington, DC. ) ".

Dissolution or diffusion of the controlled release can be accomplished by appropriate coating of tablets, capsules, pills, or granular compositions of the compound, or infusion of the compound into the appropriate matrix. The controlled release coating may be one or more of the coating materials mentioned above and / or for example shellac, beeswax, glycowax, castor wax, carnauba wax, stearyl alcohol, glyceryl monostea Latex, glyceryl distearate, glycerol palmitostearate, ethylcellulose, acrylic resin, dl-polylactic acid, cellulose acetate butyrate, polyvinyl chloride, polyvinyl acetate, vinyl pyrrolidone, polyethylene, polymethacrylate, methyl Methacrylate, 2-hydroxymethacrylate, methacrylate hydrogel, 1,3 butylene glycol, ethylene glycol methacrylate, and / or polyethylene glycol. In controlled release matrix formulations, the matrix material is for example hydrated methylcellulose, carnauba wax and stearyl alcohol, carbopol 934, silicone, glyceryl tristearate, methyl acrylate-methyl methacrylate, Polyvinyl chloride, polyethylene, and / or halogenated carbon fluoride.

The controlled release composition may also be a buoyant tablet or capsule (ie, a tablet or capsule that floats on the stomach contents for a period of time upon oral administration). Suspended tablet formulations of the compound may be prepared by grinding the antibiotic mixture with excipients and 20-75% w / w of hydrocolloids such as hydroxyethylcellulose, hydroxypropylcellulose, or hydroxypropylmethylcellulose. The granules obtained can then be compressed into tablets. In contact with gastric juices, the tablets form a gel protective film that is substantially impermeable to water on its surface. This gel protective film serves to maintain a density of 1 or less, thereby allowing the tablet to remain suspended in gastric juice. Other useful controlled release compositions are known in the art (see, eg, US Pat. Nos. 4,946,685 and 6,261,601).

The controlled release composition may consist of a compression coated core that regulates the release distribution of the antibiotic in which the geometry is embedded. By varying the geometry of the core, the distribution of antibiotic release can be adjusted to follow the zero, first, or combination order thereof. The system can also be designed to administer more advantageous formulations having different release distributions each at the same time (see, eg, US Pat. Nos. 4,111,202 and 3,279,995).

Agents targeting the tiacumicin compound of the present invention, which release in specific regions of the gastrointestinal tract, in particular OPT-80, which almost entirely consists of R- tiacumicin B, may also be prepared. The thiacumicin compounds of the present invention, in particular OPT-80, can be encapsulated in an intestinal coating to prevent gastric release attenuation and release, but dissolve easily in a weakly acidic or neutral pH environment of the small intestine. Formulations that are targeted to release antibiotics in the colon may also be used utilizing techniques such as time dependence, pH dependence, or enzymatic corrosion of polymer matrices or coatings.

The target dosage properties of the tiacumicin compounds of the invention, in particular OPT-80, comprising almost all of R-tiacumicin B, including the agent, can be adjusted by other means. For example, antibiotics can be complexed by inclusion, ionic interactions, hydrogen bonds, hydrophobic bonds, or covalent bonds. Polymers or complexes that are sensitive to enzymatic or microbial lysis may also be used as pharmaceutical administration means.

Microsphere encapsulation of the tiacumicin compounds of the invention, in particular OPT-80, almost entirely consisting of R-tiacumicin B, is another useful pharmaceutical agent for targeted antibiotic release. Antibiotic-containing microspheres may be used alone for antibiotic administration or as a component of a two stage release formulation. Suitable staged release formulations are compounds of the invention, in particular OPT-80 (almost entirely composed of R-thiacumycin B) such that antibiotics are mixed with immediate release formulations for delivery to the stomach and upper duodenum and later released in the lower gastrointestinal tract. By encapsulating it can be made of microspheres stable to the acid.

Microspheres can be made by any suitable method or from any pharmaceutically acceptable material. Particularly useful are protein analog microspheres (see, eg, US Pat. Nos. 5,601,846 or 5,792,451) and PLGA-containing microspheres (see, eg, US Pat. Nos. 6,235,224 or 5,672,659). Other polymers commonly used to form microspheres are, for example, poly-ε-caprolactone, poly (e-caprolactone-Co-DL-lactic acid), poly (DL-lactic acid), poly (DL-lactic acid- Co-glycolic acid) and poly (s-caprolactone-Co-glycolic acid) (see, eg, J. Pharm. Sci., 68: 1534, 1979 to Pitt et al.). Microspheres can be made by methods well known in the art, including spray drying, coacervation, and emulsification (eg, Microsphere and Drug Therapy, Davis et al., 1984, Elsevier; Benoit et al. Biodegradable Microspheres: Advances in Production Technologies, Chapter 3, ed.Benita, S, 1996, Dekker, New York; see US Pat. No. 6,365,187).

Powders, dispersible powders, or granules suitable for the preparation of aqueous solutions or suspensions of the thiacumicin compounds of the invention, in particular OPT-80 (mostly consisting of R-tiacumicin B), can be administered orally by addition of water. It is a convenient dosage form. Formulations, such as suspensions, provide the active ingredients in admixture with a dispersing or wetting agent, a suspending agent, and one or more preservatives. Suitable dispersing or wetting agents are, for example, naturally occurring phospholipids (eg lectinine or condensates of ethylene oxide with fatty acids, long chain aliphatic alcohols, or partial esters derived from fatty acids) and hexitol or Hexitol anhydrides (eg, polyoxyethylene stearate, polyoxyethylene sorbitol monooleate, polyoxyethylene sorbitan monooleate and analogs). Suitable suspending agents are for example sodium carboxymethylcellulose, methylcellulose, sodium alginate and analogs.

Figure 1 shows the chemical structure of the Thermal Ellipsoid Plot Program (ORTEP) of Oak Ridge of R-Tiacumicin B.

The following examples are provided by a method depicting a characteristic implementation of the invention without intending to limit in any way the scope of the invention.

Example  1: R- Tiacumicin  Precise structure of B

The exact structure of R-tiacumicin B (mainly the most active component of OPT-80) is shown in the following structural formula IV. The X-ray crystal structure of R-tiacumicin B was obtained from colorless parallel pipe-shaped crystals (0.08 x 0.14 x 0.22 mm) grown in methanol and shown as the ORTEP diagram of FIG. This X-ray structure confirms the structure shown in the following structural formula IV. The official chemical name is 3-[[[6-deoxy-4-O- (3,5-dichloro-2-ethyl-4,6-dihydroxybenzoyl) -2-O-methyl-β-D-mannopira Nosyl] oxy] -methyl] -12 (R)-[[6-deoxy-5-C-methyl-4-O- (2-methyl-1-oxopropyl) -β-D-rixo-hexopyrano Sil] oxy] -11 (S) -ethyl-8 (S) -hydroxy-18 (S)-(1 (R) -hydroxyethyl) -9,13,15-trimethyloxacyclooctadeca-3, 5,9,13,15-pentaen-2-one.

Example  2: OPT -80 and analysis of related substances

Analytical data of OPT-80 (most of which consists of the most active ingredient of OPT-80, R-thiacumiscin B) and three related compounds (S-thiacumiscin B, ripamycin A4, and C-19 ketone) Is summarized below. The structures of the three compounds are shown in formula VIII in Table 2 below.

[Table 2]

Structure of R-Tiacumicin B (most active ingredient of OPT-80) and related substances

compound X Y Z R-Tiacumicin B ethyl (R) -OH Isopropyl S-Tiacumicin B ethyl (S) -OH Isopropyl Ripiamycin A4 methyl (S) -OH Isopropyl C-19 Ketone ethyl = O Isopropyl

R- Tiacumicin  Analysis data of B

mp 166-169 ° C (white needles from isopropanol);

^{_{[α] D 20 - 6.9 (}} c 2.0, MeOH);

MS m / z (ESI) 1079.7 (M + Na) ⁺ ;

¹ H ¹ H NMR NMR (400 MHz, CD ₃ OD) δ 7.21 (d, 1H), 6.59 (dd, 1H), 5.95 (ddd, 1H), 5.83 (br s, 1H), 5.57 (t, 1H) , 5.13 (br d, 1H), 5.09 (t, 1H), 5.02 (d, 1H), 4.71 (m, 1H), 4.71 (br s, 1H), 4.64 (br s, 1H), 4.61 (d, 1H), 4.42 (d, 1H), 4.23 (m, 1H), 4.02 (pentet, 1H), 3.92 (dd, 1H), 3.73 (m, 2H), 3.70 (d, 1H), 3.56 (s, 3H ), 3.52-3.56 (m, 2H), 2.92 (m, 2H), 2.64-2.76 (m, 3H), 2.59 (heptet, 1H), 2.49 (ddd, 1H), 2.42 (ddd, 1H), 2.01 ( dq, 1H), 1.81 (s, 3H), 1.76 (s, 3H), 1.65 (s, 3H), 1.35 (d, 3H), 1.29 (m, 1H), 1.20 (t, 3H), 1.19 (d , 3H), 1.17 (d, 3H), 1.16 (d, 3H), 1.14 (s, 3H), 1.12 (s, 3H), 0.87 (t, 3H);

¹³ C NMR (100 MHz, CD ₃ OD) δ 178.4, 169.7, 169.1, 154.6, 153.9, 146.2, 143.7, 141.9, 137.1, 137.0, 136.4, 134.6, 128.5, 126.9, 125.6, 124.6, 114.8, 112.8, 108.8, 102.3, 97.2, 94.3, 82.5, 78.6, 76.9, 75.9, 74.5, 73.5, 73.2, 72.8, 71.6, 70.5, 68.3, 63.9, 62.2, 42.5, 37.3, 35.4, 28.7, 28.3, 26.9, 26.4, 20.3, 19.6, 19.2, 18.7, 18.2, 17.6, 15.5, 14.6, 14.0, 11.4.

S- Tiacumicin  Analysis data of B

To a solution of C-19 ketone (150 mg) in 3 mL of MeOH was added 3 parts of NaBH ₄ (9eq, 48 mg). After 1 h saturated NH ₄ Cl solution was added. The mixture was extracted with CHCl ₃ and then concentrated. S-Tiacumicin B was purified by YMC-pack ODS-A 75 × 30 mm ID column (H ₂ O: MeOH: AcOH 28: 72: 1) and yield yielded a pure 35 mg of pure S-Tiacumicin B. .

MS m / z 1074.5 (M + NH ₄ ) ⁺ ;

¹ H NMR (400 MHz, CDCl ₃ ) δ 7.15 (d, J = 11.4 Hz, 1H), 6.58 (dd, J = 14.1, 11.4 Hz, 1H), 5.82 (ddd, J = 14.1, 10.6, 3.5 Hz, 1H), 5.78 (s, 1H), 5.40 (dd, J = 7.8, 7.8 Hz, 1H), 5.15 (dd, J = 9.5, 9.5 Hz, 1H), 5.01 (d, J = 9.9 Hz, 1H), 5.01 (d, J = 9.9 Hz, 1H), 4.77 (ddd, J = 5.8, 5.3, 5.3 Hz, 1H), 4.68 (d, J = 11.6 Hz, 1H), 4.65 (br s, 1H), 4.62 ( br s, 1H), 4.42 (d, J = 11.6 Hz, 1H), 4.28 (br s, 1H), 4.07-3.97 (m, 2H), 3.74-3.58 (m, 4H), 3.61 (s, 3H) , 3.52 (dq, J = 9.5, 5.8 Hz, 1H), 3.08 (dq, J = 12.6, 6.1 Hz, 1H), 3.01 (dq, J = 12.6, 6.1 Hz, 1H), 2.77-2.65 (m, 2H ), 2.60 (heptet, J = 6.9 Hz, 1H), 2.55-2.44 (m, 3H), 1.95-1.84 (m, 1H), 1.80 (s, 3H), 1.76 (s, 3H), 1.66 (s, 3H), 1.34 (d, J = 5.8 Hz, 3H), 1.29-1.24 (m, 1H), 1.27 (d, J = 6.6 Hz, 3H), 1.21 (t, J = 6.1 Hz, 3H), 1.19 ( d, J = 6.9 Hz, 3H), 1.18 (d, J = 6.9 Hz, 3H), 1.15 (s, 3H), 1.10 (s, 3H), 0.84 (t, J = 7.2 Hz, 3H);

¹³ C NMR (100 MHz, CDCl ₃ ) δ 177.4, 170.1, 168.8, 157.6, 152.8, 144.4, 143.1, 141.1, 136.7, 136.2, 134.9, 133.8, 128.7, 125.7, 125.2, 123.0, 113.9, 107.5, 107.2, 101.7 , 94.9, 92.6, 80.8, 79.2, 76.6, 74.8, 73.5, 72.7, 71.9, 71.7, 70.2, 70.1, 69.5, 63.5, 62.3, 41.5, 36.6, 34.3, 29.5, 28.2, 26.2, 26.0, 19.4, 19.3, 18.9 , 18.5, 17.8, 17.3, 15.3, 14.1, 13.7, 11.1;

Ripiamycin  A ₄ Analysis data of

MS m / z 1060.5 (M + NH ₄ ) ⁺ ;

¹ H NMR (400 MHz, CDCl ₃ ) δ 7.12 (d, J = 11.6 Hz, 1H), 6.59 (dd, J = 14.1, 11.6 Hz, 1H), 5.85 (br s, 1H), 5.83 (ddd, J = 14.1, 10.6, 4.8 Hz, 1H), 5.47 (dd, J = 8.3, 8.3 Hz, 1H), 5.12 (dd, J = 9.6, 9.6 Hz, 1H), 5.00 (d, J = 10.1 Hz, 1H) , 4.98 (br d, J = 10.6 Hz, 1H), 4.75-4.69 (m, 1H), 4.68 (d, J = 11.4 Hz, 1H), 4.66 (br s, 1H), 4.62 (br s, 1H) , 4.40 (d, J = 11.4 Hz, 1H), 4.26 (br s, 1H), 4.07-4.00 (m, 1H), 4.02 (br d, J = 3.3 Hz, 1H), 3.75-3.61 (m, 4H ), 3.62 (s, 3H), 3.55 (dq, J = 9.6, 6.1 Hz, 1H), 2.82-2.45 (m, 6H), 2.60 (s, 3H), 2.07-1.97 (m, 1H), 1.92 ( s, 3H), 1.81 (s, 3H), 1.67 (s, 3H), 1.32 (d, J = 6.1 Hz, 3H), 1.30-1.22 (m, 1H), 1.21 (d, J = 6.6 Hz, 3H ), 1.19 (d, J = 7.1 Hz, 3H), 1.18 (d, J = 7.1 Hz, 3H), 1.15 (s, 3H), 1.10 (s, 3H), 0.83 (t, J = 7.2 Hz, 3H );

¹³ C NMR (100 MHz, CDCl ₃ ) δ 177.4, 170.5, 168.9, 157.8, 153.0, 144.3, 140.9, 137.7, 137.0, 136.3, 134.6, 134.4, 129.1, 127.9, 125.3, 123.2, 114.5, 107.4, 107.0, 101.8, 94.7, 92.5, 80.3, 79.6, 76.7, 74.9, 73.5, 72.7, 71.9, 71.6, 70.2, 70.1, 69.1, 63.6, 62.3, 41.9, 36.9, 34.4, 28.8, 28.2, 25.9, 20.0, 19.3, 19.0, 18.6, 18.5, 17.8, 17.2, 15.5, 13.8. 11.2;

Analytical Data of C-19 Ketones

MS m / z 1072.5 (M + NH ₄ ) ⁺ ;

¹ H NMR (400 MHz, CDCl ₃ ) δ 7.27 (d, J = 11.4 Hz, 1H), 6.61 (dd, J = 14.7, 11.4 Hz, 1H), 5.91 (ddd, J = 14.7, 9.1, 5.8 Hz, 1H), 5.83 (s, 1H), 5.31 (dd, J = 7.9, 7.9 Hz, 1H), 5.14 (dd, J = 9.7, 9.7 Hz, 1H), 5.06 (d, J = 10.6 Hz, 1H), 5.00 (d, J = 10.1 Hz, 1H), 4.98 (dd, J = 7.1, 4.8 Hz, 1H), 4.67 (d, J = 11.9 Hz, 1H), 4.66 (br s, 1H), 4.61 (br s , 1H), 4.42 (d, J = 11.9 Hz, 1H), 4.30 (br s, 1H), 4.02 (br d, J = 3.3 Hz, 1H), 3.63-3.60 (m, 4H), 3.62 (s, 3H), 3.51 (dq, J = 9.7, 6.1 Hz, 1H), 3.09 (dq, J = 14.4, 7.3 Hz, 1H), 3.03 (dq, J = 14.4, 7.3 Hz, 1H), 2.76-2.50 (m , 6H), 2.21 (s, 3H), 1.93-1.87 (m, 1H), 1.87 (s, 3H), 1.75 (s, 3H), 1.63 (s, 3H), 1.32 (d, J = 6.1 Hz, 3H), 1.27-1.22 (m, 1H), 1.21 (t, J = 7.3 Hz, 3H), 1.19 (d, J = 7.1 Hz, 3H), 1.18 (d, J = 7.1 Hz, 3H), 1.14 ( s, 3H), 1.10 (s, 3H), 0.84 (t, J = 7.3 Hz, 3H);

¹³ C NMR (100 MHz, CDCl ₃ ) δ 205.5, 177.4, 170.1, 166.9, 157.6, 152.8, 145.7, 143.1, 142.0, 137.1, 136.8, 135.5, 133.7, 128.3, 124.8, 124.0, 122.8, 113.9, 107.3, 107.2 , 101.3, 94.8, 92.4, 80.4, 77.7, 76.6, 74.7, 73.5, 72.6, 71.8, 71.7, 70.2, 70.0, 63.0, 62.3, 41.5, 36.5, 34.3, 29.6, 28.1, 26.2, 26.1, 26.0, 19.2, 18.9 , 18.5, 17.8, 17.3, 15.2, 14.0, 13.3, 11.0

Example  3: biological activity

Some C. Difficile  Measured for strains MIC  shame

OPT-80 (mostly made up of R-tiacumicin B) and related mixtures were tested for C. difficile. MIC values are presented in Table 3 below. As can be observed, OPT-80 was particularly active when compared to S-thiacumycin B and ripamycin A4.

[Table 3]

Table 3: MIC (μg / ml) vs. C. difficile strains

C. difficile strain OPT-80
(Mostly made up of R-tiacumicin B) S-Tiacumicin B Ripiamycin A4 C-19 Ketone ATCC 9689 0.03 0.125 0.06 0.06 ATCC 43255 0.125 One 0.5 0.5 ATCC 17857 0.03 0.125 0.06 nd LC # 1
(Clinical isolate) 0.125 One 0.5 0.5

Measured for various microorganisms MIC  Figures

OPT-80 (mostly made up of R-tiacumicin B) and related mixtures were tested for several other pathogens. MIC values are presented in Table 4 below. As can be observed, OPT-80 was particularly active when compared to S-thiacumycin B and ripamycin A4.

[Table 4]

MIC for other microorganisms (µg / ml)

Strain ID # organism OPT-80
(Mostly made up of R-tiacumicin B) S-Tiacumicin B Ripiamycin A4 One S. Aureus
(ATCC 29213) 4 64 8 2 S. Aureus
(MRSA) 4 64 16 3 S. Aureus
(MRSA) 4 64 8 4 E. Paecalis
(ATCC 29212) 2 8 2 5 E. Paecalis
Vancomycin resistance 4 32 16 6 E. Paecalis
Vancomycin resistance One 16 4 7 E. Paesium
Vancomycin resistance One 8 4 8 E. Paesium
Vancomycin resistance One 32 32

Example  4: C. At difficile OPT Post-Antibiotic Effect of -80

Post-antibiotic effect (PAE) of OPT-80 (mostly consisting of R-thiacumycin B) was measured for two strains C. difficile, ATCC 43255 and clinical isolate, LC3.

PAE at 4 × MIC was observed to be significantly longer: at least 24 hours for both strains. The long-term persistence of these effects prevented the accurate calculation of PAE. Vancomycin, on the other hand, had a general PAE of less than one hour when used at 4 × MIC for strain LC3.

Example  5: OPT In vitro activity of -80

In vitro efficiencies of OPT-80 (mostly made up of R-tiacumicin B), metronidazole, and vancomycin were measured for 110 genetically distinct clinical isolates of C. difficile via agar dilution. MIC data are present in Tables 5 and 6.

[Table 5]

Geometric mean, MIC range, MIC ₅₀ and MIC ₉₀ values (μg / mL) of OPT-80 for 110 C. difficile isolate, vancomycin, and metronidazole.

range Geometric mean MIC ₅₀ MIC ₉₀ OPT-80 0.015-0.25 0.08 0.125 0.125 Metronidazole 0.025-0.5 0.15 0.125 0.25 Vancomycin 0.06-4 0.8 One One

TABLE 6

C. MIC raw data (μg / mL) of OPT-80, vancomycin (VAN), and metronidazole (MTZ) for 110 clinical isolates of difficile.

Example  6: for selected anaerobic species OPT Comparative activity of -80

In vitro activity of OPT-80 was measured against 350 anaerobes. Experimental methods are presented in "Antibacterial Agents and Chemotherapy" (2004, 48: 4430-4434), which is hereby incorporated by reference in its entirety.

All organisms, including the 21 C. difficile strains, were isolated as isolates and ungrouped. Good controls of all Gram positive and negative strains recommended by NCCLS were included in each experiment. In all cases, results (if available) were within range.

The results of the MIC experiment are shown in Table 7.

[Table 7]

MICs of OPT-80 (μg / mL)

Example  7: against gut bacteria OPT Extracorporeal activity of -80

In vitro activity of OPT-80 against enteric bacteria was evaluated. Experimental methods are presented in "Antibacterial Agents and Chemotherapy" (2004, 48: 4898-4902), which is hereby incorporated by reference in its entirety.

The antimicrobial concentration range was chosen to include or exceed the values that can be obtained in the intestine, provided that the solubility limits of the drug in the test medium are within (within the limits available in this document). The concentration range of OPT-80 used during the test was 0.03 μg / mL to 1024 μg / mL.

For analysis, the bacteria tested were generally set up with at least 10 isolates in genus, species, or other groups. MIC ranges of isolates and MIC values inhibited at 50% and 90% were measured excluding organisms of less than 10 strains, which reported only the range (Table 8).

OPT-80 has good activity against most anaerobic Gram-positive spore-forming rods and anaerobic Gram-positive cocci. OPT-80 also showed good activity against enterococci and staphylococci.

[Table 8]

453 bacteria In segregation  About OPT Extracorporeal activity of -80

Etc Example

All references cited above are hereby incorporated by reference in their entirety for all purposes. While the present invention has been shown and described with preferred embodiments, it is to be understood that various changes in form and detail may be made therein without departing from the spirit and scope of the invention as defined by the appended claims. It will be understood by the technicians.

Included in this specification.

Claims (36)

Clostridium difficile ( C. difficile ) gastrointestinal tract in a mammal comprising an optically pure compound of formula IV, a compound of formula III, and a compound of formula II, or a pharmaceutically acceptable salt thereof Pharmaceutical compositions, which are oral medications for treating diarrhea caused by infection:

, And

. The method of claim 1, Wherein the pharmaceutical composition further comprises a compound of the formula:

. 3. The method according to claim 1 or 2, Pharmaceutical composition wherein the pharmaceutical composition is formulated as a tablet. 3. The method according to claim 1 or 2, Pharmaceutical composition wherein the pharmaceutical composition is formulated in a capsule. 3. The method according to claim 1 or 2, Pharmaceutical composition wherein the pharmaceutical composition is formulated in a suspension (suspension). 3. The method according to claim 1 or 2, Wherein said compound of formula IV is at least 90% by weight of the total antibiotic. 3. The method according to claim 1 or 2, Wherein said compound of formula IV does not comprise different diastereomers of that compound. 3. The method according to claim 1 or 2, Wherein said compound of formula III does not comprise different diastereomers of that compound. The method of claim 2, A pharmaceutical composition wherein the compound of the above formula does not comprise different diastereomers of that compound. delete delete delete delete delete delete delete delete delete delete delete delete delete delete delete delete delete delete delete delete delete delete delete delete delete delete delete

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